Use of a magnetic field in the manufacture of a device for the treatment of liver fibrosis

Abstract

The application discloses application of a magnetic field in preparation of a device for liver fibrosis treatment, and application in research on a liver fibrosis treatment mechanism or in construction of a liver fibrosis mouse model. In the application, through a simple and clear physical path of "applying a strong magnetic field, generating a targeted magnetic field mechanical effect (magnetic torque), and directly restructuring collagen fibers", a strategy leap from "indirect regulation of cells" to "direct reconstruction of a matrix" is realized. The device containing the magnetic field is targeted, has fast potential effect and high safety.

Description

Technical Field

[0001] This invention belongs to the field of biophysical therapy and innovative medical device technology, specifically relating to the application of a magnetic field in the preparation of a device for the treatment of liver fibrosis. Background Technology

[0002] Liver fibrosis, a necessary stage and core pathological basis for the development of cirrhosis and liver cancer, is a reversible process, making its treatment crucial. Although various anti-fibrotic drugs have been developed, such as colchicine and sorafenib, most of these drugs target biochemical signaling pathways like TGF-β, resulting in limited therapeutic efficacy and side effects such as hepatotoxicity and nephrotoxicity. Therefore, developing safe and effective anti-fibrotic treatment regimens is of significant clinical importance.

[0003] The typical pathological features of liver fibrosis include not only excessive collagen deposition, but more importantly, a significant directional spatial structure. As a major component of the extracellular matrix (ECM), collagen fibers abnormally accumulate during the progression of liver fibrosis, with type I collagen increasing by more than eightfold. Multiple clinical staging systems for liver fibrosis (such as Metavir, Scheuer, Batts-Ludwig, and Ishak) consistently indicate that in the early stages of fibrosis, collagen is deposited in the portal areas. As the disease progresses, collagen fibers extend along the hexagonal boundaries of the liver lobules, ultimately forming a directional distribution pattern of "portal area-portal area" bridging. Figure 1 This arrangement not only exacerbates mechanical tension but also provides contact guidance for cell migration, promoting the continued progression of fibrosis. Disrupting the pathological arrangement of collagen fibers holds promise for intervening in the mechanical microenvironment, offering new insights for the treatment of liver fibrosis.

[0004] Traditional strategies targeting collagen quantity in liver fibrosis treatment have frequently encountered obstacles in clinical translation. For example, simtuzumab, an antibody drug targeting lysyl oxidase (LOX), a key enzyme in collagen cross-linking, has shown anti-fibrotic effects in preclinical models, but it has proven ineffective in multiple clinical trials for patients with advanced liver fibrosis and cirrhosis, revealing the limitations of relying solely on biochemical inhibition strategies. This predicament has prompted researchers to focus on the qualitative dimension of collagen—its spatial arrangement and mechanical properties. Research by Du Yanan's team shows that the directional alignment of pathological collagen fibers is transmitted to the cell membrane via integrins, activating the liquid-liquid phase separation (LLPS) of LIM domain-containing protein 1 (LIMD1) bound to integrins. This leads to local aggregation, recruiting more focal adhesion components and actin to bind, enhancing the cell's ability to migrate along the collagen direction. Physical cutting perpendicular to collagen fibers can interrupt this process.

[0005] Magnetic fields are a non-invasive physical intervention method with high tissue penetration and outstanding biosafety, demonstrating unique potential. Collagen exhibits significant magnetic anisotropy, with the magnetic anisotropy value of a single collagen molecule being approximately -1 × 10⁻⁶. -25 J.T -2 Collagen fibers will generate torque and rotate in a uniform, strong magnetic field until their long axis is perpendicular to the direction of the magnetic field. Studies show that when type I collagen solutions are exposed to a strong magnetic field of 7T, the collagen fibers preferentially align perpendicular to the direction of the magnetic field. Figure 2 When a mixture of osteoblasts and collagen was exposed to an 8T magnetic field, collagen fibers were observed to be perpendicular to the direction of the magnetic field. This indicates that the magnetic field has the ability to directionally regulate the collagen network structure, but how the magnetic field affects collagen arrangement and regulates the liver fibrosis process remains uncertain. Summary of the Invention

[0006] The technical problem to be solved by the present invention is to provide a new application of magnetic field in the preparation of a device for the treatment of liver fibrosis, as well as a new application in the study of the treatment mechanism of liver fibrosis and in the construction of a mouse model of liver fibrosis.

[0007] The present invention solves the above-mentioned technical problems through the following technical means:

[0008] This invention proposes the application of a magnetic field in the preparation of a device for the treatment of liver fibrosis.

[0009] Preferably, the magnetic field strength is 1.5-9.4T; more preferably, it is one of 1.5T, 3T, 7T and 9.4T.

[0010] Preferably, the magnetic field is a steady-state magnetic field.

[0011] Preferably, the device for treating liver fibrosis includes exposing the person or animal with liver fibrosis to a magnetic field during the treatment process.

[0012] Preferably, the magnetic field alleviates the degree of liver fibrosis by reducing the content of liver collagen fibers and / or reducing the content of liver hydroxyproline and / or disrupting the pathological collagen fiber network structure.

[0013] Preferably, the magnetic field acts directly on the collagen fibers in the liver of humans and / or animals, inducing physical changes in the collagen fiber structure.

[0014] Preferably, the physical change includes one or more of the following: a decrease in fiber width, a shortening of fiber length, and an increase in straightness.

[0015] This invention also proposes the application of magnetic fields in studying the treatment mechanism of liver fibrosis or in constructing a mouse model of liver fibrosis.

[0016] Preferably, the magnetic field strength is 1.5-9.4T.

[0017] Preferably, the application includes exposing liver fibrotic animals to a magnetic field using the magnetic field.

[0018] Preferably, the magnetic field exposure is performed twice a week for 5 hours each time, for eight weeks, for a total of 80 hours.

[0019] Preferably, the method for constructing liver fibrosis animals includes the following steps: injecting the animal intraperitoneally with a corn oil solution containing carbon tetrachloride twice a week for eight weeks.

[0020] Preferably, the volume ratio of carbon tetrachloride to corn oil is 1:10.

[0021] Preferably, the animal is a mouse; the mouse is injected intraperitoneally with a corn oil solution containing carbon tetrachloride at a dose of 0.5 ml / kg.

[0022] Preferably, the application includes obtaining liver tissue from the subject after the magnetic field exposure has ended, and assessing the degree of liver fibrosis by Sirius red staining and / or α-SMA immunohistochemical staining.

[0023] Preferably, the application includes obtaining liver tissue from the subject after magnetic field exposure and assessing the arrangement trend of liver collagen fibers by COL1A1 immunofluorescence detection.

[0024] Preferably, the application includes using CT Fire software to extract individual collagen fibers from images and perform quantitative analysis to evaluate the physical properties of the collagen fibers after the magnetic field exposure has ended.

[0025] The beneficial effects of this invention are as follows: (1) This invention provides a novel approach for direct and active physical intervention on pathological collagen fibers, overcoming the fundamental limitations of existing technologies. By applying a strong magnetic field, the resulting magnetic field mechanical effect (with magnetic torque as the core) can directly act on the collagen fibers themselves, rather than by regulating cell function. This step makes direct mechanical reconstruction of dense fiber networks possible, overcoming the shortcomings of existing technologies that can only intervene indirectly and passively.

[0026] (2) This invention achieves targeted physical ablation and remodeling of fibrotic tissue, with a clear mechanism of action and rapid response. The key step of this invention—applying a strong magnetic field with precise and controllable intensity—can highly concentrate energy (mechanical effect) on collagen fibers with antimagnetic anisotropy while minimizing damage to surrounding normal liver tissue, thus achieving targeted treatment. Its core mechanism of action is physical magnetic torque, which does not depend on slow cell metabolism and signal transduction. Therefore, compared with drugs that require weeks or even months to show effects, this method is expected to induce immediate or rapid physical changes in collagen fiber structure (such as orientation rearrangement), thereby potentially achieving a faster onset of therapeutic effect.

[0027] (3) The magnetic field device of this invention is completely different from the traditional magnetic field intervention principle based on the chemical effect of "free radical pairs". Instead, it is based on the physical foundation of direct "magnetic field mechanical effect", contributing a brand-new technical principle and theoretical framework to the field of liver fibrosis treatment. This method does not require the introduction of any exogenous chemical substances, fundamentally avoiding the risk of drug-related systemic toxic side effects (such as hepatotoxicity and nephrotoxicity). At the same time, as a non-ionizing radiation physical factor, the strong magnetic field has excellent tissue penetration and recognized high biological safety, laying a solid foundation for the development of a safe, non-invasive, and highly compliant clinical device.

[0028] This invention achieves a leap in treatment strategy from "indirectly regulating cells" to "directly reconstructing the matrix" through a simple and physically clear path of "applying a strong magnetic field → generating a targeted magnetic field mechanical effect (magnetic torque) → directly reconstructing collagen fibers". It provides a device for a breakthrough treatment of liver fibrosis with a novel mechanism, clear targeting, potential for rapid onset of action and high safety. Attached Figure Description

[0029] Figure 1 To display the spatial distribution direction of collagen fibers during the process of liver fibrosis in a clinical staging system; Figure 2 Diagram showing how a steady-state magnetic field promotes the alignment of collagen perpendicular to the magnetic field direction; Figure 3 These are physical images of the control group and the magnetic field treatment group of this invention; Figure 4 This is a schematic diagram of the control group and the magnetic field treatment group of the present invention; Figure 5 This is a diagram for detecting the degree of liver fibrosis according to the present invention; wherein, (A) images of Sirius red and α-SMA staining; (B) statistical results of the relative amount of the ratio of the area of ​​positive area stained with Sirius red to the total area; (C) statistical results of α-SMA fluorescence intensity; (D) statistical results of liver hydroxyproline content; n=6 for each group; P<0.05, P<0.01, P<0.001, P<0.0001; Figure 6 This invention relates to the detection of physical properties of collagen fibers; wherein, (A) images of COL1A1 staining and images of collagen fibers extracted from the images using CTFire software; (B) statistical analysis results of fiber width; (C) statistical analysis results of fiber length; (D) statistical analysis results of fiber straightness; (E) magnetic anisotropy signal of type I collagen gel formed under conditions with and without a magnetic field, detected using a comprehensive physical property measurement system (PPMS); n=6 for each group in B and D; P<0.05, P<0.01, P<0.001, P<0.0001. Detailed Implementation

[0030] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Unless otherwise defined, the technical terms used below have the same meaning as understood by those skilled in the art.

[0031] Unless otherwise specified, the test materials and reagents used in the following examples are commercially available or prepared by known methods.

[0032] Example 1 Strong steady-state magnetic fields improve liver fibrosis (1) A strong magnetic field device (i.e., the device disclosed in the application document with publication number CN111304085A, entitled "A biological sample culture device suitable for static magnetic field environment and its application"). For example... Figure 3 As shown, a superconducting magnet was used to generate steady-state strong magnetic fields of 1.5T, 3T, 7T, and 9.4T. Mice were placed at the center of the magnet using an aluminum alloy frame.

[0033] Mice were placed in the center of a superconducting magnet using an aluminum alloy mouse cage, with only positions 1, 2, and 3 in the middle used. These positions generate a relatively uniform magnetic field, which can be varied from 1.5 T to 9.4 T by adjusting the intensity. Mice in the control group (sham magnetization group) were also placed in an aluminum alloy cage within a sleeve without a magnetic field to eliminate the influence of factors other than magnetic field strength. Both groups of mice were ventilated after placement to ensure normal vital functions. Figure 3 and 4 As shown; (2) Establishment of a mouse model of liver fibrosis. The mice with liver fibrosis were induced by repeated intraperitoneal injections of carbon tetrachloride, which was dissolved in corn oil (volume ratio 1:10) and injected twice a week at a dose of 0.5 ml / kg for eight weeks.

[0034] (3) Magnetic field treatment conditions. Magnetic field treatment is performed twice a week (Monday and Thursday), for 5 hours each time, for eight weeks, for a total of 80 hours.

[0035] (4) Detection of liver fibrosis degree. We performed Sirius red staining and α-SMA immunohistochemical staining on mouse livers. Figure 5 As shown in Figure A), the former is widely used to study the distribution and content of collagen fibers in histology and pathology, while the latter is a marker of hepatic stellate cell activation; both can reflect the degree of liver fibrosis. From the results of Sirius red staining, long-term administration of CCl4 significantly increased liver fibrosis in mice, with a significant increase in the stained area. Meanwhile, steady-state magnetic fields of 1.5T, 3T, 7T, and 9.4T all effectively reduced collagen fiber content and alleviated the degree of liver fibrosis in mice. Figure 5 (B). Statistical results of the α-SMA positive region area also verified the anti-fibrotic effect of the strong steady-state magnetic field. Figure 5 (C). The content of hydroxyproline in the liver also decreased significantly after treatment with a strong magnetic field, and showed a magnetic field strength dependence (C). Figure 5 (D).

[0036] (5) Detection of physical properties of collagen fibers. Immunofluorescence results of liver COL1A1 showed that type I collagen fibers in the sham magnetic exposure group had an irregular polygonal structure, consistent with previous reports. However, in the magnetic field treatment group, this polygonal structure was destroyed, and the stronger the magnetic field, the more obvious the tendency of collagen fibers to align in a certain direction. Figure 6 (A) This indicates that magnetic fields can disrupt the pathological collagen fiber network structure. Individual collagen fibers were extracted from the images using CT Fire software. Figure 6 (A) and quantitative analysis were performed. The results showed that the width, length, and straightness of the fibrous tissue in the liver of the steady-state strong magnetic field treatment group decreased, while the fibrous tissue length decreased and the straightness increased. Figure 6(BD). All the above results show that a steady-state strong magnetic field can directly affect the physical properties of collagen fibers. In in vitro experiments, we used type I rat tail collagen solution, which was diluted and neutralized to construct a collagen gel model (i.e., type I collagen (Corning, 354249) was diluted to 3 mg / ml with 1×PBS and the pH was adjusted to neutral with NaOH). The gels were placed in a magnetic field-free environment and a 7T steady-state magnetic field, respectively, and treated at 37℃ for 2 hours. The magnetic anisotropy of the two groups of samples was then precisely detected using a physical property measurement system (PPMS). The results showed that the collagen fibers self-assembled in the magnetic field-free environment had a weaker magnetic anisotropy signal, indicating that the fiber arrangement was random and the degree of magnetic orientation was low; in contrast, the collagen fibers assembled after treatment with a 7T magnetic field showed a significant magnetic anisotropy enhancement effect (…). Figure 6 The results (E) fully demonstrate that the magnetic field can effectively change the orientation of collagen fibers and significantly enhance their macroscopic magnetic orientation characteristics.

[0037] Current technologies lack direct physical intervention methods for pathological collagen fibers. Existing drug and magnetic field biological effect therapies indirectly affect collagen metabolism by regulating cell function, but cannot apply direct mechanical action to the deposited and cross-linked collagen fiber network to change its arrangement or integrity. This invention provides a non-drug method that can actively and targetedly reconstruct the structure of pathological collagen fibers. Compared to existing technologies that can only passively wait for cell response and matrix remodeling, this is a physical method that can actively apply external force to directly and precisely act on the target collagen fibers, causing them to undergo orientation changes, depolymerization, or reconstruction.

[0038] This invention relates to a technique for non-contact physical energy-targeted intervention in pathological tissues. Specifically, it involves a method and related equipment that utilizes the biomechanical effects of a strong magnetic field to directly act on the extracellular matrix, physically reconstructing and ablating abnormally deposited and arranged collagen fibers in fibrotic diseases. The fundamental problem it aims to solve is: how to transcend the traditional "cell regulation" approach and achieve "direct physical reconstruction" of the core pathological product of liver fibrosis—the pathological collagen fiber network.

[0039] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit it. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. The application of a magnetic field in the preparation of a device for the treatment of liver fibrosis.

2. The application of the magnetic field according to claim 1 in the preparation of a device for the treatment of liver fibrosis, characterized in that, The magnetic field strength is 1.5-9.4T.

3. The application of the magnetic field according to claim 1 or 2 in the preparation of a device for the treatment of liver fibrosis, characterized in that, Magnetic fields can alleviate liver fibrosis by reducing the content of collagen fibers in the liver and / or lowering the content of hydroxyproline in the liver and / or disrupting the pathological collagen fiber network structure.

4. The application of the magnetic field according to claim 1 or 2 in the preparation of a device for the treatment of liver fibrosis, characterized in that, The magnetic field acts directly on the collagen fibers in the liver of humans and / or animals, inducing physical changes in the collagen fiber structure.

5. The application of a magnetic field in studying the treatment mechanism of liver fibrosis or in constructing a mouse model of liver fibrosis.

6. The application of the magnetic field according to claim 5 in studying the therapeutic mechanism of liver fibrosis or in constructing a mouse model of liver fibrosis, characterized in that, The magnetic field strength is 1.5-9.4T.

7. The application of the magnetic field according to claim 5 in studying the therapeutic mechanism of liver fibrosis or in constructing a mouse model of liver fibrosis, characterized in that, The application includes exposing animals with liver fibrosis to the magnetic field.

8. The application of the magnetic field according to claim 7 in studying the therapeutic mechanism of liver fibrosis or in constructing a mouse model of liver fibrosis, characterized in that, The method for constructing animals with liver fibrosis includes the following steps: injecting the animals intraperitoneally with a corn oil solution containing carbon tetrachloride twice a week for eight weeks.

9. The application of the magnetic field according to claim 8 in studying the therapeutic mechanism of liver fibrosis or in constructing a mouse model of liver fibrosis, characterized in that, The animals were mice; the mice were injected intraperitoneally with a corn oil solution containing carbon tetrachloride at a dose of 0.5 ml / kg.

10. The application of the magnetic field according to any one of claims 5-9 in studying the therapeutic mechanism of liver fibrosis or in constructing a mouse model of liver fibrosis, characterized in that, The applications include, after the magnetic field exposure has ended, obtaining liver tissue from the subject and assessing the degree of liver fibrosis by Sirius red staining and / or α-SMA immunohistochemical staining; and / or, obtaining liver tissue from the subject and assessing the arrangement trend of liver collagen fibers by COL1A1 immunofluorescence detection; and / or, using CT Fire software to extract individual collagen fibers from the images and perform quantitative analysis to assess the physical properties of collagen fibers.

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