Use of curculigoside in prevention and treatment of intervertebral disc degeneration

Abstract

The application belongs to the technical field of medicine, and particularly relates to application of curculigoside in prevention and treatment of intervertebral disc degeneration. The application is for development of a non-surgical scheme for effectively treating IDD. The application finds that CUR (curculigoside) can reduce occurrence of cell apoptosis in vivo by inhibiting phosphorylation of STAT3 and up-regulating expression of BMAL1 (brain and muscle aryl hydrocarbon receptor nuclear translocator-like 1 gene), thereby reducing IVDD (intervertebral disc degeneration). It is indicated that CUR plays an important role in regulating IVDD progression, can alleviate progression of IVDD by up-regulating expression of BMAL1, and is expected to be used as a promising drug for treating IVDD. The application not only develops a new use of CUR, but also provides a new drug use strategy for prevention and treatment of IVDD.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to the application of cyclophosphamide in the prevention and treatment of intervertebral disc degeneration. Background Technology

[0002] Intervertebral disc degeneration (IVDD) is one of the leading causes of low back pain (LBP) in modern society, imposing a severe burden on people's physical, psychological, and economic well-being. The specific mechanisms underlying IVDD remain unclear. Previous research has primarily focused on biomechanics and inflammatory pathology, while recent studies have begun to investigate the link between nucleus pulposus energy metabolism and degeneration. Currently, the efficacy of non-surgical treatments is uncertain, and most patients eventually require discectomy. Since surgery inevitably involves some trauma, effectively preventing and treating IVDD has become a crucial issue in spinal surgery.

[0003] Curculigoside (CUR) is a phenolic glycoside component of Curculigo orchioides, possessing various pharmacological activities including anti-osteoporosis, antioxidant, anti-inflammatory, and anti-apoptotic effects. In the nervous system, CUR can prevent neuronal excitation, memory impairment, and depression. Furthermore, studies have shown that CUR can prevent damage caused by hydrogen peroxide (H2O2) by reducing oxidative damage. CUR can improve bone loss and oxidative stress caused by iron excess by reducing ROS production and increasing the activity of antioxidant enzymes (such as SOD and GPX4). More importantly, CUR has a significant effect on reducing apoptosis. In hepatic ischemia-reperfusion injury, CUR exerts a protective effect by inhibiting oxidative stress, inflammation, and apoptosis. It has also been reported that CUR can downregulate p53 levels and FoxO1 phosphorylation levels, enhance cell viability and autophagy, and reduce apoptosis in iron-excessive MC3T3-E1 cells. However, whether CUR can participate in the regulation of IVDD progression by mediating BMAL1 expression remains unclear, and its molecular mechanism requires further investigation. Summary of the Invention

[0004] In order to overcome the shortcomings of the prior art, the present invention has found that CUR can reduce the occurrence of apoptosis in vivo by inhibiting the phosphorylation of STAT3 and upregulating the expression of BMAL1 (brain and muscle aryl hydrocarbon receptor nuclear transporter-like protein 1 gene), thereby alleviating IVDD, and is expected to be applied to the prevention and treatment of IVDD.

[0005] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0006] The first aspect of this invention provides the use of cicroside (CUR) in the preparation of drugs for preventing and treating intervertebral disc degeneration.

[0007] Preferably, the CUR alleviates intervertebral disc degeneration (IVDD) by inhibiting STAT3 phosphorylation and upregulating BMAL1 expression.

[0008] More preferably, the inhibition of STAT3 phosphorylation is the inhibition of STAT3 phosphorylation at the TYR705 site.

[0009] This invention has shown that CUR can improve NP cell (nucleus pulposus) apoptosis by inhibiting STAT3 phosphorylation at the TYR705 site and upregulating BMAL1 expression. Further in vivo studies revealed that CUR can reduce in vivo cell apoptosis by inhibiting STAT3 phosphorylation and upregulating BMAL1 expression, thereby alleviating IVDD. This suggests that CUR holds promise for the prevention and treatment of intervertebral disc degeneration.

[0010] A second aspect of the present invention provides a drug for preventing and treating intervertebral disc degeneration, wherein the drug uses curculigoside as the main active ingredient.

[0011] Preferably, the drug further includes other drug components that can synergistically prevent and treat intervertebral disc degeneration with curculigoside. The synergistic prevention and treatment of intervertebral disc degeneration refers to using other drugs together with CUR to prevent and treat intervertebral disc degeneration, which can further enhance the preventive and treatment effect.

[0012] Preferably, the drug further includes pharmaceutically acceptable excipients. These excipients may be diluents, binders, lubricants, disintegrants, solubilizers, stabilizers, and some pharmaceutical matrices that are applicable in the pharmaceutical field; they may also be functional pharmaceutical excipients available in the pharmaceutical field, including surfactants, suspending agents, emulsifiers, and some novel pharmaceutical polymers, such as cyclodextrin, chitosan, polylactic acid (PLA), polyglycolic acid-polylactic acid copolymer (PLGA), hyaluronic acid, etc.

[0013] Preferably, the dosage form of the drug includes powder, tablets, granules, capsules, pellets, sustained-release preparations, oral liquid preparations, and injections. The above dosage forms refer to those commonly used in clinical practice. The drug preparation can be administered orally or parenterally (e.g., intravenously, subcutaneously, intraperitoneally, or locally). If certain drugs are unstable under gastric conditions, they can be prepared as enteric-coated tablets.

[0014] Compared with the prior art, the beneficial effects of the present invention are:

[0015] Intra-intra-intra-digestive disease (IVDD) is the leading cause of pulmonary embolism (LBP), and in severe cases, it can lead to quadriplegia and even death, posing a serious threat to human health. Currently, there is a lack of effective non-surgical treatment options for IDD. Therefore, this invention has discovered that CUR can reduce in vivo apoptosis by inhibiting STAT3 phosphorylation and upregulating BMAL1 expression, thereby alleviating IVDD. This indicates that CUR plays an important role in regulating IVDD progression and can alleviate IVDD progression by upregulating BMAL1 expression, showing promise as a drug for treating IVDD. This invention not only develops a new use for CUR but also provides a new drug strategy for the prevention and treatment of IVDD. Attached Figure Description

[0016] Figure 1 Inhibition of the JAK-STAT3 signaling pathway upregulates BMAL1 expression, thereby alleviating apoptosis in NP cells. (A) Sankey diagrams from RNA-seq data pathway analysis show that the JAK-STAT signaling pathway is associated with and negatively regulates BMAL1. (B and C) Transcription factor analysis indicates that STAT3 is activated in SDD NP tissues and is associated with the BMAL1 gene. (D) BMAL1 expression is significantly upregulated by treatment with the specific STAT3 inhibitor Static, and BMAL1 expression is inhibited by the JAK / STAT activator RO8191; Static reverses the reduction in BMAL1 after TNF-α treatment. (E) Molecular docking analysis predicts that CUR may bind near the phosphorylation site in STAT3. (F) Flow cytometry analysis shows that Annexin-V positivity is reduced when cells are infected with the BMAL1 expression vector, and similar results are obtained with CUR treatment. (GJ) Western blot analysis showed that BMAL1 overexpression reversed the changes in the BAX / BCL-2 ratio in NP cells and the abnormal synthesis and degradation of ECM after TNF-α treatment; CUR inhibited STAT3 phosphorylation and increased BMAL1 expression, similar to the effect of BMAL1 overexpression. (K) Cell fluorescence of NP cells showing p-STAT3 and BMAL1 (scale bar: left 100 μm, right 20 μm). *p<0.05; **p<0.01; ***p<0.001. Data are presented as mean ± standard deviation.

[0017] Figure 2(A) Molecular structure of CUR in MedChem database. (B) 3D structure of STAT3 (PDB id: 1BG1). (C) Molecular docking analysis by AutoDock predicts that CUR may bind near the phosphorylation site of STAT3. (D and E) CCK8 assay of CUR. (FI) Western blot and quantitative analysis showed that BMAL1 was upregulated by CUR in a concentration- and time-dependent manner. (J) qPCR validation of the time-dependent expression of the clock gene Bmal1, which was homogenized with dexamethasone and then transfected with TNF-α (100 ng / ml, 24 h), si-BMAL1, and Ex-BMAL1. Results were normalized to GAPDH as mean and SEM (n=3). Gray shading indicates the nighttime phase. (K) Flow cytometry of NP cells.

[0018] Figure 3 CUR alleviates IVDD by upregulating BMAL1 expression through inhibition of the JAK-STAT3 pathway; (A and B) MRI images and T2 signal intensity analysis of the three groups of mice at 8 weeks. (C and D) Micro-CT images and intervertebral disc height analysis of the three groups of mice. (EH) Quantitative analysis of H&E, S&O, and IHC staining, as well as BMAL1, ACAN, COL1, COL2, and TUNEL staining in NP tissues of CTR mice and mice treated with AFP or AFP and CUR (scale bar: 400 μm in H&E and S&O staining; scale bar: top 400 μm and bottom 100 μm in IHC staining). (I) IF staining of BMAL1 and TUNEL in NP tissues of CTR mice and mice treated with AFP or AFP and CUR (scale bar = 400 μm; high-resolution image scale bar = 100 μm). (J) Co-staining of NP tissues from IVD with ACAN, COL1, and COL2 by IF analysis (scale bar = 400 μm; scale bar for high-resolution images = 100 μm). *p<0.05; **p<0.01; ***p<0.001. Data are presented as mean ± standard deviation. Detailed Implementation

[0019] The specific embodiments of the present invention will be further described below. It should be noted that these descriptions are for the purpose of aiding understanding the present invention, but do not constitute a limitation thereof. Furthermore, the technical features involved in the various embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0020] Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods, and the experimental materials used in the following embodiments are all available through conventional commercial channels.

[0021] The materials involved in the following embodiments are as follows:

[0022] (1) siRNA sequence

[0023]

[0024] (2) Ex-BMAL1 refers to a BMAL1 overexpression lentivirus, which is coated with an overexpression plasmid constructed from ARNTL(Human) (ID:406, sequence number: NM_001178). The Ex-BMAL1 used in this invention was purchased from Ribobio.

[0025] (3) Acquisition and Detection of Intervertebral Disc Specimens of Different Degeneration Grades: Normal intervertebral disc nucleus pulposus specimens were obtained from patients who required disc removal due to trauma or other factors. Degenerated intervertebral disc nucleus pulposus specimens were obtained from patients who underwent disc removal due to disc degeneration-related diseases. Informed consent was obtained from patients before specimen acquisition. The specimen acquisition by this research group has been reviewed and approved by the Ethics Committee of the First Affiliated Hospital of Sun Yat-sen University. The acquired normal or degenerated intervertebral disc nucleus pulposus specimens were divided into a control group and a degeneration group. The specimens were graded according to the clinically used Pfirrmann criteria for disc degeneration and then grouped. The number of specimens after grouping was counted and statistically analyzed. The specimens were then embedded in paraffin, sectioned, dewaxed, and stained. Finally, they were scanned and photographed using a fully automated digital slide scanner for analysis.

[0026] (4) RT-qPCR primers for BMAL1

[0027]

[0028] (5) Antibody information (including antibody name, manufacturer, catalog number, and application concentration)

[0029]

[0030]

[0031]

[0032] (6) Reagent Information

[0033]

[0034] Example: An investigation into the therapeutic effect of curculigoside on intervertebral disc degeneration

[0035] 1. Experimental Methods

[0036] (1) Isolation and culture of human nucleus pulposus cells

[0037] Nucleus pulposus (NP) cells were isolated and cultured using human central nucleus pulposus (NP) tissue specimens (specific methods refer to "Chen F, Liu H, Wang X, et al. Melatonin activates autophagy via the NF-kappaB signaling pathway to prevent extracellular matrix degeneration in intervertebral disc. Osteoarthritis Cartilage 2020; 28(8):1121-32."). NPC cells were then placed in DMEM (Invitrogen, CA, USA) containing 10% FBS and antibiotics (1% penicillin and streptomycin) and cultured at 37°C in a humidity-controlled incubator containing 5% CO2. Cells were digested with a solution containing trypsin (0.25%) and EDTA (1 mM) (Invitrogen, CA) and then cultured further in 10 cm culture dishes. The NPCs were then transferred to 6-well culture dishes and grown to 80% density. They were then treated with CUR 5 μM (Abcam, ab143113, USA) or TNF-α 100 ng / ml (R&D, 10190-SN-050, USA) for 24 hours for subsequent experiments.

[0038] (2) Animal modeling

[0039] Referring to the method used by the research group in the previous study to establish an IVDD low back pain model by anterior needle puncture of the intervertebral disc (Chen F, Jiang G, Liu H, et al. Melatonin alleviates intervertebral disc degeneration by disrupting the IL-1beta / NF-kappaB-NLRP3 inflammasome positive feedback loop. Bone Res 2020; 8:10.), a rat acupuncture degeneration model was constructed (denoted as AFP group, i.e., annulus fibrosus puncture group). The specific steps are as follows: First, clean and shave the rats (common SD rats, sourced and raised by the Animal Breeding Center of the First Affiliated Hospital of Sun Yat-sen University). Clean the fur on the mouse's abdomen with 2% povidone-iodine. Make a longitudinal incision about 1 cm long along the midline of the abdomen, centered 0.5 cm above the line connecting the bilateral iliac joints, to expose the linea alba of the peritoneum. Use hemostats or forceps to clamp the peritoneum three times, avoiding accidental injury to the intestines, and make a longitudinal incision in the peritoneum, slightly larger than the skin incision. Then, use hemostats or cotton swabs moistened with saline to gently move the mouse's intestines and greater omentum to the right side of the abdominal cavity to facilitate subsequent operations. Locate the safety triangle area by determining the lower border of the left kidney and the upper border of the bladder. Within the safety triangle area (lower border of the left kidney, midline of the spine, and upper border of the bladder), use forceps to locate the psoas major muscle. The muscle bundles of the left psoas major muscle converge obliquely upwards. The genitofemoral nerve is visible attached to the connective tissue on the surface of the muscle; avoid damaging this nerve. Subsequently, along the convergence point of the psoas major muscle bundles on both sides, the midline connective tissue was separated, and the bilateral psoas major muscle bundles were dissected using smooth clamps to reveal the location of the intervertebral disc. The transverse vertebral artery, a branch of the abdominal aorta, was visible above the vertebral body; damage to this artery was avoided. Finally, a mouse anterior lumbar puncture model was performed using a 27G depth-limiting needle with slow puncture. Postoperatively, the peritoneum and skin incision were sutured in layers using 4-0 Vickeryl sutures.

[0040] CUR was diluted to 10 μM / mL in physiological saline, and mice in the CUR+AFP group were intraperitoneally injected with one of the following CUR concentrations: 0, 1, 3, 5, or 10 μM / kg / week. The best IVDD improvement was observed at a concentration of 5 μM mg / kg. Mice received weekly intraperitoneal injections of 5 μM / kg CUR during the first week post-surgery for a total of 3 weeks. Control group mice received no treatment post-surgery.

[0041] (3) Micro-computed tomography (Micro-CT) and magnetic resonance imaging (MRI)

[0042] Eight weeks post-surgery, before all mice were euthanized, micro-CT and T2-weighted MRI scans were performed to obtain imaging data of the intervertebral discs. Mice were anesthetized and placed in a prone position; to ensure measurement accuracy, the spine was fixed and kept straight during the scan. The micro-CT scan results were reconstructed and three-dimensional images created using NRecon, Dataviewer, and CTFox. The degree of degeneration on MRI was determined according to the Pfirrmann classification.

[0043] (4) Western Blot and Analysis

[0044] NPC proteins were extracted after treatment (including siRNA, ex-BMAL1 transfection, CUR treatment, or stimulation by inflammatory factors, etc.) and separated by 10% or 15% SDS-PAGE electrophoresis. The membrane was then blocked with 3% bovine serum albumin (BSA) and incubated with a primary antibody. After washing with PBS, the membrane was incubated with either anti-rabbit IgG (1:5,000, 7074, Cell Signaling Technology) or anti-mouse IgG (1:5,000, 7076, Cell Signaling Technology). Finally, Western blot bands were detected using an enhanced chemiluminescence assay (Invitrogen, CA, USA).

[0045] (5) Immunohistochemical and histopathological analysis

[0046] For the obtained mouse intervertebral disc tissue specimens, they are first embedded in paraffin and cut into 5-micrometer thin sections for mounting. Next, dewaxing and hydration are performed, followed by hematoxylin and eosin (H&E), safranin-fast green, and scarlet staining (S&O). Immunohistochemical staining (IHC) is also performed as needed. The specific steps are as follows: First, the antigen is repaired using 0.01M citrate, then blocked with 3% hydroperoxide and 5% normal goat serum. The sections are then incubated overnight at 4°C with primary antibodies including anti-type II collagen (1:1000) and anti-Aggrecan (1:200). After washing with PBST, incubation is performed with the corresponding HRP-labeled secondary antibody. Finally, DAB reagent is used for staining, and the cell nuclei are stained with hematoxylin. Finally, sections were observed and photographed using an Olympus BX63 microscope at magnifications of 10×, 100×, and 400×, and the percentages of SRGN+, IL-1β+, CCL3+, COL2+, and ACAN+ cells in the IVD samples were calculated using ImageJ software (National Institutes of Health, Bethesda, MD, USA). Simultaneously, histological scoring was performed according to previous methods (see “Zhang J, Wang X, Liu H, et al. TNF-alpha enhances apoptosis by promoting chop expression in nucleus pulposus cells: role of the MAPK and NF-kappaB pathways. J Orthop Res 2019; 37(3):697-705.”), including normal intervertebral discs, mildly degenerated intervertebral discs, and moderately to severely degenerated intervertebral discs.

[0047] (6) Molecular docking analysis

[0048] The structures of STAT3 (PDB id: 1BG1) and CUR were obtained from the PDB database and the Zinc database, respectively. Molecular docking of the STAT3 and CUR structures was simulated and analyzed using AutoDockTools-1.5.6, and visualized using PyMOL 2.3.2.

[0049] (7) RNA sequencing (RNA-seq) and analysis

[0050] RNA sequencing and analysis were used to investigate differential gene expression in nucleus pulposus (NP) tissues. Total RNA was extracted from nucleus pulposus tissue and analyzed using GeneRead. TMRibosomal RNA was removed using the rRNA Depletion kit (Qiagen, Germany). Then, a specific library was constructed using the VAHTS Stranded RNA-seq Library preparation kit for Illumina (Vazyme, Nanjing, China). Next, sequence reads were aligned to the human genome (version Hg38) using Hisat2 (version 2 2.1.0) and evaluated. After alignment, read counts for the genome mapping were calculated using Htseq (version 0.11.0). Expression data were normalized using FPKM (fragment count per thousand base pairs per million reads) to compare gene expression levels between different groups. After significance and FDR analysis, differentially expressed genes were filtered using the DESeq2 (1.22.2) algorithm based on the following criteria: I) log2FC > 1 and II) FDR < 0.05. Based on the differential gene analysis, volcano plots and heatmaps were generated using R software, with different colors representing log2FC and FDR values. Finally, GO and KEGG pathway enrichment analysis was performed using DAVID. These methods provide a comprehensive understanding of gene expression differences in NP tissues and lay the foundation for further exploration of their biological functions and related pathways.

[0051] (8) Statistical Analysis

[0052] Normally distributed data are expressed as mean ± standard deviation. Differences among multiple groups were analyzed using one-way ANOVA. If the ANOVA results were statistically significant, the differences between the two groups were examined using an independent samples t-test. All statistical analyses were performed using SPSS software (version 19.0) on a Windows operating system. A p-value < 0.05 was considered statistically significant.

[0053] 2. Experimental Results

[0054] (1) By inhibiting the JAK-STAT3 signaling pathway to upregulate the expression of the circadian rhythm gene (BMAL1), NP cell apoptosis is reduced.

[0055] To further investigate the molecular mechanism of BMAL1 in IVDD, potential signaling pathways in BMAL1-mediated IVD denaturation were examined using RNA-seq. GO and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed using the R package "clusterProfiler". The results showed that the JAK-STAT signaling pathway was negatively correlated with BMAL1 expression. Figure 1A). Transcription factor analysis showed that STAT3 is associated with BMAL1 transcription, and STAT3 expression was significantly upregulated in the severe intervertebral disc degeneration (SDD) model. Figure 1 (B and C). Therefore, it is hypothesized that the JAK-STAT3 pathway is involved in the negative regulation of BMAL1. To this end, NP cells were extracted from human nucleus pulposus cells and pretreated with the STAT3 inhibitor Static and the STAT3 activator RO8191, followed by TNF-α treatment (NP cells were treated with Static 10 uMol for 4 h; RO8191 5 uMol for 12 h; and TNF-α 100 ng / ml when they reached 70% confluence). Western blot analysis showed that BMAL1 expression was downregulated by TNF-α treatment or RO8191 treatment, but upregulated by Static treatment. Furthermore, compared with TNF-α treatment alone, the degree of upregulation of BMAL1 expression was greater after treatment with both TNF-α and Static. Figure 1 D).

[0056] To further investigate the roles of CUR and BMAL1 in regulating IVDD progression, the toxicological properties of CUR were first examined using the CCK8 assay. Figure 2 D and E). Then, the circadian rhythm expression of NP cells treated with CUR (5 uM, 24 h) was analyzed by RT-qPCR. The results showed that CUR treatment maintained a circadian rhythm similar to that of the CTR group (control group). Figure 2 WB analysis showed that CUR increased BMAL1 expression in a dose- and time-dependent manner. Figure 2 FI). To further examine the mechanism by which CUR upregulates BMAL1 expression, AutoDock (molecular docking analysis) was performed. The results showed that CUR may bind to STAT3 TYR705, which is the phosphorylation site of STAT3. Figure 2 AC; Figure 1 E). Therefore, it is speculated that CUR upregulates BMAL1 expression by inhibiting STAT3 phosphorylation at TYR705, thereby altering STAT3 function. Following TNF-α treatment, CUR treatment significantly increased the expression levels of apoptosis and degeneration markers in NP cells (E). Figure 2 H; Figure 1 F). Western blot analysis showed that, compared with TNF-α treatment, CUR treatment significantly inhibited TYR705 phosphorylation at STAT3 and upregulated BMAL1 expression. Figure 1 GJ). Furthermore, rhythmic expressions exhibit similar trends ( Figure 2Treatment with CUR followed by TNF-α effectively reduced STAT3 phosphorylation levels and the degree of apoptosis and degeneration in NP cells. These effects were similar to those of BMAL1 overexpression. Cell fluorescence analysis indicated that CUR could increase BMAL1 expression by reducing STAT3 phosphorylation. Figure 1 Furthermore, flow cytometry analysis showed that CUR treatment effectively reduced TNF-α-induced apoptosis in NP cells (K). Figure 2 The above results indicate that CUR can improve NP cell apoptosis by inhibiting STAT3 phosphorylation at TYR705 and upregulating BMAL1 expression.

[0057] (2) CUR alleviates IVDD in vivo by inhibiting the JAK-STAT3 pathway and upregulating BMAL1 expression.

[0058] Subsequently, the effect of CUR on IVDD in vivo was further investigated. CUR was injected intraperitoneally into IVDD model mice, and the mice were divided into CTR, AFP, and AFP+CUR groups. Eight weeks post-surgery, the T2-MRI signal in the AFP+CUR group was significantly higher than that in the AFP group (…). Figure 3 A and B). Micro-CT analysis showed that the intervertebral disc (IVD) height was also significantly increased in the AFP+CUR group. Figure 3 C and D). Furthermore, H&E, S&O, and IHC staining of mouse lumbar IVDs showed that BMAL1 expression was increased in the NP tissue of the AFP+CUR group compared to the AFP group, the number of p-STAT3(TYR705)+ cells was reduced, and the degree of degeneration was alleviated. Figure 3 EH). Immunofluorescence staining of tissues showed consistent results ( Figure 3 The above results indicate that CUR can reduce NP cell apoptosis in vivo by inhibiting STAT3 phosphorylation and upregulating BMAL1 expression, thereby alleviating IVDD.

[0059] In summary, CUR plays an important role in regulating the progression of IVDD and can alleviate the progression of IVDD by upregulating BMAL1 expression, thus showing promise as a drug for the treatment of IVDD.

[0060] The embodiments of the present invention have been described in detail above, but the present invention is not limited to the described embodiments. For those skilled in the art, various changes, modifications, substitutions, and variations can be made to these embodiments without departing from the principles and spirit of the present invention, and these variations still fall within the protection scope of the present invention.

Claims

1. Application of curculigoside in the preparation of drugs for preventing and treating intervertebral disc degeneration.

2. The application according to claim 1, characterized in that, The drug also includes pharmaceutically acceptable excipients.

3. The application according to claim 1, characterized in that, The dosage forms of the drug include powder, tablets, granules, capsules, pills, sustained-release preparations, oral liquid preparations, and injections.

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