A device for histopathological evaluation of the degree of healing of bone lesions

Abstract

The application discloses a histopathological evaluation device for the healing degree of bone injury, and relates to the technical field of medical devices, pharmacodynamics and pathological diagnosis. The device comprises a microscope configured to capture an image of a stained bone tissue section; an image processing unit connected to the microscope and configured to obtain semi-quantitative histological measurement results and microscopic bone histomorphometry measurement results based on the image of the stained bone tissue section; the semi-quantitative histological measurement results comprise the state of callus bridging and fracture line, the composition and maturity of callus, new bone formation activity and bone remodeling and structure recovery state; the microscopic bone histomorphometry measurement results comprise the bone volume fraction and the proportion of osteoid surface; and a host computer connected to the image processing unit and configured to generate the bone injury healing condition according to the two kinds of measurement results. The application combines semi-quantitative histological measurement and microscopic bone histomorphometry measurement, and can provide an objective and quantifiable statistical device and method for the healing effect of bone injury.

Description

Technical Field

[0001] This application relates to the fields of medical devices, pharmacodynamics and pathological diagnostics, and in particular to a histopathological evaluation device for the degree of bone injury healing. Background Technology

[0002] Fracture healing is a complex biological process, and accurate evaluation of its degree of healing is crucial in the development of novel bone repair materials, pharmacodynamic assessments, and animal trials related to safety evaluations. Currently, evaluation methods mainly include imaging (such as X-rays and CT scans) and histopathological assessments.

[0003] Imaging methods involve radiation and are difficult to display early and microscopic changes. Histopathology is the gold standard for evaluating fracture damage and repair, but traditional evaluations rely heavily on the subjective experience of pathologists or simple single-indicator measurements, which have the following drawbacks: (1) strong subjectivity and large differences between different observers; (2) lack of unified and comprehensive quantitative standards, making it difficult to conduct accurate longitudinal comparisons and statistical analyses; (3) inability to fully utilize the multidimensional information in the slides (such as cell activity, matrix composition, dynamic formation rate, etc.) for comprehensive judgment.

[0004] Therefore, a new method for evaluating the degree of bone injury healing is needed to address the problems of strong subjectivity and lack of standardized quantitative standards in the current fracture healing evaluation, and to achieve a more accurate, repeatable, multi-dimensional, and high-throughput evaluation. Summary of the Invention

[0005] The purpose of this application is to provide a histopathological evaluation device for the degree of bone injury healing, which, combined with semi-quantitative histological scoring or microscopic bone morphometric scoring, can provide an objective and quantifiable statistical device and method for assessing the healing effect of bone injury.

[0006] To achieve the above objectives, this application provides the following solution: This application provides a histopathological evaluation device for the degree of bone injury healing, comprising: A microscope is used to take images of stained bone tissue sections. An image processing unit, connected to the microscope, is used to obtain semi-quantitative histological measurement results and microscopic bone morphometric measurement results based on images of the stained bone tissue sections. The semi-quantitative histological measurements include: callus bridging and fracture line status, callus composition and maturity, new bone formation activity, and bone remodeling and structural recovery status. The microscopic bone morphometric measurement results include: bone volume fraction and osteoid surface area ratio. A host computer, connected to the image processing unit, is used to generate the degree of bone injury healing based on the semi-quantitative histological measurement results and / or the microscopic bone morphometric measurement results.

[0007] The status of callus bridging and fracture line is classified into four levels: level 1 is no connection, level 2 is fibrous connection, level 3 is partial bony bridging, and level 4 is complete bony bridging. The composition and maturity of the callus are divided into four levels: the first level is mainly composed of hematoma granulation tissue, the second level is mainly composed of fibrocartilage, the third level is mainly composed of woven bone, and the fourth level is mainly composed of lamellar bone. The new bone formation activity is divided into four levels: the first level is no activity, the second level is local weak activity, the third level is widespread continuous activity, and the fourth level is widespread activity with medullary cavity recanalization. Bone remodeling and structural recovery status includes four levels: Level 1 is no remodeling, Level 2 is initial absorption, Level 3 is significant coupled remodeling, and Level 4 is basic structural recovery.

[0008] Specifically, based on the semi-quantitative histological measurements, the degree of bone injury healing is generated, including: Calculate the sum of the grade values ​​for callus bridging and fracture line status, callus composition and maturity, new bone formation activity, and bone remodeling and structural restoration; If the total score is 13-16, it indicates that the bone injury has basically healed. If the total score is 5-12, it indicates that the bone injury healing has entered the middle stage; If the total score is 4, it indicates that the bone injury is in the early stage of healing or that there is a healing obstacle.

[0009] The bone volume fraction = mineralized bone matrix area / fracture area; the osteoid surface area ratio = osteoid area / fracture area.

[0010] Based on the microscopic bone morphometric measurements, the degree of bone injury healing is generated, specifically including: Multiple images of the same area that completely contain the bone injury were selected using image processing software. Calculate the bone volume fraction and osteoid surface area for each image; If both the bone volume fraction and the osteoid surface area ratio follow a normal distribution and their variances are within the preset range, then perform t-tests on the bone volume fraction and osteoid surface area ratio of stained bone tissue sections and the bone volume fraction and osteoid surface area ratio of healthy bone tissue sections, respectively; otherwise, perform Mann-Whitney U tests on the bone volume fraction and osteoid surface area ratio of stained bone tissue sections and the bone volume fraction and osteoid surface area ratio of healthy bone tissue sections, respectively. Calculate the p-value of the t-test or Mann-Whitney U test; if p ≤ 0.01, it indicates that the bone injury is in the early stage of healing or there is a healing obstacle; if p < 0.01 ≤ 0.05, it indicates that the bone injury is in the middle stage of healing; if p > 0.05, it indicates that the bone injury is basically healed.

[0011] In one embodiment of this application, the histopathological evaluation device for the degree of bone injury healing further includes: a microtome for slicing the bone tissue to be tested to obtain bone tissue slices.

[0012] Before sectioning the bone tissue to be tested, the following steps are also taken: the bone tissue to be tested is decalcified, trimmed and collected, dehydrated with alcohol and embedded in paraffin.

[0013] In one embodiment of this application, the histopathological evaluation device for the degree of bone injury healing further includes: a section staining device for staining bone tissue sections to obtain stained bone tissue sections.

[0014] Methods for staining bone tissue sections include one or more of the following: Masson-Goldner trichrome staining, H&E staining, Safranin O Fast Green staining, and TRAP staining.

[0015] The image processing software includes SPSS and / or Image J and / or GraphPad Prism.

[0016] According to the specific embodiments provided in this application, the following technical effects are disclosed: This application provides a histopathological evaluation device for the degree of bone injury healing. The device includes: a microscope for capturing images of stained bone tissue sections; an image processing unit connected to the microscope for obtaining semi-quantitative histological measurements and microscopic bone morphometric measurements based on the images of the stained bone tissue sections; the semi-quantitative histological measurements include: callus bridging and fracture line status, callus composition and maturity, new bone formation activity, and bone remodeling and structural recovery status; the microscopic bone morphometric measurements include: bone volume fraction and osteoid surface area ratio; and a host computer connected to the image processing unit for generating bone injury healing status based on the above two measurement results. This application, combining semi-quantitative histological measurements and microscopic bone morphometric measurements, provides an objective and quantifiable statistical device and method for assessing the effectiveness of bone injury healing. Attached Figure Description

[0017] To more clearly illustrate the technical solutions in the embodiments of this application or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0018] Figure 1 This is a schematic diagram of the connection relationship of a histopathological evaluation device for assessing the degree of bone injury healing in one embodiment of this application.

[0019] Figure 2 This is a flowchart illustrating the method steps corresponding to a histopathological evaluation device for assessing the degree of bone injury healing in one embodiment of this application.

[0020] Figure 3 This is a schematic diagram of the structure of a computer device provided in an embodiment of this application. Detailed Implementation

[0021] The technical solutions of the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of this application, and not all embodiments. Based on the embodiments of this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application.

[0022] To make the above-mentioned objectives, features and advantages of this application more apparent and understandable, the application will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0023] This application provides a histopathological evaluation device for the degree of bone injury healing, such as... Figure 1 As shown, it includes: A microscope is used to take images of stained bone tissue sections. An image processing unit, connected to the microscope, is used to obtain semi-quantitative histological measurement results and microscopic bone morphometric measurement results based on images of the stained bone tissue sections. The semi-quantitative histological measurements include: callus bridging and fracture line status, callus composition and maturity, new bone formation activity, and bone remodeling and structural recovery status. The microscopic bone morphometric measurement results include: bone volume fraction and osteoid surface area ratio. A host computer, connected to the image processing unit, is used to generate the degree of bone injury healing based on the semi-quantitative histological measurement results and / or the microscopic bone morphometric measurement results.

[0024] The callus bridging and fracture line status are classified into four levels: the first level is no connection, the second level is fibrous connection, the third level is partial bony bridging, and the fourth level is complete bony bridging. The composition and maturity of the callus are divided into four levels: the first level is mainly composed of hematoma granulation tissue, the second level is mainly composed of fibrocartilage, the third level is mainly composed of woven bone, and the fourth level is mainly composed of lamellar bone. The new bone formation activity is divided into four levels: the first level is no activity, the second level is local weak activity, the third level is widespread continuous activity, and the fourth level is widespread activity with medullary cavity recanalization. Bone remodeling and structural recovery status includes four levels: Level 1 is no remodeling, Level 2 is initial absorption, Level 3 is significant coupled remodeling, and Level 4 is basic structural recovery.

[0025] Specifically, the absence of connection between callus bridging and the fracture line refers to the complete separation of the fracture ends, with a clearly visible gap filled with hematoma remnants, inflammatory cells, or loose fibrous tissue, without any continuous fibrous tissue, bony, or cartilaginous tissue crossing the gap. Fiber connection refers to the gap being partially filled and connected by dense fibrous or fibrocartilaginous tissue, forming a continuous "soft tissue band" connecting the two sides of the fracture ends. Partial bony bridging means that more than half of the gap width has been bridged by newly formed mineralized bone tissue (woven bone or early lamellar bone). The bridging may be incomplete or uneven in thickness, and there may be a small amount of unclosed fibrous / cartilaginous areas. Complete bony bridging means that the fracture gap is completely and continuously bridged by new bone (lamellar or woven bone), forming a stable and continuous structure.

[0026] The composition and maturity of the callus are described as follows: Predominantly hematoma / granulation tissue means the callus area is mainly composed of unorganized erythrocytes, fibrinous exudate, numerous inflammatory cells (neutrophils, macrophages), and early proliferating capillaries (granulation tissue). Orderly extracellular matrix deposition is almost invisible. Predominantly fibrous / cartilage means the callus is primarily composed of dense fibrous or cartilage tissue. The fibrous tissue is disordered, cells are spindle-shaped, and mineralized bone content is minimal or scattered in island-like patterns. Predominantly woven bone means the callus is mainly composed of newly formed woven bone. The trabeculae of the woven bone are irregularly arranged and coarse, osteocytes are disordered, and the bone matrix stains unevenly (strongly basophilic). The osteoid margin (osteoblast rim) may be very active and thick. Cartilage and fibrous tissue have been largely replaced or encapsulated. Predominantly lamellar bone means the callus has undergone significant remodeling, with lamellar bone becoming the main component. The lamellar bone structure is orderly, the bone cells are arranged in a regular manner, and the collagen fibers are arranged in layers (showing alternating bright and dark birefringent stripes under polarized light).

[0027] Inactive bone formation refers to the absence of active osteoblasts within the callus region. The bone surface is smooth, without a osteoid layer, or only a very thin layer of inactive osteoid. No bone lining cells or osteoblast arrangement is observed. Localized weak activity refers to the presence of discontinuous, single-layered, flattened osteoblast arrangements with a thin layer of osteoid deposition only in a few areas (such as near the edge of the fracture). Most of the newly formed bone surface shows no signs of cellular activity. Extensive continuous activity refers to the presence of a continuous, abundant layer of cubic osteoblasts covering the surface of numerous newly formed bone trabeculae throughout the callus region, beneath which lies a significant, homogeneous osteoid layer. Extensive activity with medullary cavity recanalization refers to the presence of "extensive continuous activity" while a new medullary cavity has begun to form or has completely formed in the central region of the callus. The cavity is filled with hematopoietic cells or adipocytes.

[0028] The terms "no remodeling" and "structural restoration" refer to a state where the callus is quiescent with no signs of remodeling. Osteoclasts and their resorption lacunae are not visible. The new bone morphology is primitive (e.g., large woven bone), and no modification has begun. "Initial resorption" refers to the beginning of localized osteoclast-induced bone resorption. A small number of multinucleated osteoclasts are visible attached to the bone surface, forming shallow resorption lacunae. "Clearly coupled remodeling" refers to a close coupling between bone resorption and bone formation. Osteoclast-induced resorption lacunae are spatially adjacent to the osteoblasts and osteoid layer following closely around the new bone. The trabecular bone morphology changes from coarse to fine and regular. "Basic structural restoration" means the remodeling process is nearly complete, and the bone structure is nearly normal. Most of the original callus has been absorbed, and the medullary cavity is completely reopened and enlarged. The cortical bone region restores a dense, continuous lamellar bone structure with clear Haversian canals. The trabecular bone region has moderate trabecular thickness, and its orientation is consistent with the direction of mechanical load, exhibiting a normal cancellous bone structure.

[0029] Specifically, the bone volume fraction (BV / TV) = mineralized bone matrix area / fracture area; the osteoid surface ratio (OS / BS) = osteoid area / fracture area.

[0030] The semi-quantitative total score of the experimental group (such as the group using a certain bone repair material) was significantly increased, and the BV / TV and OS / BS values ​​were not significantly different from those of healthy bone tissue, indicating that the material significantly promoted fracture healing.

[0031] Specifically, based on the semi-quantitative histological measurements, the degree of bone injury healing is generated, including: Calculate the sum of the grade values ​​for callus bridging and fracture line status, callus composition and maturity, new bone formation activity, and bone remodeling and structural restoration; If the total score is 13-16, it indicates that the bone injury has basically healed; it is characterized by a firm bony bridging, high callus maturity (mainly lamellar bone), recanalization of the medullary cavity, obvious bone remodeling, and the structure is close to or restored to normal.

[0032] A total score of 5-12 indicates that bone injury healing has entered the intermediate stage. Specifically, a total score of 5-8 indicates early to mid-stage bone injury healing, with only fibrous connections or a small amount of bony connections, and the callus still mainly composed of fibrocartilage or immature woven bone. A total score of 9-12 indicates mid to late-stage bone injury healing, with partial or complete bony bridging, the callus mainly composed of woven bone and transforming into lamellar bone, with extensive new bone formation activity, and remodeling has begun.

[0033] If the total score is 4, it indicates that the bone injury is in the early stage of healing or that there is a healing disorder. There is a lack of callus bridging, with hematoma or granulation tissue dominating, and almost no osteogenic activity.

[0034] Specifically, based on the microscopic bone morphometric measurement results, the degree of bone injury healing is generated, including: Multiple images of the same area that completely contain the bone injury were selected using image processing software. Calculate the bone volume fraction and osteoid surface area for each image; If both the bone volume fraction and the osteoid surface area ratio follow a normal distribution and their variances are within the preset range, then perform t-tests on the bone volume fraction and osteoid surface area ratio of stained bone tissue sections and the bone volume fraction and osteoid surface area ratio of healthy bone tissue sections, respectively; otherwise, perform Mann-Whitney U tests on the bone volume fraction and osteoid surface area ratio of stained bone tissue sections and the bone volume fraction and osteoid surface area ratio of healthy bone tissue sections, respectively. Calculate the p-value of the t-test or Mann-Whitney U test; if p ≤ 0.01, it indicates that the bone injury is in the early stage of healing or there is a healing obstacle; if p < 0.01 ≤ 0.05, it indicates that the bone injury is in the middle stage of healing; if p > 0.05, it indicates that the bone injury is basically healed.

[0035] For example, bone volume fraction and osteoid surface area percentage should first be tested for homogeneity (Shapiro-Wilk test or Kolmogorov-Smirnov test) and homogeneity of variance (F-test or Levene test). If the data are normally distributed and homogeneous in variance, an independent samples t-test is performed; if the data are not normally distributed and / or homogeneous in variance, a Mann-Whitney U test is performed. The Mann-Whitney U test can also be used for intergroup comparisons of semi-quantitative histological scores.

[0036] In one embodiment of this application, the histopathological evaluation device for the degree of bone injury healing further includes: a microtome for slicing the bone tissue to be tested to obtain bone tissue slices.

[0037] Before sectioning the bone tissue to be tested, the following steps are also taken: the bone tissue to be tested is decalcified, trimmed and sampled, dehydrated with alcohol and embedded in paraffin.

[0038] In one embodiment of this application, the histopathological evaluation device for the degree of bone injury healing further includes: a section staining device for staining bone tissue sections to obtain stained bone tissue sections, so as to distinguish mineralized bone, osteoid and cartilage / fibrous tissue.

[0039] Methods for staining bone tissue sections include one or more of the following: Masson-Goldner trichrome staining, H&E staining, Safranin O Fast Green staining, and TRAP staining.

[0040] Among them, 1. Masson-Goldner Trichrome Stain: A special multicolor staining method that uses multiple dyes (such as Acid Fuchsin, Aniline Blue, Orange G, etc.) to sequentially stain tissue. It can stain different components with distinctly contrasting colors, hence the name "trichrome method". This method was first proposed by Masson and later improved by Goldner, hence the name Masson-Goldner trichrome method. In bone histology, this staining method is mainly used to clearly distinguish the matrix components of different stages of bone tissue maturity, and is an important tool for studying bone formation and bone remodeling. In Masson-Goldner trichrome staining of bone tissue, different bone tissue components appear in different colors: fibrous tissue / early fibrous callus: green / blue-green; osteoid / new bone matrix: bright red or orange-red; mature bone: green; cell nuclei: blue-black or purplish-black; cartilage: blue or blue-green.

[0041] 2. Hematoxylin and Eosin Staining (H&E): This is the most basic and commonly used routine staining method in histology and pathology. Hematoxylin is a basic dye that primarily stains acidic substances such as cell nuclei; eosin is an acidic dye that primarily stains basic substances such as cytoplasm and collagen fibers. Morphological description in bone tissue: H&E staining provides basic morphological and structural information about bone tissue, used to assess the overall structure, cell distribution, and morphology of bone tissue, and is the preferred initial screening method for observing bone tissue structure.

[0042] 3. Safranin O-Fast Green Stain: This is a classic staining combination used to distinguish between cartilage and bone. Safranin O is a cationic dye with a strong affinity for cartilage matrix rich in anionic glycosaminoglycans such as chondroitin sulfate; Fast Green, on the other hand, binds well to the weakly acidic mineralized bone matrix.

[0043] Different bone tissue components exhibit different colors in Safranin O-Fixed Green staining: hyaline cartilage / cartilage matrix: bright red or dark red, mineralized bone matrix: green or blue-green, calcified cartilage: pale purple or transitional color, cell nuclei: usually dark blue or black.

[0044] 4. TRAP staining (Tartrate-Resistant Acid Phosphatase Stain): This staining is a key method for specifically identifying and labeling osteoclasts and their activity. Positive osteoclasts: On the bone surface (especially in the Howship lacunae), one or more cytoplasm cells containing numerous bright red granules or diffusely bright red multinucleated giant cells can be seen. The nuclei are generally unstained or pale blue (depending on counterstaining). Negative cells: Osteoblasts, osteocytes, bone marrow cells, etc., do not show red staining. Bone resorption surface: The bone margins where positive osteoclasts attach often exhibit irregular bone surface morphology.

[0045] The image processing software includes SPSS and / or Image J and / or GraphPad Prism.

[0046] Correspondingly, such as Figure 2 As shown, the experimental steps corresponding to the device in this application are as follows: The method includes the following steps: S1: The negative control group (normal healthy animal group) and the experimental group (if a certain bone repair material is used) were evaluated. The bone tissue of the animals was decalcified with decalcification solution, trimmed into blocks and taken out, dehydrated in stages with alcohol, embedded in paraffin, sliced ​​with a microtome (about 3 μm thick), spread, picked up and dried with a slide spreader to prepare bone tissue sections. S2: The sections are stained with at least the Masson-Goldner trichrome staining method to distinguish mineralized bone, osteoid and cartilage / fibrous tissue. The staining method also includes one or more of H&E staining, Safranin O Fast Green staining and / or TRAP staining. S3: Based on the stained sections, perform semi-quantitative histological scoring, which includes at least the following four evaluation indicators: callus bridging and fracture line status (A), callus composition and maturity (B), new bone formation activity (C), and bone remodeling and structural restoration (D).

[0047] S4: Based on the stained sections, microscopic bone morphometric measurements can be selectively performed using image analysis software. These measurements include at least bone volume fraction (BV / TV) and osteoid surface area ratio (OS / BS). Statistical analysis compared with the negative control group showed that the experimental group animals had significantly higher semi-quantitative total scores (total scores in S3), and BV / TV and OS / BS values ​​were not significantly different from those of healthy bone tissue, indicating that the test material significantly promoted fracture healing.

[0048] S5: Based on the semi-quantitative histological scores and / or microscopic bone morphometric measurements, generate a comprehensive evaluation report on the degree and / or effect of fracture healing of the test material.

[0049] This invention combines semi-quantitative histological scoring and statistical analysis of microscopic bone morphometric data to comprehensively quantify and score the status of callus bridging and fracture lines, callus formation maturity, new bone formation activity, and degree of bone remodeling. It can also optionally integrate dynamic bone morphometric data. This provides an objective and quantifiable statistical evaluation method for assessing fracture repair degree and / or healing effect in the research and development, pharmacodynamic evaluation, and non-clinical safety evaluation of orthopedic medical devices. Histopathological examination is the gold standard for evaluating tissue morphology and detecting product efficacy. This invention solves the problems of strong subjectivity and lack of standardized quantitative standards in existing fracture healing evaluation technologies, enabling more accurate, repeatable, and multi-dimensional high-throughput evaluation, especially suitable for new drug development, bone repair material evaluation, and clinical prognosis assessment.

[0050] In one exemplary embodiment, a computer device is provided, which may be a server or a terminal, and its internal structure diagram may be as follows. Figure 3 As shown, this computer device includes a processor, memory, input / output (I / O) interfaces, and a communication interface. The processor, memory, and I / O interfaces are connected via a system bus, and the communication interface is also connected to the system bus via the I / O interfaces. The processor provides computational and control capabilities. The memory includes non-volatile storage media and internal memory. The non-volatile storage media stores the operating system, computer programs, and databases. The internal memory provides the environment for the operating system and computer programs stored in the non-volatile storage media to run. The I / O interfaces are used for exchanging information between the processor and external devices. The communication interface is used for communicating with external terminals via a network connection.

[0051] Those skilled in the art will understand that Figure 3The structure shown is merely a block diagram of a portion of the structure related to the present application and does not constitute a limitation on the computer device to which the present application is applied. Specific computer devices may include more or fewer components than those shown in the figure, or combine certain components, or have different component arrangements.

[0052] In one exemplary embodiment, a computer device is also provided, including a memory and a processor, wherein the memory stores a computer program, and the processor executes the computer program to implement the steps in the above-described method embodiments.

[0053] In one exemplary embodiment, a computer-readable storage medium is provided storing a computer program that, when executed by a processor, implements the steps in the above-described method embodiments.

[0054] In one exemplary embodiment, a computer program product is provided, including a computer program that, when executed by a processor, implements the steps in the above-described method embodiments.

[0055] It should be noted that the user information (including but not limited to user device information, user personal information, etc.) and data (including but not limited to data used for analysis, data stored, data displayed, etc.) involved in this application are all information and data authorized by the user or fully authorized by all parties, and the collection, use and processing of the relevant data must comply with relevant regulations.

[0056] Those skilled in the art will understand that all or part of the processes in the above embodiments can be implemented by a computer program instructing related hardware. The computer program can be stored in a non-volatile computer-readable storage medium. When executed, the computer program can include the processes of the embodiments described above. Any references to memory, databases, or other media used in the embodiments provided in this application can include at least one of non-volatile and volatile memory. Non-volatile memory can include read-only memory (ROM), magnetic tape, floppy disk, flash memory, optical memory, high-density embedded non-volatile memory, resistive random access memory (ReRAM), magnetic random access memory (MRAM), ferroelectric random access memory (FRAM), phase change memory (PCM), graphene memory, etc. Volatile memory can include random access memory (RAM) or external cache memory, etc. By way of illustration and not limitation, RAM can take many forms, such as Static Random Access Memory (SRAM) or Dynamic Random Access Memory (DRAM).

[0057] The databases involved in the embodiments provided in this application may include at least one type of relational database and non-relational database. Non-relational databases may include, but are not limited to, blockchain-based distributed databases. The processors involved in the embodiments provided in this application may be general-purpose processors, central processing units, graphics processing units, digital signal processors, programmable logic devices, quantum computing-based data processing logic devices, etc., and are not limited to these.

[0058] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0059] This document uses specific examples to illustrate the principles and implementation methods of this application. The descriptions of the above embodiments are only for the purpose of helping to understand the methods and core ideas of this application. Furthermore, those skilled in the art will recognize that, based on the ideas of this application, there will be changes in the specific implementation methods and application scope. Therefore, the content of this specification should not be construed as a limitation of this application.

Claims

1. A histopathological evaluation device for the degree of bone injury healing, characterized in that, The histopathological evaluation device for the degree of bone injury healing includes: A microscope is used to take images of stained bone tissue sections. An image processing unit, connected to the microscope, is used to obtain semi-quantitative histological measurement results and microscopic bone morphometric measurement results based on images of the stained bone tissue sections. The semi-quantitative histological measurements include: callus bridging and fracture line status, callus composition and maturity, new bone formation activity, and bone remodeling and structural recovery status. The microscopic bone morphometric measurement results include: bone volume fraction and osteoid surface area ratio. A host computer, connected to the image processing unit, is used to generate the degree of bone injury healing based on the semi-quantitative histological measurement results and / or the microscopic bone morphometric measurement results.

2. The histopathological evaluation device for the degree of bone injury healing according to claim 1, characterized in that, The status of callus bridging and fracture line is classified into four levels: level 1 is no connection, level 2 is fibrous connection, level 3 is partial bony bridging, and level 4 is complete bony bridging. The composition and maturity of the callus are divided into four levels: the first level is mainly composed of hematoma granulation tissue, the second level is mainly composed of fibrocartilage, the third level is mainly composed of woven bone, and the fourth level is mainly composed of lamellar bone. The new bone formation activity is divided into four levels: the first level is no activity, the second level is local weak activity, the third level is widespread continuous activity, and the fourth level is widespread activity with medullary cavity recanalization. Bone remodeling and structural recovery status includes four levels: Level 1 is no remodeling, Level 2 is initial absorption, Level 3 is significant coupled remodeling, and Level 4 is basic structural recovery.

3. The histopathological evaluation device for the degree of bone injury healing according to claim 2, characterized in that, Based on the semi-quantitative histological measurements, the degree of bone injury healing is calculated, specifically including: Calculate the sum of the grade values ​​for callus bridging and fracture line status, callus composition and maturity, new bone formation activity, and bone remodeling and structural restoration; If the total score is 13-16, it indicates that the bone injury has basically healed. If the total score is 5-12, it indicates that the bone injury healing has entered the middle stage; If the total score is 4, it indicates that the bone injury is in the early stage of healing or that there is a healing obstacle.

4. The histopathological evaluation device for the degree of bone injury healing according to claim 1, characterized in that, The bone volume fraction = mineralized bone matrix area / fracture area; the osteoid surface area ratio = osteoid area / fracture area.

5. The histopathological evaluation device for the degree of bone injury healing according to claim 1, characterized in that, Based on the microscopic bone morphometric measurements, the degree of bone injury healing is generated, specifically including: Multiple images of the same area that completely contain the bone injury were selected using image processing software. Calculate the bone volume fraction and osteoid surface area for each image; If both the bone volume fraction and the osteoid surface area ratio follow a normal distribution and their variances are within the preset range, then perform t-tests on the bone volume fraction and osteoid surface area ratio of stained bone tissue sections and the bone volume fraction and osteoid surface area ratio of healthy bone tissue sections, respectively; otherwise, perform Mann-Whitney U tests on the bone volume fraction and osteoid surface area ratio of stained bone tissue sections and the bone volume fraction and osteoid surface area ratio of healthy bone tissue sections, respectively. Calculate the p-value of the t-test or Mann-Whitney U test; if p ≤ 0.01, it indicates that the bone injury is in the early stage of healing or there is a healing obstacle; if p < 0.01 ≤ 0.05, it indicates that the bone injury is in the middle stage of healing; if p > 0.05, it indicates that the bone injury is basically healed.

6. The histopathological evaluation device for the degree of bone injury healing according to claim 1, characterized in that, The histopathological evaluation device for the degree of bone injury healing also includes a microtome for slicing the bone tissue to be tested to obtain bone tissue slices.

7. The histopathological evaluation device for the degree of bone injury healing according to claim 1, characterized in that, Before sectioning the bone tissue to be tested, the following steps are also taken: the bone tissue to be tested is decalcified, trimmed and collected, dehydrated with alcohol and embedded in paraffin.

8. The histopathological evaluation device for the degree of bone injury healing according to claim 1, characterized in that, The histopathological evaluation device for the degree of bone injury healing also includes: a section staining device for staining bone tissue sections to obtain stained bone tissue sections.

9. The histopathological evaluation device for the degree of bone injury healing according to claim 8, characterized in that, Methods for staining bone tissue sections include one or more of the following: Masson-Goldner trichrome staining, H&E staining, Safranin O Fast Green staining, and TRAP staining.

10. The histopathological evaluation device for the degree of bone injury healing according to claim 1, characterized in that, The image processing software includes SPSS and / or Image J and / or GraphPad Prism.

Images