Search Results (22)

Disorders of vesicular trafficking and genetic defects in autophagy play a critical role in the development of metabolic and neurometabolic diseases. These processes govern intracellular transport and lysosomal degradation, thereby maintaining cellular homeostasis. In this article, we present two siblings with a novel homozygous variant in VPS51 (Vacuolar protein sorting 51) gene (c.1511C>T; p.Thr504Met), exhibiting developmental delay, a thin corpus callosum, severe intellectual disability, epilepsy, microcephaly, hearing loss, and dysphagia. This study aimed to investigate the effects of the novel VPS51 gene variation at the RNA and protein level in fibroblasts derived from patients. A comparative proteomic analysis, which has not been previously elucidated, was performed to identify uncharacterized proteins associated with vesicular trafficking. Furthermore, the impact of disrupted pathways on mitochondria–lysosome contact sites was assessed, offering a thorough pathophysiological evaluation of GARP/EARP (Golgi Associated Retrograde Protein / Endosome Associated Retrograde Protein) complex dysfunction. An analysis of mRNA expression indicated decreased levels of the VPS51 gene, alongside modifications in the expression of autophagy-related genes (LC3B, p62, RAB7A, TBC1D15). Western blotting demonstrated a reduction in VPS51 and autophagy-related protein levels. Proteomic profiling revealed 585 differentially expressed proteins, indicating disruptions in vesicular trafficking, lysosomal function, and mitochondrial metabolism. Proteins involved in mitochondrial β-oxidation and oxidative phosphorylation exhibited downregulation, whereas pathways related to glycolysis and lipid synthesis showed upregulation. Live-cell confocal microscopy revealed a notable increase in mitochondria–lysosome contact sites in patient fibroblasts, suggesting that VPS51 protein dysfunction contributes to impaired organelle communication. The findings indicate that the novel VPS51 gene variation influences intracellular transport, autophagy, and metabolic pathways, offering new insights into its involvement in neurometabolic disorders. Full article

Benzene, a well-known carcinogenic airborne pollutant, poses significant health risks, particularly in industries such as petroleum, shoemaking, and painting. Despite strict regulations, chronic occupational exposure persists, contributing to the onset of acute myeloid leukemia (AML) and other malignancies. Benzene’s carcinogenicity stems from its metabolic activation, leading to increased oxidative stress, DNA damage, and cancer transformation. While its toxicity is well-documented, the link between genetic and epigenetic alterations and cancer susceptibility in exposed workers remains underexplored. This study aims to identify early biomarkers of benzene exposure and AML risk by analyzing gene expression and DNA methylation datasets from GEO DataSets, integrated with molecular pathway analyses, as well as miRNA-target and protein-protein network evaluations. This multi-approach led to the identification of nine deregulated genes (CRK, CXCR6, GSPT1, KPNA1, MECP2, MELTF, NFKB1, TBC1D7, ZNF331) in workers exposed to benzene, with NFKB1 showing strong discriminatory potential. Also, dose-dependent DNA methylation changes were observed in CXCR6 and MELTF, while selected miRNAs such as let-7d-5p, miR-126-3p, and miR-361-5p emerged as key post-transcriptional regulators. Furthermore, functional enrichment linked these genes to immune response, inflammation, cell proliferation, and apoptosis pathways. While network analyses highlighted NFKB1, CRK, and CXCR6 as central to benzene-associated leukemogenesis. Altogether, these findings provide novel insights into an early biomarker fingerprint for benzene exposure and AML susceptibility, supporting the future development of biomolecular-based targeted occupational health monitoring and personalized preventive strategies for at-risk workers. Full article

Ramon syndrome (MIM 266270) is an extremely rare genetic syndrome, characterized by gingival fibromatosis, cherubism-like lesions, epilepsy, intellectual disability, hypertrichosis, short stature, juvenile rheumatoid arthritis, and ocular abnormalities. Hereditary or non-syndromic gingival fibromatosis (HGF) is also rare and considered to represent a heterogeneous group of disorders characterized by benign, slowly progressive, non-inflammatory gingival overgrowth. To date, two genes, ELMO2 and TBC1D2B, have been linked to Ramon syndrome. The objective of this study was to further investigate the genetic variants associated with Ramon syndrome as well as HGF. Clinical, radiographic, histological, and immunohistochemical examinations were performed on affected individuals. Exome sequencing identified rare variants in TBC1D2B in both conditions: a novel homozygous variant (c.1879_1880del, p.Glu627LysfsTer61) in a Thai patient with Ramon syndrome and a rare heterozygous variant (c.2471A>G, p.Tyr824Cys) in a Cambodian family with HGF. A novel variant (c.892C>T, p.Arg298Cys) in KREMEN2 was also identified in the individuals with HGF. With support from mutant protein modeling, our data suggest that TBC1D2B variants contribute to both Ramon syndrome and HGF, although variants in additional genes might also contribute to the pathogenesis of HGF. Full article

Objective: Circulating exosome-enriched extracellular vesicles (EVs) have drawn considerable importance in obesity-related insulin-resistance (IR). We sought to compare the proteomics profile of serum exosomes from normal individuals and those with obesity and IR. Methods: We isolated serum exosomes from male subjects with obesity and insulin resistance (Ob-IR, HOMA-IR > 2.0) and lean/overweight insulin-sensitive (Normal (N), HOMA-IR < 2.0) individuals. The differential protein expression between the two groups was detected by a label-free quantitative mass spectrometry analysis followed by GO annotation and ingenuity pathway analysis (IPA). Results: We identified 23 upregulated and 46 downregulated proteins between Ob-IR and N groups. Some of these proteins are involved in altering insulin signaling (VPS13C, TBC1D32, TTR, and ADIPOQ), inflammation (NFκB and CRP), and B-cell proliferation/activation (IGLV4-69, IGKV1D-13, and IGHV4-28). GO analysis revealed that the differentially expressed proteins (DEPs) are mainly involved in regulating immune cell activation and are located in extracellular space. IPA analysis showed that top molecules mediating IR, inflammation and B-cell activation were upregulated in Ob-IR subjects compared to N subjects. Conclusions: Serum exosomal proteins can be used as biomarkers to identify the future risk of diabetes and a therapeutic target to prevent or slow down the progression of diabetes in high-risk individuals. Full article

Impaired skeletal muscle glucose uptake is a key feature in the development of insulin resistance and type 2 diabetes. Skeletal muscle glucose uptake can be enhanced by a variety of different stimuli, including insulin and contraction as the most prominent. In contrast to the clearance of glucose from the bloodstream in response to insulin stimulation, exercise-induced glucose uptake into skeletal muscle is unaffected during the progression of insulin resistance, placing physical activity at the center of prevention and treatment of metabolic diseases. The two Rab GTPase-activating proteins (RabGAPs), TBC1D1 and TBC1D4, represent critical nodes at the convergence of insulin- and exercise-stimulated signaling pathways, as phosphorylation of the two closely related signaling factors leads to enhanced translocation of glucose transporter 4 (GLUT4) to the plasma membrane, resulting in increased cellular glucose uptake. However, the full network of intracellular signaling pathways that control exercise-induced glucose uptake and that overlap with the insulin-stimulated pathway upstream of the RabGAPs is not fully understood. In this review, we discuss the current state of knowledge on exercise- and insulin-regulated kinases as well as hypoxia as stimulus that may be involved in the regulation of skeletal muscle glucose uptake. Full article

Oxidative stress (OS) plays a key role in autism spectrum disorder (ASD), a neurodevelopmental disorder characterized by deficits in social communication, restricted interests, and repetitive behaviors. Recent evidence suggests that the TLDc [Tre2/Bub2/Cdc16 (TBC), lysin motif (LysM), domain catalytic] domain is a highly conserved motif present in proteins that are important players in the OS response and in neuroprotection. Human proteins sharing the TLDc domain include OXR1, TLDC1, NCOA7, TBC1D24, and C20ORF118. This study was aimed at understanding whether TLDc domain-containing mRNAs together with specific microRNAs (200b-3p and 32-5p) and long noncoding RNAs (TUG1), known to target TLDc proteins, contributed to regulate the OS response in ASD. Data showed a significant increase in the levels of OXR1 and TLDC1 mRNAs in peripheral blood mononuclear cells (PBMCs) of ASD children compared to their neurotypically developing (NTD) counterparts, along with an increase in TUG1 mRNA expression levels, suggesting its possible role in the regulation of TLDc proteins. A positive correlation between the expression of some TLDc mRNAs and the Childhood Autism Rating Scale (CARS) global score as well as inflammatory gene expression was found. In conclusion, our data suggest a novel biological pathway in the OS response of ASD subjects that deserves further exploration. Full article

Glucose is an important fuel for highly active skeletal muscles. Increased adenosine monophosphate (AMP)/adenosine triphosphate (ATP) ratios during repetitive contractions trigger AMP-activated protein kinase (AMPK), indicated by phosphorylation of AMPKα^Thr172, which promotes glucose uptake to support heightened energy needs, but it also suppresses anabolic processes. Inhibition of AMPK can occur by protein kinase B (AKT)-mediated phosphorylation of AMPKα^Ser485/491, releasing its brake on growth. The influence of insulin-like growth factor I (IGF-I) on glucose uptake and its interplay with AMPK activation is not well understood. Thus, the goal of this study was to determine if increased muscle IGF-I altered AMPKα phosphorylation and activity during muscle contraction. Adult male mice harboring the rat Igf1a cDNA regulated by the fast myosin light chain promoter (mIgf1^+/+) and wildtype littermates (WT) were used in the study. mIgf1^+/+ mice had enhanced glucose tolerance and insulin-stimulated glucose uptake, but similar exercise capacity. Fatiguing stimulations of extensor digitorum longus (EDL) muscles resulted in upregulated AMPKα phosphorylation at both Thr172 and Ser485/491 in WT and mIgf1^+/+ muscles. No differences in the phosphorylation response of the downstream AMPK target TBC1D1 were observed, but phosphorylation of raptor was significantly higher only in WT muscles. Further, total raptor content was elevated in mIgf1^+/+ muscles. The results show that high muscle IGF-I can enhance glucose uptake under resting conditions; however, in contracting muscle, it is not sufficient to inhibit AMPK activity. Full article

The aim of this study was to investigate the microbial and physico-chemical characteristics of cold smoked sea bass (CSSB), a novel italian fish product. The microbiological analyses showed the presence of bacterial contamination from the raw material, the environment, and the production process. The microbial spoilage population was dominated by lactic acid bacteria (LAB) associated with Gram-negative fermenting bacteria, including Photobacterium phosphoreum and psychrotrophic Enterobacteriaceae. Brochotrix thermospacta and Aeromonas spp. were also present; in contrast, mould and yeast were not detected (<2 CFU/g). High levels (6–7 log CFU/g) of LAB and total bacteria count (TBC) were observed from day 45 of storage; however, their presence does not seem to have influenced the total volatile basic nitrogen (TVB-N), which always remained below 35 mg N/100 g. Consequently, the product is acceptable until day 60 of storage, considering that the malonaldehyde index (TBARS) was lower than 6.5 nmol/g. Pathogenic bacteria such as Salmonella spp. and Listeria monocytogenes were not detected. Currently, there is a growing demand for seafood due to its high quality and nutritional value. Cold smoked sea bass offers a source of macro- and micronutrients essential for the proper functioning of the human body. It is also rich in protein and omega-3 fatty acids. The WHO and FAO evaluated the benefits and risks and concluded that there is convincing evidence of health benefits from fish consumption, such as a reduction in the risk of heart failure and improved neurodevelopment in infants and young children when fish is consumed by the mother before and during pregnancy. The CSSB analysed in this study demonstrated to have health benefits due to long-chain omega-3 PUFAs and other nutrients, such as proteins, minerals, and vitamin D, which are sometimes difficult to obtain from other sources. The results show that CSSB has a high nutritional value and excellent microbial quality. Full article

The runt-related transcription factor 3 (RUNX3) regulates the differentiation of monocytes and their response to inflammation. However, the transcriptomic context of RUNX3 expression in blood monocytes remains poorly understood. We aim to learn about RUNX3 from its relationships within transcriptomes of bulk CD14+ cells in adults. This study used immunomagnetically sorted CD14+ cell gene expression microarray data from the Multi-Ethnic Study of Atherosclerosis (MESA, n = 1202, GSE56047) and the Correlated Expression and Disease Association Research (CEDAR, n = 281, E-MTAB-6667) cohorts. The data were preprocessed, subjected to RUNX3-focused correlation analyses and random forest modeling, followed by the gene ontology analysis. Immunity-focused differential ratio analysis with intermediary inference (DRAIMI) was used to integrate the data with protein–protein interaction network. Correlation analysis of RUNX3 expression revealed the strongest positive association for EVL (r_mean = 0.75, p_FDR-MESA = 5.37 × 10⁻¹⁴⁰, p_FDR-CEDAR = 5.52 × 10⁻⁸⁰), ARHGAP17 (r_mean = 0.74, p_FDR-MESA = 1.13 × 10⁻¹⁶⁹, p_FDR-CEDAR = 9.20 × 10⁻⁵⁹), DNMT1 (r_mean = 0.74, p_FDR-MESA = 1.10 × 10⁻¹⁶⁹, p_FDR-CEDAR = 1.67 × 10⁻⁵⁸), and CLEC16A (r_mean = 0.72, p_FDR-MESA = 3.51 × 10⁻¹⁵⁴, p_FDR-CEDAR = 2.27 × 10⁻⁵⁵), while the top negative correlates were C2ORF76 (r_mean = −0.57, p_FDR-MESA = 8.70 × 10⁻⁹⁴, p_FDR-CEDAR = 1.31 × 10⁻²⁵) and TBC1D7 (r_mean = −0.55, p_FDR-MESA = 1.36 × 10⁻⁶⁹, p_FDR-CEDAR = 7.81 × 10⁻³⁰). The RUNX3-associated transcriptome signature was involved in mRNA metabolism, signal transduction, and the organization of cytoskeleton, chromosomes, and chromatin, which may all accompany mitosis. Transcriptomic context of RUNX3 expression in monocytes hints at its relationship with cell growth, shape maintenance, and aspects of the immune response, including tyrosine kinases. Full article

Disruption of circadian rhythms is related to disorders of glucose metabolism, and the molecular clock also exists in skeletal muscle. The ChIP-derived repressor of network oscillator (Chrono) and brain and muscle ARNT-like 1 (Bmal1) are core circadian components. Chrono is considered to be the repressor of Bmal1, and the Chrono–Bmal1 pathway is important in regulating the circadian rhythm; it has been speculated that this pathway could be a new mechanism for regulating glucose metabolism. The purpose of this study was to investigate the effects of Chrono on glucose metabolism in skeletal muscle and exercise capacity by using mice with skeletal-muscle-specific overexpression of Chrono (Chrono TG) and wild-type (WT) mice as the animal models. The results of this cross-sectional study indicated that the Chrono TG mice had an impaired glucose tolerance, lower exercise capacity, and higher levels of nonfasted blood glucose and glycogen content in skeletal muscle compared to WT mice. In addition, the Chrono TG mice also showed a significant increase in the amount of Chrono bound to Bmal1 according to a co-IP analysis; a remarkable decrease in mRNA expression of Tbc1d1, Glut4, Hk2, Pfkm, Pdp1, Gbe1, and Phka1, as well as in activity of Hk and protein expression of Ldhb; but higher mRNA expression of Pdk4 and protein expression of Ldha compared with those of WT mice. These data suggested the skeletal-muscle-specific overexpression of Chrono led to a greater amount of Chrono bound to Bmal1, which then could affect the glucose transporter, glucose oxidation, and glycogen utilization in skeletal muscle, as well as exercise capacity. Full article

Insulin signaling plays a key role in glucose uptake, glycogen synthesis, and protein and lipid synthesis. In insulin-resistant states like obesity and type 2 diabetes mellitus, these processes are dysregulated. Regular physical exercise is a potential therapeutic strategy against insulin resistance, as an acute bout of exercise increases glucose disposal during the activity and for hours into recovery. Chronic exercise increases the activation of proteins involved in insulin signaling and increases glucose transport, even in insulin resistant states. Here, we will focus on the effect of acute exercise on insulin signaling and protein kinase B (Akt) pathways. Activation of proximal proteins involved in insulin signaling (insulin receptor, insulin receptor substrate-1 (IRS-1), phosphoinoside-3 kinase (PI3K)) are unchanged in response to acute exercise/contraction, while activation of Akt and of its substrates, TBC1 domain family 1 (TBC1D1), and TBC domain family 4 (TBC1D4) increases in response to such exercise/contraction. A wide array of Akt substrates is also regulated by exercise. Additionally, AMP-activated protein kinase (AMPK) seems to be a main mediator of the benefits of exercise on skeletal muscle. Questions persist on how mTORC1 and AMPK, two opposing regulators, are both upregulated after an acute bout of exercise. Full article

The health benefits of probiotics have been known for decades, but there has only been limited use of probiotics in the treatment of obesity. In this study, we describe, for the first time, the role of cell-free metabolites (CM) from Bacillus ginsengihumi-RO6 (CMRO6) in adipogenesis and lipogenesis in 3T3-L1 pre-adipocytes. The experimental results show that CMRO6 treatment effectively reduced lipid droplet accumulation and the expression of CCAAT/enhancer-binding protein α and β (C/EBPα and C/EBPβ), peroxisome proliferator-activated receptor γ (PPAR-γ), serum regulatory binding protein 1c (SREBP-1c), fatty acid-binding protein 4 (FABP4), fatty acid synthase (FAS), acetyl CoA carboxylase (ACC), phosphorylated p38MAPK, and Erk44/42. Additionally, CMRO6 treatment significantly increased glucose uptake and phosphorylated Akt (S473), AS160, and TBC1D1 protein expressions. Considering the results of this study, B. ginsengihumi may be a novel probiotic used for the treatment of obesity and its associated metabolic disorders. Full article

Chronic alcohol consumption retards lipophagy, which contributes to the pathogenesis of liver steatosis. Lipophagy-related Rab7 has been presumed as a crucial regulator in the progression of alcohol liver disease despite elusive mechanisms. More importantly, whether or not hepatoprotective quercetin targets Rab7-associated lipophagy disorder is unknown. Herein, alcoholic fatty liver induced by chronic-plus-single-binge ethanol feeding to male C57BL/6J mice was manifested by hampering autophagosomes formation with lipid droplets and fusion with lysosomes compared with the normal control, which was normalized partially by quercetin. The GST-RILP pulldown assay of Rab7 indicated an improved GTP-Rab7 as the quercetin treatment for ethanol-feeding mice. HepG2 cells transfected with CYP2E1 showed similar lipophagy dysfunction when exposed to ethanol, which was blocked when cells were transfected with siRNA-Rab7 in advance. Ethanol-induced steatosis and autophagic flux disruption were aggravated by the Rab7-specific inhibitor CID1067700 while alleviated by transfecting with the Rab7^Wt plasmid, which was visualized by immunofluorescence co-localization analysis and mCherry-GFP-LC3 transfection. Furthermore, TBC1D5, a Rab GTPase-activating protein for the subsequent normal circulation of Rab7, was downregulated after alcohol administration but regained by quercetin. Rab7 circulation retarded by ethanol and corrected by quercetin was further revealed by fluorescence recovery after photobleaching (FRAP). Altogether, quercetin attenuates hepatic steatosis by normalizing ethanol-imposed Rab7 turnover disorders and subsequent lipophagy disturbances, highlighting a novel mechanism and the promising prospect of quercetin-like phytochemicals against the crucial first hit from alcohol. Full article

Alaska pollack roe (APR) is a protein source that is usually salted and fermented, containing a high salt content. Using a combination of superheated steam roasting and smoking, we developed a new low-salt ready-to-eat APR variant, whose quality characteristics we analyzed. The optimal conditions for roasting (216 °C for 4 min) and smoking (64 °C for 14 min) were obtained from sensorial attributes using response surface methodology. Under the optimal conditions, smoke-roasted APR had an overall acceptance (OA) score of 8.89. The combination of roasting and smoking significantly increased volatile basic nitrogen (VBN, 18.6%) and decreased the total bacterial count (TBC, 38.6%), while thiobarbituric acid reactive substances (TBARS) were not affected. Smoke-roasting APR also increased its nutritional content to 30% protein with 44% essential amino acids, and more than 40% DHA and EPA in 4.3% fat. During 30 days of storage, the OA, VBN, TBARS, and TBC values significantly changed with time and storage temperature (p < 0.05). The shelf life of the product was estimated to be 24 d. In conclusion, the combination of roasting and smoking APR could improve product quality and may be an alternative to diversify processed APR. Full article

The adipokine Neuregulin 4 (Nrg4) protects against obesity-induced insulin resistance. Here, we analyze how the downregulation of Nrg4 influences insulin action and the underlying mechanisms in adipocytes. Validated shRNA lentiviral vectors were used to generate scramble (Scr) and Nrg4 knockdown (KD) 3T3-L1 adipocytes. Adipogenesis was unaffected in Nrg4 KD adipocytes, but there was a complete impairment of the insulin-induced 2-deoxyglucose uptake, which was likely the result of reduced insulin receptor and Glut4 protein. Downregulation of Nrg4 enhanced the expression of proinflammatory cytokines. Anti-inflammatory agents recovered the insulin receptor, but not Glut4, content. Proteins enriched in Glut4 storage vesicles such as the insulin-responsive aminopeptidase (IRAP) and Syntaxin-6 as well as TBC1D4, a protein involved in the intracellular retention of Glut4 vesicles, also decreased by Nrg4 KD. Insulin failed to reduce autophagy in Nrg4 KD adipocytes, observed by a minor effect on mTOR phosphorylation, at the time that proteins involved in autophagy such as LC3-II, Rab11, and Clathrin were markedly upregulated. The lysosomal activity inhibitor bafilomycin A1 restored Glut4, IRAP, Syntaxin-6, and TBC1D4 content to those found in control adipocytes. Our study reveals that Nrg4 preserves the insulin responsiveness by preventing inflammation and, in turn, benefits the insulin regulation of autophagy. Full article