Application of arabian jasmine flower lactone in preparing medicine for treating senescence caused by microgravity

Abstract

The invention provides application of arabian jasmine flower lactone in preparation of a medicine for resisting senescence caused by microgravity. The arabian jasmine flower lactone can inhibit dissolution and / or deconstruction of F-actin caused by microgravity, stabilize cytoskeleton, and further inhibit or delay the aging process under microgravity. The invention also provides application of the arabian jasmine flower lactone in preparation of medicines for preventing and / or treating space diseases. The arabian jasmine flower lactone can inhibit organ and / or cell function decline caused by weightlessness in a space environment.

Description

Technical Field

[0001] The invention discloses a medical use of a macrocyclic peptide natural product, belonging to the technical field of pharmaceutical science. Background Art

[0002] With the gradual construction of space stations, astronauts are staying in space for longer and longer periods of time. The extension of working time in space poses new challenges to the safety and health of astronauts. The impact of the space environment on the body is more adverse, including the decline of organ function and individual aging characteristics. The main manifestations are changes in cardiovascular function, muscle loss, osteoporosis, insulin resistance, melatonin reduction, decreased immunity, decreased cognitive function, optic nerve edema, anemia, etc. The space environment mainly includes cosmic radiation, magnetic field changes, microgravity environment and other factors. The mechanism of the impact of these factors on aging-like changes in the human body is still unknown.

[0003] Jasplakinolide is a natural macrocyclic peptide from the sea sponge Jaspis johnstoni (C 36 H 45 BrN4O 6， CAS No.102396-24-7), which can induce actin polymerization into microfilaments (F-actin). Jasplakinolide competes with phalloidin, an irreversible stabilizer of F-actin, for binding to F-actin, thereby stabilizing the microfilaments and promoting the filamentous assembly of actin. Currently, there are no reports on the anti-aging effects of Jasplakinolide.

[0004] The purpose of the present invention is to provide a method for overcoming the aging effects of space environmental factors such as cosmic radiation, magnetic field changes, and microgravity environment on the body, and to provide a method for the preparation and application of an anti-aging drug. Summary of the invention

[0005] Based on the above objectives, the present invention first provides the use of jasmine lactone in the preparation of a therapeutic drug for resisting microgravity-induced aging.

[0006] In a preferred embodiment, the therapeutic drug against microgravity-induced aging refers to a drug that stabilizes the cytoskeleton and inhibits cytoskeleton dissolution and / or decomposition.

[0007] In a more preferred embodiment, the therapeutic drug against microgravity-induced aging refers to a drug that stabilizes F-actin in the cytoskeleton, promotes its polymerization, and inhibits its dissolution and / or deconstruction.

[0008] More preferably, the therapeutic drug against microgravity-induced aging refers to a drug that inhibits the expression and / or activity of aging-related β-galactosidase.

[0009] In another preferred embodiment, microgravity refers to a state where the acceleration caused by gravity or other external forces does not exceed 10 -5 ~10 -4 g. Microgravity in the present invention refers to an environment in which the apparent weight of a system is much smaller than its actual weight under the effect of gravity. Environments that produce microgravity include, but are not limited to, drop towers, airplanes, rockets, and spacecraft.

[0010] Secondly, the application of the jasmine lactone of the present invention in the preparation of medicines for preventing and / or treating space diseases.

[0011] In a preferred embodiment, the space disease refers to a disease caused by organ and / or cell functional decline due to weightlessness in the space environment.

[0012] In a more preferred embodiment, the space disease includes one or more of the following symptoms: changes in cardiovascular function, muscle loss, osteoporosis, insulin resistance, decreased melatonin, decreased immunity, decreased cognitive function, optic nerve edema, and anemia.

[0013] Finally, the present invention provides a composition for preparing an anti-aging drug, wherein the composition contains jasmine lactone.

[0014] In a preferred embodiment, the composition further contains a pharmaceutically acceptable carrier.

[0015] The present invention provides the use of jasmine lactone in the preparation of anti-aging drugs. The jasmine lactone can inhibit the dissolution and / or deconstruction of F-actin that causes aging, stabilize the cytoskeleton, and thus inhibit or delay the aging process. The present invention also provides the use of jasmine lactone in the preparation of drugs for preventing and / or treating space diseases, and the jasmine lactone can inhibit the functional decline of organs and / or cells caused by weightlessness in the space environment. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 . Fluorescence microscope observation of cytoskeleton damage caused by microgravity; Figure 2 . Fluorescence microscopy observation of cytoskeleton stabilized by jasmine lactone; Figure 3 .Microscopic observation of jasmine lactone inhibiting sMG-induced cell senescence; Figure 4 . Statistical graph showing that jasmine lactone inhibits sMG-induced cell senescence. DETAILED DESCRIPTION

[0017] The present invention is further described in detail below in conjunction with specific embodiments, and the examples provided are only for illustrating the present invention, rather than for limiting the scope of the present invention. The examples provided below can be used as a guide for further improvements by those of ordinary skill in the art, and do not constitute a limitation of the present invention in any way.

[0018] The experimental methods in the following examples, unless otherwise specified, are all conventional methods, and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, etc. used in the following examples, unless otherwise specified, can all be obtained from commercial channels.

[0019] Unless otherwise specified, the quantitative tests in the following examples were performed three times and the results were averaged.

[0020] Example 1. Effects of microgravity on the cytoskeleton 1. Cultivating cells in microgravity: Dilute HeLa cells to a density of 2 × 10 5 / mL, gently blow and mix with a 5 mL pipette. Add 2 mL of cell suspension to the marked 35 mm dishes (NG (conventional control) group, sMG (microgravity simulation) group), respectively, and culture in a carbon dioxide incubator (5% CO2, 37 ℃). After 6-8 hours, when the cells are all attached to the wall, fill both 35 mm dishes with DMEM containing 10% FBS, exhaust the air in the 35 mm dishes with sterilized plastic wrap, and seal the dishes with disposable sterile sealing film and transparent tape. The sMG group was inserted into the microgravity simulator (environmentally controlled rotator, China Astronaut Center) for simulated microgravity culture (30 r / min, 37 ℃ rotation culture), and the NG group was directly placed in the environmentally controlled rotator for 48 hours.

[0021] 2. Microgravity disrupts the cytoskeleton Phalloidin staining was used to observe the changes in the F-Actin cytoskeleton of HeLa cells during simulated microgravity culture.

[0022] (1) Wash cells with PBS three times, 5 minutes each time; (2) Fix the cells with 4% paraformaldehyde solution in PBS on ice for 15 min; (3) Wash cells with PBS three times, 5 minutes each time; (4) Incubate cells with PBS solution containing 0.5% TritonX-100 for 10 minutes at room temperature; (5) Wash cells with PBS three times, 5 minutes each time; (6) Dilute the phalloidin stock solution to 20 nM with PBS, add it to the fixed cells, and leave it at room temperature for 20 minutes; (7) Wash the cells three times with PBS, 5 minutes each time. During the second wash, add DAPI to 1 ug / mL in PBS. (8) Fluorescence microscopy observation.

[0023] The results showed that the F-Actin fibers of HeLa cells in the NG group were continuously distributed throughout the cells, and thick actin bundles formed at the periphery of the cells, mostly lamellar pseudopodia. After sMG treatment, stress fibers disintegrated, and F-Actin fibers were mainly concentrated at the edge of the cells. The central actin bundles significantly depolymerized and fragmented, becoming more irregular and discontinuous, inducing the formation of filopodia at the periphery of the cells. The results showed that simulated microgravity treatment can reduce cell F-Actin polymerization, dissolve or destroy existing stress fibers ( Figure 1 ).

[0024] Example 2. Phalloidin staining to observe the effect of jasplakinolide on cytoskeleton stability Jasplakinolide is a potent actin polymerization inducer that stabilizes pre-existing actin filaments. We treated Hela cells with jasplakinolide (MedChemExpress, Cat. No. HYP0027) at a final concentration of 50 nM and stained with phalloidin. Figure 2 As shown, it was found that the F-actin fibers in the DMSO-treated cells of the control group were thinner and weaker, while the F-actin fibers in the Jasplakinolide-treated cells were significantly thicker and stronger than those in the DMSO group, indicating that Jasplakinolide can indeed stabilize the cytoskeleton.

[0025] Example 3. β-galactosidase staining to observe the inhibition of jasmine lactone on cell senescence In order to test whether Jasplakinolide has the function of antagonizing cell senescence under sMG conditions, HeLa cells were divided into four groups, namely NG+DMSO group, NG+Jasplakinolide group, sMG+DMSO group, sMG+Jasplakinolide group, and the final concentration of Jasplakinolide was 50nM. Among them, Jasplakinolide (MedChemExpress, Cat. No. HYP0027) was used to treat 50nM of HeLa cells.

[0026] HeLa cells were diluted to a density of 2 × 10 5 / mL, and gently blow and mix with a 5 mL pipette. 2 mL of cell suspension was added to the marked 35 mm dishes (NG+DMSO group, NG+Jasplakinolide group, sMG+DMSO group, sMG+Jasplakinolide group, the final concentration of Jasplakinolide was 50 nM), and cultured in a carbon dioxide incubator (5% CO2, 37°C). After 6-8 h, when the cells were all attached to the wall, the four 35 mm dishes were filled with DMEM containing 10% FBS, and the air in the 35 mm dishes was exhausted with sterile plastic wrap, and the dishes were sealed and fixed with disposable sterile sealing film and transparent tape. The two sMG groups were inserted into the microgravity simulator port for simulated microgravity culture (30 r / min, 37°C rotation culture), and the two NG groups were directly placed in the environment-controlled rotator for 48 hours.

[0027] The cultured cells were stained for senescence using the Bio-Tech Senescence Staining Kit. The Senescence β-Galactosidase Staining Kit is a kit for staining senescent cells or tissues based on the upregulation of senescence-associated β-galactosidase (SA-β-Gal) activity during senescence. The senescence of cells or tissues can be observed under an ordinary optical microscope.

[0028] Most normal cells are believed to have limited ability to divide, and enter a state of senescence when they cannot divide. At this time, the cells are still alive, but the expression profiles of the cells' genes and proteins have changed greatly. Senescent cells can no longer induce cell division under some conventional stimuli, and the cell cycle distribution of senescent cells is also quite special, which is different from some damage-induced cell dormancy and contact inhibition of cell growth. Senescent cells usually become larger in size and express β-galactosidase, which has high enzyme activity at pH 6.0. The Cell Senescence β-galactosidase Staining Kit uses X-Gal as a substrate, and generates a dark blue product under the catalysis of senescence-specific β-galactosidase. Therefore, it is easy to observe cells or tissues expressing β-galactosidase that have turned blue under an optical microscope.

[0029] 1. The experimental plan is as follows: (1) The four groups of cells were washed three times with PBS; (2) Add 1 mL of β-galactosidase staining fixative and fix at room temperature for 15 minutes (3) Remove the fixative solution and wash the cells three times with PBS, 5 minutes each time; (4) Aspirate PBS and add 1 mL of staining solution to each dish. The staining solution is prepared as follows: β-Galactosidase staining working solution A 10µl, β-Galactosidase staining working solution B 10µl, β-Galactosidase staining working solution C 930µl, X-gal solution 50 µl; (5) Incubate in a 37°C incubator overnight; (6) Observe and take photos under a microscope.

[0030] 2. Results SA-β-Gal staining revealed that only 3.01% of the cells in the DMSO-treated NG group had blue-green staining, while 59.99% of the cells in the DMSO-treated sMG group showed senescence staining, indicating that microgravity can cause cell aging. There was no significant change in senescence staining in the NG group cells treated with Jasplakinolide compared with the DMSO-treated NG group cells, with only 4.22% of the cells stained. The number of senescent cells in the sMG group cells treated with Jasplakinolide decreased significantly compared with the DMSO-treated sMG group cells, with only 8.69% of the cells senescent. This indicates that Jasplakinolide significantly inhibited cell senescence caused by sMG (Table 1, Figure 3-Figure 4 ).

[0031] Table 1. Comparison of the proportion of senescent cells in the four experimental groups .

Claims

1. Application of jasmine lactone in the preparation of therapeutic drugs against microgravity-induced aging.

2. The use according to claim 1, characterized in that: The therapeutic drug against microgravity-induced aging refers to a drug that stabilizes the cytoskeleton and inhibits the dissolution and / or deconstruction of the cytoskeleton.

3. The use according to claim 2, characterized in that: The therapeutic drug against microgravity-induced aging refers to a drug that stabilizes F-actin in the cytoskeleton, promotes its polymerization, and inhibits its dissolution and / or deconstruction.

4. The use according to claim 2, characterized in that: The therapeutic drug against microgravity-induced aging refers to a drug that inhibits the expression of aging-related β-galactosidase and / or inhibits its activity.

5. The use according to claim 1, characterized in that: Microgravity refers to a state where the acceleration caused by gravity or other external forces does not exceed 10 -5 ~10 -4 g.

6. Application of jasmine lactone in the preparation of drugs for preventing and / or treating space diseases.

7. The use according to claim 6, characterized in that: The space disease refers to a disease caused by the functional decline of organs and / or cells due to weightlessness in the space environment.

8. The use according to claim 7, characterized in that: The space diseases include one or more of the following symptoms: changes in cardiovascular function, muscle loss, osteoporosis, insulin resistance, decreased melatonin, decreased immunity, decreased cognitive function, optic nerve edema, and anemia.

9. A composition for preparing a therapeutic drug for anti-microgravity-induced aging, characterized in that: The composition contains jasmine lactone.

10. The composition according to claim 9, characterized in that The composition further contains a pharmaceutically acceptable carrier.

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