CN114437932B - Microfluidic chip, system and using method of multi-placenta vascular anastomosis model - Google Patents

Abstract

The invention discloses a microfluidic chip, a microfluidic system and a using method of a multi-placenta vascular anastomosis model. The microfluidic chip comprises a chip body, wherein a mother fluid channel and two or more fetal fluid channels are arranged on the chip body, a porous membrane is arranged between the mother fluid channel and the fetal fluid channels, and a connecting channel which can be communicated or disconnected is arranged between the fetal fluid channels. The accurate and complete microminiaturization reconstruction is carried out on the structures of multiple placenta such as twin or triplet, so that the microstructure of two different chorionic twin placenta can be simulated, and a substitution model can be provided for the research of the current twin/multiple maternal barriers and the research of the potential pathophysiology mechanism of the special complications of twin; can simulate physiological flow conditions to realize the blood convection movement of two fetuses and a mother body of the twin; the flow conditions can be varied to simulate the villus space and abnormal blood flow conditions of the fetal capillaries, as well as variations before and after the administration of therapeutic measures.

Description

Microfluidic chip, system and application method of multiple placenta vascular anastomosis model

technical field

The invention relates to the field of microfluidic technology, in particular to a microfluidic chip, a system and a use method of a multiple-fetal placenta vascular anastomosis model.

Background technique

Twin pregnancy has always been the focus of perinatal medicine and fetal medicine research. The natural pregnancy process and outcomes are diverse, especially monochorionic diamniotic twins. Because the two fetuses share a placenta, there is a 1 There are more than one vascular anastomosis in the superficial part of the placenta, and there are often arterial-arterial and venous-venous anastomoses with two-way blood flow in the superficial placenta, and vascular anastomoses in the deep placenta often show unidirectional blood flow. When there is no compensatory vascular anastomosis with superficial two-way blood flow It will lead to hemodynamic imbalance, so there may be more and more serious complications, accompanied by an increase in perinatal morbidity and mortality. Research on twin/multiple placenta is of great significance for early diagnosis of twin/multiple pregnancy, evaluation of individualized treatment and prognosis. Conventional in vitro and ex vivo twin/multiple pregnancy models have been the mainstream for studying the underlying mechanisms of normal and abnormal placentas for many years, and some in vitro single-fetal models have emerged. The static nature and rapid aging of isolated placentas limit the study of their physiological relevance, so good twin/multiple animal placenta models have not yet been established.

Microfluidics is a technology that has attracted much attention in recent years. By precisely controlling and manipulating micro-scale fluids, especially sub-micron structures, the basic operations of biology, chemistry, and medical analysis can be integrated on micron-scale chips. Fluids exhibit special properties in microfluidic chips that differ from those at the macroscale, increasing the biological complexity and relevance of in vitro models. Due to its great potential in biology, chemistry, medicine and other fields, it has developed into a new multidisciplinary research field. At present, microfluidic organ chips have been developed to study the immune response of the maternal-fetal interface, placental uptake and transportation, drug transport, etc., but its application is limited to the single-fetal maternal-fetal barrier. The mechanism is still to be explored.

Contents of the invention

The present invention aims at at least solving the technical problems existing in the prior art, and particularly innovatively proposes a microfluidic chip, system and application method of a multi-fetal placenta vascular anastomosis model.

In order to achieve the above object of the present invention, according to the first aspect of the present invention, the present invention provides a microfluidic chip for a multi-fetal placental vascular anastomosis model, including a chip body, and the chip body is provided with a maternal fluid channel and two There are one or more fetal fluid channels, a porous membrane is provided between the maternal fluid channel and the fetal fluid channel, and a connecting channel that can be connected or disconnected is provided between the fetal fluid channels.

The above-mentioned technical solution: the fluid and material exchange between the mother and the fetus is realized through the porous membrane; the superficial two-way vascular anastomosis of the placenta and the one-way vascular anastomosis of the deep placenta between the fetuses are simulated through the connecting channel. Accurately and completely miniaturized reconstruction of the placenta structure of twins or triplets through the microfluidic chip can simulate the microstructure of two different chorionic twin placenta, which can be used for the current twin/multiple birth Provides a surrogate model for the study of the maternal-fetal barrier and the potential pathophysiological mechanisms of twin/multiple birth-specific complications; allows two circulations representing two fetuses of the same mother to be cultured on the same model to simulate the intrauterine environment of twins , allowing two types of placental structures to be cultured in the same device for comparison; physiological flow conditions can be simulated to achieve convective blood movement between twin fetuses and mother, convective blood flow between two fetuses in the same model and/or unidirectional Flow; the flow conditions can be changed to simulate the abnormal blood flow environment of the villi space and fetal capillaries and the changes before and after the application of therapeutic measures.

In a preferred embodiment, the connecting channel includes a first connecting channel for two-way or multi-directional communication between fetal fluid channels.

The above technical solution: Accurately simulate the two-way (between any two fetuses) or multi-way (between two or more fetuses) vascular anastomosis of superficial twin/multiple placenta through the first connection channel.

In a preferred embodiment, the first connecting channel is provided with a first valve assembly.

The above technical solution: it is convenient to control the connection or disconnection of the first connection channel, so as to adapt to various cases to be simulated.

In a preferred embodiment, the connecting channel includes a second connecting channel that is connected to the fetal fluid channel and only allows fluid to flow in one direction.

The above technical solution: accurately simulate the unidirectional anastomosis of the placenta depth through the second connection channel.

In a preferred embodiment, one or two second connecting channels with opposite flow directions are provided between the two fetal fluid channels.

The above-mentioned technical solution: the arterial-artery/vein-vein two-way one-way vascular anastomosis between two fetuses in twin/multiple fetuses can be accurately simulated through the second connecting channels with opposite flow directions. Only one second connection channel is used to simulate the fluid flow after the comprehensive effect of the arterial-artery/vein-vein two-way vascular anastomosis in the deep part of the placenta between the two fetuses, which simplifies the chip while satisfying the experimental effect of the one-way vascular anastomosis structure.

In a preferred embodiment, the second connecting channel is provided with a second valve assembly.

The above technical solution: it is convenient to control the connection or disconnection of the second connection channel, so as to adapt to various cases to be simulated.

In a preferred embodiment, the first end of the second connection channel is connected to the fetal fluid channel, and the second end of the second connection channel is provided with an input connection port.

The above technical solution: while ensuring the experimental effect, further simplifies the chip structure, and is easier to operate and implement.

In a preferred embodiment, the fetal fluid channel is provided with a cell pool.

The above technical solution: the cell pool is used for attaching fetal tissue cells, and links the metabolism of some fetal tissues with the dynamic changes of the maternal-fetal barrier, which is conducive to further simulating the pathophysiological state of the maternal-fetal.

In a preferred embodiment, all or part of the fetal fluid channel is located on the first side of the chip body, and the maternal fluid channel is located on the second side of the chip body.

The above technical solution: the fetal fluid channel and the maternal fluid channel are respectively arranged on the upper and lower sides of the chip body, which is conducive to reducing the volume of the chip and realizing miniaturization.

In a preferred embodiment, the fetal fluid channel and the maternal fluid channel are separated by a porous membrane in the overlapping region of the fetal fluid channel and the maternal fluid channel.

The above technical solution: the porous membrane in the overlapping area is used as the maternal-fetal exchange interface between the fetal fluid channel and the maternal fluid channel, thereby realizing the accurate simulation of the placental structure and function.

In a preferred embodiment, the size of the porous membrane in the overlapping region can be adjusted.

The above technical solution: the microfluidic chip can be used to simulate cases such as insufficient blood supply caused by the reduction of the restricted fetal placental area.

In a preferred embodiment, all or part of the fetal fluid channel is located on the first side of the chip body, and the maternal fluid channel is located on the first side of the chip body.

The above technical solution: the fetal fluid channel and the maternal fluid channel are located on the same side of the core body, which is convenient for processing and experimental operations, and has the same plane as the control, which can be used for the current research on the maternal-fetal barrier of twins/multiple births and special concomitant twins/multiple births. provide a surrogate model for the study of underlying pathophysiological mechanisms of the disease.

In a preferred embodiment, the fetal fluid channel and the maternal fluid channel are separated by a porous membrane in adjacent regions of the fetal fluid channel and the maternal fluid channel.

The above technical solution: the porous membrane in the adjacent area serves as the maternal-fetal exchange interface between the fetal fluid channel and the maternal fluid channel, realizing accurate simulation of placental structure and function.

In a preferred embodiment, the size of the porous membrane in the adjacent region can be adjusted.

The above technical solution: the microfluidic chip can be used to simulate cases such as insufficient blood supply caused by the reduction of the restricted fetal placental area.

In order to achieve the above object of the present invention, according to the second aspect of the present invention, the present invention provides a multiple-fetal placenta vascular anastomosis simulation system, including the microfluidic chip described in the first aspect of the present invention and at least 3 fluid pumps, A plurality of fluid pumps are used to pump fluid into the fetal fluid channel and the maternal fluid channel respectively.

The above technical solution: In addition to the beneficial technical effect of the microfluidic chip provided by the first aspect of the present invention, it also has the advantages of realizing fluid pumping through a fluid pump, which is convenient for operation, control and implementation.

In a preferred embodiment, the fluid is pumped into the input connection port of the second connection channel by a fluid pump.

The technical solution above: realizes pumping fluid signals of any flow rate into the fetal fluid channel connected to the second connecting channel, so as to simulate various cases of hemodynamic imbalance.

In order to achieve the above object of the present invention, according to the third aspect of the present invention, the present invention provides a method for using the multi-fetal placenta vascular anastomosis simulation system described in the second aspect of the present invention, including: The first type of cells is attached to the first side of the porous membrane of the porous membrane, and the first side is the side of the porous membrane facing the fetal fluid channel; the second type of cells is attached to the second side of the porous membrane in the overlapping area or the adjacent area, The second side is the side of the porous membrane facing the maternal fluid channel; attaching a third cell in the cell pool; setting the connection state of the connecting channel; pumping fluid into the fetal fluid channel and the maternal fluid channel; and/or liquids for observation and/or testing.

The technical solution above can provide a substitute model for the current research on the maternal-fetal barrier of twins/multiple births and the research on the potential pathophysiological mechanism of special complications of twins/multiple births, and is convenient for research and implementation. Among them, the first type of cells and the second type of cells are attached to both sides of the porous membrane. Because of its own microporous structure, it is used to simulate the chorion between the fetus and the mother to improve the accuracy of the model research.

In a preferred embodiment, it also includes: changing the flow rate of the fluid pumped into the fetal fluid channel and/or the maternal fluid channel; and/or, adjusting the size of the overlapping area or adjacent area; and/or, adjusting the pumping of the maternal fluid The substance composition and/or the substance composition ratio of the fluid of the channel.

The above technical solution: realize accurate simulation research of various cases.

Description of drawings

1 is a schematic diagram of the composition of the microfluidic chip of the multiple-fetal placental vascular anastomosis model in Example 1 of the present invention;

Fig. 2 is a schematic structural view of the first layer of the microfluidic chip of the multiple-fetal placenta vascular anastomosis model in Example 1 of the present invention;

Fig. 3 is another schematic structural view of the first layer of the microfluidic chip of the multiple-fetal placenta vascular anastomosis model in Example 1 of the present invention;

4 is a schematic structural diagram of the second layer of the microfluidic chip of the multiple-fetal placenta vascular anastomosis model in Example 1 of the present invention;

Fig. 5 is a cross-sectional schematic diagram of the overlapping area of the microfluidic chip of the multiple-fetal placenta vascular anastomosis model in Example 1 of the present invention;

Fig. 6 is a schematic structural view of a microfluidic chip of the multiple-fetal placenta vascular anastomosis model in Example 2 of the present invention;

Fig. 7 is another schematic structural view of the microfluidic chip of the multiple-fetal placenta vascular anastomosis model in Example 2 of the present invention.

Reference signs:

1 chip body; 101 first layer; 102 second layer; 103 maternal fluid channel; 104 fetal fluid channel; 105 first connecting channel; 106 second connecting channel; 107 cell pool; 2 porous membrane.

Detailed ways

Embodiments of the present invention are described in detail below, examples of which are shown in the drawings, wherein the same or similar reference numerals designate the same or similar elements or elements having the same or similar functions throughout. The embodiments described below by referring to the figures are exemplary only for explaining the present invention and should not be construed as limiting the present invention.

In describing the present invention, it should be understood that the terms "longitudinal", "transverse", "upper", "lower", "front", "rear", "left", "right", "vertical", The orientations or positional relationships indicated by "horizontal", "top", "bottom", "inner", "outer", etc. are based on the orientation or positional relationships shown in the drawings, and are only for the convenience of describing the present invention and simplifying the description, rather than Nothing indicating or implying that a referenced device or element must have a particular orientation, be constructed, and operate in a particular orientation should therefore not be construed as limiting the invention.

In the description of the present invention, unless otherwise specified and limited, it should be noted that the terms "installation", "connection" and "connection" should be understood in a broad sense, for example, it can be mechanical connection or electrical connection, or two The internal communication of each element may be directly connected or indirectly connected through an intermediary. Those skilled in the art can understand the specific meanings of the above terms according to specific situations.

Example 1

This embodiment discloses a microfluidic chip for a multiple-fetal placental vascular anastomosis model, as shown in Figure 1, including a chip body 1, on which a maternal fluid channel 103 and two or more fetal fluid channels are arranged. 104 , a porous membrane 2 is provided between the maternal fluid channel 103 and the fetal fluid channel 104 , and a connecting channel that can be connected or disconnected is provided between the fetal fluid channels 104 .

In this embodiment, when there are two fetal fluid channels 104, it is used to simulate the twin placental vascular anastomosis model, when there are three fetal fluid channels 104, it is used to simulate the placental vascular anastomosis model of triplets, and so on to simulate multiple Fetoplacental vascular anastomosis model.

In this embodiment, the porous membrane 2 is preferably but not limited to porous polycarbonate membrane, polyester membrane, polytetrachlorethylene membrane, elastomeric membrane (for example, polydimethylsiloxane, PDMS, polyurethane or extracellular one of the matrix membranes). Through the porous membrane 2, fluid communication and exchange are realized in the fetal fluid channel 104 and the maternal fluid channel 103, especially in overlapping or adjacent areas of the two. The porous membrane 2 forms a semi-permeable barrier in the overlapping and adjacent areas. The first layer 101 and the second layer 102 are preferably, but not limited to, substrates made of polydimethylsiloxane (PDMS).

In this embodiment, the chip body 1 includes at least two layers, such as the first layer 101 and the second layer 102 in FIG. The second side of the body 1. All or part of the fetal fluid channel 104 is provided on the first side of the chip body 1 , specifically, it can be located inside the first layer 101 or on the upper surface or the lower surface. The maternal fluid channel 103 is provided on the second side of the chip body 1, specifically, it can be located inside or on the upper or lower surface of the second layer 102, and the fetal fluid channel 104 and the maternal fluid channel 103 have an overlapping area in the vertical direction. FIG. 5 shows a schematic cross-sectional structural diagram of the overlapping area. In FIG. 5 , the maternal fluid channel 103 is set on the upper surface of the second layer 102 , and the fetal fluid channel 104 is set on the lower surface of the first layer 101 .

In this embodiment, the shape of the fetal fluid channel 104 is preferably, but not limited to, L-shaped or back-shaped or C-shaped. Preferably, the shapes of the multiple fetal fluid channels 104 are consistent, so as to better simulate the actual situation. The shape of the parent fluid channel 103 can be set as a straight line as shown in FIG. 4 , or other shapes, such as a "several" shape, a C shape, and the like. Both ends of the fetal fluid channel 104 and the maternal fluid channel 103 are provided with connection ports, the connection port at one end can be used as an inlet, and the connection port at the other end can be used as an outlet. The inlet can be connected to a fluid pump, and the outlet can be connected to a liquid recovery device or other containers.

In this embodiment, the porous membrane 2 can be arranged between the first layer 101 and the second layer 102, and the porous membrane 2 is arranged at least in the overlapping area of the fetal fluid channel 104 and the maternal fluid channel 103 in the up-down direction, and the fetal fluid channel 104 It is separated from the parent fluid channel 103 by the porous membrane 2.

In this embodiment, the size of the porous membrane 2 may be larger than the overlapping area, such as the same size as the lower surface of the first layer 101 or the upper surface of the second layer 102 .

In this embodiment, further preferably, the size of the porous membrane 2 in the overlapping area of the fetal fluid channel 104 and the maternal fluid channel 103 in the vertical direction can be adjusted. The adjustment method is preferably but not limited to reducing the size of the porous film 2 in the overlapping area by pasting or coating a layer of impermeable film on the porous film 2 in the original overlapping area. The barrier film is preferably but not limited to plastic film or paint. The adjustment method can also be to translate the first layer 101 or the second layer 102 to change the size of the overlapping area of the fetal fluid channel 104 and the maternal fluid channel 103 in the vertical direction, and then change the size of the porous membrane 2 in the overlapping area.

In this embodiment, the connecting channel includes a first connecting channel 105 for two-way or multi-directional communication between the fetal fluid channels 104 . When the microfluidic chip is used to simulate twin placental vascular anastomosis, the first connecting channel 105 can be a bidirectional channel as shown in Figure 2 and Figure 3, preferably, a first valve assembly is set on the channel to control the communication of the channel or disconnected, the first valve assembly can be an existing manual valve or an electric valve. When the microfluidic chip is used to simulate triplet placental vascular anastomosis, the first connecting channel 105 can be a multi-directional communication channel between three or more fetal fluid channels 104. Preferably, the multi-directional communication channel includes A plurality of connecting parts and converging parts connected simultaneously with the plurality of connecting parts, the plurality of connecting parts are respectively connected with a plurality of fetal fluid channels 104 in one-to-one correspondence, preferably, a first valve assembly is provided at each connecting part for controlling The connection part is connected to or disconnected from the fetal fluid channel 104, and when the first valve components on all the connection parts are opened, the fetal fluid channel 104 connected to the connection part can communicate with other fetal fluid channels 104.

In this embodiment, a schematic diagram of a microchannel structure of the first layer 101 is shown in FIG. 2 . In FIG. There are two second connecting channels 106, wherein one second connecting channel 106 is for the right fetal fluid channel 104 to flow to the left fetal fluid channel 104, and the other second connecting channel 106 is for the left fetal fluid channel 104 to flow to the right fetal fluid Channel 104. The structure shown in Figure 2 fully simulates the two-way anastomosis of arteries-arteries/veins-veins in the placental superficial anastomosis of two fetuses in the actual twin placental anastomosis, which is different from the blood pressure and flow velocity between the two fetuses in reality. As a result, the blood flow transport phenomena are very close to each other.

In this embodiment, a schematic diagram of another microfluidic channel structure of the first layer 101 is shown in Figure 3. In Figure 3, the model shown in Figure 2 is improved according to the test results, in order to facilitate processing and facilitate Test, the second connecting channel 106 is arranged on the side of one of the fetal fluid channels 104, at this time, the first end of the second connecting channel 106 is connected with the fetal fluid channel 104 connected to the second connecting channel 106, and the second connecting channel The second end of 106 is provided with an input connection port, and a fluid pump is arranged at the input connection port to pump fluid into the second connection channel 106 to simulate the phenomenon that the connected fetal fluid channel 104 receives blood transfusion from another fetal fluid channel 104 .

In this embodiment, in order to observe the influence of hemodynamic imbalance between fetuses on fetal tissues, and the influence of changes on the maternal side on fetal tissues, preferably, as shown in Figure 1, Figure 2, and Figure 3, the fetus A cell pool 107 is provided on the fluid channel 104, and the cell pool is used for attaching fetal tissue cells. The tissue cells are preferably but not limited to embryonic stem cells, liver cells, pancreatic cells, or nerve cells. The shape of the cell pool 107 is preferably, but not limited to, oval, circular or square.

In this embodiment, preferably, an intermediate layer (intermediate layer is not shown) is further provided between the first layer 101 and the second layer 102, and the upper surface of the first layer 101 is used as the first side of the chip body 1 , the upper surface of the first layer 101 is provided with a fetal fluid channel 104, the upper surface of the second layer 102 is used as the second side of the chip body 1, and the upper surface of the second layer 102 is provided with a maternal fluid channel 103. The middle layer part corresponding to the overlapping area of the fetal fluid channel 104 and the maternal fluid channel 103 is provided with a through hole connecting the fetal fluid channel 104 and the maternal fluid channel 103, and the two ends of the through hole are covered with a porous membrane 2, through which two layers of porous membrane 2 The fetal fluid channel 104 and maternal fluid channel 103 are separated in the overlapping region.

Example 2

This embodiment provides a microfluidic chip for a multi-fetal placental vascular anastomosis model. As shown in Figures 6 and 7, the microfluidic chip includes a chip body 1, and the chip body 1 is provided with a maternal fluid channel 103 and two or more More than two fetal fluid channels 104, a porous membrane 2 is provided between the maternal fluid channel 103 and the fetal fluid channel 104, and a connecting channel that can be connected or disconnected is provided between the fetal fluid channels 104. The connecting channels include a first connecting channel 105 that enables two-way or multi-directional communication between the fetal fluid channels 104 , and a second connecting channel 106 that is connected to the fetal fluid channel 104 and allows only one-way flow of fluid.

The difference between this embodiment and Embodiment 1 is: First, this embodiment sets all or part of the fluid channel 104 and the matrix fluid channel 103 on the same side (such as the first side) of the chip body 1, as shown in Figures 6 and 7 The second is that the fetal fluid channel 104 on the same side of the chip body 1 and the adjacent area of the maternal fluid channel 103 are separated by the porous membrane 2 from the fetal fluid channel 104 and the maternal fluid channel 103 on this side, as shown in Figure 6 and Figure 7 shown.

In this embodiment, the chip body 1 is preferably, but not limited to, a polydimethylsiloxane (PDMS) material substrate. In the adjacent area of the fetal fluid channel 104 and the maternal fluid channel 103 , the fetal fluid channel 104 and the maternal fluid channel 103 communicate with each other and are separated by the porous membrane 2 . When the fetal fluid channel 104 and the maternal fluid channel 103 are close but do not communicate with each other in the adjacent area, a through hole can be opened to connect the two micro flow channels, and a porous membrane 2 is provided at both ends of the through hole to carry out fluid and material. exchange.

In this embodiment, the size of the porous membrane 2 in the adjacent area can be adjusted, which can be realized by coating or sticking a barrier film on the porous membrane 2 in the adjacent area.

In this embodiment, the fetal fluid channel 104 is provided with a cell pool 107 . One cell pool 107 is provided for each fetal fluid channel 104 .

In this embodiment, preferably, the first connecting channel 105 is provided with a first valve assembly, and the second connecting channel 106 is provided with a second valve assembly.

In this embodiment, as shown in FIG. 6 , preferably, one or two second connecting channels 106 with opposite flow directions are provided between the two fetal fluid channels 104 . Further preferably, a second valve assembly is provided on each second connecting channel 106 .

In this embodiment, as shown in FIG. 7 , preferably, the first end of the second connection channel 106 is connected to the fetal fluid channel 104 , and the second end of the second connection channel 106 is provided with an input connection port. The input connection port is connected with a fluid pump of the peripheral device, and is used for pumping fluid into the fetal fluid channel 104 connected with the second connection channel 106 to simulate hemodynamic imbalance between fetuses.

In the first application scenario of the microfluidic chip provided by the present invention, it is used to simulate the vascular anastomosis of monochorionic diamniotic twin placenta, and the first connecting channel 105 and the second connecting channel 106 are opened, respectively in the fetal fluid channel. 104 and parent fluid channel 103 to pump fluid. The second connecting channel 106 is used to simulate the one-way vascular anastomosis in the adjacent villi leaflets in the deep part of the placenta. The first connecting channel 105 is used for simulating the superficial placenta, which usually also has two-way arterial-arterial/vein-venous anastomosis. The first valve assembly is opened, and the second valve assembly on the second connecting channel 106 in two opposite directions is opened, and the first fetal fluid channel 104 representing the cord blood flow of the first fetus is connected with the cord blood flow representing the second fetus. The second fetal fluid channel 104 is set at different flow rates, using a pulse pump or a non-pulse pump to pump the culture medium, and a fluid pressure difference is generated between the two micro channels representing the two fetuses to promote blood flow through the anastomotic channel, simulating the presence of Placental model of vascular anastomosis.

In the second application scenario of the microfluidic chip provided by the present invention, it is used to simulate vascular anastomosis of two fetuses and placentas in dichorionic diamniotic sac twins. As a placenta, the blood circulation of the two fetuses is also relatively independent, the first connecting channel 105 and the second connecting channel 106 are closed, and fluid is pumped into the fetal fluid channel 104 and the maternal fluid channel 103 respectively. Close the first valve assembly and the second valve assembly on the second connecting channel 106 in opposite directions, the first fetal fluid channel 104 representing the first fetal umbilical cord blood flow and the second fetal fluid channel 104 representing the second fetal umbilical cord blood flow The flow rate of the culture medium in the two fetal fluid channels 104 is consistent, the blood circulation of the two fetuses is relatively independent, and the hemodynamics is relatively stable.

The microfluidic chip provided in Example 1 and Implementation 2 can simulate a model in which there is vascular anastomosis and a model in which each part (fetus) circulates relatively independently. It can be used to study the underlying pathophysiological mechanism under the special structure of twin placenta.

Embodiment 3 This embodiment discloses a multiple-fetal placental vascular anastomosis simulation system, including the microfluidic chip provided in Embodiment 1 or Embodiment 2 and at least 3 fluid pumps, using multiple fluid pumps to provide fetal fluid channels 104 and The parent fluid channel 103 pumps fluid.

In this embodiment, preferably, fluid is pumped into the input connection port of the second connection channel 106 through a separately provided fluid pump. The fluid pump is not the fluid pump of the fetal fluid channel 104 and the maternal fluid channel 103, but is a separate fluid pump.

In this embodiment, the fluid pumps are respectively connected to the input connection ports of the fetal fluid channel 104 , the maternal fluid channel 103 , and the second connection channel 106 to pump fluid into the channels. The fluid pump is preferably but not limited to a two-way continuous syringe pump model WH-BCSP-22. The output port of the pump is connected to the inlet of the fluid channel, and the input port of the pump is connected to the outlet of the fluid channel to realize fluid circulation.

Example 4

This embodiment discloses a method for using the multifetal placenta vascular anastomosis simulation system provided in Embodiment 3, including:

Step A, attaching the first type of cells to the first side of the porous membrane 2 in the overlapping area or the adjacent area, the first side being the side of the porous membrane facing the fetal fluid channel 104 . The first cell is preferably, but not limited to, a fetal capillary endothelial cell, such as umbilical vein endothelial cells ("HUVECs"), primary human placental villous endothelial cells ("HPVECs"), primary human endothelial cells isolated from a fetus, Fetal-derived transformed human endothelial cells, stem cell-derived endothelial cells. In practice, a device such as a straw can be used to absorb the liquid containing the first cell, and the liquid is input into the fetal fluid channel 104 (preferably but not limited to input from the inlet of the fetal fluid channel 104), and let it flow through the first cell of the porous membrane 2. side and let stand, due to the porous structure of the porous membrane 2, the first type of cells will attach to the first side of the porous membrane 2.

Step B, attaching the second type of cells on the second side of the porous membrane 2 in the overlapping area or the adjacent area, the second side is the side of the porous membrane facing the parent fluid channel 103 . The second type of cells represents trophoblast cells, preferably but not limited to chorionic (BeWo) cells, BeWo b30 clone cells, HTR8/SVneo trophoblast cells, choriocarcinoma (JEG3) cells, primary human trophoblast cells, stem cell-derived Trophoblasts, transformed human trophoblast cells. Artificial or naturally induced cases may also be included in the first cell and the second cell, the naturally induced pathology may be from a diseased placenta. Similarly, the liquid containing the second type of cells is input into the matrix fluid channel 103, the liquid is allowed to flow through the second side of the porous membrane 2 and left to stand, due to the porous structure of the porous membrane 2, the second type of cells will attach to the porous membrane 2 on the second side of the membrane 2.

Step C, attaching a third type of cells in the cell pool 107 . The third type of cells is fetal tissue cells, preferably but not limited to embryonic stem cells, liver cells, pancreatic cells, and nerve cells. The liquid containing the third cell is input into the cell pool 107 and left still. For the convenience of importing the third cell well attached to the cell pool 107, preferably, the cell pool 107 can be arranged close to the fetal fluid channel 104, so that the liquid is input into the cell pool 107 from the outlet of the fetal fluid channel 104.

Step D, setting the connection state of the connection channel. Preferably, the connection state of the connection channel is set according to the case to be simulated, for example, when simulating twin-twin transfusion syndrome, the first connection channel 105 is closed and the second connection channel 106 is opened. For example, when simulating reverse arterial perfusion syndrome of twins, the first connecting channel 105 and the second connecting channel 106 are opened. Fluid is pumped into the fetal fluid channel 104 and the maternal fluid channel 103; the fluid may be pumped by a fluid pump, preferably but not limited to culture medium.

Step E, observe and/or detect the third cell and/or liquid in the cell pool 107 .

In this embodiment, preferably, the flow rate of fluid pumped into the fetal fluid channel 104 and/or maternal fluid channel 103 is varied. The flow rate of the fluid pumped into the fetal fluid channel 104 and/or the maternal fluid channel 103 can be changed according to the simulated case. For example, when simulating the twin fetal arterial reverse perfusion syndrome, the flow rate of the fluid in one fetal fluid channel 104 should be greater than that of the other fetus The flow rate of the fluid in the fluid channel 104 . As another example, when simulating hypertension during pregnancy, the flow rate of the fluid in the maternal fluid channel 103 is increased.

In this embodiment, preferably, the size of the overlapping area or the adjacent area is adjusted. The size of the overlapping area or the adjacent area can be adjusted according to the case to be simulated. For example, when simulating selective intrauterine growth restriction, the size of the porous membrane 2 in the overlapping area or the adjacent area can be reduced.

In this embodiment, preferably, the substance composition and/or substance composition ratio of the fluid pumped into the mother fluid channel 103 is adjusted, and the substance composition and/or substance composition of the fluid pumped into the mother fluid channel 103 can be adjusted according to the case to be simulated Proportion. The material components are preferably but not limited to glucose, fatty acid, protein, bile acid concentration.

In the first application scenario of the multiple-fetal placental vascular anastomosis simulation system provided by the present invention, it is used to simulate the unique complications of monochorionic diamniotic twins, the first valve assembly on the first connecting channel 105, and the two fetal fluid channels Two unidirectional second connecting passages 106 are provided between 104, and the fluid directions of the two second connecting passages 106 are opposite, and each second connecting passage 106 is provided with a second valve assembly. Specifically:

1. Twin to twin transfusion syndrome (TTTs), the first valve assembly is closed, the second valve assembly on the second connecting channel 106 of the first one-way anastomosis is opened, and the other one-way anastomosis in the opposite direction The second valve assembly on the second connection passage 106 of the first fetal fluid passage 104 is set to be greater than the flow rate of the second fetal fluid passage 104, and the culture medium is represented by the first fetal fluid passage 104 per unit time. One fetus (donor baby) flows to the second fetus (recipient baby) represented by the second fetal fluid channel 104, and the hemodynamics of the two fetuses is unbalanced.

2. Twin reversed arterial perfusion sequence (TRAPs), the first valve assembly on the first connecting channel 105 of the two-way anastomosis channel is opened, and the first valve assembly on the second connecting channel 106 of the first one-way anastomosis The second valve assembly is opened, and the second valve assembly on the second connecting channel 106 of the opposite one-way anastomosis is opened, and the medium flow rate of the first fetal fluid channel 104 on behalf of the first fetus remains unchanged, representing the second The inlet and outlet of the second fetal fluid channel 104 of the first fetus are directly matched, and no external fluid pump is connected. The fluid pump representing the first fetus drives the blood circulation of the two microfluidic channels representing the two fetuses. Fluid dynamic imbalance.

3. Selective intrauterine growth restriction (sIUGR), on the basis of simulating the physiological structure of uncomplicated twin placenta, the first valve component is closed, the second valve component of the first second connecting channel is opened, and the second valve component is opened. The second valve assemblies of the two second connecting channels are closed, and the culture medium is transferred from the first fetus (unrestricted fetus) represented by the first fetal fluid channel 104 to the second fetal fluid channel 104 represented by the second fetal fluid channel 104 per unit time. Two fetuses (restricted fetus) flow and reduce the area where the microchannel representing the second fetus (restricted fetus) overlaps the area of maternal fluid channel 103 representing the maternal face (which can change the semipermeability of the lateral membrane of the restricted fetus Area or microchannel width) to simulate the reduced fetal placental area, resulting in insufficient blood supply, and hemodynamic imbalance between the two fetuses.

4. For single intrauterine fetal death (sIUFD), on the basis of simulating the physiological structure of uncomplicated twin placenta, the second valve component of the first second connection channel is opened, and the second second connection channel is opened. The second valve assembly of the channel is opened, the medium flow rate of the first fetal fluid channel 104 representing the first fetus remains unchanged, and the second fetal fluid channel 104 representing the second fetus closes the fluid pump to stop the blood circulation, Hemodynamic imbalance between the two fetuses.

In the first application scenario of the multi-fetal placental vascular anastomosis simulation system provided by the present invention, it is used to simulate pregnancy complications during twin pregnancy, specifically:

1. Maternal hyperglycemia: On the basis of simulating the physiological structure of uncomplicated twin placenta, increase the glucose concentration in the culture medium pumped by the maternal fluid channel 103 representing the mother.

2. Maternal hyperlipidemia: On the basis of simulating the physiological structure of uncomplicated twin placenta, increase the concentration of fatty acids in the medium pumped by the maternal fluid channel 103 representing the mother.

3. Maternal nutrition deficiency: On the basis of simulating the physiological structure of uncomplicated twin placenta, reduce the concentration of glucose, fatty acid, protein and other nutrients in the culture medium pumped by the maternal fluid channel 103 representing the mother.

4. Hypertension during pregnancy (Hypertension during pregnancy): On the basis of simulating the physiological structure of uncomplicated twin placenta, increase the fluid flow rate pumped into the maternal fluid channel 103 representing the mother, reduce the width of the microchannel, and simulate placental vasospasm , Blood pressure increased.

5. Intrahepatic cholestasis of pregnancy (intrahepatic cholestasis of pregnancy): On the basis of simulating the physiological structure of uncomplicated twin placenta, increase the bile acid concentration in the medium pumped by the maternal fluid channel 103 representing the mother.

In the first application scenario of the multifetal placental vascular anastomosis simulation system provided by the present invention, it is used to evaluate the treatment effect, after simulating twin to twin transfusion syndrome (TTTs), TTTs are treated by selective laser electrocoagulation After closing the first valve assembly and all second valve assemblies, observe the change.

The microfluidic chip of the twin placental vascular anastomosis model prepared by the present invention can use various micro-engineering techniques to reconstruct the three-dimensional microstructure of the twin placenta barrier, a micro-engineering cell culture platform for dynamic microenvironment and biological functions, and expand the capabilities of the cell culture model. range; and can simulate the microstructure of two different chorionic twin placenta, with the same plane of control, can provide a substitute model for the current study of twin maternal-fetal barrier and the potential pathophysiological mechanism of twin special complications. This microfluidic chip can allow the two circulations of two fetuses representing the same mother to be cultured on the same model to simulate the intrauterine environment of twins, and allow two types of multiple placental vascular anastomoses to be cultured in the same device to form a control; The microfluidic chip can simulate physiological flow conditions to realize convective blood movement between twin fetuses and the mother, blood convection and/or unidirectional flow between two fetuses in the same model; flow conditions can be changed to simulate the gap between villi and fetus The abnormal blood flow environment of capillaries and the changes before and after treatment; the microfluidic chip adds a cell pool representing fetal tissue, and links the metabolism of fetal tissue with the dynamic change of the maternal-fetal barrier to further simulate the pathophysiological state of the mother-fetal.

In the description of this specification, descriptions referring to the terms "one embodiment", "some embodiments", "example", "specific examples", or "some examples" mean that specific features described in connection with the embodiment or example , structure, material or characteristic is included in at least one embodiment or example of the present invention. In this specification, schematic representations of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.

Although the embodiments of the present invention have been shown and described, those skilled in the art can understand that various changes, modifications, substitutions and modifications can be made to these embodiments without departing from the principle and spirit of the present invention. The scope of the invention is defined by the claims and their equivalents.

Claims (11)

1. A method for using a multiple placenta vascular anastomosis simulation system, characterized in that, the multiple placenta vascular anastomosis simulation system includes a microfluidic chip and at least 3 fluid pumps of a multiple placenta vascular anastomosis model; The microfluidic chip of the multifetal-placental vascular anastomosis model includes a chip body (1), and the chip body (1) is provided with a maternal fluid channel (103) and two or more fetal fluid channels (104). A porous membrane (2) is provided between the maternal fluid channel (103) and the fetal fluid channel (104), and a connecting channel that can be communicated or disconnected is provided between the fetal fluid channel (104); the fetal fluid channel ( 104) is provided with a cell pool (107); using a plurality of fluid pumps to pump fluid into the fetal fluid channel (104) and the maternal fluid channel (103); All or part of the fetal fluid channel (104) is located on the first side of the chip body (1), the maternal fluid channel (103) is located on the second side of the chip body (1), and the fetal fluid channel (104) The overlapping area with the maternal fluid channel (103) is separated from the fetal fluid channel (104) and the maternal fluid channel (103) by a porous membrane (2), or, all or part of the fetal fluid channel (104) is located in the chip body (1 ), the maternal fluid channel (103) is located on the first side of the chip body (1), and the adjacent area of the fetal fluid channel (104) and the maternal fluid channel (103) is separated by a porous membrane (2). Open fetal fluid channel (104) and maternal fluid channel (103); The method of using the multifetal placenta vascular anastomosis simulation system includes: attaching the first type of cells on the first side of the porous membrane (2) in the overlapping region or the adjacent region, the first side being the side of the porous membrane facing the fetal fluid channel (104); attaching a second cell on a second side of the porous membrane (2) in the overlapping region or adjacent region, the second side being the side of the porous membrane facing the parent fluid channel (103); Attach a third cell in the cell pool (107); Set the connection status of the connection channel; pumping fluid into the fetal fluid channel (104) and the maternal fluid channel (103); Observing and/or detecting the third cell and/or liquid in the cell pool (107). 2 . The method for using the multiple-fetal placenta vascular anastomosis simulation system according to claim 1 , characterized in that, the connection channel includes a first connection channel ( 105 ) that enables two-way or multi-way communication between fetal fluid channels ( 104 ). 3. The method for using the multi-fetal placenta vascular anastomosis simulation system according to claim 2, characterized in that, the first connecting channel (105) is provided with a first valve assembly. 4. The method for using the multiple-fetal placenta vascular anastomosis simulation system as claimed in claim 1, 2 or 3, wherein the connecting channel includes a second channel that is connected to the fetal fluid channel (104) and only allows fluid one-way circulation. Connection channels (106). 5. The method for using the multi-fetal placenta vascular anastomosis simulation system according to claim 4, characterized in that, one or two second connecting channels (106) with opposite flow directions are arranged between the two fetal fluid channels (104) . 6 . The method for using the multi-fetal placenta vascular anastomosis simulation system according to claim 5 , wherein a second valve assembly is provided on the second connecting channel ( 106 ). 7. the using method of multi-fetal placenta vessel anastomosis simulation system as claimed in claim 4, is characterized in that, the first end of described second connection channel (106) is connected with fetal fluid channel (104), and the second connection The second end of the channel (106) is provided with an input connection port. 8. The method for using the multifetal placenta vascular anastomosis simulation system according to claim 1, characterized in that the size of the porous membrane (2) in the overlapping area can be adjusted. 9. The method for using the multi-fetal placenta vascular anastomosis simulation system according to claim 1, characterized in that the size of the porous membrane (2) in the adjacent area can be adjusted. 10. The method for using the multi-fetal placenta vascular anastomosis simulation system according to claim 4, characterized in that a fluid pump is used to pump fluid into the input connection port of the second connection channel (106). 11. The method for using the multiple placenta vascular anastomosis simulation system according to claim 1, wherein the method for using the multiple placenta vascular anastomosis simulation system further comprises: varying the flow rate of fluid pumped into the fetal fluid pathway (104) and/or maternal fluid pathway (103); and/or, adjusting the size of overlapping regions or adjacent regions; And/or, adjusting the substance composition and/or substance composition ratio of the fluid pumped into the parent fluid channel (103).

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