Application of miR-199a-3p in medicine for preventing and treating weight-loss-induced cardiac remodeling

Abstract

The invention provides an application of miR-199a-3p in a medicine for preventing and treating weight loss induced cardiac remodeling. According to the application disclosed by the invention, an adeno-associated virus 9 (AAV9) mediated myocardial specificity miR-199a-3p overexpression treatment experiment is carried out under a tail suspension simulation weightlessness condition, and the result shows that the miR-199a-3p can effectively relieve myocardial remodeling and functional impairment caused by weightlessness. The miR-199a-3p is used for preventing and treating weight loss induced cardiac remodeling, myocardial fibrosis can be remarkably reduced, and meanwhile the cardiac contraction function can be improved. The effects can effectively support the potential of using the miR-199a-3p as a target spot for treating weightlessness-induced cardiac remodeling, and the miR-199a-3p can be applied to prevention or treatment of weightlessness-related cardiac remodeling, and especially has a good application prospect in cardiovascular problems caused by long-term space flight.

Description

Technical Field

[0001] The invention belongs to the field of molecular biology and medical technology, and specifically relates to the use of miR-199a-3p in drugs for preventing and treating weightlessness-induced cardiac remodeling. Background Art

[0002] With the continuous advancement of human space technology, space flight, especially manned missions to Mars and civilian commercial space flight, has experienced explosive growth in recent years. This development not only gives more ordinary people the opportunity to enter space, but also makes humans face the challenge of longer space flight. In this context, the impact of microgravity environment on human health, especially the cardiovascular system, has become a key issue. During space flight, due to the lack of gravity and hydrostatic pressure gradients, the pressure and volume distribution in the cardiovascular system changes significantly, leading to fluid loss and reduced ventricular load. These changes can trigger abnormal activation of the nervous system, leading to cardiac remodeling and decreased function, seriously affecting the progress of space exploration.

[0003] In recent years, the role of non-coding RNA, especially microRNAs (miRNAs), in space flight experiments has attracted widespread attention. MiRNAs are small non-coding RNA molecules consisting of 19 to 25 nucleotides that regulate gene expression mainly by interfering with the translation of target mRNA or promoting its degradation. They play a key role in a variety of physiological and pathological processes, including cell differentiation, development, apoptosis, metabolism and proliferation. Summary of the invention

[0004] The purpose of the present invention is to provide a drug for preventing and / or treating cardiac remodeling caused by weightlessness using miR-199a-3p.

[0005] The purpose of the present invention is to adopt the following technical solutions to achieve:

[0006] Use of miR-199a-3p in drugs for preventing and treating weightlessness-induced cardiac remodeling.

[0007] Preferably, the nucleotide sequence of miR-199a-3p is:

[0008] 5'-ACAGUAGUCUGCACAUUGGUUA-3', the sequence that meets the requirements of the WIPOST.26 standard is: acagtagtctgcacattggtta.

[0009] Preferably, the cardiac remodeling is induced by the weightlessness environment during space flight.

[0010] Preferably, the drug is administered via intravenous injection of adeno-associated virus 9 (AAV9) to mediate overexpression of myocardial-specific miR-199a-3p.

[0011] Preferably, the dosage form of the drug includes injection and oral preparation.

[0012] Preferably, the encapsulation of the drug includes liposome encapsulation, nanoparticle encapsulation, microcapsule encapsulation, enteric coating encapsulation, and dendritic macromolecule encapsulation.

[0013] Compared with the prior art, the present invention has the following beneficial effects:

[0014] The present invention provides the use of miR-199a-3p in drugs for preventing and treating weightlessness-induced cardiac remodeling. The present invention conducts a myocardial-specific miR-199a-3p overexpression treatment experiment mediated by adeno-associated virus 9 (AAV9) under conditions of simulated weightlessness by tail suspension, proving that miR-199a-3p can effectively reduce myocardial remodeling and functional damage caused by weightlessness. By using miR-199a-3p as a drug for preventing and treating weightlessness-induced cardiac remodeling, myocardial fibrosis can be significantly reduced, and cardiac contractile function can be improved. The above effects can effectively support the potential of using miR-199a-3p as a target for treating weightlessness-induced cardiac remodeling, and can be used to prevent or treat weightlessness-related myocardial remodeling, especially in cardiovascular problems caused by long-term space flight. It has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 This is a flow chart of the detection of using tail vein injection of adeno-associated virus AAV9 to specifically overexpress miR-199a-3p in the mouse heart in Example 1 to resist cardiac remodeling caused by tail suspension simulating weightlessness.

[0016] Figure 2 In Example 1, the effects of tail-suspension-simulated weightlessness on the expression level of miR-199a-3p in the mouse heart and the effect of tail vein injection of adeno-associated virus to overexpress miR-199a-3p are shown.

[0017] Figure 3 In Example 1, the effect of overexpressing miR-199a-3p by tail vein injection of AAV9 on the ventricular ejection fraction (EF) of the cardiac function of mice after tail-suspension to simulate weightlessness.

[0018] Figure 4 In Example 1, the short-axis shortening rate (FS) of the cardiac function of mice after tail-suspension simulated weightlessness was affected by overexpression of miR-199a-3p via tail vein injection of AAV9.

[0019] Figure 5 This is the effect of overexpressing miR-199a-3p via tail vein injection of AAV9 on cardiac fibrosis and cardiomyocyte size in tail-suspended mice in Example 1.

[0020] Figure 6 This is the effect of AAV9 overexpression of miR-199a-3p on the cardiac index of tail-suspended mice in Example 1.

[0021] Figure 7 This is the effect of regulating miR-199a-3p on MEF2C protein level in cardiomyocytes in Example 1. DETAILED DESCRIPTION

[0022] The technical solution is further described below in conjunction with the accompanying drawings and specific embodiments to help understand the content of the present invention.

[0023] Example 1

[0024] For the sake of clarity, the following content describes the embodiments of the present invention, and these embodiments are only for the convenience of understanding the present invention, and are not intended to limit the scope thereof. Unless otherwise specified, the reagents used in the following examples are provided by conventional biochemical reagent companies, and the detection methods adopted are also standard experimental techniques. All quantitative detections were repeated three times, and averaged to ensure the reliability of the data. In addition, the recognized standard experimental procedures were strictly followed during the experiment, and the results were statistically analyzed to ensure its scientificity and credibility.

[0025] Use of miR-199a-3p in drugs for preventing and treating weightlessness-induced cardiac remodeling. The following experiments and data are used to demonstrate the effectiveness of the use of miR-199a-3p in drugs for preventing and treating weightlessness-induced cardiac remodeling.

[0026] MiR-199a-3p counteracts cardiac remodeling induced by tail-suspension simulated weightlessness:

[0027] 1. The nucleotide sequence of miR-199a-3p is: 5'-ACAGUAGUCUGCACAUUGGUUA-3', and the sequence that meets the requirements of the WIPOST.26 standard is: acagtagtctgcacattggtta.

[0028] 2. Establishment of simulated microgravity model:

[0029] The experiment selected 3-month-old mice. All mice were raised under specific pathogen-free (SPF) conditions, with a light-dark cycle of 12:12 hours and a temperature of 23°C. At the beginning of the experiment, medical tape or cloth was gently wrapped around the upper two-thirds of the mouse's tail and connected to a suspension device so that the mouse was suspended at a 30° angle, with only the forelimbs touching the ground and the hindlimbs hanging in the air. The appropriate suspension height was ensured to avoid excessive pressure on the tail, while ensuring that the mouse could move freely and obtain food and water. During the experiment, the health status and behavioral performance of the mice were monitored daily. After 6 weeks of continuous tail suspension to simulate weightlessness, it was used for subsequent experiments.

[0030] 3. Real-time quantitative PCR detection of miR-199a-3 expression level in cardiac tissue:

[0031] RNA extraction: First, take a certain amount of heart tissue and grind it. Add 1ml of TRIzol reagent to the ground tissue, mix thoroughly and let it stand for 5 minutes, then add 200μl of chloroform, shake vigorously for 15 seconds, let it stand for 3 minutes and then centrifuge. Take the supernatant (the aqueous phase where the RNA is located) to a new RNase-free centrifuge tube, add an equal volume of isopropanol and mix well, and let it stand at room temperature for 10 minutes to precipitate the RNA. After washing the RNA precipitate with 75% ethanol and drying it, dissolve the RNA with RNase-free water, and measure the RNA concentration and purity.

[0032] Reverse transcription and real-time quantitative PCR system: After the adapter is connected, reverse transcription reaction is performed, using a reaction mixture containing reverse transcriptase, and operating according to the specific reagent instructions. Next, prepare the real-time quantitative PCR reaction system: each reaction volume is 20μl, including 10μl TB Green Premix Ex Taq II, 0.8μl miR-199a-3p specific primer (10μM), 0.8μl internal reference primer (such as U6, 10μM), 2μl reverse transcribed cDNA template (1:5 dilution), and 6.4μl RNase-Free Water. The reaction mixture is amplified on a real-time quantitative PCR instrument.

[0033] Real-time quantitative PCR program and data analysis: The real-time quantitative PCR amplification program is as follows: 95°C pre-denaturation for 30 seconds, followed by 40 cycles, each cycle including 95°C denaturation for 5 seconds and 60°C annealing / extension for 30 seconds. Three technical replicates should be set up for each sample to ensure the accuracy of the data. After amplification, the cycle threshold (Ct value) was automatically calculated using the real-time PCR instrument software, and the relative expression of miR-199a-3p was calculated by the 2^-ΔΔCt method (ΔCt = Ct (miR-199a-3p) - Ct (U6 internal reference)). The difference in miR-199a-3p expression in each group can be analyzed by statistical software. During the experiment, all operations should be performed in an RNase-free environment, and a no-template control (NTC) should be used to monitor potential contamination in reverse transcription and PCR reactions, while ensuring the consistency of the concentration and volume of reagents and samples to ensure the reliability and repeatability of the experimental results.

[0034] 4. Echocardiography is used to assess cardiac function in mice:

[0035] First, mice were anesthetized with an induction dose of 3% isoflurane. After the mice lost consciousness, the anesthesia concentration was maintained at 2% to ensure that the mice were unconscious and maintained stable breathing throughout the process. Subsequently, the mice were placed in a supine position to expose the chest area for ultrasound imaging. Cardiac function was evaluated using the Vevo 1100 echocardiography system (Visual-Sonics, Toronto, Canada). Two-dimensional images of the parasternal long-axis and short-axis views were obtained with the 2D guidance of the system, and M-mode ultrasound recordings were performed in the mid-left ventricular region. Three cardiac cycles were selected to measure the average thickness of the anterior and posterior walls of the left ventricle, as well as the end-diastolic internal diameter (LVIDd) and the end-systolic internal diameter (LVIDs), and the short-axis shortening fraction (FS) and ejection fraction (EF) were calculated. All data were analyzed without the experimenters knowing the experimental groups to ensure the objectivity and accuracy of the results.

[0036] 5. Histopathological analysis:

[0037] Part of the collected heart tissue was prepared into paraffin sections, and the other part was prepared into frozen sections for subsequent analysis. Paraffin sections were used for hematoxylin-eosin (H&E) staining and Masson trichrome staining to evaluate tissue structure and fibrosis. Frozen sections were used for fluorescent staining, and the myocardial cell membrane structure was observed by FITC-labeled wheat germ agglutinin (WGA). The specific operations included: paraffin sections were dewaxed and hydrated, and then hematoxylin staining, eosin staining or trichrome staining were performed in sequence; frozen sections were fixed with 4% paraformaldehyde, stained with diluted FITC-WGA solution, and incubated in the dark. After staining, the sections were washed with PBS and sealed with a mounting medium containing Hoechst. Finally, paraffin sections and frozen sections were observed using optical microscopy and fluorescence microscopy, respectively, to obtain detailed information of tissues and cells.

[0038] 6. Analysis of protein expression changes of MEF2C by Western Blot:

[0039] Mouse heart tissue was homogenized by using a Power Gen125 tissue homogenizer (Fisher Scientific) to extract proteins. Tissue or cell samples were lysed in RIPA lysis buffer containing protease and phosphatase inhibitors and incubated on ice for 30 minutes. After lysis, the samples were centrifuged at 12,000 rpm for 10 minutes at 4°C, and the supernatant was collected. Protein concentration was determined using a BCA protein quantification kit (Thermo Fisher Scientific) according to the manufacturer's instructions. Equal amounts of protein (usually 20-40 μg) were separated on SDS-PAGE gels and then transferred to nitrocellulose membranes. After transfer, the membranes were blocked with 5% skim milk for 1 hour at room temperature. Subsequently, the membranes were incubated with specific primary antibodies overnight at 4°C. The next day, the membranes were washed three times with TBST buffer (Tris-buffered saline containing Tween-20) for 10 minutes each, followed by incubation with HRP-labeled secondary antibodies for 1 hour at room temperature. The membranes were then washed again with TBST three times for 10 minutes each. Finally, enhanced chemiluminescence (ECL) reagent was used to detect protein bands, and GAPDH was used as an internal control to normalize the protein expression level.

[0040] Figure 1 The experimental design process is presented, in which the AAV9 delivery system is used to specifically overexpress miR-199a-3p in the mouse heart to counteract the cardiac remodeling induced by tail-suspension simulated weightlessness. In the experiment, the mice were subjected to tail-suspension simulated weightlessness for 6 weeks, and echocardiography and tissue sample collection were performed in the sixth week to evaluate cardiac function and remodeling.

[0041] AAV9 injection: AAV9 is a serotype of adeno-associated virus (Adeno-Associated Virus). This viral vector is of great significance in the field of gene therapy because it can efficiently cross the blood-brain barrier and infect neuronal cells. "AAV9 injection" usually refers to the use of AAV9 as a vector for injection.

[0042] Tail Suspension: Tail suspension.

[0043] Echo: Feedback.

[0044] Collect Tissues: Collect tissue samples.

[0045] Week: Week is the time unit when mice are suspended by their tails to simulate weightlessness.

[0046] Figure 2The expression characteristics of miR-199a-3p in the hearts of wild-type mice after injection of AAV9 to overexpress miR-199a-3p were shown. The results of qPCR experiments showed that tail-hanging simulated weightlessness treatment led to a significant decrease in the expression of miR-199a-3p, and the expression level of miR-199a-3p in the heart was significantly increased after injection of AAV9--199a-3p.

[0047] Relative miR-199a-3p levels in heart(I U6): The expression levels of miR-199a-3p relative to the internal reference gene U6 in the heart tissue of experimental mice.

[0048] AAV9-NC: Adeno-associated virus type 9 (AAV9) was used as a vector and loaded with a non-coding sequence experimental control group.

[0049] AAV9-199A-3P: An experimental group using AAV9 as a vector loaded with a specific miRNA, miR-199a-3p.

[0050] Ctrl: control group.

[0051] TS: Test Sample: “TS” refers to the test sample, which is analyzed in comparison with the control group.

[0052] Figure 3 and Figure 4 The results of the analysis of cardiac function indicators EF and FS in AAV9-NC mice and AAV9-miR-199a-3p mice in the tail-hanging simulated weightlessness experiment were presented. The experimental results showed that after the tail-hanging simulated weightlessness treatment, the ventricular ejection fraction (EF) ( Figure 3 ) and fractional shortening (FS) ( Figure 4 ) decreased significantly, indicating a decrease in cardiac function. However, this decrease in cardiac function was effectively resisted in mice with AAV9-mediated cardiac-specific overexpression of miR-199a-3p.

[0053] Figure 5 The results of HE staining, MTT staining, and WGA staining of the hearts of AAV9-NC mice and AAV9-miR-199a-3p mice after tail-hanging simulated weightlessness experiments were shown. The results showed that the tail-hanging simulated weightlessness hearts of AAV9-NC mice showed obvious myocardial fibrosis and a decrease in myocardial cell area, while these changes were effectively alleviated in AAV9-miR-199a-3p mice.

[0054] WGA: refers to the use of wheat germ agglutinin (WGA) to stain heart tissue. WGA is a lectin that can bind to specific sugars on the cell surface and is often used to mark extracellular matrix and cell membranes.

[0055] MTT: refers to a cell viability assay using MTT (3-(4,5-dimethylthiazole-2)-2,5-diphenyltetrazolium bromide). MTT is a yellow dye that, when it enters living cells, is reduced by dehydrogenases in the mitochondria into a water-insoluble purple crystalline form called formazan. MTT staining is mainly used to assess cell survival and metabolic activity, and is commonly used in cytotoxicity experiments, drug screening, and cell proliferation studies.

[0056] HE: refers to "Hematoxylin and Eosin Staining", which is a conventional histological staining technique widely used in pathology and biological research.

[0057] Figure 6 The results showed that the cardiac index (heart weight / body weight) of AAV9-NC mice and AAV9-miR-199a-3p mice after tail-suspension simulated weightlessness were analyzed. The results showed that the injection of AAV9-miR-199a-3p could effectively alleviate the decrease in cardiac index caused by tail-suspension simulated weightlessness.

[0058] HW / TL (mg / cm): refers to the measurement result of "heart weight to tibia length ratio", where: HW (Heart Weight) refers to heart weight; TL (Tibia Length) refers to tibia length. This ratio is a commonly used physiological indicator used to evaluate changes in heart size relative to body size. In heart disease research, the ratio of heart weight to tibia length is often used to measure the degree of cardiac hypertrophy. Tibia length is often used as a stable indicator of growth status in animal models (such as mice or rats) because tibia length is related to body weight but is relatively stable and not easily affected by short-term changes in nutritional status.

[0059] The ratio of heart weight to tibia length is calculated as follows: HW / TL = tibia length (cm) / heart weight (mg). The significance of this ratio is:

[0060] 1. Assessment of cardiac hypertrophy: Under cardiac pathological conditions, such as hypertension and myocardial infarction, the heart will become larger to adapt to the load, which is manifested as increased heart weight. By measuring HW / TL, it is possible to assess whether the heart has hypertrophy or enlargement.

[0061] 2. Standardized indicators: Due to the differences in body weight between different animals, the simple heart weight cannot accurately reflect the relative size of the heart. By calculating the ratio of heart weight to tibia length, the influence of body weight differences between individuals can be eliminated, providing a more standardized indicator.

[0062] 3. Efficacy evaluation: When studying treatments for heart disease, changes in the HW / TL ratio can be used to evaluate whether the treatment measures are effective, such as whether cardiac hypertrophy is reduced.

[0063] Figure 7 The results showed that miR-199a-3p regulates MEF2C protein levels in cardiomyocytes. The results showed that overexpression of miR-199a-3p mimics could significantly reduce MEF2C protein levels, which is a key regulator of cardiac remodeling, indicating that miR-199a-3p can regulate myocardial remodeling by targeting MEF2C.

[0064] GAPDH: stands for "Glyceraldehyde-3-Phosphate Dehydrogenase". GAPDH is an enzyme that plays an important role in the glycolysis pathway and is present in almost all eukaryotic cells. Because its expression in cells is usually relatively constant, GAPDH is often used as an internal control (housekeeping gene) to standardize protein expression levels. In heart disease research, GAPDH can be used for the following purposes:

[0065] 1. Protein expression standardization: In protein detection techniques such as Western blot, GAPDH is often used as a loading control to ensure that the total amount of protein between different samples is consistent. By comparing the signal intensity of the target protein with that of GAPDH, differences that may be introduced during sample preparation or transfer can be corrected.

[0066] 2. Gene expression analysis: In molecular biology experiments such as real-time quantitative PCR (qPCR), GAPDH is also used as a reference gene to standardize the expression level of the target gene to evaluate the effect of experimental treatment on gene expression.

[0067] 3. Metabolic studies: Although GAPDH is most commonly used as an internal reference gene, it is also an important metabolic enzyme that participates in the glycolysis process. In some studies, the expression and activity of GAPDH may also be the subject of study, especially when exploring the association between metabolic changes and heart disease.

[0068] In conclusion, GAPDH is often used as a control in graphs and tables of heart disease research to verify the reliability and consistency of experimental results. By comparing with the GAPDH signal, the expression changes of specific proteins or genes under experimental conditions can be more accurately assessed.

[0069] MEF2C: stands for "Myocyte Enhancer Factor 2C". MEF2C is a transcription factor that is part of the MEF2 family, whose members play a key role in muscle development and function. MEF2C plays an important role in heart development, cardiomyocyte differentiation, and maintenance of heart function. The functions of MEF2C include:

[0070] 1. Heart development: MEF2C regulates the expression of multiple genes during heart development and is essential for the differentiation of cardiac progenitor cells.

[0071] 2. Cardiomyocyte differentiation: MEF2C participates in the differentiation process of cardiomyocytes and affects cardiomyocyte-specific gene expression.

[0072] 3. Regulation of cardiac function: In the mature heart, MEF2C is involved in regulating the expression of genes related to cardiac contraction and relaxation, which is very important for maintaining normal cardiac function.

[0073] 4. Heart disease: MEF2C also plays a role in heart disease. For example, under pathological conditions such as myocardial hypertrophy and heart failure, the expression and activity of MEF2C may change, affecting heart function.

[0074] In heart disease research, MEF2C is often used to indicate the expression of genes related to heart disease. Researchers may evaluate the pathological mechanism of heart disease or the effect of specific treatments by detecting changes in the expression level of MEF2C and the downstream genes it regulates. For example, by measuring the expression level of MEF2C through Western blot, qPCR and other techniques, we can understand its changing trend under different experimental conditions and further infer its role in the development of heart disease.

[0075] NC: stands for “Negative Control”.

[0076] Mimics: Refers to "simulators" or "simulators." In RNA research, especially research involving microRNA (miRNA), miRNA mimics are synthetic double-stranded RNA molecules that mimic the function of a specific miRNA. miRNA mimics are used to upregulate the expression level of a specific miRNA to study the function of that miRNA and its role in heart disease. For example, if a study finds that a certain miRNA is downregulated in heart disease, researchers might use a mimic of that miRNA to restore its expression level and observe the effects on heart cells or tissues.

[0077] HL-1: refers to a cardiac myocyte cell line, namely HL-1 cells. HL-1 cells are a cardiac myocyte cell line immortalized from AT-1 mouse atrial myocytes by Dr. William C. Claycomb and others. These cells retain many characteristics of cardiac myocytes and are therefore widely used in the study of heart diseases.

[0078] H9C2: refers to the H9C2 cell line. The H9C2 cell line is an immortalized cell line isolated and established from rat embryonic heart tissue. These cells exhibit some characteristics of cardiac myocytes, but are not fully mature cardiomyocytes. H9C2 cells are widely used in heart disease research, especially in the preliminary screening of drugs and the study of the physiological and pathological processes of cardiac myocytes.

[0079] In summary, this study revealed the role of miR-199a-3p in cardiac remodeling induced by tail-suspension simulated weightlessness in mice. The results showed that the cardiac function of mice treated with tail-suspension simulated weightlessness decreased, such as a significant decrease in cardiac ejection fraction (EF) and fractional shortening (FS), accompanied by a higher degree of myocardial fibrosis. In mice with cardiac-specific overexpression of AAV9-miR-199a-3p, the changes in these cardiac remodeling-related indicators were significantly alleviated, indicating that miR-199a-3p overexpression has a protective effect on myocardial remodeling caused by tail-suspension simulated weightlessness.

[0080] Through animal experiments, the present invention proves that overexpression of miR-199a-3p can effectively alleviate myocardial remodeling and functional decline caused by weightlessness.

[0081] miR-199a is a member of the microRNA (miRNA) gene family that produces two mature miRNA molecules, which are called miR-199a-5p and miR-199a-3p. These mature miRNAs are cleaved from different parts of a larger precursor miRNA molecule. Specifically, miR-199a-3p is a fragment cleaved from the 3' end of the precursor RNA, while the other mature form, miR-199a-5p, is the part cleaved from the 5' end of the precursor RNA.

[0082] Although miR-199a-3p and miR-199a-5p are produced from the same precursor, they usually have different sequences and may have different target genes and functions. This means that they may regulate different signaling pathways or biological processes in cells. For example:

[0083] miR-199a-3p has been shown to play an important regulatory role in a variety of cancers. As mentioned above, it can act as a tumor suppressor gene in esophageal squamous cell carcinoma, and may also participate in the formation and development of bladder cancer by regulating the PI3K-Akt-mTOR signaling pathway. In addition, it may also participate in regulating the sugar metabolism of testicular tumor cells.

[0084] miR-199a-5p has also been found to be associated with cancer in some studies. For example, in myocardial fibrosis, it promotes myocardial fibrosis by targeting Sirt1.

[0085] Although they are both produced from the same precursor miRNA, the functions of miR-199a-3p and miR-199a-5p are not the same, and may even be opposite. This is because they recognize different mRNA sequences, thereby regulating different gene expressions. Therefore, when studying the function of miRNA, it is very important to distinguish between specific miRNAs (such as -5p or -3p). Each miRNA has its own unique function and mechanism of action, which is crucial for understanding the biological basis of the disease and finding new therapeutic targets.

[0086] miR-199a is a miRNA gene that can produce two main mature forms: miR-199a-5p and miR-199a-3p. These two mature miRNA molecules are not the same, they have the following differences:

[0087] Sequence differences: Although they are generated from the same precursor miRNA, they originate from different ends of this precursor. miR-199a-5p comes from the 5' end of the precursor, while miR-199a-3p comes from the 3' end. Because of their different sequences, they each have different pairing rules to recognize target mRNAs.

[0088] Target genes: Because miR-199a-5p and miR-199a-3p have different sequences, they tend to bind to different mRNA targets, which means they can regulate different gene expressions and thus participate in different biological processes or signaling pathways.

[0089] Functional differences: Due to their different target genes, miR-199a-5p and miR-199a-3p may also have different functions within the cell. For example, miR-199a-3p may function as a tumor suppressor gene in some types of cancer, while miR-199a-5p may have different functions in other contexts.

[0090] Expression pattern: miR-199a-5p and miR-199a-3p may have different expression levels in different tissues or cell types, depending on specific physiological or pathological conditions.

[0091] In summary, although miR-199a-5p and miR-199a-3p are generated from the same miRNA precursor molecule, they are different miRNA molecules with their own characteristics, targets and functions. Therefore, in scientific research, it is very important to distinguish these two mature miRNA molecules in order to accurately understand their respective roles in biological processes.

[0092] Through tail-suspension simulated weightlessness conditions, adeno-associated virus 9 (AAV9)-mediated myocardial-specific miR-199a-3p overexpression therapy experiments were conducted, and it was found that miR-199a-3p can effectively reduce myocardial remodeling and functional damage caused by weightlessness. The results showed that miR-199a-3p significantly reduced myocardial fibrosis and improved cardiac contractile function. These results support the potential of miR-199a-3p as a therapeutic target, which can be used to prevent or treat myocardial remodeling related to weightlessness, especially in cardiovascular problems caused by long-term space flight.

[0093] Example 2

[0094] miR-199a-3p can be used to screen and develop drugs to prevent or treat cardiac remodeling induced by weightlessness.

[0095] Furthermore, the cardiac remodeling is caused by weightlessness conditions during space flight.

[0096] Furthermore, the drug is administered via intravenous injection of adeno-associated virus 9 (AAV9) to mediate overexpression of myocardial-specific miR-199a-3p.

[0097] The nucleotide sequence of the miR-199a-3p is: 5'-ACAGUAGUCUGCACAUUGGUUA-3', and the sequence that meets the requirements of the WIPOST.26 standard is: acagtagtctgcacattggtta.

[0098] The dosage forms of the drug include injection and oral preparation.

[0099] The encapsulation of the drug includes liposome encapsulation, nanoparticle encapsulation, microcapsule encapsulation, enteric coating encapsulation, and dendritic macromolecule encapsulation.

[0100] Furthermore, miR-199a-3p inhibited cardiac remodeling and protected cardiac function by targeting the MEF2C pathway.

[0101] The above are merely embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention are included in the scope of the claims of the present invention.

Claims

1. Use of miR-199a-3p in the prevention and treatment of weightlessness-induced cardiac remodeling.

2. The use of miR-199a-3p in a drug for preventing and treating weightlessness-induced cardiac remodeling according to claim 1, characterized in that: The nucleotide sequence of miR-199a-3p is: 5'-ACAGUAGUCUGCACAUUGGUUA-3'.

3. The use of miR-199a-3p in a drug for preventing and treating weightlessness-induced cardiac remodeling according to claim 1, characterized in that: The cardiac remodeling is induced by the weightless environment of space flight.

4. The use of miR-199a-3p in a drug for preventing and treating weightlessness-induced cardiac remodeling according to claim 1, characterized in that: The drug is administered intravenously via adeno-associated virus 9 (AAV9) to mediate overexpression of myocardial-specific miR-199a-3p.

5. The use of miR-199a-3p in a drug for preventing and treating weightlessness-induced cardiac remodeling according to claim 1, characterized in that: The dosage forms of the drug include injection and oral preparation.

6. The use of miR-199a-3p in a drug for preventing and treating weightlessness-induced cardiac remodeling according to claim 1, characterized in that: The encapsulation of the drug includes liposome encapsulation, nanoparticle encapsulation, microcapsule encapsulation, enteric coating encapsulation, and dendritic macromolecule encapsulation.

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