Increased let-7d-5p in non-alcoholic fatty liver promotes insulin resistance and is a potential blood biomarker for diagnosis.

BACKGROUND AND AIMS: The molecular mechanisms driving non-alcoholic fatty liver disease (NAFLD) are poorly understood; however, microRNAs might play a key role in these processes. We hypothesize that let-7d-5p could contribute to the pathophysiology of NAFLD and serve as a potential diagnostic biomarker. METHODS: We evaluated let-7d-5p levels and its targets in liver biopsies from a cross-sectional study including patients with NAFLD and healthy donors, and from a mouse model of NAFLD. Moreover, the induction of let-7d-5p expression by fatty acids was evaluated in vitro. Further, we overexpressed let-7d-5p in vitro to corroborate the results observed in vivo. Circulating let-7d-5p and its potential as a NAFLD biomarker was determined in isolated extracellular vesicles from human plasma by RT-qPCR. RESULTS: Our results demonstrate that hepatic let-7d-5p was significantly up-regulated in patients with steatosis, and this increase correlated with obesity and a decreased expression of AKT serine/threonine kinase (AKT), insulin-like growth factor 1 (IGF1), IGF-I receptor (IGF1R) and insulin receptor (INSR). These alterations were corroborated in a NAFLD mouse model. In vitro, fatty acids increased let-7d-5p expression, and its overexpression decreased AKT, IGF-IR and IR protein expression. Furthermore, let-7d-5p hindered AKT phosphorylation in vitro after insulin stimulation. Finally, circulating let-7d-5p significantly decreased in steatosis patients and receiver operating characteristic (ROC) analyses confirmed its utility as a diagnostic biomarker. CONCLUSIONS: Our results highlight the emerging role of let-7d-5p as a potential therapeutic target for NAFLD since its overexpression impairs hepatic insulin signalling, and also, as a novel non-invasive biomarker for NAFLD diagnosis.

Oxidative Stress Modulation by ncRNAs and Their Emerging Role as Therapeutic Targets in Atherosclerosis and Non-Alcoholic Fatty Liver Disease.

Atherosclerosis and non-alcoholic fatty liver disease (NAFLD) are pathologies related to ectopic fat accumulation, both of which are continuously increasing in prevalence. These threats are prompting researchers to develop effective therapies for their clinical management. One of the common pathophysiological alterations that underlies both diseases is oxidative stress (OxS), which appears as a result of lipid deposition in affected tissues. However, the molecular mechanisms that lead to OxS generation are different in each disease. Non-coding RNAs (ncRNAs) are RNA transcripts that do not encode proteins and function by regulating gene expression. In recent years, the involvement of ncRNAs in OxS modulation has become more recognized. This review summarizes the most recent advances regarding ncRNA-mediated regulation of OxS in atherosclerosis and NAFLD. In both diseases, ncRNAs can exert pro-oxidant or antioxidant functions by regulating gene targets and even other ncRNAs, positioning them as potential therapeutic targets. Interestingly, both diseases have common altered ncRNAs, suggesting that the same molecule can be targeted simultaneously when both diseases coexist. Finally, since some ncRNAs have already been used as therapeutic agents, their roles as potential drugs for the clinical management of atherosclerosis and NAFLD are analyzed.

Implication of miR-155-5p and miR-143-3p in the Vascular Insulin Resistance and Instability of Human and Experimental Atherosclerotic Plaque.

(1) Background: Cardiovascular diseases (CVDs) are the main cause of death in developed countries, being atherosclerosis, a recurring process underlying their apparition. MicroRNAs (miRNAs) modulate the expression of their targets and have emerged as key players in CVDs; (2) Methods: 18 miRNAs were selected (Pubmed and GEO database) for their possible role in promoting atherosclerosis and were analysed by RT-qPCR in the aorta from apolipoprotein E-deficient (ApoE-/-) mice. Afterwards, the altered miRNAs in the aorta from 18 weeks-ApoE-/- mice were studied in human aortic and carotid samples; (3) Results: miR-155-5p was overexpressed and miR-143-3p was downregulated in mouse and human atherosclerotic lesions. In addition, a significant decrease in protein kinase B (AKT), target of miR-155-5p, and an increase in insulin-like growth factor type II receptor (IGF-IIR), target of miR-143-3p, were noted in aortic roots from ApoE-/- mice and in carotid plaques from patients with advanced carotid atherosclerosis (ACA). Finally, the overexpression of miR-155-5p reduced AKT levels and its phosphorylation in vascular smooth muscle cells, while miR-143-3p overexpression decreased IGF-IIR reducing apoptosis in vascular cells; (4) Conclusions: Our results suggest that miR-155-5p and miR-143-3p may be implicated in insulin resistance and plaque instability by the modulation of their targets AKT and IGF-IIR, contributing to the progression of atherosclerosis.

Concerted regulation of non-alcoholic fatty liver disease progression by microRNAs in apolipoprotein E-deficient mice.

The prevalence of non-alcoholic fatty liver disease (NAFLD) is constantly increasing, and altered expression of microRNAs (miRNAs) fosters the development and progression of many pathologies, including NAFLD. Therefore, we explored the role of new miRNAs involved in the molecular mechanisms that trigger NAFLD progression and evaluated them as biomarkers for diagnosis. As a NAFLD model, we used apolipoprotein E-deficient mice administered a high-fat diet for 8 or 18 weeks. We demonstrated that insulin resistance and decreased lipogenesis and autophagy observed after 18 weeks on the diet are related to a concerted regulation carried out by miR-26b-5p, miR-34a-5p, miR-149-5p and miR-375-3p. We also propose circulating let-7d-5p and miR-146b-5p as potential biomarkers of early stages of NAFLD. Finally, we confirmed that circulating miR-34a-5p and miR-375-3p are elevated in the late stages of NAFLD and that miR-27b-3p and miR-122-5p are increased with disease progression. Our results reveal a synergistic regulation of key processes in NAFLD development and progression by miRNAs. Further investigation is needed to unravel the roles of these miRNAs for developing new strategies for NAFLD treatment. This article has an associated First Person interview with the joint first authors of the paper.

Severe Hepatic Insulin Resistance Induces Vascular Dysfunction: Improvement by Liver-Specific Insulin Receptor Isoform A Gene Therapy in a Murine Diabetic Model.

BACKGROUND: Cardiovascular dysfunction is linked to insulin-resistant states. In this paper, we analyzed whether the severe hepatic insulin resistance of an inducible liver-specific insulin receptor knockout (iLIRKO) might generate vascular insulin resistance and dysfunction, and whether insulin receptor (IR) isoforms gene therapy might revert it. METHODS: We studied in vivo insulin signaling in aorta artery and heart from iLIRKO. Vascular reactivity and the mRNA levels of genes involved in vascular dysfunction were analyzed in thoracic aorta rings by qRT-PCR. Finally, iLIRKO mice were treated with hepatic-specific gene therapy to analyze vascular dysfunction improvement. RESULTS: Our results suggest that severe hepatic insulin resistance was expanded to cardiovascular tissues. This vascular insulin resistance observed in aorta artery from iLIRKO mice correlated with a reduction in both PI3K/AKT/eNOS and p42/44 MAPK pathways, and it might be implicated in their vascular alterations characterized by endothelial dysfunction, hypercontractility and eNOS/iNOS levels' imbalance. Finally, regarding long-term hepatic expression of IR isoforms, IRA was more efficient than IRB in the improvement of vascular dysfunction observed in iLIRKO mice. CONCLUSION: Severe hepatic insulin resistance is sufficient to produce cardiovascular insulin resistance and dysfunction. Long-term hepatic expression of IRA restored the vascular damage observed in iLIRKO mice.

miRNA Dysregulation in the Development of Non-Alcoholic Fatty Liver Disease and the Related Disorders Type 2 Diabetes Mellitus and Cardiovascular Disease.

According to the World Health Organization, the continuing surge in obesity pandemic creates a substantial increase in incidences of metabolic disorders, such as non-alcoholic fatty liver disease (NAFLD), type 2 diabetes mellitus, and cardiovascular disease. MicroRNAs (miRNAs) belong to an evolutionarily conserved class of short (20-22 nucleotides in length) and single-stranded non-coding RNAs. In mammals, miRNAs function as critical post-transcriptional negative regulators involved not only in many biological processes but also in the development of many diseases such as NAFLD and comorbidities. More recently, it has been described that cells can secrete miRNAs in extracellular vesicles, transported by body fluids, and uptaken by other tissues regulating gene expression. Therefore, this could be a mechanism of signaling involved not only in physiological pathways but also in the development of diseases. The association of some miRNA expression profiles with certain disorders has made them very interesting molecules for diagnosis, prognosis, and disease management. The finding of specific miRNA signatures to diagnose NAFLD and related diseases could anticipate the risk of development of related complications and, actually, it is the driving force of present health strategies worldwide. In this review, we have included latest advances in knowledge about the miRNAs involved in the development of NAFLD and related diseases and examined how this knowledge could be used to identify new non-invasive biomarkers and new pharmacological interventions.

The Interplay between Oxidative Stress and miRNAs in Obesity-Associated Hepatic and Vascular Complications.

Nowadays, the obesity pandemic is one of the most relevant health issues worldwide. This condition is tightly related to comorbidities such as non-alcoholic fatty liver disease (NAFLD) and cardiovascular diseases (CVDs), namely atherosclerosis. Dysregulated lipid metabolism and inflammation link these three diseases, leading to a subsequent increase of oxidative stress (OS) causing severe cellular damage. On the other hand, microRNAs (miRNAs) are short, single-stranded, non-coding RNAs that act as post-transcriptional negative regulators of gene expression, thus being involved in the molecular mechanisms that promote the development of many pathologies including obesity and its comorbidities. The involvement of miRNAs in promoting or opposing OS in disease progression is becoming more evident. Some miRNAs, such as miR-200a and miR.421, seem to play important roles in OS control in NAFLD. On the other hand, miR-92a and miR-133, among others, are important in the development of atherosclerosis. Moreover, since both diseases are linked to obesity, they share common altered miRNAs, being miR-34a and miR-21 related to OS. This review summarizes the latest advances in the knowledge about the mechanisms of oxidative stress (OS) generation in obesity-associated NAFLD and atherosclerosis, as well as the role played by miRNAs in the regulation of such mechanisms.