MicroRNA-721 regulates gluconeogenesis via KDM2A-mediated epigenetic modulation in diet-induced insulin resistance in C57BL/6J mice.

BACKGROUND: Aberrant gluconeogenesis is considered among primary drivers of hyperglycemia under insulin resistant conditions, with multiple studies pointing towards epigenetic dysregulation. Here we examine the role of miR-721 and effect of epigenetic modulator laccaic acid on the regulation of gluconeogenesis under high fat diet induced insulin resistance. RESULTS: Reanalysis of miRNA profiling data of high-fat diet-induced insulin-resistant mice model, GEO dataset (GSE94799) revealed a significant upregulation of miR-721, which was further validated in invivo insulin resistance in mice and invitro insulin resistance in Hepa 1-6 cells. Interestingly, miR-721 mimic increased glucose production in Hepa 1-6 cells via activation of FOXO1 regulated gluconeogenic program. Concomitantly, inhibition of miR-721 reduced glucose production in palmitate induced insulin resistant Hepa 1-6 cells by blunting the FOXO1 induced gluconeogenesis. Intriguingly, at epigenetic level, enrichment of the transcriptional activation mark H3K36me2 got decreased around the FOXO1 promoter. Additionally, identifying targets of miR-721 using miRDB.org showed H3K36me2 demethylase KDM2A as a potential target. Notably, miR-721 inhibitor enhanced KDM2A expression which correlated with H3K36me2 enrichment around FOXO1 promoter and the downstream activation of the gluconeogenic pathway. Furthermore, inhibition of miR-721 in high-fat diet-induced insulin-resistant mice resulted in restoration of KDM2A levels, concomitantly reducing FOXO1, PCK1, and G6PC expression, attenuating gluconeogenesis, hyperglycemia, and improving glucose tolerance. Interestingly, the epigenetic modulator laccaic acid also reduced the hepatic miR-721 expression and improved KDM2A expression, supporting our earlier report that laccaic acid attenuates insulin resistance by reducing gluconeogenesis. CONCLUSION: Our study unveils the role of miR-721 in regulating gluconeogenesis through KDM2A and FOXO1 under insulin resistance, pointing towards significant clinical and therapeutic implications for metabolic disorders. Moreover, the promising impact of laccaic acid highlights its potential as a valuable intervention in managing insulin resistance-associated metabolic diseases.

Wound-healing effect of topical nanoemulsion-loaded cream and gel formulations of Hippophae rhamnoides L. (sea buckthorn) fruit oil and their acute dermal toxicity study on female SD rats.

OBJECTIVE: The objective of this study was to evaluate the efficacy and safety of topical nanoemulsion (NE)-loaded cream and gel formulations of Hippophae rhamnoides L. (sea buckthorn [SBT]) fruit oil for wound healing. MATERIALS AND METHODS: The NE-loaded cream and gel formulations of H. rhamnoides L. (SBT) fruit oil (IPHRFH) were prepared and evaluated for their wound-healing activity on female Sprague-Dawley (SD) rats. They were further divided into groups (seven) and the wound-healing activity was determined by measuring the area of the wound on the wounding day and on the 0th, 4th, 8th, and 10th days. The acute dermal toxicity of the formulations was assessed by observing the erythema, edema, and body weight (BW) of the rats. RESULTS: The topical NE cream and gel formulations of H. rhamnoides L. (SBT) fruit oil showed significant wound-healing activity in female SD rats. The cream formulation of IPHRFH showed 78.96%, the gel showed 72.59% wound contraction on the 8th day, whereas the positive control soframycin (1% w/w framycetin) had 62.29% wound contraction on the 8th day. The formulations also showed a good acute dermal toxicity profile with no changes significantly affecting BW and dermal alterations. CONCLUSIONS: The results of this study indicate that topical NE-loaded cream and gel formulation of H. rhamnoides L. (SBT) fruit oil are safe and effective for wound healing. The formulations showed no signs of acute dermal toxicity in female SD rats.

Adipose tissue macrophage-derived microRNA-210-3p disrupts systemic insulin sensitivity by silencing GLUT4 in obesity.

Management of chronic obesity-associated metabolic disorders is a key challenge for biomedical researchers. During chronic obesity, visceral adipose tissue (VAT) undergoes substantial transformation characterized by a unique lipid-rich hypoxic AT microenvironment which plays a crucial role in VAT dysfunction, leading to insulin resistance (IR) and type 2 diabetes. Here, we demonstrate that obese AT microenvironment triggers the release of miR-210-3p microRNA-loaded extracellular vesicles from adipose tissue macrophages, which disseminate miR-210-3p to neighboring adipocytes, skeletal muscle cells, and hepatocytes through paracrine and endocrine actions, thereby influencing insulin sensitivity. Moreover, EVs collected from Dicer-silenced miR-210-3p-overexpressed bone marrow-derived macrophages induce glucose intolerance and IR in lean mice. Mechanistically, miR-210-3p interacts with the 3'-UTR of GLUT4 mRNA and silences its expression, compromising cellular glucose uptake and insulin sensitivity. Therapeutic inhibition of miR-210-3p in VAT notably rescues high-fat diet-fed mice from obesity-induced systemic glucose intolerance. Thus, targeting adipose tissue macrophage-specific miR-210-3p during obesity could be a promising strategy for managing IR and type 2 diabetes.

L-Methionine accentuates anti-tumor action of Gefitinib in Gefitinib-resistant lung adenocarcinoma: Role of EGFR/ERK/AKT signaling and histone H3K36me2 alteration.

Adenocarcinoma, the predominant subtype of non-small cell lung cancer (NSCLC), poses a significant clinical challenge due to its prevalence and aggressive nature. Gefitinib, an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor is often susceptible to development of resistance despite being the preferred treatment option for NSCLC. In this study, we investigated the potential of L-Methionine in enhancing the cytotoxicity of Gefitinib and preventing resistance development. In vitro experiment employing the H1975 cell line demonstrated a notable enhancement in cytotoxic efficacy when L-Methionine (10 mM) was combined with Gefitinib, as indicated by a substantial reduction in IC50 values (155.854 ± 1.87 µM vs 45.83 ± 4.83 µM). Complementary in vivo investigations in a lung cancer model corroborated these findings. Co-administration of L-Methionine (100 mg/kg and 400 mg/kg) with Gefitinib (15 mg/kg) for 21 days exhibited marked improvements in therapeutic efficacy, which was observed by macroscopic and histopathological assessments. Mechanistic insights revealed that the enhanced cytotoxicity of the combination stemmed from the inhibition of the EGFR, modulating the downstream cascade of ERK/AKT and AMPK pathways. Concurrently inhibition of p-AMPK-α by the combination also disrupted metabolic homeostasis, leading to the increased production of reactive oxygen species (ROS). Notably, L-Methionine, functioning as a methyl group donor, elevated the expression of H3K36me2 (an activation mark), while reducing the p-ERK activity. Our study provides the first evidence supporting L-Methionine supplementation as a novel strategy to enhance Gefitinib chemosensitivity against pulmonary adenocarcinoma.

Synthesis and mechanistic study of ultrashort peptides that inhibits Alzheimer's Aß-aggregation-induced neurotoxicity.

Misfolding/aggregation of ß-amyloid peptide lead to the formation of toxic oligomers or accumulation of amyloid plaques, which is a seminal step in the progression of Alzheimer's disease (AD). Despite continuous efforts in the development of therapeutic agents, the cure for AD remains a major challenge. Owing to specific binding affinity of structure-based peptides, we report the synthesis of new peptide-based inhibitors derived from the C-terminal sequences, Aß38-40 and Aß40-42. Preliminary screening using MTT cell viability assay and corroborative results from ThT fluorescence assay revealed a tripeptide showing significantly effective inhibition towards Aß1-42 aggregation and induced toxicity. Peptide 3 exhibited excellent cell viability of 94.3 % at 2 µM and of 100 % at 4 µM and 10 µM. CD study showed that peptide 3 restrict the conformation transition of Aß1-42 peptide towards cross-ß-sheet structure and electron microscopy validated the absence of Aß aggregates as indicated by the altered morphology of Aß1-42 in the presence of peptide 3. The HRMS-ESI, DLS and ANS studies were performed to gain mechanistic insights into the effect of inhibitor against Aß aggregation. This Aß-derived ultrashort motif provides impetus for the development of peptide-based anti-AD agents.

l-Methionine potentiates anticancer activity of Sorafenib by epigenetically altering DUSP3/ERK pathway in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the third most common cancer-related cause of death worldwide. Although Sorafenib is the standard systemic therapy for treating HCC, but it develops resistance very quickly, leading to poor prognosis. The current study was planned to explore the effect of l-methionine on the anticancer activity of Sorafenib in HCC. Ten millimolar of l-methionine treatment significantly reduced the IC50 of Sorafenib from 5.513 ± 0.171 to 0.8095 ± 0.0465 µM in HepG2 cell line. It also resulted in concomitant increase in oxidative stress and deactivation of ERK/AMPK/AKT pathway. Additionally, it also resulted in the increased expression of dual specificity phosphatase 3 (DUSP3). In a rat model of sorafenib-resistant HCC induced by diethylnitrosamine (DEN) (100 mg/L/day) and Sorafenib (10 mg/kg), l-methionine (300 and 500 mg/kg/day) supplementation overcame the drug resistance, as indicated by the reduced formation of surface tumor nodules, prevention of cellular hypertrophy, hyperplasia and inflammation, and improved animal survival. Furthermore, l-methionine in combination with Sorafenib also inhibited AMPK/AKT and ERK pathway. At chromatin level, l-methionine supplementation prevented global methylation of H3K27me3, an inactivation mark, and demethylation of H3K36me2, an activation mark. Interestingly, our findings suggest that inhibition of the ERK pathway via increased activity of DUSP3 is epigenetically regulated. Besides, chromatin immunoprecipitation data exhibited augmented H3K36me2 (an activation mark) levels on the DUSP3 promoter region. To the best of our knowledge, we are the first to report that l-methionine supplementation improves the chemosensitivity in Sorafenib-resistant HCC via modulating the epigenetic landscape and can be a potential therapeutic strategy.

Comparative quantitative analysis of fruit oil from Hippophae rhamnoides (seabuckthorn) by qNMR, FTIR and GC-MS.

Objective: To develop a qNMR method for quantitative analysis of triacylglycerols in fruit oil of Hippophae rhamnoides (seabuckthorn, SBT) and analyze commercial samples of SBT oils using GC-MS and FTIR. Methods: SBT fruit oil (IPHRFH) was extracted with hexane and the triglyceride (TAG) was isolated by vacuum liquid chromatography. Six different branded SBT oils purchased from e-commerce suppliers (Amazon) and in-house prepared SBT oil was analyzed by qNMR and fatty acyl composition of TAGs determined by using NMR. In-house oil was also analysed by GC-MS and FTIR spectroscopy. Results: The qNMR results showed that the oil contained 80.3% of triacylglycerol (TAG). The SBT oil TAGs comprised of linolenate 6.6%, palmitoleate/oleate 65.4%, and total saturated fatty acyl chain including palmitate 28% as determined by qNMR. GC-MS analysis revealed that the major acyl functionalities present in the TAG were palmitoleic acid 36.5%, oleic acid 12.9%, palmitic acid 21.2%, and linoleic acid 18%. Of the six commercial samples analyzed, samples from only one supplier (SW) were fruit oil; All others were the seed oils or mix of fruit oil and seed oil. The labels for samples except for the SW did not indicate whether it was fruit oil or seed oil. Conclusion: The results suggest that SBT oil should be analyzed by combination of GC-MS, FTIR and qNMR for factual content of free fatty acid or TAGs, which are chemically different in nature and affect the quality of oil. GC-MS showed the content of omega free fatty acids after hydrolysis, while qNMR and FTIR showed the content of TAGs. The major acyl functionalities found in SBT fruit oil TAGs are palmitoleate/palmitate/oleate, while linoleate and linonelate make up a minor fraction. Furthermore, analysis of commercial samples showed discrepancies between label claims and actual content.

Development and characterization of fused human arginase I for cancer therapy.

Recombinant human arginase I (rhArg I) have emerged as a potential candidate for the treatment of varied pathophysiological conditions ranging from arginine-auxotrophic cancer, inflammatory conditions and microbial infection. However, rhArg I have a low circulatory half-life, leading to poor pharmacokinetic and pharmacodynamic properties, which necessitating the rapid development of modifications to circumvent these limitations. To address this, polyethylene glycol (PEG)ylated-rhArg I variants are being developed by pharmaceutical companies. However, because of the limitations associated with the clinical use of PEGylated proteins, there is a dire need in the art to develop rhArg I variant(s) which is safe (devoid of limitations of PEGylated counterpart) and possess increased circulatory half-life. In this study, we described the generation and characterization of a fused human arginase I variant (FHA-3) having improved circulatory half-life. FHA-3 protein was engineered by fusing rhArg I with a half-life extension partner (domain of human serum albumin) via a peptide linker and was produced using P. pastoris expression system. This purified biopharmaceutical (FHA-3) exhibits (i) increased arginine-hydrolyzing activity in buffer, (ii) cofactor - independency, (iii) increased circulatory half-life (t1/2) and (iv) potent anti-cancer activity against human cancer cell lines under in vitro and in vivo conditions.

Target validation and structure-based virtual screening to Discover potential lead molecules against the oncogenic NSD1 histone methyltransferase.

The aim of the study was to validate Nuclear receptor-binding SET Domain NSD1 as a cancer drug target followed by the design of lead molecules against NSD1. TCGA clinical data, molecular expression techniques were used to validate the target and structure-based virtual screening was performed to design hits against NSD1. Clinical data analysis suggests the role of NSD1 in metastasis, prognosis and influence on overall survival in various malignancies. Furthermore, the mRNA and protein expression profile of NSD1 was evaluated in various cell lines. NSD1 was exploited as a target protein for in silico design of inhibitors using two major databases including ZINC15 and ChemDiv by structure-based virtual screening approach. Virtual screening was performed using the pharmacophore hypothesis designed with a protein complex S-adenosyl-l-methionine (SAM) as an endogenous ligand. Subsequently, a combined score was used to distinguish the top 10 compounds from the docking screened compounds having high performance in all four scores (docking score, XP, Gscore, PhaseScreenScore, and MMGBSA delta G Bind). Finally, the top three Zinc compounds were subjected to molecular dynamic simulation. The binding MMGBSA data suggests that ZINC000257261703 and ZINC000012405780 can be taken for in vitro and in vivo studies as they have lesser MMGBSA energy towards the cofactor binding site of NSD1 than the sinefungin. Our data validates NSD1 as a cancer drug target and provides promising structures that can be utilized for further lead optimization and rational drug design to open new gateways in the field of cancer therapeutics. Supplementary Information: The online version contains supplementary material available at 10.1007/s40203-023-00158-0.

Type 2 diabetes alters the viscoelastic behavior and macromolecular composition of vertebra.

Type 2 diabetes (T2D) affects the functional behavior of vertebra bone by altering its structural and mechanical properties. The vertebral bones are responsible to carry the body weight and it remains under prolonged constant load which results to viscoelastic deformation. The effect of T2D on the viscoelastic behavior of vertebral bone is not well explored yet. In this study, the effects of T2D on the creep and stress relaxation behavior of vertebral bone are investigated. Also, this study established a correlation between T2D associated alteration in macromolecular structure and viscoelastic behavior of vertebra. In this study T2D female rat SD model was used. The obtained results demonstrated a significant reduction in the amount of creep strain (p ≤ 0.05) and stress relaxation (p ≤ 0.01) in T2D specimens than the control. Also, the creep rate was found significantly lower in T2D specimens. On the other hand, molecular structural parameters such as mineral-to-matrix ratio (control vs T2D: 2.93 ± 0.78 vs 3.72 ± 0.53; p = 0.02), and non-enzymatic cross link ratio (NE-xL) (control vs T2D: 1.53 ± 0.07 vs 3.84 ± 0.20; p = 0.01) were found significantly altered in T2D specimens. Pearson linear correlation tests show a significant correlation; between creep rate and NE-xL (r = -0.94, p < 0.01), and between stress relaxation and NE-xL (r = -0.946, p < 0.01). Overall this study explored the understanding about the disease associated alteration in viscoelastic response of vertebra and its correlation with macromolecular composition which can help to understand the disease related impaired functioning of the vertebrae body.

Pharmacological inhibition of DNMT1 restores macrophage autophagy and M2 polarization in Western diet-induced nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) is associated with an increased ratio of classically activated M1 macrophages/Kupffer cells to alternatively activated M2 macrophages, which plays an imperative role in the development and progression of NAFLD. However, little is known about the precise mechanism behind macrophage polarization shift. Here, we provide evidence regarding the relationship between the polarization shift in Kupffer cells and autophagy resulting from lipid exposure. High-fat and high-fructose diet supplementation for 10 weeks significantly increased the abundance of Kupffer cells with an M1-predominant phenotype in mice. Interestingly, at the molecular level, we also observed a concomitant increase in expression of DNA methyltransferases DNMT1 and reduced autophagy in the NAFLD mice. We also observed hypermethylation at the promotor regions of autophagy genes (LC3B, ATG-5, and ATG-7). Furthermore, the pharmacological inhibition of DNMT1 by using DNA hypomethylating agents (azacitidine and zebularine) restored Kupffer cell autophagy, M1/M2 polarization, and therefore prevented the progression of NAFLD. We report the presence of a link between epigenetic regulation of autophagy gene and macrophage polarization switch. We provide the evidence that epigenetic modulators restore the lipid-induced imbalance in macrophage polarization, therefore preventing the development and progression of NAFLD.

Synthesis, biological evaluation and mechanistic studies of 4-(1,3-thiazol-2-yl)morpholine-benzimidazole hybrids as a new structural class of antimicrobials.

In spite of several attempts to develop newer pharmacophores as potential antimicrobial agents, the benzimidazole scaffold is still considered as one of the most sought after structural component towards the design of compounds that act against a wide spectrum of microbes. Herein, we report the design and synthesis of a new structural class of 4-(1,3-thiazol-2-yl)morpholine-benzimidazole hybrids as antimicrobial agents. The most potent analog, 6g shows IC50 of 1.3 µM, 2.7 µM, 10.8 µM, 5.4 µM and 10.8 µM against Cryptococcus neoformans, Candida albicans, Candida parapsilosis, Escherichia coli and Staphylococcus aureus, respectively. Interestingly 6g exhibits selectivity towards the cryptococcal cells with fungicidal behavior. Propidium iodide uptake study shows permeabilization of pathogenic cells in the presence of 6g. Flow cytometric analysis confirms that cell death is predominantly due to apoptosis. Moreover, electron microscopic analysis specifies that it shrinks, disrupts and initiate pore(s) formation in the cell membrane leading to cell lysis.

Laccaic acid restores epigenetic alterations responsible for high fat diet induced insulin resistance in C57BL/6J mice.

Laccaic acid, the major constituent of the food colouring agent-lac dye, possesses antioxidant and anti-inflammatory properties. Here we have evaluated the effects of laccaic acid on the high-fat diet induced insulin resistance in C57BL/6J mice. Insulin resistance was developed in mice by feeding high-fat diet for 12 weeks. 6 week treatment with laccaic acid showed significant improvement in the morphometric, biochemical parameters and liver function. Western blotting experiments showed, laccaic acid increased phosphorylation of IRS1/2/AKT/GSK3ß which is suppressed under insulin-resistant conditions in liver. Furthermore, it also attenuated the inflammatory ERK/NFκB signalling, thereby reducing the expression of inflammatory cytokines- TNFα, IL-1ß and IL-6. Concomitantly, laccaic acid increased AMPK/AKT-mediated phosphorylation of FOXO1, preventing its nuclear translocation and transcriptional activation of gluconeogenic genes (G6PC and PCK1). Interestingly, treatment with laccaic acid also prevented high-fat diet induced alterations of histone methylation (H3K27me3 and H3K36me2) at global level. Our chromatin-immunoprecipitation data shows high-fat diet induced loss of inactivation mark H3K27me3 at FOXO1 promoter was regained upon laccaic acid treatment. Additionally, the expression of the H3K27 methylating enzyme EZH2 was also upregulated by laccaic acid. Together it all results in the downregulation of FOXO1 gene expression. To the best of our knowledge, we provide first evidence that laccaic acid either directly or indirectly modulates the epigenetic landscape of genes responsible for high-fat diet induced insulin resistance.

A small molecule potent IRAK4 inhibitor abrogates lipopolysaccharide-induced macrophage inflammation in-vitro and in-vivo.

Increasing evidence supports vanillin and its analogs as potent toll-like receptor signaling inhibitors that strongly attenuate inflammation, though, the underlying molecular mechanism remains elusive. Here, we report that vanillin inhibits lipopolysaccharide (LPS)-induced toll-like receptor 4 activation in macrophages by targeting the myeloid differentiation primary-response gene 88 (MyD88)-dependent pathway through direct interaction and suppression of interleukin-1 receptor-associated kinase 4 (IRAK4) activity. Moreover, incubation of vanillin in cells expressing constitutively active forms of different toll-like receptor 4 signaling molecules revealed that vanillin could only able to block the ligand-independent constitutively activated IRAK4/1 or its upstream molecules-associated NF-κB activation and NF-κB transactivation along with the expression of various proinflammatory cytokines. A significant inhibition of LPS-induced IRAK4/MyD88, IRAK4/IRAK1, and IRAK1/TRAF6 association was evinced in response to vanillin treatment. Furthermore, mutations at Tyr262 and Asp329 residues in IRAK4 or modifications of 3-OMe and 4-OH side groups in vanillin, significantly reduced IRAK4 activity and vanillin function, respectively. Mice pretreated with vanillin followed by LPS challenge markedly impaired LPS-induced IRAK4 activation and inflammation in peritoneal macrophages. Thus, the present study posits vanillin as a novel and potent IRAK4 inhibitor and thus providing an opportunity for its therapeutic application in managing various inflammatory diseases.

Synthetic amino acids-based short amphipathic peptides exhibit antifungal activity by targeting cell membrane disruption.

Availability of a limited number of antifungal drugs created a necessity to develop new antifungals with distinct mode of action. Investigation on a new series of peptides led us to identify Boc-His-Trp-His[1-(4-tert-butylphenyl)] (10g) as the most promising inhibitor exhibiting IC50 value of 4.4 µg/mL against Cryptococcus neoformans. Analog 10g exhibit high selectivity to fungal cells and was nonhemolytic and noncytotoxic at its minimum inhibitory concentration. 10g produced fungicidal effect on growing cryptococcal cells and displayed synergistic effect with amphotericin B. Overall cationic character of 10g resulted in interaction with negatively charged fungal membrane while hydrophobicity enhanced penetration inside the cryptococcal cells causing hole(s) formation and disruption to the membrane as evident by the scanning electron microscopy, transmission electron microscopy, and confocal laser scanning microscopy analyses. Flow cytometric investigation revealed rapid death of fungal cells by apopotic pathway.

Anticryptococcal activity and mechanistic studies of short amphipathic peptides.

Cryptococcus neoformans, an opportunistic fungal pathogen, causes cryptococcosis in immunocompromised persons. A series of modified L-histidines-containing peptides are synthesized that exhibit promising activity against C. neoformans. Analog 11d [L-His(2-adamantyl)-L-Trp-L-His(2-phenyl)-OMe] produced potency with an IC50 of 3.02 µg/ml (MIC = 5.49 µg/ml). This peptide is noncytotoxic and nonhaemolytic at the MIC and displays synergistic effects with amphotericin B at subinhibitory concentration. Mechanistic investigation of 11d using microscopic tools indicates cell wall and membrane disruption of C. neoformans, while flow cytometric analysis confirms cell death by apoptosis. This study indicates that 11d exhibits antifungal potential and acts via the rapid onset of action.

Appraising the electrocatalytic performance of beta-cyclodextrin embellished supramolecular recognition system for pernicious food colorants.

The current research presents the evaluation of supramolecular proficiency of the designed platform for electrocatalytic determination of pernicious food colorants, amaranth and fast green. The approach involving surface modification of glassy carbon electrode with beta cyclodextrin decorated strontium ferrite reduced graphene oxide nanocomposite (SFrGO-ßCD) to ensure fast and reversible electro-oxidation of hydroxyl groups of the colorant molecules. The synergy between SF and rGO facilitated the sensor with enhanced surface area and conductivity through faradic redox reaction. Tremendous decrease in the obtained values of peak separation potential and impedance as manifested in CV and EIS analysis, enabled by electrostatic interactions between surface functionalities of rGO and ßCD has resulted in the significant augmentation of sensitivity. The value of charge transfer coefficient, number of electrons involved, nature of electron transport process at electrode electrolyte interface during the analysis of electrochemical detection were explored through CV experiments. Food samples analysis (without spiking) utilizing screen printed electrode manifested the sensor as portable device for real time monitoring. Outstanding detection limit (0.022 nM for amaranth and 0.051 nM for fast green), excellent regenerability (Relative standard deviation less than 3%) and apparent recovery rate (above 90%) of the modified electrode presented a colossal potential for the development of sustainable and commercially competitive electrochemical sensor in food sector.

Selenium-Enriched Probiotic Alleviates Western Diet-Induced Non-alcoholic Fatty Liver Disease in Rats via Modulation of Autophagy Through AMPK/SIRT-1 Pathway.

Current study was aimed to investigate the ability of L.acidophilus SNZ 86 to biotransform inorganic selenium to a more active organic form, resulting in trace element enrichment. Selenium-enriched L. acidophilus SNZ 86 has been shown to be effective in the treatment of a variety of gastrointestinal illnesses, indicating the need for additional research to determine the full potential of this therapeutic strategy in the treatment of metabolic disorders. Herein, we employed the western style diet-induced model of non-alcoholic fatty liver disease (NAFLD) to explore the therapeutic effect of selenium-enriched probiotic (SP). Male Sprague Dawley rats (160-180 g) were fed a high-fat (58% Kcal of fat) and high-fructose (30% w/v) diet for 12 weeks to develop an animal model mimicking NAFLD. High-fat and High-fructose diet-fed rats exhibited hyperglycemia, hyperlipidemia, insulin resistance, abnormal liver function test, increased hepatic oxidative stress, and steatosis. SP was then administered orally (L acidophilus 1 × 109 CFU/ml containing 0.4 g Se/day; p.o.) for 8 weeks. The selenium enrichment within L. acidophilus SNZ 86 was validated by TEM, which allowed for visualisation of the selenium deposition and size distribution in the probiotic. In NAFLD control rats, the expression of autophagy proteins (LC-3 A/B and Beclin), AMPK, and SIRT-1 was significantly reduced indicating downregulation of autophagy. However, supplementation of SP ameliorates hepatic steatosis as evidenced by improved biochemical markers and autophagic activation via upregulation of the AMPK and SIRT-1 pathway showing the relevance of autophagy in the disease aetiology. Collectively, these findings provide us with a better understanding of the role of SP in the treatment of hepatic steatosis and establish a therapeutic basis for potential clinical application of SP in the prevention of NAFLD and associated pathological conditions.

miR-210-3p Promotes Obesity-Induced Adipose Tissue Inflammation and Insulin Resistance by Targeting SOCS1-Mediated NF-κB Pathway.

Under the condition of chronic obesity, an increased level of free fatty acids along with low oxygen tension in the adipose tissue creates a pathophysiological adipose tissue microenvironment (ATenv), leading to the impairment of adipocyte function and insulin resistance. Here, we found the synergistic effect of hypoxia and lipid (H + L) surge in fostering adipose tissue macrophage (ATM) inflammation and polarization. ATenv significantly increased miR-210-3p expression in ATMs which promotes NF-κB activation-dependent proinflammatory cytokine expression along with the downregulation of anti-inflammatory cytokine expression. Interestingly, delivery of miR-210-3p mimic significantly increased macrophage inflammation in the absence of H + L co-stimulation, while miR-210-3p inhibitor notably compromised H + L-induced macrophage inflammation through increased production of suppressor of cytokine signaling 1 (SOCS1), a negative regulator of the NF-κB inflammatory signaling pathway. Mechanistically, miR-210 directly binds to the 3'-UTR of SOCS1 mRNA and silences its expression, thus preventing proteasomal degradation of NF-κB p65. Direct delivery of anti-miR-210-3p LNA in the ATenv markedly rescued mice from obesity-induced adipose tissue inflammation and insulin resistance. Thus, miR-210-3p inhibition in ATMs could serve as a novel therapeutic strategy for managing obesity-induced type 2 diabetes.

The multiscale characterization and constitutive modeling of healthy and type 2 diabetes mellitus Sprague Dawley rat skin.

In type 2 diabetes mellitus (T2DM), elevated glucose level impairs the biochemistry of the skin which may result in alteration of its mechanical and structural properties. The several aspects of structural and mechanical changes in skin due to T2DM remain poorly understood. To fill these research gaps, we developed a non-obese T2DM rat (Sprague Dawley (SD)) model for investigating the effect of T2DM on the in vivo strain stress state, mechanical and structural properties of skin. In vivo strain and mechanical anisotropy of healthy and T2DM skin were measured using the digital imaging correlation (DIC) technique and DIC coupled bulge experiment, respectively. Fluorescence microscopy and histology were used to assess the collagen and elastin fibers microstructure whereas nanoscale structure was captured through atomic force microscopy (AFM). Based on the microstructural observations, skin was modeled as a multilayer membrane where in and out of plane distribution of collagen fibers and planar distribution of elastin fibers were cast in constitutive model. Further, the state of in vivo stresses of healthy and T2DM were measured using model parameters and in vivo strain in the constitutive model. The results showed that T2DM causes significant loss in in vivo stresses (p < 0.01) and increase in anisotropy (p < 0.001) of skin. These changes were found in good correlation with T2DM associated alteration in skin microstructure. Statistical analysis emphasized that increase in blood glucose concentration (HbA1c) was the main cause of impaired biomechanical properties of skin. The presented data in this study can help to understand the skin pathology and to simulate the skin related clinical procedures. STATEMENT OF SIGNIFICANCE: Our study is significant as it presents findings related to the effect of T2DM on the physiologic stress strain, structural and mechanical response of SD rat skin. In this study, we developed a non-obese T2DM SD rat model which mimics the phenotype of Asian type 2 diabetics (non-obese). Several structural and mechanical characterization techniques were explored for multiscale characterization of healthy and T2DM skin. Further, based on microstructural information, we presented the constitutive models that incorporate the real microstructure of skin. The presented results can be helpful to simulate the realistic mechanical response of skin during various clinical trials.