Magnesium isoglycyrrhizinate regulates oxidative stress and apoptosis via PI3K/AKT1 pathway to improve myocardial remodeling in mice

doi:10.21203/rs.3.rs-3596412/v1

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Magnesium isoglycyrrhizinate regulates oxidative stress and apoptosis via PI3K/AKT1 pathway to improve myocardial remodeling in mice

https://doi.org/10.21203/rs.3.rs-3596412/v1

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The role of magnesium isoglycyrrhizinate (MgIG) in myocardial remodeling is being investigated. We evaluated the result of MgIG on isoproterenol (ISO) -enticed myocardial remodeling in mice by activating the PI3K/AKT1 pathway. The heart function of mice was tested by echocardiography, and it was found that MgIG could improve the left ventricular function. Pathological staining analysis showed that MgIG could decrease the degree of myocardial injury caused by ISO. The serum data detected by ELISA showed that MgIG could reduce the content of CK-MB, MDA and LDH, and increase the activity of GSH-Px. Western blotting showed that the protein expressions of Collagen Ⅰ, BNP, Bax, Cleaved caspase-3, p-PI3K and p-AKT1 were decreased, while the protein expressions of Bcl-2, COX2 and SOD1 were increased. Meanwhile activation of the PI3K activator (740Y-P) reverses the cardioprotective effect of MgIG. These findings suggest that the myocardial remodeling induced by ISO could be improved by MgIG, and its mechanism may be associated with inhibite PI3K/AKT1 pathway to regulate apoptosis and oxidative stress.

Magnesium isoglycyrrhizinate

Isoproterenol

Myocardial remodeling

PI3K/AKT1

Apoptosis

Oxidative stress

With the continuous enhancement of people's life level and the gradual change of work and rest rules, cardiovascular diseases are becoming younger and more frequent, and the mortality rate is likewise increasing (Foron et al., 2018). Therefore, reversed myocardial remodeling and improved cardiac function are the focus of clinical research. Cardiac hypertrophy is a familiar pathological change in the development of various cardiovascular problems, and its mechanism is complex and changeable, including apoptosis (Sha et al., 2023), oxidative stress ( Shen et al., 2022), inflammatory response and fibrosis (Zhao et al., 2023). When myocardial injury occurs, the increase of oxygen radical in vivo leads to the breakdown of redox balance, which causes oxidative stress damage to cell macromolecules in vivo. The production of oxygen free radicals can also damage mitochondrial membrane, increase membrane permeability, and cause mitochondrial swelling and breakage, resulting in cell death and fibrosis formation (Vuda et al., 2016). In addition, excessive generation of reactive oxygen species (ROS) in cardiac tissue can accelerate lipid peroxidation and apoptosis (Park et al., 2014). The defense mechanism of the body can improve the harm caused by ROS and reduce the damage to cardiomyocytes. Therefore, the restoration of endogenous defense mechanisms is an important target for the treatment of cardiomyopathy.

Phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway adjusts a number of essential cellular processes, including cell growth, transcription, translation, cell proliferation and glycogen metabolism (Mayer et al., 2015; Zhao et al., 2023). Among them, it can play a powerful role in adjusting apoptosis and oxidative stress, which is an important mechanism of oxidative stress injury in various organs. SOD1 and Bcl-2 gene are the main target proteins of PI3K/AKT1 in adjusting apoptosis and oxidative stress. PI3K/AKT signaling axis plays an significant role in adjusting oxidative stress, mediating apoptosis, reducing mitochondrial damage, regulating Ca^{2 +} influx and regulating cell growth, and is an indispensable signaling pathway for anti-apoptotic response (Cantley, 2002).

Glycyrrhiza is a leguminous plant, which has the functions of invigorating spleen and qi, relieving cough and moistening lung, and resisting inflammation. Magnesium isoglycyrrhizinate is a compound synthesized by structural modification of glycyrrhetinic acid extracted from licorice (Ma et al., 2017). Magnesium isoglycyrrhizinate is considered to be a safe and well-tolerated drug for use in clinical studies because of its acceptable pharmacokinetic profile and the absence of serious adverse effects after intravenous administration in healthy young volunteers (Rinaldi et al., 2010). MgIG plays an anti-inflammatory and anti-apoptotic role in acute kidney injury to alleviate nephrotoxicity (Wei et al., 2022). MgIG can suppress hepatotoxicity by suppressing oxidative stress (Liu et al., 2021). However, the relationship between MgIG and cardiac remodeling necessitates further investigation. For this study, we structured the myocardial injury model induced by ISO to discuss the defensive effect and mechanism of MgIG on myocardial damage.

2.1 Reagents and antibodies

Magnesium isoglycyrrhizinate (MgIG, H20051942, Chia Tai Tianqing Pharmaceutical Co. Ltd.), Isoproterenol (Cat#1129810, Aladdin), PI3K activator (740Y-P, Cat#M00988-FQC, Beijing BioLab Technology Co. Ltd.). Lactate dehydrogenase detection kit (A020-2-2, Nanjing Jiancheng), Creatine kinase isoenzyme detection kit (H197-1-1, Nanjing Jiancheng), Glutathione Peroxidase Assay Kit (A005-1-2, Nanjing Jiancheng), Lipid oxidation detection kit (A003-1-2, Nanjing Jiancheng), Masson Trichrome Stain Kit (G1340, Solarbio), HE staining kit (G1120, Solarbio), Antibodies: BNP (Cat#AF6336), ANP (Cat#AG3623), CollagenⅠ (Cat#AG1656), TGF-β (Cat#AF0297), COX2 (Cat#AF1924), Cat(Cat#AF0084), SOD1 (Cat#AF8028), cleaved caspase-3 (Cat#AC033), Bax (Cat#AF1270), Bcl-2 (Cat#AB112), AKT (Cat#AF0045), PI3K (Cat#AF7749), p-AKT (Cat#AF1546), p-PI3K (Cat#AF5905), GAPDH (Cat#AF1186) (Biyuntian biotechnology)

2.2 Experimental mice and design

Hunan Slack Jingda Laboratory Animal Co. Ltd. purchased 10-week-old male SPF mice. This experiment meets the requirements of Animal Experiment Ethics Committee of Yichun University (YXY1042-2022). After dosing, blood and cardiac tissue samples were collected for analysis.

2.2.1 Effects of MgIG on Myocardial Remodeling Induced by ISO

Hypertrophy model was established by intraperitoneal injection of ISO (5.0 mg/kg) for 14 days. After established cardiac remodeling by ISO, the mouse were divided into 4 groups with 10 mouse in each group: blank group (Con), positive control group (Con + MgIG), model group (ISO) and administration group (ISO + MgIG). Con group and ISO group were intraperitoneally injected with 50 mg/kg of 0.9% NaCl, and Con + MgIG group and ISO + MgIG group were intraperitoneally injected with 50 mg/kg of MgIG. The mice were administered every other day for 14 days, and the condition of the mice was recorded 24 hours after each administration..

2.2.2 Effects of MgIG based on activation PI3K/AKT1 by 740Y-P

The experimental modeling method is consistent with 2.2.1. The mouse were divided into 6 groups with 10 mouse in each group: blank group (Con), positive control group 1 (Con + MgIG), positive control group 2 (Con + MgIG + 740Y-P), model group (ISO), administration group (ISO + MgIG) and activator group (ISO + MgIG + 740Y-P). Con and ISO groups were intraperitoneally injected with 50 mg/kg of 0.9% NaCl, and Con + MgIG and ISO + MgIG groups were intraperitoneally injected with 50 mg/kg of MgIG. Con + MgIG + 740Y-P group and ISO + MgIG + 740Y-P group were treated with MgIG 50 mg/kg and 740Y-P 40 mg/kg. The mice were administered every other day for 14 days, and the condition of the mice was recorded 24 hours after each administration..

2.3 Heart ultrasonic detection method

The heart function of mouse was detected by small animal ultrasound. After the mouse were anesthetized with isoflurane, the hair was shorn from the neck to the chest with depilatory cream, and the couplant was smeared on the skin. The left ventricular fractional shortening (LVFS) and left ventricular ejection fraction (LVEF) of the mice were observed by ultrasound with the corresponding probe.

2.4 Enzyme activity assay

Blood samples collected from the eyeballs of mice were rapidly centrifuged (5000 rpm, 10 min), and the supernatant was gathered. The levels of LDH, GSH-Px, CK-MB and MDA were determined by the corresponding detection kit.

2.5 Histological examination

The mouse heart tissue was cut and embedded in the embedding box, which was fixed in 10% paraformaldehyde for 24 hours, dehydrated with 75%, 85%, 95% and 100% ethanol concentration gradient, washed with xylene, embedded in paraffin, and prepared into 5 µm sections for HE and Masson staining. After staining, neutral gum was dropped on the sections for sealing. The pathological changes of mouse heart tissue were observed under microscope (Zeiss microscope, Axioplan 2 Imaging, AxioVision software).

2.6 Western blotting

Mouse heart tissue (0.1 g) was shredded pieces with ophthalmic scissors, added with 1 mL of normal saline and ground in a manual homogenization tube to obtain tissue homogenate. The tissue was centrifuged in a low-temperature high-speed centrifuge (12000 rpm, 30 min, 4℃), and the precipitate was discarded. Extract 1 mL of supernatant for BCA protein concentration determination, add 250 µL of protein loading buffer, shake and mix, and heat in a water bath at 95℃ for 5 minutes to denature. The prepared samples were kept in reserve at -80℃ for subsequent Western blot analysis. After the sample solution was added, the protein was separated by electrophoresis, Here, the membrane is cut horizontally, we get the target protein through the pre-stained Mark, and cut the gel position where the target protein is located, and then the gel was transferred to the activated PVDF membrane for membrane transfer treatment. After the membrane transfer was completed, it was put into 5% skim milk powder for blocking incubation for 1 h, and then washed with TBST (5 min/time, 3 times). Rabbit anti-mouse primary antibodies (1:1000) were added and incubated overnight at 4℃. 3 times), Sheep anti-rabbit secondary antibody was added and incubated for 1 hour, the secondary antibody was recovered and washed by TBST (5 min/time, 3 times), the PVDF membrane was soaked in ultra-hypersensitive ECL chemiluminescence solution for 1 min for development, and finally the image was processed by Image J.

2.7 Statistical analysis

SPSS20.0 software was used to analyze the statistical data, and the results were expressed as mean ± standard deviation (x̄ ± s). One-way analysis of variance was used to compare the data of each group, and P < 0.05 was statistically significant.

3.1 MgIG repaired pathological condition of myocardium and reduce cardiac damage

As shown in Figure. 1(A, B), MgIG can improve the myocardial injury induced by ISO in mice. After successful ISO modeling, the heart volume, heart weight ratio and lung weight ratio were meaningfully rose, which were prevented by MgIG treatment. The left ventricular function of mouse in each group was examined by ultrasonic diagnostic instrument, and the left ventricular fractional shortening (LVFS) and left ventricular ejection fraction (LVEF) were recorded (Figure. 1C). As shown in Figure. 1(D, E), there was no significant difference in the morphological structure of the control group, however, ISO group showed noticeably the cells edema, necrosis, infiltration of inflammatory cells, derangement of myocardial cells and enlargement of myocardial fiber bundles. Compared with ISO group, after administration of MgIG, the myocardial injury was gradually repaired.

The degree of myocardial hypertrophy and myocardial fibrosis was detected. As shown in Figure. 2(A-F), western blot detection showed that the expression of BNP, ANP, β-MHC, CollagenⅠ and TGF-β in ISO group was sensibly higher than that in Con group (P < 0.05). Compared with ISO group, the expression of BNP, ANP, β-MHC, CollagenⅠ and TGF-β in ISO + MgIG group was significantly decreased (P < 0.05).

3.2 Pharmacological effects of MgIG on myocardial remodeling induced by ISO in mice

CK-MB, GSH-Px, LDH and MDA are the markers of myocardial injury in serum, which together constitute the defense mechanism in vivo. As shown in Figure. 3A, compared with Con group, the contents of CK-MB, LDH and MDA in ISO group were enhanced, while the content of GSH-Px was decreased (P < 0.05). Compared with ISO group, the contents of CK-MB, LDH and MDA in ISO + MgIG group were decreased, while the content of GSH-Px was rose (P < 0.05).As antioxidant protein, COX2, CAT and SOD1 play an irreplaceable role in that body. As shown in Figure. 3(B-E), the protein expression of COX2, CAT and SOD1 in ISO group was noticeably lower than that in Con group (P < 0.05). Compared with ISO group, the protein expressions of COX2 and SOD1 were up-regulated in ISO + MgIG group (P < 0.05).

As shown in Figure. 4(A-D), compared with Con group, the expression of Bax and Cleaved caspase-3 in ISO group was significantly rose, while the expression of Bcl-2 was reduced (P < 0.05). Compared with ISO group, the protein expression of Bax and Cleaved caspase-3 in ISO + MgIG group was decreased, and the protein expression of Bcl-2 was increased (P < 0.05). As shown in Figure. 4(E-G), compared with Con group, the protein expression of p-PI3K and p-AKT1 was significantly up-regulated in ISO group (P < 0.05). Compared with ISO group, the protein expression of p-PI3K and p-AKT1 was decreased in ISO + MgIG group (P < 0.05).

3.3 Activation of the PI3K/AKT pathway reverses the protective action of MgIG on cardiac function

The above results indicate that MgIG could ameliorate the adverse effects of myocardial remodeling in mice and can inhibit phosphorylation of PI3K/AKT1 pathway, so we hypothese that PI3K/AKT1 was potential targets for MgIG effects. For futher research, we used 740Y-P to activite the effects of PI3K/AKT1 pathway. As shown in Figure. 5(A-C), compared with Con group, heart weight ratio and lung weight ratio in ISO group were noticeably rose (P < 0.05), and the left ventricular function was noticeably reduced (P < 0.05). After the administration of MgIG, the myocardium of mice was improved (P < 0.05), but after the combination of 740Y-P, the improvement of MgIG was reversed (P < 0.05). Through the pathological analysis of the mouse heart Figure. 5(D,E), compared with the Con group, the ISO group showed myocardial edema, cell necrosis, disordered arrangement, and thickened cell fiber bundles, while MgIG could play a role in improving the pathological phenomenon, but after adding 740Y-P, it could antagonize the action of MgIG on the myocardium.

3.4 Activation of that PI3K/AKT1 pathway reverse the effects of MgIG.

Serum myocardial injury markers were detected (Figure. 6A), compared with Con, the contents of CK-MB, LDH and MDA in ISO group were noticeably rose (P < 0.05), and the avidity of GSH-Px was noticeably reduced (P < 0.05). Myocardial injury markers were significantly improved in the ISO + MgIG group compared with the ISO group (P < 0.05). Compared with ISO + MgIG group, ISO + MgIG + 740Y-P group reversed the action of MgIG. Through the determination of phenotypic proteins of the myocardium, as shown in Figure. 6(B-G), the protein expression of BNP, ANP, β-MHC, Collagen Ⅰ and TGF-β in ISO group was significantly higher than that in Con group (P < 0.05). Compared with ISO group, the expression of these proteins was significantly decreased in ISO + MgIG group (P < 0.05). Compared with ISO + MgIG group, 740Y-P could reverse the improvement of MgIG (P < 0.05).

3.5 Activation of PI3K/AKT1 pathway reverses the antiapoptotic and antioxidant effects of MgIG

As shown in Figure. 7(A-G), MgIG can regulate apoptosis and oxidative stress through PI3K/AKT1, and 740Y-P can reverse the effect of MgIG. Western blotting results showed that the protein expressions of p-PI3K, p-AKT1 and Bax in ISO group were noticeably higher than those in Con group (P < 0.05), while the protein expressions of Bcl-2, CAT and SOD1 were significantly lower than those in Con group (P < 0.05). Compared with ISO group, the expressions of p-PI3K, p-AKT1 and Bax in ISO + MgIG group were snoticeably decreased (P < 0.05), while the expressions of Bcl-2, CAT and SOD1 in ISO + MgIG group were noticeably rose (P < 0.05). Compared with ISO + MgIG group, 740Y-P could reverse the improvement of MgIG (P < 0.05).

Cardiac hypertrophy is the adaptive change of myocardium when the volume or pressure load increases, which is the response of cardiomyocytes to various physiological or pathological stimuli (Oldfield et al., 2020). However, the progression of myocardial hypertrophy can lead to myocardial fibrosis, which can aggravate cardiovascular disease (Rai et al., 2017). Therefore, reduction of heart fibrosis is a significant factor in the treatment of cardiomyopathy (Zhao et al., 2021). The research have shown that the selection of ISO model can result in cardiac hypertrophy and heart failure, and then cause cell necrosis and interstitial cell fibrosis (Benjamin et al., 1989). Zhao demonstrated that the expression of hypertrophic genes ANP and BNP can be increased by ISO to establish cardiac hypertrophy model (Yuan et al., 2021). Mounting evidence suggest that the construction of models through ISO leads to myocardial fibrosis, which is manifested by collagen deposition (Li et al., 2021). Therefore, ISO was also used to establish cardiac hypertrophy model in this study.

Several recent Studies showed that MgIG can increase the proliferative activity of hepatocytes and has antioxidant and anti-apoptotic effects in the treatment of hepatic ischemia-reperfusion (Zheng et al., 2015). MgIG can reduce inflammation and apoptosis, treat liver damage and reinforce liver function (Rong et al., 1963). Jiang found that MgIG plays a protective role in cyclophosphamide-induced liver damage model by reducing oxidative stress and anti-apoptosis (Jiang et al., 2017). And myocardial injury can be improved by inhibiting oxidative stress (Zhou et al., 2018). The expression of pro-apoptosis protein Bax and Caspase-3 increased, but the level of anti-apoptosis protein Bcl-2 decreased, it can lead to the aggravation of myocardial injury (Guo et al., 2010). The pharmacological effects of MgIG on myocardium were detected, and the myocardial hypertrophy model was founded by ISO. Homologous results were found in this study, through the determination of the damage substance in the serum, ISO can increase the content of CK-MB, LDH and MDA, decrease the activity of GSH-Px, and reduce the protein expression of COX2, CAT and SOD1, thus destroy the oxidative defense mechanism in vivo. By detecting Bax, Bcl-2 and Cleaved caspase-3 proteins, it was found that ISO could accelerate apoptosis. However, MgIG can effectively improve this series of adverse reactions of cardiomyocytes induced by ISO. It is suggested that MgIG has a protective action on myocardial remodeling leaded by ISO.

The regulation of relevant signaling pathways is essential for the prevention and treatment of pathological heart hypertrophy. The research have shown that cardiac hypertrophy is regulated by multiple pathways, such as JAK/STAT, MAPK and PI3K (Varghese et al., 2022; Shen et al., 2021). However, PI3K is a critical enzyme in the induction of heart hypertrophy, interstitial fibrosis and cardiac dysfunction, while AKT1 is the main target of PI3K signaling and plays an important role in improving cardiac hypertrophy (Cheng et al., 2021). Inhibition of PI3K/AKT1 pathway can effectively reduce cardiac hypertrophy (Soudeh et al., 2022) and improve the anti-apoptotic ability of cells (Zhang et al., 2023). In this study, we speculate that MgIG can alleviate the pathological condition of myocardial hypertrophy leaded by ISO. This may be relevant to the excessive activation of PI3K/AKT1 pathway by ISO and the inhibition of PI3K/AKT1 pathway. Based on the above effects, we detected the PI3K/AKT1 pathway and found that ISO could over-activate the PI3K/AKT1 pathway and increase its phosphorylation, but MgIG could reduce the phosphorylation expression of PI3K/AkT1 pathway. Based on this phenomenon, we added 740Y-P to verify the mechanism of MgIG.

Liang confirmed that 740Y-P can activate PI3K/AKT to reverse the suppressive effect of vitexin on endometrial cancer (Liang et al., 2023). Meanwhile, the previous study reported that 740Y-P can up-regulate the expression of p-PI3K and p-AKT (Jiang et al., 2015). Therefore, by adding 740Y-P, we validated for the first time that MgIG alleviated ISO-leaded myocardial remodeling through PI3K/AKT1 signaling. This study shows that ISO can over-activate PI3K channels and lead to myocardial remodeling. 740Y-P could reverse the effect of MgIG and cause persistent dysfunction of left ventricle. HE/Masson pathological staining showed that 740Y-P could increase the severity of cardiac pathology, aggravate the soak of inflammatory cells and the necrosis of myocardial cells, and thicken the myocardial fiber bundles. 740Y-P also aggravated myocardial fibrosis and hypertrophy by increasing the protein expression of BNP, β-MHC, ANP,,TGF-β and CollagenⅠ. These results indicate that 740Y-P can reverse the relieving effect of MgIG on myocardium.

PI3K/AKT1 pathway defends the myocardium by suppressing or strengthening the expression of downstream related target proteins. AKT1 is activated by activating PI3K, and then apoptosis is mediated by regulating Bcl-2 factor (Yu et al., 2023). Sun proposed that Bcl-2 family proteins regulate ISO-induced cardiac apoptosis by down-regulating Bcl-2 and up-regulating Bax (Sun et al., 2006). In this study, after MgIG treatment, the expression of Bax was decreased and the expression of Bcl-2 was increased, which reduced the necrosis of cardiomyocytes caused by ISO. However, 740Y-P treatment reversed the anti-apoptotic effect of MgIG. Lipid peroxidation induced by excessive oxidative stress can lead to the necrosis of myocardial cells, and then lead to the damage of cardiac tissue. MDA is the stable product of lipid peroxidation of myocardial cell membrane, which has certain biological toxicity, and its content directly reflects the degree of oxidative damage of myocardial tissue (Yin et al., 2019). In this project, we discovered that MgIG could ameliorate the oxidative stress damage caused by ISO. However, 740Y-P reversed the antioxidant effect of MgIG. Finally, we detected the expression of PI3K/AKT1 pathway protein phosphorylation and found that MgIG could effectively reduce the expression of PI3K and AKT1 hyperphosphorylation induced by ISO, while 740Y-P reversed the effect of MgIG.

In summary, MgIG can effectively alleviate ISO-induced myocardial remodeling in mice by adjusting oxidative stress and apoptosis. After the addition of PI3K activator, 740Y-P could effectively reverse the therapeutic effect of MgIG. Therefore, we found that the therapeutic mechanism of MgIG may be related to the suppression of PI3K/AKT1 pathway. Finally MgIG may be a prospective agent for the treatment of myocardial damage.

Ethical Approval

The animal study was reviewed and approved by Medical College of YiChun University.

Consent to Participate

All authors have their consent to participate.

Consent to Publish

All authors have their consent to publish their work.

Authors Contributions

The authors contributed equally to this work. Xingyu Zhou: Methodology, Data analysis, Investigation, Writing original draf. Kai Qian: Validation, Investigation, Writing review & Editing. Dan Fu: Data analysis, Investigation. Saige Sun: Writing review & editing. Jia Shi: Investigation, Validation. Lei Li: Funding Acquisition, Writing review. All authors contributed to the article and approved the submitted version.

Funding

This study was supported by the Natural Science Foundation of Zhejiang Province China (LZ23H020001), and Natural Foundation of Jiangxi Provincial China (20202BABL216078).

Competing Interests

The authors declare no competing interests.

Availability of data and material

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

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FigS1.png
Figure S1. Effective dose of MgIG ameliorates cardiac dysfunction induced by isoproterenol in mice. (A): Cardiac morphology in 6 groups; (B): Statistical results of HW/BW and LW/BW; (C) Western blotting was used to determine the expression levels of BNP, ANP, β-MHC, CollagenⅠ and TGF-β. (D-H) Quantitative analysis of BNP, ANP, β-MHC, CollagenⅠ and TGF-β protein expression. *Significant difference compared with the Con group, p<0.05; #significant difference compared with the ISO group, p<0.05; ^significant difference compared with the ISO+MgIG-25 group, p<0.05; ^vsignificant difference compared with the ISO+MgIG-50 group, p<0.05. n=3 per group.

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Magnesium isoglycyrrhizinate regulates oxidative stress and apoptosis via PI3K/AKT1 pathway to improve myocardial remodeling in mice

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Abstract

Figures

1. Introduction

2. Materials and Methods

2.1 Reagents and antibodies

2.2 Experimental mice and design

2.2.1 Effects of MgIG on Myocardial Remodeling Induced by ISO

2.2.2 Effects of MgIG based on activation PI3K/AKT1 by 740Y-P

2.3 Heart ultrasonic detection method

2.4 Enzyme activity assay

2.5 Histological examination

2.6 Western blotting

2.7 Statistical analysis

3. Results

3.1 MgIG repaired pathological condition of myocardium and reduce cardiac damage

3.2 Pharmacological effects of MgIG on myocardial remodeling induced by ISO in mice

3.3 Activation of the PI3K/AKT pathway reverses the protective action of MgIG on cardiac function

3.4 Activation of that PI3K/AKT1 pathway reverse the effects of MgIG.

3.5 Activation of PI3K/AKT1 pathway reverses the antiapoptotic and antioxidant effects of MgIG

4. Discussion

5. Conclusions

Declarations

References

Supplementary Files

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