Use of recombinant serpin b1 protein in the preparation of a medicament for the prevention and treatment of myocardial hypertrophy and heart failure

Abstract

The application provides application of recombinant serine protease inhibitor B1 protein in preparation of drugs for preventing and treating myocardial hypertrophy. The myocardial hypertrophy in the application is pathological myocardial hypertrophy caused by hypertension, aortic valve stenosis, cardiomyopathy and the like. The application adopts subcutaneous injection or intravenous infusion of serine protease inhibitor B1 0.3-0.6 mg / kg of body weight, 1-5 times / week. Experimental results show that the recombinant serine protease inhibitor B1 protein can significantly reduce pathological myocardial hypertrophy induced by aortic constriction, effectively relieve heart dysfunction and cardiac remodeling, and improve the treatment of cardiovascular diseases characterized by or leading to pathological myocardial hypertrophy.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to the application of recombinant serine protease inhibitor B1 protein in the preparation of drugs for the prevention and treatment of myocardial hypertrophy and heart failure. Background Technology

[0002] Myocardial hypertrophy is a common and important pathological basis for the development of heart failure in various cardiovascular diseases such as hypertension, myocardial infarction, valvular heart disease, and cardiomyopathy. It also increases the risk of cardiovascular events such as arrhythmias and sudden cardiac death. Therefore, preventing and delaying the development of myocardial hypertrophy has become a fundamental strategy for the prevention and treatment of heart failure. Although numerous scholars both domestically and internationally have long been dedicated to studying the pathogenesis of myocardial hypertrophy and have discovered a large number of therapeutic targets, effective prevention and treatment strategies and drugs are still lacking.

[0003] Serine protease inhibitor B1 (SerpinB1) is a highly differentiated superfamily of protease inhibitors with conserved amino acid sequences and spatial structures. It plays a crucial role in inhibiting serine protease cascade activation pathways and maintaining homeostasis in organisms. In addition to its regulatory role in inflammatory cells, SerpinB1 can circulate in the bloodstream via endocrine pathways, affecting the regeneration of pancreatic β-cells, suggesting that SerpinB1 may have potential regulatory effects on various tissues and organs.

[0004] Currently, there are no reports on the effects of serine protease inhibitor B1 on pathological myocardial hypertrophy. Summary of the Invention

[0005] In view of this, the purpose of this invention is to provide the application of recombinant serine protease inhibitor B1 protein in the preparation of drugs for the prevention and treatment of myocardial hypertrophy and heart failure. This invention has verified that recombinant serine protease inhibitor B1 protein has a significant effect in inhibiting pathological myocardial hypertrophy, alleviating cardiac function deterioration, and promoting cardiac remodeling; therefore, it can provide an effective drug for the treatment of cardiovascular diseases characterized by or causing pathological myocardial hypertrophy.

[0006] This invention also provides the use of recombinant serine protease inhibitor B1 protein in the preparation of drugs for the prevention and treatment of hypertrophic cardiomyopathy and chronic heart failure.

[0007] This invention also provides the application of recombinant serine protease inhibitor B1 protein in the preparation of drugs for preventing and treating ventricular remodeling.

[0008] Furthermore, the myocardial hypertrophy mentioned refers to pathological myocardial hypertrophy caused by conditions such as hypertension, aortic stenosis, and cardiomyopathy.

[0009] Furthermore, the drug contains recombinant serine protease inhibitor B1 protein, or a splice variant or variant thereof with its activity.

[0010] Furthermore, the amino acid sequence of the recombinant serine protease inhibitor B1 protein is shown in SEQ NO.1.

[0011] Furthermore, the drug is administered subcutaneously or intravenously at a dose of 0.3-0.6 mg / kg body weight, 1-5 times per week.

[0012] SEQ NO.1: MEQLSSANTLFALELFQTLNESSPTGNIFFSPFSISSALAMVILGAKGSTAAQLSKTFHFDSVEDIHSRFQSLNAEVSKRGASHTLKLANRLYGEKTYNFLPEYLASTQKMYGADLAPVDFLHASEDARKEINQWVKGQTEGKIPELLSVGVVDSMTKLVLVNAIYFKGMWEEKFMTEDTTDAPFRLSKK DTKTVKMMYQKKKFPFGYISDLKCKVLEMPYQGGELSMVILLPKDIEDESTGLKKIEKQITLEKLLEWTKRENLEFIDVHVKLPRFKIEESYTLNSNLGRLGVQDLFSSSKAADLSGMSGSRDLFISKIVHKSFVEVNEEGTEAAAATGGIATFCMLLPEEEFTVDHPFIFFIRHNPTSNVLFLGRVCSP.

[0013] Compared with the prior art, the beneficial effects of the present invention are as follows: 1. This invention provides the application of recombinant serine protease inhibitor B1 protein in the preparation of drugs for the prevention and treatment of myocardial hypertrophy and heart failure.

[0014] 2. This invention also provides the application of recombinant serine protease inhibitor B1 protein in the preparation of drugs for the prevention and treatment of hypertrophic cardiomyopathy and chronic heart failure.

[0015] 3. This invention also provides the application of recombinant serine protease inhibitor B1 protein in the preparation of drugs for preventing and treating ventricular remodeling.

[0016] 4. The application of the recombinant serine protease inhibitor B1 protein provided by this invention in the preparation of drugs can effectively inhibit pathological myocardial hypertrophy, alleviate the deterioration of cardiac function and cardiac remodeling, and provide an effective drug for the treatment of cardiovascular diseases characterized by or caused by pathological myocardial hypertrophy. Attached Figure Description

[0017] To more clearly illustrate the implementation techniques of this invention, the accompanying drawings used in the description of the embodiments or prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of this invention; those skilled in the art can derive other drawings from these drawings without any creative effort.

[0018] Figure 1 The effect of recombinant serine protease inhibitor B1 (rSerpinB1) protein on pathological myocardial hypertrophy induced by aortic constriction; Wherein: A: is a schematic diagram of the experimental procedure for verifying the present invention; B1-B5: Detection of left ventricular systolic function in mice; (Sham group was the sham-operated group, TAC group was the aortic coarctation-operated group;) B1: An echocardiogram of left ventricular systolic function in mice; B2: Statistical results of left ventricular end-diastolic diameter (LVEDd); B3: Statistical results of left ventricular end-systolic diameter (LVESd); B4: Statistics on left ventricular ejection fraction (LVEF); B5: Statistical results of left ventricular fractional shortening (LVFS); C1-C2: Comparison of mouse heart size (TAC is the aortic coarctation surgery group, Sham is the sham surgery group); C1: Comparison photos of heart size; C2: Statistical results of heart mass to body weight ratio (HW / BW); D1-D2: Cross-sectional area of ​​mouse myocardium (TAC was the aortic coarctation surgery group, and Sham was the sham surgery group;) D1: Comparison of WGA-stained images of cardiac sections; D2: Statistical results of myocardial cross-sectional area; E1-E2: represents the area of ​​cardiac fibrosis in mice (TAC is the aortic coarctation surgery group, Sham is the sham surgery group); E1: Comparison of images of heart sections stained with masson stain; E2: Statistics on the area of ​​cardiac fibrosis. Detailed Implementation

[0019] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the specific implementation of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. All other implementation methods obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the protection scope of the present invention.

[0020] Example 1 Recombinant serine protease inhibitor B1 protein alleviates aortic coarctation-induced cardiac dysfunction. I. Modeling of pathological myocardial hypertrophy and treatment with recombinant serine protease inhibitor B1 protein.

[0021] A myocardial hypertrophy model was established using aortic coarctation (TAC) surgery. Mice were anesthetized with sodium pentobarbital (40 mg / kg) via intraperitoneal injection and fixed on the operating table. They were then connected to a ventilator via endotracheal intubation to maintain respiration. The surgical area was prepared and disinfected, and the skin was incised layer by layer. The pectoralis major and pectoralis minor muscles were bluntly dissected. An opening was made between the 2nd and 3rd ribs, the thymus was dissected, and the aortic arch was exposed. A suture was threaded, and the aorta was ligated to a needle with an outer diameter of 0.8 mm. The needle was removed, all tissues were returned to their original positions, the chest was closed, the skin was sutured, and the mice were returned to their cages for continued rearing. The sham-operated group (Sham) underwent only chest opening and closing without aortic coarctation. Post-operatively, the experimental group received intraperitoneal injections of recombinant serine protease inhibitor B1 protein (rSerpinB1) (0.5 mg / kg, twice a week), while the control group received injections of PBS (Vehicle).

[0022] II. Cardiac function testing.

[0023] On day 28 post-TAC surgery, mice were randomly numbered and, following a double-blind approach, their cardiac structure and function were assessed using a Vevo 2100 ultrasound imaging platform. Specifically, after isoflurane-induced anesthesia, mice were fixed on foam boards, and ultrasound probes were used to capture long-axis images of the heart parasternally in M-mode. Left ventricular ejection fraction was calculated using the Teicholz formula, with each data point repeated three times, and the average value used as the statistical result.

[0024] Results: Treatment with recombinant serine protease inhibitor B1 protein had no significant effect on cardiac function in Sham mice, but it significantly alleviated cardiac dysfunction in TAC mice, manifested as decreased left ventricular end-diastolic diameter (LVEDd) and end-systolic diameter (LVESd), while significantly increased left ventricular ejection fraction (LVEF) and fractional shortening (LVFS). Figure 1 B1-5).

[0025] Example 2 I. Detection of the effect of recombinant serine protease inhibitor B1 protein on aortic coarctation-induced myocardial hypertrophy and remodeled cardiac morphology Mice were sacrificed on day 35 after TAC surgery, and heart tissue was collected for comparison of overall heart size. Heart mass and body weight were measured, and the heart mass / body weight ratio (HW / BW) was calculated.

[0026] II. Cardiac WGA staining Mice were sacrificed on day 35 after TAC surgery. Heart tissue was fixed in 4% paraformaldehyde, embedded in paraffin, and prepared into paraffin sections. After dewaxing, the tissue was stained with WGA staining solution labeled with Alexa Fluor 488, followed by washing with PBS three times for 5 min each time. Finally, DAPI staining solution was added, and the tissue was incubated at 25°C for 20 min, followed by washing with PBS three times for 5 min each time. Fluorescence scanning was performed using an Olympus laser confocal microscope, and the cross-sectional area of ​​the myocardium was calculated.

[0027] III. Detection of cardiac fibrosis by Marson staining Mice were sacrificed on day 35 after TAC surgery. Heart tissue was fixed in 4% paraformaldehyde, embedded in paraffin, and sectioned. After dewaxing, nuclei were stained with Weigert hematoxylin and eosin for 5 min, thoroughly washed with water, stained with Masson's Ponceau S acid fuchsin for 5 min, rinsed with 2% glacial acetic acid solution for 30 s, differentiated with 1% phosphomolybdic acid solution for 3 min, stained with aniline blue or light green solution for 5 min, rinsed with 0.2% glacial acetic acid solution for 30 s, cleared with 95% ethanol, anhydrous ethanol, xylene, and mounted with neutral resin. Images were taken using a fully automated digital sectioning system, and the left ventricular fibrotic area was calculated.

[0028] Results: Treatment with recombinant serine protease inhibitor B1 protein significantly improved the inhibition of TAC-induced myocardial hypertrophy, manifested as a reduction in heart size. Figure 1 C1-2), myocardial cross-sectional area ( Figure 1 D1-2) and fibrosis area ( Figure 1 E1-2).

[0029] It should be noted that the specific embodiments described above enable those skilled in the art to more fully understand the present invention, but do not limit the present invention in any way. Therefore, although the present invention has been described in detail with reference to the accompanying drawings and embodiments, those skilled in the art should understand that modifications or equivalent substitutions can still be made to the present invention without departing from the spirit and scope of the present invention, and all such modifications or substitutions should be covered within the scope of the claims of the present invention.

Claims

1. Application of recombinant serine protease inhibitor B1 protein in the preparation of drugs for treating aortic coarctation-induced myocardial hypertrophy.

2. Application of recombinant serine protease inhibitor B1 protein in the preparation of drugs to prevent aortic constriction-induced ventricular remodeling.

3. The application according to any one of claims 1-2, characterized in that, The amino acid sequence of the recombinant serine protease inhibitor B1 protein is shown in SEQ NO.

1.

4. The application according to any one of claims 1-2, characterized in that, The drug is administered subcutaneously or intravenously at a dose of 0.3-0.6 mg / kg, 1-5 times per week.

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