Motion intervention heart failure repair method based on Lgmn signal regulation and control

Abstract

The invention discloses a motion intervention heart failure repair method based on Lgmn signal regulation and control, and relates to the technical field of heart failure treatment. According to the method, through aerobic exercise training (AET) with specific parameters, expression of Legumain (Lgmn) genes in macrophages is regulated in a targeted mode, the interburial effect of the macrophages is enhanced, then cell communication between the macrophages and cardiac muscle cells is enhanced, removal of the vertical death cardiac muscle cells is promoted, myocardial fibrosis and inflammatory response are relieved, and the effect of treating myocardial fibrosis is achieved. Finally, ventricular remodeling and heart functions of chronic heart failure (CHF) mice are improved. Experimental verification shows that the exercise intervention method can significantly improve the left ventricular ejection fraction (LVEF) and shortening fraction (FS) of CHF model animals, reduce the left ventricular end-of-systole inner diameter (LVESD), reduce myocardial cell apoptosis and necrosis, and promote inflammation regression. Drug intervention is not needed, safety is high, operation is easy and convenient, and a new effective way is provided for clinical rehabilitation treatment of heart failure.

Description

Technical Field

[0001] This invention belongs to the field of heart failure treatment technology, and in particular relates to a method for repairing heart failure through exercise intervention based on Lgmn signal regulation. Background Technology

[0002] Chronic heart failure (CHF) is a severe manifestation or advanced stage of various heart diseases, characterized by ventricular remodeling, myocardial fibrosis, cardiomyocyte apoptosis and necrosis, and persistent inflammatory responses. Current clinical treatment primarily relies on pharmacological interventions; however, long-term use easily leads to drug resistance and side effects, and it is difficult to reverse existing myocardial structural damage. Therefore, developing safe and effective non-pharmacological interventions is of significant clinical value.

[0003] Aerobic exercise training (AET), as a non-pharmacological intervention, has been shown to improve heart failure to some extent, but its specific molecular mechanisms are not yet fully understood. Legumain (Lgmn) is a cysteine ​​protease that plays a key role in the phagocytosomal degradation process of macrophages and is closely related to inflammation regulation. Existing studies have shown that Lgmn gene expression is reduced in heart failure, and aerobic exercise may exert a cardioprotective effect by regulating the Lgmn signaling pathway, but the relevant mechanisms and standardized exercise protocols remain unclear.

[0004] Therefore, a method for repairing heart failure through exercise intervention based on Lgmn signal regulation is proposed. Summary of the Invention

[0005] The purpose of this invention is to provide a method for repairing heart failure through exercise intervention based on Lgmn signal regulation. By standardizing aerobic exercise training, the Lgmn signaling pathway is targeted and regulated to achieve efficient repair of heart failure and overcome the limitations of existing treatment methods.

[0006] To solve the above-mentioned technical problems, the present invention is achieved through the following technical solution: This invention is a method for repairing heart failure through exercise intervention based on Lgmn signal regulation, comprising the following steps: performing phased aerobic exercise training on heart failure model animals, thereby promoting the clearance of dying cardiomyocytes and improving ventricular remodeling and cardiac function by regulating the expression of the Lgmn gene in macrophages.

[0007] In one embodiment, the parameters of the aerobic exercise training are: first, perform 5 minutes of adaptation training at a speed of 7 m / min, then perform 30 minutes of formal training at a speed of 12 m / min, train 5 days a week, and continue training for 4 weeks.

[0008] In one embodiment, the heart failure model animal was constructed by ligating the left anterior descending (LAD) coronary artery using 6-week-old male C57BL / 6J mice.

[0009] In one embodiment, the exercise intervention enhances the macrophage burial function by increasing the expression level of the Lgmn gene in macrophages, thereby enhancing intercellular communication between macrophages and cardiomyocytes and improving the macrophages' ability to phagocytose and clear dying cardiomyocytes.

[0010] In one embodiment, the exercise intervention can upregulate the expression of anti-apoptotic genes Bcl2, Mcl1, and Sirt1, and downregulate the expression of apoptotic genes Fas, Casp4, Casp8, and necrosis genes Mb, Ldha, and Ckm.

[0011] In one embodiment, the exercise intervention can promote the expression of anti-inflammatory mediators IL-10 and TGF-β1, inhibit the expression of pro-inflammatory mediators IL-1β, TNF-α, and IL-6, and accelerate the resolution of inflammation.

[0012] In one embodiment, the method achieves heart failure repair through the following mechanism: aerobic exercise training induces high expression of the Lgmn gene in macrophages, enhances the cytotoxicity of macrophages, reduces the accumulation of dying cardiomyocytes, alleviates myocardial fibrosis and inflammatory response, and improves ventricular remodeling and cardiac systolic and diastolic function.

[0013] The present invention has the following beneficial effects: 1. This invention improves cardiac function in heart failure through standardized aerobic exercise training, assists conventional drug intervention, reduces drug side effects and drug resistance, and is safer.

[0014] 2. This method has a clear target and plays a role in several key aspects by regulating the Lgmn signaling pathway, such as enhancing macrophage burial, clearing dying cardiomyocytes, inhibiting inflammatory response, and reducing myocardial fibrosis, resulting in a significant cardioprotective effect.

[0015] 3. The exercise program is simple to operate, low in cost, highly repeatable, and easy to translate into clinical applications, providing a new and effective approach for the rehabilitation treatment of patients with heart failure.

[0016] 4. Experimental verification shows that this method can significantly improve LVEF and FS in heart failure model animals, reduce LVESD, reduce heart weight and myocardial fibrosis, and improve immune regulation function, thus having a comprehensive cardioprotective effect.

[0017] Of course, any product implementing this invention does not necessarily need to achieve all of the advantages described above at the same time. Attached Figure Description

[0018] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0019] Figure 1 Figure showing the correlation between AET and ventricular remodeling and cardiac function in CHF mice; Figure 2 Figure showing the results of AET promoting the clearance of dying cardiomyocytes in CHF mice; Figure 3 Figure showing the results of AET enhancing cell communication between cardiomyocytes and macrophages in CHF mice; Figure 4 Figure showing the key role of Lgmn in enhancing macrophage burial and alleviating CHF inflammation; Figure 5 The image shows the verification results of Lgmn overexpression in CHF mouse macrophages; Figure 6 Figure showing the results of Lgmn overexpression improving cardiac function in CHF mice; Figure 7 Figure showing the results of Lgmn overexpression enhancing the clearance of dying cardiomyocytes and promoting inflammation resolution; Figure 8 Figure showing the results of eliminating the repair effect of AET by knocking out the Lgmn gene in macrophages on CHF mice. Detailed Implementation

[0020] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0021] In the description of this invention, it should be understood that the terms "upper," "middle," "outer," "inner," etc., which indicate orientation or positional relationship, are only for the convenience of describing this invention and simplifying the description, and do not indicate or imply that the components or elements referred to must have a specific orientation, or be constructed and operated in a specific orientation, and therefore should not be construed as limiting this invention.

[0022] In the description of this invention, it should be noted that, unless otherwise explicitly specified and limited, the terms "installed," "equipped with," "connected," etc., should be interpreted broadly. For example, "connection" can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection or an electrical connection; it can be a direct connection or an indirect connection through an intermediate medium; it can be a connection within two components. Those skilled in the art can understand the specific meaning of the above terms in this invention based on the specific circumstances. Example 1

[0023] Please see Figures 1-8 As shown, this invention is a method for repairing heart failure through exercise intervention based on Lgmn signal modulation, comprising the following steps: Heart failure model establishment: Six-week-old male C57BL / 6J mice were anesthetized by intraperitoneal injection of 1.25% tribromoethanol, then intubated and connected to a ventilator for assisted breathing. An incision was made between the third and fourth intercostal spaces, and the left anterior descending (LAD) coronary artery was permanently ligated with 7-0 nylon suture to establish a heart failure model. The sham-operated group underwent only the surgical procedure without coronary artery ligation.

[0024] Aerobic exercise training protocol: After successful model establishment, mice with heart failure underwent phased aerobic exercise training. The training was divided into an adaptation phase and a formal phase. The adaptation phase consisted of 5 minutes of exercise on a treadmill at a speed of 7 m / min on flat ground, and the formal phase consisted of 30 minutes of exercise at a speed of 12 m / min. Training was conducted 5 days a week for 4 weeks. Control group mice were fed routinely and did not receive any specific exercise training.

[0025] Repair mechanism: The above-mentioned exercise training can significantly upregulate the expression level of Lgmn gene in macrophages of mice with heart failure, enhance the cytotoxicity of macrophages, thereby enhancing intercellular communication between macrophages and cardiomyocytes and improving the phagocytic clearance efficiency of macrophages on dying cardiomyocytes; at the same time, it can upregulate the expression of anti-apoptotic genes and downregulate the expression of apoptosis and necrosis-related genes, reducing cardiomyocyte apoptosis and necrosis; in addition, it can promote the secretion of anti-inflammatory mediators, inhibit the release of pro-inflammatory mediators, accelerate the resolution of inflammation, thereby reducing myocardial fibrosis and ventricular remodeling, and improving cardiac systolic and diastolic function.

[0026] Example 2 (Construction of a Heart Failure Model and Exercise Intervention) 1. Experimental animals: Six-week-old male C57BL / 6J mice (Jiangsu Jicui Yaokang Biotechnology Co., Ltd.) were selected and kept in an environment with a temperature of 22-25℃, humidity of 50-60%, a circadian rhythm of 12h / 12h, and free access to food and water.

[0027] 2. Model Establishment: Mice were anesthetized with 1.25% tribromoethanol via intraperitoneal injection, then intubated and connected to an HX-101E ventilator for assisted breathing. The skin and chest wall muscles were incised in the third and fourth intercostal spaces to expose the heart. A permanent ligation was performed using 7-0 nylon suture 2 mm below the origin of the left anterior descending coronary artery. The chest wall and skin were then sutured layer by layer. Mice that died within 24 hours post-surgery were discarded, and the remaining mice were used as the heart failure model group (WT-HF group). The sham-operated group (WT-Ctrl group) underwent the same procedure, but without coronary artery ligation.

[0028] 3. Grouping and Intervention: Mice in the WT-HF group were randomly divided into an exercise intervention group (WT-HF-EX group) and a non-exercise group (WT-HF group), with 10 mice in each group. The WT-HF-EX group underwent 4 weeks of aerobic exercise training according to the above exercise protocol, while the WT-HF group and the WT-Ctrl group were fed routinely without exercise training.

[0029] Example 3 (Effect Verification) 1. Cardiac Function Testing: Transthoracic echocardiography was performed on mice in each group before and after exercise intervention using a Vevo2100 ultrasound system. Mice were lightly anesthetized with 0.5% isoflurane, and parasternal long-axis and short-axis images were acquired. LVEF, FS, and LVESD were calculated. Results showed that compared with the WT-HF group, the WT-HF-EX group mice had significantly increased LVEF and FS, and significantly decreased LVESD, indicating a significant improvement in cardiac function.

[0030] 2. Myocardial fibrosis detection: Masson's trichrome staining method was used to stain the heart tissue of mice in each group to observe the myocardial fibrosis. The results showed that the proportion of myocardial fibrosis area in the WT-HF-EX group was significantly lower than that in the WT-HF group, indicating that exercise intervention can reduce myocardial fibrosis.

[0031] 3. Detection of dying cardiomyocytes: TUNEL staining and Western blot analysis were used to detect cardiomyocyte apoptosis and necrosis. The results showed that the proportion of TUNEL-positive cardiomyocytes in WT-HF-EX mice was significantly reduced, and the expression levels of apoptosis genes Fas, Casp4, Casp8 and necrosis genes Mb, Ldha, Ckm were significantly downregulated, while the expression levels of anti-apoptotic genes Bcl2, Mcl1 and Sirt1 were significantly upregulated.

[0032] 4. Lgmn gene expression detection: The expression of the Lgmn gene in macrophages was detected by snRNA-seq, Western blotting, and immunofluorescence staining. The results showed that compared with the WT-HF group, the expression level of Lgmn in the macrophages of mice in the WT-HF-EX group was significantly increased, and Lgmn was mainly located in CD68-positive macrophages.

[0033] 5. Detection of inflammatory factors: The expression of inflammatory factors in cardiac tissue and blood was detected by RT-qPCR and ELISA. The results showed that the expression of anti-inflammatory mediators IL-10 and TGF-β1 was significantly increased in the WT-HF-EX group mice, while the expression of pro-inflammatory mediators IL-1β, TNF-α, and IL-6 was significantly decreased.

[0034] Example 4 (Verification of the function of the Lgmn gene) 1. Lgmn overexpression experiment: An AAV2 / 6-CD68-Lgmn adeno-associated virus vector was constructed and injected orally into the myocardium of mice with heart failure to achieve macrophage-specific Lgmn overexpression. The results showed that the LVEF and FS of the Lgmn-overexpressing group were significantly higher than those of the control group, the degree of myocardial fibrosis was reduced, the clearance efficiency of dying cardiomyocytes was improved, and the inflammatory response was alleviated.

[0035] 2. Lgmn Gene Knockout Experiment: Lgmnflox / flox mice were crossed with Lyz2-Cre transgenic mice to construct a macrophage-specific Lgmn gene knockout mouse model of heart failure. Aerobic exercise training of this model revealed that Lgmn gene knockout did not improve cardiac function, nor did it alleviate myocardial fibrosis or inflammatory responses, indicating that the Lgmn gene is a key molecule in the repair function of exercise in heart failure.

[0036] The above embodiments demonstrate that the aerobic exercise training method of the present invention can effectively improve cardiac function in heart failure by regulating the Lgmn signaling pathway in macrophages, and has important clinical application value.

[0037] In the description of this specification, references to terms such as "an embodiment," "example," and "specific example" indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the invention. In this specification, illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0038] The preferred embodiments of the present invention disclosed above are merely illustrative of the invention. These preferred embodiments do not exhaustively describe all details, nor do they limit the invention to the specific implementations described. Clearly, many modifications and variations can be made based on the content of this specification. This specification selects and specifically describes these embodiments to better explain the principles and practical applications of the invention, thereby enabling those skilled in the art to better understand and utilize the invention. The invention is limited only by the claims and their full scope and equivalents.

Claims

1. A method for repairing heart failure through exercise intervention based on Lgmn signal modulation, characterized in that, Includes the following steps: Phased aerobic exercise training in heart failure model animals can enhance macrophage burial by regulating the expression of the Lgmn gene in macrophages, promote the clearance of dying cardiomyocytes, and improve ventricular remodeling and cardiac function.

2. The method for repairing heart failure through exercise intervention based on Lgmn signal regulation according to claim 1, characterized in that, The parameters for the aerobic exercise training are as follows: first, perform 5 minutes of adaptation training at a speed of 7 m / min, then perform 30 minutes of formal training at a speed of 12 m / min, train 5 days a week, and continue training for 4 weeks.

3. The method for repairing heart failure through exercise intervention based on Lgmn signal regulation according to claim 1, characterized in that, The heart failure model animals were constructed by ligating the left anterior descending (LAD) coronary artery, using 6-week-old male C57BL / 6J mice.

4. The method for repairing heart failure through exercise intervention based on Lgmn signal regulation according to claim 1, characterized in that, The exercise intervention enhances the expression level of the Lgmn gene in macrophages, thereby increasing the macrophage burial function, enhancing intercellular communication between macrophages and cardiomyocytes, and improving the macrophages' ability to phagocytose and clear dying cardiomyocytes.

5. The method for repairing heart failure through exercise intervention based on Lgmn signal regulation according to claim 1, characterized in that, The exercise intervention can upregulate the expression of anti-apoptotic genes Bcl2, Mcl1 and Sirt1, and downregulate the expression of apoptotic genes Fas, Casp4, Casp8 and necrosis genes Mb, Ldha and Ckm.

6. The method for repairing heart failure through exercise intervention based on Lgmn signal regulation according to claim 1, characterized in that, The exercise intervention can promote the expression of anti-inflammatory mediators IL-10 and TGF-β1, inhibit the expression of pro-inflammatory mediators IL-1β, TNF-α, and IL-6, and accelerate the resolution of inflammation.

7. The method for repairing heart failure through exercise intervention based on Lgmn signal regulation according to claim 1, characterized in that, The method achieves heart failure repair through the following mechanism: aerobic exercise training induces high expression of the Lgmn gene in macrophages, enhances the cytotoxicity of macrophages, reduces the accumulation of dying cardiomyocytes, alleviates myocardial fibrosis and inflammatory response, and improves ventricular remodeling and cardiac systolic and diastolic function.

Images