Skin-like chip simulation platform based on microfluidic technology

Abstract

The invention relates to a skin-like chip simulation platform based on a microfluidic technology. The skin-like chip simulation platform sequentially comprises a skin-like layer, a skin-like layer, a cytoplasm matrix simulation layer and an interstitial fluid circulation layer from top to bottom, a hole array is arranged on the imitated skin layer; an interstitial fluid enrichment cavity communicated with the hole array is formed in the simulated cortex, and the interstitial fluid enrichment cavity is provided with an enrichment outlet and an enrichment inlet; an interstitial fluid micro-channel is arranged in the cytoplasm matrix simulation layer, one end of the interstitial fluid micro-channel is communicated to the enrichment inlet, and the other end of the interstitial fluid micro-channel is communicated to the interstitial fluid flowing layer; an interstitial fluid storage cavity is formed in the interstitial fluid circulation layer and is provided with a storage cavity outlet and a storage cavity inlet; the device further comprises an interstitial fluid circulation channel. The skin-like chip simulation platform designed by the invention can accurately simulate the human skin environment, provides a simulation test platform for novel extraction electrode design, parameter optimization and safety evaluation of a counter ion electroosmosis technology, and remarkably reduces the preclinical research risk and cost of new technology development.

Description

TECHNICAL FIELD

[0001] The application relates to the field of microfluidic technology, and particularly relates to a skin-like chip simulation platform based on microfluidic technology. BACKGROUND

[0002] In the field of biomedical detection and health monitoring, interstitial fluid (ISF) extraction is an important technical direction for continuous health monitoring. As the liquid in the intercellular space of human tissues, interstitial fluid contains rich physiological and pathological information, and through analysis of the interstitial fluid, real-time monitoring of the health status of the human body and early warning of diseases can be realized. At present, the counter-ion electroosmosis method is a relatively mature non-invasive ISF extraction method, which extracts ions and small molecular substances in the interstitial fluid to the skin surface by applying an electric field on the skin surface and using electroosmosis, so as to realize the collection of the interstitial fluid. However, the optimization and verification of this technology highly depend on human experiments.

[0003] In traditional human verification experiments, there are problems of strict ethical norms. Human experiments need to follow strict ethical guidelines to ensure the safety and rights of the subjects, which to some extent limits the scale and repeatability of the experiments. At the same time, due to the large individual differences of the human body, the verification of the technical path faces many difficulties, and it is difficult to accurately evaluate the effectiveness and reliability of the technology.

[0004] In addition, most of the existing simulation verification platforms are used for sweat extraction and detection simulation, and the development of interstitial fluid simulation and detection platforms is still in the blank. For example, Tianjin University simulates human pores by using pig skin in related research, and adjusts the flow rate of the injection pump to simulate the flow rate of sweat actively exuding from the pig skin; the Institute of Nanotechnology and Nanobionics of the Chinese Academy of Sciences proposes a double-layer structure simulated by etching film and machining punching to simulate the device for active sweating. However, these schemes are all for sweat extraction simulation, and there are differences between interstitial fluid and sweat in physiological characteristics, extraction mechanism and other aspects, and the existing sweat simulation platform cannot meet the needs of optimization and verification of interstitial fluid extraction technology.

[0005] In summary, the optimization and verification of the counter-ion electroosmosis method for extracting interstitial fluid in the prior art face the problems of ethical constraints due to the dependence on human experiments and the lack of a special interstitial fluid simulation and detection platform. Therefore, there is an urgent need for a simulation platform that can efficiently simulate the process of human interstitial fluid extraction and reduce the dependence on human experiments, to solve the problems of ethical constraints and technical verification difficulties faced by related research institutions. SUMMARY

[0006] The application aims to solve the problem of lack of reliable simulation platform for interstitial fluid detection and dependence on biological experiments.

[0007] To achieve the above object, in a first aspect, the application provides a skin-like chip simulation platform based on microfluidic technology, which comprises, from top to bottom, a skin-mimicking layer, a simulated skin layer, a cytoplasm matrix simulation layer and an interstitial fluid flow layer. The skin-mimicking layer is provided with a hole array. The simulated skin layer is provided with an interstitial fluid enrichment cavity communicating with the hole array, the interstitial fluid enrichment cavity having an enrichment outlet and an enrichment inlet communicating with the cytoplasm matrix simulation layer. The cytoplasm matrix simulation layer is provided with an interstitial fluid microchannel, one end of the interstitial fluid microchannel communicating with the enrichment inlet and the other end communicating with the interstitial fluid flow layer. The interstitial fluid flow layer is provided with an interstitial fluid storage cavity, the interstitial fluid storage cavity having a storage cavity outlet communicating with the other end of the interstitial fluid microchannel and a storage cavity inlet. Further comprising an interstitial fluid circulation channel, one end of the interstitial fluid circulation channel communicating with the storage cavity inlet and the other end communicating with the enrichment outlet.

[0008] Further specifically, the skin-mimicking layer is made of a flexible material, which can be selected from any one of polyimide (PI), polyethylene terephthalate (PET), polymethyl methacrylate (PMMA), PDMS, pigskin and agarose gel.

[0009] Further specifically, the cross section of the interstitial fluid microchannel is circular or square.

[0010] Further specifically, the flow channel wall of the interstitial fluid microchannel is provided with a plurality of groove structures.

[0011] Further specifically, the plurality of groove structures have uneven distribution spacing or have different sizes.

[0012] Further specifically, the interstitial fluid flows in a Cassie state or a Wenzel state on the surface of the interstitial fluid microchannel.

[0013] Further specifically, the surface contact angle of the flow channel wall is 80°-115°.

[0014] In a second aspect, the application provides the use of the above skin-like chip simulation platform in simulating interstitial fluid extraction.

[0015] In a third aspect, the application provides an interstitial fluid extraction method based on counter-ion electroosmosis, which uses the above skin-like chip simulation platform, and the extraction method comprises the following steps: Making the interstitial fluid circulate in the skin-like chip simulation platform and the interstitial fluid circulation channel; An electrode sheet is arranged on the upper surface of the skin-mimicking layer, and a voltage is arranged on the electrode sheet, the interstitial fluid is extracted and partially exuded from the hole array.

[0016] Further specifically, the positive electrode sheet and the negative electrode sheet are both flexible electrodes.

[0017] The beneficial effects of the present application are as follows: The skin-mimicking chip simulation platform designed in the present application can simulate the epidermis layer, dermis layer and subcutaneous tissue layer of human skin, and through the internal microfluidic channel design, the flow environment of interstitial fluid in the intercellular space can be simulated, the precise simulation of human skin is realized, the dependence of interstitial fluid extraction experiment on real human experiment is greatly reduced, and the ethical and health problems existing in real human experiment are reduced. BRIEF DESCRIPTION OF DRAWINGS

[0018] Figure 1 is an exploded schematic view of the skin-mimicking chip simulation platform in the present application; Figure 2 is a schematic view of the skin-mimicking layer in the present application; Figure 3 is a schematic view of the skin-mimicking layer in the present application; Figure 4 is a schematic view of the skin-mimicking layer in the present application; Figure 5 is a schematic view of the cytoplasmic matrix simulation layer in the present application; Figure 6 is a schematic view of the cytoplasmic matrix simulation layer in the present application; Figure 7 is a schematic view of the cytoplasmic matrix simulation layer in the present application; Figure 8 is a schematic view of the cytoplasmic matrix simulation layer in the present application; Figure 9 is a schematic view of the cytoplasmic matrix simulation layer in the present application; Figure 10 is a schematic view of the cytoplasmic matrix simulation layer in the present application; Figure 11 is a schematic view of the cytoplasmic matrix simulation layer in the present application; Figure 12 is a schematic view of the cytoplasmic matrix simulation layer in the present application; Figure 13 is a schematic view of the cytoplasmic matrix simulation layer in the present application; Figure 14 is a schematic view of the cytoplasmic matrix simulation layer in the present application;

[0019] In the figure: 10, skin-like layer; 11, interstitial fluid through hole; 20, artificial skin layer; 21, enrichment inlet; 22, interstitial fluid enrichment cavity; 23, enrichment outlet; 30, cytoplasm matrix simulation layer; 31, interstitial fluid microchannel; 32, flow channel outlet; 33, flow channel inlet; 34, groove structure; 40, interstitial fluid flow layer; 41, storage cavity inlet; 42, interstitial fluid storage cavity; 43, storage cavity outlet; 50, interstitial fluid circulation channel; 60, electrode sheet. DETAILED DESCRIPTION

[0020] In order to make the purposes, technical solutions and advantages of the present application clearer, the specific embodiments of the present application are described clearly and completely below. Obviously, the described embodiments are part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor fall within the scope of protection of the present application.

[0021] In the description of the present application, it should be noted that the orientations or positional relationships indicated by the terms "center", "upper", "lower", "left", "right", "vertical", "horizontal", "inner", "outer" and the like are based on the orientations or positional relationships shown in the drawings, and are only for the convenience of describing the present application and simplifying the description, and therefore cannot be understood as indicating or implying that the devices or elements referred to must have a particular orientation, be constructed and operated in a particular orientation, and therefore cannot be understood as limiting the present application. In addition, the terms "first", "second", "third" are only for the purpose of description, and cannot be understood as indicating or implying relative importance.

[0022] In the description of the present application, it should be noted that unless otherwise explicitly specified and limited, the terms "mounting", "connection", "connection" should be understood broadly, for example, it can be fixedly connected, or it can be detachably connected, or integrally connected; it can be mechanically connected, or it can be electrically connected; it can be directly connected, or it can be indirectly connected through an intermediate medium; it can be the communication inside two elements. For those skilled in the art, the specific meaning of the above terms in the present application can be understood according to the specific circumstances. In addition, the technical features involved in the different embodiments of the present application described below can be combined with each other as long as they do not conflict with each other.

[0023] Interstitial fluid is an extracellular fluid located between cells and blood vessels. Interstitial fluid fills the intercellular space of tissues and is one of the important liquid environments for maintaining the normal function of cells. Interstitial fluid contains biomarkers such as glucose, and exchanges substances between the interstitial fluid and the plasma through the small pores of the capillary wall, so that the concentration of these markers in the interstitial fluid has a strong correlation with the concentration in the blood, thereby having potential value for biological information analysis.

[0024] The present application is to simulate the physiological environment of interstitial fluid in vivo, and to design a skin-like chip simulation platform based on microfluidic technology, which imitates the skin epidermis, dermis and subcutaneous tissue. As shown in Figure 1 The skin-like chip simulation platform includes four layers from top to bottom: skin-like layer 10, simulated skin layer 20, cytoplasmic matrix simulation layer 30 and interstitial fluid flow layer 40. The skin-like layer 10 is provided with a hole array. The simulated skin layer 20 is provided with an interstitial fluid enrichment cavity 22 communicating with the hole array, the interstitial fluid enrichment cavity 22 has an enrichment outlet 23 and an enrichment inlet 21 communicating with the cytoplasmic matrix simulation layer 30. The cytoplasmic matrix simulation layer 30 is provided with an interstitial fluid microchannel 31, one end of the interstitial fluid microchannel 31 communicates with the enrichment inlet 21, and the other end communicates with the interstitial fluid flow layer 40. The interstitial fluid flow layer 40 is provided with an interstitial fluid storage cavity 42, the interstitial fluid storage cavity 42 has a storage cavity outlet 43 communicating with the other end of the interstitial fluid microchannel 31 and a storage cavity inlet 41.

[0025] The skin-like chip simulation platform also includes an interstitial fluid circulation channel 50, one end of the interstitial fluid circulation channel 50 communicates with the storage cavity inlet 41, and the other end communicates with the enrichment outlet 23. The interstitial fluid circulation channel 50 can be provided with a peristaltic pump, starting the peristaltic pump can make the interstitial fluid of the interstitial fluid circulation channel 50 from the storage cavity inlet 41 into the interstitial fluid storage cavity 42, then from the storage cavity outlet 43 to the one end of the interstitial fluid microchannel 31 into the cytoplasmic matrix simulation layer 30, then from the other end of the interstitial fluid microchannel 31 to the enrichment inlet 21 into the interstitial fluid enrichment cavity 22, and finally from the enrichment outlet 23 back to the interstitial fluid circulation channel 50 and continuously circulate.

[0026] As shown in Figure 2 The skin-like layer 10 is provided with a hole array, the pore size of the interstitial fluid through hole 11 in the hole array is 1-100 μm, and the distance between adjacent interstitial fluid through holes 11 is 500-1000 μm. The skin-like layer 10 is made of flexible material, such as polyimide (PI), polyethylene terephthalate (PET), polymethyl methacrylate (PMMA), PDMS material, pigskin, agarose gel. Preferably, in order to make the skin-like layer 10 have softness and toughness closer to the epidermis of human skin, and in order to make the interstitial fluid through hole 11 have morphological stability closer to the skin sweat gland through hole, the skin-like layer 10 is made of polyimide material. The skin-like layer 10 designed in the present application can simulate the epidermis of human skin, and under the condition of no external force, the interstitial fluid will not flow out of the interstitial fluid through hole 11, and the internal interstitial fluid can be extracted from the interstitial fluid through hole 11 by using reverse iontophoresis or vacuum negative pressure / suction method on the skin-like layer 10.

[0027] As shown in Figure 3 and Figure 4As shown, the interstitial fluid enrichment cavity 22 is arranged in the simulation skin layer 20, and corresponds to the hole array region of the upper simulation skin layer 10. The interstitial fluid in the interstitial fluid enrichment cavity 22 can be extracted from the interstitial fluid through hole 11 under the driving force. The interstitial fluid enrichment cavity 22 is connected to the cytoplasm matrix simulation layer 30 below through the enrichment inlet 21, and is connected to the interstitial fluid circulation channel 50 through the enrichment outlet.

[0028] As shown in Figure 5 and Figure 6 , the cytoplasm matrix simulation layer 30 is provided with an interstitial fluid microchannel 31. One end of the interstitial fluid microchannel 31 is connected to the enrichment inlet 21, and the other end is connected to the interstitial fluid flow layer 40. The cross section of the interstitial fluid microchannel 31 can be circular or square. The interstitial fluid microchannel 31 can simulate the flow environment of interstitial fluid in the intercellular space. On the basis of the interstitial fluid circulation speed controlled by the peristaltic pump, the interstitial fluid microchannel 31 can regulate the enrichment speed of the interstitial fluid in the interstitial fluid enrichment cavity 22, thereby providing an experimental platform closer to the physiological environment for the counter-ion electrophoresis method for extracting interstitial fluid.

[0029] In further improvements of the present application, in order to further simulate the wetting heterogeneity of the extracellular matrix to the interstitial fluid, as shown in Figure 7 , the flow channel wall of the interstitial fluid microchannel 31 is provided with a groove structure 34. When the cross section of the interstitial fluid microchannel 31 is square, the preferred groove structure 34 is arranged at the bottom of the flow channel; when the cross section of the interstitial fluid microchannel 31 is circular, the groove structure 34 is arranged around the flow channel wall.

[0030] In some embodiments, several groove structures 34 have the same size, but the distance between adjacent groove structures 34 is different. From one end of the interstitial fluid microchannel 31 to the other end (i.e. the flow direction of the interstitial fluid), the distance between adjacent grooves gradually decreases in gradient. In other embodiments, adjacent groove structures 34 have the same distance, but the width of adjacent groove structures 34 (the length of the groove structure 34 is the width of the interstitial fluid microchannel 31) is different. The width of the groove structure 34 gradually decreases from one end of the interstitial fluid microchannel 31 to the other end. As shown in Figure 13As shown, the groove structure 34 designed in the present application can make the interstitial fluid have a changing wetting state, i.e. Cassie state (suspended state) or Wenzel state (penetrating state), when flowing in the interstitial fluid microchannel 31. In the flow direction of the interstitial fluid, the interstitial fluid is in the Wenzel state on the groove structure 34 at the end close to the inlet of the interstitial fluid microchannel 31, and the interstitial fluid is in the Cassie state on the groove structure 34 at the end close to the outlet of the interstitial fluid microchannel 31. By setting the non-uniform groove structure 34, the present application forms a surface topography with continuously changing contact ratio, so that the interstitial fluid realizes controllable conversion between Cassie state (contact angle 115°→85°) and Wenzel state (contact angle 80°→95°) on the surface of the interstitial fluid microchannel 31, and builds a wetting gradient from low hydrophobicity to high hydrophobicity (contact angle change range 80°→115°), accurately simulating the wetting heterogeneity of the extracellular matrix.

[0031] As shown in Figure 8 and Figure 9 , the interstitial fluid flow layer 40 is provided with an interstitial fluid storage cavity 42, the storage cavity inlet 41 of the interstitial fluid storage cavity 42 is connected with the interstitial fluid circulation channel 50, and the storage cavity outlet 43 of the interstitial fluid storage cavity 42 is connected with the interstitial fluid microchannel 31 of the upper cell matrix simulation layer 30.

[0032] The skin-like chip simulation platform designed in the present application can be prepared by using photosensitive resin materials (such as PDMS, PI, etc.). As shown in Figure 10 , the high-precision SLA printer is used to print the simulation skin layer 20, the cell matrix simulation layer 30 and the interstitial fluid flow layer 40 layer by layer using photosensitive resin; then the printed part is placed in an isopropanol solution for ultrasonic cleaning for 10 minutes to completely remove the unhardened resin residue; then the material is subjected to secondary curing treatment under ultraviolet light for 30 minutes to significantly improve the mechanical properties and stability of the material; finally, the skin-like chip simulation platform is formed by adhering the simulation skin layer 10 with a hole array and each layer of the printed part.

[0033] The device can extract interstitial fluid by using reverse ion electrophoresis or vacuum negative pressure / suction method (such as microneedle extraction method). As shown in Figure 11 and Figure 12 , the reverse ion electrophoresis method specifically includes: setting electrode sheets 60 on the surface of the simulation skin layer 10, connecting the two electrode sheets 60 to the power supply device, when the electrode sheets 60 have voltage, the positive electrode sheet 60 can attract negatively charged ions (such as Cl - ) and neutral molecules (such as water, glucose) in the interstitial fluid, and the negative electrode sheet 60 can attract positively charged ions (such as Na + ). Start the peristaltic pump to make the interstitial fluid continuously circulate in the device. The interstitial fluid is extracted to the surface of the simulation skin layer 10 under the action of the electric field force.

[0034] In further improvements of the present application, by setting the skin-like layer 10 as a convex shape with a certain curvature, and at the same time, the mass liquid enrichment cavity 22 has a bottom convex corresponding to the skin-like layer 10, the device can further simulate the real topography of the uneven skin surface. When using the counter-ion electrophoresis method to extract interstitial fluid, an electrode sheet 60 with the same convex structure as the surface of the device can be used. In some specific embodiments, the main material of the electrode sheet 60 is polyimide, and by depositing a Ti layer on the surface of the polyimide and then depositing a pt layer, a flexible electrode sheet with excellent mechanical flexibility and fatigue resistance is obtained.

[0035] In some specific embodiments, the pore array on the surface of the skin-like layer 10 has a pore diameter of 40 μm, and the distance between adjacent pores is 750 μm. By starting the peristaltic pump to circulate the interstitial fluid and applying a voltage of 5.0 V on the electrode sheet 60, the change curve of the interstitial fluid extraction amount and time is recorded as shown in Figure 14 The interstitial fluid extraction efficiency is about 34 μL / min. The present application can also obtain the desired interstitial fluid extraction efficiency by adjusting the voltage on the surface of the electrode sheet 60 and the circulation speed of the interstitial fluid, which can meet the needs of various interstitial fluid research experiments.

[0036] The present application realizes the whole system simulation from the skin barrier, extracellular matrix to the interstitial fluid dynamics, provides a simulation test platform for the design of new extraction electrodes, parameter optimization and safety evaluation of counter-ion electrophoresis technology, and significantly reduces the preclinical research risk and cost of new technology development.

[0037] The above describes the embodiments of the present application in detail, but the present application is not limited to the described embodiments. For those skilled in the art, various changes, modifications, replacements and variations can be made to these embodiments without departing from the principles and spirits of the present application, and still fall within the protection scope of the present application.

Claims

1. A skin-like chip simulation platform based on microfluidic technology, characterized in that, From top to bottom, it includes: a skin-like layer, a simulated dermis, a cytoplasmic matrix simulation layer, and a stromal fluid circulation layer; The simulated skin layer is provided with a pore array; The simulated cortex contains an interstitial fluid enrichment cavity with an array of interconnected pores. The interstitial fluid enrichment cavity has an enrichment outlet and an enrichment inlet connected to the simulated cytoplasmic matrix layer. The cytoplasmic matrix simulation layer is provided with interstitial fluid microchannels, one end of which is connected to the enrichment inlet and the other end is connected to the interstitial fluid flow layer. The interstitial fluid flow layer is provided with an interstitial fluid storage cavity, which has a storage cavity outlet and a storage cavity inlet that connect to the other end of the interstitial fluid microchannel. It also includes an interstitial fluid circulation channel, one end of which is connected to the reservoir inlet and the other end is connected to the enrichment outlet.

2. The skin-like chip simulation platform according to claim 1, characterized in that, The skin-like layer is made of a flexible material.

3. The skin-like chip simulation platform according to claim 1, characterized in that, The cross-section of the interstitial fluid microchannel is circular or square.

4. The skin-like chip simulation platform according to claim 3, characterized in that, The interstitial fluid microchannel has several groove structures on its channel wall.

5. The skin-like chip simulation platform according to claim 4, characterized in that, The groove structures have unevenly spaced distributions or have not identical dimensions.

6. The skin-like chip simulation platform according to claim 5, characterized in that, When the interstitial fluid flows on the surface of the interstitial fluid microchannel, it exhibits a Cassie state or a Wenzel state.

7. The skin-like chip simulation platform according to claim 5, characterized in that, The contact angle of the flow channel wall surface is 80°~115°.

8. The application of the skin-like chip simulation platform according to any one of claims 1 to 7 in the simulation of interstitial fluid extraction.

9. A method for extracting interstitial fluid based on reverse ion electroosmosis, characterized in that, The steps of using the skin-like chip simulation platform according to any one of claims 1 to 7 include: The interstitial fluid is circulated within the skin-like chip simulation platform and the interstitial fluid circulation channel. An electrode sheet is disposed on the upper surface of the skin-like layer, and a voltage is applied to the electrode sheet, so that the interstitial fluid is extracted and partially seeps out from the pore array.

10. The method for extracting interstitial fluid according to claim 9, characterized in that, The electrode sheet is a flexible electrode sheet.

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