Aims: Obesity is characterized by a low-grade systemic inflammatory state and adipose tissue (AT) dysfunction, which predispose individuals to the development of insulin resistance (IR) and metabolic disease. However, a subset of obese individuals, referred to as metabolically healthy obese (MHO) individuals, are protected from obesity-associated metabolic abnormalities. Here, we aim at identifying molecular factors and pathways in adipocytes that are responsible for the progression from the insulin-sensitive to the insulin-resistant, metabolically unhealthy obese (MUHO) phenotype. Results: Proteomic analysis of paired samples of adipocytes from subcutaneous (SC) and omental (OM) human AT revealed that both types of cells are altered in the MUHO state. Specifically, the glutathione redox cycle and other antioxidant defense systems as well as the protein-folding machinery were dysregulated and endoplasmic reticulum stress was increased in adipocytes from IR subjects. Moreover, proteasome activity was also compromised in adipocytes of MUHO individuals, which was associated with enhanced accumulation of oxidized and ubiquitinated proteins in these cells. Proteasome activity was also impaired in adipocytes of diet-induced obese mice and in 3T3-L1 adipocytes exposed to palmitate. In line with these data, proteasome inhibition significantly impaired insulin signaling in 3T3-L1 adipocytes. Innovation: This study provides the first evidence of the occurrence of protein homeostasis deregulation in adipocytes in human obesity, which, together with oxidative damage, interferes with insulin signaling in these cells. Conclusion: Our results suggest that proteasomal dysfunction and impaired proteostasis in adipocytes, resulting from protein oxidation and/or misfolding, constitute major pathogenic mechanisms in the development of IR in obesity. Antioxid. Redox Signal . 23, 597–612.

Glycerol is an important metabolite for the control of lipid accumulation in white adipose tissue (WAT) and liver. We aimed to investigate whether exogenous administration of leptin improves features of non-alcoholic fatty liver disease (NAFLD) in leptin-deficient ob/ob mice via the regulation of AQP3 and AQP7 (glycerol channels mediating glycerol efflux in adipocytes) and AQP9 (aquaglyceroporin facilitating glycerol influx in hepatocytes). Twelve-week-old male wild type and ob/ob mice were divided in three groups as follows: control, leptin-treated (1 mg/kg/d) and pair-fed. Leptin deficiency was associated with obesity and NAFLD exhibiting an AQP3 and AQP7 increase in WAT, without changes in hepatic AQP9. Adipose Aqp3 and hepatic Aqp9 transcripts positively correlated with markers of adiposity and hepatic steatosis. Chronic leptin administration (4-weeks) was associated with improved body weight, whole-body adiposity and hepatosteatosis of ob/ob mice and to a down-regulation of AQP3, AQP7 in WAT and an up-regulation of hepatic AQP9. Acute leptin stimulation in vitro (4-h) induced the mobilization of aquaglyceroporins towards lipid droplets (AQP3) and the plasma membrane (AQP7) in murine adipocytes. Our results show that leptin restores the coordinated regulation of fat-specific AQP7 and liver-specific AQP9, a step which might prevent lipid overaccumulation in WAT and liver in obesity.

Context: Molecular mechanisms associated with physiological variations in adipose tissue (AT) are not fully recognized. The most recent reports highlight the critical relevance of microRNAs (miRNAs) found in AT. Objective: To identify changes in messenger RNA (mRNA) and miRNA expressions and their interaction in human AT before and after surgery-induced weight loss. Research Design and Setting: Genome-wide mRNA and miRNA expressions were assessed by microarrays in abdominal subcutaneous AT of 16 morbidly obese women before and 2 years after laparoscopic Roux-en-Y gastric bypass. The association of changes in miRNAs with their respective mRNA targets was studied. The results were replicated in publicly available microarray datasets. Validation was made by real-time polymerase chain reaction in additional fat samples from 26 age-matched lean women and in isolated human adipocytes. Results: A total of 5018 different mRNA probe sets and 15 miRNAs were differentially expressed after surgery-induced weight loss. The clustering of similar expression patterns for gene products with related functions revealed molecular footprints that elucidate significant changes in cell cycle, development, lipid metabolism, and the inflammatory response. The participation of inflammation was demonstrated by results assessed in isolated adipocytes. Interestingly, when transcriptomes were analyzed taking into account the presence of miRNA target sites, miRNA target mRNAs were upregulated in obese AT (P value = 2 × 10⁻¹⁸¹) and inflamed adipocytes (P value = 4 × 10⁻⁶¹), according to the number of target sites harbored by each transcript. Conclusions: Current findings suggest impaired miRNA target gene expression in obese AT in close association with inflammation, both improving after weight loss.

Chitosan scaffolds based on blending polymers are a common strategy used in tissue engineering. The objective of this study was evaluation the properties of scaffolds based on a ternary blend of chitosan (Chi), gelatin (Ge), and polyvinyl alcohol (PVA) (Chi/Ge/PVA), which were prepared by cycles of freeze-thawing and freeze-drying. It then was used for three-dimensional BRIN-BD11 beta-cells culturing. Weight ratios of Chi/Ge/PVA (1:1:1, 2:2:1, 2:3:1, and 3:2:1) were proposed and porosity, pore size, degradation, swelling rate, compressive strength, and cell viability analyzed. All ternary blend scaffolds structures are highly porous (with a porosity higher than 80%) and interconnected. The pore size distribution varied from 0.6 to 265 μm. Ternary blends scaffolds had controllable degradation rates compared to binary blend scaffolds, and an improved swelling capacity of the samples with increasing chitosan concentration was found. An increase in Young’s modulus and compressive strength was observed with increasing gelatin concentration. The highest compressive strength reached 101.6 Pa. The MTT assay showed that the ternary blends scaffolds P3 and P4 supported cell viability better than the binary blend scaffold. Therefore, these results illustrated that ternary blends scaffolds P3 and P4 could provide a better environment for BRIN-BD11 cell proliferation.

Methicillin-resistant Staphylococcus aureus (MRSA) is a prevailing bacterial pathogen linked to superficial skin and soft tissue infections (SSTIs). Rifampicin (RIF), a potent antibiotic against systemic and localised staphylococcal infections, faces limitations due to its low solubility. This constraint hampers its therapeutic potential for MRSA-induced SSTIs. To address this, an advanced liposomal system was designed for efficient dermal RIF delivery. Rifampicin-loaded liposomes (LipoRIF) were embedded within polymeric dissolving microneedles (DMNs) to enable targeted intradermal drug delivery. A robust Design of Experiment (DoE) methodology guided the systematic preparation and optimisation of LipoRIF formulations. The optimal LipoRIF formulation integrated within polymeric DMNs. These LipoRIF-DMNs exhibited favourable mechanical properties and effective skin insertion characteristics. Notably, in vitro assays on skin deposition unveiled a transformative result - the DMN platform significantly enhanced LipoRIF deposition within the skin, surpassing LipoRIF dispersion alone. Moreover, LipoRIF-DMNs displayed minimal cytotoxicity toward cells. Encouragingly, rigorous in vitro antimicrobial evaluations demonstrated LipoRIF-DMNs' capacity to inhibit MRSA growth compared to the control group. LipoRIF-DMNs propose a potentially enhanced, minimally invasive approach to effectively manage SSTIs and superficial skin ailments stemming from MRSA infections.