Connecting device and method for obtaining SVF by treating fat through mechanical method

Abstract

The invention provides a connecting device and method for processing fat through a mechanical method to obtain SVF. The connecting device comprises a shell, a connector pipe, a wheel disc, a rolling wheel, a window and a limiting block. The window is arranged on the shell; the connector pipes are arranged at the two ends of the shell. The wheel disc is rotationally connected into the shell through a shaft, and the roller protrudes out of the shell and is meshed with the wheel disc; the limiting block is fixed in the shell and makes rolling contact with the side face of the wheel disc. Limiting structures and processing holes are arranged on the wheel discs, the wheel discs can be connected in a pressing mode through the processing holes, and the processing holes penetrate through the wheel discs and can be communicated with the connector pipes. The limiting structure is in rolling fit with the limiting block. The connecting device provided by the invention has a convertible aperture, so that the aperture shape and number between injectors can be adjusted, and the purpose of cutting and emulsifying fat at different degrees is achieved. Meanwhile, the aperture size and the shape of the connecting device can be customized according to the separation and preparation process of the SVF.

Description

Technical Field

[0001] The present invention relates to the technical field of medical devices, and particularly relates to a connecting device and method for mechanically treating fat to obtain SVF. Background Art

[0002] Human fat is rich in stromal vascular fraction (SVF), which is a mixture formed by a variety of cells with repair functions and cytokines. It includes adipose-derived stem cells (ADSCs), hematopoietic stem cells, endothelial progenitor cells, endothelial cells, vascular smooth muscle cells, fibroblasts, lymphocytes, macrophages, mast cells, platelets, etc. Especially ADSCs among them have good immune regulation ability and pro-angiogenic ability, and there have been many clinical application reports in assisted fat transplantation and osteoarthritis treatment.

[0003] Most of the fat used for preparing SVF is obtained by liposuction surgery, which contains the tumescent fluid added during the liposuction process. Usually, a centrifugation step is carried out before SVF preparation. After centrifugation, the fat obtained from liposuction surgery is divided into four layers: oil phase, fat layer, tumescent fluid, and tissue fragments. The fat layer among them (the fat obtained in this step is generally called "Coleman fat") is taken for subsequent treatment. The subsequent treatment can generally be divided into two types: enzymatic method and mechanical method. The enzymatic method uses collagenase to digest and break down the extracellular matrix components, enabling the SVF-related cells to be collected by centrifugation. The mechanical separation method emulsifies the fat by means of ultrasound, oscillation, etc., destroys the extracellular matrix components, and separates the SVF-related cells. However, the disadvantages of the enzymatic method are that it takes a long time and the operation is complex; using collagenase brings a risk of residue; additional operations need to be carried out outside the surgical environment, and the risk of contamination caused by the operation is higher. The advantages of mechanical separation are that it takes a short time and the operation is simple; there is no risk of residue caused by the introduction of foreign substances; part of the extracellular matrix components are retained, presenting a gel-like state, which is more suitable for tissue filling. Generally speaking, the characteristics of short time-consuming and simple operation of the mechanical separation method are particularly suitable for operation in the operating room, that is, SVF can be prepared shortly after the liposuction surgery is completed for fat backfilling or other treatment surgeries. The patient does not need to undergo a second operation, which has advantages in terms of treatment convenience and surgical risk control. However, at present, the yield and viability of SVF obtained by mechanical separation are not high.

[0004] In summary, the following problems exist in the prior art: how to improve the yield and viability of SVF obtained by mechanical separation. Summary of the Invention

[0005] The present invention provides a connection device and method for mechanically treating fat to obtain SVF, and the technical problem to be solved is how to improve the yield and viability of SVF obtained by mechanical separation.

[0006] To achieve the above object, on the one hand, the present invention proposes a connection device for mechanically treating fat to obtain SVF, including:

[0007] A housing, interface pipes provided at both ends of the housing, a disk rotatably connected inside the housing, rollers meshing with the disk, a viewing window opened on the housing, and a limiting block fixed inside the housing;

[0008] A plurality of the disks are provided, the disks are rotatably connected inside the housing through a shaft, a limiting structure and treatment holes are provided on the disks, the disks can be tightly connected through the treatment holes, and the treatment holes penetrate through the disks and can communicate with the interface pipes;

[0009] The limiting structure is provided on the side surface of the disk, so that the limiting block is in rolling contact with the limiting structure;

[0010] The rollers protrude from the housing, and the disks are rotated by rolling the rollers, so that the treatment holes communicate with the interface pipes.

[0011] Specifically, there is a cavity between the disks; three disks are provided, including: a first disk; a second disk; a third disk;

[0012] Three rollers are provided, including: a first roller, a second roller, and a third roller;

[0013] The first roller meshes with the first disk, the second roller meshes with the second disk, and the third roller meshes with the third disk.

[0014] Specifically, the limiting structure is provided on both sides of the disk, and the limiting structure includes: a limiting ring groove and a limiting groove;

[0015] The limiting ring groove is opened on two side surfaces of the disk with the shaft as the center;

[0016] The limiting groove is hemispherical, the limiting groove is provided on the limiting ring groove, and the diameter of the limiting groove is greater than the width of the limiting ring groove.

[0017] Specifically, a limiting spring and a limiting ball are provided inside the limiting block; the limiting ball can protrude from the limiting block;

[0018] The limiting spring and the limiting ball are in pressing contact, and the limiting ball is in rolling contact with the limiting structure on the disk.

[0019] Specifically, the processing holes of the first turntable include: a first cutting hole, a second cutting hole, and a third cutting hole;

[0020] The processing holes of the second turntable include: a first extrusion hole, a second extrusion hole, and a third extrusion hole.

[0021] Specifically, a well-shaped wire mesh is arranged in the first cutting hole; a cross-shaped wire mesh is arranged in the third cutting hole; and a plurality of silk threads are arranged radially in the second cutting hole, and both ends of the silk threads are fixed on the inner wall of the second cutting hole.

[0022] Specifically, the aperture of the first extrusion hole is smaller than that of the second extrusion hole, and the aperture of the second extrusion hole is smaller than that of the third extrusion hole.

[0023] Specifically, the limiting blocks include: a first limiting block, a second limiting block, a third limiting block, and a fourth limiting block;

[0024] The first limiting block is fixed on the inner wall of the housing, and the first limiting block is in rolling contact with the limiting structure on one side of the first turntable;

[0025] The second limiting block is arranged in the cavity between the first turntable and the second turntable, and the second limiting block is fixed on the shaft. The second limiting block is in rolling contact with the limiting structure on the other side of the first turntable, and the second limiting block is in rolling contact with the limiting structure on one side of the second turntable;

[0026] The third limiting block is arranged in the cavity between the second turntable and the third turntable, and the third limiting block is fixed on the shaft. The third limiting block is in rolling contact with the limiting structure on the other side of the second turntable; the third limiting block is in rolling contact with the limiting structure on one side of the third turntable;

[0027] The fourth limiting block is fixed on the inner wall of the housing, and the fourth limiting block is in rolling contact with the limiting structure on the other side of the third turntable.

[0028] Specifically, the interface pipe is connected to an injection device.

[0029] On the other hand, the present invention provides a method for mechanically treating fat to obtain SVF. Using the connecting device for mechanically treating fat to obtain SVF as described above, connect the interface pipe of the connecting device to an injection device;

[0030] Rotate the turntable to select a corresponding processing hole to communicate with the interface pipe;

[0031] Push the injection device so that the fat passes from one injection device through the connecting device for obtaining SVF from mechanically processed fat into the injection device at the other end;

[0032] Obtain SVF from the processed fat.

[0033] Connecting two syringes using the connecting device of the present invention can ensure complete sealing. The connecting device of the present invention has a convertible aperture, which can adjust the aperture shape and quantity between the syringes, achieving the purpose of cutting and emulsifying fat to different extents. When adjusting the aperture, there is no need to replace the connector, which can ensure that the fat in the syringe will not be exposed multiple times, reducing the risk of contamination of SVF during the operation. At the same time, the aperture size and shape of the connecting mechanism can be customized according to the separation and preparation process of SVF. Description of the Drawings

[0034] Figure 1 is a schematic structural view of a connecting device for obtaining SVF from mechanically processed fat according to an embodiment of the present invention;

[0035] Figure 2 is a schematic view of the installation and use of a connecting device for obtaining SVF from mechanically processed fat according to an embodiment of the present invention;

[0036] Figure 3 is a top view of a connecting device for obtaining SVF from mechanically processed fat according to an embodiment of the present invention;

[0037] Figure 4 is a side view of a connecting device for obtaining SVF from mechanically processed fat according to an embodiment of the present invention;

[0038] Figure 5 is a front view of the internal structure section of a connecting device for obtaining SVF from mechanically processed fat according to an embodiment of the present invention;

[0039] Figure 6 is a side view of the internal structure section of a connecting device for obtaining SVF from mechanically processed fat according to an embodiment of the present invention;

[0040] Figure 7 is a schematic structural view of the first turntable according to an embodiment of the present invention;

[0041] Figure 8 is a schematic structural view of the second turntable according to an embodiment of the present invention;

[0042] Figure 9 is a front view of the second turntable according to an embodiment of the present invention;

[0043] Figure 10 is a schematic diagram of the percentage of SVF in Coleman fat in the examples and comparative examples;

[0044] Figure 11 Typical fluorescence images of AO / PI staining for the examples and comparative examples;

[0045] Figure 12 Schematic diagrams of the cell viability of SVF in the examples and comparative examples;

[0046] Figure 13 Cell density graphs of each group of SVF in the examples and comparative examples;

[0047] Figure 14 Cell morphology graph in Example 3;

[0048] Figure 15 The first flow cytometry sample detection result graph of the examples of the present invention;

[0049] Figure 16 The second flow cytometry sample detection result graph of the examples of the present invention;

[0050] Figure 17 The third flow cytometry sample detection result graph of the examples of the present invention;

[0051] Figure 18 The fourth flow cytometry sample detection result graph of the examples of the present invention;

[0052] Figure 19 The fifth flow cytometry sample detection result graph of the examples of the present invention;

[0053] Figure 20 Flow cytometry sample detection statistical graph of the examples of the present invention;

[0054] Figure 21 The first flow cytometry detection result graph of the isotype control of the examples of the present invention;

[0055] Figure 22 The second flow cytometry detection result graph of the isotype control of the examples of the present invention;

[0056] Figure 23 The third flow cytometry detection result graph of the isotype control of the examples of the present invention;

[0057] Figure 24 The fourth flow cytometry detection result graph of the isotype control of the examples of the present invention;

[0058] Figure 25 The fifth flow cytometry detection result graph of the isotype control of the examples of the present invention;

[0059] Figure 26 Flow cytometry detection statistical graph of the isotype control of the examples of the present invention.

[0060] Description of the attached drawing reference numerals:

[0061] 1. Housing; 2. Injection device; 11. Roller; 12. Interface pipe; 13. Window; 14. Disc; 15. Limit block; 16. Shaft; 17. Fixed rod; 21. Push rod; 111. First roller; 112. Second roller; 113. Third roller; 131. First window; 132. Second window; 133. Third window; 141. First disc; 142. Second disc; 143. Third disc; 1411. First cutting hole; 1412. Second cutting hole; 1413. Third cutting hole; 1414. First prompt area; 1415. Second prompt area; 1416. Third prompt area; 1421. First extrusion hole; 1422. Second extrusion hole; 1423. Third extrusion hole; 1424. Fourth prompt area; 1425. Fifth prompt area; 1426. Sixth prompt area; 1417. First limit ring groove; 1441. Second limit ring groove; 1442. Limit groove; 151. First limit block; 152. Second limit block; 153. Third limit block; 154. Fourth limit block; 155. Limit ball; 145. Shaft hole. Detailed implementation manners

[0062] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0063] An embodiment of the present invention provides a connection device for mechanically treating fat to obtain SVF, as Figure 1 shown, including:

[0064] A housing 1, interface pipes 12 provided at both ends of the housing 1, a disc 14 rotatably connected inside the housing 1, rollers 11 engaged with the disc 14, a window 13 opened on the housing 1, and a limit block 15 fixed inside the housing 1;

[0065] The window 13 is provided on the housing 1; the window 13 is used to display the aperture type or size on the disc 14. The interface pipes 12 are provided at both ends of the housing 1, as Figure 2 shown, the interface pipes 12 are connected to the injection device 2, and the connection method is preferably a threaded connection.

[0066] The disc 14 is rotatably connected inside the housing 1 through a shaft 16, as Figure 4As shown, the roller 11 protrudes from the housing 1 and the roller 11 meshes with the disc 14; the disc 14 is rotated by rolling the roller 11, so that the processing hole communicates with the interface pipe 12. The limiting block 15 is fixed inside the housing 1 and is in rolling contact with the side surface of the disc 14; the limiting structure and the processing hole are provided on the disc 14, and the discs 14 can be tightly connected through the processing holes. The processing holes penetrate the disc 14 and can communicate with the interface pipe 12; the limiting structure is in rolling cooperation with the limiting block 15. A marking area is provided on the disc 14, and the position of the marking area can correspond to the position of the window 13. When the marking area rotates to the position of the window 13, the operator can distinguish which processing hole is communicating with the interface pipe 12 by seeing the corresponding marking area.

[0067] There is a cavity between the discs 14; there are 3 discs 14, including: the first disc 141; the second disc 142; the third disc 143; preferably, the number of discs 14 is 2, 4, or 5. The roller 11 is used to adjust the internal aperture of the connecting device, where, as Figure 3 shown, there are 3 rollers 11, including: the first roller 111, the second roller 112, and the third roller 113; among them, the first roller 111 meshes with the first disc 141, the second roller 112 meshes with the second disc 142, and the third roller 113 meshes with the third disc 143. By rotating the roller 11, the roller 11 drives the disc 14 to rotate, and the processing hole communicating with the interface pipe 12 can be changed. That is to say, rotating the roller 11 can select the required type or size of the processing hole.

[0068] The limiting structure on the disc 14 is arranged on both sides of the disc 14, and the limiting structure includes: a limiting ring groove and a limiting groove 1442; the limiting ring groove is opened on the two side surfaces of the disc 14 with the axis 16 as the center; the limiting groove is hemispherical, the limiting groove is arranged on the limiting ring groove, and the diameter of the limiting groove is larger than the width of the limiting ring groove.

[0069] The limiting block 15 is internally provided with a limiting spring and a limiting ball 155; the limiting ball can protrude from the limiting block 15; the limiting spring and the limiting ball are in pressing contact, and the limiting ball is in rolling contact with the limiting structure on the disc 14. The limiting block 15 is used to limit the rotational displacement of the disc, so that the holes of the disc correspond to the orifices of the other two discs in the connecting device.

[0070] As Figure 5 shown, the limiting block 15 includes: the first limiting block 151, the second limiting block 152, the third limiting block 153, and the fourth limiting block 154;

[0071] The first limiting block 151 is fixed on the inner wall of the housing 1, and the first limiting block 151 is in rolling contact with the limiting structure on one side of the first disc 141;

[0072] The second limiting block 152 is arranged in the cavity between the first disc 141 and the second disc 142, and the second limiting block 152 is fixed on the shaft 16 through a fixing rod. The second limiting block 152 is in rolling contact with the limiting structure on the other side of the first disc 141, and the second limiting block 152 is in rolling contact with the limiting structure on one side of the second disc 142;

[0073] The third limiting block 153 is arranged in the cavity between the second disc 142 and the third disc 143, and the third limiting block 153 is fixed on the shaft 16 through a fixing rod 17. The third limiting block 153 is in rolling contact with the limiting structure on the other side of the second disc 142; the third limiting block 153 is in rolling contact with the limiting structure on one side of the third disc 143;

[0074] The fourth limiting block 154 is fixed on the inner wall of the housing 1, and the fourth limiting block 154 is in rolling contact with the limiting structure on the other side of the third disc 143.

[0075] As Figure 5 , the leftmost and rightmost sliding limiting blocks are fixed on the housing 1 of the connecting device, and the middle two limiting blocks are fixed on the shaft 16. The spring that is always in a compressed state inside the limiting block provides a force for the limiting ball (ball), and at the same time, the outer shell of the limiting block restricts the displacement of the ball, so that only the tip leaks out of the outer shell and contacts the limiting ring groove. When the disc rotates and the selectable aperture corresponds to the connection port, the ball will enter a deeper limiting groove, realizing the displacement limitation of the disc component. When it is necessary to replace the selectable aperture, rotate the roller with a greater force, and the ball will slide out of the deeper limiting groove into the limiting ring groove. When the next selectable aperture corresponds to the interface, the ball will enter the next deeper limiting groove. When the ball enters the limiting groove, the currently corresponding aperture mark connected to the interface 12 will be displayed on the window 13. As Figure 3 , the window 13 is successively the window 131, the window 132, and the window 133 from left to right. For example, as Figure 9 , when the limiting ball 155 in the second limiting block 152 enters the limiting groove 1442 of the second disc 142, the currently corresponding aperture mark connected to the interface pipe 12 (interface) will be displayed on the window 132.

[0076] As Figure 7 As shown, the processing holes of the first disc 141 include: a first cutting hole 1411, a second cutting hole 1412, and a third cutting hole 1413; a well-shaped wire mesh is arranged in the first cutting hole 1411; a cross-shaped wire mesh is arranged in the third cutting hole 1413; a plurality of silk threads are arranged radially in the second cutting hole 1412, and both ends of the silk threads are fixed on the inner wall of the second cutting hole 1412.

[0077] As Figure 8As shown, the second wheel disc 142 is provided with three through-hole structures, and the through-holes have different diameters. The through-holes are used as processing holes to process fat, and the fat that has been mechanically divided by the apertures of the wire mesh on the left and right wheels is squeezed and emulsified. Specifically, the processing holes of the second wheel disc 142 include: a first extrusion hole 1421, a second extrusion hole 1422, and a third extrusion hole 1423. The aperture of the first extrusion hole 1421 is smaller than the aperture of the second extrusion hole 1422, and the aperture of the second extrusion hole 1422 is smaller than the aperture of the third extrusion hole 1423. Preferably, the first extrusion hole 1421 is a hole with a diameter of 1.4 mm, the second extrusion hole 1422 is a hole with a diameter of 2 mm, and the third extrusion hole 1423 is a hole with a diameter of 2.4 mm. On the outer edge of the wheel near the through hole, there is a character sticker marking the through hole near the through hole for displaying on the connection mechanism window, prompting the number of the through hole currently corresponding to the interface pipe 12 of the connection device, that is, the fourth prompt area 1424 mark corresponds to the first extrusion hole 1421, the fifth prompt area 1425 mark corresponds to the second extrusion hole 1422, and the sixth prompt area 1426 mark corresponds to the third extrusion hole 1423. When the limiting ball 155 in the limiting block rolls into the limiting groove in the second limiting ring groove 1441, the mark of the aperture currently connected to the interface pipe 12 will be displayed on the window 132.

[0078] There is a hole in the center of the wheel disc, which is an axle hole 145, and the axle 16 passes through the axle hole 145 to connect the three wheel discs. Figure 6 As shown, the wheel disc can rotate on the shaft 16 when the roller 11 is turned. Figure 5 As shown, Figure 5 This is a front view of the internal structure of the connection device, in which the rollers correspond to the left wheel disc, the middle wheel disc, and the right wheel disc in the adjustment connection mechanism. The center of the three wheels is passed by the shaft. The sliding limit mechanism at the leftmost and rightmost ends is fixed on the connection mechanism housing, and the two middle limit blocks are fixed on the shaft. There are annular grooves on both sides of the wheel disc. There is a corresponding deeper circular groove on the annular groove on the same radius connecting line as the optional aperture. The limit block 15 is a sliding limit mechanism. The limit ball 155 in the limit block 15 can roll in the annular groove. When the optional aperture corresponds to the interface tube 12, the limit ball 155 slides into the circular groove and is blocked from rotating and provides mechanical feedback to achieve the purpose of positioning. When it is necessary to continue to change the selected aperture, use a slightly larger force to move the roller (limit ball 155) corresponding to the wheel disc, which can continue to rotate to the target aperture and achieve positioning. The limit structure on the first wheel disc 141 and the third wheel disc 143 is the same as the second wheel disc 142.

[0079] The type of treatment holes provided on the third turntable 143 is the same as that of the first turntable 141, both having wire mesh blades within the pore diameter structure, capable of physically cutting the passing fat. The middle turntable, i.e., the second turntable 142, has different pore diameters. The first turntable 141, the second turntable 142, and the third turntable 143 within the connecting device are respectively adjusted by the first roller 111, the second roller 112, and the third roller 113. When in use, two injection devices 2 (syringes) are respectively connected to the interface tubes 12 of the connecting device, and the push rods 21 of the syringes are alternately pushed, so that the fat is first cut by the wire mesh blades when passing through the connecting device, and then mechanically emulsified by being squeezed by the fluid through the pore diameter structure.

[0080] Connecting two syringes using the connecting device of the present invention can ensure complete sealing. The connecting device of the present invention has convertible pore diameters, capable of adjusting the pore diameter shape and quantity between the syringes, achieving the purpose of cutting and emulsifying fat to different degrees. When adjusting the pore diameter, there is no need to replace the connector, which can ensure that the fat in the syringe will not be exposed multiple times, reducing the risk of contamination to SVF during the operation process. At the same time, the pore diameter size and shape of the connecting mechanism can be customized according to the separation and preparation process of SVF.

[0081] That is, through the pore diameter design and transformation of the connector device, the Coleman fat is first divided into smaller fat blocks by the wire mesh in the wire mesh pore diameter, and then emulsified through the large pore diameter and small pore diameter of the connector, so as to reduce the resistance generated during the fat emulsification process, facilitate the operation; at the same time, reduce the damage to SVF, improve the SVF yield, and increase the cell concentration in SVF.

[0082] In addition, the embodiment of the present invention also provides a method for mechanically treating fat to obtain SVF, using the connecting device for mechanically treating fat to obtain SVF as described above, connecting the interface tube of the connecting device to an injection device;

[0083] Select a corresponding treatment hole on the rotating turntable to communicate with the interface tube;

[0084] Push the injection device so that the fat passes from one injection device through the connecting device for mechanically treating fat to obtain SVF and enters the injection device at the other end; obtain SVF from the treated fat.

[0085] For example: Transfer the fat into a syringe, connect the syringe to one end of the above-mentioned connecting device through a Luer interface, and then connect another empty syringe to the other end of the connecting device. Then, the holes corresponding to the turntable and the interface tube are, from left to right in sequence, the crosshair hole, the 2.4 mm hole, and the crosshair hole. Quickly inject the syringe to make the fat pass through the above-mentioned connecting device and enter the syringe at the other end. Then continue to operate the syringe to inject the fat back into the syringe at the initial end through the above-mentioned connecting device, which is recorded as 1 cycle. Repeat this process for multiple cycles. Thereby, the fat is cut and emulsified. Finally, SVF is obtained.

[0086] As Figure 2 shown, the connecting device has multiple rollers 11 that can be used to adjust the inner aperture of the connecting device. Preferably, 3 rollers are provided, namely the first roller 111, the second roller 112, and the third roller 113, which respectively correspond to adjusting the left, middle, and right turntables inside the connecting device; the two interface tubes 12 of the connecting device are connected to the syringes, and the connection method is threaded connection, or they can also be connected by other connection methods, such as sliding connection; the connecting device has a window 13 that can display the aperture number of the currently used turntable. Among them, the first window 131, the second window 132, and the third window 133 respectively correspond to adjusting the left, middle, and right turntables inside the connecting device. The left turntable, the first turntable 141, and the right turntable, the third turntable 143, have wire mesh blades inside their aperture structures, which can physically cut the passing fat. The middle turntable (the second turntable 142) has different aperture structures. The left, middle, and right turntables inside the connecting device are respectively adjusted by the rollers 11. When in use, connect the two syringes 2 to the interface tubes 12 of the connecting device respectively, and alternately push the push rods 21 of the syringes, so that when the fat passes through the connecting device, it is first cut by the wire mesh blade, and then mechanically emulsified by the fluid extrusion through the aperture structure. Figure 7 It is the structure of the left turntable (the first turntable 141) inside the connecting mechanism. There are limit grooves 1417 on the turntable for cooperating with the limiting mechanism (the limiting block 15) to limit the rotational displacement of the turntable, so that the holes of the turntable correspond to the two ends of the connecting mechanism. There are three through-hole structures on the first turntable 141, and different-shaped wire meshes are provided inside the through-holes for mechanically dividing the passing fat. The diameters of the three apertures are all 2 - 4 mm, preferably 2.4 mm. Among them, the first cutting hole 1411 is a well-shaped wire mesh, the second cutting hole 1412 is a star-shaped wire mesh, and the third cutting hole 1413 is a cross-shaped wire mesh. There is a prompt area on the outer edge of the turntable near the processing hole, where a character sticker marking the nearby processing hole can be pasted, and it can be displayed on the window of the connecting mechanism to prompt the number of the through-hole corresponding to the interface tube 12 of the connecting mechanism currently.

[0087] The roulette 14 is provided with a sliding limit mechanism, i.e., a limit block 15, which is composed of a spring, a ball and an outer housing. The outer housings of the sliding limit mechanisms at the leftmost and rightmost ends are fixed on the inner wall of the housing 1 of the connecting device, and the middle two sliding limit mechanisms are fixed on the shaft 16. The spring that is always in a compressed state provides a force for the ball, and at the same time the housing restricts the displacement of the ball so that only its tip leaks out of the housing and contacts the annular groove (limit ring groove). When the roulette rotates and the selectable aperture corresponds to the connection port of the interface tube, the ball will enter a deeper circular groove (limit groove 1442), realizing the displacement limit of the roulette component. When it is necessary to replace the selectable aperture, rotate the roller with a greater force, and the ball will slide out of the deeper circular groove 1442 into the annular groove. When the next selectable aperture corresponds to the interface tube, the ball will enter the next deeper circular groove. When the ball enters the circular groove, a mark of the aperture currently corresponding to the interface tube 12 will be displayed on the window 13. For example, when the limit ball (ball) rolls from the first limit ring groove 1417 into the circular groove of the first roulette 141, a mark of the aperture currently corresponding to the interface tube 12 will be displayed on the window 131, that is, the mark of the first prompt area 1414 corresponds to the first cutting hole 1411, the mark of the second prompt area 1415 corresponds to the second cutting hole 1412, and the mark of the third prompt area 1416 corresponds to the third cutting hole 1413.

[0088] The material of the connecting device of the two syringes can be metal, such as pure titanium, TC4 titanium alloy, 316L stainless steel, etc., or can be a polymer material, such as polyethylene, polypropylene, polyacetate, polymethyl methacrylate, etc. Each component can be made of different materials and then assembled together.

[0089] The material of the syringe should be able to withstand one or several sterilization methods such as high temperature and high pressure, radiation sterilization, ethylene oxide sterilization, etc. Figure 2 The interface tube 12 shown in the figure is the connection port between the connecting device and the syringe. The connection method shown here is a threaded interface, and it can also be connected by other connection methods, such as a sliding interface.

[0090] Only one form of the connecting device for connecting two syringes for preparing SVF is shown in the drawings, and there are only 3 selectable apertures and wire mesh shapes on each roulette structure. However, the present invention does not limit the shape and number of the apertures. For example, there can also be 2, 4 or other numbers of apertures on the roulette. The shape and size of the apertures are not limited to those shown in the figures.

[0091] Example 1: Use the connecting device of the present invention to prepare SVF.

[0092] Step 1: Centrifuge the fat obtained by liposuction under the conditions of 1200×g (1200 times the acceleration of gravity) for 3 minutes. Take the fat layer (Coleman fat) for further processing.

[0093] Step 2: Transfer the Coleman fat into a syringe, connect the syringe to one end of this device through a Luer interface, and then connect another empty syringe to the other end of this device.

[0094] Step 3: Adjust the holes corresponding to the connector wheel of this device and the interface to be "crosshair hole - 2.4mm hole - crosshair hole" from left to right in sequence. Quickly inject the syringe to make the Coleman fat pass through this connection mechanism and enter the syringe at the other end. Then continue to operate the syringe to inject the Coleman fat back to the syringe at the initial end through this connection mechanism, which is recorded as 1 cycle. Repeat this process for 10 - 30 cycles.

[0095] Step 4: Adjust the holes corresponding to the connector wheel of this device and the interface to be "star line hole - 2mm hole - star line hole" from left to right in sequence. According to the steps in 3, reciprocally inject the Coleman fat through the connector of this device for 10 - 30 cycles.

[0096] Step 5: Adjust the holes corresponding to the connector wheel of this device and the interface to be "grid line hole - 1.4mm hole - grid line hole" from left to right in sequence. According to step 3, reciprocally inject the Coleman fat through the connector of this device for 10 - 30 cycles.

[0097] Step 6: Centrifuge the emulsified fat, discard the oil phase and the liquid layer containing fragmented tissue components, and retain the component rich in SVF.

[0098] Example 2: Conduct a comparative study on the differences in SVF volume, cell density, and cell viability between the SVF obtained using the above preparation method and the SVF obtained only through one - sized pore treatment.

[0099] First, centrifuge the fat obtained by liposuction under the conditions of 1200×g for 3 minutes. Take the fat layer (Coleman fat) for further processing.

[0100] Method A: Use the method described in Example 1 for preparation. When performing steps 3, 4, and 5, the number of cycles of the Coleman fat passing through the connection mechanism is 10 times each.

[0101] Method B: Pass the Coleman fat only through the existing syringe - to - syringe connector (hole diameter is 2mm), and the number of cycles passing through the connector is 30 times.

[0102] Method C: Coleman fat is passed only through the existing syringe connectors (hole diameter: 1.4 mm) currently available, and the number of cycles through the connector is 30 times.

[0103] Subsequently, the processed fat is centrifuged respectively, and the oil phase and the liquid layer containing fragmented tissue components are discarded, and the SVF-rich components are retained, denoted as SVF A, SVF B, and SVF C respectively.

[0104] In the three treatment methods A, B, and C during the treatment process, the total number of times Coleman fat passes through the device described in this patent is the same, all 30 times. Each treatment method is carried out 3 times to obtain 3 sample data. The three types of SVF components A, B, and C obtained are analyzed as follows

[0105] Volume ratio: Calculate the percentage of the obtained SVF in Coleman fat, and draw a chart for comparison as Figure 10 shown.

[0106] Cell density: Take 1 ml samples from SVF A, SVF B, and SVF C respectively, mix them evenly with 1 ml of phosphate buffer solution, and then filter them through a cell sieve with a pore size of 100 microns. Take 20 microliters of the filtrate, mix it with 20 microliters of acridine orange / propidium iodide (AO / PI) dye for staining (the staining principle is that AO can pass through the intact cell membrane and embed in the nuclei of living and dead cells, showing green fluorescence; PI can only pass through the incomplete cell membrane, that is, the cell membrane of dead cells, and embed in the nuclei of all dead cells, showing red fluorescence), and use a fluorescence automatic cell counter to count the stained samples to obtain the cell viability and cell density. According to the detection results of the cell density, the original cell density in SVF A, SVF B, and SVF C is calculated. A typical fluorescence picture of AO / PI staining is as Figure 11 shown, and the cell viability of each group of SVF is as Figure 12 shown, and the cell density of each group of SVF is as Figure 13As shown in the figure. It can be seen from the above results that the percentage of the volume of SVF prepared by using the device (Method A) of the present invention in Coleman fat is between Method B and Method C. The reason is that in Method B, only a relatively large fixed pore size is used, and the emulsification degree of Coleman fat is limited, and some fat cells are not completely destroyed; in Method C, only a relatively small fixed pore size is used, and the shear force received by Coleman fat is always large, and the degree of destruction of fat cells is higher. This result is confirmed in the detection result of cell viability. The cell viability of SVF obtained by Method C is the lowest among these three methods, and the cell viability of SVF prepared by using the device (Method A) of the present invention is significantly improved. In addition, since the preparation of SVF by using the device (Method A) of the present invention causes less damage to cells (high cell viability), and at the same time has a higher emulsification degree of fat (greater volume compression), the obtained SVF cell density is the highest. That is, under the condition of the same volume of SVF, the SVF prepared by using the device (Method A) of the present invention has more effective cells.

[0107] Example 3: The SVF prepared in Example 1 was subjected to adherent culture, and the mesenchymal stem cells therein were directionally amplified.

[0108] The specific method is to wash the SVF prepared by using the method described in this patent (Example 1) with phosphate buffer solution (PBS), that is, mix 1 ml of the SVF with 9 ml of PBS and shake evenly, centrifuge at 300×g for 5 minutes, discard the supernatant in the centrifuge tube, and obtain a cell pellet.

[0109] The cell pellet was mixed with 5 ml of complete medium (α-MEM basal medium + 10% fetal bovine serum), placed in a T75 culture flask, and the liquid was spread over the entire culture flask. The culture flask was placed in an incubator for culture (incubator conditions: 5% CO2, humidity 95%, constant temperature at 37°C).

[0110] On the 3rd day of culture, 5 ml of complete medium was added to the culture flask.

[0111] On the 5th day of culture, it was observed that there were obvious adherent cells in the culture flask, and the confluence of the adherent cells reached more than 70%. The complete medium in the culture flask was aspirated and discarded, 3 ml of trypsin digestion solution was added thereto, and it was treated at 37°C for 2 minutes. It was observed under the microscope that the adherent cells had completely detached from the bottom surface, and then 7 ml of complete medium was added to terminate the digestion.

[0112] The mixed liquid of trypsin digestion solution, complete medium and adherent cells was collected into a centrifuge tube, centrifuged at 300×g for 5 minutes, the supernatant in the centrifuge tube was discarded, and a cell pellet was obtained.

[0113] Resuspend the cell pellet with complete medium and count the cell suspension using AO / PI dye. Add 3.75×10 5 cells to a new T75 culture flask and supplement the volume of complete medium to 10 ml.

[0114] On the 4th day of culture, obvious adherent cells were observed in the culture flask, and the confluence of the adherent cells reached more than 70%, then take a picture. The cell morphology is as Figure 14 shown, it can be seen that the cells adhere to the bottom of the culture flask and grow, and the cells show a spindle shape.

[0115] Aspirate and discard the complete medium in the culture flask, add 3 ml of trypsin digestion solution to it, treat at 37 °C for 2 minutes, observe under the microscope that the adherent cells have completely detached from the bottom surface, and then add 7 ml of complete medium to terminate the digestion.

[0116] Collect the mixed liquid of trypsin digestion solution, complete medium and adherent cells into a centrifuge tube, centrifuge at 300×g for 5 minutes, discard the supernatant in the centrifuge tube to obtain a cell pellet.

[0117] Use the MSC Marker flow cytometry kit (BD 562245) to stain the cells for surface markers according to the instructions, and then use a flow cytometer (BD Canto II) to analyze the cells, as Figure 15 、 Figure 16 、 Figure 17 、 Figure 18 、 Figure 19 、 Figure 20 、 Figure 21 、 Figure 22 、 Figure 23 、 Figure 24 、 Figure 25 、 Figure 26 shown, the analysis results of the surface markers of the cells are as follows:

[0118] Table 1 Data table of relevant detection of flow cytometry

[0119]

[0120] According to the detection results, it can be seen that there are cells capable of adherent culture in the SVF prepared by the method described in this application (Example 1), and the proportions of positive staining of CD73, CD90, and CD105 in the surface markers of these cells capable of adherent culture are all greater than 95%, and the results of positive staining of the surface markers of CD11b, CD19, CD34, CD45, and HLA-DR are all less than 2%. It can be explained that these cells are adipose-derived mesenchymal stem cells.

[0121] The present invention has the following beneficial effects:

[0122] Connecting two syringes using the connecting device of the present invention can ensure complete sealing. The connecting device of the present invention has a convertible aperture, which can adjust the aperture shape and quantity between syringes to achieve the purpose of cutting and emulsifying fat to different degrees. When adjusting the aperture, there is no need to replace the connector, which can ensure that the fat in the syringe will not be exposed multiple times, reducing the risk of contamination of SVF during the operation. At the same time, the aperture size and shape of the connecting device can be customized according to the separation and preparation process of SVF.

[0123] The above are only illustrative specific embodiments of the present invention and are not intended to limit the scope of the present invention. The components of the present invention can be combined with each other under the condition of no conflict. Any equivalent changes and modifications made by any person skilled in the art without departing from the concept and principle of the present invention shall fall within the scope of protection of the present invention.

Claims

1. A connecting device for mechanically treating fat to obtain SVF, characterized in that, Comprising: A housing (1), interface pipes (12) provided at both ends of the housing (1), a turntable (14) rotatably connected inside the housing (1), rollers (11) engaged with the turntable (14), a viewing window (13) opened on the housing (1), and a limiting block (15) fixed inside the housing (1); A plurality of turntables (14) are provided. The turntables (14) are rotatably connected inside the housing (1) through a shaft (16). The turntables (14) are provided with limiting structures and processing holes. The turntables (14) can be tightly connected through the processing holes. The processing holes penetrate through the turntables (14) and can communicate with the interface pipes (12); The limiting structure is provided on the side surface of the turntable (14) so that the limiting block (15) is in rolling contact with the limiting structure; The rollers (11) protrude from the housing (1). By rolling the rollers (11), the turntable (14) is rotated to communicate the processing holes with the interface pipes (12).

2. The connecting device for mechanically treating fat to obtain SVF according to claim 1, characterized in that There is a cavity between the turntables (14). Three turntables (14) are provided, including: a first turntable (141); a second turntable (142); a third turntable (143); Three rollers (11) are provided, including: a first roller (111), a second roller (112), and a third roller (113); The first roller (111) is engaged with the first turntable (141), the second roller (112) is engaged with the second turntable (142), and the third roller (113) is engaged with the third turntable (143).

3. The connecting device for obtaining SVF by mechanically treating fat according to claim 1, characterized in that, The limiting structure is provided on both sides of the turntable (14). The limiting structure includes: a limiting ring groove and a limiting groove; The limiting ring groove is opened on two side surfaces of the turntable (14) with the shaft (16) as the center; The limiting groove is hemispherical. The limiting groove is provided on the limiting ring groove. The diameter of the limiting groove is larger than the width of the limiting ring groove.

4. A connection device for obtaining SVF by mechanically treating fat according to claim 1, characterized in that, A limiting spring and a limiting ball are provided inside the limiting block (15); the limiting ball can protrude from the limiting block (15); The limiting spring and the limiting ball are in pressing contact, and the limiting ball is in rolling contact with the limiting structure on the turntable (14).

5. A connecting device for mechanically treating fat to obtain SVF according to claim 2, characterized in that, The processing holes of the first turntable (141) include: a first cutting hole (1411), a second cutting hole (1412), and a third cutting hole (1413); The processing holes of the second turntable (142) include: a first extrusion hole (1421), a second extrusion hole (1422), and a third extrusion hole (1423).

6. A connection device for mechanically processing fat to obtain SVF according to claim 5, characterized in that, A well-shaped wire mesh is provided in the first cutting hole (1411); a cross-shaped wire mesh is provided in the third cutting hole (1413); a plurality of silk threads are arranged radially in the second cutting hole (1412), and both ends of the silk threads are fixed on the inner wall of the second cutting hole (1412).

7. A connection device for mechanically treating fat to obtain SVF according to claim 6, characterized in that, The aperture diameter of the first extrusion hole (1421) is smaller than that of the second extrusion hole (1422), and the aperture diameter of the second extrusion hole (1422) is smaller than that of the third extrusion hole (1423).

8. A connecting device for mechanically treating fat to obtain SVF according to claim 2, characterized in that, The limiting block (15) includes: a first limiting block (151), a second limiting block (152), a third limiting block (153), and a fourth limiting block (154); The first limiting block (151) is fixed on the inner wall of the housing (1), and the first limiting block (151) is in rolling contact with the limiting structure on one side of the first disk (141); The second limiting block (152) is disposed in the cavity between the first disk (141) and the second disk (142), and the second limiting block (152) is fixed on the shaft (16). The second limiting block (152) is in rolling contact with the limiting structure on the other side of the first disk (141), and the second limiting block (152) is in rolling contact with the limiting structure on one side of the second disk (142); The third limiting block (153) is disposed in the cavity between the second disk (142) and the third disk (143), and the third limiting block (153) is fixed on the shaft (16). The third limiting block (153) is in rolling contact with the limiting structure on the other side of the second disk (142); the third limiting block (153) is in rolling contact with the limiting structure on one side of the third disk (143); The fourth limiting block (154) is fixed on the inner wall of the housing (1), and the fourth limiting block (154) is in rolling contact with the limiting structure on the other side of the third disk (143).

9. The connecting device for obtaining SVF by mechanically treating fat according to claim 1, characterized in that, The interface pipe (12) can be connected to the injection device (2).

10. A method for obtaining SVF by mechanically treating fat, characterized in that, Using the connecting device for mechanically treating fat to obtain SVF according to any one of claims 1-9, connect the interface pipe of the connecting device to the injection device; Rotate the disk to select the corresponding treatment hole to communicate with the interface pipe; Push the injection device so that the fat passes from one injection device through the connecting device for mechanically treating fat to obtain SVF and enters the injection device at the other end; Obtain SVF from the treated fat.

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