Effect of low-level laser therapy on experimental wounds of hard palate mucosa in mice

Under general anesthesia and sterile conditions, incision wound was induced in the hard palate mucosa of adult male mice. The wounds of groups 1 and 2 were irradiated daily with He-Ne laser at 3 and 7.5 J/cm2 for 120 and 300 s, respectively, while the incision wound of group 3 not exposed served as controls. On day 3 of injury, the laser-treated wounds contained significantly lower neutrophils than the wounds in the control group. By day 7 after injury, the laser-treated wounds contained significantly more fibroblasts and at the same time contained significantly fewer macrophages. In conclusion, an acceleration of the wound healing process of experimental wounds in the hard palate mucosa of mice at low-level laser therapy with a He-Ne laser at energy densities of 3 and 7.5 J/cm2 was observed.     

Constraints on the evolution of toxin-resistant Na,K-ATPases have limited dependence on sequence divergence

A growing body of theoretical and experimental evidence suggests that intramolecular epistasis is a major determinant of rates and patterns of protein evolution and imposes a substantial constraint on the evolution of novel protein functions. Here, we examine the role of intramolecular epistasis in the recurrent evolution of resistance to cardiotonic steroids (CTS) across tetrapods, which occurs via specific amino acid substitutions to the α-subunit family of Na,K-ATPases (ATP1A). After identifying a series of recurrent substitutions at two key sites of ATP1A that are predicted to confer CTS resistance in diverse tetrapods, we then performed protein engineering experiments to test the functional consequences of introducing these substitutions onto divergent species backgrounds. In line with previous results, we find that substitutions at these sites can have substantial background-dependent effects on CTS resistance. Globally, however, these substitutions also have pleiotropic effects that are consistent with additive rather than background-dependent effects. Moreover, the magnitude of a substitution’s effect on activity does not depend on the overall extent of ATP1A sequence divergence between species. Our results suggest that epistatic constraints on the evolution of CTS-resistant forms of Na,K-ATPase likely depend on a small number of sites, with little dependence on overall levels of protein divergence. We propose that dependence on a limited number sites may account for the observation of convergent CTS resistance substitutions observed among taxa with highly divergent Na,K-ATPases (See S1 Text for Spanish translation).     

Phenolics Impart Au3+-Stress Tolerance to Cowpea by Generating Nanoparticles

While evaluating impact of Au nanoparticles on seed germination and early seedling growth of cowpea, HAuCl₄ was used as control. Seedlings of cowpea raised in HAuCl₄, even at concentration as high as 1 mM, did not show any suppression in growth. Accordingly, Au³⁺, despite being a heavy metal, did not alter levels of stress markers (viz. proline and malondialdehyde) in cowpea. Interestingly, cowpea turned clear pale yellow HAuCl₄ solutions colloidal purple during the course of seed germination and seedling growth. These purple colloidal suspensions showed Au-nanoparticle specific surface plasmon resonance band in absorption spectra. Transmission electron microscopic and powder X-ray diffraction investigations confirmed presence of crystalline Au-nanoparticles in these purple suspensions. Each germinating seed of cowpea released ∼35 nmoles of GAE of phenolics and since phenolics promote generation of Au-nanoparticles, which are less/non toxic compared to Au³⁺, it was contemplated that potential of cowpea to withstand Au³⁺ is linked to phenolics. Of the different components of germinating seed of cowpea tested, seed coat possessed immense power to generate Au-nanoparticles, as it was the key source of phenolics. To establish role of phenolics in generation of Au-nanoparticles (i) seed coat and (ii) the incubation medium in which phenolics were released by germinating seeds, were tested for their efficacy to generate Au-nanoparticles. Interestingly, incubation of either of these components with Au³⁺ triggered increase in generation of Au-nanoparticles with concomitant decrease in phenolics. Accordingly, with increase in concentration of Au³⁺, a proportionate increase in generation of Au-nanoparticles and decrease in phenolics was recorded. In summary, our findings clearly established that cowpea possessed potential to withstand Au³⁺-stress as the phenolics released by seed coat of germinating seeds possess potential to reduce toxic Au³⁺ to form non/less toxic Au-nanoparticles. Our investigations also pave a novel, simple, green and economically viable protocol for generation of Au-nanoparticles.     

Targeting the microRNA binding domain of argonaute 2: rational inhibitor design and study of mutation effects on protein-ligand interaction

Abstract

Based on the accumulative evidences during recent decades, miRNAs have been found overexpressed in several human cancer types and also in Down syndrome patients, contributing to the neuropathology of Down syndrome. From this point of view, investigations on the structure and dynamic mechanisms related to the Argonaute 2 miRNAs binding in which silencing of the mRNA occurs, have inspired many clinical researchers to target this complex to inhibit the silencing process. In the current research, we have virtually screened the OTAVA_CNS_library to introduce new inhibitor compounds for the Ago2/miRNA complex. Ten hit compounds were obtained, with just one of them nominated as the best compound. Following the interaction analysis, by utilizing molecular dynamics (MD) simulations, effects of two mutations (Thr526 to isoleucine and Gln545 to alanine) on the dynamic properties of Ago2 in the complex with the best inhibitor compound were investigated. RMSD, RMSF and h-bond number beside other analyses, highlighted the importance of the Thr526 and Gln545 mutations for the stability and flexibility of the (Ago2)/ligand complex.

Communicated by Ramaswamy H. Sarma     

Generation of new human embryonic stem cell lines with diploid and triploid karyotypes

Human pluripotent embryonic stem cells (hESC) have great promise for research into human developmental biology and the development of cell therapies for the treatment of diseases. To meet the increased demand for characterized hESC lines, we present the derivation and characterization of five hESC lines on mouse embryonic fibroblast cells. Our stem cell lines are characterized by morphology, long-term expansion, and expression profiles of a number of specific markers, including TRA-1-60, TRA-1-81, alkaline phosphatase, connexin 43, OCT-4, NANOG, CXCR4, NODAL, LEFTY2, THY-1, TDGF1, PAX6, FOXD3, SOX2, EPHA2, FGF4, TAL1, AC133 and REX-1. The pluripotency of the cell line was confirmed by spontaneous differentiation under in vitro conditions. Whereas all of the cell lines expressed all the characteristics of undifferentiated pluripotent hESC, two of the cell lines carried a triploid karyotype.     

FLT3-ITDs Instruct a Myeloid Differentiation and Transformation Bias in Lymphomyeloid Multipotent Progenitors

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Nitric oxide generation from heme/copper assembly mediated nitrite reductase activity

Nitric oxide (NO) as a cellular signaling molecule and vasodilator regulates a range of physiological and pathological processes. Nitrite (NO₂ ^?) is recycled in vivo to generate nitric oxide, particularly in physiologic hypoxia and ischemia. The cytochrome c oxidase binuclear heme a ₃/Cu_B active site is one entity known to be responsible for conversion of cellular nitrite to nitric oxide. We recently reported that a partially reduced heme/copper assembly reduces nitrite ion, producing nitric oxide; the heme serves as the reductant and the cupric ion provides a Lewis acid interaction with nitrite, facilitating nitrite (N–O) bond cleavage (Hematian et al., J. Am. Chem. Soc. 134:18912–18915, 2012). To further investigate this nitrite reductase chemistry, copper(II)–nitrito complexes with tridentate and tetradentate ligands were used in this study, where either O,O′-bidentate or O-unidentate modes of nitrite binding to the cupric center are present. To study the role of the reducing ability of the ferrous heme center, two different tetraarylporphyrinate–iron(II) complexes, one with electron-donating para-methoxy peripheral substituents and the other with electron-withdrawing 2,6-difluorophenyl substituents, were used. The results show that differing modes of nitrite coordination to the copper(II) ion lead to differing kinetic behavior. Here, also, the ferrous heme is in all cases the source of the reducing equivalent required to convert nitrite to nitric oxide, but the reduction ability of the heme center does not play a key role in the observed overall reaction rate. On the basis of our observations, reaction mechanisms are proposed and discussed in terms of heme/copper heterobinuclear structures.     

DEN+2-AAF-induced multistep hepatotumorigenesis in Wistar rats: supportive evidence and insights

Diethylnitrosamine (DEN), found in many commonly consumed foods, has been reported to induce cancers in animals and humans. Several models have been developed to study multistage carcinogenesis in rat liver; these include the Solt–Farber-resistant hepatocyte model. In the Solt–Farber model, the initiation consists of either a necrogenic dose of a hepatocarcinogen or a non-necrogenic dose in conjunction with partial hepatectomy (PH). We report a novel protocol for tumor induction in liver which eliminates the need for PH. Male Wistar rats were injected with single i.p. dose of DEN (200 mg/kg body weight), controls received saline only. After 1 week of recovery, the DEN-treated animals were administered with the repeated doses of 2-acetyamino fluorine (150 mg/kg body weight) orally in 1 % carboxymethyl cellulose that served as promoting agent. Thirty days after the DEN administration, hepatocellular damage was observed as evident by histopathological analysis. The marker enzyme analysis showed elevated levels of serum AST, ALT, and alkaline phosphatase and a decrease in the levels of liver superoxide dismutase and catalase. The oxidative stress in liver was confirmed by elevated levels of lipid peroxidation and a decrease in antioxidant parameters.     

High Level Activity of Recombinant Lysostaphin After Computer Simulation and Additive-Based Refolding

International Journal of Peptide Research and Therapeutics - Lysostaphin is a peptidoglycan hydrolase, produced by Staphylococcus simulans, which has illustrated significant bactericidal activities...