Application of RING finger protein 130 in the treatment of calcific aortic valve disease

Abstract

The present invention discloses the application of RING finger protein 130 (RNF130) in the treatment of calcific aortic valve disease, and belongs to the field of biomedicine technology. The present invention specifically relates to the application of RNF130 and shRNA that targets and knocks down the RNF130 gene in valve cells in the preparation of anti-calcific aortic valve disease (CAVD) drugs. The present invention found that RNF130 has obvious calcified lesions in the aortic valve of CAVD patients, and the expression of RNF130 is significantly increased, and the shRNA of RNF130 can effectively reverse the osteogenic phenotype conversion and calcium deposition of hVICs induced by calcification culture medium, indicating that RNF130 has the potential function of becoming a pharmaceutical target for the treatment of CAVD.

Description

Technical Field

[0001] The invention provides application of RING finger protein 130 (RNF130) in treating calcific aortic valve disease, belonging to the technical field of biomedicine. Background Art

[0002] Calcific aortic valve disease (CAVD) is one of the most common degenerative aortic valve diseases. It refers to the aortic valve fibrosis and calcium salt deposition leading to valve stiffness and calcification, and the resulting left ventricular outflow tract obstruction. With the decrease in the incidence of rheumatic heart disease and the increase in age, CAVD has become the third most common cardiovascular disease after coronary heart disease and hypertension. The early manifestations of calcific aortic valve are mainly aortic valve sclerosis, without left ventricular outflow tract obstruction caused by thickening, calcification and stiffness of the valve leaflets. Its incidence rate is about 25% in people over 65 years old. As the disease progresses, late-stage calcific aortic valve stenosis will lead to left ventricular outflow tract obstruction, and its incidence rate can reach 2% in people over 65 years old.

[0003] CAVD is the most common type of valvular disease. To date, the pathogenesis of CAVD has not been fully elucidated, but histopathological studies have shown that CAVD involves a variety of pathological processes, including high circulatory resistance and abnormal valve tension, chronic inflammatory response, extracellular matrix remodeling, lipid metabolism disorders, increased osteogenic factors or decreased activity of calcification inhibitors, neovascularization and even genetics. Although the diagnosis and treatment of CAVD have undergone tremendous changes in the past fifty years, surgical aortic valve replacement is still the only effective treatment, and drugs can only palliatively delay symptoms. However, due to the advanced age of patients, heart failure or other underlying diseases, nearly 1 / 3 of patients cannot undergo surgery. Therefore, finding drug treatment targets for CAVD and exploring their mechanisms of action, intervening in key targets to improve aortic valve calcification and stenosis may become one of the important strategies for delaying and treating CAVD.

[0004] Currently, the treatment of calcific aortic valve disease is still challenging, especially in early prevention and treatment. In clinical practice, some drugs have been used to treat this disease, but these drugs still have some defects and shortcomings.

[0005] Commonly used drugs are lipid-lowering drugs, such as statins. These drugs slow down the process of arterial calcification by lowering cholesterol levels, thereby improving the function of the aortic valve to a certain extent. However, lipid-lowering drugs cannot completely prevent or reverse calcification that has already occurred. In addition, long-term use of lipid-lowering drugs may have some adverse reactions, such as muscle pain and muscle damage, elevated liver enzymes or other liver function abnormalities, gastrointestinal discomfort, etc., which require close monitoring. Another class of drugs is anti-calcification drugs, such as antioxidants and anti-inflammatory drugs. The mechanism of action of these drugs is mainly to reduce oxidative stress and inflammatory response, thereby slowing down or inhibiting the process of arterial calcification. However, there is currently insufficient clinical trial evidence to support the effectiveness and safety of these drugs in the treatment of calcific aortic valve disease.

[0006] Although some drugs have been tried in the treatment of calcific aortic valve disease, there are still many challenges, such as: the pathogenesis of calcific aortic valve disease is not fully understood, resulting in insufficient targeted treatment strategies; for CAVD, existing drugs have limited effectiveness in treating early-stage disease and preventing progression. Summary of the invention

[0007] RING finger protein 130 (RNF130) is a protein belonging to the RING finger protein family. It plays the role of E3 ubiquitin ligase in cells and participates in regulating the ubiquitination modification process in cells, thereby regulating protein degradation and signal transduction, and further affecting a variety of biological processes. Although some breakthroughs have been made in the existing RNF130 research, there are still many unresolved issues. For example, whether the expression of RNF130 in other diseases is abnormal and its role in the interaction with other signaling pathways are worthy of further in-depth exploration. In addition, how to make full use of the characteristics of RNF130 to develop innovative therapeutic strategies and its potential value in clinical applications require further in-depth research and exploration. In response to the above problems, the present invention uses RNF130 as a pharmaceutical target for the treatment of CAVD for the first time, and designs targeted shRNA with the function of treating CAVD based on RNF130. The present invention is of breakthrough significance and provides new ideas for finding effective CAVD treatment methods and drug preparation.

[0008] The first object of the present invention is to provide a use of RNF130 in the preparation of a drug for preventing, treating or diagnosing calcific aortic valve disease.

[0009] In one embodiment, the protein ID of RNF130 is NP_001267730.11 and the gene ID is 55819.

[0010] In one embodiment, the calcific aortic valve disease comprises calcific aortic stenosis or calcific aortic regurgitation or calcific aortic valvulopathy.

[0011] In one embodiment, the medicament reduces the expression of RNF130 protein or gene.

[0012] In one embodiment, the drug contains an inhibitor of RNF130 protein.

[0013] The second object of the present invention is to provide a method for studying the gene function of calcific aortic valve disease, or screening drugs for preventing or treating calcific aortic valve disease, or constructing a calcific aortic valve disease model, or analyzing gene therapy for calcific aortic valve disease, the method comprising using an adeno-associated virus (AAV) shRNA interference vector containing a specific aortic valve RNF130 gene to reduce the expression of RNF130.

[0014] In one embodiment, the sequence of shRNA is shown as SEQ ID NO.1-2.

[0015] In one embodiment, the method can prevent or treat calcific aortic valve disease in in vitro cells, or in vivo animal models, or in drug screening.

[0016] The third object of the present invention is to provide a drug for preventing or treating calcific aortic valve disease, wherein the drug can inhibit the expression of RNF130 gene or protein.

[0017] In one embodiment, the medicament prevents or treats calcific aortic valve disease by regulating the RNF130 signaling pathway or bone differentiation marker protein.

[0018] In one embodiment, the drug is a pharmaceutically acceptable carrier or excipient.

[0019] Optionally, the drug is a vector for gene therapy, including vectors for gene editing, gene expression regulation or gene delivery; or the drug is a vector for cell therapy, including stem cells, immune cells or repair cells.

[0020] Optionally, the dosage form of the drug includes oral solution, injection, tablet, pill, dispersant, capsule, pellet, granule, suspension, emulsion.

[0021] Beneficial effects:

[0022] (1) The present invention uses proteomics technology to detect that the protein expression of RNF130 in the aortic valve of CAVD patients changes. CAVD patients have obvious calcification lesions in the aortic valve area, and the expression of RNF130 is significantly increased;

[0023] (2) The expression of RNF130 was significantly upregulated in CAVD mouse models and human aortic valve interstitial cells (hVICs) treated with calcification medium;

[0024] (3) RNF130 shRNA can effectively reverse the osteogenic phenotype conversion and calcium deposition of hVICs induced by calcification medium, indicating that RNF130 has the function of becoming a pharmaceutical target for the treatment of CAVD. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 Upregulated RNF130 expression and calcification in aortic valves of patients with CAVD. Figure 1 A is a heat map showing different protein expression patterns in human aortic valve samples; Figure 1 B is Western blot detection of the protein levels of RNF130, Runx2, BMP-2, Osterix, and Osteocalcin in the aortic valve of CAVD patients; Figure 1 C is qPCR detection of RNF130 mRNA levels in aortic valves of CAVD patients; Figure 1 D shows the aortic valves of the Non-CAVD and CAVD groups stained with Alizarin Red (ARS), Von Kossa (VK), DAPI, RNF130 and α-SMA, as well as the merged images (DAPI, RNF130 and α-SMA).

[0026] Figure 2 When the calcification environment is upregulated, RNF130 expression in cells is upregulated. Figure 2 A and 2B are RT-PCR and western blot to detect the mRNA and protein expression of RNF130 in hVICs incubated with OGM calcification medium for 0, 3, 5, 7, and 10 days, respectively; Figure 2 C is immunofluorescence observation of RNF130 expression in hVICs treated with OGM calcification medium for 7 days; Figure 2 D is the qPCR detection of RNF130 mRNA level after hVICs were incubated in OGM calcification medium for 7 days; Figure 2 E is qPCR detection of 3 mM Na 3 PO 4 The mRNA level of RNF130 after incubation of hVICs for 7 days; Figure 2F is Western blot detection of ApoE - / - Protein expression of RNF130 in mouse aortic valve; Figure 2 G is Western blot detection of LDLR - / - Protein expression of RNF130 in mouse aortic valve.

[0027] Figure 3 Inhibition of RNF130 can alleviate calcification by inhibiting the conversion of hVICs to an osteoblast-like phenotype. Figure 3 A is RT-PCR detection of the mRNA levels of Runx2, BMP-2, Osterix, and Osteocalcin after RNF130 shRNA interference; Figure 3 B is Western blot detection of the protein levels of Runx2, BMP-2, Osterix, and Osteocalcin after RNF130 shRNA interference; Figure 3 C is the ALP activity kit to detect ALP activity in hVICs; Figure 3 D. Calcium content kit to detect the calcium content in hVICs; Figure 3 E: Alizarin red mineralized nodules stained to observe calcium deposition in hVICs. DETAILED DESCRIPTION

[0028] 1. Human aortic valve sample collection and processing:

[0029] (1) Patient selection and preparation: CAVD and non-CAVD patients who required surgery were selected. Before collecting valve tissue samples, informed consent was obtained from the patients, and the sample collection process and research purpose were informed;

[0030] (2) Sample processing: After collecting the heart valve tissue samples, they are quickly placed in liquid nitrogen to prevent protein degradation and denaturation, and sent to the laboratory as soon as possible for follow-up experiments.

[0031] 2. Proteomics:

[0032] Mass spectrometry is one of the most commonly used methods in proteomics, which can be used to identify and quantify proteins in samples. In protein mass spectrometry analysis, heart valve tissue samples are first subjected to liquid chromatography protein separation technology, and proteins are enzymatically cleaved into small fragments (peptides). These peptides are analyzed by a mass spectrometer to generate a mass spectrum. By matching the mass spectrometry data of known proteins in the database, proteins in heart valve tissue samples can be identified.

[0033] 3. Construction of RNF130 shRNA and AAV viral vector

[0034] (1) Designing and synthesizing RNF130 gene-specific shRNA sequences (RNF130 shRNA sequence, SEQ ID NO.1: Forward: 5'-GGTTCTTTGTCCCTCCTAATA-3'; SEQ ID NO.2: Reverse: 5'-TATTAGGAGGGACAAAGAACC-3');

[0035] (2) constructing a shRNA vector including a promoter suitable for hVICs and a marker gene for screening;

[0036] (3) Clone the synthesized shRNA sequence into the vector and verify the accuracy of the cloning;

[0037] (4) preparing RNF130 gene knockdown AAV viral vector and using it to infect hVICs;

[0038] (5) Use molecular biological methods such as Western blotting and PCR to verify whether the expression level of the RNF130 gene has been successfully knocked down and evaluate the degree of calcification.

[0039] 4. Experimental animals and animal models:

[0040] Strictly refer to international authoritative magazines and purchase ApoE directly - / - Mice and LDLR - / - Mice, 8-week-old ApoE - / - Mouse or LDLR - / - Mice were fed a high cholesterol diet for 24 weeks to maintain chow-fed ApoE - / - Mouse or LDLR - / - Mice served as controls.

[0041] 5. In vitro cell calcification model and grouping:

[0042] The cell line is human aortic valve interstitial cells (hVICs). Two in vitro cell calcification models were constructed with strict reference to international authoritative journals.

[0043] (1) Cell calcification model induced by calcification medium. hVICs in the calcification group were incubated with OGM (0.25 mM L-ascorbic acid, 10 mM glycerophosphate and 10 nM dexamethasone) purchased from Sigma for 7 days, while cells in the control group were incubated with an equal volume of phosphate buffered saline (PBS).

[0044] (2) High phosphorus-induced cell calcification model. Calcification group hVICs were added with 3 mM inorganic sodium phosphate (Na 3 PO 4 ) for 7 days, and the control group cells were incubated with an equal volume of PBS. According to the added drugs, the cells were divided into a control group and a calcification group, and the two cell calcification models were grouped in the same way.

[0045] 6. Alizarin Red S staining:

[0046] (1) Sample preparation: a. Human aortic valve sample; b. Fix the tissue with 4% formaldehyde solution at room temperature for about 30 minutes; c. Rinse with PBS to remove the fixative;

[0047] (2) Alizarin Red S staining: a. Prepare Alizarin Red S staining solution; b. Prepare 0.1% Alizarin Red S solution in PBS buffer; c. Place the fixed tissue sample in the Alizarin Red S solution and incubate at room temperature for about 30 minutes to 1 hour to allow it to react with calcium ions; d. Wash the sample with PBS buffer to remove unbound stain;

[0048] (3) Observation and image acquisition: Place the stained sample on a microscope slide and observe the stained sample under a microscope. Alizarin Red S complexes with calcium to produce orange-red to red deposits that can be observed with the naked eye or a microscope.

[0049] 7. Western blot:

[0050] Western blot is a commonly used protein analysis technique used to detect the presence and quantification of specific proteins in complex samples. This technique is based on the molecular weight and immunological properties of proteins. It separates proteins by electrophoresis, transfers them to a membrane, and uses specific antibodies for detection. The specific operation method is as follows:

[0051] (1) Sample preparation: a. Collection of cell or valve tissue samples; b. Tissue lysis: Place the collected samples in a suitable lysis buffer, grind the tissue completely to release proteins;

[0052] (2) Protein extraction: a. Centrifugation: Centrifuge the lysed sample to remove solid residues such as cell debris and cell nuclei, and collect the supernatant, which contains the lysed protein; b. Protein concentration determination: Use the BCA protein quantification method to determine the protein concentration, and dilute the sample to make the protein concentration suitable for subsequent experiments according to experimental needs;

[0053] (3) SDS-PAGE gel electrophoresis: a. Sample preparation: Mix the extracted protein sample with SDS-PAGE sample buffer and heat it at an appropriate temperature for denaturation; b. Gel preparation: Prepare polyacrylamide gel of appropriate concentration and select the appropriate gel percentage according to the protein size and resolution requirements; c. Sample loading: Load the denatured protein sample into the gel wells and load the molecular weight marker as a reference; d. Electrophoresis: Place the gel in an electrophoresis tank, add appropriate electrophoresis buffer, and apply an electric field for electrophoresis. Set the appropriate current and time according to the gel size and protein size;

[0054] (4) Transfer: a. Prepare the membrane: Transfer the protein from the gel to a protein blotting membrane (such as polyvinylidene fluoride or nitrocellulose membrane). Prepare a membrane of appropriate size and pretreat the membrane before transfer; b. Transfer device: Stack the gel and membrane in sequence, place them in the protein transfer device, and add appropriate transfer buffer. Make sure there are no bubbles between the gel and the membrane and that they are in full contact; c. Transfer: Apply appropriate current and time to transfer the protein from the gel to the membrane;

[0055] (5) Immunoblotting: a. Blocking: Place the protein blotting membrane in a blocking buffer to block nonspecific binding sites and reduce the nonspecific binding of subsequent antibodies; b. Antibody binding: Add the specific primary antibody (primary antibody) to the antibody binding buffer, incubate the membrane with the primary antibody, and allow the primary antibody to bind to the target protein; c. Washing: Perform multiple washing steps on the membrane to remove unbound primary antibody and other nonspecific binding substances; d. Secondary antibody binding: Add fluorescently labeled or enzyme-labeled secondary antibodies (such as HRP or AP-labeled anti-rabbit / anti-mouse IgG), incubate the membrane with the secondary antibody, and allow the secondary antibody to bind to the primary antibody; e. Washing: Perform multiple washing steps on the membrane again to remove unbound secondary antibodies and other nonspecific binding substances; f. Signal detection: Use chemiluminescence to detect the signal of the protein on the membrane according to the secondary antibody and detection method used; g. Data analysis: Quantitatively analyze the protein blotting images using tools such as image analysis software to obtain the relative expression level of RNF130 protein.

[0056] 8. Reverse transcription polymerase chain reaction (RT-PCR):

[0057] RT-PCR is a commonly used molecular biology technique for detecting and amplifying RNA molecules. It combines the two steps of reverse transcription and polymerase chain reaction, which can transcribe RNA into corresponding DNA and amplify it, thereby realizing the detection and quantitative analysis of RNA. The specific operation method is as follows:

[0058] (1) Sample preparation: Collect the RNA sample to be tested, pay attention to avoid RNA degradation and protect the integrity of RNA;

[0059] (2) Reverse transcription: Reverse transcription is the process of transcribing RNA into a corresponding DNA molecule. First, the RNA template is added to the reaction system together with reverse transcriptase and primers. Primers are DNA fragments that can complement and pair with specific regions of the target RNA sequence. Reverse transcriptase binds the primer to the end of the RNA template and begins to synthesize the DNA chain at the starting point of the primer. Reverse transcriptase synthesizes the corresponding complementary DNA chain according to the nucleotide sequence on the RNA template to form an RNA-DNA hybrid molecule;

[0060] (3) Heat denaturation: In order to remove the RNA template, the reverse transcription reaction mixture is subjected to heat denaturation treatment. This step will dissolve the RNA-DNA hybrid molecules at high temperature, denature the RNA template, but retain the synthesized DNA chain;

[0061] (4) PCR amplification: The heat-denatured reaction mixture is added to the PCR reaction system for DNA amplification. The PCR reaction system contains DNA polymerase, primers, and four nucleotides. The primers will complement each other with specific regions of the synthesized DNA chain, and the DNA polymerase will start synthesizing a new DNA chain at the starting point of the primer. Through a series of cyclic reactions, including high-temperature denaturation, primer annealing, and DNA synthesis, specific DNA fragments can be amplified;

[0062] (5) Result analysis: The amplified DNA product can be used to detect the presence of the target RNA by gel electrophoresis, and to perform quantitative analysis on it.

[0063] 9. Determination of phenotypic conversion of hVICs:

[0064] Western blot was used to detect osteogenic differentiation marker proteins such as Runx2, BMP-2, Osterix and Osteocalcin.

[0065] 10. Von Kossa staining:

[0066] A staining method for detecting calcification deposits in tissues is used to detect calcification in tissues. The basic steps are as follows:

[0067] (1) Preparation of tissue sections: including steps such as aortic valve tissue fixation, embedding, and cutting;

[0068] (2) Degreasing: Degrease the tissue sections in xylene to remove fat and wax from the sections;

[0069] (3) Hydration: The slices were transferred from xylene to alcohol solutions of different concentrations for gradual dehydration;

[0070] (4) Silver salt deposition: Place the slices in a solution containing 5% silver nitrate, and then expose the slices to sunlight or ultraviolet light for 20-30 minutes to allow the silver salt to react with calcium cations in the tissue to form silver salt deposition;

[0071] (5) Removal of unreacted silver salt: Transfer the slices to a solution containing 5% sodium thiosulfate to remove unreacted silver salt;

[0072] (6) Washing: Wash the sections in water to remove the remaining reagents;

[0073] (7) Nuclear staining: For contrast display, nuclear staining can be performed on the sections after washing, for example, using eosin staining;

[0074] (8) Dehydration and fixation: The sections are transferred from water to solutions with increasing alcohol concentrations, and then the sections are soaked and embedded with a transparent agent (such as lignanol ether);

[0075] (9) Microscope observation: Finally, the slices are placed under a microscope for observation. Silver salt deposition will appear black or brown in the calcified deposition area, thus showing the calcification phenomenon.

[0076] 11.ALP activity detection:

[0077] Collect cells or aortic valve samples, follow the instructions of the kit, and measure ALP activity at a wavelength of 520 nm.

[0078] 12. Determination of intracellular calcium content:

[0079] Collect cells or aortic valve samples and press QuantiChrom TM The calcium content was determined at a wavelength of 612 nm according to the Calcium Assay Kit instructions.

[0080] 13. Immunofluorescence (IF):

[0081] The cells were seeded in a 6-well plate and fixed with 4% paraformaldehyde for 15 min; washed with PBS for 5 min, incubated with 0.2% Tritonx-100 for 15 min, washed with PBS for 5 min, incubated with 5% BSA at room temperature for 1 h, added with primary antibody, and incubated at 4°C overnight; washed with PBS three times, 5 min each time, added with secondary antibody, incubated at room temperature for 1 h, and finally stained the cell nucleus with DAPI and sealed, and photographed under a fluorescence microscope.

[0082] Example 1: RNF130 is upregulated in CAVD

[0083] The experimental techniques involved in Example 1 include: human aorta sample collection and processing, proteomics, animal model construction, cell model construction, Alizarin Red S staining, Von Kossa staining, immunofluorescence, Western blot, reverse transcription polymerase chain reaction (RT-PCR) and data statistical analysis.

[0084] 1. RNF130 expression is elevated in CAVD patients

[0085] The present invention has carried out testing on CAVD patients, and the results are as follows Figure 1 As shown in A, proteomic analysis of human aortic samples was used to screen targets that affect calcific aortic valve disease, and it was initially found that RNF130 protein expression was increased in the aorta of CAVD patients; Figure 1 B The levels of RNF130 and classic bone differentiation marker proteins (Runx-2, BMP-2, Osterix and Osteocalcin) in human aortic samples were further verified by western blot, showing that the aortic valve of CAVD patients was significantly calcified and the level of RNF130 was significantly increased; Figure 1 C shows that the mRNA level of RNF130 is also significantly increased, proving that the protein synthesis and expression of RNF130 are abnormally increased in CAVD patients; Figure 1 D shows the results of Alizarin Red S, Von Kossa and immunofluorescence staining of human aorta samples, showing obvious valvular calcification and increased expression of RNF130 in CAVD patients.

[0086] 2. RNF130 expression is elevated in CAVD mice and cell models

[0087] In order to further study the changes of RNF130 in the process of CAVD, the present invention constructed ApoE - / - , LDLR - / - Mouse CAVD model and in vitro CAVD cell model. Figure 2 As shown in A and 2B, RT-PCR and western blot were used to detect the mRNA and protein expression of RNF130 in hVICs incubated with OGM calcification medium for 0, 3, 5, 7, and 10 days, respectively. It can be seen that the synthesis and expression of RNF130 protein in calcified hVICs were significantly increased; Figure 2 C. The expression of RNF130 was detected by immunofluorescence after hVICs were treated with OGM calcification medium for 7 days. It can be seen that the fluorescence content of RNF130 increased significantly, proving that the expression of RNF130 was significantly upregulated; Figure 2D, 2E are PCR detection of RNF130 mRNA levels in calcified hVICs, and it can be seen that the mRNA levels of RNF130 in both models were significantly increased; Figure 2 F ApoE was detected by western blot - / - Mice and LDLR - / - The protein expression of RNF130 in the aortic valve of mice showed that RNF130 in the aortic valve of two CAVD mice was significantly increased. The above experimental results show that the expression levels of RNF130 gene and protein in mice and cells of CAVD model are increased.

[0088] Example 2: Construction of shRNA targeting RNF130 and its application in the treatment of CAVD

[0089] The experimental techniques involved in Example 2 include: construction of RNF130 shRNA and AAV viral vector, construction of cell model, RT-PCR, western blot, ALP activity detection, determination of intracellular calcium content, Alizarin Red S staining and data analysis.

[0090] In order to verify the conclusion obtained in Example 1, the present invention constructs shRNA to knock down the expression of RNF130 to detect the therapeutic effect of targeted inhibition of RNF130 on CAVD. Figure 3 A, 3B RT-PCR and western blot showed that targeted inhibition of RNF130 protein synthesis could restore the abnormally elevated mRNA levels of bone differentiation marker proteins (Runx-2, BMP-2, Osterix, and Osteocalcin) in calcified hVICs to normal; Figure 3 C is the result of ALP activity test kit to detect ALP activity in hVICs, which shows that targeted inhibition of RNF130 protein synthesis can reverse abnormally activated ALP; Figure 3 D is the result of the calcium content test kit for hVICs. It can be seen that the targeted inhibition of RNF130 protein synthesis can reverse the abnormally increased calcium content. Figure 3 E is Figure 3 D is the Alizarin Red S staining result of hVICs cells, showing that targeted inhibition of RNF130 protein synthesis can reverse the abnormally increased calcium deposition.

[0091] The above experimental results show that RNF130 protein plays a key role in regulating abnormal bone differentiation and calcium metabolism in calcified hVICs. Targeted inhibition of RNF130 protein synthesis can reverse abnormal bone differentiation and calcium deposition, thereby preventing and treating CAVD.

[0092] Although the present invention has been disclosed as above in the form of a preferred embodiment, it is not intended to limit the present invention. Anyone familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be based on the definition of the claims.

Claims

1. Use of an RNF130 inhibitor in the preparation of a drug for preventing or treating calcific aortic valve disease, characterized in that: The RNF130 inhibitor is an adeno-associated virus shRNA interference vector of the RNF130 gene; wherein the nucleotide sequence of the shRNA is shown in SEQ ID NO.1-2.

2. The use according to claim 1, characterized in that: The drug can reduce the expression of RNF130 protein or gene.

3. The use according to claim 1, characterized in that: The calcific aortic valve disease includes calcific aortic valve stenosis or calcific aortic valve insufficiency or calcific aortic valve lesions.

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