The innate immune Toll-like receptors (TLRs) 2/4 are important players in chronic low-grade inflammation called metabolic inflammation in obesity and type-2 diabetes (T2D). While TLR2/4 expression changes associated with metabolic inflammation are known, the adipose tissue expression of endocytic TLR8, which is expressed by all major macrophage subsets, remain unclear. We, therefore, determined the TLR8 mRNA/protein expression in the adipose tissue samples from lean, overweight, and obese individuals with or without T2D. Subcutaneous fat biopsy samples were collected from 49 non-diabetic (23 obese, 17 overweight, and nine lean) and 45 T2D (32 obese, ten overweight, and three lean) individuals. TLR8 gene expression was determined using real-time RT-PCR and TLR8 protein expression was assessed by both immunohistochemistry and confocal microscopy. The changes in TLR8 expression were compared with those of macrophage markers, proinflammatory cytokines/chemokines, and surface TLRs/adapter proteins. The data were analyzed using t-test/Mann-Whitney U-test, Pearson's correlation, and multiple regression test. The data show that in obese non-diabetic/T2D individuals, TLR8 gene expression was significantly upregulated as compared with lean individuals which correlated with body mass index (BMI) and body fat percentage in non-diabetic population (P < 0.05). As expected, TLR8 adipose tissue protein expression in non-diabetic/T2D obese individuals was also higher than that of overweight/lean counterparts. In non-diabetic/T2D individuals, TLR8 gene expression associated (P < 0.05) with the expression of CD68, CD11c, CD86, and CD163 macrophage markers. Also, in these individuals, TLR8 gene expression correlated positively (P < 0.05) with adipose tissue expression of TNF-α, IL-18, and IL-8 as well as with systemic CRP levels (in non-diabetics). TLR8 expression was also associated with TLR4/TLR2 and MyD88 expression in the adipose tissue. The elevated adipose tissue expression of TLR8 in obesity/T2D has consensus with inflammatory signatures and may thus represent an immune marker of metabolic inflammation.
Abstract Introduction The proinflammatory cytokine IL‐18 is involved in the pathogenesis of metabolic syndrome. While the changes in IL‐18 are known, IL‐18R expression and relationship with IL‐18 and other inflammatory markers in the adipose tissue in obesity/type‐2 diabetes (T2D) remain unclear. Methods We, therefore, determined the adipose tissue expression of IL‐18R and IL‐18 mRNA/protein in lean, overweight, and obese individuals with and without T2D, 15 each, using qRT‐PCR, immunohistochemistry, and confocal microscopy. Data (mean ± SEM) were analyzed using unpaired t ‐test and Pearson's correlation ( r ); all P values ≤0.05 were considered statistically significant. Results We found the upregulated gene/protein expression of IL‐18R and IL‐18 in non‐diabetic obese/overweight as compared with lean individuals ( P < 0.05). BMI correlated positively ( P < 0.05) with the adipose tissue expression of IL‐18R (mRNA: r = 0.90 protein: r = 0.84) and IL‐18 (mRNA: r = 0.84 protein: r = 0.80). Similarly, in T2D individuals, gene and protein expression of IL‐18R/IL‐18 was significantly higher in obese as compared with overweight/lean individuals. The BMI was associated with the changes in both IL‐18R (mRNA: r = 0.55 protein: r = 0.50) and IL‐18 (mRNA: r = 0.53 protein: r = 0.57) expression. IL‐18R/IL‐18 gene expression in the adipose tissue was positively associated ( P < 0.05) with local gene expression of other inflammatory markers including CD11c, CD86, CD68, CD163, TNF‐α, and CCL5. Homeostatic model assessment of insulin resistance (HOMA‐IR) was higher in diabetic/non‐diabetic obese and it correlated with BMI ( P < 0.05). IL‐18R and IL‐18 mRNA/protein expression in obesity was associated with HOMA‐IR only in non‐diabetics. Conclusions The adipose tissue IL‐18R/IL‐18 expression is enhanced in obesity which associates with proinflammatory gene signature and insulin resistance in these individuals.
1 A nomogram and a digital computer program have been developed to calculate dosage schedules of gentamicin for individual patients. The minimum input data consist of the patients' age, sex, body weight and serum creatinine concentration. 2 These prescribing aids have been evaluated in 36 patients with severe Gram negative infections. Renal function ranged from normal to complete anuria. Nomogram dosage schedules gave serum concentrations of gentamicin within the chosen therapeutic limits. Physician dosage schedules gave serum concentrations which sometimes exceeded and sometimes fell below these limits. The validity of the computer program was demonstrated by its ability to predict serum concentrations of gentamicin whatever the dosage schedule. 3 Half the patients recovered from the bacterial infection but seven remained infected and eleven died. Pseudomonas aeruginosa was the most difficult organism to eradicate. 4 Four of the patients who survived developed ataxia and two developed hearing loss at high frequencies. The risk of ototoxicity was a function of mean trough serum gentamicin concentration and duration of treatment. Ototoxicity was only detected in patients with serum creatinine concentrations above 3 mg/100 ml who tended to have higher trough concentrations. When treatment was prolonged beyond 8-10 days the risk of ototoxicity was increased without evidence of further substantial therapeutic benefit.
Abstract The emergence of effective vaccines for COVID-19 has been welcomed by the world with great optimism. Given their increased susceptibility to COVID-19, the question arises whether individuals with type-2 diabetes mellitus (T2DM) and other metabolic conditions can respond effectively to the mRNA-based vaccine. We aimed to evaluate the levels of anti-SARS-CoV-2 IgG and neutralizing antibodies in people with T2DM and/or other metabolic risk factors (hypertension and obesity) compared to those without. This study included 262 people that took two doses of BNT162b2 (Pfizer–BioNTech) mRNA vaccine. Both T2DM and non-diabetic individuals had a robust response to vaccination as demonstrated by their high antibody titers. However, both SARS-CoV-2 IgG and neutralizing antibodies titers were lower in people with T2DM. Their levels were 154±49.1 vs. 138±59.4BAU/mL for IgG and 87.1±11.6 vs. 79.7±19.5% for neutralizing antibodies in individuals without diabetes compared to those with T2DM, respectively. In a multiple linear regression adjusted for individual characteristics, comorbidities, previous COVID-19 infection and duration since second vaccine dose, diabetics had 13.86 BAU/ml (95%CI: -27.08 to -0.64BAU/ml, p=0.041) less IgG antibodies and 4.42% (95%CI: -8.53 to -0.32%, p=0.036) less neutralizing antibodies than non-diabetics. Hypertension and obesity did not show significant changes in antibody titers. Taken together, both type-2 diabetic and non-diabetic individuals elicited strong immune responses to SARS-CoV-2 BNT162b2 mRNA vaccine; nonetheless, lower levels were seen in people with diabetes. Continuous monitoring of the antibody levels might be a good indicator to guide personalized needs for further booster shots to maintain adaptive immunity.
This study unveils verapamil’s compelling cytoprotective and proliferative effects on pancreatic β-cells amidst diabetic stressors, spotlighting its unforeseen role in augmenting cholecystokinin (CCK) expression. Through rigorous investigations employing MIN6 β-cells and zebrafish models under type 1 and type 2 diabetic conditions, we demonstrate verapamil’s capacity to significantly boost β-cell proliferation, enhance glucose-stimulated insulin secretion, and fortify cellular resilience. A pivotal revelation of our research is verapamil’s induction of CCK, a peptide hormone known for its role in nutrient digestion and insulin secretion, which signifies a novel pathway through which verapamil exerts its therapeutic effects. Furthermore, our mechanistic insights reveal that verapamil orchestrates a broad spectrum of gene and protein expressions pivotal for β-cell survival and adaptation to immune-metabolic challenges. In vivo validation in a zebrafish larvae model confirms verapamil’s efficacy in fostering β-cell recovery post-metronidazole infliction. Collectively, our findings advocate for verapamil’s reevaluation as a multifaceted agent in diabetes therapy, highlighting its novel function in CCK upregulation alongside enhancing β-cell proliferation, glucose sensing, and oxidative respiration. This research enriches the therapeutic landscape, proposing verapamil not only as a cytoprotector but also as a promoter of β-cell regeneration, thereby offering fresh avenues for diabetes management strategies aimed at preserving and augmenting β-cell functionality.
Granulocyte–macrophage colony-stimulating factor (GM-CSF) is a monomeric glycoprotein that has been implicated in the tumor growth and progression of different types of cancer. GM-CSF is produced by various non-immune cells including MDA-MB-231 in response to various stimuli. However, the role of lipopolysaccharide (LPS) in the regulation of GM-CSF in MDA-MB-231 breast cancer cells so far remains unclear. Herein, we asked whether LPS could induce GM-CSF production in MDA-MB-231 cells, and if so, which signaling pathway was involved. MDA-MB-231 cells were treated with LPS or tumor necrosis factor alpha (TNF-α; positive control), and GM-CSF expression levels were determined by qRT-PCR, ELISA, and confocal microscopy. Phosphorylation of the mitogen-activated protein kinases (MAPKs) and nuclear factor-κB (NF-kB) signaling proteins were evaluated by flow cytometry. Our results show that LPS induces GM-CSF expression at both mRNA and protein levels in MDA-MBA-231 cells. Inhibition of acyl-CoA synthetase 1 (ACSL1) activity in the cells with triacsin C significantly reduces the secretion of GM-CSF. Furthermore, the inhibition of ACSL1 activity significantly blocks the LPS-mediated phosphorylation of p38 MAPK, MEK1/2, extracellular signal-regulated kinase (ERK)1/2, c-Jun NH2-terminal kinase (JNK), and nuclear factor-κB (NF-kB) in the cells. These findings provide the first evidence that LPS induces ACSL1-dependent GM-CSF gene expression in MDA-MB-231 breast cancer cells, which requires the activation of p38 MAPK, MEK1/2, ERK1/2, JNK, and NF-kB.
For many diabetics in end-stage renal failure, the initial therapy they receive will be continuous ambulatory peritoneal dialysis (CAPD) together with i.p. insulin. To date, all published protocols rely on empirical dosages based upon predialysis insulin requirements. A practical regimen for the institution of i.p. insulin administration during CAPD is described. The only endpoints used to determine insulin dosage were fasting plasma glucose 5 to 10 mmol/L and 2 h postprandial plasma glucose 8 to 15 mmol/L. An initial protocol related to body weight, dextrose content, volume, and timing of dialysate was based on a retrospective analysis of the results in our first 10 patients. Subsequently, a prospective assessment in an additional 22 patients confirmed the effectiveness of the regimen. The following protocol is recommended for the institution of i.p. insulin therapy in patients undergoing CAPD: Preprandial exchanges 1.36% dextrose-0.175 U/L dialysate/kg body weight 3.86% dextrose-0.25 U/L/kg Overnight exchanges 1.36% dextrose-0.1 U/L/kg 3.86% dextrose-0.15 U/L/kg Further adjustment of insulin dosage is then made on the basis of four hourly plasma glucose profiles. Self-monitoring of capillary blood glucose is recommended.
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