Collagen type vi for inhibiting premature ovarian failure and use thereof

Abstract

This invention discloses a type VI collagen that inhibits premature ovarian failure and its application, belonging to the technical field of collagen and its applications. The type VI collagen that inhibits premature ovarian failure contains a type VI collagen peptide with the amino acid sequence SEQ ID NO:1. This invention obtains a collagen peptide capable of improving premature ovarian failure from type VI collagen COL6A3, exhibiting high safety. The collagen peptide is composed of 29 amino acids, has a small molecular weight, and can be synthesized in vitro via a solid-phase polypeptide method, facilitating industrial production, simplifying quality control, and enabling formulation development and quality standardization. The type VI collagen in this invention can act on ovarian tissue, improve the uterine microenvironment, promote the recovery of hormone levels in the ovaries, inhibit ovarian granulosa cell apoptosis, repair ovarian tissue morphology, reduce follicular atresia, significantly enhance ovarian reserve function and fertility, alleviate ovarian function decline, and achieve the prevention and treatment of premature ovarian failure.

Description

Technical Field

[0001] This invention belongs to the field of collagen and its application technology, specifically relating to a type VI collagen that inhibits premature ovarian failure and its application. Background Technology

[0002] Women's reproductive health is the cornerstone of population health. The ovaries, which secrete hormones and produce oocytes, directly determine a woman's fertility and overall health throughout her life. Premature ovarian failure (POF) is a condition characterized by amenorrhea, infertility, low estrogen levels, and high gonadotropin levels in women under the age of 40. Due to a combination of factors, including environmental changes, unhealthy lifestyle habits, and increased socio-psychological stress, the number of women with declining ovarian reserve and POF is increasing, and the trend is towards younger ages. Ovarian dysfunction not only directly leads to decreased or even lost fertility but also causes long-term complications such as osteoporosis, cardiovascular disease, and cognitive decline due to long-term estrogen deficiency, severely impacting women's quality of life and healthy lifespan. Coupled with the current societal trend of women having children at a later age, older women are more prone to infertility, miscarriage, and adverse pregnancy outcomes due to the gradual decline in egg quality and ovarian reserve.

[0003] Currently, traditional treatments for premature ovarian failure are very limited and their effectiveness is not ideal. While hormone replacement therapy can alleviate symptoms caused by estrogen deficiency to some extent, it cannot fundamentally restore ovarian function, and long-term use may lead to serious risks such as blood clots and breast cancer. Assisted reproductive technologies such as in-vitro fertilization also have extremely low success rates due to ovarian failure, a low number of follicles, or poor follicle quality, making it difficult to meet patients' fertility needs.

[0004] Given that conventional hormone replacement therapy cannot reverse ovarian function decline and there are safety concerns regarding long-term use, developing safe, long-term-use drugs to alleviate premature ovarian failure has become a research hotspot and important direction in the field of ovarian protection. Summary of the Invention

[0005] To address the issues that conventional hormone replacement therapy cannot reverse ovarian function decline and that long-term use poses safety concerns, this invention provides a type VI collagen that inhibits premature ovarian failure and its application. The type VI collagen (peptide) that inhibits premature ovarian failure can act on ovarian tissue, improve the uterine microenvironment, promote the recovery of hormone levels in the ovaries, inhibit ovarian granulosa cell apoptosis, repair ovarian tissue morphology, reduce follicular atresia, significantly improve ovarian reserve function and fertility, alleviate ovarian function decline, and achieve the prevention and treatment of premature ovarian failure.

[0006] This invention is achieved through the following technical solution: This invention provides a type VI collagen protein that inhibits premature ovarian failure, wherein the type VI collagen protein that inhibits premature ovarian failure contains a type VI collagen peptide with the amino acid sequence SEQ ID NO:1: SEQ ID NO: 1:VFIPAEFRAAPLQGMLPGLLAPLRTLSGT.

[0007] This invention also provides a type VI collagen peptide that inhibits premature ovarian failure, the amino acid sequence of which is shown in SEQ ID NO:1. Furthermore, the type VI collagen peptide is a truncated protein of type VI collagen COL6A3.

[0008] Furthermore, the type VI collagen peptide is prepared by solid-phase synthesis or recombinant genetic engineering expression.

[0009] This invention also provides the application of the aforementioned type VI collagen or type VI collagen peptide that inhibits premature ovarian failure in the preparation of drugs for the prevention or treatment of premature ovarian failure.

[0010] Furthermore, the levels of type VI collagen or type VI collagen peptides that inhibit premature ovarian failure decrease, while the levels of follicle-stimulating hormone and luteinizing hormone increase.

[0011] Furthermore, the type VI collagen or type VI collagen peptides that inhibit premature ovarian failure repair ovarian tissue morphology, increase follicle reserve, and reduce the proportion of follicle atresia.

[0012] Furthermore, the type VI collagen or type VI collagen peptides that inhibit premature ovarian failure improve the level of ovarian and uterine fibrosis.

[0013] Furthermore, the aforementioned drugs for the prevention or treatment of premature ovarian failure also contain pharmaceutically acceptable excipients.

[0014] Furthermore, the excipients are selected from any one or more of excipients, diluents, lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents, or preservatives.

[0015] Furthermore, the dosage form of the drug for preventing or treating premature ovarian failure is any one of injection, oral liquid, capsule, tablet or granule.

[0016] Compared with the prior art, the beneficial effects achieved by the present invention are as follows: (1) The present invention obtains type VI collagen peptides that can improve premature ovarian failure from type VI collagen COL6A3, which has high safety; the collagen peptides are composed of 29 amino acids, have a small molecular weight, and can be synthesized in vitro by solid phase polypeptide method, which is conducive to industrial production, simple quality control, and easy to develop formulations and standardize quality.

[0017] (2) The type VI collagen or type VI collagen peptide in this invention can act on ovarian tissue, improve the uterine microenvironment, promote the recovery of hormone levels in the ovary, inhibit ovarian granulosa cell apoptosis, repair ovarian tissue morphology, reduce follicular atresia, significantly improve ovarian reserve function and fertility, alleviate ovarian function decline, and achieve prevention and treatment of premature ovarian failure. Attached Figure Description

[0018] Figure 1 The images show the ovarian morphology of SD rats in the blank control group and the model group. A represents the blank control group, and B represents the model group. Figure 2 HE staining images of ovarian tissue from SD rats: A is the blank control group, B is the model group, and C is the medium-dose group. Figure 3 The image shows a quantitative 2x magnification of uterine tissue fibrosis in SD rats. A represents the blank control group, B represents the model group, and C represents the medium-dose group. Detailed Implementation

[0019] The present invention is further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods not specifically described in the following examples are generally performed under conventional conditions or as recommended by the manufacturer.

[0020] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of skill in the art. All reagents and materials used in this invention are readily available through conventional means, and unless otherwise specified, they shall be used in accordance with conventional methods in the art or as per the product instructions.

[0021] Example 1 The type VI collagen in this invention that inhibits premature ovarian failure contains a type VI collagen peptide with the amino acid sequence shown in SEQ ID NO:1 (VFIPAEFRAAPLQGMLPGLLAPLRTLSGT). This type VI collagen peptide is derived from type VI collagen COL6A3 (UNIPROTID: P12111) and consists of 29 amino acids, making it suitable for Fmoc solid-phase chemical synthesis. The peptide shown in SEQ ID NO:1 was synthesized using Fmoc solid-phase chemical synthesis, and subsequent tests were performed.

[0022] Example 2 (1) Modeling: SPF-grade female SD rats (200-220g) were selected as experimental subjects. They were allowed free access to food and water and were acclimatized for one week before the experiment. The experimental SD rats were randomly divided into 6 groups (n=10 per group). Except for the blank control group, the rats were intraperitoneally injected with cyclophosphamide at a dose of 50mg / kg, once a week for two weeks, to establish the premature ovarian failure model. The blank control group was injected with an equal volume of physiological saline. The ovarian morphology of the mice in the modeling group and the blank control group is shown in the figure below. Figure 1 As shown, A is the blank control group and B is the model group. Figure 1 It was found that after cyclophosphamide treatment, the ovaries in the model group exhibited characteristic pathological changes of premature ovarian failure, such as atrophy, pale and dark color, and reduced follicles, indicating that the cyclophosphamide-induced premature ovarian failure model was successfully established. Except for the blank control group and the model group, other modeling groups were treated with the following drugs: the positive control group and type VI collagen peptide were prepared or diluted with physiological saline to the prescribed concentration. The concentration of the positive control drug estradiol was 0.02 mg / mL, and the concentrations of type VI collagen peptide were 10 μg / mL (low-dose group), 50 μg / mL (medium-dose group), and 100 μg / mL (high-dose group), respectively. These were administered via tail vein injection at a dose of 0.2 mL / animal / day. The blank control group and the model group were injected with the same dose of physiological saline. The treatment was continued for 28 days. The grouping and drug administration details are shown in Table 1 below. (Blank control group and model group are listed below.) Table 1 Grouping and Dosing Information Twenty-four hours after the last administration, blood was drawn from the orbital follicle to detect the levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E2) using ELISA; the results are shown in Table 2 below. Table 2 Serum reproductive hormone levels in SD rats Table 2 shows that the levels of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in the model group rats were significantly higher than those in the blank control group, while the level of estradiol (E2) was significantly lower. This indicates that cyclophosphamide administration damaged ovarian granulosa cells, leading to increased production of FSH and LH and decreased estradiol secretion. However, after intervention with the positive control drug and type VI collagen peptides, the levels of FSH, LH, and E2 gradually returned to normal, indicating that the positive control drug and type VI collagen peptides had a good effect on improving hormonal imbalances caused by premature ovarian failure. The medium-dose group of type VI collagen peptides was slightly better than the positive control drug, while the high-dose and medium-dose groups had comparable levels. The low-dose group showed limited improvement, and the medium-dose group could be selected for future experiments.

[0023] Example 3 Example 2: After collecting blood from the orbit, rats were euthanized by cervical dislocation. Ovarian tissue was collected, fixed, embedded, sectioned, and stained with hematoxylin and eosin (HE). The number of primordial follicles, secondary follicles, small antral follicles, and antral follicles was observed and counted, and the average value was taken. The results are shown in Table 3 below: Table 3 Statistical data of rat ovarian oocytes Table 3 shows that compared with the blank control group, the model group had a significantly increased number of atretic follicles and a decreased number of granulosa cells, indicating insufficient follicular reserve and reduced ovarian function. After intervention with positive control drugs and type VI collagen peptides, the number of atretic follicles was relatively reduced compared with the model group, while the number of follicles increased. The medium-dose group of type VI collagen peptides was slightly better than that of the positive control drugs, while the high-dose and medium-dose groups were comparable. The low-dose group showed limited improvement, consistent with the technical effects of Example 2.

[0024] Comparing the HE staining results of rat ovaries in the blank control group, model group, and medium-dose group, as shown in the figure. Figure 2 As shown, A is the blank control group, B is the model group, and C is the medium-dose group. Figure 2 It can be seen that follicles at different developmental stages were visible in the blank control group, with granulosa cells arranged regularly; the number of atretic follicles was significantly increased in the model group, the number of granulosa cells in the follicles was reduced, and the proportion of atretic follicles in each intervention group was reduced compared with the model group, while the number of follicles was increased, indicating that ovarian function has recovered to some extent.

[0025] Example 4 Pathological evaluation of the uterus: In Example 2, after taking blood from the orbit, the rat was euthanized by cervical dislocation. Uterine tissue was collected from the rat and stained with HE and Masson staining. Then, the level of fibrosis was compared. The results are as follows: Figure 3 As shown, A is the blank control group, B is the model group, and C is the medium-dose group. Figure 3 It can be seen that, compared with the model group, the deposition of uterine collagen fibers in the medium-dose group of type VI collagen peptide was reduced, and the proportion of fibrosis area showed a decreasing trend to varying degrees. This indicates that type VI collagen has a certain improving effect on the uterine stromal microenvironment and has the potential to improve the level of uterine fibrosis.

[0026] The above embodiments are merely preferred embodiments of the present invention and should not be construed as limiting the scope of protection of the present invention. Any non-substantial changes and substitutions made by those skilled in the art based on the present invention shall fall within the scope of protection claimed by the present invention.

Claims

1. A type VI collagen protein that inhibits premature ovarian failure, characterized in that, The type VI collagen that inhibits premature ovarian failure contains a type VI collagen peptide with the amino acid sequence SEQ ID NO:

1.

2. A type VI collagen peptide that inhibits premature ovarian failure, characterized in that, The amino acid sequence of the type VI collagen peptide that inhibits premature ovarian failure is shown in SEQ ID NO:

1.

3. The type VI collagen peptide for inhibiting premature ovarian failure according to claim 1 or the type VI collagen peptide for inhibiting premature ovarian failure according to claim 2, characterized in that, The type VI collagen peptide is a truncated protein of type VI collagen COL6A3.

4. The type VI collagen peptide for inhibiting premature ovarian failure according to claim 1 or the type VI collagen peptide for inhibiting premature ovarian failure according to claim 2, characterized in that, The type VI collagen peptides are prepared by solid-phase synthesis or recombinant genetic engineering expression.

5. The use of the type VI collagen peptide for inhibiting premature ovarian failure as described in claim 1 or the type VI collagen peptide for inhibiting premature ovarian failure as described in claim 2 in the preparation of drugs for the prevention or treatment of premature ovarian failure.

6. The application according to claim 5, characterized in that, The levels of type VI collagen or type VI collagen peptides decreased, while the levels of follicle-stimulating hormone and luteinizing hormone increased.

7. The application according to claim 5, characterized in that, The type VI collagen or type VI collagen peptides mentioned above repair the morphology of ovarian tissue, increase follicle reserve, and reduce the proportion of follicle atresia.

8. The application according to claim 5, characterized in that, The type VI collagen or type VI collagen peptides described herein improve the level of fibrosis in the ovaries and uterus.

9. The application according to claim 5, characterized in that, The aforementioned drugs for the prevention or treatment of premature ovarian failure also contain pharmaceutically acceptable excipients.

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