Virological Characteristics of Acute Hepatitis B in Eastern India: Critical Differences with Chronic Infection

Hepatitis B Virus (HBV) manifests high genetic variability and is classifiable into ten genotypes (A-J). HBV infection can lead to variable clinical outcomes, ranging from self-limiting acute hepatitis to active chronic hepatitis, cirrhosis and hepatocellular carcinoma. The present study characterizes HBV strains circulating among patients with acute (AHB) and chronic HBV infection (CHB). Among a total of 653 HBsAg positive cases, 40 manifested acute infection. After sequencing the surface(S), basal core promoter/pre-core(BCP/PC) and the X gene regions, phylogenetic tree was constructed using MEGA4 by neighbor-joining method. Statistical robustness was established with bootstrap analysis. Nucleotide diversity was determined by Shannon entropy per site using the Entropy program of the Los Alamos National Laboratories. Analyses of acute patients revealed that HBV/D2 is the major circulating sub-genotype and commonly associated with sexual promiscuity and the age group between15-30 years. Comparison of AHB and CHB patients revealed that HBeAg positivity, ALT levels and genotype D were significantly high in AHB, whereas CHB patients were predominantly male, had a high viral load, and were commonly associated with genotype C. The frequencies of mutations in the S, BCP/PC, and X gene were low in AHB as compared to CHB. Drug resistant mutations were not detectable in the polymerase gene of AHB. Average nucleotide diversity in AHB was considerably low as compared to CHB. Further, the highest average ΔH (average difference in entropy between chronic and acute infection) was observed in the BCP/PC region implying that this region was most vulnerable to mutations upon HBV persistence, especially in case of genotype C. Additionally, among all substitutions, the A1762T and G1764A BCP mutations were the strongest indicators of chronicity. In conclusion, the study exhibits a general portrait of HBV strains circulating among acute hepatitis B patients in Eastern India and their intricate differences with chronic patients which should be useful from the clinical point of view.     

Structural organization of the toe pads in the amphibian Philautus annandalii (Boulenger, 1906)

We describe the morphology of toe pads in the Himalayan tree frog Philautus annandalii. These are expanded tips of digits and show modifications of their ventral epidermis for adhesion. The outer cells of toe pad epidermis (TPE) bear surface microstructures (0.7 × 0.2 μm), which are keratinized. Their cytoplasm contains no organelles, but pleomorphic nuclei and mucous granules (0.4–0.5 μm) that glue the keratin filaments. In the intermediate cell layer of TPE, similar keratinized microstructures as in the outer cells are present, so that when the outer layer is shed, it is ready with features for adhesion. These cells contain more keratin than the outer cells. The basal cell layer contains thin keratin bundles and usual cell organelles. The dermis contains mucous‐secreting glands, whose ducts open in the outer epidermal cell layer in channels. The dorsal epidermal cells lack surface microstructures and keratin bundles. Ultrastructural features suggest that toe pads utilize the surface microstructures for adhesion aided by mucus, in which the intermediate cell layer seems to bear the shear stress generated during locomotion. Further, TPE can expand and fit into an increased contact area of the substrate. The long, surface microstructures may also help in mechanical interlocking with rough surfaces on plants.     

ZnCl2-Modified Activated Carbon from Biomass Coir Pith for the Removal of 2-Chlorophenol by Adsorption Process

Coir pith, a waste biomass, from coconut coir industry was used to prepare activated carbon with ZnCl₂ and employed for the removal of 2-chlorophenol (2-CP) from aqueous solution and wastewater. Zinc chloride–activated coir pith carbon (ZnCPC) was prepared by mixing coir pith with ZnCl₂ in the ratio 2:1 and carbonized at 700°C for 1 h. ZnCPC was characterized using standard physicochemical methods, Brunaver, Emmett, and Teller (BET) surface area, and scanning electron microscopy (SEM) studies. Batch mode adsorption studies were carried out to evaluate the effect of contact time, initial concentration, adsorbent dose, pH, and temperature. The Langmuir adsorption capacity Q ₀ was found to be 149.3 mg g^?1. Kinetic studies showed that the adsorption obeyed second-order and Bangham's model. Equilibrium adsorption data fit better into Langmuir, Freundlich, and D-R isotherms. pH effect and desorption studies showed that ion-exchange mechanism was involved in the adsorption process. Effect of temperature was not significant. Quantitative removal of 2-CP from synthetic wastewater was also achieved. ZnCPC is economically effective compared to commercial activated carbon, because the raw material is abundantly and freely available and it can be used effectively in the treatment of water contaminated with 2-CP.     

Identification,purification and partial characterization of a 70 kDa inhibitor protein of Na+/K+-ATPase from cytosol of pulmonary artery smooth muscle

AimsWe sought to identify, purify and partially characterize a protein inhibitor of Na⁺/K⁺-ATPase in cytosol of pulmonary artery smooth muscle.Main methods(i) By spectrophotometric assay, we identified an inhibitor of Na⁺/K⁺-ATPase in cytosolic fraction of pulmonary artery smooth muscle; (ii) the inhibitor was purified by a combination of ammonium sulfate precipitation, diethylaminoethyl (DEAE) cellulose chromatography, hydroxyapatite chromatography and gel filtration chromatography; (iii) additionally, we have also purified Na⁺/K⁺-ATPase α₂β₁ and α₁β₁ isozymes for determining some characteristics of the inhibitor.Key findingsWe identified a novel endogenous protein inhibitor of Na⁺/K⁺-ATPase having an apparent mol mass of ～ 70 kDa in the cytosolic fraction of the smooth muscle. The IC₅₀ value of the inhibitor towards the enzyme was determined to be in the nanomolar range. Important characteristics of the inhibitor are as follows: (i) it showed different affinities toward the α₂β₁ and α₁β₁ isozymes of the Na⁺/K⁺-ATPase; (ii) it interacted reversibly to the E₁ site of the enzyme; (iii) the inhibitor blocked the phosphorylated intermediate formation; and (iv) it competitively inhibited the enzyme with respect to ATP. CD studies indicated that the inhibitor causes an alteration of the conformation of the enzyme. The inhibition study also suggested that the DHPC solubilized Na⁺/K⁺-ATPase exists as (αβ)₂ diprotomer.SignificanceThe inhibitor binds to the Na⁺/K⁺-ATPase at a site different from the ouabain binding site. The novelty of the inhibitor is that it acts in an isoform specific manner on the enzyme, where α₂ is more sensitive than α₁.     

Non-enzymatic glycation induces structural modifications of myoglobin

Increased glucose concentration in diabetes mellitus causes glycation of several proteins, leading to changes in their properties. Although glycation-induced functional modification of myoglobin is known, structural modification of the protein has not yet been reported. Here, we have studied glucose-modified structural changes of the heme protein. After in vitro glycation of metmyoglobin (Mb) by glucose at 25°C for 6 days, glycated myoglobin (GMb) and unchanged Mb have been separated by ion exchange (BioRex 70) chromatography, and their properties have been compared. Compared to Mb, GMb exhibits increased absorbance around 280 nm and enhanced fluorescence emission with excitation at 285 nm. Fluorescence quenching experiments of the proteins by acrylamide and KI indicate that more surface accessible tryptophan residues are exposed in GMb. CD spectroscopic study reveals a change in the secondary structure of GMb with decreased α-helix content. 1-anilino-naphthaline-8-sulfonate (ANS) binding with Mb and GMb indicates that glycation increases hydrophobicity of the heme protein. GMb appears to be less stable with respect to thermal denaturation and differential calorimetry experiments. Heme-globin linkage becomes weaker in GMb, as shown by spectroscopic and gel electrophoresis experiments. A correlation between glycation-induced structural and functional modifications of the heme protein has been suggested.     

Ethics of care and HIV: a case for rural women in India

Recent literature shows that ethics of care can be used as a theoretical basis to add a new, important dimension to social issues. This paper argues for a similar extension of the theoretical support from ethics of care to an area in bioethics. Specifically, it contends that a justification based ethics of care can be constructed to argue for a moral obligation to give some priority in the HIV-related initiatives to one of most vulnerable groups; namely, the rural women in India. In an epidemic situation this care-based approach has certain advantages as a moral justification over the usual traditional approaches.     

Identification, purification, and characterization of a secretory serine protease in an Indian strain of Leishmania donovani

An aprotinin sensitive serine protease was identified in the culture supernatant of the Indian strain of Leishmania donovani (MHOM/IN/1983/AG83). The protease was subsequently purified and characterized. The apparent molecular mass of the enzyme was 115 kDa in SDS-PAGE under non-reducing condition, while on reduction it showed a 56 kDa protein band indicating that the protease is a dimeric protein. The purified enzyme was optimally active at the pH and temperature of 7.5 and 28 degrees C, respectively. Assays of thermal stability indicated that the enzyme preserved 59% of activity even after pretreatment at 42 degrees C for 1 h. The purified protease was not glycosylated and its isoelectric pI was 5.0. N-alpha-p-tosyl-L-arginine methylester (TAME) appeared to be relatively better substrate among the commonly used synthetic substrates. The enzyme was inhibited by Ca(2+) and Mn(2+), but activated by Zn(2+). The protease could play important role(s) in the pathogenesis of visceral leishmaniasis or kala-azar.     

Role of Ca2+-dependent metalloprotease-2 in stimulating Ca2+ ATPase activity under peroxynitrite treatment in bovine pulmonary artery smooth muscle membrane

We have determined effect of the oxidant peroxynitrite (ONOO-) on Ca2+-dependent matrix metalloprotease-2 (MMP-2) activity and the role of the protease on Ca2+ ATPase activity in bovine pulmonary vascular smooth muscle plasma membrane under ONOO- -triggered conditions. The smooth muscle plasma membrane possesses a 72-kDa protease activity in a gelatin-containing zymogram. The 72-kDa protease activity has been found to be inhibited by tissue inhibitor of metalloprotease-2 (TIMP-2), indicating that the protease is the matrix metalloprotease-2 (MMP-2). Treatment of the membrane suspension with ONOO- caused stimulation of the MMP-2 activity (as evidenced by 14C-gelatin degradation) and also increased Ca2+ ATPase activity. The ONOO- -triggered protease activity and the Ca2+ ATPase activity were found to be inhibited by the antioxidants: vitamin E, thiourea, and mannitol. Pretreatment with catalase and superoxide dismutase did not significantly alter ONOO- -stimulated MMP-2 activity and Ca2+ATPase activity, indicating that peroxide and superoxide are not present in appreciable amount in ONOO-. Under both basal and ONOO- triggered conditions, the MMP-2 activity and the Ca2+ ATPase activity were also inhibited by EGTA, 1:10-phenanthroline, and TIMP-2. However, the ONOO- -stimulated MMP-2 activity and the Ca2+ ATPase activity were found to be insensitive to phenylmethylsulfonylfluoride, Bowman-Birk inhibitor, chymostatin, leupeptin, antipain, N-ethylmaleimide, and pepstatin. These results suggest that ONOO- caused stimulation of MMP-2 activity and that the increased MMP-2 activity subsequently played a pivotal role in stimulating Ca2+ ATPase activity in bovine pulmonary vascular smooth muscle plasma membrane.