Role of CD45 Signaling Pathway in Galactoxylomannan-Induced T Cell Damage

Previously, we reported that Galactoxylomannan (GalXM) activates the extrinsic and intrinsic apoptotic pathways through an interaction with the glycoreceptors on T cells. In this study we establish the role of the glycoreceptor CD45 in GalXM-induced T cell apoptosis, using CD45^+/+ and CD45^−/− cell lines, derived from BW5147 murine T cell lymphoma. Our results show that whereas CD45 expression is not required for GalXM association by the cells, it is essential for apoptosis induction. In CD45^+/+ cells, CD45 triggering by GalXM reduces the activation of Lck, ZAP70 and Erk1/2. Conversely, in CD45^−/− cells, Lck was hyperphosphorylated and did not show any modulation after GalXM stimulation. On the whole, our findings provide evidence that the negative regulation of Lck activation occurs via CD45 engagement. This appears to be related to the capacity of GalXM to antagonize T cell activation and induce T cell death. Overall this mechanism may be responsible for the immune paralysis that follows GalXM administration and could explain the powerful immunosuppression that accompanies cryptococcosis.     

Potassium channel-oxidative phosphorylation relationship in durum wheat mitochondria from control and hyperosmotic-stressed seedlings

Durum wheat mitochondria (DWM) possess an ATP-inhibited K(+) channel, the plant mitoK(ATP) (PmitoK(ATP) ), which is activated under environmental stress to control mitochondrial ROS production. To do this, PmitoK(ATP) collapses membrane potential (ΔΨ), thus suggesting mitochondrial uncoupling. We tested this point by studying oxidative phosphorylation (OXPHOS) in DWM purified from control seedlings and from seedlings subjected both to severe mannitol and NaCl stress. In severely-stressed DWM, the ATP synthesis via OXPHOS, continuously monitored by a spectrophotometric assay, was about 90% inhibited when the PmitoK(ATP) was activated by KCl. Contrarily, in control DWM, although PmitoK(ATP) collapsed ΔΨ, ATP synthesis, as well as coupling [respiratory control (RC) ratio and ratio between phosphorylated ADP and reduced oxygen (ADP/O)] checked by oxygen uptake experiments, were unaffected. We suggest that PmitoK(ATP) may play an important defensive role at the onset of the environmental/oxidative stress by preserving energy in a crucial moment for cell and mitochondrial bioenergetics. Consistently, under moderate mannitol stress, miming an early stress condition, the channel may efficiently control reactive oxygen species (ROS) generation (about 35-fold from fully open to closed state) without impairing ATP synthesis. Anyway, if the stress significantly proceeds, the PmitoK(ATP) becomes fully activated by decrease of ATP concentration (25-40%) and increase of activators [free fatty acids (FFAs) and superoxide anion], thus impairing ATP synthesis.     

Malaria seroprevalence in blood bank donors from endemic and non-endemic areas of Venezuela

In Venezuela, a total of 363,466 malaria cases were reported between 1999-2009. Several states are experiencing malaria epidemics, increasing the risk of vector and possibly transfusion transmission. We investigated the risk of transfusion transmission in blood banks from endemic and non-endemic areas of Venezuela by examining blood donations for evidence of malaria infection. For this, commercial kits were used to detect both malaria-specific antibodies (all species) and malaria antigen (Plasmodium falciparum only) in samples from Venezuelan blood donors (n = 762). All samples were further studied by microscopy and polymerase chain reaction (PCR). The antibody results showed that P. falciparum-infected patients had a lower sample/cut-off ratio than Plasmodium vivax-infected patients. Conversely, a higher ratio for antigen was observed among all P. falciparum-infected individuals. Sensitivity and specificity were higher for malarial antigens (100 and 99.8%) than for antibodies (82.2 and 97.4%). Antibody-positive donors were observed in Caracas, Ciudad Bolívar, Puerto Ayacucho and Cumaná, with prevalences of 1.02, 1.60, 3.23 and 3.63%, respectively. No PCR-positive samples were observed among the donors. However, our results show significant levels of seropositivity in blood donors, suggesting that more effective measures are required to ensure that transfusion transmission does not occur.     

The Marine Isolate <Emphasis Type="Italic">Novosphingobium</Emphasis> sp. PP1Y Shows Specific Adaptation to Use the Aromatic Fraction of Fuels as the Sole Carbon and Energy Source

Novosphingobium sp. PP1Y, isolated from a surface seawater sample collected from a closed bay in the harbour of Pozzuoli (Naples, Italy), uses fuels as its sole carbon and energy source. Like some other Sphingomonads, this strain can grow as either planktonic free cells or sessile-aggregated flocks. In addition, this strain was found to grow as biofilm on several types of solid and liquid hydrophobic surfaces including polystyrene, polypropylene and diesel oil. Strain PP1Y is not able to grow on pure alkanes or alkane mixtures but is able to grow on a surprisingly wide range of aromatic compounds including mono, bi, tri and tetracyclic aromatic hydrocarbons and heterocyclic compounds. During growth on diesel oil, the organic layer is emulsified resulting in the formation of small biofilm-coated drops, whereas during growth on aromatic hydrocarbons dissolved in paraffin the oil layer is emulsified but the drops are coated only if the mixtures contain selected aromatic compounds, like pyrene, propylbenzene, tetrahydronaphthalene and heterocyclic compounds. These peculiar characteristics suggest strain PP1Y has adapted to efficiently grow at the water/fuel interface using the aromatic fraction of fuels as the sole carbon and energy source.     

The effects of ATP and sodium chloride on the cytochrome c-cardiolipin interaction: the contrasting behavior of the horse heart and yeast proteins

In cells a portion of cytochrome c (cyt c) (15–20%) is tightly bound to cardiolipin (CL), one of the phospholipids constituting the mitochondrial membrane. The CL-bound protein, which has nonnative tertiary structure, altered heme pocket, and disrupted Fe(III)-M80 axial bond, is thought to play a role in the apoptotic process. This has attracted considerable interest in order to clarify the mechanisms governing the cyt c–CL interaction. Herein we have investigated the binding reaction of CL with the c-type cytochromes from horse heart and yeast. Although the two proteins possess a similar tertiary architecture, yeast cyt c displays lower stability and, contrary to the equine protein, it does not bind ATP and lacks pro-apoptotic activity. The study has been performed in the absence and in the presence of ATP and NaCl, two compounds that influence the (horse cyt c)-CL binding process and, thus, the pro-apoptotic activity of the protein. The two proteins behave differently: while CL interaction with horse cyt c is strongly influenced by the two effectors, no effect is observed for yeast cyt c. It is noteworthy that NaCl induces dissociation of the (horse cyt c)–CL complex but has no influence on that of yeast cyt c. The differences found for the two proteins highlight that specific structural factors, such as the different local structure conformation of the regions involved in the interactions with either CL or ATP, can significantly affect the behavior of cyt c in its reaction with liposomes and the subsequent pro-apoptotic action of the protein.     

From chronic kidney disease to transplantation: the roles of obestatin

This study was designed to investigate the possible effect of sitagliptin on renal damage induced by renal ischemia reperfusion (I/R) in diabetic rats. T2DM in rats was induced by the administration of nicotinamide (230 mg/kg, i.p.), 15 min prior to a single dose of streptozotocin (65 mg/kg, i.v.). In vivo renal I/R was performed in both T2DM and normal rats. Each protocol comprised ischemia for 30 min followed by reperfusion for 24 h and a treatment period of 14 days before induction of ischemia. Sitagliptin treated diabetic rats that underwent renal I/R demonstrated significant decrease in the serum concentrations of aspartate aminotransferase (p < 0.01), urea nitrogen (p < 0.01) and creatinine (p < 0.001) compared to renal I/R in diabetic rats. Lipid peroxidation, xanthine oxidase activity, myeloperoxidase activity and nitric oxide level in renal tissue were significantly (p < 0.05, p < 0.001, p < 0.01, p < 0.05 respectively) decreased after renal I/R in sitagliptin treated rats compared to diabetic rats. Antioxidant enzymes like glutathione (p < 0.05), glutathione peroxidase (p < 0.001), superoxide dismutase (p < 0.05) and catalase (p < 0.001) were significantly increased after renal I/R in sitagliptin treated diabetic rats compared to non treated diabetic rats. The typical DNA laddering was observed when renal I/R performed in diabetic rats, which indicates cell apoptosis. Sitagliptin treated rats demonstrated a decrease in DNA fragmentation and apoptosis. Furthermore, renal histopathology preserved in sitagliptin treated rats verified protection against renal I/R in diabetes. The results of present investigation established sitagliptin treatment attenuated renal damage induced by renal I/R in diabetic rats.     

SslE Elicits Functional Antibodies That Impair In Vitro Mucinase Activity and In Vivo Colonization by Both Intestinal and Extraintestinal Escherichia coli Strains

SslE, the Secreted and surface-associated lipoprotein from Escherichia coli, has recently been associated to the M60-like extracellular zinc-metalloprotease sub-family which is implicated in glycan recognition and processing. SslE can be divided into two main variants and we recently proposed it as a potential vaccine candidate. By applying a number of in vitro bioassays and comparing wild type, knockout mutant and complemented strains, we have now demonstrated that SslE specifically contributes to degradation of mucin substrates, typically present in the intestine and bladder. Mutation of the zinc metallopeptidase motif of SslE dramatically impaired E. coli mucinase activity, confirming the specificity of the phenotype observed. Moreover, antibodies raised against variant I SslE, cloned from strain IHE3034 (SslE_IHE3034), are able to inhibit translocation of E. coli strains expressing different variants through a mucin-based matrix, suggesting that SslE induces cross-reactive functional antibodies that affect the metallopeptidase activity. To test this hypothesis, we used well-established animal models and demonstrated that immunization with SslE_IHE3034 significantly reduced gut, kidney and spleen colonization by strains producing variant II SslE and belonging to different pathotypes. Taken together, these data strongly support the importance of SslE in E. coli colonization of mucosal surfaces and reinforce the use of this antigen as a component of a broadly protective vaccine against pathogenic E. coli species.     

Body Mass Index and the Prevalence of Hypertension and Dyslipidemia

Objective: To describe and evaluate relationships between body mass index (BMI) and blood pressure, cholesterol, high‐density lipoprotein‐cholesterol (HDL‐C), and hypertension and dyslipidemia. Research Methods and Procedures: A national survey of adults in the United States that included measurement of height, weight, blood pressure, and lipids (National Health and Nutrition Examination Survey III 1988–1994). Crude age‐adjusted, age‐specific means and proportions, and multivariate odds ratios that quantify the association between hypertension or dyslipidemia and BMI, controlling for race/ethnicity, education, and smoking habits are presented. Results: More than one‐half of the adult population is overweight (BMI of 25 to 29.9) or obese (BMI of ≥30). The prevalence of high blood pressure and mean levels of systolic and diastolic blood pressure increased as BMI increased at ages younger than 60 years. The prevalence of high blood cholesterol and mean levels of cholesterol were higher at BMI levels over 25 rather than below 25 but did not increase consistently with increasing BMI above 25. Rates of low HDL‐C increased and mean levels of HDL‐C decreased as levels of BMI increased. The associations of BMI with high blood pressure and abnormal lipids were statistically significant after controlling for age, race or ethnicity, education, and smoking; odds ratios were highest at ages 20 to 39 but most trends were apparent at older ages. Within BMI categories, hypertension was more prevalent and HDL‐C levels were higher in black than white or Mexican American men and women. Discussion: These data quantify the strong associations of BMI with hypertension and abnormal lipids. They are consistent with the national emphasis on prevention and control of overweight and obesity and indicate that blood pressure and cholesterol measurement and control are especially important for overweight and obese people.     

8-Hydroxydeoxyguanosine in DNA from leukocytes of healthy adults: relationship with cigarette smoking, environmental tobacco smoke, alcohol and coffee consumption

8-Hydroxydeoxyguanosine (8-OHdG) has been widely used as a biomarker of oxidative DNA damage in both animal and human studies. However, controversial data exist on the relationship between 8-OHdG formation and age, sex and tobacco smoking in humans, while few or no data are available on other exposures such as environmental tobacco smoke, alcohol, coffee and tea consumption. We investigated the level of 8-OHdG in DNA from peripheral leukocytes among 102 healthy adults living in Brescia province, North Italy, aged 25-45 (mean: 35.2 years), of which 51 were males. 8-OHdG levels expressed as a ratio to total deoxyguanosine (8-OHdG/106 dG) in DNA showed wide interindividual variation, the highest value (63.8) being 6. 2-fold greater than the lowest (10.3). Current smokers showed lower mean 8-OHdG values than subjects who never smoked (29.3 and 34.0, respectively, p<0.05), and an inverse relationship was found between 8-OHdG and lifetime smoking, which was independent of age, sex and body mass index. An inverse relationship was also found with coffee drinking while no association was observed with alcohol and tea consumption, exposure to environmental tobacco smoke and use of vitamins in all subjects, and with use of oral contraceptives in females. The inverse relationship between smoking status and 8-OHdG levels could be explained by the presence of efficient repair processes for the oxidative damage induced by smoking. In this study, the smokers were relatively young (77% were less than 40 years) and only 7% smoked 30 or more cigarettes a day. In conclusion, it would appear that 8-OHdG levels in leukocytes may not provide a sensitive marker of exposure to tobacco smoking.