Glucose-6-phosphate-dehydrogenase deficiency as a risk factor in proliferative disorder development

Glucose-6-phosphate dehydrogenase (G6PD) is an important site of metabolic control in the pentose phosphate pathway (PPP) which provides reducing power (NADPH) and pentose phosphates. The former is mainly involved in the detoxification of chemical reactive species; the latter in the regulation of cell proliferation. G6PD deficiency is the most common enzymopathy in the human population, characterized by decreased G6PD activity, mainly in red blood cells, but actually also in nucleated cells. This decreased activity is not due to enzyme synthesis impairment, but rather to reduced enzyme stability, which leads to a shortening of its half-life. Therefore, a major problem is to understand the underlying mechanisms linking G6PD deficiency to oxidative stress and cell proliferation. In order to address this issue, in the present study we utilized, as an experimental model, fibroblasts isolated from pterygium, an ocular proliferative lesion, from G6PD normal and deficient (PFs+ and PFs-, respectively) patients. Our choice was determined by the fact that pterygium is believed to be caused by chronic oxidative stress induced by UV exposure, and that pterygium fibroblasts resemble a tumorigenic phenotype. As controls we utilized fibroblasts isolated from conjunctiva from G6PD normal and deficient patients (NCFs+ and NCFs-, respectively) who had undergone cataract surgery. 
Growth rate analysis revealed that PFs grow faster than NCFs, but while NCFs- grow more slowly than NCFs+, PFs- and PFs+ grow at the same rate. This was associated with significantly lower G6PD activity in NCFs+ compared to NCFs-, while no significant differences in the G6PD activity of PFs+ and PFs- were noted. This result was supported by the finding that in PFs-, G6PD mRNA levels were significantly higher than in PFs+. Another interesting finding of this study was increased green autofluorescence in both NCFs- and PFs- compared to corresponding positive cells, indicative of pronounced oxidative stress in deficient cells. Finally, abnormal accumulation of neutral lipids, mainly cholesterol esters was observed both in PFs- and PFs+ compared to NCFs- and NCFs+. Though further studies are necessary for better understanding the exact mechanism which links G6PD to oxidative stress and cell proliferation, our data allow to speculate on the role of G6PD on tumorigenesis, and to consider G6PD-deficient subjects at major risk to develop common and dreaded proliferative disorders, such as atherosclerosis and cancer. 
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Impact of Environmental Factors on Bacteriocin Promoter Activity in Gut-Derived Lactobacillus salivarius

peer-reviewedBacteriocin production is regarded as a desirable probiotic trait that aids in colonization and persistence in the gastrointestinal tract (GIT). Strains of Lactobacillus salivarius, a species associated with the GIT, are regarded as promising probiotic candidates and have a number of associated bacteriocins documented to date. These include multiple class IIb bacteriocins (salivaricin T, salivaricin P, and ABP-118) and the class IId bacteriocin bactofencin A, which show activity against medically important pathogens. However, the production of a bacteriocin in laboratory media does not ensure production under stressful environmental conditions, such as those encountered within the GIT. To allow this issue to be addressed, the promoter regions located upstream of the structural genes encoding the L. salivarius bacteriocins mentioned above were fused to a number of reporter proteins (green fluorescent protein [GFP], red fluorescent protein [RFP], and luciferase [Lux]). Of these, only transcriptional fusions to GFP generated signals of sufficient strength to enable the study of promoter activity in L. salivarius. While analysis of the class IIb bacteriocin promoter regions indicated relatively weak GFP expression, assessment of the promoter of the antistaphylococcal bacteriocin bactofencin A revealed a strong promoter that is most active in the absence of the antimicrobial peptide and is positively induced in the presence of mild environmental stresses, including simulated gastric fluid. Taken together, these data provide information on factors that influence bacteriocin production, which will assist in the development of strategies to optimize in vivo and in vitro production of these antimicrobials.This work was funded by a SFI PI award “Obesibiotics” (11/PI/1137) to PD

Human microglial cells synthesize albumin in brain

Albumin has been implicated in Alzheimer's disease since it can bind to and transport amyloid beta, the causative agent; albumin is also a potent inhibitor of amyloid beta polymerization. In a pilot phase study of Human Brain Proteome Project, we found evidence that albumin may be synthesized in immortalized human microglial cells, human primary microglial cells, and human fetal and adult brain tissues. We also found the synthesis and secretion is enhanced upon microglial activation by Amyloid [beta]~1-42~, lipopolysaccharide treatment or human Alzheimer's brain

The disintegrin echistatin in combination with doxorubicin targets high-metastatic human osteosarcoma overexpressing ανβ3 integrin in chick embryo and nude mouse models.

Echistatin, a cyclic RGD peptide, which is an antagonist of αvβ3 integrin (disintegrin), inhibited human osteosarcoma in the chick chorioallontoic membrane (CAM) model and tumor growth and pulmonary metastases in a nude mouse orthotopic model. A high-metastatic variant of human osteosarcoma, 143B-LM4, overexpressing αvβ3 integrin was used. Tumor angiogenesis by high-metastatic variant 143B-LM4 cells in the CAM was significantly inhibited by echistatin (P<0.05) as was overall growth. A doxorubicin (DOX)-echistatin combination inhibited orthotopic tumor growth compared to untreated control (P<0.01) or DOX alone (P<0.05) in nude mice. Tumor-bearing mice treated with the DOX-echistatin combination survived longer than those treated with DOX alone or control PBS (P<0.01 and P<0.01, respectively). Echistatin also inhibited experimental lung metastasis of 143B-LM4 cells in nude mice. These results suggest that DOX in combination with a disintegrin has potential to treat osteosarcoma and that αvβ3 integrin may be a target for osteosarcoma

Angiotensin type-2 (AT-2)-receptor activation reduces renal fibrosis in cyclosporine nephropathy: evidence for blood-pressure independent effect

Compound 21 (C21), selective agonist of AT2 receptors, shows antinflammatory effects in hypertension and nephroprotection in diabetes. The aim of this study was to evaluate the effects of C21 in cyclosporine nephropathy, which is characterized mainly by tubulo-interstitial fibrosis. Ten days before and during the experimental periods, low-salt diet was administered to Sprague Dawley rats. Cyclosporine-A (15mg/kg/day, i.p.) and cyclosporine-A plus C21 (0.3 mg/kg /day, i.p) were administered for 1 and 4 weeks. Control groups was left without any treatment. Blood pressure (plethysmographic method) and 24 hour albuminuria were measured once a week. At the end of the experiments, the kidneys were excised for histomorphometric analysis of renal fibrosis and for immunohistochemical evaluation of inflammatory infiltrates and type I and IV collagen expression.After 1 and 4 weeks, the rats treated with cyclosporine showed a significant increase (p <0.01) in blood pressure, no significant changes in albuminuria, a significant increase (p <0.01) in glomerular and tubulo-interstitial fibrosis and inflammatory infiltrates as compared to the control rats. Treatment with C21 did not modify the cyclosporine dependent increase of blood pressure, which was higher than in control rats, but after 4 weeks of treatment significantly reduced (p <0.01) glomerular and tubulo-interstitial fibrosis, type 1 collagen expression and macrophage infiltration, as compared to rats treated with cyclosporine.The administration of C21 showed a protective effect on cyclosporine nephropathy, decreasing renal fibrosis and macrophage infiltration. These data suggest that C21 may counteract tubulo-interstitial fibrosis, the most potent predictor of the progression of renal diseases

Spectral tracing of deuterium for imaging glucose metabolism.

Cells and tissues often display pronounced spatial and dynamical metabolic heterogeneity. Common glucose-imaging techniques report glucose uptake or catabolism activity, yet do not trace the functional utilization of glucose-derived anabolic products. Here we report a microscopy technique for the optical imaging, via the spectral tracing of deuterium (STRIDE), of diverse macromolecules derived from glucose. Based on stimulated Raman-scattering imaging, STRIDE visualizes the metabolic dynamics of newly synthesized macromolecules, such as DNA, protein, lipids and glycogen, via the enrichment and distinct spectra of carbon-deuterium bonds transferred from the deuterated glucose precursor. STRIDE can also use spectral differences derived from different glucose isotopologues to visualize temporally separated glucose populations using a pulse-chase protocol. We also show that STRIDE can be used to image glucose metabolism in many mouse tissues, including tumours, brain, intestine and liver, at a detection limit of 10 mM of carbon-deuterium bonds. STRIDE provides a high-resolution and chemically informative assessment of glucose anabolic utilization