Method and apparatus for tissue grafting and copying

Abstract

To provide an apparatus and method for obtaining one or more portions of biological tissue (micrografts) to form grafts.SOLUTION: A hollow tube 1010 can be inserted into tissue at a donor site, and a pin 1040 provided within the tube can facilitate controlled removal of micrografts from the tube. Micrografts can be harvested and directly implanted into an overlying biocompatible matrix through coordinated motion of the tube and pin. A needle can be provided around the tube to facilitate a direct implantation of a micrograft into a remote recipient site or matrix. An exemplary apparatus 1100 can include a plurality of such tubes and pins for simultaneously harvesting and / or implanting a plurality of micrografts. The harvested micrografts can have a small diameter, e.g., less than about 1 mm, which can promote healing of the donor site and / or viability of the harvested tissue.SELECTED DRAWING: Figure 11

Description

Technical Field

[0001] Cross - Reference to Related Applications This application is related to and claims priority from U.S. Provisional Patent Application No. 61 / 332,230, filed May 7, 2010; U.S. Provisional Patent Application No. 61 / 373,498, filed Aug. 13, 2010; and U.S. Provisional Patent Application No. 61 / 437,507, filed Jan. 28, 2011. The disclosures of those applications are hereby incorporated by reference in their entireties.

[0002] The present invention relates to exemplary embodiments of methods and apparatuses for providing tissue grafts and for replicating tissue structures, for example, using tissue from a donor site.

Background Art

[0003] An autograft may refer to tissue transplanted from one part of an individual's body (e.g., a "donor site") to another part (e.g., a "recipient site"). Autografts can be used, for example, to replace lost skin and other tissues and / or to promote healing from trauma, wounds, burns, surgical, and birth abnormalities. The availability of tissue for autografting can be limited by the characteristics of the candidate donor site, including the number and / or overall area of transplantable tissue pieces, the healing behavior of the donor site, the similarity between the donor site and the recipient site, and aesthetic considerations.

[0004] Skin grafts can be performed surgically. For example, a conventional autograft procedure can include the excision or surgical removal of tissue damaged by burns, the selection of a donor site that can be an area removed from healthy skin used to cover the clean burn area, and, if the graft can be removed from the donor site, the collection using a device similar to, for example, an electric shaver. Such a device (e.g., a dermatome) can be structured to gently shave a tissue piece that can be about 10 / 1000 inches thick for split-thickness skin grafting from the skin of the non-burned donor site for use as a skin graft. The skin graft can then be placed on the clean wound site so that it can heal. The donor skin tissue can be removed to a depth such that the donor site can heal naturally in a process similar to the healing of a second-degree burn.

[0005] Two conventional types of autografts that can be used for covering permanent wounds include sheet grafts and mesh grafts. A sheet graft can mean a small piece of skin tissue removed from a non-burned donor site of the body in a process called collection. The size of the small piece of donor skin used can be approximately the same size as the damaged area. The sheet graft can be applied over the excised wound site and can be held or fixed in place. The donor skin tissue used for the sheet graft cannot stretch significantly, and a sheet graft slightly larger than the damaged area to be covered can be obtained because, often, after collection, the transplanted tissue can shrink slightly.

[0006] Sheet grafts can provide an improved appearance of the repaired tissue site. For example, when damaged, sheet grafts can be used on large areas of the face, neck, and hands, so many visible parts of the body can appear to have few scars after healing. Sheet grafts can be used, for example, when the damaged site is narrow, to cover the entire area of the burn or damaged skin. The narrow area of the sheet graft can be damaged after placement because, after placement of the sheet graft, the accumulation of body fluids (e.g., a hematoma) can occur under the sheet graft.

[0007] The sheet graft is a full-thickness skin graft or a split-thickness skin graft. For example, split-thickness skin grafting can be used to cover wounds in burn and skin ulcer patients. Conventional split-thickness grafts can be formed, for example, by collecting a sheet of epidermal and epithelial tissue from the donor site in a manner similar to peeling an apple. The split-thickness skin graft can then be placed at the site of the burn or ulcer. The skin tissue can then usually grow back to its original state at the donor site after a long healing time. Split-thickness grafting may be more suitable than full-thickness grafting. This is because removing a large amount of full-thickness skin tissue from the donor site can cause scarring, a long healing time, and a high risk of infection at the donor site. However, the tissue removed from the donor site for a split-thickness autograft may contain only a thin epithelial layer and may lack certain elements of the dermis that improve structural stability and normal appearance at the recipient site.

[0008] A full-thickness skin graft can be formed using a sheet of tissue that includes the entire epidermal layer and dermal components of various thicknesses. Since the dermal components can be preserved in a full-thickness graft, many of the characteristics of normal skin can be maintained after the grafting procedure. A full-thickness graft can contain a greater amount of collagen, the dermal vascular network, and epidermal appendages compared to a split-thickness graft. However, a full-thickness graft may require more precise conditions for survival because it requires more tissue to revascularize.

[0009] Full-thickness skin grafts may be suitable, for example, for repairing visible areas of the face for grafting procedures that cannot be treated by local skin flaps or for which local skin flaps are contraindicated. Such full-thickness skin grafts can retain many of the characteristics of normal skin, such as, for example, color, texture, and thickness, compared to split-thickness skin grafts. Full-thickness skin grafts also can hardly shrink during healing. These characteristics can be important in many visible areas such as the face and hands. Furthermore, full-thickness grafts in children may have a high potential to grow with the individual. However, the application of conventional full-thickness skin grafts can be limited to relatively small, uncontaminated, well-vascularized wounds, and thus may not be as suitable for as many types of grafting procedures as split-thickness grafts. Additionally, donor sites for full-thickness grafts may require new skin using surgical sutures or split-thickness grafts.

[0010] For example, due to the lack of sufficient areas of healthy donor sites, mesh skin grafts can be used to cover open wounds of large areas that may be difficult to cover using sheet grafts. The mesh of a skin graft can facilitate expansion to cover a large area with skin tissue from the donor site. It also can facilitate the drainage of blood and body fluids from under the skin graft when placed on a wound and can help prevent graft loss. The expansion ratio of a mesh graft (e.g., the ratio of the non-expanded graft area to the expanded graft area) can typically be between about 1:1 and 1:4. For example, the donor skin may be meshed at a ratio of about 1:1 to 1:2, but larger expansion ratios may result in more fragile grafts, scarring of the mesh graft as it heals, and / or longer healing times.

[0011] Conventional graft mesh procedures can involve subjecting donor skin tissue to a machine that cuts incisions into the tissue and can facilitate expansion in a pattern similar to a fishnet or wire mesh fence. Healing can occur as the graft expands through the spaces between the meshes, which may be referred to as gaps or interstices that are filled with new epithelial growth. However, mesh grafts can result in grafts that are less durable than sheet grafts, and large meshes may result in permanent scarring after graft healing.

[0012] To assist in the healing and safety of the graft, the area of the graft may preferably be immobilized (e.g., not moved) for at least about 5 days after each surgery. During this immobilization period, blood vessels can grow from the underlying tissue into the skin graft and help bind the two tissue layers together. Approximately 5 days after the graft is placed, physical therapy programs, bathing, and other normal daily activities can often be resumed. Deep second-degree and full-thickness burns may require skin grafting surgery to heal quickly and with minimal scarring. The size of a large burn may lead to more than one graft procedure during hospitalization and may require long-term immobilization for healing.

[0013] As an alternative to autografting, skin tissue obtained from recently deceased individuals (e.g., a homograft, allograft, or cadaver skin as it may be referred to) can be used as temporary skin for a debrided wound area. The non-meshed cadaver skin can be placed over the excised wound at a given location and immobilized. After surgery, the cadaver skin can be covered with a dressing. The wound coverage using cadaver allograft can then be removed prior to a permanent autograft.

[0014] A xenograft or heterograft can refer to skin obtained from various animals, such as parts of pigs. Skin tissue from a xenograft can also be used for temporary coverage of an excised wound until a sufficient permanent autograft can be placed, which can be used due to the limited availability and / or high cost of human skin tissue. In some cases, religious, financial, or cultural objections to using human cadaver skin can also be a factor leading to the use of xenografts. Wound coverage using xenografts or allografts is usually a temporary measure that can be used until an autograft can be harvested and placed.

[0015] Accessory epithelial organs can be regenerated after transplantation. For example, hair may tend to grow from full-thickness grafts rather than split-thickness grafts, but such hair growth may be undesirable based on the location of the wound. Thus, donor sites for full-thickness grafts can be carefully selected, for example, based in part on the pattern of hair growth at the time of surgery. Additionally, certain hair follicles may not be oriented perpendicular to the skin surface, and they can be transected if the incisions given to remove the transplanted tissue are not properly oriented.

[0016] Sweat glands and sebaceous glands located in the transplanted tissue may initially degenerate after transplantation. These structures may tend to regenerate in full-thickness grafts rather than split-thickness grafts. This is because full-thickness grafts can be transferred as complete functional units. For example, sweat gland regeneration may depend in part on nerve regeneration of the skin graft along with the recipient's basic sympathetic nerve fibers. Once such internal growth occurs, the skin graft may assume the sweating properties of the recipient site rather than retaining the properties of the donor site. In contrast, sebaceous gland regeneration may be independent of transplanted nerve regeneration and may retain the properties of the donor site. Prior to regeneration, the skin graft tissue may lack the normal lubrication by these glands, making such grafts more sensitive to injury.

[0017] Generally, the transplantation procedure can be limited by the amount of tissue that can be removed from the donor site without causing excessive side effects. Full-thickness grafts can provide improved tissue quality at the wound site, but the donor site may be damaged quite severely as described above. Split-thickness grafts can result in a compromise between healing time and the aesthetic and functional characteristics of the donor and recipient sites, whereas meshes can provide a more extensive graft coverage at the expense of visible scarring.

[0018] The collection of transplanted tissue from the donor site may generally cause undesired large-scale tissue damage to the donor site. On the other hand, a narrow area of skin wound adjacent to healthy tissue can exhibit good tolerance and can heal rapidly. The healing of such minor wounds can be achieved in techniques such as "fractional photothermolysis" or "fractional resurfacing", where a damage pattern having small dimensions can be generated in the skin tissue. These exemplary techniques are described, for example, in Patent Document 1 and Patent Document 2. The small-scale damage pattern can heal rapidly by the regeneration of healthy tissue and can further provide desired effects such as skin tightening without causing visible scarring.

Prior Art Documents

Patent Documents

[0019]

Patent Document 1

Patent Document 2

Summary of the Invention

Problems to be Solved by the Invention

[0020] In view of the drawbacks of the above-described procedures for tissue transplantation, it may be desirable to provide exemplary embodiments of methods and apparatuses that can provide tissue suitable for transplantation while minimizing undesired damage to the donor site, for example.

Means for Solving the Problems

[0021] Exemplary embodiments of the present disclosure provide a method and apparatus for obtaining a small portion of transplant tissue that can be accompanied by rapid healing of the donor site. For example, an exemplary embodiment of this method can be provided for obtaining a skin transplant tissue by collecting a small portion of tissue, such as a micrograft, from the donor site. Such micrografts can be used to form grafts or "replicated" tissue to generate a larger tissue structure from a small tissue sample.

[0022] Such micrografts can include, for example, skin tissue that can include epidermal and dermal tissue and / or tissue obtained from other body organs. The micrografts can have at least one diameter that is relatively small, for example, less than about 1 mm, or less than about 0.5 mm, or less than about 0.3 mm, or about 0.2 mm as required. Such an exemplary small diameter of the micrograft can promote both the survival of the micrograft by enabling healing of the donor site after collection and more extensive diffusion of nutrients to the micrograft tissue. The small area of damage at the donor site caused by the removal of the tissue portion can heal rapidly with little or no visible scarring. Micrografts obtained from skin tissue can include, for example, epidermal and dermal tissue and may also include stem cells located proximate to the dermal / fat layer boundary. Micrografts may also be obtained from other types of tissue, such as various internal organs and the like.

[0023] The proportion of dermal tissue removed from the donor site is, for example, less than about 70%, or less than about 50%, although other proportions may be used. The collected tissue portion may be in a shape such as a cylindrical shape, an elongated piece, or other shapes that can include at least one small dimension. In certain exemplary embodiments, a portion of the tissue at the donor site can be frozen or partially frozen. Such freezing can facilitate the cutting, removal, and / or survival of the tissue portion to be collected.

[0024] In a further exemplary embodiment of the present disclosure, an apparatus for collecting micro-grafts may be provided that includes at least one hollow tube with pins provided at least partially within the tube, the pins being controllably movable in a direction along the longitudinal axis of the tube. The diameter of the pins may be substantially the same as the inner diameter of the lumen of the tube or may be slightly smaller.

[0025] Such a tube and corresponding pins may be mechanically attached to a substrate having at least one hole therethrough. A plurality of linear actuators may be provided to controllably position and / or move at least one tube and pin relative to the substrate. For example, the actuator may be configured to position at least one tube through a hole in the substrate and control the distance that the distal end of the tube protrudes from the lower surface of the substrate. The actuator may further be configured to independently control the position of the pins within the tube. The substrate may be configured to be placed on the surface of the tissue to facilitate the collection of micro-grafts from the tissue and / or the implantation of micro-grafts into the tissue.

[0026] In a further exemplary embodiment of the present disclosure, a plurality of tubes and corresponding pins may be provided to facilitate the collection of a plurality of micro-grafts. A plurality of actuators or mechanical configurations may be provided in communication with the proximal ends of the tubes and / or pins to facilitate the precise positioning and movement of the plurality of tubes and pins relative to each other and to the tissue. The actuator may be configured to move all of the tubes and / or pins simultaneously or, if desired, to move a particular one of the tubes or pins independently of the other tubes. A vibration configuration may be attached to the apparatus to facilitate the insertion of the tubes into the donor and / or recipient sites.

[0027] Exemplary micro-grafts can be placed in a biocompatible matrix, for example, to form grafts or larger replicas at the donor site, or can be directly implanted into the tissue at the recipient site. The biocompatible matrix can be formed using collagen, polylactic acid, hyaluronic acid, and / or other substances that can support the collected micro-graft tissue portions and promote their growth. The matrix may optionally contain nutrients, growth factors, and / or other substances to promote tissue growth or survival. The collected tissue portions may be bound to the matrix using techniques such as photochemical tissue bonding to provide structural stability. The matrix can then be applied to the recipient site and can promote the growth and revascularization of the tissue portions to form a continuous sheet of transplanted tissue. Optionally, the matrix can be placed in a suitable environment to promote the growth of the micro-grafts and then form a larger portion of the "replicated" tissue from the donor site.

[0028] Exemplary micro-grafts may also be assembled in a small structure to form a transplant tissue that can be directly applied to the recipient site. Exemplary micro-grafts may also be directly inserted into the tissue at the recipient site, for example, using the exemplary hollow tubes described herein, at the recipient site such as scar tissue. In certain exemplary embodiments, the micro-grafts may be collected from donor site tissue that is different from the tissue type at the recipient site to form various types of xenografts.

[0029] Even more exemplary embodiments of the present disclosure can provide methods for extracting or collecting micro-grafts from tissue and, optionally, directly placing them in a matrix material. For example, such exemplary methods can utilize an exemplary device comprising at least one tube and a pin provided for collecting tissue as follows. A) A portion or sheet of the matrix material can be placed at the donor tissue site, and the distal ends of the tube and the pin provided within the tube can each be placed in proximity to the upper surface of the matrix material. B) The tube and the pin can be moved together so as to penetrate the matrix and can be positioned adjacent to the surface of the donor site. C) Then, with the distal end of the pin remaining on the surface of the donor site, the tube can be moved downward into the tissue of the donor site to cut a portion of the tissue from the surrounding tissue. D) Then, the tube and the pin can be retracted simultaneously until the distal end of the tube is adjacent to the lower surface of the matrix material or within the matrix material, and the micro-graft containing the cut tissue remains within the distal portion of the tube. E) Then, while the position of the pin is held substantially fixed relative to the matrix, the tube can be further retracted from the matrix, whereby when the tube is retracted from around the micro-graft, the micro-graft is held within the matrix material by the fixed pin and remains within the matrix.

[0030] In yet another exemplary embodiment of the present disclosure, a perforated needle may be provided around at least a portion of a hollow tube and a pin. An apparatus comprising at least one hollow tube, a pin, and a perforated needle may be configured to place micro grafts contained within the distal portion of the hollow tube into tissue located at a recipient site. The perforated needle may be inserted into the tissue at the recipient site. The hollow tube containing the micro grafts collected at the distal ends of the pin and the hollow tube may be advanced through the perforated tube such that the distal end of the hollow tube is proximate to the surface of the recipient site, or beneath the surface. While holding the hollow tube fixed, the perforated needle may be withdrawn from the recipient site. Then, while holding the pin within the tube fixed relative to the recipient site, the hollow tube may be withdrawn. In this way, the micro grafts may remain within the tissue of the recipient site, for example, being held in the recipient site by the pin when the hollow tube is withdrawn. The pin may subsequently be withdrawn such that the micro grafts are left within the recipient site where the tissue was first separated by the perforated needle. A substrate may be provided with at least one opening therethrough, the substrate being configured to facilitate positioning of the needle, tube, and / or pin relative to the tissue disposed on and being treated on the tissue surface, and / or to facilitate mechanical stability of the tissue during treatment.

[0031] These and other objects, features and advantages of the present disclosure will become apparent upon reading the following detailed description of the exemplary embodiments of the present disclosure in conjunction with the appended claims.

[0032] Further objects, features and advantages of the present disclosure will become apparent from the following detailed description in conjunction with the accompanying drawings showing exemplary embodiments of the present disclosure, results and / or features of the exemplary embodiments.

Brief Description of the Drawings

[0033]

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DETAILED DESCRIPTION OF THE INVENTION

[0034] Throughout the drawings, unless otherwise stated, the same reference numerals and characters are used to denote similar features, elements, components, or parts of the illustrated embodiments. Further, the present disclosure is described in detail herein with reference to the drawings, which are described in connection with the illustrated embodiments, and the present invention is not limited to the specific embodiments illustrated in the drawings.

[0035] According to exemplary embodiments of the present disclosure, methods and apparatuses for generating autograft, in particular, methods and apparatuses that can promote more rapid healing of the donor site while providing improved tissue characteristics at the recipient site can be provided. Exemplary embodiments of the present disclosure may include a plurality of small-scale tissue portions (e.g., micrografts) that can be used to provide autograft. Such grafts can provide transplant tissue that can heal rapidly and regenerate skin tissue having desirable characteristics at the recipient site while avoiding permanent damage to the donor site.

[0036] In an exemplary embodiment of the present disclosure, as shown in FIG. 1A, a method of making an autograft can be provided in which a tissue portion (e.g., micrograft) having at least one small dimension is collected from an exemplary donor site 100. The holes 110 shown in FIG. 1A represent the regions of the exemplary donor site 100 from which the tissue portion (e.g., micrograft) is removed. These exemplary holes 110 may have a generally circular cross-sectional shape, although other shapes may be used.

[0037] An exemplary donor site 110 after healing of the collected tissue has occurred is shown in FIG. 1B. The small region 100 of damage created at the donor site by the removed tissue can heal rapidly and / or have no visible scarring. For example, the remaining pattern of the healed donor site 100 shown in FIG. 1B is not readily perceptible to the naked eye under normal visual conditions.

[0038] For example, FIG. 1C shows an exemplary micrograft 120 that can be formed by collecting or removing a portion of tissue from a donor site 100 to form a hole 110 in the donor site 100. The exemplary micrograft 120 may have an elongated shape that can be substantially cylindrical. The micrograft 120 may include both epidermal tissue 130 and dermal tissue 140 from the exemplary donor site 100. For example, the exemplary micrograft 120 may be about 3 mm in length, which may correspond to the entire depth of a typical skin layer (e.g., the epidermal and dermal layers). Different lengths may be used based on the specific skin or tissue characteristics of the donor site 100. Generally, since it may be preferable to avoid collecting a significantly large amount of subcutaneous tissue, the collected micrograft 120 may mainly include epidermal tissue 130 and dermal tissue 140. The lower portion 150 of the exemplary micrograft 120 may also include stem cells that may be present in the lower part of the dermis of the donor site 100 (e.g., proximate to the dermal / fat layer boundary).

[0039] The width or diameter of the hole 110 that occurs during collection, which may correspond to approximately the diameter of the portion of the micrograft 120 being collected, may be less than about 2 mm, or less than about 1 mm. In certain exemplary embodiments of the present disclosure, the diameter or width of the micrograft 120 may be less than about 0.5 mm, less than about 0.3 mm, or less than about 0.2 mm. The size of the exemplary hole 110 may be selected, for example, based on the effect of being able to heal quickly and / or create a small diameter area in the donor site 100 without scarring, and being small enough to promote survival when transplanted or placed in a growth medium, and large enough to form a sufficient amount of transplanted and / or captured tissue that may be present in the donor tissue.

[0040] For example, living tissue may be provided with nutrients by diffusion transport over a distance of about 0.1 mm. Thus, exemplary micro-grafts 120 having at least one diameter that is less than about 0.5 mm, such as less than about 0.3 mm or, for example, about 0.2 mm in size, may exhibit improved survival, have a high likelihood of survival, and, when used in transplantation, they can grow. Such exemplary micro-grafts 120 can receive more nutrients (e.g., including oxygen) when placed at the recipient site prior to vascularization of the tissue.

[0041] Larger micro-grafts 120, for example having a width of about 1 - 2 mm, may also benefit from such diffusion transport of nutrients and may also have a higher likelihood of survival than significantly larger portions of transplanted tissue (e.g., conventional full-thickness, split-thickness, or mesh grafts). These larger sizes may be suitable for tissues that are heterologous and contain specific structures that can be preserved within a single micro-graft 120. For example, skin tissue has specific structures such as hair follicles, sebaceous glands, etc., and collecting micro-grafts 120 that are somewhat larger from the skin can help preserve their tissue structure when collected and transplanted. On the other hand, smaller micro-grafts, for example those less than about 0.5 mm or less than about 0.2 mm in width, may be suitable for relatively homogeneous tissues such as muscle tissue that have few or no large structures to be preserved in the tissue.

[0042] The proportion of surface tissue removed from the donor site 100 by collection (which may correspond to the proportion of the surface area of the exemplary donor site 100 occupied by the hole 110) may be less than about 70%, or more preferably less than about 50%. The proportion of tissue removed may be large enough to provide a sufficiently sized collected micro-graft 120 to generate a suitable sized graft therefrom, but small enough to promote rapid healing at the donor site 100 based on growth from the remaining undamaged tissue. Other proportions of tissue may be removed from the donor site 100 depending on factors such as the specific characteristics of the donor site 100, the size of the grafts required, and the total amount of donor site tissue available.

[0043] In a further exemplary embodiment of the present disclosure, as shown, for example, in FIG. 2A, the graft 200 may be provided by embedding or inserting a plurality of micro-grafts 120 into a biocompatible matrix 210. The exemplary matrix 210 containing the micro-grafts 120 may be exposed to nutrients to promote the growth of the collected micro-grafts 120 in order to form a continuous or substantially continuous layer of tissue in the graft 200 after growth has occurred. As shown in FIG. 2B, an exemplary graft 200, which may include a matrix 210 and micro-grafts 120, may be placed directly on a recipient site 220 (e.g., a clean wound area). The exemplary micro-grafts 120 may also contain stem cells, as described herein, and those stem cells may also promote healing and the integration of the exemplary micro-grafts 120 when transplanted into the recipient site 220. The recipient site 220 can provide nutrients and / or promote angiogenesis of the collected micro-grafts 120, thereby further improving their growth through the matrix 210 to ultimately fill the space separating the micro-grafts 120. For example, FIG. 2B shows the micro-grafts 120 after they have begun to grow within the surrounding matrix 210.

[0044] In one exemplary embodiment of the present disclosure, the micro-grafts 120 may be arranged in the matrix 210 at substantially the same intervals (e.g., similar areal density) when they are removed from the donor site 100. With this exemplary configuration, after the micro-grafts 120 grow and fill the space between them and the new tissue, an amount of transplanted tissue can be generated that can be approximately the same size as the entire collected area of the donor site 100. The average interval of the micro-grafts 120 within the matrix 210 may also be increased to form a transplanted tissue larger than the entire area of the collected donor site 100. The specific interval of the micro-grafts 120 in a particular graft 200 may be selected based on factors such as, for example, the size and percentage of damage of the donor site 100, the size of the recipient site 220 covered by the skin graft 200, the time for the micro-grafts 120 to regrow and generate a continuous tissue layer, the desired appearance of the transplanted recipient site, and the like. For example, the exemplary micro-grafts 120 may be spaced apart from a particular graft, which can provide a larger transplant area, but may also require a longer healing time and there may be a possibility of some visible scarring or texture in the healed graft 200.

[0045] In a further exemplary embodiment of the present disclosure, as shown in FIG. 3C, the tissue portion 320 may be collected in an elongated, narrow strip shape. One or more exemplary tissue pieces 320 may include both epidermal tissue 130 and dermal tissue 140, which is similar to the micro-grafts 120 in FIG. 1C. For example, the height of the exemplary tissue piece 320 may be about 3 mm or another length corresponding to the local depth of the dermal layer at the donor site 100. A deeper and / or shorter dermal layer depth may also be selected, for example, when collecting the tissue piece 320 based on the donor site and recipient site, the characteristics of the wound to be repaired by the transplant.

[0046] For example, as shown in FIG. 3A, by collecting such an exemplary tissue piece 320, a long and narrow groove 310 can be left in the donor area 100. The width of the groove 310 (and thus the width of the collected tissue piece 320) can be less than about 1 mm, or less than about 0.5 mm. In certain exemplary embodiments, the width of such a tissue piece can be less than about 0.3 mm or less than about 0.2 mm. As described herein, such small dimensions can facilitate the diffusive transport of nutrients to the transplanted tissue and can improve the survival of the collected tissue. The depth of the groove 310 from the skin surface can correspond to the height of the collected piece 320.

[0047] The percentage of the surface area of the exemplary donor site 310 removed from the tissue piece 320 can be less than about 70%, or less than about 50%. The factors that affect the selection of the parameters associated with the collected elongated tissue piece 320 (such as the width and the percentage of the area removed from the donor site) can be similar to those described above for the substantially cylindrical micro-graft 120. The length of the collected piece 320 can be selected based on factors such as, for example, the ease of cutting, removing, and treating the thin tissue piece 320, and the location of the donor site 100. As shown in FIG. 3B, the elongated groove 310 formed in the donor site can also heal quickly with little or no visible scarring due to the small side dimensions and the presence of adjacent healthy tissue that can support local tissue regeneration.

[0048] The collected pieces 320 can be placed in a biocompatible matrix similar to the matrix 210 shown, for example, in FIG. 2. The tissue pieces 320 may be arranged in a substantially parallel configuration, for example corresponding to the configuration of the grooves 310 of the donor site being removed. The spacing between the pieces 320 may alternatively be increased or decreased as desired compared to the spacing of the grooves 310 of the donor site 100, for example providing a larger overall area or a more densely packed graft of the transplanted tissue respectively. Such collected tissue pieces 320 may be used for a particular transplantation procedure. Because the large dimensions can promote angiogenesis and preserve the structure in the collected skin tissue that can improve the healing of the graft formed from the tissue pieces 320.

[0049] The tissue portions collected may be removed from donor sites of other shapes, including a tile pattern or a fractal-like shape. Generally, each removed tissue piece (and each corresponding hole or void in the donor site, for example) may have at least one small dimension of less than about 1 mm, or less than 0.5 mm. In certain exemplary embodiments, this small dimension may be less than about 0.3 mm, or about 0.2 mm.

[0050] In further exemplary embodiments of the present disclosure, the collected tissue portions can be placed in the recipient site in a high-density configuration. For example, FIG. 4A shows a schematic top view of a plurality of substantially cylindrical micro-grafts 120 that can be collected in an exemplary high-density arrangement where adjacent ones of the exemplary micro-grafts 120 are at least partially in direct contact with each other. FIG. 4B shows a schematic side view of the micro-grafts 120 shown in FIG. 4A. This exemplary high-density configuration can provide grafts that are smaller than the entire area of the donor site 100 being collected, but may tend to heal more quickly and result in less visible scarring than grafts formed using the tissue portions 120, 320 collected at intervals. An exemplary configuration of similarly collected tissue may also be formed using tissue pieces 320 as shown, for example, in FIG. 3C.

[0051] The exemplary biocompatible matrix 210 may be formed using one or more materials structured to provide mechanical stability and / or support the collected micro grafts 200 and / or promote tissue regeneration. Examples of materials that may be used to form the matrix 210 include polylactic acid (PLA), collagen (such as a collagen sponge), low melting point agarose (LMA), hyaluronic acid (such as hyaluranon), or inactivated or cadaver skin. The matrix 210 may be formed, for example, from allogeneic skin, such as by freezing and thawing a portion of donor skin tissue several times. For example, about 7 freeze / thaw cycles may be performed to effectively kill the cells at the donor site for use as the matrix 210. The frozen and thawed tissue may then be washed with a detergent or other composition to remove dead cells, debris, and the like.

[0052] In further exemplary embodiments of the present disclosure, a living donor skin tissue sample can be treated with radiation to form the matrix material 210. For example, the donor skin tissue can be treated with a lethal dose of x-rays, such as gamma rays, that can keep the cells intact. Thus, the cells in the donor matrix material can remain alive for a specific time, such as 48 to 72 hours, after radiation exposure before drying. Such a matrix 210 containing short-lived cells can initially support the growth of the transplanted micro graft 120, but then die before a significant adverse interaction, such as an autoimmune reaction, occurs against the growth and / or survival of the micro graft 120. Alternatively, transplantation of such a radiation-irradiated matrix 210 with viable micro grafts 120 can be delayed for a sufficient time, such as 72 hours or more, after radiation exposure to avoid an autoimmune reaction between the micro graft 120 and the matrix 210. The matrix 210 can also be made from donor skin, for example, by detaching the cells with detergents and acidic / basic solutions. Exemplary protocols for making such a matrix or scaffold are described, for example, in Alsberg et al., Proc Natl Acad Sci USA. September 17, 2002; 99(19): 12025-12030.

[0053] In further exemplary embodiments of the present disclosure, a portion of the matrix material can be mixed with the micro graft 120 in a configuration similar to that shown in FIGS. 4A and 4B, except that a portion of the pillar is formed from the matrix material, as described herein. The micro graft 120 and the matrix pillars can be adhered using, for example, adhesives, photochemical binders, and the like. The relative sizes and numbers of the portions of the micro graft 120 and the matrix material can be varied to produce a composition having a specific proportion of micro graft material.

[0054] Nutrients or other additives may also be provided to the matrix 210 to further promote tissue regeneration. Such additives may include, for example, one or more growth factors, stem cells. Examples of such growth factors include, but are not limited to, vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF), transforming growth factor beta (TGF-β), and fibroblast growth factor (FGF), which can improve or promote angiogenesis of the graft. Epidermal growth factor (EGF) and keratinocyte growth factor may also be used, which can increase the recruitment and differentiation of specific skin cells such as keratinocytes and fibroblasts. Platelet-rich crystals (which can be made from the patient's own blood using, for example, a commercially available system) may also be used to provide specific growth factors. Such plasma may require greater effort to produce than exogenous growth factors, but can be closer to the natural wound healing environment. Such growth factors can be introduced into the matrix 210, such as an LMA / collagen mix, at an appropriate temperature at which the matrix remains in liquid form. The LMA / collagen matrix can then be formed by adjusting the pH value of the collagen solution before mixing with the heated LMA. For the coagulation of the matrix 210, for example, the mixture can be stored at a temperature of about 37 °C for about 20 minutes before cooling to about 4 °C.

[0055] Stem cell sources may include, for example, stem cells derived from adipose tissue and / or mesenchymal stem cells derived from bone marrow. Micrografts obtained or collected from skin tissue may also contain stem cells derived from hair follicles. Such stem cells or other cells may be introduced into the matrix by culturing the cells with the matrix prior to transplantation of the micrograft. Low-level light therapy (LLLT) may also be used to promote the growth and survival of micrografts. For example, red or near-infrared light may be used to irradiate the donor site and / or recipient site after tissue collection and placement of the transplanted tissue to further promote tissue healing and / or growth.

[0056] Growth factors and / or other additives may be introduced into the matrix by, for example, immersing the matrix material in a solution containing the growth factor prior to implanting the micro grafts into the matrix material. Growth factors may also be introduced using a controlled release mechanism by, for example, directly moving the growth factors from a reservoir within the matrix over time or pumping them, or by encapsulating the growth factors in a biodegradable polymer that can be introduced into the matrix, and releasing them into the matrix over time. Specific growth factors or additives may also be bound to the matrix material by a chemical bond that can be cleaved over time, thereby gradually releasing the growth factor into the matrix.

[0057] In certain exemplary embodiments of the present disclosure, certain techniques such as photochemical tissue bonding may be used to improve the mechanical stability of the micro grafts 120 and / or tissue pieces 320 within the matrix 210. For example, techniques related to photochemical tissue bonding are described in U.S. Patent No. 7,073,510. This technique involves applying a photosensitizing agent to the tissue, followed by irradiation with electromagnetic energy to create a tissue seal. A photosensitizing agent such as Rose Bengal can be applied to the matrix 210 containing the exemplary micro pieces 120 and / or tissue pieces 320, and subsequently, the matrix can be exposed to green light for about 2 minutes. Photochemical tissue bonding can catalyze a polymerization reaction that can promote a stronger bond of the micro grafts 120 and / or tissue pieces 320 to the matrix 210, which may include proteins such as hyaluronic acid or collagen, for example.

[0058] In further exemplary embodiments of the present disclosure, the matrix material may be provided in a liquid or gel precursor form that can be mixed with the micro graft 120. The matrix material may then be solidified or gelled (either before or after introducing the mixture of the micro graft 120 and the matrix material into, for example, the recipient site). The solidification of the matrix material to form the matrix 210 can be achieved using various procedures based on the matrix material used. For example, a collagen gel (such as that shown in FIG. 16) may be provided in a liquid form at a low temperature and may be solidified by raising the temperature up to about 37°C. Other matrices 210 may be formed using polymers that can be solidified by a chemical cross-linking mechanism. Such a mechanism can be activated, catalyzed, and / or initiated, for example, by changes in temperature, light (similar to photo tissue bonding), pH, etc.

[0059] If the micro graft 120 is provided first in the matrix 210 in a liquid form, the orientation of the micro graft 120 within the matrix 210 can be set and / or maintained in several ways. For example, a material with a high liquid content can be applied to the upper end or top of the micro graft 120 (e.g., the epidermal surface of the micro graft 120 obtained from skin tissue), so the tip of the micro graft 120 tends to float relative to the surface of the liquid matrix material, and thus, the upper end of the micro graft 120 is disposed relative to the upper surface of the matrix 210. Alternatively, a metal coating may be applied to the upper part of the micro graft 120, and then a magnetic field may be applied in proximity to the graft material to align the micro graft 120 within the matrix material. Such exemplary alignment procedures can be performed before and / or during the solidification or gelation of the matrix material to produce a matrix 210 having micro grafts 120 with a specific orientation. Alternatively, in certain exemplary embodiments of the present disclosure, the micro graft 120 may be provided in the matrix 210 without requiring a specific preferred orientation.

[0060] In yet another exemplary embodiment of the present disclosure, as shown in FIG. 5A, an apparatus 500 is provided that can facilitate the collection of an exemplary micrograft 120 from a donor site 100 described herein. The exemplary apparatus 500 can include a hollow tube 510 that can be formed from a metal or another structurally rigid material. For example, the tube 510 can be formed using a stainless steel tube, a biopsy needle, or a similar structure. The tube 510 may be coated with a lubricant or a low friction material, such as Teflon®, to further facilitate the movement of the tube 510 through the donor site tissue 100.

[0061] The inner diameter of the tube 510 can be selected or structured to substantially correspond to the specific diameter of the micrograft 120 removed from the donor site described herein. According to one exemplary embodiment, the inner diameter of the tube 510 can be less than about 1 mm. For example, an 18 or 20 gauge biopsy needle (e.g., having inner diameters of 0.838 mm and 0.564 mm, respectively) may be used to form the tube 510. Biopsy tubes with larger gauges (and smaller inner diameters) may also be used. Based on the interaction with the tube, the width or diameter of the micrograft 120 collected may be slightly smaller than the inner diameter of the apparatus 500 used for collection.

[0062] The distal end of the tube 510 can be made to be pointed to form a plurality of tip portions. For example, as shown in FIG. 5A, two exemplary tip portions or extensions can be formed by polishing opposite sides of the tube 510 at an angle with respect to the long axis of the tube 510. In a further exemplary embodiment shown in FIG. 5B, an exemplary device 550 may be provided that includes a tube 510 having three tip portions or extensions 520 provided at the distal end. This exemplary configuration can be formed, for example, by polishing three tip portions of the tube 510 at an angle with respect to the long axis of the tube 510, where the three tip portions are spaced apart by approximately 120° around the tube 510. In still further exemplary embodiments, an apparatus for collecting micro-grafts including a tube having more than three tip portions or extensions 520 provided at the distal end may be provided (e.g., the tube 510 has 4, 5, 6, 7, or 8 tip portions 520).

[0063] The exemplary tip portions or extensions 510 can facilitate insertion of the devices 500, 550 into the tissue at the donor site 100. For example, the exemplary tip portions or extensions 520, which can be formed by polishing a portion of the distal end of the tube 510, may also have angled ends along their sides, thereby further facilitating insertion of the devices 500, 550 into the donor site tissue. For example, tip portions or extensions 520 that form a narrow angle at their tips can be inserted into the tissue using less force compared to tip portions 520 having a larger tip angle, although this force needs to be applied over a longer distance and / or time to achieve complete insertion of the tube 510 into the tissue than is used with a tip having a wider tip angle and thereby a shorter length of the angled tip region. The initial force on the tube 510 required to penetrate the tissue can be approximately proportional to the number of tip portions 520 if the angle of each tip portion or extension 520 is kept constant. Providing a greater number of tip portions, extensions 520 at the distal end of the tube can improve the mechanical stability of the tube 510 and / or the shape control of the tissue being cut, although it may require using a greater force to penetrate the tissue.

[0064] The exemplary device 500 may also include a collar or stop 540 provided on the outer surface of the tube 510. The exemplary stop 540 may be attached to the tube 510 at a specific distance from the end of the tip 520, or this distance may be adjustable over a range of lengths, for example, by moving the stop 540 along the axis of the tube 510.

[0065] FIG. 6A shows an exemplary device 500 after insertion into the tissue at the donor site 100, for example, until the stop 540 contacts the surface of the donor site 100. A portion of the tissue 600 may be present within the lower portion of the tube 510. The sides of this tissue portion 600 may be severed or separated from the surrounding tissue by the distal end and / or the tip or extension 520 of the tube 510 when the tube 510 penetrates the donor site tissue 100. Such tissue 600 may be maintained within the tube 510 and, for example, when the tube 510 is removed from the donor site 100 as shown in FIG. 6B, may be separated from the donor site 100 to form the micro-graft 120. The exemplary micro-graft 120 thus formed may include both epidermal tissue 130 and dermal tissue 140.

[0066] The exemplary micro-graft 120 may be removed from the device 500 by applying pressure, for example, through the opening 620 at the proximal end of the tube 510 as shown in FIG. 6C. Such pressure may be mechanical, hydraulic, pneumatic, etc. For example, the pressure may be applied, for example, by blowing air into the opening, by squeezing a flexible spherical portion attached to the opening, by opening a valve communicating from a high-pressure source such as a small pump. Alternatively, the exemplary micro-graft 120 may be collected by inserting the exemplary device 500 into a plurality of positions at the donor site 100. Each micro-graft 120 within the tube 510 may then push any one of the micro-grafts up the tube 510 towards the opening. Once the tube 510 is sufficiently filled with the collected tissue, each further insertion of the exemplary device 500 into the donor site 100 may facilitate pushing the uppermost micro-graft 120 within the tube 510 outwards from the proximal opening 620.

[0067] The exemplary device 500 can be inserted into the donor site tissue 100 to a depth corresponding to approximately the desired length of the harvested micrografts 120. Such a distance can be determined and / or controlled, for example, by appropriately positioning or adjusting the stop 540 on the exemplary device 500. For example, the exemplary device 500 can be configured or structured such that the tip or extension 520 extends to a position at or near the dermal / adipose layer junction 610, as shown in FIG. 6A. For example, the micrografts 120 can be removed from the donor site 100 by removing the device 500 from the donor site without rotating the tube 510 about its axis. In contrast, conventional biopsy needles and the like may require rotation about the long axis to facilitate removal of tissue samples from surrounding tissue. The tip or extension 520 provided on the exemplary device 500 can facilitate such removal of the micrografts 120 from the surrounding tissue at the donor site 100.

[0068] In certain exemplary embodiments of the present disclosure, some or all of the tissue at the donor site can be cooled, frozen, or partially frozen prior to collecting the micrografts 120. Such freezing can facilitate cutting, removal, manipulation, and / or survival of the micrografts 120. The donor site tissue 100 can be cooled or frozen using conventional cooling techniques such as applying a cryopspray or contacting the surface of the donor site 100 with a cooled object for an appropriate period of time. The exemplary device 500 may also be cooled prior to collecting the micrografts 120. Such cooling and / or freezing can increase their mechanical stability, for example, when the micrografts 120 are collected and / or placed in the matrix 210.

[0069] Exemplary micro-grafts 120 can be provided within matrix 210 using a variety of techniques. For example, individual micro-grafts 120 can be inserted into specific locations of matrix 210 using, for example, tweezers. As shown in FIG. 6B, an exemplary device 500 containing the micro-grafts 120 to be collected can also be inserted into the location of matrix 210, and pressure can be applied to proximal opening 620 to push the micro-grafts 120 into matrix 210. The exemplary device 500 may then be removed from matrix 210, and the procedure may be repeated to place multiple micro-grafts 120 within matrix 210. The proximal opening 620 may be covered while the device 500 is inserted into matrix 210 to prevent the micro-grafts 120 from being further pushed up within the device 500. For example, the upper portion of tube 510 may be filled with a fluid, such as water or saline, to provide a non-compressible volume that can prevent the micro-grafts 120 from being further pushed up within tube 510. Such a fluid may also facilitate removal of the micro-grafts 120 from the exemplary device 500 by applying pressure to the proximal opening 620.

[0070] In a further exemplary embodiment of the present disclosure, as shown in FIG. 7, an exemplary apparatus 700 may be provided. The apparatus 700 may include, for example, a plurality of tubes 510 that are attached to or mechanically coupled to a base 710. The tubes 510 may be provided in various configurations, such as in a linear arrangement or in any one of various two-dimensional patterns along the base 710. The number of tubes 510 provided in the exemplary apparatus 700 may be, for example, at least six tubes 510, more than about ten tubes 510, or more than about twenty tubes 510. For example, the apparatus 700 may include about three tubes 510, a linear arrangement of at least six tubes 510, or about twelve tubes 510, such as a rectangular 3×4 arrangement (array) or a 2×6 arrangement (array). Larger arrangements (arrays) of the tubes 510 may also be provided in further exemplary embodiments of the present disclosure. The spacing of the tubes 510 may be somewhat non-uniform or varied in order to avoid forming a pattern that is recognizable at the donor and / or recipient sites. A large number of tubes 510 may be suitable for more rapidly collecting and / or implanting many micro-grafts 120. However, a large number of tubes 510 may also be able to withstand the large forces used to simultaneously insert the tubes 510 into tissue or a matrix, which may be undesirable if the forces become very large. Also, the mechanical complexity of the apparatus 700 may be increased by a large number of tubes 510.

[0071] A conduit 720 may be provided in communication with a proximal opening 620 of the tube 510. The conduit 720 may also be provided in communication with, for example, a pressure source 730. For example, the pressure source 730 may include a pump or a deformable sphere, etc. The pressure source 730 may include, for example, a flexible membrane in communication with the conduit 720, and when the membrane is deformed, a high pressure may be applied into the conduit 720. Such a configuration may facilitate applying pressure to the proximal opening 620 to remove and / or insert the micro-grafts 120 that may be collected within the tubes 510 described herein.

[0072] A vibration configuration 740 may be provided to the device 700 as needed. To facilitate the insertion of the tube 510 into the tissue or matrix material for collecting or placing the micro grafts 120, the vibration configuration 740 may be mechanically coupled to the base 710 and / or the tube 510. The vibration configuration 740 may have an amplitude of vibration in the range of about 50 - 500 μm, or about 100 - 200 μm. The frequency of the induced vibration may be about 10 Hz to about 10 kHz, or about 500 Hz to about 2 kHz, or even about 1 kHz. Specific vibration parameters may be selected based on, for example, the size, average spacing, and material of the tube 510 in the exemplary device 700, the number of tubes 510, and / or the tissue being processed. The vibration configuration 740 may include a circuit configured to adjust the amplitude and / or frequency of the vibration.

[0073] The exemplary device 700 may be used to simultaneously obtain a plurality of micro grafts 120 in one or more of the tubes 510. An exemplary procedure for obtaining and removing such micro grafts 120 using the exemplary device 700 may be similar to the procedures described herein for obtaining a single micro graft 120 using the exemplary device 500 shown in FIGS. 6A - 6C.

[0074] Vibration can also help cut the tissue proximate to the distal end of the tube 510 after the tube 510 is fully inserted into the donor site 100. This can facilitate the separation and / or extraction of tissue portions from the donor site 100 into the tube 510. These tissue portions can also be retained by friction within the tube 510 when the tube 510 is withdrawn from the donor site 100.

[0075] In further exemplary embodiments of the present disclosure, the tissue at the donor site can be pre-cooled prior to insertion of the tube 510 using convection or conduction techniques such as, for example, applying a cooling spray or contacting the tissue surface with a cooled object. When the tube 510 is inserted into the donor site tissue 100, cooling of the donor site 100 can reduce pain sensation and also make the tissue 100 more rigid, facilitating more accurate cutting of tissue portions (e.g., micro-grafts 120) by the tube 510.

[0076] The position and spacing of the tubes 510 in the exemplary device 700 can be determined based on, for example, the characteristics of the resulting micro-grafts 120, the damage pattern to the donor site 100, and / or other factors described hereinabove. The number of tubes 510 provided in the exemplary device 700 can be selected based on various factors. For example, a greater number of tubes 510 may be desired to enable collection of more micro-grafts 120 from the donor site 100 simultaneously. Such an exemplary configuration can facilitate a more effective collection process. A smaller number of tubes 510 can be easier to insert simultaneously into the donor site tissue 100. Additionally, an exemplary device 500 having a very large number of tubes 510 can be difficult to manufacture and / or maintain.

[0077] The collected tissue portions can be deposited directly from the tubes 510 into the biocompatible matrix material 210. The tubes 510 and the tissue portions contained therein can be cooled prior to removal of the tissue portions. This hardens the tissue portions within the tubes 510, making them easier to manipulate and place.

[0078] In further exemplary embodiments of the present disclosure, an apparatus may be provided that includes a plurality of substantially parallel blades. A particular one end of adjacent blades may be connected or closed, for example, to provide a narrow rectangular opening between adjacent blades. Such an exemplary apparatus may be used, for example, to form tissue piece 320 as shown in FIG. 3C. The spacing, length, and other characteristics of such an exemplary apparatus may be selected based on factors similar to those described herein for exemplary apparatuses 500, 700.

[0079] In further exemplary embodiments of the present disclosure, the exemplary methods and apparatuses described herein may be applied to tissues other than skin tissue, such as visceral organs such as the liver or heart. Thus, grafts may be formed for various tissues while creating transplant tissue suitable for placement at the recipient site with little damage to the donor site and promoting their rapid healing.

[0080] An image of the distal end of an exemplary apparatus including two tip portions is shown in FIG. 8A. Such an exemplary apparatus is similar to exemplary apparatus 500 shown in FIG. 5A, for example. An image of such an exemplary apparatus further rotated is shown in FIG. 8B. Such an exemplary apparatus was formed using a tube having an outer diameter of about 1 mm and an inner diameter of about 0.5 mm. The tip portion or extension was formed by polishing two opposing sides of the distal end of the tube at an appropriate angle with respect to the axis of the tube. The angles used in the exemplary apparatuses shown in FIGS. 8A and 8B were about 30°, although other angles may be used. The angled ends of the tube can be seen along the sides of the tip portion or extension. The shape of these tip portions can facilitate the insertion of the apparatus into the tissue at the donor site and / or the separation of a portion of the micro-transplant tissue from the donor site, as described in more detail herein. For example, such a micro-graft can be separated and removed from the donor site by inserting it and pulling the apparatus out of the donor site without rotating the tube along the axis of the tube.

[0081] The shape (with holes open) of the distal end of an exemplary tube 510 including two tips 520 can be characterized by an angle α that represents the angle between each of the opposing sides of the tube 510 that form the tip or extension 520 and the longitudinal axis of the tube 510. Each of the angled sides at the distal end of the tube 510 can be shaved or cut at this angle α with respect to the axis of the tube 510, for example, to form an inclined structure at the distal end of the tube 510. As viewed from the side, the angle formed at the tip or extension 520 can thus be represented by an angle 2α, as shown in FIG. 8C. For example, the exemplary tip angle of about 30° shown in FIGS. 8A and 8B corresponds to an angle α of about 15°.

[0082] Additional inclined surfaces may be provided, if desired, in a direction perpendicular to the main inclination characterized by the angle α shown in FIG. 8C. This second inclination angle can be characterized by an angle β that represents the angle in each of the opposing sides of the tube 510 that can be shaved or cut with respect to the longitudinal axis of the tube 510, as shown in FIG. 8D. This second bevel angle may be provided to reduce the size or width of the acute angle of the tip 520 formed at the end of the tube 510.

[0083] FIGS. 8E, 8F, and 8G show exemplary inclined ends of the tube 510 where the main bevel angle α can be about 6°. For example, in FIG. 8E there is no second bevel angle formed, and the tip or extension 520 has a flat cut end having a length equal to the wall thickness of the tube 510. FIGS. 8F and 8G include a second bevel angle where the angle β corresponding to the second bevel angle is also 6°. The second bevel angle is relatively shallow in the exemplary tip region shown in FIG. 8F that provides a short flat cut end at the tip of the tip or extension 520. The second bevel angle is deeper in the exemplary tip region shown in FIG. 8G, and the tip of the tip or extension 520 forms a sharp tip that can more readily facilitate penetration into tissue.

[0084] Figures 8H, 8I, and 8J show exemplary inclined ends of tube 510 that are similar to those shown in Figures 8E - 8G, but the main bevel angle α is 12°. There is no second bevel angle in Figure 8H, the narrow second bevel angle shown in Figure 8I has an angle β of about 6°, and the deep second bevel angle shown in Figure 8J again has an angle β of, for example, about 6°.

[0085] The various shapes and tip portions 520 shown above and in Figures 8A - 8F can be used in any of the exemplary embodiments of the present disclosure with respect to, for example, the various devices and methods described herein. For example, the hollow tube 510 is provided with a bevel angle α of less than about 15°, for example about 12°. Such a sharp tip angle can provide a sharp tip of the tip portion or extension 520 that can easily penetrate biological tissue or matrix material. A narrower tip angle α, such as about 6°, as shown in Figures 8E - 8G, may be suitable for collecting and / or implanting micro - grafts within dense or firm tissue or matrix material, and the narrow tip of the tip portion or extension 520 can be configured to more easily cut through tissue or matrix material when the tube 510 is inserted. A second bevel angle having an angle β, such as the exemplary tips of the tip portions or extensions 520 shown in Figures 8F, 8G, 8I, and 8J, is smaller and can further facilitate the insertion of the distal end of the tube 510 into various materials by providing a smaller, sharper tip of the tip portion or extension 520. However, a sharper and / or narrower tip of the tip portion or extension 520, for example, having a small tip angle α and / or a second bevel angle having an angle β, may also be more prone to wear, bending, or other deformations when the tube 510 is repeatedly inserted into tissue or matrix. Thus, the tip shape selected for a particular application can be chosen based on the type of material or tissue, the device used, and the desired lifespan of the tube 510.

[0086] FIG. 9 shows an exemplary image of a plurality of micro-grafts obtained from a donor site of in vitro skin tissue using the exemplary device shown in FIGS. 8A-8B. The micro-grafts are elongated and substantially of the same shape, although the details of the shape may be somewhat irregular. The upper portions of these micro-grafts contain epidermal tissue, and the lower portions of these micro-grafts contain dermal tissue removed from the donor site. The width of these micro-grafts is slightly smaller than the inner diameter of the tube 510 shown in FIGS. 8A-8B used to collect them.

[0087] The micro-grafts shown in FIG. 9 were removed from the device by inserting the exemplary device into the donor site a plurality of times until the tube was filled with the collected tissue. Then, by successively inserting the device into the donor site tissue, the uppermost micro-grafts were pushed out from the proximal end of the tube and collected individually for analysis. Such micro-grafts may also be removed by applying pressure to the proximal end of the tube containing the micro-grafts so as to release it from the distal end of the tube as described herein.

[0088] In still further exemplary embodiments of the present disclosure, as shown in FIG. 10A, an apparatus 1000 may be provided that facilitates the collection of exemplary micro-grafts 120 from a donor site 100 and, if desired, disposes them in a biocompatible matrix 210 as described hereinafter. The exemplary apparatus 1000 may include a hollow tube 1010 that may be similar to the hollow tube 510 shown in FIGS. 5A and 5B and may include a plurality of tip portions 1020 at the distal end as described herein.

[0089] In one exemplary embodiment of the present disclosure, the tube 1010 may be provided with two tip portions 1020, which may be formed by polishing the opposite side of the distal end of the hollow tube 1010 at an acute angle, for example, about 6° with respect to the longitudinal axis of the tube 1010. Such a sharp tip angle, for example, less than about 12°, may be particularly effective for penetrating and cutting biological tissue to remove small biopsy specimens 120. Such a tube provided with two tip portions 1020 can use approximately twice the force associated with a single tip needle of the same diameter for penetrating tissue or another material.

[0090] For example, FIG. 19A shows an exemplary sequence of insertion and withdrawal or pulling of a collection needle device from a tissue layer. The exemplary needle device may be similar to the hollow pointed tube 510 shown in FIG. 5A or may be similar to the exemplary pointed tube 1010 shown in FIG. 10A. A plot of the position of the needle relative to the tissue as a function of time is shown in FIG. 19B together with a plot of the corresponding force applied to the exemplary needle (e.g., along the longitudinal axis) to perform the insertion and withdrawal sequence. The needle used to obtain the data shown in FIG. 19B was formed from a 25-gauge tube having a tip bevel angle α of 6° and a side bevel angle β of 6°, similar to the exemplary needle shapes shown in FIGS. 8C, 8D, and 8F.

[0091] As shown in the graph of FIG. 19B, the needle can be rapidly inserted through the tissue layer in about 6 seconds, held there for about 1 second, and then stably pulled to a position outside the tissue layer in about 0.8 seconds. After insertion into the tissue, there may be a perceived delay in the force required to maintain the needle at a particular depth and the remaining force in the needle after bringing the needle to a position outside the tissue layer. During insertion and pulling of a needle formed using a 25-gauge tube, the maximum and minimum forces observed on the needle were about -0.2 N and 0.2 N, respectively. The delay during moving the needle into and out of the tissue and the forces obtained on the needle result, at least in part, from tissue deformation when the needle is inserted and pulled and / or tissue adhesion to the needle when the needle moves relative to the tissue. The small forces seen at the start and end of the sequence plotted in FIG. 19B appear to be related to a vertical load that detects some frictional and / or adhered components.

[0092] In one exemplary embodiment of the present disclosure, the tube 1010 is formed using a 25-gauge thin-walled needle having an exemplary outer diameter of about 0.51 mm and an exemplary inner diameter of about 0.31 mm. This exemplary needle size can be used to collect the micro-grafts 120 having a width or diameter of about 0.2 mm. The tube 1010 can be formed from any sufficiently strong material that is preferably biocompatible and inert to living tissue, such as 304 stainless steel, surgical stainless steel, etc. Further finishing processes such as electropolishing to increase material strength may be applied to the tube 1010. Such finishing processes can increase the effectiveness of cutting and / or improve the effective service life of the needle 1010.

[0093] The tube 1010 may be slidably attached to the substrate 1030. As shown in FIG. 10A, the tube 1010 moves through a hole provided in the substrate 1030. The position of the tube 1010 relative to the substrate 1030 can be controlled, for example, by arranging a configuration that can controllably move the tube 1010 relative to the substrate 1030 substantially along the longitudinal axis of the tube 1010. In this way, the distance that the distal end of the tube 1010 projects through the lower surface of the substrate 1030 can be controllably changed.

[0094] The exemplary device 1000 further includes a pin 1040 within a central cavity or opening of the tube 1010. The diameter of the pin 1040 can be substantially the same as or slightly smaller than the inner diameter of the tube 1010, such that the pin 1040 can move along the axis of the tube 1010 while filling or blocking most or all of the lumen of the tube 1010. The pin 1040 can be formed from a low-friction material or can be coated with a low-friction material such as Teflon (registered trademark) to facilitate movement of the pin 1040 within the tube 1010 and / or to prevent accumulation or adhesion of biological material to the pin 1040. The distal end of the pin 1040 can be substantially flat to facilitate movement of the micro-grafts 120 within the tube 1010 when the pin 1040 moves.

[0095] In an exemplary embodiment of the present disclosure, the tube 1010 can be formed using a 25-gauge thin-walled needle having an exemplary outer diameter of about 0.51 mm and an inner diameter of about 0.31 mm. This exemplary needle size can be used to collect micro-grafts 120 having a width or diameter of about 0.2 mm. The pin 1040 that can be used with the tube 1010 of this size can have an outer diameter of about 0.24 mm. The difference between the inner diameter of the tube 1010 and the diameter of the pin 1040 can facilitate movement of the pin 1040 within the tube 1010, while the pin is wide enough to push the micro-grafts 120 out of the interior of the tube 1010. The tube 1010 and / or the pin 1040 can preferably be formed from any sufficiently strong material that is biocompatible and inert to biological tissue, such as 304 stainless steel, surgical stainless steel, etc.

[0096] For example, a further positioning configuration can be provided for the pin 1040 that enables controllable movement of the pin 1040 relative to the tube 1010 substantially along the longitudinal axis of the tube 1010. In this way, the position of the distal end of the tube 1010 relative to the distal end of the pin 1040 can be controllably changed. For example, the positions of the distal ends of both the tube 1010 and the pin 1040 relative to the lower surface of the substrate 1030 can preferably be controllably and independently selected and changed.

[0097] An exemplary diagram of the tube 1010 and pin 1040 described in this specification is shown in FIG. 10B, which shows the pin 1040 disposed with respect to the tube 1010, and their distal ends are substantially aligned. Coatings or surface treatments may be provided on portions of the pin 1040 and / or tube 1010, as needed, to reduce friction between them and / or between the components and living tissue. Exemplary coatings that may be used include plastics or polymers, such as nylon or polyethylene, polished metal alloys, and the like.

[0098] A schematic diagram of an exemplary device 1100 that may be used to facilitate collection of exemplary micro-grafts 120 from the donor site 100 and, if needed, place them in the biocompatible matrix 210 is shown in FIG. 11. The device 1100 may include a tube 1010, a pin 1040, and a substrate 1030, as shown in FIG. 10A. A plate 1110 or other support structure may be fixed to the substrate 1030, or may be provided on the substrate 1030 as a single unit component, as needed.

[0099] The first actuator 1120 may be attached to the plate 1110 and may be mechanically coupled to the tube 1010 using, for example, a first connection arm 1125. Similarly, the second actuator 1130 may be attached to the plate 1110 and may be mechanically coupled to the pin 1040 using, for example, a second connection arm 1135. The first actuator 1120 and the second actuator 1130 may be conventional linear actuators or the like. Such actuators 1120, 1130 may be provided with appropriate control devices, and the positions of the connection arms 1125, 1135 with respect to the plate 1110 (and substrate 1130) are controllably arranged and may be independently changed. For example, the linear range of movement of the actuators 1120, 1130 may be less than about 2 cm for many typical uses of the device 1100 for collecting micro-grafts of skin tissue and placing them in a matrix, as shown hereinafter in this specification.

[0100] In the exemplary schematic structure of the device 1100 shown in FIG. 11, the movement or placement range of the actuator 1120 can be selected such that the distal end of the tube 1010 can rise above the lower surface of the substrate 1030 and can descend so that the distal end projects up to a maximum distance of about 1 to 2 cm below the lower surface of the substrate 1030. Similarly, the placement range of the actuator 1130 can be selected such that the distal end of the pin 1040 can rise above the distal end of the tube 1010 by a distance of about 1 to 2 cm and the distal end of the pin 1040 can descend so as to be substantially aligned with the distal end of the tube 1010 (e.g., aligned with the tip of the tip portion 1020). The movement ranges of the actuators 1120 and 1130 may also be greater than these exemplary distances, for example, when the donor tissue to be collected is relatively thick. Alternatively, these movement ranges may be somewhat smaller when the micro grafts are collected from a thin layer and placed in a thin matrix as needed.

[0101] Exemplary sequences for collecting the micro grafts 120 from the donor site 100 and placing them in the matrix 210 according to embodiments of the present disclosure are shown in FIGS. 12A-12E. As in the initial state, the surface of the donor site can be cleaned, sterilized, shaved, and / or otherwise prepared, and as shown in FIG. 12A, a portion or sheet of the matrix material 210 can be placed thereon. The distal ends of the tube 1010 and the pin 1040 can be aligned by respective actuators (not shown) and placed close to the upper surface of the matrix 210. The tube 1010 and the pin 1040 can be moved so that their distal ends can penetrate or pass through the thickness of the matrix 210 and are placed close to the surface of the donor site 100 as shown in FIG. 12B. The tube 1010 and the pin 1040 can displace a small amount of matrix material such as viscous, flexible, elastic when they pass through the matrix 210.

[0102] Next, the tube 1010 can be moved downward so as to penetrate into the tissue of the donor site 100. However, since the position of the pin 1040 can be held substantially fixed relative to the donor site 100, its distal end remains disposed proximal to the surface of the donor site 100 as shown in FIG. 12C. When the distal end of the tube 1010 penetrates the donor site, a portion of the tissue can be cut from the surrounding tissue at the donor site. Thus, a portion of the tissue from the donor site 100 can be located within the distal end of the lumen of the tube 1010.

[0103] The depth of penetration of the tube 1010 into the donor site 100 can be controlled by the tube actuator 1120 as shown in FIG. 11. The tube 1010 can be moved so that its distal end is positioned at a specific depth within the donor site tissue. For example, if the donor site is skin tissue, the distal end of the tube 1010 can be extended to penetrate slightly into the underlying adipose tissue, for example, as shown in FIG. 12C, so as to be close to the lower end of the dermis layer.

[0104] Next, as shown in FIG. 12D, the tube 1010 and the pin 1040 can be lifted or withdrawn simultaneously so that they maintain their relative positions substantially until the distal end of the tube 1010 is close to or slightly above the surface of the donor site 100. A portion of the tissue cut from the donor site 100 by the tube 1010, which can be used as the micro-graft 120, can also be held within the distal end of the tube 1010. Thus, when the tube 1010 is withdrawn from the donor site 100, the tube 1010 is lifted or removed from the donor site 100. When the tissue to be collected is skin tissue, the removal of the micro-graft 120 from the donor site can be facilitated if the tube 1010 first penetrates at least the upper surface of the subcutaneous fat layer. The lower end of the micro-graft 120 can be more easily separated or peeled off from the adjacent adipose tissue than from the dermal tissue.

[0105] As shown in FIG. 12D, the tube 1010 and the pin 1040 can be withdrawn together from the donor site 100, and the micro-graft 120 can be placed within the layer of the matrix material 210. For example, the tube 1010 and the pin 1040 can be lifted substantially simultaneously such that the distal end of the tube 1010 is close to the lower surface of the matrix material 210 or disposed within the matrix material 210. The tube 1010 may then be further retracted from the matrix 210 while holding the position of the pin 1040 substantially fixed relative to the matrix 210, as shown in FIG. 12E. This exemplary procedure facilitates the placement of the micro-graft 120 within the material of the matrix 210 using the pin 1040 when the tube 1010 is withdrawn from around the micro-graft 120. Thus, the micro-graft 120 can be placed and held within the matrix 210, whereas the tube 1010 and the pin 1040 are completely removed from the donor site 100 and the matrix 210.

[0106] The exemplary apparatus shown in FIG. 11 and the order of collecting the tissues shown in FIGS. 12A - 12D and described herein provide several advantages for collecting the micro-grafts 120 from the donor site and placing them in the matrix 210 to facilitate their use in transplantation or autotransplantation procedures. For example, the micro-grafts 120 can be placed within the matrix 120 without being exposed to the outside air (or another intermediate environment) to reduce the chance of contamination, biological stress, etc. The various penetration depths can be selected or adjusted based on the desired depth of the micro-grafts to be collected, the thickness of the matrix material 210 used, etc. The micro-grafts 120, which can be difficult to handle due to their small size and / or soft tissue consistency, can be placed in the matrix 210 in substantially the same orientation as when they were at the donor site.

[0107] When present, the substrate 1030 shown in FIG. 11 can provide mechanical stability to the surfaces of the matrix 210 and the donor site 100 when the device 1100 is disposed on a layer of the matrix material 210. When present, by disposing the substrate 1030 on the matrix 210 and / or the donor site 100, movement and / or deformation of the tissues of the matrix 210 and the donor site 100 can be prevented while the micro-grafts 120 are being harvested.

[0108] In further exemplary embodiments of the present disclosure, the substrate 1310 can include a plurality of strips or slats 1315, as shown in FIG. 13B, having elongated openings therebetween. The black dots shown in FIG. 13B are micro-grafts harvested from the black donor site and are transplanted to the recipient site shown in this figure. Both the donor and recipient sites were porcine skin tissue that resulted in autografts. The substrate 1030 can be disposed on the donor tissue 100 as described above to stabilize the tissue 100, whereas the collection tubes 510, 1010 can be inserted into the tissue 100 between the slats 1315. The substrate 1310 can also be used, for example, to facilitate the placement of the tubes 510, 1010 relative to the donor tissue for repeated insertion and withdrawal of the tubes 510, 1010 from a region of the donor tissue 100. The substrate 1030 can also be provided in other geometric shapes and configurations of openings, such as a plurality of openings having an oval, triangular, square, or other shape, or combinations thereof.

[0109] In a further exemplary embodiment of the present disclosure, as shown in FIG. 13, the substrate 1030 can be disposed directly on the surface of the donor site 100 to provide mechanical stability and reduce movement of the tissue at the donor site 100. The substrate 1030 can be provided with one or more through-holes 1310. For example, as described above and shown in FIGS. 12A-12E, one or more tubes 1010 can be configured to pass through the holes 1310 to cut and extract the micro-grafts 120 from the donor site. As shown in FIG. 13A, the matrix 210 can be provided on top of the substrate 1030. Optionally, a bandage material 1320 or the like can be disposed on the upper surface of the matrix 210 to further stabilize the matrix 210 and / or facilitate handling of the matrix 210. When the substrate 1030 is provided between the matrix 210 and the donor site, as shown in FIG. 13A, the travel distance or height of the tubes 1010 and pins 1030 shown in the order of FIGS. 12A-E can be appropriately controlled such that the micro-grafts 120 are collected from the donor site 100 and placed on the matrix 210 on the substrate 1030.

[0110] In a further exemplary embodiment of the present disclosure, as shown in FIG. 14, two substrates 1400, 1410 can be used to provide further stabilization. The lower substrate 1410 having one or more through-holes 1420 can be provided on the surface of the donor site 100, similar to the substrate 1030 shown in FIG. 13A. The matrix 210 can be provided on the lower substrate 1410. The upper substrate 1400 can be provided on the matrix 210 or, if present, on the dressing material 1320. Such an upper substrate 1400 can be provided with one or more through-holes 1425 that can correspond to and / or be aligned with the holes 1420 provided in the lower substrate 1410. These aligned holes 1420, 1425 can facilitate the movement of the tube 1010 into and out of the matrix 210 and the donor site tissue 100 while providing mechanical stability to both the donor site 100 and the matrix 210. Such aligned holes 1420, 1425 in the lower substrate 1410 and the upper substrate 1400 can also provide further mechanical stability and an improved arrangement of one or more tubes 1010 when the tube 1010 is vertically moved through the holes 1420, 1425.

[0111] In yet a further exemplary embodiment of the present disclosure, as shown in FIG. 14, a barrier 1430, such as a side wall, etc., can be provided between the lower substrate 1410 and the upper substrate 1400 in proximity to the perimeter of one or both of the substrates. The lower substrate 1410, the upper substrate 1400, and the barrier 1430 can together form an enclosure around the matrix 210. This exemplary configuration can facilitate the containment of the matrix 210, for example, when the matrix 210 is formed from a viscous or easily deformable material. Thus, the exemplary embodiment of the present disclosure shown in FIG. 14 can facilitate the placement of the micro-grafts 120 into the matrix 210 that may not have mechanical rigidity or stability. The exemplary embodiment shown in FIG. 14 may be used in conjunction with various other embodiments of the present disclosure described herein.

[0112] The exemplary device 1100 can be used to collect a plurality of micro-grafts 120 from the donor site 100 and, if necessary, place them in the matrix 210. For example, the substrate 1030 is provided with a plurality of spaced-apart holes therethrough. The plate 1110 shown in FIG. 11, or another part of the device 1100 that supports the tube 1010 and the pin 1040, can be configured or structured to move on the substrate 1030, so that the tube 1010 and the pin 1040 can be placed in a plurality of holes in the substrate 1030. Such movement can be, for example, one-dimensional (e.g., linear) or two-dimensional over a particular surface area of the substrate 1030. In this exemplary method, while the substrate 1030 and the exemplary device 1100 are maintained in a single position relative to the donor site 100, a plurality of micro-grafts 120 can be collected from the donor site 100 and, for example, placed in a plurality of positions of the matrix 210 proximate to the plurality of holes in the substrate 1030.

[0113] In a further exemplary embodiment of the present disclosure, the device 1100 shown in FIG. 11 can be provided with a plurality of tubes 1010 and pins 1040. All of the tubes 1010 can be movable together using a single first actuator 1120, or a particular one of the tubes 1010 can be movable simultaneously and / or continuously using a plurality of first actuators 1120 and appropriate first connection arms 1125. Similarly, the plurality of pins 1040 can be movable together using a single second actuator 1130, or a particular one of the pins 1040 can be movable simultaneously and / or continuously using a plurality of second actuators 1130 and appropriate second connection arms 1135. Typically, each movement of the tubes 1010 can preferably cooperate with its associated pin 1040 (i.e., the pin 1040 provided in the lumen of a particular tube 1010). In this exemplary method, each combination of the tubes 1010 and their associated pins 1040 can be controlled to perform the exemplary collection and implantation sequence shown in FIGS. 12A - 12E.

[0114] A matrix containing one or more micro-grafts 120 described herein can be used as a graft material that can be placed over a recipient site of damaged tissue that is to be appropriately repaired. Typically, this graft material includes a plurality of micro-grafts 120 provided in a matrix 210. The spacing of the micro-grafts 120 within the matrix 210 can be selected to facilitate the growth of the micro-grafts within and through the matrix 120, and ultimately provide sufficient coverage and / or repair of the damaged area. The micro-grafts 120 can be arranged within the matrix 210 in a uniform pattern, randomly, or in any other desired spatial configuration. In certain exemplary embodiments of the present disclosure, the density or spacing of the micro-grafts 120 being transplanted can be varied in different regions of the matrix 210. For example, a higher density and / or a narrower spacing of the micro-grafts 120 can be provided closer to the ends of the matrix 210 to improve the peripheral integration and / or vascular reconstruction of the grafts. A sterile dressing, etc., can be placed over the graft material after the graft material has been placed in the damaged area of the tissue. Such a dressing can be attached to the graft material to facilitate handling and placement of the graft material at the recipient site.

[0115] In further exemplary embodiments of the present disclosure, the collected micro-grafts 120 can be introduced or transplanted directly, for example, substantially into the entire tissue at the recipient site. For example, the micro-grafts 120 can be collected from a donor site 100 that may contain melanocytes and inserted directly into the tissue at the recipient site that lacks sufficient melanocytes. Such an exemplary procedure can be used to repigment skin tissue, for example, to treat vitiligo or similar conditions. The tissue at the recipient site 100 can also be frozen or partially frozen prior to insertion of the micro-grafts 120, as described herein.

[0116] Exemplary device 1500 for implanting micro-grafts 120 into recipient site 1510 is shown in FIGS. 15A - 15E. Device 1500 can include a hollow tube 1010 that can include a plurality of tip portions 1020 at its distal end, similar to the device shown in FIG. 10A, and a pin 1040 provided in the central cavity or opening of tube 1010. Exemplary device 1500 can further include a hollow, perforated needle 1520 that can be provided around tube 1010, as shown in FIG. 15, and tube 1010 can advance and / or retract within the perforated needle 1520. The perforated needle 1520 can include a single tip portion 1525 configured to pierce and penetrate biological tissue. The perforated needle 1520 may be manually controlled or may be controlled using an actuator similar to actuators 1120, 1130 in exemplary device 1100 shown, for example, in FIG. 11.

[0117] In an exemplary method of the present disclosure, tube 1010 and pin 1040 can be used to collect tissue micro-grafts 120 using, for example, the exemplary collection sequence shown in FIGS. 12A - D. When tube 1010 and pin 1040 are completely withdrawn from donor site 100, micro-grafts 120 can be retained in tube 1010. Since tube 1010 and pin 1040 can be positioned within the perforated tube 1520, as shown in FIG. 15A, the distal end of tube 1010 is within the perforated needle 1520.

[0118] Next, the perforated needle 1520 can enter into the recipient site 1510, and the tip 1525 of the perforated needle 1520 separates a portion of the tissue at the recipient site 1510, as shown in FIG. 15B. Together with the micro-graft 120, the tube 1010 and the pin 1040 can advance forward together with the perforated needle 1520, so that the distal end of the tube 1010 containing the micro-graft 120 is positioned beneath or proximate to the surface of the recipient site 1510, as shown in FIG. 15B. Since the perforated needle 1520 can be withdrawn from the recipient site while substantially fixing the tube 1010 with respect to the recipient site 1510 and holding it, the distal end of the tube 1010 containing the micro-graft 120 is positioned within the separated tissue of the recipient site 1510, as shown in FIG. 15C.

[0119] Next, the tube 1010 can be withdrawn from the recipient site 1510 while substantially fixing the tube 1010 with respect to the recipient site 1510 and holding the pin 1040, so that when the tube 1010 is withdrawn, as shown in FIG. 15D, the micro-graft 120 remains within the separated tissue of the recipient site 1510. When the device 1500 is removed from the recipient site 1510, as shown in FIG. 15E, the micro-graft 120 can remain within the recipient site 1510 in a known orientation.

[0120] Such direct injection can be used, for example, for the normalization of tissue for treating vitiligo by transplanting micro-grafts 120 containing melanin directly into the depigmented recipient site 220. Exemplary micro-grafts 120 can also be collected from a healthy donor site 100 using the exemplary methods and apparatuses shown in FIGS. 15A - 5E and placed directly into a recipient site 1510 containing scar tissue to promote the growth of healthy tissue in the scar. In a further exemplary embodiment of the present disclosure, tissue portions can be removed from the recipient site 1510 before placing the micro-grafts 120 into holes formed in the recipient site 1510 by removing these tissue portions. The holes can be approximately the same size as the micro-grafts 120 inserted therein or slightly larger. The holes can be formed in the recipient site, for example, by removing or excising tissue using an ablation laser or the like, for example, using one or more tubes 510 described herein.

[0121] The exemplary methods and apparatuses shown in FIGS. 15A - 15D can be used for various treatments, including transplantation between and within various tissues other than the skin, such as muscle tissue, organ tissue, etc. "Hybrid" or xenograft grafts containing different tissues can also be generated using the exemplary methods and apparatuses described herein. For example, micro-grafts 120 from a donor site having a first tissue type can be placed in a second tissue type at the donor site. Such exemplary transplantation procedures can be used for many different applications. For example, micro-grafts 120 from an endocrine organ can be placed in a donor site 1510 including the skin. For example, micro-grafts of pancreatic tissue can be placed in skin tissue to provide insulin secretion. As another example, smooth muscle tissue can be introduced into the digestive tract. Micro-grafts 120 obtained from other functional tissues can be placed in donor sites having different characteristics.

[0122] In certain exemplary embodiments of the present disclosure, the perforated needle 1520 can be provided around at least a portion of the tubes 510, 1010 without having a sharp distal tip. The distal end of the perforated needle 1520 may be flat, for example, the outside of the distal end may be widened or flanged as needed. Such a perforated needle 1520 can function as a guide and / or support for the tubes 510, 1010, and can reduce or prevent bending, distortion, breakage, etc. of the tubes 510, 1010 when the tubes are inserted into and withdrawn from the donor tissue site. It can also facilitate control of the insertion depth of the tube 1010 and / or the pin 1040 (if present) in the tissue.

[0123] In further exemplary embodiments of the present disclosure, for example, a conduit can be provided that communicates with the lumens of the tubes 510, 1010 described herein and is connected to the proximal ends of the tubes 510, 1010. Such a conduit can be configured similarly to the conduit shown in FIG. 7. The conduit can also be provided in communication with a low and / or high pressure source, such as a vacuum configuration and / or a pressurized gas or liquid source. For example, the diameter of the pin 1040 may be sized slightly smaller than the diameter of the lumen of the tube 1010. Such a configuration can facilitate the passage of a fluid, such as a gas, through the lumen of the tube 1010 around the pin 1040, and can further propagate a pressure differential through the lumen of the tube 1010. The controllable application of low pressure from the conduit to the lumen of the tube 1010 can facilitate the separation of the micro graft 120 from the surrounding tissue. Similarly, the application of high pressure from the conduit to the lumen of the tube 1010 can facilitate the removal or expulsion of the micro graft from the tube 1010 after collection of the micro graft.

[0124] In further exemplary embodiments of the present disclosure, for example, fluid can be provided within tube 510 shown in FIGS. 5A and 5B or tube 1010 shown in FIG. 10A such that a portion of the fluid is present between tube 1010 and pin 1040. The fluid can reduce friction between pin 1040 and tube 1010. The fluid can also reduce or prevent the collection or accumulation of biological tissue proximate to the distal end of tube 1010 when tube 1010 is used to collect a plurality of tissue micro-grafts 120. The fluid can also facilitate the release of micro-grafts 110 from the tube by providing the fluid at low or high pressure during appropriate steps of, for example, the exemplary micro-graft manipulation sequences shown in FIGS. 6A - 6C, 12A - 12E, and / or 15A - 15D for the retention of micro-grafts 120 within tube 1010. For example, the fluid can improve the accuracy of placement of micro-grafts 120 at the donor site tissue 1510 or within matrix 210 described herein. Such accuracy in placing the collected micro-grafts 120 can reduce or prevent cyst formation or other undesirable outcomes, for example, if micro-grafts 120 can grow after collection.

[0125] Such fluid can be provided through a conduit, for example, similar to conduit 720 shown in FIG. 7, that is provided in communication with the proximal end of tube 100. Alternatively, the fluid can be provided through an opening formed in the side of tube 1010. When an open needle 1520 as provided in the exemplary device 1500 shown in FIG. 15 is used, the fluid can also or alternatively be provided between open needle 1520 and tube 1010.

[0126] Exemplary fluids that can be used are biocompatible and inert with respect to living tissue and the like. Such fluids preferably cause no side effects even when in contact with tissues 100, 1510. For example, the fluid may include physiological saline, glycerol, and the like. It may be buffered and may contain one or more additional components, such as anticoagulants, antibacterial agents, coagulants, and the like. One or more growth factors may also be added to this fluid to expose the micro graft 120 to such growth factors before implanting the micro graft 120 into the matrix 120 or directly into the recipient site 220, thereby enhancing the viability of the micro graft 120.

[0127] To improve the control of the exemplary collection and / or matrix implantation procedures described herein, force sensors, optical sensors, and / or position sensors may be provided in communication with actuators 1120, 1130 and / or tubes 510, 1010 and / or pins 1040 as needed. For example, such sensors can be used to detect the penetration depth and / or penetration resistance of tubes 101 and / or pins 1040 so as to assist in collecting and / or implanting specific layers of the tissue and / or size of the micro graft 120.

[0128] Sensors may also be provided for detecting the presence of micro grafts within tubes 510, 1010. Such sensors may include, for example, a small current source provided in tubes 510, 1010 configured to detect a change in electrical resistance or resistance within tubes 510, 1010. For example, when a small current flows through needles 510, 1010 (e.g., in an electrode configuration), the detected resistance can indicate whether the needles 510, 1010 are empty or whether the micro graft 120 is present therein. Alternatively, a laser fiber and a photodetector may be provided within the needle to optically detect a change in scattered light indicating whether the micro graft 120 is present or not.

[0129] For example, if a plurality of such micro grafts are processed by two-dimensional scanning or traversing of the donor site 100 and / or the recipient site 1510, such micro graft sensors can be used to determine the number or proportion of actual micro grafts collected and / or implanted by the tubes 510, 1010. A relatively small number of "lost" micro grafts may be acceptable in a particular procedure, whereas a large proportion of "lost" micro grafts may indicate, for example, that the needles 510, 1010 need to be replaced, that the graft material is not viable or has not been accepted, and / or that the procedure needs to be repeated or continued. If the detected "lost" micro grafts are localized in the donor site 100, this may indicate that the tissue can be structurally different in a particular region, for example, that a mole or small scar may be present in the donor site 100.

[0130] A set of exemplary images of micro grafts collected from porcine skin and placed in a collagen gel is shown in FIG. 16. The epidermal portion of the micro grafts is the darker region at the lower end of the micro grafts in these images. Even at a short time of 12 hours (the second image from the left in FIG. 16), live cells migrating from the micro grafts into the surrounding gel matrix can be observed, as indicated by the bright region around the dark micro grafts. This migration is continuously observed for 72 hours after placing the micro grafts in the collagen gel (the rightmost image in FIG. 16), indicating that, according to an exemplary embodiment of the present disclosure, the micro grafts collected and placed in the matrix can survive for a long time and can provide viable graft material.

[0131] In further exemplary embodiments of the present disclosure, the micro-grafts 120 can be transplanted into a cleaned wound area 220 that has no matrix. For example, FIG. 17 shows a series of exemplary images showing the healing of a wound created in a nude (hairless) mouse, where micro-grafts obtained from the skin of a black mouse are transplanted into the wound area and then the wound can heal. After about 6 weeks, the wound appears to be well healed and tufts of black hair can be observed. These tufts of black hair at the nude recipient site suggest that at least a portion of the micro-grafts survived during the healing process and that functional hair follicles were successfully transplanted into the nude recipient.

[0132] FIG. 18A shows an exemplary wound formed in a porcine subject by surgically removing a substantially square region of full-thickness skin tissue (dermis below the epidermis and subcutaneous fat layer). The size of this wound is approximately 1.5 cm × 1.5 cm. Micro-grafts were collected from the donor site of the porcine subject using an apparatus similar to apparatus 500 shown in FIG. 5A according to the exemplary embodiments described herein. The micro-grafts were transplanted directly (without using a matrix) into one such wound. A second similar wound was also created in the subject and was able to heal without transplanting the micro-grafts.

[0133] The upper row of FIG. 18B shows a series of exemplary images depicting the four-week healing progression for the wound shown in FIG. 18A where the micro-grafts were implanted. The lower row of FIG. 18B shows exemplary images of the four-week healing progression for wounds without micro-grafts. The amount of wound reduction observed appears to be substantially decreased in wounds with micro-grafts implanted at the wound site. In contrast, wounds without micro-grafts appear to shrink more significantly during the healing process. Skin tissue reduction as wound healing is generally undesirable. For example, reduction of the skin around a joint can decrease the range of motion of the joint and can be painful when the joint is bent or extended. In some cases, the joint can be substantially or completely immobilized (e.g., severe tissue reduction around the temporomandibular joint can prevent the subject from opening their mouth and may require a liquid diet). Thus, implantation of the micro-grafts described herein into the wound area can reduce tissue shrinkage during wound healing, thereby reducing or avoiding the detrimental side effects of such shrinkage.

[0134] The exemplary methods and devices described herein may also be used to collect other types of biological tissue using the exemplary methods and devices described herein and need not be limited to skin. Embodiments of the present disclosure can facilitate the collection of small tissue portions (e.g., micro-grafts 120) from various organ or tissue types while reducing or avoiding the generation of damage at the donor site. The collected tissue portions can provide viable tissue that can be used in various transplantation or culturing processes.

[0135] The foregoing merely illustrates the principles of the present disclosure. Various modifications and changes to the described embodiments will be apparent to those skilled in the art in view of the teachings herein. The various exemplary embodiments described herein may be used interchangeably. Thus, those skilled in the art will be able to devise many techniques that, although not specifically described herein, embody the principles of the present disclosure. Therefore, they are within the spirit and scope of the present disclosure. All patents and publications cited herein are hereby incorporated by reference in their entirety.

Claims

1. 1. An apparatus for obtaining at least a portion of at least one biological tissue, comprising: at least one hollow tube having at least two tips disposed at a distal end of the hollow tube; a pin provided at least partially within a central lumen of at least one of said tubes; Equipped with At least one of the tubes has an inner diameter of less than about 1 mm; at least one portion of the hollow tube is structured for insertion into at least one of the biological tissues at a donor site, and at least one portion of the tube is withdrawn from the donor site to remove at least a portion of the donor site as transplant tissue from the donor site; the pin is controllably movable in a direction along a longitudinal axis of at least one of the tubes; The pin is configured to facilitate removal of at least a portion of the implant from at least one of the vessels.

2. The apparatus of claim 1 , wherein an inner diameter of at least one of the tubes is less than about 0.5 mm.

3. The device of claim 1 , wherein at least one of the tubes includes at least three tips disposed at a distal end of the tube.

4. The apparatus of claim 1 , further comprising a positioning arrangement configured to control a position of the pin within at least one of the tubes.

5. 10. The apparatus of claim 1, further comprising a substrate including at least one opening therethrough, wherein at least one of the tubes is slidably mounted to the substrate and configured to move at least partially through the at least one opening.

6. 6. The apparatus of claim 5, further comprising a first actuator configured to control a position of at least one of the tubes relative to the substrate, and a second actuator configured to control a position of the pin within the at least one of the tubes.

7. The apparatus of claim 6 , wherein the apparatus comprises a plurality of tubes.

8. The apparatus of claim 6 , wherein the apparatus comprises at least six tubes.

9. The apparatus of claim 8 , wherein the tubes are provided in at least one of a rectangular arrangement or a linear configuration relative to one another.

10. The apparatus of claim 1 , further comprising at least one sensor arrangement configured to detect the presence of at least one portion of said implant tissue within at least one of said vessels.

11. The device of claim 1 , wherein the angle between the side of the tip of at least one of the tubes and the longitudinal axis is less than about 15°.

12. 10. The device of claim 1, further comprising a hollow needle provided around at least a portion of the at least one hollow tube, the hollow needle structured to facilitate insertion of the at least one hollow tube into at least one of the at least one biological tissue or matrix material.

13. 13. The device of claim 12, further comprising a first actuator configured to control a position of at least one of the hollow tubes relative to a position of at least one of the hollow needles, and a second actuator configured to control a position of the pin relative to a position of the at least one of the hollow tubes.

14. 14. The apparatus of claim 13, further comprising a substrate including at least one opening therethrough, the hollow needle being slidably mounted to the substrate and structured to move at least partially through the at least one opening.

15. The apparatus of claim 14 , further comprising a third actuator configured to control a position of the hollow needle relative to a position of the substrate.

16. 1. A method for obtaining at least a portion of at least one biological tissue, comprising: providing a matrix material over the donor tissue site; placing a distal end of at least one hollow tube adjacent to a top surface of the matrix material; disposing a pin within a lumen of at least one of the tubes such that a distal end of the pin is proximate a distal end of the at least one of the tubes; lowering the pin and the at least one tube substantially together through a thickness of the matrix material such that distal ends of the at least one tube and the pin are adjacent a surface of the donor tissue site; advancing at least one of the tubes into the donor site to sever at least a portion of the at least one biological tissue from surrounding tissue, with a distal end of the pin remaining adjacent a surface of the donor tissue site; simultaneously lifting the at least one tube and the pin at substantially the same speed until a distal end of the at least one tube is proximate a lower surface of the matrix material; withdrawing at least one of the tubes from the matrix while maintaining the position of the pin substantially fixed relative to the matrix material, leaving at least a portion of at least one of the biological tissues within the matrix material; The method includes:

17. The method of claim 16 further comprising providing the matrix material with at least one growth factor.

18. 17. The method of claim 16, wherein the matrix material comprises at least one of collagen, low melting point agarose, or devitalized biological tissue.

19. 1. A method for harvesting and transplanting at least a portion of at least one biological tissue, comprising: positioning a distal end of at least one hollow tube adjacent to a top surface of at least one of said body tissues; disposing a pin within the lumen of at least one of said tubes such that a distal end of said pin is provided a predetermined distance rearward of a distal end of said at least one of said tubes; advancing the at least one tube into the at least one biological tissue to sever at least a portion of the at least one biological tissue from surrounding tissue, a distal end of the pin being positioned proximate a top surface of the at least one biological tissue; simultaneously lifting the at least one tube and the pin until the at least one tube is removed from the at least one living tissue, leaving at least a portion of the at least one living tissue within the at least one tube; inserting a hollow needle into a recipient material to a specific depth; providing at least one hollow tube containing at least a portion of at least one of the biological tissues within a lumen of the hollow needle such that a distal end of the hollow needle is adjacent to a distal end of the at least one hollow tube; retracting the hollow needle from the recipient material while maintaining the position of at least one of the hollow tubes within the recipient material; withdrawing at least one of the hollow tubes from the recipient material while maintaining the position of the pin substantially fixed relative to the recipient material such that at least a portion of the at least one biological tissue remains within the recipient material; The method includes:

20. 20. The method of claim 19, wherein the recipient material comprises at least one of a biologically compatible matrix or additional biological tissue.

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