Influence of Antioxidants on the Peroxidative Swelling of Mitochondria in vitro

To clarify the role of prooxidative processes during in vitro swelling of freshly isolated rat liver mitochondria, the influence of different antioxidants and free-radical scavengers was tested. Ascorbate below 10 mmol/L without externally added Fe²⁺ acted as a prooxidant and enhanced swelling. Higher concentrations in the presence of Fe²⁺ showed antioxidant properties and a decrease in swelling and lipid peroxidation. Swelling was abolished by -tocopherol and reduced to 50% by butylated hydroxytoluene. Glutathione supplementation decreased both swelling and lipid peroxidation. Oxidized glutathione caused swelling without any effect on peroxidation. Hydrogen peroxide, cumene hydroperoxide and t-butyl hydroperoxide caused progressive decreases in glutathione and reduced niacinamide coenzyme levels, suggesting prooxidative changes. Dithiothreitol was found to abolish this effect. Thus, antioxidants reverse superoxide-induced mito chondrial swelling and lipid peroxidation in vitro.     

Synthesis,anticancer activity and mitochondrial mediated apoptosis inducing ability of 2,5-diaryloxadiazole–pyrrolobenzodiazepine conjugates

A series of 2,5-diaryloxadiazole linked pyrrolo[2,1-c][1,4]benzodiazepine conjugates have been prepared and evaluated for their anticancer activity. These conjugates have shown promising activity with GI₅₀ values ranging from <0.1 to 0.29 μΜ. It is observed that some of these conjugates particularly 4a, 4d, 4i and 4l exhibit significant anticancer activity. Some detailed biological assays relating to the cell cycle aspects associated to Bax and caspases have been examined with a view to understand the mechanism of action of these conjugates.     

Comparison of Effects of p53 Null and Gain-of-Function Mutations on Salivary Tumors in MMTV-Hras Transgenic Mice

p53 is an important tumor suppressor gene which is mutated in ~50% of all human cancers. Some of these mutants appear to have acquired novel functions beyond merely losing wild-type functions. To investigate these gain-of-function effects in vivo, we generated mice of three different genotypes: MMTV-Hras/p53^+/+, MMTV-Hras/p53^-/-, and MMTV-Hras/p53^R172H/R172H. Salivary tumors from these mice were characterized with regard to age of tumor onset, tumor growth rates, cell cycle distribution, apoptotic levels, tumor histopathology, as well as response to doxorubicin treatment. Microarray analysis was also performed to profile gene expression. The MMTV-Hras/p53^-/- and MMTV-Hras/p53^R172H/R172H mice displayed similar properties with regard to age of tumor onset, tumor growth rates, tumor histopathology, and response to doxorubicin, while both groups were clearly distinct from the MMTV-Hras/p53^+/+ mice by these measurements. In addition, the gene expression profiles of the MMTV-Hras/p53^-/- and MMTV-Hras/p53^R172H/R172H tumors were tightly clustered, and clearly distinct from the profiles of the MMTV-Hras/p53^+/+ tumors. Only a small group of genes showing differential expression between the MMTV-Hras/p53^-/- and MMTV-Hras/p53^R172H/R172H tumors, that did not appear to be regulated by wild-type p53, were identified. Taken together, these results indicate that in this MMTV-Hras-driven salivary tumor model, the major effect of the p53 R172H mutant is due to the loss of wild-type p53 function, with little or no gain-of-function effect on tumorigenesis, which may be explained by the tissue- and tumor type-specific properties of this gain-of-function mutant of p53.     

Genetic Association of SNPs near ATOH7, CARD10, CDKN2B,CDC7 and SIX1/SIX6 with the Endophenotypes of Primary Open Angle Glaucoma in Indian Population

Primary open angle glaucoma (POAG) belonging to a group of optic neuropathies, result from interaction between genetic and environmental factors. Study of associations with quantitative traits (QTs) is one of the successful strategies to understand the complex genetics of POAG. The current study attempts to explore the association of variations near/in genes like ATOH7, SIX1/SIX6 complex, CDKN2B, CARD10, and CDC7 with POAG and its QTs including vertical cup to disc ratio (VCDR), central corneal thickness (CCT), intra ocular pressure (IOP), and axial length (AL). Case-control study design was carried out in a sample size of 97 POAG cases and 371 controls from South India. Model-based (additive, recessive, dominant) association of the genotypes and their interaction was carried out between cases and controls using chi-square, linear and logistic regression methods. Nominal significance (P<0.05) was observed for QTs like i) VCDR with SNPs rs1900004 (ATOH7); rs1192415 (CDC7); rs10483727 (SIX1/SIX6), rs9607469 (CARD10); ii) CCT with rs1192415; iii) IOP with rs1900004 and iv) AL with rs1900004 and rs1063192 (CDKN2B). We were able to replicate previously known interactions between ATOH7-SIX6 and SIX6-CDKN2B along with few novel interactions between ATOH7—CDC7 and SIX6 with genes including CARD10 and CDC7. In summary, our results suggest that a probable interaction among the candidate genes for QTs, play a major role in determining the individual’s susceptibility to POAG.     

Chronic Endotoxemia in Subjects with Type-1 Diabetes Is Seen Much before the Onset of Microvascular Complications

Background

Lipopolysaccharide (LPS)/Endotoxin is hypothesized to play an important role in chronic inflammation associated with Type-1 diabetes (T1DM) and its complications. Endotoxin core antibodies (EndoCAb), LPS binding protein (LBP) and soluble CD14 (sCD14) act as modulators of LPS induced activation of innate immune system in vivo. For the present study we estimated the levels of LPS and its translocation markers in T1DM subjects with and without microvascular complications (MVC) and correlate them with clinical parameters of T1DM and serum inflammatory cytokine levels (TNF-α, IL-6, IL-1β and GM-CSF).

Methods

A total of 197 subjects (64 normal glucose tolerance (NGT) subjects, 97 T1DM subjects without MVC and 36 with MVC) were included in this study and the levels of serum LPS, its translocation markers and cytokines measured by immunoassays.

Results

Compared to NGT, T1DM subjects (both with and without MVC) had significantly higher levels of LPS, reduced levels of LBP and EndoCAb along with significant increase in the levels of IL-1β, IL-6, TNF-α and GM-CSF (p<0.05). No significant change was seen in the levels of these biomarkers between T1DM subjects with and without MVC.

Conclusions

Decreased levels of EndoCAb and LBP suggest sustained endotoxin activity in T1DM subjects even before the onset of microvascular complications.     

Basal insulin, glucagon, and growth hormone replacement

Conclusions drawn from the pancreatic (or islet) clamp technique (suppression of endogenous insulin, glucagon, and growth hormone secretion with somatostatin and replacement of basal hormone levels by intravenous infusion) are critically dependent on the biological appropriateness of the selected doses of the replaced hormones. To assess the appropriateness of representative doses we infused saline alone, insulin (initially 0.20 mU.kg(-1).min(-1)) alone, glucagon (1.0 ng.kg(-1).min(-1)) alone, and growth hormone (3.0 ng.kg(-1).min(-1)) alone intravenously for 4 h in 13 healthy individuals. That dose of insulin raised plasma insulin concentrations approximately threefold, suppressed glucose production, and drove plasma glucose concentrations down to subphysiological levels (65 +/- 3 mg/dl, P < 0.0001 vs. saline), resulting in nearly complete suppression of insulin secretion (P < 0.0001) and stimulation of glucagon (P = 0.0059) and epinephrine (P = 0.0009) secretion. An insulin dose of 0.15 mU.kg(-1).min(-1) caused similar effects, but a dose of 0.10 mU.kg(-1).min(-1) did not. The glucagon and growth hormone infusions did not alter plasma glucose levels or those of glucoregulatory factors. Thus, insulin "replacement" doses of 0.20 and even 0.15 mU.kg(-1).min(-1) are excessive, and conclusions drawn from the pancreatic clamp technique using such doses may need to be reassessed.