Enhanced oxidative stress contributes to the pathogenesis of diabetes and its complications. Peroxiredoxin 6 (PRDX6) is a key regulator of cellular redox balance, with the peculiar ability to neutralize peroxides, peroxynitrite, and phospholipid hydroperoxides. In the current study, we aimed to define the role of PRDX6 in the pathophysiology of type 2 diabetes (T2D) using PRDX6 knockout (2/2) mice. Glucose and insulin responses were evaluated respectively by intraperitoneal glucose and insulin tolerance tests. Peripheral insulin sensitivity was analyzed by euglycemic-hyperinsulinemic clamp, and molecular tools were used to investigate insulin signaling. Moreover, inflammatory and lipid parameters were evaluated. We demonstrated that PRDX6 2/2 mice developed a phenotype similar to early-stage T2D caused by both reduced glucose-dependent insulin secretion and increased insulin resistance. Impaired insulin signaling was present in PRDX6 2/2 mice, leading to reduction of muscle glucose uptake. Morphological and ultrastructural changes were observed in islets of Langerhans and livers of mutant animals, as well as altered plasma lipid profiles and inflammatory parameters. In conclusion, we demonstrated that PRDX6 is a key mediator of overt hyperglycemia in T2D glucose metabolism, opening new perspectives for targeted therapeutic strategies in diabetes care.A large body of evidence supports a pivotal role for oxidative stress in the etiopathogenesis of insulin resistance (IR) and diabetes (1). Oxidative stress is characterized by an imbalance between reactive oxygen species (ROS) production and antioxidant defense systems. Among all body tissues, pancreatic b-cells are very sensitive to oxidative stress because of their low expression of antioxidant enzymes like superoxide dismutase (SOD) and glutathione peroxidase (2). Moreover, hyperglycemia by itself induces IR, increasing oxidative stress injuries, which lead to overt type 2 diabetes (T2D) (3). Interestingly, a relatively new family of antioxidant proteins, the peroxiredoxins (PRDXs), is more highly expressed in pancreatic b-cells (4). Among the six members of this non-seleno peroxidase family, PRDX6 is present in the cytoplasm and is unique because it has peroxidase and also phospholipase A 2 activity (5). Several findings demonstrate the importance of PRDX6 in maintaining redox homeostasis, as follows: lack of PRDX6, in fact, increases the susceptibility to oxidative stress in different tissues (6,7). Nevertheless, data on the relationship between PRDX6 and the pathogenesis of IR and T2D are not available (8). Therefore, we hypothesized that, in terms of physiological status, PRDX6 may play a role in the etiology of IR and diabetes conditions through tissue redox levels. In the current study, we tested our hypothesis in a model of PRDX6 knockout mice (PRDX6 2/2). RESEARCH DESIGN AND METHODS Animal ModelsC57BL/6J wild-type (WT) mice weighing 18-20 g were obtained from The Jackson Laboratory (Bar Harbor, ME), while PRDX6 2/2 mice of mixed background (C57BL6/129SvJ)...

Cetuximab and panitumumab bind the human epidermal growth factor receptor (EGFR). Although the chimeric cetuximab (IgG1) triggers antibody‐dependent‐cellular‐cytotoxicity (ADCC) of EGFR positive target cells, panitumumab (a human IgG2) does not. The inability of panitumumab to trigger ADCC reflects the poor binding affinity of human IgG2 Fc for the FcγRIII (CD16) on natural killer (NK) cells. However, both human IgG1 and IgG2 bind the FcγRII (CD32A) to a similar extent. Our study compares the ability of T cells, engineered with a novel low‐affinity CD32A131R‐chimeric receptor (CR), and those engineered with the low‐affinity CD16158F‐CR T cells, in eliminating EGFR positive epithelial cancer cells (ECCs) in combination with cetuximab or panitumumab. After T‐cell transduction, the percentage of CD32A131R‐CR T cells was 74 ± 10%, whereas the percentage of CD16158F‐CR T cells was 46 ± 15%. Only CD32A131R‐CR T cells bound panitumumab. CD32A131R‐CR T cells combined with the mAb 8.26 (anti‐CD32) and CD16158F‐CR T cells combined with the mAb 3g8 (anti‐CD16) eliminated colorectal carcinoma (CRC), HCT116FcγR+ cells, in a reverse ADCC assay in vitro. Crosslinking of CD32A131R‐CR on T cells by cetuximab or panitumumab and CD16158F‐CR T cells by cetuximab induced elimination of triple negative breast cancer (TNBC) MDA‐MB‐468 cells, and the secretion of interferon gamma and tumor necrosis factor alpha. Neither cetuximab nor panitumumab induced Fcγ‐CR T antitumor activity against Kirsten rat sarcoma (KRAS)‐mutated HCT116, nonsmall‐cell‐lung‐cancer, A549 and TNBC, MDA‐MB‐231 cells. The ADCC of Fcγ‐CR T cells was associated with the overexpression of EGFR on ECCs. In conclusion, CD32A131R‐CR T cells are efficiently redirected by cetuximab or panitumumab against breast cancer cells overexpressing EGFR.

During ischemia/reperfusion (I/R), cardiomyocytes are exposed to sudden lack of nutrients and successively to radical oxygen species (ROS). In the present study, we used the HL-5 cardiac atrial myocyte cell line exposed to serum/glucose depletion added or not in H(2)O(2) to mimic ROS during ischemia, then replaced in their standard culture medium to simulate reperfusion. We investigated the effects of serum/glucose depletion combined or not to ROS exposure on AKT and MAP kinases activation to address the role of each event with respect to apoptosis. We demonstrate that serum/glucose depletion per se did not induce apoptosis when compared to ROS exposure. In particular, ROS recruited p38MAPK and JNK pathways. SB202190 preventing p38MAPK activity, partially protected HL-5 from apoptosis while blocking JNK, thanks to JNKI, further enhanced apoptosis. Blocking phosphatidylinositol (PI) 3-kinase with LY294002 or ERKs with U0126 was without consequence on apoptosis. Finally, BCL-2 and BCL-X(L/S) expression levels were analyzed in cells exposed to 1 h ischemia followed by 12-h reperfusion in the presence or not of SB202190; BCL-2, but not BCL-X(L/S), expression was decreased in ROS treated cells but SB202190 failed to restore BCL-2 level. Our data suggest that p38MAPK activation primarily mediates ROS-induced apoptosis while concomitant JNK activation would represent a scavenger pathway for cells trying to escape apoptosis.

BackgroundThe efficacy of Phosphodiesterase 5 (PDE5) inhibitors to re-establish endothelial function is reduced in diabetic patients. Recent evidences suggest that therapy with PDE5 inhibitors, i.e. sildenafil, may increase the expression of nitric oxide synthase (NOS) proteins in the heart and cardiomyocytes. In this study we analyzed the effect of sildenafil on endothelial cells in insulin resistance conditions in vitro.Methodology/Principal FindingsHuman umbilical vein endothelial cells (HUVECs) were treated with insulin in presence of glucose 30 mM (HG) and glucosamine 10 mM (Gluc-N) with or without sildenafil. Insulin increased the expression of PDE5 and eNOS mRNA assayed by Real time-PCR. Cytofluorimetric analysis showed that sildenafil significantly increased NO production in basal condition. This effect was partially inhibited by the PI3K inhibitor LY 294002 and completely inhibited by the NOS inhibitor L-NAME. Akt-1 and eNOS activation was reduced in conditions mimicking insulin resistance and completely restored by sildenafil treatment. Conversely sildenafil treatment can counteract this noxious effect by increasing NO production through eNOS activation and reducing oxidative stress induced by hyperglycaemia and glucosamine.Conclusions/SignificanceThese data indicate that sildenafil might improve NOS activity of endothelial cells in insulin resistance conditions and suggest the potential therapeutic use of sildenafil for improving vascular function in diabetic patients.

Senescence is a phenomenon characterized by a progressive decline of body homeostasis. Premature senescence acts when the cellular system is not able to adequately respond to noxious stimuli by synthesis of stressor molecules. Among those, serum-and-glucocorticoidinducible kinase-1 (SGK-1) dramatically increases under typical physiopathological conditions, such as glucocorticoid or mineralcorticoids exposure, inflammation, hyperglycemia, and ischemia. SGK-1 has been implicated in mechanism regulating oxidative stress, apoptosis, and DNA damage, which are all leading to a state of accelerating aging. Moreover, SGK-1-sensitive ion channels participate in the regulation of renal Na(+)/K(+) regulation, blood pressure, gastric acid secretion, cardiac action potential, and neuroexcitability. Recently, we demonstrated in endothelial cells as an increase in SGK-1 activity and expression reduces oxidative stress, improves cell survival and restores insulin-mediated nitric oxide production after hyperglycemia. Moreover, we showed as SGK-1 delays the onset of senescence by increasing telomerase activity, significantly decreasing reactive oxygen species (ROS) production, and by directly interacting with hTERT. Therefore, SGK-1 may represent a specific target to further develop novel therapeutic options against chronic diseases such as diabetes typical of aging. SGK-1 has been also associated with cancer, neurodegenerative diseases, and cardiovascular disease, among other age-related diseases. However, to date, the data available on SGK-1 and aging, are sparse, controversial, and only from C. elegans experimental models. In this review we sought to discuss the possible implication of SGK-1 in mechanisms regulating senescence and age-related diseases. Moreover, we aimed to discuss and identify the possible role of SGK-1 as possible molecular target to counteract and prevent aging.