Impaired Surfactant Production by Alveolar Epithelial Cells in a SCID-hu Lung Mouse Model of Congenital Human Cytomegalovirus Infection

Human cytomegalovirus (HCMV) is the leading viral cause of birth defects and life-threatening lung-associated diseases in premature infants and immunocompromised children. Although the fetal lung is a major target organ of the virus, HCMV lung pathogenesis has remained unexplored, possibly as a result of extreme host range restriction. To overcome this hurdle, we generated a SCID-hu lung mouse model that closely recapitulates the discrete stages of human lung development in utero. Human fetal lung tissue was implanted into severe combined immunodeficient (CB17-scid) mice and inoculated by direct injection with the VR1814 clinical isolate of HCMV. Virus replication in the fetal lung was assessed by the quantification of infectious virus titers and HCMV genome copies and the detection of HCMV proteins by immunohistochemistry and Western blotting. We show that HCMV efficiently replicated in the lung implants during a 2-week period, forming large viral lesions. The virus productively infected alveolar epithelial and mesenchymal cells, imitating congenital infection of the fetal lung. HCMV replication triggered apoptosis near and within the viral lesions and impaired the production of surfactant proteins in the alveolar epithelium. Our findings highlight that congenital and neonatal HCMV infection can adversely impact lung development, leading to pneumonia and acute lung injury. We have successfully developed a small-animal model that closely recapitulates fetal and neonatal lung development and provides a valuable, biologically relevant tool for an understanding of the lung pathogenesis of HCMV as well as other human respiratory viruses. Additionally, this model would greatly facilitate the development and testing of new antiviral therapies for HCMV along with select human pulmonary pathogens.     

Design,green synthesis and pharmacological evaluation of novel 5,6-diaryl-1,2,4-triazines bearing 3-morpholinoethylamine moiety as potential antithrombotic agents*

The aim of this research work was to investigate a series of novel 5,6-diaryl-1,2,4-triazines (3a–3q) containing 3-morpholinoethylamine side chain, and to address their antiplatelet activity by in vitro, ex vivo and in vivo methods. All compounds were synthesized by environment benign route and their structures were unambiguously confirmed by spectral data. Compounds (3l) and (3m) were confirmed by their single crystal X-ray structures. Out of all the synthesized compounds, 10 were found to be more potent in vitro than aspirin; six of them were found to be prominent in ex vivo assays and one compound (3d) was found to have the most promising antithrombotic profile in vivo. Moreover, compound (3d) demonstrated less ulcerogenicity in rats as compared to aspirin. The selectivity of the most promising compound (3d) for COX-1 and COX-2 enzymes was determined with the help of molecular docking studies and the results were correlated with the biological activity.     

Lactoferrin-melanin interaction and its possible implications in melanin polymerization: crystal structure of the complex formed between mare lactoferrin and melanin monomers at 2.7-A resolution

The concentration of melanin determines the intensity of colors of the skin and hair of animals. Melanin pigments are tyrosine-based polymers formed in melanocytes within specialized organelles called melanosomes. In order to understand the mechanism of melanin polymerization, lactoferrin, a basic protein with a pI value of 9.0, has been used to produce melanin. Lactoferrin is a monomeric iron-binding protein with a molecular weight of 80 kDa. The crystals of lactoferrin were soaked in a solution containing dihydroxyphenylalanine (DOPA) and tyrosinase enzyme. These crystals were used for X-ray intensity data collection. The intensity data were collected to 2.7-A resolution to an overall completeness of 91% with an R(sym) of 0.071. The crystals belong to orthorhombic space group P2(1)2(1)2(1) with cell dimensions: a = 85.0 A, b = 99.8 A, c = 103.4 A. The structure was determined by molecular replacement method, using the model of diferric mare lactoferrin, and refined to an R-factor 0.215 (R(free) = 0.287) for all the data to 2.7-A resolution. The final model comprises 5,281 protein atoms from 689 amino acids, 2Fe(3+), 2CO(2-)(3) ions, 2 indole-5,6-quinone molecules (IQ), and 73 water molecules. Two IQ molecules, one in each lobe, bind to lactoferrin. In the C-lobe, the IQ binds in the iron-binding cleft, whereas in the N-lobe, it is located in the side pocket between two alpha-helices, filled with solvent molecules in the native iron-saturated mare lactoferrin. The IQ molecules interact with protein molecule mainly through glutamic acid in both lobes, without significant perturbation to the protein structure. The orientation of N- and C-lobes in the present structure is similar to that observed in the native iron-saturated protein. However, as a result of the binding of IQ molecules, the orientations of the domains N1, N2 and C1, C2 in the two cases differ slightly.     

Synthesis of a new 4-aza-2,3-didehydropodophyllotoxin analogues as potent cytotoxic and antimitotic agents

A series of novel conjugates of 4-aza-2,3-didehydropodophyllotoxins (11a-w) were synthesized by a straightforward one-step multicomponent synthesis that demonstrated cytotoxicity against five human cancer cell lines (breast, oral, colon, lung and ovarian). All the twenty three compounds (11a-w) have been examined for the inhibition of tubulin polymerization. Among these compounds, 11a, 11k and 11p exhibited inhibition of polymerization tubulin comparable to podophyllotoxin apart from disruption of microtubule organization within the cells. Flow cytometric analysis showed that these compounds (11a, 11k and 11p) arrested the cell cycle in the G2/M phase of cell cycle leading to caspase-3 dependent apoptotic cell death.     

Reduction of 8-iso-prostaglandin F2α in the first week after Roux-en-Y gastric bypass surgery

Obesity is associated with increased markers of oxidative stress. We examined whether oxidative stress is reduced within the first week after Roux‐en‐Y gastric bypass (RYGB) surgery and could be related to changes in adipose tissue depots. The reactive oxygen species (ROS) marker 8‐iso‐prostaglandin F2α (8‐iso‐PGF2α) and activity of antioxidant glutathione peroxidases (GPX) in plasma were compared before and ～1 week after RYGB. The effects of RYGB on subcutaneous adipose tissue and interstitial fluid 8‐iso‐PGF2α levels and subcutaneous adipose tissue expression of GPX‐3 were also assessed. Levels of 8‐iso‐PGF2α in subcutaneous and visceral adipose tissue were determined. Plasma 8‐iso‐PGF2α levels decreased (122 ± 75 to 56 ± 15 pg/ml, P = 0.001) and GPX activity increased (84 ± 18 to 108 ± 25 nmol/min/ml, P = 0.003) in the first week post‐RYGB. RYGB also resulted in reductions of 8‐iso‐PGF2α in subcutaneous adipose tissue (1,742 ± 931 to 1,132 ± 420 pg/g fat, P = 0.046) and interstitial fluid (348 ± 118 to 221 ± 83 pg/ml, P = 0.046) that were comparable to plasma (26–33%, P = 0.74). Adipose GPX‐3 expression was increased (6.7 ± 4.7‐fold, P = 0.004) in the first postoperative week. The improvements in oxidative stress occurred with minimal weight loss (2.4 ± 3.4%, P = 0.031) and elevations in plasma interleukin‐6 (18.0 ± 46.8 to 28.0 ± 58.9 pg/ml, P = 0.004). Subcutaneous and visceral adipose tissues express comparable 8‐iso‐PGF2α levels (1,204 ± 470 and 1,331 ± 264 pg/g fat, respectively; P = 0.34). These data suggest that RYGB affects adipose tissue leading to the restoration of adipose redox balance within the first postoperative week and that plasma 8‐iso‐PGF2α is primarily derived from subcutaneous adipose tissue.     

Photoelectrochemical Behavior of Planar and Microwire‐Array Si|GaP Electrodes

Gallium phosphide exhibits a short diffusion length relative to its optical absorption length, and is thus a candidate for use in wire array geometries that allow light absorption to be decoupled from minority carrier collection. Herein is reported the photoanodic performance of heteroepitaxially grown gallium phosphide on planar and microwire‐array Si substrates. The n‐GaP|n‐Si heterojunction results in a favorable conduction band alignment for electron collection in the silicon. A conformal electrochemical contact to the outer GaP layer is produced using the ferrocenium/ferrocene (Fc⁺/Fc) redox couple in acetonitrile. Photovoltages of ～750 mV under 1 sun illumination are observed and are attributed to the barrier formed at the (Fc⁺/Fc)|n‐GaP junction. The short‐circuit current densities of the composite microwire‐arrays are similar to those observed using single‐crystal n‐GaP photoelectrodes. Spectral response measurements along with a finite‐difference‐time‐domain optical model indicate that the minority carrier diffusion length in the GaP is ～80 nm. Solid‐state current–voltage measurements show that shunting occurs through thin GaP layers that are present near the base of the microwire‐arrays. The results provide guidance for further studies of 3D multi‐junction photoelectrochemical cells.     

Interaction of melanin with proteins--the importance of an acidic intramelanosomal pH.

Melanin is a highly irregular heteropolymer consisting of monomeric units derived from the enzymatic oxidation of the amino acid tyrosine. The process of melanin formation takes place in specialized acidic organelles (melanosomes) in melanocytes. The process of melanin polymerization requires an alkaline pH in vitro, and therefore, the purpose of an acidic environment in vivo remains a mystery. It is known that melanin is always bound to protein in vivo. It is also seen that polymerization in vitro at an acidic pH necessarily requires the presence of proteins. The effect of various model proteins on melanin synthesis and their interaction with melanin was studied. It was seen that many proteins could increase melanin synthesis at an acidic pH, and that different proteins resulted in the formation of different states of melanin, i.e., a precipitate or a soluble, protein-bound form. We also present evidence to show that soluble protein-bound melanin is present in vivo (in B16 cells as well as in B16 melanoma tissue). An acidic pH appeared to be necessary to ensure the formation of a uniform, very high molecular weight melano-protein complex. The interaction between melanin and proteins appears to be largely charge-dependent as evidenced by zeta potential measurements, and this interaction is also increased in an acidic pH. Thus, it appears that an acidic intramelanosomal pH is essential to ensure maximum interaction between protein and melanin, and also to ensure that all the melanin formed is protein-bound.     

Eco-friendly biodegradation of a reactive textile dye Golden Yellow HER by Brevibacillus laterosporus MTCC 2298

Brevibacillus laterosporus MTCC 2298 showed 87% decolorization of Golden Yellow HER within 48 h under static condition at the concentration 50 mg l^?1; however no significant change in the decolorization performance was observed under shaking condition. Decolorization performance was maximum (74%) at the pH 7.0 and 30 °C. TLC and HPLC analysis confirmed the biodegradation of Golden Yellow HER. Biodegradation pathway was proposed using GC–MS and FTIR spectral analysis. Mainly elected metabolites are the 2,5-Dichloro-4 (3-hydrazino-2-hydroxy cyclopentylamino-) dibenzene-sulfonic acid (peak 1, m/z = 526), 4-(3-hydrazino-2-hydroxy cyclopentylamino)-benzene-sulfonic acid (peak 2, m/z = 455), 4-(3-amino-2-hydroxy-cyclopentylamino)-benzene-sulfonic acid and 5-amino-cyclohex-3-ene-sulfonic acid (peak 3, m/z = 183). Phytotoxicity results suggested that degradation products of Golden Yellow HER are non-toxic to the common crops such as Sorghum vulgare and Phaseolus mungo. Also, degradation products are non-toxic to B. laterosporus as well as ecologically important bacteria like Pseudomonas aeruginosa and Azotobacter vinelandii.