Contraction force measuring device, contraction force measuring system, and method for measuring the contraction force of annular tissue

Abstract

This technology provides a method for quantitatively evaluating the radial (concentric) contraction force of annular tissues. [Solution] The shrinkage force measuring device comprises a base portion, a plurality of pillars erected on the upper surface of the base portion, having a flexible and conductive resistive layer formed thereon and arranged in a ring shape, and a pair of electrodes electrically connected to each end of the plurality of pillars, wherein the plurality of pillars form a ring-shaped array and can contact the inner circumferential surface of an annular structure fitted onto the outside of the ring-shaped array, and an external circuit can detect the change in the electrical characteristics of the resistive layer due to the bending of the plurality of pillars, and the radial shrinkage force of the annular structure can be calculated based on the detection result.

Description

Technical Field

[0001] The present invention relates to a contraction force measuring device, a contraction force measuring system, and a method for measuring the contraction force of a circular tissue.

Background Art

[0002] Conventionally, for the purpose of understanding vascular functions and disease mechanisms, vascular models in an in vitro environment, such as a two-layered vascular model (Non-Patent Document 1), triple coaxial cell printing (Non-Patent Document 2), Vessel-on-a-chip (Non-Patent Document 3), etc., have been proposed. In these studies, a hierarchical structure including both smooth muscle cells and endothelial cells has been constructed, and a model close to an actual blood vessel has been realized.

Prior Art Documents

Non-Patent Documents

[0003]

Non-Patent Document 1

Non-Patent Document 2

Non-Patent Document 3

Summary of the Invention

Problems to be Solved by the Invention

[0004] However, conventional models were too large and lacked a sensor-integrated platform capable of real-time monitoring, and there was no suitable method for measuring contractile force, thus preventing the quantitative evaluation of smooth muscle contractile force.

[0005] To solve the above problems, the present invention provides a technology that can quantitatively evaluate the radial (concentric) contraction force of annular tissue. [Means for solving the problem]

[0006] To solve the above problems, the present invention employs the following configuration. In other words, the gist of the present invention is as follows.

[0007] [1] A shrinkage force measuring device comprising a base portion, a plurality of pillars erected on the upper surface of the base portion and having a flexible and conductive resistive layer formed thereon, arranged in a ring shape, and a pair of electrodes electrically connected to each end of the plurality of pillars, wherein the plurality of pillars form a ring arrangement and can contact the inner surface of an annular structure fitted to the outside of the ring arrangement, and an external circuit can detect the change in the electrical characteristics of the resistive layer due to the bending of the plurality of pillars, and can calculate the radial shrinkage force of the annular structure based on the detection result. [2] The contraction force measuring device according to [1], wherein the pillar has a pair of columnar parts and a connecting part that connects the upper ends thereof, and the lower ends of the pair of columnar parts are each connected to the electrodes. [3] The shrinkage force measuring device according to [1] or [2], wherein the pillar comprises a flexible core material and the resistive layer formed on the surface of the core material, and the change in the resistance value of the resistive layer occurs in accordance with the bending of the pillar. [4] The contraction force measuring device according to any one of [1] to [3], wherein the pair of electrodes are electrically insulated from each other by grooves formed on the upper surface of the base portion. [5] The contraction force measuring device according to any one of [1] to [4], wherein the plurality of pillars are arranged at equal intervals in the circumferential direction. A contraction force measuring system comprising: a contraction force measuring device described in any of [6] [1] to [5]; a bridge circuit that converts the change in the resistance value of the pillar into a voltage; and an information processing device that converts the voltage into a force acting on the pillar. [7] The contraction force measurement system according to [6], wherein the information processing device calculates the force from the voltage by integrated calibration based on displacement-force and displacement-voltage calibration. A method for measuring the contraction force of an annular tissue, comprising: fitting an annular tissue onto the annular arrangement of the contraction force measuring device described in any of [8], [1] to [5], bringing it into contact with each pillar; detecting the change in electrical properties associated with the bending of the pillars; and calculating the contraction force of the annular tissue based on the change in electrical properties. [Effects of the Invention]

[0008] According to the present invention, it is possible to quantitatively evaluate the radial contraction force of annular tissue. [Brief explanation of the drawing]

[0009] [Figure 1] Figure 1 is an overall perspective view of the contraction force measuring device 100 according to an embodiment. [Figure 2] Figure 2 is a plan view of the contraction force measuring device 100 according to an embodiment. [Figure 3] Figure 3 is an enlarged view of the strain gauge 10 in the contraction force measuring device 100 according to the embodiment. [Figure 4] Figure 4 is an overall perspective view showing the measurement of the contractile force of an annular tissue T1 using the contractile force measuring device 100 according to the embodiment. [Figure 5] Figure 5 is a plan view showing the state of measuring the contractile force of an annular tissue T1 using the contractile force measuring device 100 according to the embodiment. [Figure 6] Figure 6 is a diagram illustrating the measurement principle. [Figure 7] Figure 7 is a schematic diagram of a measurement system 1000 equipped with a contraction force measuring device 100 according to an embodiment. [Figure 8]FIG. 8 is a diagram for explaining a method of manufacturing the contraction force measuring device 100 according to the embodiment. [Figure 9] FIG. 9 is a conceptual diagram of a 3D flexible variable resistor (contraction force measuring device) capable of evaluating the concentric contraction force of an annular tissue. [Figure 10] FIG. 10(a) is a diagram showing a method of manufacturing a 3D variable resistor, and FIG. 10(b) is a conceptual diagram of an experimental setup. [Figure 11] FIG. 11(a) is a diagram showing immunostaining and orientation of an annular smooth muscle tissue, and FIG. 11(b) is a diagram showing a computational simulation and the displacement-force gradient between six pillars. [Figure 12] FIG. 12 is a diagram for explaining the voltage-force calibration of a pillar. FIG. 12(a) shows the relationship between the displacement-force and the displacement-voltage of the pillar, and FIG. 12(b) shows the voltage-force relationship of the pillar. [Figure 13] FIG. 13 is a diagram showing a real-time electrical reading of the spontaneous contraction force generated by an annular smooth muscle tissue on a single pillar. FIG. 13(a) is a top image showing the measurement state, FIG. 13(b) is a cross-sectional image, and FIG. 13(c) is a diagram showing the measurement results of the contraction force. [Figure 14] FIG. 14 is a conceptual diagram of a 3D printed flexible device capable of evaluating the concentric contraction force of an annular smooth muscle tissue. [Figure 15] FIG. 15 is a diagram showing a method of manufacturing a 3D printed flexible device with an annular smooth muscle tissue attached. [Figure 16] FIG. 16 is a photographic image of a 3D printed flexible device. FIG. 16(a) is a top image, and FIG. 16(b) is a cross-sectional image. FIG. 16(b-i) shows the state without applying force, and FIG. 16(b-ii) shows the state with force applied. [Figure 17] FIG. 17(a) is a photographic image showing the contraction of a smooth muscle tissue. FIG. 17(b) is a diagram showing an equation related to force and displacement. [Figure 18] FIG. 18 is a diagram showing the area change due to the contraction of a smooth muscle tissue. [Figure 19]FIG. 19(a) is a photographic image showing immunostaining of smooth muscle tissue. FIG. 19(b) is a diagram showing the orientation of smooth muscle tissue. FIG. 19(c) is a diagram showing the passive contractile force of smooth muscle tissue. FIG. 19(d) is a diagram showing the active contractile force of smooth muscle tissue. [Figure 20] FIG. 20 is a conceptual diagram of a 3D printed flexible device capable of evaluating the concentric contractile force of circular smooth muscle tissue. [Figure 21] FIG. 21 is a conceptual diagram of an experimental setup. [Figure 22] FIG. 22 is a diagram showing a method for manufacturing a 3D printed flexible device using circular smooth muscle tissue. [Figure 23] FIG. 23 is a diagram showing the characteristics of smooth muscle tissue. [Figure 24] FIG. 24 is a diagram showing the characteristics of smooth muscle tissue. [Figure 25] FIG. 25 is a diagram showing the contraction of smooth muscle tissue. [Figure 26] FIG. 26 is a diagram showing the contraction of smooth muscle tissue. [Figure 27] FIG. 27 is a diagram showing a sensing experiment using Cr / Au film-coated pillars.

MODE FOR CARRYING OUT THE INVENTION

[0010] Hereinafter, embodiments of the present disclosure will be described with reference to the drawings. However, such description should not be construed in a limiting sense and does not limit the subject matter described in the claims. Also, various changes, substitutions, and modifications can be made without departing from the spirit and scope of the present disclosure. Further, different embodiments can be appropriately combined.

[0011] In this specification, the numerical range represented by "~" means a range including the numerical values described before and after "~" as the lower limit value and the upper limit value. For example, "A~B" means A or more and B or less.

[0012] In this specification, "tissue" refers to living tissue that exhibits a contractile response spontaneously or induced. Specifically, in addition to muscle tissue (smooth muscle tissue, skeletal muscle tissue, etc.), it also includes skin tissue, connective tissue (including fibroblasts), vascular endothelial tissue, etc., which exert contractile force during the tissue formation process. These tissues have been confirmed to exhibit contractile force even in an in vitro environment and are targets that can be measured by the contractile force measuring device of the present invention.

[0013] In this specification, "annular (ring-shaped)" means a closed contour or similar shape formed along the circumferential direction, and includes not only forms in which the constituent elements are continuous in the circumferential direction, but also forms in which the constituent elements are arranged intermittently (intermittently) in the circumferential direction. Furthermore, "annular" also includes cylindrical forms in which the ring is extended in the axial direction. Accordingly, "annular tissue" refers to an annular tissue that is formed along the circumferential direction and exhibits contractile force, and a three-dimensional tissue structure having an annular cross-sectional shape and a certain height in the axial direction is also included in "annular tissue". In addition, "arranged in an annular shape" or "arranged along the circumferential direction" does not necessarily mean that the elements are strictly arranged on a perfect circular orbit, but also includes forms in which they are generally arranged along the circumferential direction. That is, even if the arranged constituent elements are lined up in a shape close to a circle, or arranged in a manner similar to a circle, they are included in "arranged in an annular shape" or "arranged along the circumferential direction".

[0014] Furthermore, "radial direction (concentric direction)" refers to the direction perpendicular to the axial direction of the annular member, and "diameter" refers to the dimension in the radial direction. Also, "diameter" refers to the dimension in the direction perpendicular to the axial direction of a cylindrical or annular shape, and regardless of the cross-sectional shape, it shall indicate the maximum width or representative dimension of the cross-section.

[0015] In this specification, "change in electrical characteristics" refers to the change in the electrical resistance of the resistive layer that occurs when the pillar is bent, and this change in resistance is output as a differential voltage signal via a bridge circuit. Therefore, "change in electrical characteristics" includes not only the change in resistance itself, but also the change in the voltage signal caused by it.

[0016] In this specification, "external circuit" refers to a circuit for detecting changes in the electrical characteristics (changes in resistance) of the pillar, and mainly includes a bridge circuit. If necessary, the external circuit may also include signal processing circuits such as amplifiers that amplify and bandwidth-limit the detected signal.

[0017] Figure 1 is a perspective view showing the entire shrinkage force measuring device 100 (hereinafter sometimes simply referred to as "device 100") according to the embodiment. Figure 2 is a plan view of the shrinkage force measuring device 100 according to the embodiment. Figure 3(A) is an enlarged view of the strain gauge 10 in the shrinkage force measuring device 100 according to the embodiment. Figure 3(B) is a diagram showing a cross-section of the pillar 2 according to the embodiment that is perpendicular to the extension direction. Figure 4 is an overall perspective view showing the state of measuring the shrinkage force of an annular tissue using the shrinkage force measuring device 100 according to the embodiment. Figure 5 is a plan view showing the state of measuring the shrinkage force of an annular tissue using the shrinkage force measuring device 100 according to the embodiment. Figure 6 is a diagram illustrating the measurement principle. Figure 7 is a configuration diagram of the shrinkage force measuring system 1000 (hereinafter sometimes simply referred to as "measurement system 1000") equipped with the device 100 according to the embodiment. Figure 8 is a diagram illustrating the manufacturing method of the shrinkage force measuring device 100 according to the embodiment. Hereinafter, the device 100, the measurement system 1000, and the measurement method will be described with reference to the drawings.

[0018] [Contraction force measuring device] As shown in Figure 1, the device 100 includes a base portion 1 that serves as a base, a plurality of pillars 2 erected on the upper surface 1A of the base portion 1 and arranged in a ring shape, and a pair of electrodes 3, 4 provided on the upper surface 1A of the base portion 1 and connected to each end of the plurality of pillars 2. Each pillar 2 and the pair of electrodes 3, 4 constitute a strain gauge 10 (variable resistor). The ring-shaped structure composed of the plurality of pillars 2 is referred to as a ring array 20. The axial direction of the ring array 20 coincides with the vertical direction of the device 100. In this embodiment, six strain gauges 10 are provided, but the number is not limited to this.

[0019] As shown in Figures 4 and 5, the device 100 can measure the radial contractile force of tissue T1 when the annular tissue T1 is fitted onto the annular arrangement 20 from the outside and the inner circumferential surface of tissue T1 is in contact with the pillar 2. Examples of tissue T1 that the device 100 can measure the contractile force of include, but are not limited to, muscle tissue (smooth muscle tissue, skeletal muscle tissue, etc.), skin tissue, connective tissue, and vascular endothelial tissue.

[0020] [Base section 1] The base portion 1 is an insulating substrate that supports the strain gauge 10 and has a regular hexagonal shape in plan view. However, the shape of the base portion 1 is not limited to this. The material of the base portion 1 is not particularly limited, but an insulating material such as a biocompatible resin is preferred. A photocurable resin can be used as an example of the material of the base portion 1, and the base portion 1 can be integrally molded by 3D printing or the like. A groove 11 is formed on the upper surface 1A of the base portion 1, outlining a pair of electrodes 3 and 4. The groove 11 ensures electrical insulation between the pair of electrodes 3 and 4. The surface of the base portion 1 may be coated with parylene to improve the adhesion of the Cr / Au film and reduce cytotoxicity.

[0021] [Pillar 2] As shown in Figure 3(A), the pillar 2 has a shape in which a rod-shaped member with a circular cross-section is bent into an inverted U-shape. Specifically, the pillar 2 includes a pair of upright columnar sections 21 and 22, and a connecting section 23 that connects the upper ends of the pair of columnar sections 21 and 22, and is folded back at the connecting section 23. The lower ends of the pair of columnar sections 21 and 22 are electrically and mechanically connected to electrodes 3 and 4, respectively. Specifically, columnar section 21 is erected on electrode 3, and columnar section 22 is erected on electrode 4. As a result, the pillar 2 is erected straddling the pair of electrodes 3 and 4.

[0022] Here, Figure 3(A) shows the axial direction, circumferential direction (i.e., the direction of arrangement of the multiple pillars 2) and radial direction of the annular arrangement 20. As shown in Figure 3(A), the pair of columnar parts 21 and 22 are arranged along the circumferential direction of the annular arrangement 20.

[0023] As shown in Figure 3(B), the pillar 2 is composed of a flexible core material 2A and a conductive resistor layer 2B formed on the surface of the core material 2A. This allows the pillar 2 to function as a three-dimensional variable resistor element that is flexible and whose electrical resistance changes with bending. The material of the core material 2A is not particularly limited, but a resin material such as a flexible biocompatible resin is preferred. A photocurable resin can be used as an example of the core material 2A, and the core material 2A may be integrally molded from the same material as the base part 1. The material of the resistor layer 2B is not particularly limited, but for example, a thin metal film such as Cr / Au, a conductive polymer, or an alloy thin film can be used. Furthermore, the surface of the pillar 2 may be coated with parylene to improve the adhesion of the Cr / Au film and reduce cytotoxicity. As shown in Figure 3(B), since the cross-section of the core material 2A is circular, the pillar 2 has a circular cross-section. In addition to flexibility, the pillar 2 may also be pliable. As a result, pillar 2 can flex to respond with greater flexibility and sensitivity to even minute displacements associated with the contraction of the annular tissue, contributing to the precise detection of contraction force.

[0024] As shown in Figures 4 and 5, the multiple pillars 2 are positioned inside the annular tissue T1 during the measurement of contractile force, and each pillar 2 contacts the inner circumferential surface of the tissue T1. At this time, as described above, since the pair of columnar portions 21 and 22 are arranged along the circumferential direction, the columnar portions 21 and 22 contact the tissue T1 evenly. The height of the pillars 2 can be appropriately changed according to the height (length in the axial direction) of the tissue T1, and the method is also applicable when the tissue T1 is a cylindrical shape that is extended in the axial direction.

[0025] As shown in Figure 6, when measuring the shrinkage force of tissue T1, a component of the shrinkage force acts on each pillar 2 from the radial outside as tissue T1 shrinks. That is, the inner surface of tissue T1 comes into contact with multiple pillars 2, and the shrinkage force of the entire tissue T1 is distributed to each pillar and acts locally. At this time, when a component of the shrinkage force of tissue T1 acts from the radial outside on a pillar 2 containing a flexible core material 2A and a resistive layer 2B, the columnar portions 21 and 22 of the pillar 2 are displaced so as to curve radially inward, and strain is generated in the resistive layer 2B. The resistance value of the resistive layer 2B changes in accordance with this strain.

[0026] In this embodiment, six pillars 2 are arranged at equal intervals in the circumferential direction, but the number of pillars 2 is not limited to this. Multiple pillars 2 can be arranged in a ring, and the number of pillars 2 is preferably three or more, more preferably six or more.

[0027] [Electrodes 3,4] The pair of electrodes 3 and 4 are formed by a conductive pattern provided on the upper surface 1A of the base portion 1. The electrodes 3 and 4 are conductive and are insulated from each other by grooves 11 formed on the upper surface 1A of the base portion 1. The pair of electrodes 3 and 4 are electrically continuous with the resistive layer of each pillar 2 and are connected by wires to an external circuit (bridge circuit 300 described later). The material of electrodes 3 and 4 is not particularly limited, but like the resistive layer 2B, for example, a thin metal film such as Cr / Au, a conductive polymer, or an alloy thin film can be used.

[0028] [Measurement System 1000] Next, a measurement system 1000 comprising the device 100 will be described. As shown in Figure 7, the measurement system 1000 comprises the device 100, a bridge circuit 300 (an example of an external circuit), an amplifier 400, a data logger 500, and an information processing device 600.

[0029] Multiple bridge circuits 300 are arranged to correspond to each of the multiple strain gauges 10, and are connected by wires to the electrodes 3 and 4 of each strain gauge 10. In a Wheatstone bridge configuration, the small resistance changes occurring in the resistive layer 2B of the pillar 2 are output as differential voltage signals.

[0030] Amplifier 400 is a differential amplifier for low-noise measurement, which amplifies and band-limits the bridge output voltage output from the bridge circuit 300 within a desired bandwidth. Multiple amplifiers 400 are arranged to correspond to each of the multiple strain gauges 10.

[0031] The data logger 500 records the voltage signal output from the amplifier 400 in a time series at a predetermined sampling period (e.g., 1 Hz to 100 Hz). The data logger 500 has multiple channels corresponding to each of the multiple strain gauges 10, and detects the voltage signal output from each amplifier 400 from each channel. In this way, the data logger 500 records the voltage signals corresponding to the multiple strain gauges 10 in a time series simultaneously. In addition to the recording function, the data logger 500 may also be configured as an information processing device that has voltage signal processing and conversion functions.

[0032] The information processing device 600 is a computer device that includes, for example, a CPU (a processor), memory, a communication interface (IF), and storage. The information processing device 600 acquires voltage signals from multiple strain gauges 10 output from the data logger 500 and converts them into forces acting on each pillar 2 based on the correspondence between voltage and force. As described above, the force acting on each pillar 2 when measuring contraction force is a component force that acts locally on each pillar 2 when the contraction force of the entire tissue T1 is distributed to each pillar 2. The information processing device 600 calculates the total radial contraction force of the entire annular tissue by summing the measured values ​​of the component forces acting on each pillar 2.

[0033] The information processing device 600 performs force conversion based on experimental calibration of displacement-force and displacement-voltage. Specifically, it obtains in advance the relationship between the displacement (Δx) and force (F) of pillar 2 when a known external force is applied to pillar 2, and the relationship between the displacement and voltage (V), and integrates these to construct a calibration formula for calculating force from voltage. The displacement Δx is obtained, for example, by analyzing images of pillar 2 before and after deformation.

[0034] The measurement system 1000 may also be configured to calculate the contraction force acting on each pillar 2 by displacement-force conversion based on material mechanics equations. In that case, the measurement system 1000 may be equipped with a camera that takes images of the pillar 2 before and after deformation, and the displacement amount Δx of the pillar 2 may be obtained by analyzing the images.

[0035] [Method for measuring contractile force] Next, a method for measuring the contractile force of annular tissue using the apparatus 100 will be described.

[0036] First, as shown in Figures 4 and 5, the cultured tissue T1 is fitted onto the annular arrangement 20, which consists of multiple pillars 2, and brought into contact with the pillars 2. As a result, the multiple pillars 2 are positioned inside the tissue T1, and the columnar portions 21 and 22 are in contact with the inner circumferential surface of the tissue T1.

[0037] Spontaneous contraction or pharmacological stimulation (e.g., histamine, high potassium) + When tissue T1 contracts due to the force applied, the columnar portions 21 and 22 of each pillar 2 bend, and the resistance values ​​of the strain gauges 10 change. As a result, the output voltage of the bridge circuit 300 changes over time. At this time, the information processing device 600 acquires the voltage signals from multiple strain gauges 10 output from multiple data loggers 500 and converts them into component forces of the contraction force acting on each pillar 2 based on the correspondence between voltage and force. The information processing device 600 calculates the total radial contraction force of tissue T1 by summing the measured values ​​of these component forces. In this way, the contraction force of tissue T1 can be measured.

[0038] Alternatively, instead of fitting the cultured tissue T1 onto the annular array 20, the tissue T1 may be cultured on the upper surface 1A of the device 100 so as to surround the annular array 20, and its contractile force may be measured. Furthermore, the device 100 can also be applied to measuring contractile force during the tissue formation process.

[0039] [Manufacturing method] Next, the manufacturing method for the apparatus 100 will be described. In the manufacturing method described below, as an example, the base part 1 and the core material 2A of the pillar 2 are integrally molded, and then the resistor layer 2B and electrodes 3 and 4 are formed by Cr / Au deposition.

[0040] Specifically, first, a base body 30 is formed by integrating the base part 1 and the core material 2A of the pillar 2, for example, using a 3D printer. Next, as shown in Figure 8, the mask 200 is attached to the base body 30. At this time, the frame 201 of the mask 200 is fitted into the groove 11 of the base body to cover the surface that does not require vapor deposition. Next, a Cr / Au vapor-deposited film is formed on predetermined areas of the pillar surface and the upper surface 1A to form the resistor layer 2B and electrodes 3 and 4. This manufactures a device 100 equipped with a strain gauge 10 composed of the pillar 2 and electrodes 3 and 4.

[0041] [Effects / Effects] As described above, the apparatus 100 according to the embodiment comprises a base portion 1, a plurality of pillars 2 erected on the upper surface 1A of the base portion 1, having a flexible and conductive resistor layer 2B formed on them and arranged in an annular shape, and a pair of electrodes 3 and 4 electrically connected to each end of the plurality of pillars 2. The plurality of pillars 2 form an annular array 20 and can contact the inner circumferential surface of an annular tissue T1 fitted onto the outside of the annular array 20. An external circuit (bridge circuit 300) can detect changes in the electrical characteristics of the resistor layer 2B due to the bending of the plurality of pillars 2, and the radial shrinkage force of the tissue T1 can be calculated based on the detection result. As a result, since local forces due to the shrinkage of the tissue T1 can be detected by the plurality of pillars 2 arranged in an annular shape, the shrinkage force of the entire tissue T1 can be measured in real time and quantitatively.

[0042] Furthermore, in the apparatus 100 according to the embodiment, the pillar 2 has a pair of columnar portions 21 and 22 and a connecting portion 23 that connects their upper ends, and the lower ends of the columnar portions 21 and 22 are connected to electrodes 3 and 4, respectively. With this, the strain gauge 10 can be constructed by the pillar 2 and electrodes 3 and 4.

[0043] Furthermore, the pillar 2 includes a flexible core material 2A and a resistive layer 2B formed on the surface of the core material 2A, and the resistance value of the resistive layer 2B changes in accordance with the bending of the pillar 2. As a result, a highly sensitive electrical response can be obtained to minute force changes, and the contraction force of the tissue T1 can be detected with high accuracy.

[0044] The resistive layer 2B is a thin metal film. This provides high conductivity and stable strain response characteristics, thereby improving signal accuracy and reliability.

[0045] The pair of electrodes 3 and 4 are electrically insulated from each other by grooves formed on the upper surface of the base portion 1. This prevents interference between adjacent terminals and circuits, thereby improving the separation and accuracy of the measurement signal.

[0046] There are six pillars 2, arranged at equal intervals in the circumferential direction. This allows for the uniform and multi-point detection of the contractile force of tissue T1, making it possible to calculate the total contractile force with less variability.

[0047] Furthermore, the contraction force measurement system 1000 according to this embodiment includes a contraction force measuring device 100, a bridge circuit 300 that converts the change in resistance of the pillar 2 into a voltage, and an information processing device 600 that converts the voltage into a force acting on the pillar 2. This allows for integrated processing from the detection to the conversion of the contraction force, enabling real-time and automatic force evaluation.

[0048] Furthermore, the information processing device 600 according to this embodiment calculates force from voltage through integrated calibration based on experimental calibration of displacement Δx - force F and displacement Δx - voltage V. This enables highly accurate force conversion and improves the reliability of the measurement.

[0049] Furthermore, the method for measuring the shrinkage force of tissue T1 according to the embodiment involves fitting the annular tissue T1 onto the annular arrangement 20 of the shrinkage force measuring device 100 and bringing it into contact with the pillars 2, measuring the change in the electrical characteristics of the resistor layer 2B as the pillars 2 are bent, and calculating the shrinkage force of tissue T1 based on the change in electrical characteristics. As a result, since the local force associated with the shrinkage of tissue T1 can be detected by the multiple pillars 2 arranged in an annular shape, the shrinkage force of the entire tissue T1 can be measured in real time and quantitatively.

[0050] <Examples of application> The contraction force measuring device and measuring system of the present invention can be suitably applied to the following uses.

[0051] [Application as a drug discovery model for cardiovascular diseases] Because it allows for real-time and quantitative evaluation of muscle tissue contraction responses, pharmacological stimuli (e.g., histamine, hyperpotassium) can be used. + By measuring the response to ), this can be used for screening cardiovascular disease-related drugs and analyzing their mechanisms of action.

[0052] [Assessment of Vascular Function Characteristics] By detecting and aggregating the contractile force of tissue T1 at multiple points via pillars arranged in a ring, it is possible to evaluate the mechanical response, such as the contractile function and elastic properties of blood vessels, with high accuracy.

[0053] [Analysis of the regulatory mechanism of vasoconstriction function by smooth muscle] This invention allows for the direct measurement of the function of smooth muscle tissue responsible for the contraction of blood vessel walls, and contributes to the elucidation of the physiological and pathological mechanisms involved in the contraction and dilation of blood vessels.

[0054] [Use as an evaluation system for cultured smooth muscle tissue] By embedding a ring-shaped tissue made from human-derived smooth muscle cells into this device, functional evaluation in an in vitro environment becomes possible, and applications in the fields of regenerative medicine and tissue engineering are expected. [Examples]

[0055] The present invention will be described in more detail below with reference to examples applied to annular smooth muscle tissue. However, the present invention is not limited to the following examples, unless it deviates from its essence, and can be applied to measuring the contractile force of annular tissue as described above.

[0056] (Example 1) Figure 9 is a conceptual diagram of a 3D flexible variable resistor (contraction force measuring device) capable of evaluating the concentric contraction force of annular tissue. Figure 10(a) shows the manufacturing method of the 3D variable resistor, and Figure 10(b) is a conceptual diagram of the experimental setup. Figure 11(a) shows immunostaining and orientation of annular smooth muscle tissue, and Figure 11(b) shows a calculation simulation and the displacement-force gradient between six pillars. Figure 12 explains the voltage-force calibration of the pillars, with Figure 12(a) showing the displacement-force relationship and displacement-voltage relationship of the pillars, and Figure 12(b) showing the voltage-force relationship of the pillars. Figure 13 shows the real-time electrical readout of the spontaneous contraction force generated by annular smooth muscle tissue on a single pillar, with Figure 13(a) being a top view showing the measurement, Figure 13(b) being a cross-sectional view, and Figure 13(c) being a diagram showing the contraction force measurement results.

[0057] This embodiment describes a contraction force measuring device using a 3D-printed variable resistor for electrically reading the concentric contraction force of annular smooth muscle tissue in real time. By using an annular mold, smooth muscle tissue in which cells are oriented concentrically was formed. A three-dimensional flexible variable resistor was fabricated by depositing a Cr / Au film on a pillar structure formed on a flexible substrate (base). Parylene coating was applied to improve the adhesion of the Cr / Au film and reduce cytotoxicity.

[0058] This variable resistor was connected to a low-noise amplifier circuit, and the voltage output was recorded by a data logger, enabling real-time readout of the contractile force. Human aortic smooth muscle cells (HAoSMC) were mixed with a collagen substrate and injected into a ring-shaped mold. After substrate hardening, the formed ring-shaped smooth muscle tissue was transferred onto the 3D variable resistor, and the contractile force was continuously monitored for two days after placement (Figure 10).

[0059] Immunostaining of α-smooth muscle actin (α-SMA) confirmed that smooth muscle cells are oriented concentrically (Figure 11(a)). Furthermore, by comparing simulation results using COMSOL with experimental results, it was confirmed that the variable resistor is effective in measuring concentric contractile force (Figure 11(b)). By integrating the displacement-force and displacement-voltage measurement results, voltage-force calibration data was established, demonstrating that contractile force can be quantitatively calculated from the voltage output (Figure 12).

[0060] The contractile force increased rapidly over approximately 10 hours as the tissue oriented and mechanical tension formed, reaching an average of 400 μN per column on the second day. This corresponds to a concentric contractile force of approximately 2.4 mN for all six columns combined (Figure 13).

[0061] These results demonstrate that the contractile force measuring device of the present invention can evaluate the concentric contractile force of annular tissue in real time and quantitatively, making it a useful tool in biomedical research.

[0062] (Example 2) The embodiments of the present invention will be described below in the following paper format.

[0063] Figure 14 is a conceptual diagram of a 3D-printed flexible device capable of evaluating the concentric contraction force of annular smooth muscle tissue. Figure 15 is a diagram showing a method for manufacturing a 3D-printed flexible device with annular smooth muscle tissue attached. Figure 16 is a photographic image of the 3D-printed flexible device. Figure 16(a) is a top view image, and Figure 16(b) is a cross-sectional view image. Figure 16(bi) shows the state without applied force, and Figure 16(b-ii) shows the state with applied force. Figure 17(a) is a photographic image showing the contraction of smooth muscle tissue. Figure 17(b) is a diagram showing the equations relating force and displacement. Figure 18 is a diagram showing the change in area due to the contraction of smooth muscle tissue. Figure 19(a) is a photographic image showing immunohistochemical staining of smooth muscle tissue. Figure 19(b) is a diagram showing the orientation of smooth muscle tissue. Figure 19(c) is a diagram showing the passive contraction force of smooth muscle tissue. Figure 19(d) is a diagram showing the active contraction force of smooth muscle tissue.

[0064] This study describes a method for evaluating the concentric contractile force of annular smooth muscle tissue using a flexible device fabricated by 3D printing. By using an annular template, smooth muscle tissue with cells arranged concentrically was created. The contractile force of this smooth muscle tissue was evaluated using a flexible device fabricated by 3D printing. When histamine was added, the smooth muscle tissue responded with a contractile force of approximately 400 μN, demonstrating that the fabricated tissue possesses functional characteristics. Further development of this technology is expected to enable precise control and measurement of smooth muscle tissue, leading to more advanced applications in biomedical research.

[0065] To reproduce the shape and function of blood vessels, in vitro vascular models such as bilayer vascular models [Reference 1] and triple coaxial cell printing [Reference 2] have been proposed. These studies have realized hierarchical structures containing both smooth muscle cells and endothelial cells. However, these studies have not been able to quantitatively evaluate the contractile force of smooth muscle due to the large size of the models and the lack of measurement methods. Therefore, in this study, we propose a flexible device fabricated by 3D printing that can evaluate the concentric contractile force of annular smooth muscle tissue (Figure 14).

[0066] Human aortic smooth muscle cells (HAoSMCs) were mixed with a collagen substrate and 10x concentrated DPBS in a volume ratio of 9:1, and the mixture was poured into an annular mold. After the substrate solidified in 20-30 minutes, the annular smooth muscle tissue was transferred to a flexible device fabricated by 3D printing (Figure 15). The smooth muscle tissue was then cultured in the device for a further 2 days to stabilize the concentric alignment of the tissue. This 3D printed device was designed to have a flexible column that bends appropriately to allow for displacement measurement, as shown in Figure 16. Assuming a uniform load distribution, the contractile force was calculated from the observed displacement based on material mechanics [Reference 3] (Figure 17). To minimize toxicity to cells, biocompatible resin was used to fabricate the device.

[0067] Contraction of the smooth muscle tissue resulted in an area change of approximately 20%, which continued until day 5 (Figure 18). To confirm the shape of the smooth muscle cells and the arrangement of the tissue, immunostaining for α-smooth muscle actin (α-SMA) was performed (Figure 19(a)). As shown in Figure 19(b), a peak in brightness was observed at a 90-degree angle to the horizontal axis, confirming that the smooth muscle cells were aligned in a 90-degree direction and arranged concentrically. On day 5, a contractile force of approximately 1.8 mN was generated by passive contraction (Figure 19(c)). To evaluate active contraction, histamine was added to the culture medium to induce contraction. No significant change in force was observed in the control sample, but in the histamine-added sample, two distinct peak responses with a difference of approximately 400 μN were observed (Figure 19(d)). These results confirm that the 3D-printed device and annular smooth muscle tissue we propose not only satisfy the morphological characteristics of smooth muscle but also allow for the quantitative evaluation of its contractile function.

[0068] Figure 20 is a conceptual diagram of a 3D-printed flexible device capable of evaluating the concentric contraction force of annular smooth muscle tissue. Figure 21 is a conceptual diagram of the experimental setup. Figure 22 shows a diagram illustrating the manufacturing method of a 3D-printed flexible device using annular smooth muscle tissue. Figures 23 and 24 show the properties of smooth muscle tissue. Figures 25 and 26 show the contraction of smooth muscle tissue. Figure 27 shows a sensing experiment using a Cr / Au film-deposited pillar.

[0069] (References) 1. Y. Shimazu, et al., Journal of Bioscience and Bioengineering, vol. 127, pp 114-120, 2019 2. G. Gao and H. Kim, et al., Appl. Phys. Rev. vol 6, 041402, 2019 3. M. Bernini, et al., PLOS ONE; vol 18(8): e0283492, 2023 [Explanation of Symbols]

[0070] 100: Contraction force measuring device 1: Base section 1A:Top surface 11: Groove 10: Strain gauge 2: Pillar 21: Pillar part 22: Pillar part 23: Connection part 3: Electrode 4: Electrode 200: Mask 201: Frame 300: Bridge Circuit 400: Amplifier 500: Data logger 600: Information Processing Device 1000: Measurement System

Claims

1. The base part, Multiple pillars are erected on the upper surface of the base portion, and are flexible and have a conductive resistor layer formed on them, arranged in a ring shape. The plurality of pillars are each electrically connected to a pair of electrodes, The plurality of pillars form an annular arrangement and are capable of contacting the inner circumferential surface of an annular structure that is fitted onto the outside of the annular arrangement. The change in the electrical characteristics of the resistive layer due to the bending of the plurality of pillars can be detected by an external circuit, and the radial contraction force of the annular structure can be calculated based on the detection results. Contraction force measuring device.

2. The pillar has a pair of columnar sections and a connecting section that connects their upper ends. The lower ends of the pair of columnar sections are each connected to the electrodes. The contraction force measuring device according to claim 1.

3. The pillar includes a flexible core material and a resistive layer formed on the surface of the core material, wherein the resistance value of the resistive layer changes in accordance with the bending of the pillar. The contraction force measuring device according to claim 1.

4. The pair of electrodes are electrically insulated from each other by grooves formed on the upper surface of the base portion. The contraction force measuring device according to claim 1.

5. The plurality of pillars are arranged at equal intervals in the circumferential direction. The contraction force measuring device according to claim 1.

6. A contraction force measuring device according to any one of claims 1 to 5, A bridge circuit that converts the change in the resistance of the pillar into a voltage, The system includes an information processing device that converts the voltage into a force acting on the pillar, Contraction force measurement system.

7. The information processing device calculates the force from the voltage by integrated calibration based on displacement-force and displacement-voltage calibration. The contractile force measurement system according to claim 6.

8. An annular structure is fitted onto the annular arrangement of the contraction force measuring device according to any one of claims 1 to 5 and brought into contact with each pillar. The change in electrical characteristics associated with the bending of the pillar is detected, Based on the change in the electrical characteristics, the contractile force of the annular tissue is calculated. A method for measuring the contractile force of a ring-shaped tissue.

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