Modular nozzle for coaxial extrusion

Abstract

The present invention relates to a nozzle for coaxial extrusion-based bioprinting, the nozzle comprising a plurality of separate modules, the modules comprising an inlet, an extrusion channel and an outlet. The modules are adapted to be assembled and reassembled and configured such that when the nozzle is formed, the extrusion channels are substantially coaxially and concentrically aligned along the longitudinal axis of the first module.

Description

Technical Field

[0001] The present invention relates to a nozzle for coaxial extrusion according to the preambles of the independent claims and in particular to a nozzle for coaxial extrusion based bioprinting. Background Art

[0002] Bioengineered or bioprinted tissue and organ-like structures have been designed as next-generation solutions for the lack of organ transplants and ultimately provide functionality in engineered tissue-based regenerative therapies. However, one of the major limitations in exploiting the development of tissues associated with size and function is the incorporation of vascular networks within these constructs. The complexity of these interconnected networks is difficult to achieve even with existing technologies. However, their presence in engineered tissues is critical to ensuring access to oxygen and nutrients and successful waste disposal. In addition, biofabrication of patient-specific vascular-like structures suitable for use as vascular grafts can help address the limitations faced by currently available autologous grafts and synthetic grafts.

[0003] Various nozzles for co-extruding materials are known. For example, WO2018 / 106705A1 shows a coaxial 3D printing nozzle for printing polymers, wherein the nozzle can be connected to a material reservoir. CN210174208U shows a coaxial bioprinting nozzle that can be assembled and disassembled using a spring / clamping mechanism to help maintain the coaxiality of the parts. US2016288414A1 discloses a bioprinter having a nozzle for bioprinting a tubular structure in a manner where one material is arranged around another material. US2019008998A1 and US20200164109A1 disclose a variety of coaxial bioprinting needles. JP6601931 shows a system utilizing coaxial extrusion of cell fibers. WO2020 / 056517A1 discloses a print head for producing a hollow fiber structure.

[0004] Bioprinting of coaxial constructs, such as tubular constructs with multiple layers containing different materials, requires a bioprinting nozzle capable of printing different materials in layers around a central core. An example of a nozzle capable of printing a tubular construct with multiple layers containing different materials is shown in Silva C, Cortés-Rodriguez C, Hazur J et al., 2020, Rational Design of a Triple-Layered Coaxial Extruder System: in silico and in vitro Evaluations Directed Toward Optimizing Cell Viability, Int J Bioprint, 6(4): 282. The nozzle shown is a three-layer coaxial nozzle suitable for attachment to a commercial bioprinter. Although the nozzle has shown some positive results specifically for bioprinting, the inventors have recognized the need for further improvements.

[0005] The common element in all bioprinting is that the use of biological materials requires specialized handling, as the materials are often sensitive to changes in temperature and pressure, and they often need to be handled under sterile or cleanroom-like conditions to avoid contamination of the material. For example, biological materials containing living cells must be handled so that cell viability is optimized and the introduction of other living materials is avoided.

[0006] Accordingly, the inventors of the present invention have recognized a need for an improved nozzle for extrusion-based bioprinting. Summary of the invention

[0007] It is an object of the present invention to provide a nozzle for coaxial bioprinting that provides adaptability in terms of the number of different layers that can be printed simultaneously in a tubular structure.

[0008] Another object of the present invention is to provide a nozzle for coaxial bioprinting that provides improved centering and alignment when assembling the nozzle.

[0009] Another object is to provide a nozzle for coaxial bioprinting that provides temperature control of the material used at least in the inlet of the nozzle.

[0010] Yet another object is to provide a nozzle for coaxial bioprinting that helps reduce internal pressure distribution and improve viability in 3D bioprinted constructs.

[0011] Furthermore, another object is to provide a nozzle which is easy to assemble, disassemble and clean all parts of the nozzle.

[0012] The above objects are achieved by the invention according to the independent claims. Preferred embodiments are set forth in the dependent claims.

[0013] According to the present invention, the nozzle for coaxial extrusion includes at least two or more modules for coaxial extrusion. Each module in the module has an inlet, an extrusion channel and an outlet. The inlet is suitable for the inflow and application of the material to be extruded via the extrusion channel. The extrusion channel connects the inlet and the outlet. In the first module of the two or more modules, the extrusion channel is arranged centrally along the longitudinal axis of the module. In the subsequent module, the extrusion channel is a tubular channel, which is suitable for being arranged along the longitudinal axis of the first module around the extrusion channel of the first module. The first module and the subsequent module are suitable for being installed together so that the extrusion channel is substantially coaxially and concentrically aligned along the longitudinal axis of the first module and the outlet of each module is coaxially arranged, for coaxial printing of a single-layer, double-layer or multi-layer tubular structure when the modules are connected. Two or more modules are constructed as separate components suitable for being assembled together by a user, i.e., forming a common unit for producing a nozzle. Two or more modules are also suitable for being disassembled from each other by a user and optionally reassembled with the same or different number of modules.

[0014] In addition, a method for bioprinting a coaxial tubular construct is disclosed, the method comprising the steps of providing a nozzle as disclosed herein; assembling 2, 3, 4, 5, 6 or more modules to form a nozzle; connecting at least two module inlets of the nozzle to separate compartments containing materials to be used in printing the construct; arranging the outlets of the connected nozzles above the print bed; and printing the tubular construct by extruding the material through the nozzle.

[0015] The present disclosure will become apparent from the detailed description given below. The detailed description and specific examples disclose preferred embodiments of the present disclosure only by way of illustration. Those skilled in the art can understand from the guidance in the detailed description that changes and modifications can be made within the scope of the present disclosure.

[0016] Therefore, it should be understood that the disclosure disclosed herein is not limited to the specific component parts of the described device or the steps of the described method, because such devices and methods can change. It should also be understood that the terms used herein are only used for the purpose of describing specific embodiments, and are not intended to be restrictive. It should be noted that, as used in the specification and the appended claims, unless the context clearly indicates otherwise, the articles "one", "a kind of", "the" and "said" are intended to represent one or more elements in the element. Therefore, for example, the "unit" or "the unit" mentioned may include several devices, etc. In addition, the words "include", "comprise", "contain", "contain" and similar words do not exclude other elements or steps. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1a A first aspect of a nozzle for coaxial extrusion-based bioprinting is shown. Figure 1b A printed construct produced by such a nozzle is illustrated.

[0018] Figure 2a and Figure 2b Another aspect of a nozzle for coaxial extrusion-based bioprinting is shown.

[0019] Figure 3 Shows Figure 2a and Figure 2b The nozzle module.

[0020] Figure 4a , Figure 4b and Figure 4c Various views of yet another aspect of a nozzle for coaxial extrusion-based bioprinting are shown.

[0021] Figure 4d A cross-sectional view of another aspect of a nozzle for coaxial extrusion-based bioprinting is shown, and Figure 4e An exploded view of another aspect of the nozzle is illustrated.

[0022] Figure 4f A further nozzle is shown when provided with a tube connected to the inlet and a temperature box system.

[0023] Figure 5a An exemplary configuration of a nozzle connected to a bioprinter is illustrated. Figure 5b Another exemplary configuration of a nozzle connected to a bioprinter is illustrated.

[0024] Figure 6 Examples of bioprinted 3D constructs printed using coaxial extrusion with temperature management-based bioprinting using any of the nozzles of the present disclosure are shown. DETAILED DESCRIPTION

[0025] Figure 1a and Figure 1b A schematic diagram of a first aspect of a nozzle 10 for coaxial extrusion-based bioprinting is shown. The illustrated nozzle can simultaneously extrude tubular constructs made of a total of six different materials. Figure 1a The top of the figure shows a cross-sectional view of the nozzle along the longitudinal axis L, and Figure 1a The bottom of shows a cross-sectional view of the nozzle along the plane AA of the top view across the longitudinal axis L. The central extrusion channel is coaxially surrounded by the second to sixth extrusion channels, each of which has an increased radius compared to the previous extrusion channel.

[0026] As will become apparent below, the nozzle 10 includes at least two or more modules, each of which has an inlet 12, an extrusion channel 13 and an outlet 14. The nozzle 10 can be provided with 2, 3, 4, 5 or 6 or more modules, depending on how many material layers are required in the bioprinted construct produced. Two or more modules are constructed as separate components, which are suitable for being assembled together by the user before use, that is, together forming a common unit as a nozzle. Therefore, when the modules are assembled to form the nozzle 10, the inner cavity of the extrusion channel of the second module and subsequent modules will form a tubular shape around the core extrusion channel of the first module. In addition, when the modules are assembled into the nozzle 10, the outlet 14 is preferably arranged substantially in the same plane across the longitudinal axis L, so that when the material is extruded through the nozzle, the printed or extruded construct produced will be a tubular structure having a central core and one or more coaxial tubular material layers surrounding the central core. Therefore, the assembled nozzle has a separate inlet 12 for each extrusion channel, and preferably has an aligned outlet 14. The inlet 12 is adapted for the inflow and application of material to be extruded via the extrusion channel 13. The material to be extruded may be applied to the nozzle inlet by connection to a material container or reservoir of a suitable type.

[0027] exist Figure 1b The corresponding view shows the Figure 1a Example of a construct extruded or printed by a nozzle. Figure 1b The top of FIG. 1 shows a cross-sectional view of the extruded tubular construct along the longitudinal axis L, and FIG. Figure 1b The bottom portion shows a cross-sectional view of the nozzle along plane BB of the top view across the longitudinal axis.

[0028] Thus, in particular, the nozzle described herein is suitable for being assembled and disassembled by the user, for example, just before starting extrusion, and can therefore be provided to the user as an unassembled set of modules. The user then selects the number of modules to be assembled into the nozzle according to the number of tubular layers with a central core of extruded material to be printed. In a non-limiting example, if a structure with three coaxial tubular material layers of a central core and a surrounding core is to be extruded, the user can assemble four modules into the nozzle. In another non-limiting example, if a structure with five coaxial tubular material layers of a central core and a surrounding core is to be extruded, the user can assemble six modules into the nozzle.

[0029] In other non-limiting examples, during the extrusion process, the program can be briefly paused and the nozzle reconfigured to include fewer or more modules to accommodate the number of material layers being extruded. Modules are easily removed from the nozzle, such as from each other, and additional modules are easily added to form new nozzle configurations. For example, after a structure with three tubular material layers of a central core and a surrounding core is printed for a certain period of time, the program can be paused and another module is added. When extrusion is resumed, subsequent extrusions will produce a structure with four material layers of a central core and a surrounding core. Alternatively, modules can be removed accordingly.

[0030] Thus, one advantage of a module of a nozzle adapted for assembly and disassembly by a user is that the number of material layers required for a particular extrusion can be quickly and easily adapted. Additionally, as the extrusion process of the tubular construct progresses, the user can change the number of layers in a single extruded construct to a greater or lesser number. Another advantage of being adapted for assembly and disassembly by a user is that the nozzle can be removed to easily clean the module after use, which is important when using biological materials.

[0031] To further illustrate the versatility of the disclosed nozzle, Figure 2a Another schematic diagram of an exemplary nozzle 20 in a semi-assembled configuration is illustrated in FIG. Figure 2b Another schematic diagram of an exemplary nozzle 20 in a fully assembled configuration is illustrated in FIG. Figure 2a and Figure 2b , a cross-sectional view along the longitudinal axis L is shown. Figure 1a The nozzle 20 has six modules 11. Figure 3A single module 11 is illustrated, for example, as provided prior to being assembled into the nozzle 20. As mentioned, the modules 11 are constructed as separate components adapted to be assembled together by a user to collectively form the nozzle 20, and the number of modules to be assembled can be selected by the user. When assembled, the first module provides an internal core extrusion channel. As modules are added, each of the subsequent modules 11 will have a separate extrusion channel 13 that surrounds the extrusion channel of the previous module and provides a tubular channel along the longitudinal axis L of the module 11. Thus, in the assembled nozzle, all extrusion channels of the installed modules will be substantially coaxially and concentrically aligned along the longitudinal axis L of the first module. In addition, the outlet 14 of each module is coaxially arranged for coaxial printing of single-layer, double-layer or multi-layer tubular structures.

[0032] The modules can preferably be assembled together using a centering attachment mechanism, such as by aligning each module 11 with an adjacent module 11 using an alignment member 15. The centering attachment mechanism ensures that the extrusion channels of different modules in the assembled nozzle are always arranged substantially coaxially and aligned concentrically along the longitudinal axis L of the first module. In addition, the centering attachment mechanism can further ensure that there is a specific distance between the extrusion channels of adjacent modules, so that a specific layer thickness is ensured during extrusion.

[0033] exist Figure 3 An example of a centering attachment mechanism is shown in , where the alignment member 15 is arranged as a raised ridge positioned around at least a portion of the length of the outer surface of the extrusion channel of the module in a semi-helical arrangement. However, other arrangements are possible as long as the alignment member helps to center and align each module relative to adjacent modules when assembling the nozzle so that the extrusion channels 13 of the assembled modules are substantially coaxial and concentrically aligned along the same longitudinal axis L when assembled. The use of the alignment member allows the user to easily assemble and disassemble the required number of modules for a particular situation. When the modules are assembled into a nozzle, the centering attachment mechanism improves the centering and alignment between the modules.

[0034] Figure 2a and Figure 2b It is illustrated that the inlets 12 may further be arranged to be staggered in the longitudinal direction, and wherein the inlets may also optionally extend in different radial directions from the longitudinal axis L, thereby allowing easy access to each inlet and the ability to arrange the inlets in a suitable configuration according to specific needs.

[0035] During use, the inlet of each module is connected to a separate material reservoir. This connection can be established before assembly into the nozzle and / or can be actively connected / disconnected during assembly / disassembly of the nozzle. The connection of the inlet to the material container or reservoir will be further detailed and illustrated in the following section.

[0036] In general, the modular construction of the nozzle as disclosed herein allows assembly, disassembly and reassembly to be performed quickly and easily, making the nozzle extremely adaptable. The number of modules used in the nozzle can be easily changed according to the specific construct that the user wants to print. For example, for a construct having only a core and one surrounding layer, the user can use two modules, or for a multilayer structure, the user can use 3, 4, 5, 6 or more modules. In one example, the nozzle includes 4, 5 or 6 modules, thereby providing a nozzle for printing a tubular construct having a central core and three tubular layers surrounding the core. In addition, as mentioned, the user can pause the extrusion program to rearrange the number of modules so that different numbers of layers can be extruded in different parts of the printed construct.

[0037] In addition, modules of different sizes can be provided, which are fitted together in different combinations. For example, modules with extrusion channels of different diameters suitable for fitting together to form a nozzle can be provided, so that the user can customize the thickness of each extruded layer by assembling the modules into a nozzle. Therefore, by selecting specific modules to be combined into a nozzle, both the number of layers and the thickness of the layers extruded separately can be changed according to the needs of a specific desired construct. As previously mentioned, the extrusion program can be suspended to replace one, two or more modules with other modules that provide different extrusion channel inner diameters, so that continued extrusion will provide one or more material layers of new thickness.

[0038] As mentioned above, another advantage of the modular construction is the ease of cleaning of all parts of the nozzle, making it suitable for extrusion of materials such as biomaterials that require clean, preferably sterile and controlled conditions. Cleaning in this sense includes any one or a combination of manual or automatic cleaning (e.g., with water and / or cleaning solutions), sterilizing and drying the module.

[0039] Bioprinting and the use of biological materials, such as materials containing cells, require not only a clean environment, but also typically a temperature-controlled environment. Figure 4a , Figure 4b and Figure 4c Another example of a nozzle is shown in . Figure 4a is a cross-sectional view of the nozzle 30, Figure 4b is a perspective view of the nozzle 30, and Figure 4c3 is a top view of a nozzle 30. The nozzle 30 has all the features described above, such as having individual modules 11, each of which has an inlet 12, an extrusion channel 13 and an outlet 14. The modules are all centrally aligned along a common longitudinal axis when assembled into the nozzle, so that the extrusion channels 13 are substantially coaxial and concentrically aligned. In addition, when the modules are assembled together, the outlets 14 are preferably arranged substantially in the same plane across the longitudinal axis L of the nozzle.

[0040] Figure 4d Another view of a nozzle 40 according to the present disclosure is illustrated in FIG. 4 , which shows a partial cross-sectional view of an assembled nozzle. The nozzle 40 has all the features described above, such as having separate modules 11, each of which has an inlet 12, an extrusion channel 13, and an outlet 14. Figure 4d In the view of FIG. 1 , not all modules in the nozzle are visible. The modules are all centrally aligned along a common longitudinal axis when assembled into the nozzle, so that the extrusion channels 13 are substantially coaxially and concentrically aligned. Furthermore, when the modules are assembled together, the outlets 14 are preferably arranged substantially in the same plane across the longitudinal axis L of the nozzle. Details of the arrangement of the extrusion channels 13 and outlets 14 are described in detail in FIG. Figure 1a or Figure 2a The outlets 14 may optionally be slightly staggered relative to each other.

[0041] Figure 4e Picture shows Figure 4d An exploded view of the nozzle 40, that is, the modules are separated from each other, similar to the way the modules are provided to the user before assembly.

[0042] Figure 4a , Figure 4b , Figure 4c and Figure 4d The nozzles are shown in an assembled state. However, as in the previous aspects, the modules 11 may be provided to the user as individual modules, for example in a kit of modules 11, so that the user may assemble a desired number of modules for a particular printing program. Figure 4e Examples of nozzles provided as separate parts can be seen in . Furthermore, the module can be provided as two or more modules intended for various layers, and depending on the combination selected, the modules will then provide various alternatives for the extruded layer thickness in the assembled nozzle. The modules are adapted so that a ready-to-use nozzle can be obtained with two modules or with three, four, five, six or more nozzles.

[0043] Each module 11 of the nozzle 30 also preferably has a thermally insulating enclosure 31 suitable for thermally insulating the material to be extruded through the extrusion channel (13) from external or internal temperature variations. Figure 4a and Figure 4d 1 and 2. It will be seen that two examples of insulating enclosures 31 are shown. When the nozzle is assembled, or in other words, when at least two or more modules 11 are assembled into a common nozzle, the insulating enclosures 31 are adapted to thermally insulate at least the material to be extruded or being extruded, i.e. any such material present in the nozzle. Preferably, the insulating enclosures 31 are adapted to substantially thermally insulate any extruded material in at least the extrusion channel of any individual module, and in some aspects, even when the modules are not assembled into a nozzle.

[0044] The insulating enclosure 31 of the nozzle is preferably suitable for insulating at least the extrusion channel 13 of the corresponding module 11 from external or internal temperature changes, and is preferably also suitable for insulating at least a portion of the extrusion channel of any other module in the same nozzle when the modules are assembled.

[0045] In other aspects, the enclosure 31 of the nozzle is preferably adapted to insulate the inlet 12 of the corresponding module 11 from external or internal temperature changes. In some aspects, as described above, the enclosure 31 is also adapted to insulate the extrusion channels of the corresponding module at least when the module is assembled with at least one other module to form a nozzle. The enclosure 31 of the module forming the nozzle can also be adapted so that when two or more modules are assembled to form a nozzle, all extrusion channels of the nozzle are insulated.

[0046] The enclosure 31 may be constructed in a variety of ways, such as from any suitable insulating material, so as to protect the materials within the module from the effects of the surrounding ambient temperature. Figure 4a and Figure 4d Two examples of enclosures 31 suitable for insulating at least each inlet 12 are shown, comprising a system 32 of channels arranged in the insulating material or enclosure 31, wherein the channels are suitable for containing a fluid for controlling the temperature of the inlet. The channels can be connected to means for generating a flow of the fluid through the channels. Figure 4f and Figure 5b Two such examples are illustrated in FIG.

[0047] Figure 4f A perspective view of an assembled nozzle 50 is shown, which is similar to the nozzle described above. The inlet of each module is provided with respective attachment ports and tubes ready for connection to a material reservoir. In addition, a temperature regulating member 55 can be connected to an inlet port 56 of the nozzle and an outlet port 56 from the nozzle via a tube so that a suitable temperature regulating fluid can be circulated through the system 32 of channels arranged in the insulating material or enclosure 31. The temperature regulating member 55 can include a pump and optionally include a power source for powering the pump.

[0048] exist Figure 5bIn another example shown, the temperature regulating member 65 can also be a temperature regulating tool head 65 for a bioprinter, wherein the power is supplied via the bioprinter itself. The nozzle 60 illustrated here is similar to the aforementioned nozzle disclosed herein. The temperature regulating tool head 65 can include a pump and is preferably connected via an inlet / outlet 56 device using a tube. The temperature regulating tool head 65 circulates a suitable fluid for controlling the temperature through a system 32 of channels arranged in the insulating enclosure of the nozzle 60 as previously described. Such a system can also include a controller adapted to control the temperature of the material to be extruded by controlling the fluid circulating through the system 32 of channels.

[0049] Thus, by controlling the temperature of the fluid circulating through the channel system 32, the extruded material within the nozzle module can be cooled, heated or maintained at a specific temperature. This is particularly important when extruding biological materials such as materials with living cells to ensure the viability of the material throughout the extrusion process.

[0050] As mentioned, the enclosure 31 can be arranged to insulate the extrusion channel 13 and / or the inlet 12 of at least the corresponding module 11, so that the temperature of all materials passing through the inlet 12 and the extrusion channel 13 can be controlled. The use of fluids arranged in the channels or other similar controllable isolation devices allows the materials within the modules to be cooled and heated. In some aspects, the enclosure 31 of the nozzle can be configured for separate temperature control of each inlet or module, so that the enclosure 31 can provide different temperatures for different materials used in the nozzle 30. An example of such separate temperature control connects separate devices for generating the flow of cooling or heating fluids through the above-mentioned channels to different modules, so that each module can be provided with an optimal temperature for the selected extrusion material.

[0051] The ability to provide a nozzle adapted to the specific requirements of both a clean / sterile environment and a temperature-controlled environment makes the nozzle 30 particularly suitable for bioprinting, for example to ensure high viability of living cells in the final construct.

[0052] Furthermore, compared to other nozzles, the nozzles disclosed herein offer a more compact design, even with built-in temperature regulation, and thus achieve a smaller volume. This makes the nozzles easier to connect and use, for example, in a bioprinter.

[0053] Figure 5a It illustrates how a nozzle 10 , 20 , 30 , 40 , 50 , 60 as disclosed herein may be attached to a separate printhead or tool head 110 of a bioprinter 100 , such that each material compartment of the printhead 110 is connected to a unique inlet 12 of the nozzle 30 via a dedicated tube.

[0054] As in Figure 5b As an alternative illustrated in , the nozzles 10, 20, 30, 40, 50, 60 disclosed herein can be mounted directly on one of the tool heads 110 of the bioprinter 100, for example a tool head containing material for the core of the desired construct, and the inlet 12 for the core module 11 is directly connected to the material container of the tool head. The remaining inlets 12 can then be connected to the other tool heads 110 by tubes. It is also conceivable to connect the nozzle to only a single tool head. In this case, preferably, the tool head has at least two different material compartments that can be connected to the inlet of the module in the nozzle. As described above, the temperature regulating member 65, for example a temperature regulating tool head 65 for a bioprinter, circulates a suitable fluid for controlling the temperature through a system 32 of channels in the insulating enclosure 31 of the nozzle.

[0055] A method for bioprinting coaxial tubular constructs using the disclosed nozzle, the method comprising the steps of: assembling 2, 3, 4, 5, 6 or more modules as described above to form a nozzle; connecting at least two module inlets of the nozzle to separate compartments, such as in a printer tool head, which contain materials to be used in the final construct; arranging the outlets of the connected nozzles above the print bed; and printing the tubular construct by extruding the material through the nozzle.

[0056] As mentioned, use of the disclosed nozzles allows for printing of multi-layer tubular structures. Such constructs can be used in a variety of applications. For example, each layer can have different materials, or can have the same material in different concentrations or ratios to produce gradients and / or layered structures. In other aspects, the multi-layer configuration allows for alternating patterns of printed materials.

[0057] Typically, a sacrificial ink is used for the central core so that it can be removed after printing to obtain a pourable annular structure. Figure 6 Some examples of printed tubular structures and microfluidic constructs made of biomaterials and cells are shown in Figure 1. This shows that complex cylindrical tissues, such as those made up of many different cells and layers, such as veins, blood vessels or arteries, can be printed, for example for transplantation. For research purposes, it can be used to print other complex tissues such as hair or muscle fibers.

[0058] On the other hand, a nozzle assembly is disclosed, which includes any of the above-mentioned nozzles and also includes a pressure regulation configuration. For example, the extrusion pressure can be adjusted by connecting a pressure regulator to one or more of the module inlets via appropriate valves and tubes. Such a system may include a controller suitable for controlling extrusion through the nozzle by means of operating the pressure regulator. Therefore, the extrusion pressure can be varied in each individual extrusion channel to adapt to changes in material type and / or changes in print size. As described in detail below, experiments have shown that the nozzle of the present invention is particularly suitable for the specific needs of biomaterials and bioprinting. Compared with commercial coaxial nozzles currently available on the market, significant improvements in pressure distribution and temperature dynamic management have been demonstrated and measured.

[0059] The currently disclosed nozzles can be used in the FRESH (Freeform Reversible Embedding of Suspended Hydrogels) printing method, a 3D bioprinting method designed to address many of the limitations faced when using soft biomaterials for tissue engineering applications.

[0060] The tip extender can be attached to the currently disclosed nozzle to deposit material into the vat of material when it is used for suspension printing.Using the disclosed nozzle allows easy printing of tubular structures using a variety of materials, even when using softer materials.

[0061] The presently disclosed nozzles may be combined with a material selector mixing system to dispense materials in various gradients, ratios, or rapid switching modes.

[0062] The currently disclosed nozzle can be coupled to a high voltage tuning system for cell electrowriting.Using the disclosed nozzle allows for easy printing of tubular structures using a variety of materials.

[0063] In an example, a multilayer vascular-like structure is produced by coaxial bioprinting using a nozzle according to the present disclosure. The materials used include living cells and printable hydrogel biomaterials. The natural architecture of large and medium diameter blood vessels is simulated by manufacturing double-layer vascular-like structures and perfusable channels by means of direct 3D bioprinting and embedded 3D printing. The known bioprinter is customized to include four complete functional print heads, which have coaxial nozzles as disclosed herein and mainly use four modules in the nozzle. The print head is basically a tool head as a material box for a bioprinter, and each material box is connected to a separate module inlet. The configuration is similar to the configuration shown in Figure 5, except that only four channels / modules are used. Several different coaxial nozzles with four extrusion channels of different diameters are used, and in each specific combination of modules, the size is adapted to fit together and produce different layers of a specific thickness in the desired construct. In various combinations, the smallest (inner) channel had an inner diameter of the extruded channel between 0.30 mm and 0.84 mm, while the largest (outer) channel had an inner diameter of the extruded channel between 3.00 mm and 3.55 mm.

[0064] In the bioprinting experiments, the inner (core) channel used the sacrificial ink CELLINK PLU RONICS, and in some of the subsequent layers human foreskin fibroblasts HFF-1 ( SRCR-1041 TM ) and primary human umbilical vein endothelial cells (HUVEC) ( PCS-100-010). A cross-linking agent was used in the outermost layer to cross-link the vascular construct immediately after extrusion. In some cases, the vascular-like construct was printed embedded in a supporting hydrogel. As observed from the microscope images, the results show that a double-layer vascular-like construct was successfully manufactured. In addition, it was demonstrated that the extrusion pressure is a key printing parameter for cell viability on these multi-layer tissue constructs manufactured by means of a coaxial extrusion system. The bioprinting procedure was performed under the following circumstances: using four designed coaxial extrusion systems, and while changing the extrusion pressure of the print head for the channels containing living cells, separate pressure regulators were used, which allowed the control of pressurized air from the external pressure compressor to each inlet of the module of the nozzle. The results showed that increasing the extrusion pressure to more than 20% above the standard extrusion pressure would have a significant negative impact on cell viability. However, when the layer area and layer thickness of different channels were changed, no significant effect on cell viability was seen. Therefore, the inlet pressure must be carefully set to an optimal value in order to maintain cell viability.

[0065] The present invention is not limited to the preferred embodiments described above. Alternatives, modifications and equivalents may be used. Therefore, the above embodiments should not be considered to limit the scope of the present invention, which is defined by the appended claims.

Claims

1. A nozzle (10, 20, 30, 40, 50, 60) for coaxial extrusion, the nozzle (10, 20, 30, 40, 50, 60) comprising: At least two or more modules (11) for coaxial extrusion, each of the modules (11) comprising an inlet (12), an extrusion channel (13) and an outlet (14), the inlet (12) being suitable for the inflow or application of material to be extruded via the extrusion channel (13), and the extrusion channel (13) connecting the inlet (13) and the outlet (14), wherein, in a first module of the two or more modules (11), the extrusion channel (13) is arranged centrally along the longitudinal axis (L) of the module (11), wherein, in the subsequent module, the extrusion channel (13) is a tubular channel, which is suitable for being arranged around the extrusion channel (13) of the first module along the longitudinal axis (L) of the first module, wherein the first module and the subsequent modules are adapted to be mounted together such that the extrusion channels (13) are substantially coaxially and concentrically aligned along the longitudinal axis (L) of the first module and the outlets (14) of each module are coaxially arranged for coaxially printing a single-layer, double-layer or multi-layer tubular structure when the modules (11) are connected, wherein two or more of the modules are constructed as separate components suitable for being assembled together by a user to jointly form the nozzle (10, 20, 30, 40, 50, 60), Wherein, two or more of the modules (11) are also suitable for being disassembled from each other by the user and optionally reassembled with the same or different number of modules (11).

2. The nozzle (10, 20, 30, 40, 50, 60) according to any of the preceding claims, wherein Two or more of said modules (11) are adapted to be assembled together using a centred attachment mechanism.

3. The nozzle (10, 20, 30, 40, 50, 60) according to claim 3, wherein: The centering attachment mechanism includes an alignment member (15).

4. The nozzle (10, 20, 30, 40, 50, 60) according to any of the preceding claims, wherein The number of at least two or more modules (11) for coaxial extrusion is 2, 3, 4, 5 or 6 or more modules.

5. The nozzle (10, 20, 30, 40, 50, 60) according to any of the preceding claims, further comprises at least one insulating enclosure (31), which is arranged to insulate the material to be extruded through the extrusion channel (13) so as to protect the material from external or internal temperature changes.

6. The nozzle (10, 20, 30, 40, 50, 60) according to claim 5, wherein: The thermally insulating enclosure (31) is suitable for thermally insulating at least the extrusion channel (13) of the corresponding module (11) to protect the extrusion channel (13) from external or internal temperature changes.

7. The nozzle (10, 20, 30, 40, 50, 60) according to claim 5 or 6, wherein: The insulating enclosure (31) is suitable for insulating the inlet (12) of the corresponding module (11) to protect the inlet (12) from external or internal temperature changes.

8. The nozzle (10, 20, 30, 40, 50, 60) according to any one of claims 5 to 7, wherein: The enclosure (31) of the nozzle is configured for individual temperature control of each respective module.

9. The nozzle (10, 20, 30, 40, 50, 60) according to any one of claims 5 to 8, wherein: The enclosure (31) comprises a system of channels (32) arranged to contain a fluid for controlling the temperature of at least the inlet (12) and / or the extrusion channel (13) of the corresponding module (11).

10. A nozzle assembly for bioprinting, the nozzle assembly comprising a nozzle according to any one of claims 1 to 9, the nozzle assembly also comprising a temperature regulating member (55, 65), the temperature regulating member (55, 65) being arranged to circulate a fluid through the system (32) of the channel so as to control the temperature of the material to be extruded.

11. The nozzle assembly according to claim 10, wherein: The temperature regulating member (65) is a temperature regulating tool head for a bio-printer.

12. A nozzle assembly for bioprinting, the nozzle assembly comprising a nozzle according to any one of claims 1 to 9 or a nozzle assembly according to claim 10 or 11, the nozzle assembly also comprising at least one pressure regulator, the at least one pressure regulator being arranged to change the extrusion pressure in at least one of the modules.

13. A system for coaxial extrusion, comprising a nozzle assembly according to claim 10 or 11, the system further comprising a controller adapted to control the temperature of the material to be extruded by controlling a fluid circulating through the system (32) of channels.

14. A system for coaxial extrusion, the system comprising the nozzle assembly of claim 12, the system further comprising a controller adapted to control extrusion through the nozzle by operating the at least one pressure regulator.

15. The system of claim 13 or 14, further comprising two or more tool heads, the tool heads being material cartridges for a bioprinter, and each material cartridge being adapted to be connected to a separate module inlet of the nozzle.

16. A method for bioprinting a coaxial tubular construct, the method comprising the steps of: Providing a nozzle according to any one of claims 1 to 9 or a nozzle assembly according to any one of claims 10 to 12; Assembling 2, 3, 4, 5, 6 or more modules to form the nozzle; connecting at least two module inlets of the nozzle to separate compartments containing materials to be used in printing the construct; Arranging the outlet of the connected nozzle above a print bed; as well as The tubular construct is printed by extruding material through the nozzle.

Images