Introduction
Autophagy is a basic cell survival mechanism that allows cells to respond to metabolic stress by degrading and recycling intracellular components, in order to generate macromolecular precursors and energy. The autophagy system is essential for the development, maintenance of cellular and tissue homeostasis, immunity, and disease prevention in humans. Autophagy defects have been linked to cancer, neurodegeneration, muscle and cardiac disease, infectious disease, and aging. Although autophagy has once been considered to have a passive quality control and general housekeeping function, new data has revealed that this is an active process that regulates the metabolic condition of cells. Adult stem cells have unique metabolic requirements, since these are long-lived cells with the ability to self-renew and develop into specialized cells throughout the body. Autophagy has key functions in stem cell quiescence, activation, differentiation and self-renewal. Furthermore, autophagy is a crucial regulator of stem cell activity. Impaired autophagy in stem cells contributes to degenerative illness, aging, and the formation of cancer stem cells. Thus, the use of autophagy as a regenerative medicine technique can improve stem cell activity and stem cell-based therapeutics.1
Right heart ventricular dysfunction would consecutively lead to abnormal liver function.1 Even in the absence of concomitant heart illness, chronic liver diseases can have an impact on cardiac function. Therefore, improving cardiac dysfunction is essential to improve hepatic dysfunction. A severe increase in alanine aminotransferase (ALT), aspartate aminotransferase (AST) and lactate dehydrogenase (LDH) levels would lead to cardiac ischemic hepatitis within 5–10 days of onset.2
The synthetic catecholamine, novodrin, can induce infarct-like lesions in animal models. Novodrin overdose can affect the liver’s metabolism by disrupting the equilibrium between its oxidant and antioxidant levels. The production of harmful reactive oxygen species (ROS), such as OH•, O2• and H2O2, can damage the liver. Novodrin is an α-adrenergic agonist and synthetic catecholamine, and it was previously noted that this can cause severe myocardial dysfunction, which results in hepatic injury. Furthermore, novodrin can rapidly oxidize, and the oxidative byproducts are the cause of hepatic dysfunction.3
Beclin-1 is important for mammalian autophagy. Furthermore, Beclin-1 is a type III phosphatidylinositol 3-kinase complex component necessary for the production of autophagic vesicles. Human melanoma, colon, ovarian, liver and brain malignancies have been linked to abnormal Beclin-1 expression.4
Both the liver and the large intestine consistently express X-box binding protein 1 (XBP-1). XBP-1 possesses specific RNA polymerase II-enzyme binding activity, and promotes DNA-binding transcription factor and PI3K regulatory subunit binding. This has a vital role in several functions, including lipid homeostasis, and cellular response to hexose stimulation, in addition to controlling the macromolecule metabolism. Furthermore, this can affect a number of activities, including the cellular response to the formation of the exocrine system, the leukemia inhibitory factor, and the positive control of the proteasomal protein catabolic process. XBP1 deficit triggers liver pyroptosis by limiting mitophagy activation of mtDNA/cGAS/STING signaling in macrophages, supplying potential therapeutic targets for liver injury. According to a study conducted XBP-1 can be utilized to examine inflammatory bowel illness and liver dysfunction.5
Mesenchymal stem cells (MSCs) have the ability to clonally differentiate into adipocytes, chondrocytes, hepatocytes and osteoblasts.6 Furthermore, MSCs are suitable for transplantation, contributing to its multi-lineage potential capacity, and preventing immune system recognition after transplantation and facile multiplication.7,8
Liver transplantation has been regarded as the sole curative option for late-stage liver injury.9 However, liver transplantation is not considered a practical alternative, due to its limitations, the danger of immunological rejection, and the dependence on donor supply. Consequently, investigations are being conducted in a range of therapeutic models, including stem cell transplantation.
MSCs are present in various tissues, including umbilical cord, bone marrow, placenta, adipocyte, adult, and embryonic tissues.10 Due to its significant therapeutic potential, MSCs have gained great consideration in the arena of regenerative medication. A modest amount of aspirated bone marrow can be used to separate and cultivate bone marrow-derived mesenchymal stem cells (BM-MSCs) without reducing its potency. Numerous researches have revealed that BM-MSCs can develop into tissues that resemble hepatocytes. These cells interact with different immune cells, have minimal inherent immunogenicity, and may modulate immunological responses, overall increasing the safety of employing MSCs in liver therapy.6 By performing the functional action of mature hepatocytes, which are engaged in supportive functions necessary for regenerative therapy practice, MSCs can enhance liver function.11
The present study aims to assess the liver dysfunction correlated with myocardial infarction (MI) following novodrin hepatotoxicity. In addition, the present study was expanded to compare the autophagy genomic pathways in rats that received MSCs as treatment.
Discussion
Autophagy is a highly conserved cellular mechanism that sequesters damaged or toxic cytoplasmic components in autophagosomes, which eventually fuse with the lysosome for destruction. Due to the potential to self-replicate and give rise to any specialized cell type, MSCs have become extremely significant resources for cell-based medicinal interventions, and these have opened various interesting avenues for research on human disease. It has been proposed that autophagy is essential for the preservation of cellular homeostasis in MSCs, and the regulation of MSC self-renewal and differentiation.6
Furthermore, autophagy is a highly conserved procedure regulated by complicated signaling networks, including the AKT/PKB system, the PI3K inositol pathway, the p52 route, endoplasmic reticulum stress, and AMP-activated protein kinase (AMPK) and ROS production. Furthermore, autophagy is induced when the AMPK pathway is activated. The AMPK pathway is activated by metabolic stress, culminating in the phosphorylation of p27, which is a cyclin-dependent kinase inhibitor. The phosphorylation of p27 improves its stability, and this allows cells to survive the growth factor withdrawal through autophagy.6
In addition, the autophagy process has been recently identified as the fundamental mechanism by which MSCs achieve its precise morphology and function by controlling the protein turnover. Recent research has revealed that autophagy activation is required for MSC self-renewal and differentiation.17 Furthermore, autophagy can efficiently transport sets of adhesion molecules, transcription factors, or released factors, in response to environment stimulation and hormone activation, and all of these are critical for stem cell self-renewal and differentiation.17
Previous research indicated that the liver’s regenerating ability is primarily due to resident hepatic progenitor cells (HPCs), which are defined as cells that give rise to both biliary epithelial cells and hepatocytes at the post-liver injury. The role of autophagy in hepatocyte regulation has been extensively investigated in liver regeneration,6 and the maintenance of metabolic balance in the liver. It has been observed that the knockdown of the critical autophagy gene Beclin-1 reduced the clonogenic and proliferative capacity of HPCs. Furthermore, it has been demonstrated that autophagy deficiency enhances the accumulation of damaged mitochondria and mitochondrial ROS, and blocks the homologous recombination pathway for DNA damage repair in HPCs.10 These results demonstrate that autophagy plays an indispensable role in MSC-associated expansion. In the present study, comparisons were conducted to investigate the effectiveness of transplanted MSCs in reducing rat liver injury induced by novodrin, and the potential autophagy pathways involved.
In the present study, novodrin increased the ALT and AST levels, when compared to the controls. As indicated by the novodrin-induced hepatic injury and plasma membrane leakiness, the elevated liver biomarkers were linked to the loss of functional integrity of the liver cell membrane, and cellular leakage. In previous research, carbon tetrachloride was employed to establish the liver cirrhosis model,18 and the BM-MSC group presented with significantly lower levels of ALT, alkaline phosphatase, LDH, AST, and liver function indicators (bilirubin, albumin, and total protein), when compared to the carbon tetrachloride-treated group. This result is consistent with the results of other studies, which reported that transplanted MSCs may considerably reduce liver injury by restoring blood-liver function biomarkers and liver enzyme levels.10,19
It may be considered that the novodrin-prompted lipid peroxide (LPO) and production of ROS are responsible for the hepatic membrane injury, and leakage of LDH, ALT, and AST in serum.12 Oxidative stress, which results from the increase in free radical production and decreases in antioxidant levels in hepatocytes, is a crucial factor in the development of hepatic disorders, such as necrosis. The production of harmful ROS, such as O2•, H2O2 and OH•, would damage the hepatocytes. Hepatotoxicity has been linked to the β-adrenergic agonist, novodrin, which rapidly goes through auto-oxidation, and it was proposed that alterations in hepatocytes are caused by the oxidative byproducts of catecholamines.15
In this regard, the present study revealed that the novodrin intoxication significantly increased the hepatic MDA level, demonstrating that the liver of novodrin-intoxicated rats experienced severe oxidative stress. A well-known mechanism of cellular damage is lipid peroxidation, which is a marker for oxidative stress that contributes to MI and is linked to liver dysfunction. Novodrin-intoxicated rats with MI have been shown to present with LPO-mediated liver injury.20
Previous studies have reported that hepatic dysfunction, inflammatory cascades, and histological alterations, as the result of novodrin injections, markedly decreased the glutathione, superoxide dismutase and catalase activity, and enhanced the vascular responsiveness to different catecholamines. The pathogenic stage that makes liver tissue more vulnerable to oxidative injury appears to be the increase in lipid peroxidation. Novodrin quinone metabolites may cause oxidative stress by interacting with antioxidant enzymes to synthesize O2• and other ROS.15
The stem cell transplantation led to a substantial decrease in MDA levels, as a result of the increase in hepatic antioxidants and glutathione levels. Kumar et al. reported that the main molecular pathway behind the curative impact of MSCs in hepatotoxicity is the antioxidant and ROS quenching properties.21–23
Novodrin significantly upregulated the PTEN, STAT5A, PI3K and AKT1 genes, and these were modulated by the post-stem cell treatment. Meanwhile, MSCs elevated the PIP3 and p-AKT protein expression. This was highlighted by the study conducted by Chen et al., which revealed that BM-MSCs resemble immune cells, and react to various stress signals induced by injured areas at post-inflammation, similar to the case of hepatic and autoimmune illnesses.24 The ability of MSCs to discriminate into hepatocytes, and the immune-modulatory and antioxidant capabilities are all factors that may contribute to its therapeutic index in liver injury.25 For its capacity for multipotent discrimination, self-regeneration and low immunogenicity, MSCs exhibit a great prospective for therapeutic utility.26 Numerous reports have revealed how MSCs can diminish liver injury.6,10
PIP3 is produced by a lipid kinase, which is known as PI3K. Tris-phosphate is a secondary messenger and is a vital AKT1 plasma membrane transporter, which is activated and phosphorylated. According to the studies conducted by Wang et al. and Záleák et al., the activation of AKT1 is essential for cell survival and proliferation.27,28 MSCs release a range of physiologically active chemicals, including growth factors, chemokines and cytokines, and together, this is referred to as the MSC secretome, which in physiopathological circumstances, is similar to ischemia. Recently, the idea of the MSC secretome has been considered to be crucial to mechanisms that involve repair.29 The role of the PI3K/AKT pathway in MSC survival was previously reported through the overexpression of certain pathway-specific genes. Rat heart transplantation can result in improved MSC survival, when AKT1 is overexpressed in the transplanted cells. Furthermore, anti-apoptotic protein, B-cell leukemia/lymphoma-2 (Bcl-2), was upregulated in MSCs that overexpressed AKT1, while Bax, which is a pro-apoptotic protein, decreased. It has been demonstrated that MSC survival in MI therapy increased after overexpressing Bcl-2.30 Cell apoptosis may be impacted by statins (STAT) through lowering anti-apoptotic biomarkers Bcl-xL and Bcl-2, and increasing the activity of pro-apoptotic biomarkers, caspase 3, 8 and 9, Bax and Bad, and some tumors have been treated using these effects.31,32 In general, the PI3K/AKT pathway functions as a survival mechanism that regulates cell division, migration, proliferation and apoptosis. The results for the overexpression of several pathway-specific genes present the role of the PI3K/AKT pathway in MSC survival.33
BM-MSCs may lower acute liver inflammation, and its subsequent hepatocyte damage by secreting various soluble compounds and trophic factors, such as growth factors, some specific cytokines, and chemokines, which have significant therapeutic roles in regenerative medicine.34 Furthermore, MSCs control the production of inflammatory cytokines, which produce hepato-protection and inflammatory biomarkers. Moreover, in addition to limiting cell death, inflammation and fibrosis in injured tissues, these promote tissue regeneration and angiogenesis.35,36
The novodrin intoxication significantly downregulated the gene expression of autophagy biomarkers, including Beclin-1 and XBP-1, when compared to the controls. However, these biomarkers were considerably upregulated in rats that received stem cell treatment.
Recent research revealed that autophagy, which is a conserved cellular response to stress, plays a part in human malignancies. Beclin-1, which is a crucial autophagic gene, has been reported to have an aberrant expression in a number of human malignancies. Furthermore, a previous study determined the correlation between Beclin-1, and cell apoptosis and proliferation or prognosis in liver carcinoma. Hepatic carcinoma pathophysiology and development are linked to diminished autophagy. The Bax and Beclin-1 expression in hepatic carcinoma have a synergistic impact in reducing the growth, invasion, metastasis, and angiogenesis of a disease. The expression of Beclin-1 can be a useful prognostic indicator for hepatocellular carcinoma.
XBP-1 is a transcription factor from the cAMP-response element binding protein/Cyclic AMP-dependent transcription factor family, which is highly expressed in hepatocellular carcinomas. Furthermore, XBP-1 is necessary for the growth of the liver. XBP-1 deficient mice presented with hypoplastic fetal livers, and the diminished hematopoiesis induced anemia, which led to mortality. However, there were no cell-autonomous differentiation deficits in XBP-1-deficient hematopoietic progenitors. Instead, the hepatocyte formation was substantially influenced by two factors: the high rate of apoptosis, and the decreased rate of growth. Thus, XBP-1 is a vital transcription factor required for the proliferation of hepatocytes.37 XBP-1 knockdown may cause organ-specific inflammation, offering a mechanistic explanation for the onset of pro-inflammatory illnesses.5