Effect of vitamin E supplementation with standard treatment on oxidant-antioxidant status in chronic obstructive pulmonary disease.

BACKGROUND & OBJECTIVES: Chronic oxidant burden and depletion of endogenous antioxidants have been proposed to play a key role in the pathogenesis of chronic obstructive pulmonary disease (COPD). Exogenous antioxidants have potential therapeutic implications and their role has not been explored in COPD. The objective of this study was to investigate the effect of supplementation of standard treatment (inhaled long-acting beta(2) agonists, anticholinergics and corticosteroids) with vitamin E on oxidant-antioxidant balance in patients with COPD. METHODS: The study was carried out in the outpatient setting. Patients were divided into two groups: group A- placebo group (n=14), receiving only standard therapy, and group B- vitamin E-supplemented group (n=10), receiving 400 IU of vitamin E capsules twice daily in addition to standard therapy. Spirometry and clinical assessment were carried out at the start and completion of 8 wk treatment along with measurements of several biochemical parameters of oxidant-antioxidant status in plasma, leukocytes and red cells separated from venous blood. RESULTS: Leukocyte superoxide generation was decreased in both the groups. Vitamin E-supplemented group had significantly increased levels of plasma sulphydryls and red cell catalase while the placebo group had decreased levels of plasma nitrates and nitrites. No significant differences were observed in red cell superoxide dismutase and glutathione peroxidase activities, total blood glutathione, and plasma total antioxidant capacity, lipid peroxides and glutathione peroxidase activity in either group. There was a similar degree of lung function and clinical improvement in both the groups. INTERPRETATION & CONCLUSION: Our findings showed that an 8 wk supplementation of standard treatment with 400 IU twice daily of vitamin E did not provide any additional clinical benefit although it augmented certain endogenous antioxidants in patients with COPD.

Targeting protein acetylation for improving cancer therapy.

Acetylation is one of the most important post-translational modification of proteins determining the structure, function and intracellular localization that plays an important role in the signal transduction pathways related to diverse cell functions, both during unstimulated and stress conditions. Protein acetylation in cells is regulated by a co-ordinated action of histone acetyl transferases (HAT) and histone deacetylases(HDAC) that ensures the maintenance of homeostasis and execution of activities related to damage response viz. DNA repair, cell cycle delay, apoptosis and senescence. Since inhibition of histone deacetylation, stalls the progress of many nuclear events including proliferation and damage response events on the one hand and the levels of deacetylases are elevated in many tumours on the other. Histone deacetylase has been among the targets for the development of anticancer drugs and adjuvant. The recent observation showing acetylation of proteins by calreticulin (an endoplasmic reticulum resident protein) with a high efficiency when polyphenolic acetates are the acetyl group donating molecules and acetyl CoA as weak substrate extends the realm of protein acetylation beyond HAT/HDAC combination. Elucidation of the relative roles of HAT/HDAC mediated acetylation viz. a calreticulin mediated acetylation in cell function under a variety of stress conditions would hold key to the design of drugs targeting protein acetylation system.

Effect of sensitization on membrane ion fluxes & intracellular calcium in guineapigs.

BACKGROUND & OBJECTIVES: The biochemical mechanisms underlying the development of sensitization-induced airway hyperresponsiveness (AHR) in asthma are poorly defined. Alterations in the regulation of intracellular calcium may play an important role in its pathogenesis. We carried out this study to see the effect of sensitization with ovalbumin on membrane ion fluxes and intracellular calcium in a guinea pig model. METHODS: Airway reactivity to inhaled histamine was measured initially and after sensitization with ovalbumin in 28 guineapigs. Intracellular calcium [Ca(2+)]i was measured in tracheal smooth muscle cells and peripheral leukocytes using fluorescent dye FURA 2AM. Calcium and sodium ion influx across the cell membrane was measured in leukocytes. Ouabain-sensitive Rubidium ((86)Rb) influx was measured in tracheal smooth muscles cells. The activities of Na(+), K(+) ATPase and Ca(2+) ATPase were measured in tracheal smooth muscle cells. Lipid peroxides were measured in plasma. RESULTS: Airway responsiveness was significantly (P<0.001) increased after sensitization along with an increase in [Ca2+]i levels in leukocytes and tracheal smooth muscle cells, higher rates of (45)Ca and (22)Na influx in leukocytes and higher (86)Rb influx rates in tracheal smooth muscle cells, and increased levels of lipid peroxides in plasma. INTERPRETATION & CONCLUSION: In guineapig model of asthma sensitization to allergen increased the membrane permeability to calcium and sodium, and intracellular calcium levels. These alterations may play a role in the pathogenesis of airway hyper-responsiveness following sensitization.

Suicidal oxidative stress induced by certain antioxidants.

Well known antioxidants-coumarins (7,8-dihydroxy-4-methyl coumarin-DHMC and 7,8-diacetoxy-4-methyl coumarin-DAMC) and flavonoids (quercetin-Q and quercetin penta-acetate-QPA) were investigated for their pro-oxidant effects in two human tumor cell lines. The breast carcinoma cell line (MDA-MB-468) was found to be more sensitive to treatment by the drugs-DAMC, Q and QPA at 10 microM than the glioma cell line (U-87MG), while DHMC was non toxic in both cell lines at this concentration. In MDA-MB-468 distinct growth inhibition was observed by 48 hr post treatment. Paradoxically, an increase in the formazan production was revealed by MTT assay at this time indicating an increase in the production of free radicals. An increase in the levels of reactive oxygen species (ROS) was also confirmed by DCFH-DA assay. In cells treated with DAMC, Q and QPA an increase in the percentage of cells with the hypodiploid DNA content was suggestive of apoptotic cell death. Taken together, these results suggest that an increase in oxidative stress caused by the pro-oxidant action of these drugs is responsible for cell death.

Buffalo plasma fibronectin: a physico-chemical study.

Plasma fibronectin (FN) of buffalo (Babulis babulis) was purified to apparent homogeneity, using gelatin-Sepharose and heparin-Sepharose affinity columns. It was found to have two subunits of molecular mass 246 kDa and 228 kDa, on SDS-gel. Its immunological cross-reactivity with anti-human plasma FN was confirmed by Western blotting. The amino acid composition was found to be similar to that of human and bovine plasma FNs. Buffalo plasma FN contained 2.23% neutral hexoses and 1.18% sialic acids. No titrable sulfhydryl group could be detected in the absence of denaturant. Reaction with DTNB indicated 3.4 sulfhydryl groups in the molecule, whereas BDC-OH titration gave a value of 3.8 -SH groups in buffalo plasma FN. Stoke's radius, intrinsic viscosity, diffusion coefficient and frictional ratio indicated that buffalo plasma FN did not have a compact globular conformation at physiological pH and ionic strength. Molecular dimensions (average length, 120 nm; molar mass to length ratio, 3950 nm(-1) and mean diameter, 2.4 nm) as revealed by rotary shadowing electron microscopy further supported the extended conformation of buffalo plasma FN. These results show that buffalo plasma FN has similar properties as that of human plasma FN.

Long-term stability of bronchial reactivity in guinea pigs.

Measurement of lung function and bronchial reactivity are widely used as outcome parameters to assess the efficacy of therapeutic interventions. In order to interpret the results correctly, it is necessary that the outcome parameters are themselves stable over time so that any significant changes measured may be attributed to the interventions. Specific airway conductance (SGaw) and airway reactivity to histamine are two commonly used parameters in animal models such as guinea pigs. Although short-term variability of these parameters has been investigated, there has been no study of long-term stability. In the present paper, SGaw and bronchial reactivity to histamine were measured in 111 conscious guinea pigs using a non-invasive, whole body plethysmograph. Baseline values of SGaw and ED35 histamine were measured and followed for eight weeks at weekly intervals. At baseline, mean SGaw in guinea pigs was 0.17 +/- 0.055 sec-1 cm H2O-1 and ED35 histamine ranged from 0.064 to more than 10 mg/ml. The distribution of ED35 histamine values was gaussian. We observed that the changes in SGaw and ED35 histamine recorded using this technique are highly reproducible over eight weeks. The reactivity varied by less than a doubling dose of histamine over any two consecutive weeks. Thus, the technique described in this paper is quick, easily learned, reproducible, independent of temperature-humidity artifact and highly suitable for studies of repeated measurements as in the study of dietary interventions and evaluation of effect of drugs.