A steady-state electrochemical model of vascular smooth muscle cells

A model of the steady-state electrochemical response of vascular smooth muscle cells to external stimuli is presented, which accounts for K, Na, and Ca fluxes. The results of the model are broadly in accordance with experimental data 1), at various transmural pressures; 2), with channel and pump blockade; and 3), under manipulation of external ionic concentrations. The model exhibits dual stable states which sometimes coexist, and abrupt transitions between these states may account for nongraded responses in arteries as external potassium or pressure is varied. The simulations suggest that changes in the intracellular sodium concentration ([Na]i) often accompany smooth muscle responses. For example, [Na]i values vary threefold over the range of pressures from 10 to 100 mmHg.     

Fusarium proliferatum, an endophytic fungus from Dysoxylum binectariferum Hook.f, produces rohitukine, a chromane alkaloid possessing anti-cancer activity

Rohitukine is a chromane alkaloid possessing anti-inflammatory, anti-cancer and immuno-modulatory properties. The compound was first reported from Amoora rohituka (Meliaceae) and later from Dysoxylum binectariferum (Meliaceae) and Schumanniophyton problematicum (Rubiaceae). Flavopiridol, a semi-synthetic derivative of rohitukine is a potent CDK inhibitor and is currently in Phase III clinical trials. In this study, the isolation of an endophytic fungus, Fusarium proliferatum (MTCC 9690) from the inner bark tissue of Dysoxylum binectariferum Hook.f (Meliaceae) is reported. The endophytic fungus produces rohitukine when cultured in shake flasks containing potato dextrose broth. The yield of rohitukine was 186 μg/100 g dry mycelial weight, substantially lower than that produced by the host tissue. The compound from the fungus was authenticated by comparing the LC–HRMS and LC–HRMS/MS spectra with those of the reference standard and that produced by the host plant. Methanolic extract of the fungus was cytotoxic against HCT-116 and MCF-7 human cancer cell lines (IC₅₀ = 10 μg/ml for both cancer cell lines).     

Simple, rapid and cost-effective method for high quality nucleic acids extraction from different strains of Botryococcus braunii

This study deals with an effective nucleic acids extraction method from various strains of Botryococcus braunii which possesses an extensive extracellular matrix. A method combining freeze/thaw and bead-beating with heterogeneous diameter of silica/zirconia beads was optimized to isolate DNA and RNA from microalgae, especially from B. braunii. Eukaryotic Microalgal Nucleic Acids Extraction (EMNE) method developed in this study showed at least 300 times higher DNA yield in all strains of B. braunii with high integrity and 50 times reduced working volume compared to commercially available DNA extraction kits. High quality RNA was also extracted using this method and more than two times the yield compared to existing methods. Real-time experiments confirmed the quality and quantity of the input DNA and RNA extracted using EMNE method. The method was also applied to other eukaryotic microalgae, such as diatoms, Chlamydomonas sp., Chlorella sp., and Scenedesmus sp. resulting in higher efficiencies. Cost-effectiveness analysis of DNA extraction by various methods revealed that EMNE method was superior to commercial kits and other reported methods by >15%. This method would immensely contribute to area of microalgal genomics.     

The Socioeconomic and Institutional Determinants of Participation in India’s Health Insurance Scheme for the Poor

The Rashtriya Swasthya Bima Yojana (RSBY), which was introduced in 2008 in India, is a social health insurance scheme that aims to improve healthcare access and provide financial risk protection to the poor. In this study, we analyse the determinants of participation and enrolment in the scheme at the level of districts. We used official data on RSBY enrolment, socioeconomic data from the District Level Household Survey 2007–2008, and additional state-level information on fiscal health, political affiliation, and quality of governance. Results from multivariate probit and OLS analyses suggest that political and institutional factors are among the strongest determinants explaining the variation in participation and enrolment in RSBY. In particular, districts in state governments that are politically affiliated with the opposition or neutral parties at the centre are more likely to participate in RSBY, and have higher levels of enrolment. Districts in states with a lower quality of governance, a pre-existing state-level health insurance scheme, or with a lower level of fiscal deficit as compared to GDP, are significantly less likely to participate, or have lower enrolment rates. Among socioeconomic factors, we find some evidence of weak or imprecise targeting. Districts with a higher share of socioeconomically backward castes are less likely to participate, and their enrolment rates are also lower. Finally, districts with more non-poor households may be more likely to participate, although with lower enrolment rates.     

Energy dissipation mechanisms in the FCPb light-harvesting complex of the diatom Cyclotella meneghiniana

Transient absorption spectroscopy has been applied to investigate the energy dissipation mechanisms in the nonameric fucoxanthin-chlorophyll-a,c-binding protein FCPb of the centric diatom Cyclotella meneghiniana. FCPb complexes in their unquenched state were compared with those in two types of quenching environments, namely aggregation-induced quenching by detergent removal, and clustering via incorporation into liposomes. Applying global and target analysis, in combination with a fluorescence lifetime study and annihilation calculations, we were able to resolve two quenching channels in FCPb that involve chlorophyll-a pigments for FCPb exposed to both quenching environments. The fast quenching channel operates on a timescale of tens of picoseconds and exhibits similar spectral signatures as the unquenched state. The slower quenching channel operates on a timescale of tens to hundreds of picoseconds, depending on the degree of quenching, and is characterized by enhanced population of low-energy states between 680 and 710?nm. The results indicate that FCPb is, in principle, able to function as a dissipater of excess energy and can do this in vitro even more efficiently than the homologous FCPa complex, the sole complex involved in fast photoprotection in these organisms. This indicates that when a complex displays photoprotection-related spectral signatures in vitro it does not imply that the complex participates in photoprotection in vivo. We suggest that FCPa is favored over FCPb as the sole energy-regulating complex in diatoms because its composition can more easily establish the balance between light-harvesting and quenching required for efficient photoprotection.     

Purification and characterization of a novel plant-type carbonic anhydrase from Bacillus subtilis

Carbonic anhydrase enzyme, one of the fastest known enzymes, remains largely unexplored in prokaryotes when compared to its mammalian counterparts despite its ubiquity. In this study, the enzyme has been purified from Bacillus subtilis SA3 using sequential Sephadex G-75 chromatography, DEAE cellulose chromatography, and sepharose-4B-L-tyrosinesulphanilamide affinity chromatography and characterized to provide additional insights into its properties. The apparent molecular mass of carbonic anhydrase obtained by SDS-PAGE was found to be approximately 37 kDa. Isoelectric focusing of the purified enzyme revealed an isoelectric point (pI) of around 6.1 when compared with marker. The presence of metal ions such as Zn²⁺, Co²⁺, Cu²⁺, Fe³⁺, Mg²⁺, and anion SO₄⁻ increased enzyme activity while strong inhibition was observed in the presence of Hg²⁺, Cl⁻, HCO₃⁻, and metal chelator EDTA. The optimum pH and temperature for the enzyme were found to be 8.3 and 37°C, respectively. Enzyme kinetics with p-nitrophenyl acetate as substrate at pH 8.3 and 37°C determined the V_max and K_m values of the enzyme to be 714.28 μmol/mg protein/min and 9.09 mM, respectively. The K_i value for acetazolamide was 0.22 mM, compared to 0.099 mM for sulphanilamide. The results from N-terminal amino acid sequencing imply the purified protein is a putative beta-carbonic anhydrase with close similarities to CAs from plants, microorganisms.