CN103740590A - Three-dimensional cell-mechanical-gradient loading platform - Google Patents

Abstract

A three-dimensional cell mechanical gradient loading platform, including a mold composed of two cantilever beams or two cantilever beam arrays, and a three-dimensional tissue module or three-dimensional tissue module wrapped in cells is sandwiched between the two cantilever beams or the two cantilever beam arrays The array, the three-dimensional tissue module and the contact part of the cantilever beam are connected with 502 glue, the end of the cantilever beam is provided with a through hole, the threaded column passes through the through holes of two cantilever beams, and the threaded column on both sides of the through hole of a cantilever beam is provided The first nut and the second nut, the threaded posts on both sides of the through hole of the other cantilever beam are provided with the third nut and the fourth nut; or the ends of the two cantilever beams are respectively arranged on the one-dimensional mobile platform The control clip is connected, the control clip is connected with the motor; or the ends of multiple cantilever beam arrays are respectively connected and fixed by upper and lower two-layer flat plates, threaded columns and nuts. The present invention has the advantages of low cost, simple preparation, convenient use, controllable size, mechanical Adjustable gradient, high throughput and other advantages.

Description

A three-dimensional cell mechanics gradient loading platform

technical field

The invention relates to the technical field of tissue mechanics stimulation research, in particular to a three-dimensional cell mechanics gradient loading platform.

Background technique

In the body, cells are in the dynamic mechanical environment provided by the body and are subjected to various mechanical stimuli from the cellular microenvironment, in which stress (strain) has a significant impact on the structure, shape and function of the tissue. However, when culturing tissues in vitro, traditional cell culture methods and culture conditions do not include these important mechanical stimuli, and it is difficult to meet the growth requirements of three-dimensional tissues and organs, causing them to lose their normal shape and function. Appropriate stress (strain) stimulation is conducive to the construction of functional three-dimensional biological tissues in vitro. Studies have shown that mechanical stress (strain) has become an important external stimulus affecting cell structure and function, and appropriate mechanical stimulation conditions can induce and promote cell proliferation in vitro. Therefore, constructing an in vitro mechanical loading system to simulate mechanical stimuli in the in vivo environment is of great significance for constructing functional tissues in vitro.

Cell culture is the basic link in tissue engineering research. Body cells grow in a complex microenvironment. Conventional in vitro two-dimensional cell culture cannot provide the environmental conditions required for normal growth and development of tissues, and deviations in the growth environment will block growth signals. , chemical signals, and stress signal transmission, leading to apoptosis, loss of normal cell morphology or physiological function. Living cells in the body exist in a three-dimensional tissue structure. The three-dimensional space structure is not only the place for cell growth and metabolism, but also the basis for cell differentiation to form new tissues and organs with specific shapes and functions. The existing in vitro mechanical loading systems are almost all for mechanical loading of two-dimensional cultured cells. The response of cells in two-dimensional culture and in vivo environment to mechanical stimuli is very different, and the research results cannot reflect the real situation of cells growing in three-dimensional state in vivo. Moreover, the two-dimensional loading system cannot be used to construct three-dimensional tissue-organ systems. Therefore, the in vitro three-dimensional mechanical loading system has greater advantages than the two-dimensional mechanical loading system.

At present, the widely used cell loading devices mainly include rotary bioreactors, perfusion bioreactors, and basement membrane stretching loading devices. The key to the experimental study of cell mechanics lies in the design of loading methods to deform cells. At present, most of the existing mechanical loading studies are based on complex and expensive equipment (bioreactors and mechanical loading equipment), and mechanical loading is achieved by adjusting the amplitude, frequency, duration, and intensity of mechanical stress. However, these instruments can only achieve uniform mechanical stimulation, and in vivo tissues are often in a non-uniform extracellular matrix. Considering that the existing in vitro methods for studying mechanical stimulation of cells and tissues have many limitations and deficiencies, such as complex operation, high cost, the need to use precision instruments, uniform and single mechanical stimulation, and low throughput, it is urgent to establish a simpler and more effective, Reliable new mechanical loading method.

To sum up, it is very important for tissue engineering to study the microkinetic culture environment that simulates tissue growth under physiological conditions in vivo. Therefore, the development of a high-throughput three-dimensional cell mechanical mechanical loading platform with low cost, simple preparation, convenient use, controllable size, and adjustable mechanical gradient has become a new trend in the development of tissue mechanical stimulation research.

Contents of the invention

In order to overcome the above-mentioned shortcomings of the prior art, the purpose of the present invention is to provide a three-dimensional cell mechanical gradient loading platform, which is low in cost, simple in preparation, convenient in use, controllable in size, adjustable in mechanical gradient, and can be programmed arbitrarily.

In order to achieve the above object, the technical scheme that the present invention takes is:

A three-dimensional cell mechanical gradient loading platform, including a mold composed of two cantilever beams 7 or two arrays of cantilever beams 7, and a three-dimensional tissue module wrapped in cells is sandwiched between the two cantilever beams 7 or between the two cantilever beam 7 arrays 6 or a three-dimensional tissue module array, the contact parts of the three-dimensional tissue module 6 and the cantilever beam 7 are connected with 502 glue.

The end of described cantilever beam 7 is provided with through hole, and threaded column 5 passes through the through hole of two cantilever beams 7, and the threaded column 5 on both sides of the through hole of a cantilever beam 7 is provided with first nut 1 and the second A nut 2, a third nut 3 and a fourth nut 4 are arranged on the threaded columns 5 on both sides of the through hole of the other cantilever beam 7 .

The ends of the two cantilever beams 7 are respectively connected to the control clamps 8 provided on the one-dimensional moving platform 9 , and the two control clamps 8 are respectively connected to the first motor 10 and the second motor 11 .

The upper and lower ends of a cantilever beam array of the two cantilever beam arrays are respectively provided with a first flat plate 12 and a second flat plate 13, and the upper and lower ends of the other cantilever beam array are respectively provided with a third flat plate 14 and a fourth flat plate 15. , a through hole is provided in the middle of the plate, and a first nut 1 and a second nut 2 are arranged on the threaded columns 5 on both sides of the through hole of the first plate 12 and the second plate 13 clamping a cantilever beam 7 arrays. A third nut 3 and a fourth nut 4 are provided on the threaded columns 5 on both sides of the through holes of the third plate 14 and the fourth plate 15 of the other cantilever beam 7 array.

The cantilever beam 7 is made of organic or inorganic materials and has a specific geometric configuration. In the computer, CorelDRAW is used to draw the plane figure of the cantilever beam 7, and the two-dimensional figure is processed into the desired configuration by a laser cutting machine. The surface of the beam 7 in contact with the three-dimensional tissue module 6 wrapping the cells is straight, zigzag, wavy or convex.

The length of the cantilever beam 7 is 0.5-25 cm, the width is 0.1-50 mm, and the thickness is 0.1-15 mm.

The distance between the two cantilever beams 7 is 1-20 mm.

The two cantilever beams 7 are parallel or have an included angle, and the included angle ranges from 0 to 179°.

The cell-encapsulated three-dimensional tissue module 6 is composed of a mixture of biomaterials and cells, and the biomaterials are gel or porous scaffold materials.

The three-dimensional cell gradient mechanical loading experiment platform has the advantages of low cost, simple preparation, convenient use, controllable size, and adjustable mechanical gradient. Cutting harder organic or inorganic materials by simple equipment such as laser cutting machines, such as cutting polydimethylsiloxane (PDMS) or organic glass (PMMA) out of molds consistent with the geometric configuration, without complex , expensive technology; by constructing a through hole on the PDMS or PMMA bracket, the displacement between the two cantilever beams can be controlled by using the threaded column and the movement of the paired nuts inside and outside the cantilever beam, which overcomes the complicated operation of traditional mechanical loading equipment , high cost and other defects; use 502 glue to connect the mechanical loading device and the contact part of the three-dimensional tissue module wrapped in cells. On the one hand, it can prevent the three-dimensional cell mechanical loading module from sliding off after being stressed, and on the other hand, it can also facilitate mechanical pulling. By connecting the threaded column and the nut device of the two cantilever beams, the cantilever beam nut can be rotated outward to achieve the mechanical static tension effect, and the cantilever beam nut can be rotated inward to achieve the mechanical static compression effect; by adjusting the two cantilever beams The displacement change can adjust the tensile or compressive strain to meet different experimental requirements. In addition to static loading, dynamic mechanical loading of the device can also be achieved by using simple motors, one-dimensional moving platforms, and splints. Through the array design of the loading device, a high-throughput three-dimensional cell mechanical gradient loading device and an experimental platform can be formed. Due to the strong flexibility of computer graphics, laser cutting organic and inorganic materials and other substrates, the loading device can be cut into any shape, and a three-dimensional cell mechanical gradient loading device and experimental platform with adjustable mechanical gradient can be formed.

Description of drawings

FIG. 1 is a schematic diagram of the structure of the three-dimensional cytomechanical gradient static loading platform prepared in Example 1.

Fig. 2a is a schematic diagram of three-dimensional cell mechanical tensile gradient loading in Example 1, and Fig. 2b is a graph of internal strain distribution under tensile gradient loading.

Fig. 3a is a schematic diagram of three-dimensional cell mechanical compression gradient loading in Example 1, and Fig. 3b is a graph of internal strain distribution under compression gradient loading.

Fig. 4 is a schematic diagram of the structure of the three-dimensional cell mechanical gradient dynamic loading platform prepared in Example 2.

Fig. 5 is a schematic diagram of the structure of the high-throughput three-dimensional cell mechanical gradient static loading platform prepared in Example 3.

FIG. 6 a is a schematic structural diagram of the zigzag cantilever beam 7 in Embodiment 4, and FIG. 6 b is a curve diagram of the internal strain distribution of the zigzag cantilever beam 7 under loading.

Detailed ways

The present invention will be described in detail below in conjunction with the accompanying drawings and embodiments.

Example 1

Referring to Figure 1, use a computer to design two cantilever beams 7 in a rectangular shape (length 25mm, width 2.5mm, thickness 5mm), and a three-dimensional cell gradient mechanical loading mold configuration with a distance of 5mm between the two cantilever beams 7. The mechanical loading device consists of The organic material PMMA is cut to obtain a rectangular cantilever beam mold, and a through hole with the same diameter as the threaded column is punched at the ends of the two PMMA support cantilever beams 7 with a laser cutting machine. The diameter of the threaded column 5 is 1.5mm. A three-dimensional tissue module 6 wrapped in cells is sandwiched between the two cantilever beams 7, and the contact parts of the three-dimensional tissue module 6 and the cantilever beams 7 are bonded with 502 glue. Hole, diameter 1.5mm, threaded column 5 passes through the through hole of two cantilever beams 7, the threaded column 5 on both sides of the through hole of one cantilever beam 7 is provided with first nut 1 and second nut 2, and the other cantilever A third nut 3 and a fourth nut 4 are provided on the threaded posts 5 on both sides of the through hole of the beam 7 .

In the structure of Example 1, the nut 1 and the nut 2 are rotated and moved upwards, and the displacement is 0.5mm. Rotate and move the nut 3 and the nut 4 downward at the same time, and the displacement is 0.5 mm. As shown in FIG. 2 a , the three-dimensional tissue module 6 produces gradient tensile deformation. The internal strain distribution curve is shown in Figure 2b. The results show that there is a tensile stress gradient inside the three-dimensional tissue module 6, which proves that the device can generate a tensile strain gradient;

In the structure of Example 1, the nut 1 and the nut 2 are rotated downward, and the displacement is 0.25 mm. Rotate and move nut 3 and nut 4 upward at the same time, and the displacement is 0.25mm. As shown in Figure 3a, the three-dimensional tissue module 6 produces gradient compression deformation, and the internal strain distribution curve is shown in Figure 3b. The results show that there is Compressive stress gradients, demonstrating that the device can generate compressive strain gradients.

Example 2

Referring to Fig. 4, a three-dimensional cell mechanical gradient dynamic loading platform includes a mold composed of two cantilever beams 7, and a three-dimensional tissue module 6 wrapped in cells is sandwiched between the two cantilever beams 7, and the three-dimensional tissue module 6 and the cantilever beam 7 The contact parts are connected with 502 glue, the ends of the two cantilever beams 7 are respectively connected with the control clips 8 on the one-dimensional mobile platform 9, and the two control clips 8 are connected with the first motor 10 and the second motor 11 respectively. The computer can control the motor 10 and the motor 11 to move on a mobile platform, and the displacement is transmitted to the two cantilever beams 7 through the two control clamps 8 to generate a strain gradient.

Example 3

Referring to Figure 5, a high-throughput three-dimensional cell mechanical gradient static loading platform, a mold composed of two cantilever beams with 7 arrays, a three-dimensional tissue module array wrapped with cells is sandwiched between the cantilever beam arrays, the three-dimensional tissue module array and the cantilever beam array The contact parts are connected by 502 glue, and the upper and lower ends of one cantilever beam array of the two cantilever beam arrays are respectively provided with a first flat plate 12 and the second flat plate 13, and the upper and lower ends of the other cantilever beam array are respectively provided with a third flat plate 14 and the fourth flat plate 15, a through hole is provided in the middle of the flat plate, and the first nut 1 and the second nut 1 are arranged on the threaded posts 5 on both sides of the through hole of the first flat plate 12 and the second flat plate 13 clamping a cantilever beam 7 arrays. A third nut 3 and a fourth nut 4 are arranged on the threaded posts 5 on both sides of the through holes of the third flat plate 14 and the fourth flat plate 15 clamping another cantilever beam 7 array. The distance between the cantilever beam arrays is controlled by adjusting the positions of the nuts 1, 2, 3, and 4, so that the high-throughput three-dimensional tissue module array applies a mechanical gradient.

Example 4

Use a computer to design two cantilever beams 7 and the three-dimensional tissue module 6 that wraps the cells. Gradient mechanical loading mold configuration, the mechanical loading device is cut from the organic material PDMS, and a through hole with the same diameter as the threaded column is punched at the end of the two PDMS support cantilever beams with a laser cutting machine, and the threaded column (diameter 1.5mm). A cell-wrapped three-dimensional tissue module 6 is sandwiched between the two cantilever beams 7, and the contact parts of the cell-wrapped three-dimensional tissue module 6 and the cantilever beam 7 are bonded with 502 glue, and the two cantilever beams 7 are arranged in parallel. The end of 7 is provided with through hole, diameter 1.5mm, and threaded column 5 passes through the through hole of two cantilever beams 7, and the threaded column 5 on both sides of the through hole of a cantilever beam 7 is provided with first nut 1 and second nut. The cap 2 is provided with a third nut 3 and a fourth nut 4 on the threaded columns 5 on both sides of the through hole of the other cantilever beam 7 , as shown in FIG. 6 a .

Rotate and move nut 1 and nut 2 downward, and the displacement is 1mm. Rotate and move nut 3 and nut 4 upward at the same time, and the displacement is 1mm. The three-dimensional tissue module 6 produces gradient compression deformation. The internal strain distribution curve is shown in Fig. 6b, and the results show that the compressive stress gradient existing inside the three-dimensional tissue module 6 can be regulated by controlling the shape of the cantilever beam.

The working principle of the present invention is: illustrated with the structure of embodiment 1, by adjusting nut 1, nut 2, nut 3, nut 4 to adjust and control the distance between the ends of two cantilever beams 7, to form between the two cantilever beams The gradient distance, that is, the displacement of the gradient, when the ends of the two cantilever beams 7 move to the outside, the effect of mechanical stretching is achieved; when the ends of the cantilever beams 7 are moved to the inside, the effect of mechanical compression is achieved, because the wrapped cells between the two cantilever beams 7 The three-dimensional tissue module 6 produces gradient deformation after undergoing gradient displacement, and forms gradient strain inside, thereby affecting cell behavior.

The above-mentioned embodiments only express several implementation modes of the present invention, and the descriptions thereof are relatively specific and detailed, but should not be construed as limiting the patent scope of the present invention. It should be pointed out that those skilled in the art can make several modifications and improvements without departing from the concept of the present invention, and these all belong to the protection scope of the present invention.

Claims (9)

1. a three-dimensional cell mechanical gradient weighted platform, comprise the mould being formed by two socle girders (7) or two socle girders (7) array, it is characterized in that: between two socle girders (7) or between two socle girders (7) array, accompany three-dimensional tissue's module (6) or three-dimensional tissue's module array of parcel cell, three-dimensional tissue's module (6) is connected with 502 glue with the contact site of socle girder (7).

2. a kind of three-dimensional cell mechanical gradient weighted platform according to claim 1, it is characterized in that: the end of described socle girder (7) is provided with through hole, threaded column (5) is through the through hole of two socle girders (7), the threaded column (5) on the through hole both sides of a socle girder (7) is provided with the first nut (1) and the second nut (2), and the threaded column (5) on the through hole both sides of another socle girder (7) is provided with the 3rd nut (3) and the 4th nut (4).

3. a kind of three-dimensional cell mechanical gradient weighted platform according to claim 1, it is characterized in that: the end of described two socle girders (7) is connected with the control folder (8) being located on one-dimensional movement platform (9) respectively, two control folder (8) respectively with the first motors (10), the second motors (11) are connected.

4. a kind of three-dimensional cell mechanical gradient weighted platform according to claim 1, it is characterized in that: the upper and lower ends of a cantilever array of described two socle girders (7) array is respectively equipped with the first flat board (12), the second flat board (13), the upper and lower ends of another cantilever array is respectively equipped with the 3rd flat board (14) He Siping City plate (15), in the middle of dull and stereotyped, be provided with through hole, the threaded column (5) of clamping first flat board (12) of a socle girder (7) array and the through hole both sides of the second flat board (13) is provided with the first nut (1) and the second nut (2), the threaded column (5) of clamping the 3rd flat board (14) He Siping City plate (15) through hole both sides of another socle girder (7) array is provided with the 3rd nut (3) and the 4th nut (4).

5. a kind of three-dimensional cell mechanical gradient weighted platform according to claim 1, it is characterized in that: described socle girder (7) is to obtain having particular geometric configuration by the manufacture of organic or inorganic material, in computer, with CorelDRAW, draw socle girder (7) planar graph, X-Y scheme is become to required configuration by laser cutting machine by materials processing, and socle girder (7) is linear pattern, spination, wavy or convex with the surface that three-dimensional tissue's module (6) of parcel cell contacts.

6. a kind of three-dimensional cell mechanical gradient weighted platform according to claim 1, is characterized in that: the length 0.5～25cm of described socle girder (7), wide 0.1～50mm, thick 0.1～15mm.

7. a kind of three-dimensional cell mechanical gradient weighted platform according to claim 1, is characterized in that: the spacing of described two socle girders (7) is 1～20mm.

8. a kind of three-dimensional cell mechanical gradient weighted platform according to claim 1, is characterized in that: described two socle girders (7) are parallel or have angle, angular range is 0～179 °.

9. a kind of three-dimensional cell mechanical gradient weighted platform according to claim 1, is characterized in that: three-dimensional tissue's module (6) of described parcel cell is comprised of biomaterial and cytomixis, and biomaterial adopts gel or porous support materials.

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