Transcriptomic and Proteomic Analysis Reveals the Potential Role of RBMS1 in Adipogenesis and Adipocyte Metabolism.

Adipocytes play a critical role in maintaining a healthy systemic metabolism by storing and releasing energy in the form of fat and helping to regulate glucose and lipid levels in the body. Adipogenesis is the process through which pre-adipocytes are differentiated into mature adipocytes. It is a complex process involving various transcription factors and signaling pathways. The dysregulation of adipogenesis has been implicated in the development of obesity and metabolic disorders. Therefore, understanding the mechanisms that regulate adipogenesis and the factors that contribute to its dysregulation may provide insights into the prevention and treatment of these conditions. RNA-binding motif single-stranded interacting protein 1 (RBMS1) is a protein that binds to RNA and plays a critical role in various cellular processes such as alternative splicing, mRNA stability, and translation. RBMS1 polymorphism has been shown to be associated with obesity and type 2 diabetes, but the role of RBMS1 in adipose metabolism and adipogenesis is not known. We show that RBMS1 is highly expressed during the early phase of the differentiation of the murine adipocyte cell line 3T3-L1 and is significantly upregulated in the adipose tissue depots and adipocytes of high-fat-fed mice, implying a possible role in adipogenesis and adipose metabolism. Knockdown of RBMS1 in pre-adipocytes impacted the differentiation process and reduced the expression of some of the key adipogenic markers. Transcriptomic and proteomic analysis indicated that RBMS1 depletion affected the expression of several genes involved in major metabolic processes, including carbohydrate and lipid metabolism. Our findings imply that RBMS1 plays an important role in adipocyte metabolism and may offer novel therapeutic opportunity for metabolic disorders such as obesity and type 2 diabetes.

Whole genome transcriptomic reveals heat stroke molecular signatures in humans.

An evolutionary heat shock response (HSR) protects most living species, including humans, from heat-induced macromolecular damage. However, its role in the pathogenesis of heat stroke is unknown. We examined the whole genome transcriptome in peripheral blood mononuclear cells of a cohort of subjects exposed to the same high environmental heat conditions, who developed heat stroke (n = 19) versus those who did not (n = 19). Patients with heat stroke had a mean rectal temperature at admission of 41.7 ± 0.8°C, and eight were in deep coma (Glasgow Coma Score = 3). The transcriptome showed that genes involved in more than half of the entire chaperome were differentially expressed relative to heat stress control. These include the heat shock protein, cochaperone, and chaperonin genes, indicating a robust HSR. Differentially expressed genes also encoded proteins related to unfolded protein response, DNA repair, energy metabolism, oxidative stress, and immunity. The analysis predicted perturbations of the proteome network and energy production. Cooling therapy attenuated these alterations without complete restoration of homeostasis. We validated the significantly expressed genes by a real-time polymerase chain reaction. The findings reveal the molecular signature of heat stroke. They also suggested that a powerful HSR may not be sufficient to protect against heat injury. The overwhelming proteotoxicity and energy failure could play a pathogenic role. KEY POINTS: Most living species, including humans, have inherent heat stress response (HSR) that shields them against heat-induced macromolecular damage. The role of the HSR in subjects exposed to environmental heat who progressed to heat stroke versus those that did not is unknown. Our findings suggest that heat stroke induces a broad and robust HSR of nearly half of the total heat shock proteins, cochaperones, and chaperonin genes. Heat stroke patients exhibited inhibition of genes involved in energy production, including oxidative phosphorylation and ATP production. Significant enrichment of neurodegenerative pathways, including amyloid processing signalling, the Huntington's and Parkinson's disease signalling suggestive of brain proteotoxicity was noted. The data suggests that more than a powerful HSR may be required to protect against heat stroke. Overwhelming proteotoxicity and energy failure might contribute to its pathogenesis.

Profiling of G-Protein Coupled Receptors in Adipose Tissue and Differentiating Adipocytes Offers a Translational Resource for Obesity/Metabolic Research.

G protein-coupled receptors (GPCRs) are expressed essentially on all cells, facilitating cellular responses to external stimuli, and are involved in nearly every biological process. Several members of this family play significant roles in the regulation of adipogenesis and adipose metabolism. However, the expression and functional significance of a vast number of GPCRs in adipose tissue are unknown. We used a high-throughput RT-PCR panel to determine the expression of the entire repertoire of non-sensory GPCRs in mouse white, and brown adipose tissue and assess changes in their expression during adipogenic differentiation of murine adipocyte cell line, 3T3-L1. In addition, the expression of GPCRs in subcutaneous adipose tissues from lean, obese, and diabetic human subjects and in adipocytes isolated from regular chow and high-fat fed mice were evaluated by re-analyzing RNA-sequencing data. We detected a total of 292 and 271 GPCRs in mouse white and brown adipose tissue, respectively. There is a significant overlap in the expression of GPCRs between the two adipose tissue depots, but several GPCRs are specifically expressed in one of the two tissue types. Adipogenic differentiation of 3T3-L1 cells had a profound impact on the expression of several GPCRs. RNA sequencing of subcutaneous adipose from healthy human subjects detected 255 GPCRs and obesity significantly changed the expression of several GPCRs in adipose tissue. High-fat diet had a significant impact on adipocyte GPCR expression that was similar to human obesity. Finally, we report several highly expressed GPCRs with no known role in adipose biology whose expression was significantly altered during adipogenic differentiation, and/or in the diseased human subjects. These GPCRs could play an important role in adipose metabolism and serve as a valuable translational resource for obesity and metabolic research.

Transcriptome Changes and Metabolic Outcomes After Bariatric Surgery in Adults With Obesity and Type 2 Diabetes.

Context: Bariatric surgery has been shown to be effective in inducing complete remission of type 2 diabetes in adults with obesity. However, its efficacy in achieving complete diabetes remission remains variable and difficult to predict before surgery. Objective: We aimed to characterize bariatric surgery-induced transcriptome changes associated with diabetes remission and the predictive role of the baseline transcriptome. Methods: We performed a whole-genome microarray in peripheral mononuclear cells at baseline (before surgery) and 2 and 12 months after bariatric surgery in a prospective cohort of 26 adults with obesity and type 2 diabetes. We applied machine learning to the baseline transcriptome to identify genes that predict metabolic outcomes. We validated the microarray expression profile using a real-time polymerase chain reaction. Results: Sixteen patients entered diabetes remission at 12 months and 10 did not. The gene-expression analysis showed similarities and differences between responders and nonresponders. The difference included the expression of critical genes (SKT4, SIRT1, and TNF superfamily), metabolic and signaling pathways (Hippo, Sirtuin, ARE-mediated messenger RNA degradation, MSP-RON, and Huntington), and predicted biological functions (ß-cell growth and proliferation, insulin and glucose metabolism, energy balance, inflammation, and neurodegeneration). Modeling the baseline transcriptome identified 10 genes that could hypothetically predict the metabolic outcome before bariatric surgery. Conclusion: The changes in the transcriptome after bariatric surgery distinguish patients in whom diabetes enters complete remission from those who do not. The baseline transcriptome can contribute to the prediction of bariatric surgery-induced diabetes remission preoperatively.

Free Fatty Acid Receptors (FFARs) in Adipose: Physiological Role and Therapeutic Outlook.

Fatty acids (FFAs) are important biological molecules that serve as a major energy source and are key components of biological membranes. In addition, FFAs play important roles in metabolic regulation and contribute to the development and progression of metabolic disorders like diabetes. Recent studies have shown that FFAs can act as important ligands of G-protein-coupled receptors (GPCRs) on the surface of cells and impact key physiological processes. Free fatty acid-activated receptors include FFAR1 (GPR40), FFAR2 (GPR43), FFAR3 (GPR41), and FFAR4 (GPR120). FFAR2 and FFAR3 are activated by short-chain fatty acids like acetate, propionate, and butyrate, whereas FFAR1 and FFAR4 are activated by medium- and long-chain fatty acids like palmitate, oleate, linoleate, and others. FFARs have attracted considerable attention over the last few years and have become attractive pharmacological targets in the treatment of type 2 diabetes and metabolic syndrome. Several lines of evidence point to their importance in the regulation of whole-body metabolic homeostasis including adipose metabolism. Here, we summarize our current understanding of the physiological functions of FFAR isoforms in adipose biology and explore the prospect of FFAR-based therapies to treat patients with obesity and Type 2 diabetes.

Functional Characterization of the Obesity-Linked Variant of the ß3-Adrenergic Receptor.

Adrenergic receptor ß3 (ADRß3) is a member of the rhodopsin-like G protein-coupled receptor family. The binding of the ligand to ADRß3 activates adenylate cyclase and increases cAMP in the cells. ADRß3 is highly expressed in white and brown adipocytes and controls key regulatory pathways of lipid metabolism. Trp64Arg (W64R) polymorphism in the ADRß3 is associated with the early development of type 2 diabetes mellitus, lower resting metabolic rate, abdominal obesity, and insulin resistance. It is unclear how the substitution of W64R affects the functioning of ADRß3. This study was initiated to functionally characterize this obesity-linked variant of ADRß3. We evaluated in detail the expression, subcellular distribution, and post-activation behavior of the WT and W64R ADRß3 using single cell quantitative fluorescence microscopy. When expressed in HEK 293 cells, ADRß3 shows a typical distribution displayed by other GPCRs with a predominant localization at the cell surface. Unlike adrenergic receptor ß2 (ADRß2), agonist-induced desensitization of ADRß3 does not involve loss of cell surface expression. WT and W64R variant of ADRß3 displayed comparable biochemical properties, and there was no significant impact of the substitution of tryptophan with arginine on the expression, cellular distribution, signaling, and post-activation behavior of ADRß3. The obesity-linked W64R variant of ADRß3 is indistinguishable from the WT ADRß3 in terms of expression, cellular distribution, signaling, and post-activation behavior.

Obesity and COVID-19: what makes obese host so vulnerable?

The disease (COVID-19) novel coronavirus pandemic has so far infected millions resulting in the death of over a million people as of Oct 2020. More than 90% of those infected with COVID-19 show mild or no symptoms but the rest of the infected cases show severe symptoms resulting in significant mortality. Age has emerged as a major factor to predict the severity of the disease and mortality rates are significantly higher in elderly patients. Besides, patients with underlying conditions like Type 2 diabetes, cardiovascular diseases, hypertension, and cancer have an increased risk of severe disease and death due to COVID-19 infection. Obesity has emerged as a novel risk factor for hospitalization and death due to COVID-19. Several independent studies have observed that people with obesity are at a greater risk of severe disease and death due to COVID-19. Here we review the published data related to obesity and overweight to assess the possible risk and outcome in Covid-19 patients based on their body weight. Besides, we explore how the obese host provides a unique microenvironment for disease pathogenesis, resulting in increased severity of the disease and poor outcome.

Free fatty acids receptors 2 and 3 control cell proliferation by regulating cellular glucose uptake.

BACKGROUND: Colorectal cancer (CRC) is a worldwide problem, which has been associated with changes in diet and lifestyle pattern. As a result of colonic fermentation of dietary fibres, short chain free fatty acids are generated which activate free fatty acid receptors (FFAR) 2 and 3. FFAR2 and FFAR3 genes are abundantly expressed in colonic epithelium and play an important role in the metabolic homeostasis of colonic epithelial cells. Earlier studies point to the involvement of FFAR2 in colorectal carcinogenesis. AIM: To understand the role of short chain FFARs in CRC. METHODS: Transcriptome analysis console software was used to analyse microarray data from CRC patients and cell lines. We employed short-hairpin RNA mediated down regulation of FFAR2 and FFAR3 genes, which was validated using quantitative real time polymerase chain reaction. Assays for glucose uptake and cyclic adenosine monophosphate (cAMP) generation was done along with immunofluorescence studies to study the effects of FFAR2/FFAR3 knockdown. For measuring cell proliferation, we employed real time electrical impedance-based assay available from xCELLigence. RESULTS: Microarray data analysis of CRC patient samples showed a significant down regulation of FFAR2 gene expression. This prompted us to study the FFAR2 in CRC. Since, FFAR3 shares significant structural and functional homology with FFAR2, we knocked down both these receptors in CRC cell line HCT 116. These modified cell lines exhibited higher proliferation rate and were found to have increased glucose uptake as well as increased level of glucose transporter 1. Since, FFAR2 and FFAR3 signal through G protein subunit (Gαi), knockdown of these receptors was associated with increased cAMP. Inhibition of protein kinase A (PKA) did not alter the growth and proliferation of these cells indicating a mechanism independent of cAMP/PKA pathway. CONCLUSION: Our results suggest role of FFAR2/FFAR3 genes in increased proliferation of colon cancer cells via enhanced glucose uptake and exclude the role of PKA mediated cAMP signalling. Alternate pathways could be involved that would ultimately result in increased cell proliferation as a result of down regulated FFAR2/FFAR3 genes. This study paves the way to understand the mechanism of action of short chain FFARs in CRC.

Proteomic and Molecular Assessment of the Common Saudi Variant in ACADVL Gene Through Mesenchymal Stem Cells.

Very-long-chain acyl-coenzyme A dehydrogenase (VLCAD) is a coenzyme encoded by ACADVL that converts very-long-chain fatty acids into energy. This process is disrupted by c.65C > A; p.Ser22∗ mutation. To clarify mechanisms by which this mutation leads to VLCAD deficiency, we evaluated differences in molecular and cellular functions between mesenchymal stem cells with normal and mutant VLCAD. Saudi Arabia have a high incidence of this form of mutation. Stem cells with mutant VLCAD were isolated from skin of two patients. Metabolic activity and proliferation were evaluated. The Same evaluation was repeated on normal stem cells introduced with same mutation by CRISPR. Mitochondrial depiction was done by electron microscope and proteomic analysis was done on patients' cells. Metabolic activity and proliferation were significantly lower in patients' cells. Introducing the same mutation into normal stem cells resulted in the same defects. We detected mitochondrial abnormalities by electron microscopy in addition to poor wound healing and migration processes in mutant cells. Furthermore, in a proteomic analysis, we identified several upregulated or downregulated proteins related to hypoglycemia, liver disorder, and cardiac and muscle involvement. We concluded experimental assays of mutant ACADVL (c.65C > A; p.Ser22∗) contribute to severe neonatal disorders with hypoglycemia, liver disorder, and cardiac and muscle involvement.

Heatstroke.

Heatstroke is an acute medical emergency that is always fatal if left untreated. The diagnosis of heatstroke should be considered in any hyperthermic patient with altered mental status during a heat wave or following vigorous muscle exertion. Heat can damage the structure and function of essential macromolecules, including proteins, membrane lipids, and nucleic acids, and thereby lead to multiple-organ failure, culminating in death. The cytotoxic effect of heat is a function of degree and duration of hyperthermia; thus, an early diagnosis and prompt initiation of cooling are paramount to halt progression to tissue damage and death.

TXNIP in Metabolic Regulation: Physiological Role and Therapeutic Outlook.

Background & Objective: Thioredoxin-interacting protein (TXNIP) also known as thioredoxin binding protein-2 is a ubiquitously expressed protein that interacts and negatively regulates expression and function of Thioredoxin (TXN). Over the last few years, TXNIP has attracted considerable attention due to its wide-ranging functions impacting several aspects of energy metabolism. TXNIP acts as an important regulator of glucose and lipid metabolism through pleiotropic actions including regulation of ß-cell function, hepatic glucose production, peripheral glucose uptake, adipogenesis, and substrate utilization. Overexpression of TXNIP in animal models has been shown to induce apoptosis of pancreatic ß-cells, reduce insulin sensitivity in peripheral tissues like skeletal muscle and adipose, and decrease energy expenditure. On the contrary, TXNIP deficient animals are protected from diet induced insulin resistance and type 2 diabetes. SUMMARY: Consequently, targeting TXNIP is thought to offer novel therapeutic opportunity and TXNIP inhibitors have the potential to become a powerful therapeutic tool for the treatment of diabetes mellitus. Here we summarize the current state of our understanding of TXNIP biology, highlight its role in metabolic regulation and raise critical questions that could help future research to exploit TXNIP as a therapeutic target.