In silico identification of compounds from Piper sarmentosum Roxb leaf fractionated extract inhibit interleukin-6 to prevent rheumatoid arthritis.

Objective: Medicinal herbs with a phytonutrient background has been applied globally as major alternatives to ameliorate the continuous increase in rheumatoid arthritis cases worldwide. We herein aimed to critically examine the bioactive components of the medicinal herb Piper sarmentosum Roxb leaf fractionated extract for its potential to inhibit the influx of interleukin-6 (IL-6) in rheumatoid arthritis. Methods: The Schrödinger platform was employed as the main computational acumen for the screening of bioactive compounds identified and reference compounds subjected to molecular simulation (MDS) for analyzing the stability of docked complexes to assess fluctuations and conformational changes during protein-ligand interactions. Results: The values of the simulatory properties and principal component analysis (PCA) revealed the good stability of these phytochemicals in the active pocket of interleukin-6 (IL-6). Discussion: Our findings reveal new strategies in which these phytochemicals are potential inhibitory agents that can be modified and further evaluated to develop more effective agents for the management of rheumatoid arthritis, thereby providing a better understanding and useful model for the reproduction and/or discovery of new drugs for the management of rheumatoid arthritis and its complications.

Brown Fat and Nutrition: Implications for Nutritional Interventions.

Brown and beige adipocytes are renowned for their unique ability to generate heat through a mechanism known as thermogenesis. This process can be induced by exposure to cold, hormonal signals, drugs, and dietary factors. The activation of these thermogenic adipocytes holds promise for improving glucose metabolism, reducing fat accumulation, and enhancing insulin sensitivity. However, the translation of preclinical findings into effective clinical therapies poses challenges, warranting further research to identify the molecular mechanisms underlying the differentiation and function of brown and beige adipocytes. Consequently, research has focused on the development of drugs, such as mirabegron, ephedrine, and thyroid hormone, that mimic the effects of cold exposure to activate brown fat activity. Additionally, nutritional interventions have been explored as an alternative approach to minimize potential side effects. Brown fat and beige fat have emerged as promising targets for addressing nutritional imbalances, with the potential to develop strategies for mitigating the impact of metabolic diseases. Understanding the influence of nutritional factors on brown fat activity can facilitate the development of strategies to promote its activation and mitigate metabolic disorders.

Reconstructing human brown fat developmental trajectory in vitro.

Brown adipocytes (BAs) represent a specialized cell type that is able to uncouple nutrient catabolism from ATP generation to dissipate energy as heat. In humans, the brown fat tissue is composed of discrete depots found throughout the neck and trunk region. BAs originate from a precursor common to skeletal muscle, but their developmental trajectory remains poorly understood. Here, we used single-cell RNA sequencing to characterize the development of interscapular brown fat in mice. Our analysis identified a transient stage of BA differentiation characterized by the expression of the transcription factor GATA6. We show that recapitulating the sequence of signaling cues identified in mice can lead to efficient differentiation of BAs in vitro from human pluripotent stem cells. These precursors can in turn be efficiently converted into functional BAs that can respond to signals mimicking adrenergic stimuli by increasing their metabolism, resulting in heat production.

U0126 Compound Triggers Thermogenic Differentiation in Preadipocytes via ERK-AMPK Signaling Axis.

In recent years, thermogenic differentiation and activation in brown and white adipose tissues have been regarded as one of the major innovative and promising strategies for the treatment and amelioration of obesity. However, the pharmacological approach towards this process has had limited and insufficient commitments, which presents a greater challenge for obesity treatment. This research evaluates the effects of U0126 compound on the activation of thermogenic differentiation during adipogenesis. The results show that U0126 pretreatment primes both white and brown preadipocytes to upregulate thermogenic and mitochondrial genes as well as enhance functions during the differentiation process. We establish that U0126-mediated thermogenic differentiation induction occurs partially via AMPK activation signaling. The findings of this research suggest U0126 as a promising alternative ligand in pursuit of a pharmacological option to increase thermogenic adipocyte formation and improve energy expenditure. Thus it could pave the way for the discovery of therapeutic drugs for the treatment of obesity and its related complications.

Mitochondrial defects in pancreatic beta-cell dysfunction and neurodegenerative diseases: Pathogenesis and therapeutic applications.

Mitochondria malfunction is linked to the development of ß-cell failure and a variety of neurodegenerative disorders. Pancreatic ß-cells are normally configured to detect glucose and other food secretagogues in order to adjust insulin exocytosis and maintain glucose homeostasis. As a result of the increased glucose level, mitochondria metabolites and nucleotides are produced, which operate in concert with cytosolic Ca2+ to stimulate insulin secretion. Furthermore, mitochondria are the primary generators of adenosine triphosphate (ATP), reactive oxygen species (ROS), and apoptosis regulation. Mitochondria are concentrated in synapses, and any substantial changes in synaptic mitochondria location, shape, quantity, or function might cause oxidative stress, resulting in faulty synaptic transmission, a symptom of various degenerative disorders at an early stage. However, a greater understanding of the role of mitochondria in the etiology of ß-cell dysfunction and neurodegenerative disorder should pave the way for a more effective approach to addressing these health issues. This review looks at the widespread occurrence of mitochondria depletion in humans, and its significance to mitochondria biogenesis in signaling and mitophagy. Proper understanding of the processes might be extremely beneficial in ameliorating the rising worries about mitochondria biogenesis and triggering mitophagy to remove depleted mitochondria, therefore reducing disease pathogenesis.

Disruption of mitochondria-associated ER membranes impairs insulin sensitivity and thermogenic function of adipocytes.

The prevalence and healthcare burden of obesity and its related metabolic disorders such as type 2 diabetes (T2D) are increasing rapidly. A better understanding of the pathogenesis of these diseases helps to find the therapeutic strategies. Mitochondria and endoplasmic reticulum (ER) are two important organelles involved in the maintenance of intracellular Ca2+ and ROS homeostasis. Their functional defects are thought to participate in the pathogenesis of insulin resistance or T2D. The proper structure and function of the mitochondria-associated ER membranes (MAMs) is required for efficient communication between the ER and mitochondria and defects in MAMs have been shown to play a role in metabolic syndrome and other diseases. However, the detailed mechanism to link MAMs dysfunction and pathogenesis of insulin resistance or T2D remains unclear. In the present study, we demonstrated that the proteins involved in .MAMs structure are upregulated and the formation of MAMs is increased during adipogenic differentiation of 3T3-L1 preadipocytes. Disruption of MAMs by knocking down GRP75, which is responsible for connecting ER and mitochondria, led to the impairment of differentiation and ROS accumulation in 3T3-L1 preadipocytes. Most importantly, the differentiated 3T3-L1 adipocytes with GRP75 knockdown displayed inactivation of insulin signaling pathway upon insulin stimulation. Moreover, GRP75 knockdown impaired thermogenesis and glucose utilization in brown adipocytes, the adipocytes with abundant mitochondria that regulate whole-body energy homeostasis. Taken together, our findings suggest that MAMs formation is essential for promoting mitochondrial function and maintaining a proper redox status to enable the differentiation of preadipocytes and normal functioning such as insulin signaling and thermogenesis in mature adipocytes.

Brown adipose tissue-derived MaR2 contributes to cold-induced resolution of inflammation.

Obesity induces chronic inflammation resulting in insulin resistance and metabolic disorders. Cold exposure can improve insulin sensitivity in humans and rodents, but the mechanisms have not been fully elucidated. Here, we find that cold resolves obesity-induced inflammation and insulin resistance and improves glucose tolerance in diet-induced obese mice. The beneficial effects of cold exposure on improving obesity-induced inflammation and insulin resistance depend on brown adipose tissue (BAT) and liver. Using targeted liquid chromatography with tandem mass spectrometry, we discovered that cold and ß3-adrenergic stimulation promote BAT to produce maresin 2 (MaR2), a member of the specialized pro-resolving mediators of bioactive lipids that play a role in the resolution of inflammation. Notably, MaR2 reduces inflammation in obesity in part by targeting macrophages in the liver. Thus, BAT-derived MaR2 could contribute to the beneficial effects of BAT activation in resolving obesity-induced inflammation and may inform therapeutic approaches to combat obesity and its complications.

Hydrophilic-Hydrophobic Nanohybrids of AuNP-Immobilized Two-Dimensional Nanomica Platelets as Flexible Substrates for High-Efficiency and High-Selectivity Surface-Enhanced Raman Scattering Microbe Detection.

A flexible hybrid substrate was developed by affixing gold nanoparticles (AuNPs) onto the surface of two-dimensional nanomica platelets (NMPs). The substrate was successfully used in biosensors with high efficiency and high selectivity through surface-enhanced Raman scattering (SERS). By controlling the amphiphilicity of the hybrid substrate, the flexible substrate was made highly selective toward biomolecules. Four different SERS substrate systems were constructed, including intercalated mica, exfoliated NMPs, hydrophilic exfoliated NMPs, and hydrophobic exfoliated NMPs. NMPs were only 1 nm thick. AuNPs adsorbed on both sides of NMPs and thus created excellent three-dimensional hot junction effects in the z-axis direction. For the detection of adenine in DNA, a satisfactory Raman enhancement factor (EF) of up to 8.9 × 106 was achieved with the detection limit as low as 10-8 M. Subsequently, the AuNP/NMP hybrids were adopted to rapidly detect hydrophilic Staphylococcus hominis and hydrophobic Escherichia coli. The AuNP/PIB-POE-PIB/NMP nanohybrid was concurrently hydrophilic and hydrophobic. This amphiphilic property greatly enhanced the detection selectivity and signal intensity for hydrophilic or hydrophobic bacteria. Overall, AuNPs/PIB-POE-PIB/NMPs developed as SERS substrates enable rapid, sensitive biodetection.

Therapeutic efficacy of Clompanus pubescens leaves fractions via downregulation of neuronal cholinesterases/Na+-K+ATPase/IL-1 ß, and improving the neurocognitive and antioxidants status of streptozotocin-induced diabetic rats.

The increasing global burden of diabetes mellitus has called for the search for a therapeutic alternative that offers better activities and safety than conventional chemotherapy. Herein, we evaluated the neuroprotective and antioxidant properties of different fractions (ethyl acetate, N-butanol and residual aqueous) of Clompanus pubescens leaves in streptozotocin (STZ)-induced diabetic rats. Our results revealed a significant elevation in the levels of blood glucose, pro-inflammatory cytokines, lipid peroxidation, neuronal activities of acetylcholinesterase, butyrylcholinesterase, nitric oxide, epinephrine, norepinephrine, and Na+/K+-ATPase in diabetic non treated rats. In addition, decreased levels of enzymatic and non-enzymatic antioxidants were observed. Treatment with different fractions of C. pubescens leaves resulted in significant reversal of the biochemical alteration and improved the neurocognitive deficit in STZ induced diabetic rats. However, the ethyl-acetate fraction demonstrated higher activities than the other fractions and was characterized for its phytoconstituents, revealing the presence of Gallic acid (713.00 ppm), catechin (0.91 ppm), ferulic acid (0.98 ppm), rutin (59.82 ppm), quercetin (3.22 ppm) and kaempferol (4.07 ppm). Our molecular docking analysis revealed that these compounds exhibited different binding affinities and potentials for targeting BChE/AChE/ IL-1 ß/Na+ -K+ -ATPase. However, only Kampferol and ferulic exhibited good drug-like, ADMET, and permeability properties suitable for use as a neuronal drug target agent. Hence, the ethyl-acetate fraction of C. pubescens leaves could be considered as a source of promising bioactive metabolite for the treatment and management of cognitive impairments related to type II diabetes mellitus.

The evolving view of thermogenic adipocytes - ontogeny, niche and function.

The worldwide incidence of obesity and its sequelae, such as type 2 diabetes mellitus, have reached pandemic levels. Central to the development of these metabolic disorders is adipose tissue. White adipose tissue stores excess energy, whereas brown adipose tissue (BAT) and beige (also known as brite) adipose tissue dissipate energy to generate heat in a process known as thermogenesis. Strategies that activate and expand BAT and beige adipose tissue increase energy expenditure in animal models and offer therapeutic promise to treat obesity. A better understanding of the molecular mechanisms underlying the development of BAT and beige adipose tissue and the activation of thermogenic function is the key to creating practical therapeutic interventions for obesity and metabolic disorders. In this Review, we discuss the regulation of the tissue microenvironment (the adipose niche) and inter-organ communication between BAT and other tissues. We also cover the activation of BAT and beige adipose tissue in response to physiological cues (such as cold exposure, exercise and diet). We highlight advances in harnessing the therapeutic potential of BAT and beige adipose tissue by genetic, pharmacological and cell-based approaches in obesity and metabolic disorders.

Therapeutic Perspectives of Thermogenic Adipocytes in Obesity and Related Complications.

There is a rapidly increasing prevalence of obesity and related metabolic disorders such as type 2 diabetes worldwide. White adipose tissue (WAT) stores excess energy, whereas brown and beige adipose tissues consume energy to generate heat in the process of thermogenesis. Adaptive thermogenesis occurs in response to environmental cues as a means of generating heat by dissipating stored chemical energy. Due to its cumulative nature, very small differences in energy expenditure from adaptive thermogenesis can have a significant impact on systemic metabolism over time. Targeting brown adipose tissue (BAT) activation and converting WAT to beige fat as a method to increase energy expenditure is one of the promising strategies to combat obesity. In this review, we discuss the activation of the thermogenic process in response to physiological conditions. We highlight recent advances in harnessing the therapeutic potential of thermogenic adipocytes by genetic, pharmacological and cell-based approaches in the treatment of obesity and metabolic disorders in mice and the human.

CRISPR-engineered human brown-like adipocytes prevent diet-induced obesity and ameliorate metabolic syndrome in mice.

Brown and brown-like beige/brite adipocytes dissipate energy and have been proposed as therapeutic targets to combat metabolic disorders. However, the therapeutic effects of cell-based therapy in humans remain unclear. Here, we created human brown-like (HUMBLE) cells by engineering human white preadipocytes using CRISPR-Cas9-SAM-gRNA to activate endogenous uncoupling protein 1 expression. Obese mice that received HUMBLE cell transplants showed a sustained improvement in glucose tolerance and insulin sensitivity, as well as increased energy expenditure. Mechanistically, increased arginine/nitric oxide (NO) metabolism in HUMBLE adipocytes promoted the production of NO that was carried by S-nitrosothiols and nitrite in red blood cells to activate endogenous brown fat and improved glucose homeostasis in recipient animals. Together, these data demonstrate the utility of using CRISPR-Cas9 technology to engineer human white adipocytes to display brown fat-like phenotypes and may open up cell-based therapeutic opportunities to combat obesity and diabetes.

Roles of Mitochondrial Sirtuins in Mitochondrial Function, Redox Homeostasis, Insulin Resistance and Type 2 Diabetes.

Mitochondria are the metabolic hubs that process a number of reactions including tricarboxylic acid cycle, ß-oxidation of fatty acids and part of the urea cycle and pyrimidine nucleotide biosynthesis. Mitochondrial dysfunction impairs redox homeostasis and metabolic adaptation, leading to aging and metabolic disorders like insulin resistance and type 2 diabetes. SIRT3, SIRT4 and SIRT5 belong to the sirtuin family proteins and are located at mitochondria and also known as mitochondrial sirtuins. They catalyze NAD+-dependent deacylation (deacetylation, demalonylation and desuccinylation) and ADP-ribosylation and modulate the function of mitochondrial targets to regulate the metabolic status in mammalian cells. Emerging evidence has revealed that mitochondrial sirtuins coordinate the regulation of gene expression and activities of a wide spectrum of enzymes to orchestrate oxidative metabolism and stress responses. Mitochondrial sirtuins act in synergistic or antagonistic manners to promote respiratory function, antioxidant defense, insulin response and adipogenesis to protect individuals from aging and aging-related metabolic abnormalities. In this review, we focus on the molecular mechanisms by which mitochondrial sirtuins regulate oxidative metabolism and antioxidant defense and discuss the roles of their deficiency in the impairment of mitochondrial function and pathogenesis of insulin resistance and type 2 diabetes.

The Perlman syndrome DIS3L2 exoribonuclease safeguards endoplasmic reticulum-targeted mRNA translation and calcium ion homeostasis.

DIS3L2-mediated decay (DMD) is a surveillance pathway for certain non-coding RNAs (ncRNAs) including ribosomal RNAs (rRNAs), transfer RNAs (tRNAs), small nuclear RNAs (snRNAs), and RMRP. While mutations in DIS3L2 are associated with Perlman syndrome, the biological significance of impaired DMD is obscure and pathological RNAs have not been identified. Here, by ribosome profiling (Ribo-seq) we find specific dysregulation of endoplasmic reticulum (ER)-targeted mRNA translation in DIS3L2-deficient cells. Mechanistically, DMD functions in the quality control of the 7SL ncRNA component of the signal recognition particle (SRP) required for ER-targeted translation. Upon DIS3L2 loss, sustained 3'-end uridylation of aberrant 7SL RNA impacts ER-targeted translation and causes ER calcium leakage. Consequently, elevated intracellular calcium in DIS3L2-deficient cells activates calcium signaling response genes and perturbs ESC differentiation. Thus, DMD is required to safeguard ER-targeted mRNA translation, intracellular calcium homeostasis, and stem cell differentiation.

Parental metabolic syndrome epigenetically reprograms offspring hepatic lipid metabolism in mice.

The prevalence of nonalcoholic fatty liver disease (NAFLD) is increasing worldwide. Although gene-environment interactions have been implicated in the etiology of several disorders, the impact of paternal and/or maternal metabolic syndrome on the clinical phenotypes of offspring and the underlying genetic and epigenetic contributors of NAFLD have not been fully explored. To this end, we used the liver-specific insulin receptor knockout (LIRKO) mouse, a unique nondietary model manifesting 3 hallmarks that confer high risk for the development of NAFLD: hyperglycemia, insulin resistance, and dyslipidemia. We report that parental metabolic syndrome epigenetically reprograms members of the TGF-ß family, including neuronal regeneration-related protein (NREP) and growth differentiation factor 15 (GDF15). NREP and GDF15 modulate the expression of several genes involved in the regulation of hepatic lipid metabolism. In particular, NREP downregulation increases the protein abundance of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) and ATP-citrate lyase (ACLY) in a TGF-ß receptor/PI3K/protein kinase B-dependent manner, to regulate hepatic acetyl-CoA and cholesterol synthesis. Reduced hepatic expression of NREP in patients with NAFLD and substantial correlations between low serum NREP levels and the presence of steatosis and nonalcoholic steatohepatitis highlight the clinical translational relevance of our findings in the context of recent preclinical trials implicating ACLY in NAFLD progression.

FGF6 and FGF9 regulate UCP1 expression independent of brown adipogenesis.

Uncoupling protein-1 (UCP1) plays a central role in energy dissipation in brown adipose tissue (BAT). Using high-throughput library screening of secreted peptides, we identify two fibroblast growth factors (FGF), FGF6 and FGF9, as potent inducers of UCP1 expression in adipocytes and preadipocytes. Surprisingly, this occurs through a mechanism independent of adipogenesis and involves FGF receptor-3 (FGFR3), prostaglandin-E2 and interaction between estrogen receptor-related alpha, flightless-1 (FLII) and leucine-rich-repeat-(in FLII)-interacting-protein-1 as a regulatory complex for UCP1 transcription. Physiologically, FGF6/9 expression in adipose is upregulated by exercise and cold in mice, and FGF9/FGFR3 expression in human neck fat is significantly associated with UCP1 expression. Loss of FGF9 impairs BAT thermogenesis. In vivo administration of FGF9 increases UCP1 expression and thermogenic capacity. Thus, FGF6 and FGF9 are adipokines that can regulate UCP1 through a transcriptional network that is dissociated from brown adipogenesis, and act to modulate systemic energy metabolism.

Analyzing Mitochondrial Function in Brown Adipocytes with a Bioenergetic Analyzer.

Brown adipocytes are a cell type with high mitochondrial content and bioenergetic capacity. A critical means to measure mitochondrial function, macromolecule fuel usage, and other important phenotypes is with a bioenergetic analyzer. Here, we describe how to isolate, culture, and differentiate brown preadipocytes into mature adipocytes. We also explain how to perform a mitochondrial (mito) stress test, using the bioenergetic analyzer. The mito stress test is able to give researchers a plethora of insights into mitochondrial function, including basal respiration, proton leak, ATP production, maximal respiration, and reserve capacity, making it a powerful tool for analyzing brown adipocytes.

Cell-autonomous light sensitivity via Opsin3 regulates fuel utilization in brown adipocytes.

Opsin3 (Opn3) is a transmembrane heptahelical G protein-coupled receptor (GPCR) with the potential to produce a nonvisual photoreceptive effect. Interestingly, anatomical profiling of GPCRs reveals that Opn3 mRNA is highly expressed in adipose tissue. The photosensitive functions of Opn3 in mammals are poorly understood, and whether Opn3 has a role in fat is entirely unknown. In this study, we found that Opn3-knockout (Opn3-KO) mice were prone to diet-induced obesity and insulin resistance. At the cellular level, Opn3-KO brown adipocytes cultured in darkness had decreased glucose uptake and lower nutrient-induced mitochondrial respiration than wild-type (WT) cells. Light exposure promoted mitochondrial activity and glucose uptake in WT adipocytes but not in Opn3-KO cells. Brown adipocytes carrying a defective mutation in Opn3's putative G protein-binding domain also exhibited a reduction in glucose uptake and mitochondrial respiration in darkness. Using RNA-sequencing, we identified several novel light-sensitive and Opn3-dependent molecular signatures in brown adipocytes. Importantly, direct exposure of brown adipose tissue (BAT) to light in living mice significantly enhanced thermogenic capacity of BAT, and this effect was diminished in Opn3-KO animals. These results uncover a previously unrecognized cell-autonomous, light-sensing mechanism in brown adipocytes via Opn3-GPCR signaling that can regulate fuel metabolism and mitochondrial respiration. Our work also provides a molecular basis for developing light-based treatments for obesity and its related metabolic disorders.

Role of p110a subunit of PI3-kinase in skeletal muscle mitochondrial homeostasis and metabolism.

Skeletal muscle insulin resistance, decreased phosphatidylinositol 3-kinase (PI3K) activation and altered mitochondrial function are hallmarks of type 2 diabetes. To determine the relationship between these abnormalities, we created mice with muscle-specific knockout of the p110α or p110ß catalytic subunits of PI3K. We find that mice with muscle-specific knockout of p110α, but not p110ß, display impaired insulin signaling and reduced muscle size due to enhanced proteasomal and autophagic activity. Despite insulin resistance and muscle atrophy, M-p110αKO mice show decreased serum myostatin, increased mitochondrial mass, increased mitochondrial fusion, and increased PGC1α expression, especially PCG1α2 and PCG1α3. This leads to enhanced mitochondrial oxidative capacity, increased muscle NADH content, and higher muscle free radical release measured in vivo using pMitoTimer reporter. Thus, p110α is the dominant catalytic isoform of PI3K in muscle in control of insulin sensitivity and muscle mass, and has a unique role in mitochondrial homeostasis in skeletal muscle.