Application of thyroid hormone T3 in preparation of medicine for delaying or treating intervertebral disc degeneration

Abstract

The invention relates to application of thyroid hormone T3 in preparation of a medicine for delaying or treating intervertebral disc degeneration, and the thyroid hormone T3 can inhibit glycolysis capability of degeneration nucleus pulposus cells and reduce intervertebral disc lactic acid accumulation, so that nucleus pulposus cell senescence and apoptosis are relieved, and intervertebral disc degeneration is delayed or treated. Compared with the prior art, the technology for delaying or treating intervertebral disc degeneration by applying the thyroid hormone T3 provides new application of the thyroid hormone T3, the application value of the thyroid hormone T3 is greatly improved, the application range of the thyroid hormone T3 is expanded, the new field of application of the thyroid hormone T3 is expanded, and the thyroid hormone T3 has wide application prospects and commercial value.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, and in particular to application of thyroid hormone T3 in the preparation of a medicine for delaying or treating intervertebral disc degeneration. Background Art

[0002] Low back pain is a common disease with a high incidence and wide coverage in the modern population. With the development of society and changes in people's lifestyles, the incidence of low back pain has increased year by year, and the patients tend to be younger. A large number of studies have shown that intervertebral disc degeneration is one of the main causes of low back pain. At present, the efficacy of non-surgical treatment in clinical practice is not clear. Most patients still need to undergo nucleus pulposus removal surgery in the later stage, and the surgery is traumatic. Therefore, how to delay intervertebral disc degeneration has become an important topic in the field of spinal surgery.

[0003] Patent CN108606969A discloses the use of aspirin in the preparation of drugs for treating intervertebral disc degeneration. The invention constructs an intervertebral disc degeneration model by using LPS to induce oxidative stress, observes the therapeutic effect of aspirin on intervertebral disc degeneration, evaluates the therapeutic effect of aspirin on intervertebral disc degeneration by observing various indicators of oxidative stress and intervertebral disc degeneration, and uses Western blot to prove the brief mechanism of aspirin reducing oxidative stress and alleviating intervertebral disc degeneration. Oxidative stress in the nucleus pulposus tissue of the intervertebral disc causes intervertebral disc degeneration, and aspirin delays intervertebral disc degeneration by inhibiting the oxidative stress of the nucleus pulposus tissue and reducing the decomposition of cell matrix and type II collagen.

[0004] Patent CN108635349A discloses the use of melatonin in the preparation of drugs for treating intervertebral disc degeneration. The invention establishes a rat coccygeal disc degeneration model by needle puncture, observes the therapeutic effect of melatonin on disc degeneration, evaluates the therapeutic effect of Mel on disc degeneration by observing various indicators of disc degeneration, and observes the expression of Mell-type receptors and RANKI on the cartilage endplates of rat coccygeal discs by immunohistochemical staining and TRAP staining to illustrate the brief mechanism of Mel in delaying disc degeneration. The destruction of the disc cartilage endplates and the increased loss of subchondral bone may be important mechanisms causing disc degeneration. Mel delays disc degeneration by inhibiting the expression of RANKI by endplate chondrocytes, thereby reducing the formation of osteoclasts in the cartilage endplate area.

[0005] Patent CN115068463B discloses the use of a specific metabolic kinase PFKFB3 inhibitor (KAN) in products for improving intervertebral disc degeneration. KAN can partially restore the anaerobic glycolysis energy metabolism phenotype of over-activated nucleus pulposus cells under inflammatory stimulation, thereby reducing the excretion of lactic acid into the extracellular matrix, improving the acidification of the extracellular environment and its effects on nucleus pulposus cells and extracellular matrix, and improving intervertebral disc degeneration.

[0006] The intervertebral disc is the largest avascular tissue in the human body, and blood vessels are mainly located at the edge of the intervertebral disc. Nucleus pulposus cells mainly obtain oxygen and nutrients from surrounding tissues through diffusion. The oxygen tension in the nucleus pulposus area is extremely low (<1%), so the nucleus pulposus cells are in a hypoxic environment for a long time. This environment forces the nucleus pulposus cells to rely mainly on glycolysis to produce energy. Lactic acid is the main metabolite of glycolysis and is usually transported outside the cell, resulting in an acidic nucleus pulposus microenvironment. Nucleus pulposus cells rely on glycolysis, leading to lactic acid accumulation, further aggravating the local acidic environment, which is not conducive to the maintenance of cell function and may accelerate degeneration. Therefore, improving the microenvironment of nucleus pulposus cells (such as reducing lactic acid accumulation or alleviating the acidic environment) can help slow down intervertebral disc degeneration.

[0007] Thyroid hormones (THs), namely 3,5,3'-triiodo-L-tyrosine (3,5,3'-triiodo-L-thyronine, T3) and 3,5,3',5'-tetraiodo-L-tyrosine (3,5,3',5'-Tetraiodo-L-tyrosine, T4), have been considered to be involved in a variety of physiological and pathological processes, including cell growth, cell metabolism, tumor occurrence and development. T4 is the main hormone secreted by the thyroid gland, accounting for more than 90% of the total thyroid hormone secretion. T4 is converted into the more active T3 in the body, so T4 is generally considered a precursor hormone. T3 is the active form of thyroid hormone, and its physiological effect is 4-5 times stronger than that of T4.

[0008] At present, inventions related to thyroid hormones are mostly concentrated on thyroid hormone receptor agonists, such as a polycyclic thyroid hormone beta receptor agonist and its use (CN202210999229.5), the use of thyroid hormone beta receptor agonists in the preparation of drugs for improving intestinal flora (CN202410212000.1), etc.

[0009] Thyroid hormone has a wide range of pharmacological activities, and its core role is to regulate metabolism, promote growth and development, and maintain the nervous system and cardiovascular function. It plays an important role in the treatment of endocrine diseases (such as hypothyroidism and hyperthyroidism), and also shows important clinical application value in metabolic diseases, neurological diseases and reproductive disorders. For example, patent CN111419834A discloses the use of thyroid hormone and its analogs in the preparation of treatment for sickle cell disease; patent CN111388459A discloses the use of thyroid hormone and its analogs in the preparation of treatment for α* thalassemia.

[0010] Yang Chuncheng et al. studied the mechanism by which thyroid hormone inhibits the progression of liver cancer by regulating thyroid hormone response protein and found that thyroid hormone T3 can enhance the anti-tumor effect of the multi-kinase inhibitor lenvatinib.

[0011] Regarding the application of thyroid hormone, there is no application to delay or treat intervertebral disc degeneration. Summary of the invention

[0012] The purpose of the present invention is to overcome the defects of the above-mentioned prior art and to provide a method for preparing a thyroid hormone T3 for the preparation of a drug for delaying or treating intervertebral disc degeneration. The thyroid hormone T3 is used to inhibit the glycolytic ability of aging nucleus pulposus cells, reduce the accumulation of lactic acid in the intervertebral disc, thereby reducing the aging and apoptosis of nucleus pulposus cells, and thus use the thyroid hormone T3 to delay or treat intervertebral disc degeneration.

[0013] The purpose of the present invention can be achieved by the following technical solutions:

[0014] The first purpose of the present invention is to protect the use of thyroid hormone T3 in the preparation of a drug for delaying or treating intervertebral disc degeneration, with thyroid hormone T3 as a component that exerts a pharmacological effect.

[0015] Furthermore, the use of thyroid hormone T3 in the preparation of drugs for delaying intervertebral disc degeneration.

[0016] Furthermore, the use of thyroid hormone T3 in the preparation of drugs for treating intervertebral disc degeneration.

[0017] Furthermore, the intervertebral disc refers to a lumbar intervertebral disc.

[0018] Furthermore, the intervertebral disc degeneration includes one or more of the degeneration of the intervertebral disc annulus fibrosus, the degeneration of the intervertebral disc nucleus pulposus, the degeneration of the cartilage endplates, osteophyte formation, etc., and more preferably, the intervertebral disc degeneration is the degeneration of the intervertebral disc nucleus pulposus.

[0019] Furthermore, the thyroid hormone T3 is used in the preparation of a drug for inhibiting the glycolytic capacity of aging nucleus pulposus cells, reducing lactic acid accumulation in the intervertebral disc, and alleviating aging and apoptosis of nucleus pulposus cells.

[0020] Furthermore, the thyroid hormone T3 is used in the preparation of a drug for delaying or treating intervertebral disc degeneration caused by strong glycolysis capacity of aging nucleus pulposus cells, lactic acid accumulation, and apoptosis of nucleus pulposus cells.

[0021] Furthermore, the medicine is an oral preparation or an injectable preparation.

[0022] More preferably, the drug is an injectable preparation.

[0023] Furthermore, the medicine comprises a therapeutically effective amount of thyroid hormone T3 and pharmaceutically acceptable excipients.

[0024] Further preferably, the pharmaceutically acceptable excipients include one or more of solvents, carriers, and excipients.

[0025] Further preferably, the pharmaceutically acceptable excipients specifically include one or more of ethanol, glycerol, exosomes, etc.

[0026] Furthermore, the dosage of the drug to exert its pharmacodynamic effect is 1 to 100 nM in terms of thyroid hormone T3.

[0027] More preferably, the dosage of the drug to exert its pharmacodynamic effect is 10 nM calculated as thyroid hormone T3.

[0028] Furthermore, the thyroid hormone T3 is triiodothyronine, and the structural formula is as follows:

[0029]

[0030] The second purpose of the present invention is to protect a drug for delaying or treating intervertebral disc degeneration, said drug comprising the above-mentioned thyroid hormone T3.

[0031] Furthermore, the drug uses thyroid hormone T3 (abbreviated as T3) as the only component that exerts its pharmacological effect.

[0032] Furthermore, the dosage of the drug to exert its pharmacodynamic effect is 1 to 100 nM (nM represents nanomoles per liter, 1 nM = 10 -9 mol / L).

[0033] Furthermore, the dosage of the drug to exert its pharmacodynamic effect is 1 to 10 nM based on thyroid hormone T3.

[0034] More preferably, the dosage of the drug to exert its pharmacodynamic effect is 10 nM calculated as thyroid hormone T3.

[0035] Furthermore, the medicine also includes pharmaceutically acceptable excipients.

[0036] Further preferably, the pharmaceutically acceptable excipients include one or more of solvents, carriers, and excipients.

[0037] Further preferably, the pharmaceutically acceptable excipient specifically includes one of ethanol, glycerol, exosomes, and the like.

[0038] Furthermore, the specific dosage of thyroid hormone T3 in the medicine is 10 mg / kg body weight.

[0039] The present invention has found through research that thyroid hormone T3 as a pharmaceutical ingredient can delay or treat intervertebral disc degeneration. Specifically, the present invention has found through research that thyroid hormone T3 has the ability to inhibit the glycolysis of aging nucleus pulposus cells in cell experiments, reducing the accumulation of lactic acid in the intervertebral disc, thereby reducing the aging and apoptosis of nucleus pulposus cells. At the same time, the present invention has also found through animal experiments that injection of thyroid hormone T3 into the caudal intervertebral disc of rats can effectively delay or treat intervertebral disc degeneration.

[0040] Compared with the prior art, the present invention has the following advantages:

[0041] 1) The drug prepared by the present invention uses thyroid hormone T3 as a component to exert its pharmacological effect, which can inhibit the glycolytic ability of aging nucleus pulposus cells, reduce the accumulation of lactic acid in the intervertebral disc, thereby alleviating the aging and apoptosis of nucleus pulposus cells, and delaying or treating intervertebral disc degeneration.

[0042] 2) The present invention uses the technology of thyroid hormone T3 to delay or treat intervertebral disc degeneration, provides a new use of thyroid hormone T3, greatly improves its application value, expands the application scope of thyroid hormone T3, and develops a new field for the application of thyroid hormone T3, which has broad application prospects and commercial value. BRIEF DESCRIPTION OF THE DRAWINGS

[0043] Figure 1 This is a graph showing the results of detecting the activity of nucleus pulposus cells after treatment with different concentrations of thyroid hormone T3 using the CCK8 kit in Example 1 of the present invention.

[0044] Figure 2 The figure is a Western blotting detection result showing that the thyroid hormone T3 in Example 1 of the present invention inhibits the expression of glycolysis-related proteins pyruvate dehydrogenase kinase 1 (PDK1) and lactate dehydrogenase isozyme 5 (LDH5) in aging nucleus pulposus cells.

[0045] Figure 3 This is a graph showing the results of the thyroid hormone T3 in Example 1 of the present invention reducing lactic acid accumulation in aging nucleus pulposus cells.

[0046] Figure 4 This is a SA-β-gal staining result diagram showing that the thyroid hormone T3 in Example 1 of the present invention alleviates the senescence of nucleus pulposus cells.

[0047] Figure 5 This is a flow cytometry test result showing that the thyroid hormone T3 in Example 1 of the present invention reduces the apoptosis of nucleus pulposus cells.

[0048] Figure 6 This is a graph showing the results of HE staining and histological scoring of the rat coccyx in Example 2 of the present invention in delaying or treating intervertebral disc degeneration in vivo using the thyroid hormone T3.

[0049] Figure 7 This is a diagram showing the mechanism of action of the thyroid hormone T3 in the present invention in delaying or treating intervertebral disc degeneration. DETAILED DESCRIPTION

[0050] The present invention is described in detail below in conjunction with specific drawings and embodiments. The following embodiments will help those skilled in the art to further understand the present invention, but are not intended to limit the present invention in any form. It should be noted that, for those of ordinary skill in the art, several variations and improvements may be made without departing from the concept of the present invention. These all belong to the protection of the present invention.

[0051] Any features such as preparation methods, materials, structures or composition ratios that are not clearly described in this technical solution shall be deemed as common technical features disclosed in the prior art.

[0052] The present invention is described in detail below with reference to the accompanying drawings and specific embodiments.

[0053] In this embodiment, thyroid hormone T3 was purchased from Yisheng Biotechnology (Shanghai) Co., Ltd. (Article No.: 54574ES). Normal human nucleus pulposus cells were purchased from Wuhan Punosai Life Science Co., Ltd. (Article No.: CP-H097), cultured with DMEM / F12 medium containing 10% fetal bovine serum and 1% double antibody by volume, and placed in a cell culture incubator at 37°C and 5% CO2 by volume. The culture medium was replaced every other day, and the cell growth state was observed under a microscope. The healthy human nucleus pulposus cells used in the experiment were cells that were passaged to the 2nd-3rd generation (P2-3), and the aging human nucleus pulposus cells were obtained by passage to the 6th-8th generation (P6-8). SD rats were purchased from the Experimental Animal Center of Fudan University.

[0054] Example 1

[0055] The present embodiment provides an experimental process in which the thyroid hormone T3 inhibits the glycolytic ability of nucleus pulposus cells in vitro, reduces lactic acid accumulation, and alleviates the aging and apoptosis of nucleus pulposus cells. In the present embodiment, three experimental groups are set up, namely: a control group (Control group, P2-3 nucleus pulposus cells without any treatment, i.e., normal cells), an aging group (Aged group, P6-8 nucleus pulposus cells without any treatment, i.e., aging cells), and an aging+T3 group (T3 group, aging human nucleus pulposus cells are seeded in a 6 cm culture dish and cultured for 24 hours until the cells adhere to the wall. After washing the cells once with serum-free culture medium, a culture medium containing 10 nM T3 is added, and the cells are further cultured for 48 hours, i.e., aging cells+T3).

[0056] The glycolytic capacity, lactic acid accumulation, cell senescence and apoptosis of the nucleus pulposus cells in the above three experimental groups were detected respectively.

[0057] The CCK8 kit was used to detect the viability of nucleus pulposus cells after treatment with different concentrations of T3. The viability of aging nucleus pulposus cells decreased significantly, and the cell viability was dose-dependent with the T3 concentration. When the T3 concentration exceeded 10 nM, the cell viability did not increase much, so the concentration of 10 nM was selected for the experiment ( Figure 1 ).

[0058] Western blotting was used to detect glycolysis-related proteins in the three groups of cells. Pyruvate dehydrogenase kinase 1 (PDK1) can inhibit the entry of glucose into the mitochondrial tricarboxylic acid cycle metabolism, thereby enhancing the anaerobic glycolysis pathway; lactate dehydrogenase isoenzyme 5 (LDH5) can preferentially convert pyruvate into lactate. The results showed that the expression levels of PDK1 and LDH5 in aged nucleus pulposus cells were higher than those in normal cells, indicating that their glycolysis level was high. After T3 treatment, the expression levels of both decreased, and glycolysis was inhibited ( Figure 2 ).

[0059] The lactate detection kit was used to detect the accumulation of lactate in the culture medium. The results showed that compared with the normal cells in the control group, the degenerated nucleus pulposus cells in the aging group had lactate accumulation, while compared with the aging group, the lactate production in the T3 group decreased after using T3 ( Figure 3 ).

[0060] SA-β-gal staining was used to detect cell senescence. The results showed that compared with the normal cells in the control group, the degenerated nucleus pulposus cells in the aging group were significantly aged, while compared with the aging group, the T3 group could reduce the senescence level of nucleus pulposus cells after T3 treatment ( Figure 4 ).

[0061] Flow cytometry was used to detect cell apoptosis. The results showed that compared with the normal cells in the control group, the degenerated nucleus pulposus cells in the aging group had a higher apoptosis rate, and compared with the aging group, T3 treatment could reduce the apoptosis level of nucleus pulposus cells ( Figure 5 ).

[0062] Through this example, it is found that thyroid hormone has the ability to inhibit the glycolysis of aging nucleus pulposus cells, reduce the accumulation of lactic acid in the intervertebral disc, thereby alleviating the aging and apoptosis of nucleus pulposus cells, and delaying or treating intervertebral disc degeneration.

[0063] Example 2

[0064] This example provides an experimental process of using thyroid hormone to delay or treat intervertebral disc degeneration in vivo. Thirty 8-week-old SD rats were taken. Three experimental groups were set up in this example, namely: a control group (Control, no intervention was implemented), a degeneration group (IVDD, a group with intervertebral disc degeneration caused by acupuncture), and a degeneration + T3 group (T3, a group injected with 10nM T3 after acupuncture modeling).

[0065] The specific experiments of the above experimental groups are as follows:

[0066] After general anesthesia, the rats were placed in a prone position. Under fluoroscopic guidance, a 20-gauge needle was used to puncture the 8th-9th segment intervertebral disc (Co8 / 9). The needle passed through the center of the intervertebral disc to the opposite side, rotated 180°, and held for 10 seconds. The degeneration + T3 group was injected with 2 μL of 10nM T3 with a 33-gauge needle immediately after modeling. After modeling, the wound was covered with gauze and standard postoperative treatment was performed. After surgery, the health status of the rats was monitored daily, and they were allowed to move freely and eat and drink water normally.

[0067] All rats were killed 8 weeks after modeling, and the coccyx was taken for HE staining and histological scoring. The histological score of the intervertebral disc in the degeneration group was the highest, and the sections showed disordered nucleus pulposus tissue structure; while the injection of T3 in the degeneration + T3 group could better preserve the structure of the intervertebral disc ( Figure 6 ).

[0068] Through this example, it was found that the injection of thyroid hormone into the caudal intervertebral disc of the degeneration group rats can effectively delay or treat intervertebral disc degeneration. The mechanism of action of thyroid hormone in delaying or treating intervertebral disc degeneration is as follows: Figure 7 shown.

[0069] The above description of the embodiments is to facilitate the understanding and use of the invention by those skilled in the art. It is obvious that those skilled in the art can easily make various modifications to these embodiments and apply the general principles described herein to other embodiments without creative work. Therefore, the present invention is not limited to the above embodiments, and improvements and modifications made by those skilled in the art based on the disclosure of the present invention without departing from the scope of the present invention should be within the scope of protection of the present invention.

Claims

1. A use of thyroid hormone T3 in the preparation of a drug for delaying or treating intervertebral disc degeneration, characterized in that: The thyroid hormone T3 is used as the component that exerts its pharmacological effect.

2. The use of a thyroid hormone T3 according to claim 1 in the preparation of a drug for delaying or treating intervertebral disc degeneration, characterized in that: The thyroid hormone T3 is used in the preparation of a drug for inhibiting the glycolysis ability of aging nucleus pulposus cells, reducing lactic acid accumulation in the intervertebral disc, and alleviating aging and apoptosis of nucleus pulposus cells.

3. Use of a thyroid hormone T3 according to claim 1 in the preparation of a drug for delaying or treating intervertebral disc degeneration, characterized in that: The medicine is an oral preparation or an injection preparation.

4. The use of a thyroid hormone T3 according to claim 1 in the preparation of a drug for delaying or treating intervertebral disc degeneration, characterized in that: The medicine comprises a therapeutically effective amount of thyroid hormone T3 and pharmaceutically acceptable excipients.

5. The use of a thyroid hormone T3 according to claim 4 in the preparation of a drug for delaying or treating intervertebral disc degeneration, characterized in that: The pharmaceutically acceptable excipients include one or more of solvents, carriers, and excipients.

6. Use of a thyroid hormone T3 according to claim 1 in the preparation of a drug for delaying or treating intervertebral disc degeneration, characterized in that: The dosage of the drug to exert its pharmacodynamic effect is 1 to 100 nM based on thyroid hormone T3.

7. Use of a thyroid hormone T3 according to claim 6 in the preparation of a drug for delaying or treating intervertebral disc degeneration, characterized in that: The dosage of the drug to exert its pharmacodynamic effect is 10 nM based on thyroid hormone T3.

8. A drug for delaying or treating intervertebral disc degeneration, characterized in that: The drug comprises the thyroid hormone T3 as described in any one of claims 1 to 7.

9. The drug for delaying or treating intervertebral disc degeneration according to claim 8, characterized in that: The drug uses thyroid hormone T3 as the only component that exerts its pharmacological effect.

10. The drug for delaying or treating intervertebral disc degeneration according to claim 8, characterized in that: The medicine also includes pharmaceutically acceptable excipients; the pharmaceutically acceptable excipients include one or more of solvents, carriers, and excipients.

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