Obesity Differs from Diabetes Mellitus in Antibody and T Cell Responses Post COVID-19 Recovery.

Obesity and type 2 diabetes (DM) are risk factors for severe COVID-19 outcomes, which disproportionately affect South Asian populations. This study aims to investigate the humoral and cellular immune responses to SARS-CoV-2 in adult COVID-19 survivors with obesity and DM in Bangladesh. In this cross-sectional study, SARS-CoV-2-specific antibody and T cell responses were investigated in 63 healthy and 75 PCR-confirmed COVID-19 recovered individuals in Bangladesh, during the pre-vaccination first wave of the COVID-19 pandemic in 2020. In COVID-19 survivors, SARS-CoV-2 infection induced robust antibody and T cell responses, which correlated with disease severity. After adjusting for age, sex, DM status, disease severity, and time since onset of symptoms, obesity was associated with decreased neutralising antibody titers, and increased SARS-CoV-2 spike-specific IFN-γ response along with increased proliferation and IL-2 production by CD8+ T cells. In contrast, DM was not associated with SARS-CoV-2-specific antibody and T cell responses after adjustment for obesity and other confounders. Obesity is associated with lower neutralising antibody levels and higher T cell responses to SARS-CoV-2 post COVID-19 recovery, while antibody or T cell responses remain unaltered in DM.

Arsenic exposure during pregnancy and postpartum maternal glucose tolerance: evidence from Bangladesh.

BACKGROUND: Arsenic exposure has been associated with gestational diabetes mellitus. However, the extent to which arsenic exposure during pregnancy is associated with postpartum glucose intolerance is unknown. METHODS: We studied 323 women in Bangladesh. We assessed arsenic exposure in early pregnancy via toenail and water samples. We measured fasting glucose and insulin in serum at a mean (SD) of 4.0 (3.5) weeks post-delivery. We ran covariate-adjusted, linear regression models to examine associations of arsenic concentrations with HOMA-IR, a marker of insulin resistance, and HOMA-ß, a marker of beta cell function. RESULTS: Median (IQR) arsenic concentration was 0.45 (0.67) µg/g in toenails and 2.0 (6.5) µg/L in drinking water. Arsenic concentrations during pregnancy were not associated with insulin resistance or beta cell function postpartum. HOMA-IR was 0.07% (- 3.13, 3.37) higher and HOMA-ß was 0.96% (- 3.83, 1.99) lower per IQR increment in toenail arsenic, but effect estimates were small and confidence intervals crossed the null. CONCLUSIONS: Although arsenic exposure during pregnancy has been consistently associated with gestational diabetes mellitus, we found no clear evidence for an adverse effect on postpartum insulin resistance or beta cell function.

Expression of neutrophil elastase and myeloperoxidase mRNA in patients with newly diagnosed type 2 diabetes mellitus.

BACKGROUND AND AIMS: Neutrophil elastase and myeloperoxidase enzymes protect us from infection by killing pathogens. However, exaggerated activities of these enzymes can induce tissue damage, inflammation and oxidative stress. The present study was aimed to explore the expressions of neutrophil elastase and myeloperoxidase mRNA in the peripheral blood leukocytes (PBL) in patients with newly diagnosed type 2 diabetes mellitus. METHODS: In this cross-sectional study, 104 participants including 65 normoglycemic control subjects and 39 newly diagnosed type 2 diabetes patients were recruited. Glycemic and metabolic markers were evaluated from fasting blood samples. The mRNA levels of neutrophil elastase and myeloperoxidase genes in the PBL were quantified by real-time quantitative PCR. RESULTS: Compared to control subjects, diabetes patients showed a significant down regulation of both neutrophil elastase (p = 0.039) and myeloperoxidase (p = 0.023) mRNA expressions in the PBL. The neutrophil elastase and myeloperoxidase mRNA levels showed a negative trend with fasting glucose levels but did not show any significant correlations with HbA1c, insulin level, insulin resistance or sensitivity status. CONCLUSIONS: It was concluded that type 2 diabetes mellitus is associated with a decrease in neutrophil elastase and myeloperoxidase gene expression in the PBL.

Spike protein D614G and RdRp P323L: the SARS-CoV-2 mutations associated with severity of COVID-19.

The severity of coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), greatly varies from patient to patient. In the present study, we explored and compared mutation profiles of SARS-CoV-2 isolated from mildly affected and severely affected COVID-19 patients in order to explore any relationship between mutation profile and disease severity. Genomic sequences of SARS-CoV-2 were downloaded from Global Initiative on Sharing Avian Influenza Data (GISAID) database. With the help of Genome Detective Coronavirus Typing Tool, genomic sequences were aligned with the Wuhan seafood market pneumonia virus reference sequence and all the mutations were identified. Distribution of mutant variants was then compared between mildly and severely affected groups. Among the numerous mutations detected, 14,408C>T and 23,403A>G mutations resulting in RNA-dependent RNA polymerase (RdRp) P323L and spike protein D614G mutations, respectively, were found predominantly in severely affected group (>82%) compared with mildly affected group (<46%, p < 0.001). The 241C>T mutation in the non-coding region of the genome was also found predominantly in severely affected group (p < 0.001). The 3,037C>T, a silent mutation, also appeared in relatively high frequency in severely affected group compared with mildly affected group, but the difference was not statistically significant (p = 0.06). We concluded that spike protein D614G and RdRp P323L mutations in SARS-CoV-2 are associated with severity of COVID-19. Further studies will be required to explore whether these mutations have any impact on the severity of disease.

Pharmacogenetic Variants in MTHFR Gene are Significant Predictors of Methotrexate Toxicities in Bangladeshi Patients With Acute Lymphoblastic Leukemia.

BACKGROUND: The objective of this pharmacogenetic study was to investigate the relationship of methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms with methotrexate (MTX)-induced toxicities and plasma homocysteine level in patients with acute lymphoblastic leukemia (ALL) from Bangladesh. Several polymorphisms result in reduced MTHFR activity that causes impaired remethylation of homocysteine to methionine and abnormal MTX metabolism, especially in tissues with high turnover. Therefore, the risk of elevated plasma homocysteine as well as MTX-induced toxicities become higher with MTHFR polymorphisms. PATIENTS AND METHODS: We recruited 160 patients with ALL receiving MTX containing chemotherapeutic protocol, and they were genotyped for MTHFR C677T and A1298C polymorphisms with polymerase chain reaction-restriction fragment length polymorphism. We also measured the plasma homocysteine level of 51 patients by the AxSYM homocysteine assay method. RESULTS: We found 68.1% CC, 26.3% CT, and 5.6% TT genotype for MTHFR C677T polymorphism and 39.3% AA, 46.9% AC, and 13.8% CC genotype for MTHFR A1298C polymorphism in patients with ALL. Our study suggested that MTX-induced mucositis and diarrhea are significantly associated with MTHFR C677T as well as MTHFR A1298C polymorphisms (P < .05). CONCLUSION: The risk of elevated plasma homocysteine level was 5 to 6 times higher for both polymorphisms. This study may help to identify the patients who are at higher risk for MTX-related toxicities.

Neutrophil elastase and myeloperoxidase mRNA expression in overweight and obese subjects.

Neutrophil elastase and myeloperoxidase enzymes have been implicated in high-fat diet-induced obesity, insulin resistance (IR) and atherosclerosis in animal models. The aim of the present study was to explore neutrophil elastase and myeloperoxidase mRNA expressions in the peripheral blood leukocytes (PBL) in overweight and obese subjects, and to correlate those mRNA expressions with BMI, IR and cardiovascular biomarkers. In this cross-sectional study, 74 apparently healthy subjects including 22 lean, 27 overweight and 25 obese subjects were recruited. Cardiovascular and metabolic biomarkers were evaluated from fasting blood samples. The mRNA levels of neutrophil elastase and myeloperoxidase genes in the PBL were quantified by real-time PCR. Compared to lean group, the overweight and obese groups showed significant upregulation of both neutrophil elastase (p < 0.001) and myeloperoxidase (p < 0.03) mRNA expressions in the PBL. But no difference was found between overweight and obese groups. The neutrophil elastase and myeloperoxidase mRNA levels showed significant positive correlation with BMI, serum triglyceride, atherogenic index of plasma and 10-year risk of developing cardiovascular disease. But no correlation was found with glucose, insulin or IR. It was concluded that the neutrophil elastase and myeloperoxidase genes are up-regulated in both overweight and obese subjects and are associated with BMI and markers of cardiovascular disease.

Homeostatic nuclear RAGE-ATM interaction is essential for efficient DNA repair.

The integrity of genome is a prerequisite for healthy life. Indeed, defects in DNA repair have been associated with several human diseases, including tissue-fibrosis, neurodegeneration and cancer. Despite decades of extensive research, the spatio-mechanical processes of double-strand break (DSB)-repair, especially the auxiliary factor(s) that can stimulate accurate and timely repair, have remained elusive. Here, we report an ATM-kinase dependent, unforeseen function of the nuclear isoform of the Receptor for Advanced Glycation End-products (nRAGE) in DSB-repair. RAGE is phosphorylated at Serine376 and Serine389 by the ATM kinase and is recruited to the site of DNA-DSBs via an early DNA damage response. nRAGE preferentially co-localized with the MRE11 nuclease subunit of the MRN complex and orchestrates its nucleolytic activity to the ATR kinase signaling. This promotes efficient RPA2S4-S8 and CHK1S345 phosphorylation and thereby prevents cellular senescence, IPF and carcinoma formation. Accordingly, loss of RAGE causatively linked to perpetual DSBs signaling, cellular senescence and fibrosis. Importantly, in a mouse model of idiopathic pulmonary fibrosis (RAGE-/-), reconstitution of RAGE efficiently restored DSB-repair and reversed pathological anomalies. Collectively, this study identifies nRAGE as a master regulator of DSB-repair, the absence of which orchestrates persistent DSB signaling to senescence, tissue-fibrosis and oncogenesis.

Effect of aqueous extract of Aegle marmelos fruit and leaf on glycemic, insulinemic and lipidemic status of type 2 diabetic model rats.

BackgroundAegle marmelos is a popular fruit plant in the Indian subcontinent, various parts of which are traditionally used against various illnesses including diabetes mellitus (DM). However, the underlying mechanisms of the antidiabetic effects of the plant are not clear, especially in type 2 DM. The present study was undertaken to investigate the effect of aqueous extracts of A. marmelos fruits (AMFE) and leaves (AMLE) on glycemic, lipidemic, insulinemic, insulin resistance and ß-cell functional status of type 2 diabetic model rats. Methods An interventional study was designed using 20 type 2 diabetic rats. Type 2 DM was induced in Long Evans rats by a single intra-peritoneal injection of streptozotocin (90 mg/kg body weight) to 48 h old pups. Three months after induction of diabetes, the rats were divided into three independent groups: water-treated control group (n=6), AMLE-treated group (n=7) and AMFE-treated group (n=7). The rats were fed with extracts or water for 21 consecutive days and blood samples were collected at days 0 and 21 after an overnight fast. Data were expressed as mean±SD and analyzed by paired t-test or ANOVA as appropriate. Results There were significantly lower blood glucose values in AMLE and AMFE groups at Endpoint compared to Baseline (mmol/l, mean±SD, Baseline vs. Endpoint, 7.04±1.0 vs. 6.06±0.92; p=0.032 and 7.04±0.97 vs. 5.87±0.93; p=0.047). There were also significantly lower serum insulin levels in AMLE and AMFE groups at Endpoint compared to Baseline (µIU/mL, mean±SD, Baseline vs. Endpoint, 14.02±5.48 vs. 7.57±2.90; p=0.026 and 11.54±4.83 vs. 6.58±4.36; p=0.008). Insulin resistance (HOMA-IR) was significantly improved both in AMLE and AMFE groups at Endpoint compared to Baseline (mean±SD, Baseline vs. Endpoint, 4.22±1.68 vs. 2.05±0.90; p=0.021 and 3.69±1.79 vs. 1.69±1.61; p=0.013). However, ß-cell function or lipid profile did not show any significant alteration at Endpoint compared to Baseline in AMLE and AMFE groups. Conclusions Aqueous extracts of A. marmelos leaf and fruit have hypoglycemic property which seem to be mediated by lowering of insulin resistance. These findings highlight the therapeutic potential of the extracts of A. marmelos in human type 2 DM and provides strong impetus for further studies.

Serine 307 on insulin receptor substrate 1 is required for SOCS3 and TNF-α signaling in the rMC-1 cell line.

PURPOSE: To establish the key insulin receptor substrate 1 (IRS-1) structural elements required in this insulin regulatory pathway, we investigated the effects of substituting alanine for serine 307 in IRS-1 on the ability of tumor necrosis factor-α (TNF-α) and a related mediator, suppressor of cytokine signaling 3 (SOCS3), to phosphorylate IRS-1 and regulate insulin signaling in the rat retinal Müller cell (rMC-1) cell line. METHODS: rMC-1 cells were grown in normal (5 mM) or high (25 mM) glucose medium and transfected with either normal IRS-1(Ser307)plasmid or a mutated IRS-1(Ser307Ala) plasmid. Cells were also treated with recombinant TNF-α or SOCS3 to induce increased levels of these proteins. RESULTS: In cells with IRS-1(Ser307Ala), TNF-α and SOCS3 failed to phosphorylate IRS-1. Likewise, resulting downstream effects, including changes in phosphorylation of insulin receptor(Tyr960), antiapoptotic Akt phosphorylation, and proapoptotic cleavage of caspase 3 were also blocked. We also report for the first time that SOCS3 and TNF-α are reciprocally stimulatory leading to a mutual enhancement of levels of both factors, thus forming a potential positive feedback loop that contributes to insulin receptor resistance. CONCLUSIONS: Increases in TNF-α and SOCS3 are triggered by high glucose and through reciprocal stimulation of expression of these two factors, which in turn could be major drivers of insulin resistance and related cell death. The demonstration that a single phosphorylation site is key for these pathways suggests that drugs targeted to this site might be effective in protecting against diabetic damage to the retina.

Sp1-dependent regulation of PPARα in bone metabolism.

PURPOSE: Successful repair and regeneration in bone tissue engineering vastly depends on proper interaction between the tissue-engineered construct and the recipient's immune system. In clinical application, adverse responses to bioartificial implants may result in chronic inflammation and loss of the implant. It is known that prolonged inflammation linked to NF-κB inflammatory pathways inhibits bone-forming activity of osteoblast cells. Contributing to orchestrate inflammatory processes, the ligand-activated transcription factor peroxisome proliferator-activated receptor alpha (PPARα) holds inhibitory effects on NF-κB and CEBß activity. Sp1, a widely expressed transcription factor, has been linked to PPAR pathways, cellular homeostasis, and responsiveness to environmental perturbation. Formerly not being characterized, the role of PPARα in inflammatory-mediated bone loss requires further investigation. The aim of the present study was to identify regulatory transcription factor binding sites (TFBS) on the PPAR alpha promoter and to assess the role of Sp1 and associated proteins in its regulation. MATERIALS AND METHODS: In a first set of experiments, polymerase chain reaction assessed the presence of PPARα gene expression in isolated murine bone tissue. Deletion mutagenesis was performed on the human PPARα (hPPARα) promoter gene, and the deletion constructs were transiently transfected to murine osteoblasts to identify important TFBS. PPARα promoter-driven reporter gene expression was monitored in response to overexpression and repression of Sp1 to analyze functional transcription factor recruitment to the PPARα promoter. RESULTS: This study could demonstrate that the full-length hPPARα promoter contains inhibiting promoter regions and that hPPARα basal expression can be significantly increased by deletion mutagensis. Sp1 TFBS proved functional in the regulation of PPARα promoter activity, and the first five Sp1 motifs on the PPARα promoter were sufficient to significantly increase PPARα expression. Additional transient co-transfection experiments could not detect any direct effect of NF-κB/IκB downstream pathway on the regulation of PPARα promoter activity. Taken together, we could demonstrate that Sp1 plays a key role in transcriptional regulation of PPARα promoter activity and gene expression. CONCLUSION: This study provides further insight on Sp1-dependent PPARα regulatory mechanisms and suggests that Sp1-regulated PPARα expression plays a key role in inflammatory mediated bone loss.

Serum soluble receptor for advanced glycation end products (sRAGE) is independently associated with cigarette smoking in non-diabetic healthy subjects.

This study was designed to explore the relationship between serum levels of soluble receptor for advanced glycation end products (sRAGE) and cigarette smoking in non-diabetic healthy subjects. A total of 98 non-diabetic, otherwise healthy male subjects were recruited. A fasting blood sample and medical history including detail history of cigarette smoking was collected. The serum sRAGE levels were found significantly higher (p=0.002) in cigarette smokers (1475±422 pg/ml, n=45) compared with non-smokers (1165±350 pg/ml, n=53). Moreover, among the cigarette smokers, serum sRAGE levels were found significantly correlated with number of cigarettes smoked per day (r=0.60, p<0.001). In bivariate analysis in the total population, sRAGE positively correlated with smoking habit (r=0.37, p=0.002) and negatively correlated with systolic (r=-0.32, p=0.01) and diastolic blood pressure (r=-0.36, p=0.003). However, in stepwise multivariate linear regression model, sRAGE showed a significant independent association with smoking habit (b=0.32, p=0.007, R2=0.23). In conclusion, this study for the first time shows a significant elevation of serum sRAGE in cigarette smokers compared with non-smokers, a strong correlation between sRAGE and number of cigarettes smoked per day and an independent association of sRAGE with smoking habit in non-diabetic healthy subjects.

Diabetic retinal neurodegeneration is associated with mitochondrial oxidative stress and is improved by an angiotensin receptor blocker in a model combining hypertension and diabetes.

OBJECTIVE: Diabetic retinopathy displays the features of a neurodegenerative disease. Oxidative stress is involved in the pathogenesis of diabetic retinopathy. This investigation sought to determine whether hypertension exacerbates the oxidative stress, neurodegeneration, and mitochondrial dysfunction that exists in diabetic retinopathy and whether these changes could be minimized by the angiotensin II type 1 (AT(1)) receptor blocker (ARB) losartan. RESEARCH DESIGN AND METHODS: Diabetes was induced in spontaneously hypertensive rats (SHRs) and normotensive Wistar-Kyoto (WKY) rats. The diabetic SHRs were assigned to receive or not receive losartan. RESULTS: The level of apoptosis in the retina was higher in diabetic WKY rats than in the control group, and higher levels were found in diabetic SHRs. The apoptotic cells expressed neural and glial markers. The retinal glial reaction was more evident in diabetic WKY rats and was markedly accentuated in diabetic SHRs. Superoxide production in retinal tissue increased in diabetic WKY rats, and a greater increase occurred in diabetic SHRs. Glutathione levels decreased only in diabetic SHRs. As a consequence, the levels of nitrotyrosine and 8-hydroxy 2'-deoxyguanosine, markers of oxidative stress, were elevated in diabetic groups, mainly in diabetic SHRs. Mitochondrial integrity was dramatically affected in the diabetic groups. The ARB treatment reestablished all of the above-mentioned parameters. CONCLUSIONS: These findings suggest that concomitance of hypertension and diabetes exacerbates oxidative stress, neurodegeneration, and mitochondrial dysfunction in the retinal cells. These data provide the first evidence of AT(1)blockage as a neuroprotective treatment of diabetic retinopathy by reestablishing oxidative redox and the mitochondrial function.

Antioxidant SOD mimetic prevents NADPH oxidase-induced oxidative stress and renal damage in the early stage of experimental diabetes and hypertension.

AIMS: The presence of hypertension increases renal oxidative stress by increasing NADPH oxidase-dependent superoxide production and by decreasing antioxidant defense in the early stage of experimental diabetes mellitus (DM). In the present study, we investigated whether the administration of an antioxidant mimetic of the superoxide dismutase (SOD) (tempol) corrects the oxidative imbalance and oxidative stress-induced renal injury in the presence of DM and hypertension. METHODS: DM was induced in spontaneously hypertensive rats (SHR) by streptozotocin at 4 weeks of age. The diabetic rats either did or did not receive tempol for 20 days. Oxidative-stress parameters and indices of renal injury were evaluated. RESULTS: Tempol reestablished the imbalance in redox status induced by DM. It elevated the expression of renal antioxidant extracellular SOD, p < 0.0001; decreased (p = 0.049) the production of renal NADPH-dependent superoxide production, and diminished (p = 0.016) a marker of oxidative stress-induced DNA damage, 8-hydroxy-2'-deoxyguanosine. Reduction of oxidative stress markers was associated with reduction in renal damage parameters associated with DN. DM-induced albuminuria and elevation in renal expression of collagen IV were reduced to the level observed in control rats. CONCLUSION: We conclude that an imbalance in renal redox status is associated with markers of renal injury in the early stage of DM and hypertension. Antioxidant treatment reestablished the redox status and prevented oxidative stress-induced renal damage.

Green tea (Camellia sinensis) attenuates nephropathy by downregulating Nox4 NADPH oxidase in diabetic spontaneously hypertensive rats.

Green tea (GT), through its antioxidant properties, may be useful to treat or prevent human diseases. Because several lines of evidence suggest that oxidative stress contributes to the pathogenesis of diabetic nephropathy, we tested the hypothesis that GT prevents diabetes and hypertension-related renal oxidative stress, attenuating renal injury. Spontaneously hypertensive rats (SHR) with streptozotocin-induced diabetes and nondiabetic SHR were treated daily with tap water or freshly prepared GT (13.3 g/L). After 12 wk, the systolic blood pressure did not differ between treated and untreated nondiabetic or diabetic rats. However, body weight was less (P < 0.05) and glycemia was greater in diabetic SHR rats than in nondiabetic rats. Renal oxidative stress variables such as 8-hydroxy-2'-deoxyguanosine (8-OHdG) and nitrotyrosine expression, NADPH oxidase-dependent superoxide generation, and the expression of renal cortex Nox4 were greater (P < 0.05) in diabetic rats that received water (DW) than in nondiabetic rats that received water (CW). The 8-OHdG and NADPH oxidase-dependent superoxide generation were significantly less in rats treated with GT. Nitrotyrosine and Nox4 expression were significantly less in diabetic rats that received GT (DGT) than in DW. Likewise, the indices of renal injury, albuminuria, and renal expression of collagen IV were significantly greater in DW than in CW. These differences were significantly less in DGT than in DW. GT reestablished the redox state and reduced the indicators of nephropathy without altering glycemia and blood pressure levels in diabetic SHR. These findings suggest that the consumption of GT may ameliorate nephropathy in diabetic hypertensive patients.

Hypertension increases pro-oxidant generation and decreases antioxidant defense in the kidney in early diabetes.

AIMS: The combination of hypertension and diabetes exacerbates renal oxidative stress. The aim of the present study was therefore to evaluate the pro-oxidant and antioxidant mechanisms responsible for the induction of renal oxidative stress in the presence of hypertension and diabetes mellitus. METHODS: Diabetes was induced in spontaneously hypertensive rats (SHR) and their genetically normotensive control Wistar-Kyoto (WKY) rats by streptozotocin at 12 weeks of age. After 10 days, pro-oxidant, antioxidant and oxidative stress parameters were evaluated in the renal tissue. RESULTS: NADPH oxidase-dependent superoxide generation in the renal cortex was significantly elevated in WKY and SHR diabetic (D) groups compared to the respective control (C) groups (p < 0.005, n = 5). However, the highest level of superoxide generation was observed in the SHR-D group compared to all other groups. The expression of the gp91phox subunit of NADPH oxidase was significantly elevated in the SHR-D (p < 0.05, n = 5), but not in the WKY-D group, compared to the respective control groups. The renal cortical extracellular-superoxide dismutase level was found to be markedly decreased in the SHR groups compared to the WKY groups (p < 0.05, n = 5). The antioxidant glutathione level was found to be lower in the SHR-D (p = 0.03, n = 15), but not in the WKY-D group, compared to the respective control groups. Finally, nitrotyrosine and 8-hydroxy-2'-deoxyguanosine, markers of oxidative stress, were found to be similar in the kidneys of WKY-C and WKY-D, but were elevated in the SHR-D compared to the SHR-C group. CONCLUSION: We therefore conclude that hypertension increases pro-oxidant generation and decreases antioxidant defense, and thereby induces renal oxidative stress in early diabetes.