Inferences of evolutionary history of a widely distributed mangrove species,Bruguiera gymnorrhiza,in the Indo‐West Pacific region

Inference of genetic structure and demographic history is fundamental issue in evolutionary biology. We examined the levels and patterns of genetic variation of a widespread mangrove species in the Indo‐West Pacific region, Bruguiera gymnorrhiza, using ten nuclear gene regions. Genetic variation of individual populations covering its distribution range was low, but as the entire species it was comparable to other plant species. Genetic differentiation among the investigated populations was high. They could be divided into two genetic clusters: the West and East clusters of the Malay Peninsula. Our results indicated that these two genetic clusters derived from their ancestral population whose effective size of which was much larger compared to the two extant clusters. The point estimate of speciation time between B. gymnorrhiza and Bruguiera sexangula was two times older than that of divergence time between the two clusters. Migration from the West cluster to the East cluster was much higher than the opposite direction but both estimated migration rates were low. The past Sundaland and/or the present Malay Peninsula are likely to prevent gene flow between the West and East clusters and function as a geographical or land barrier.     

Mechanisms of Sodium Transport at the Blood-Brain Barrier Studied with In Situ Perfusion of Rat Brain

Abstract: The mechanism of unidirectional transport of sodium from blood to brain in pentobarbital-anesthetized rats was examined using in situ perfusion. Sodium transport followed Michaelis-Menten saturation kinetics with a V _max of 50.1 nmol/g/min and a K _m of 17.7 m M in the left frontal cortex. The kinetic analysis indicated that, at a physiologic sodium concentration, ∼26% of sodium transport at the blood-brain barrier (BBB) was carrier mediated. Dimethylamiloride (25 µ M ), an inhibitor of Na⁺/H⁺ exchange, reduced sodium transport by 28%, whereas phenamil (25 µ M ), a sodium channel inhibitor, reduced the transfer constant for sodium by 22%. Bumetanide (250 µ M ) and hydrochlorothiazide (1.5 m M ), inhibitors of Na⁺-K⁺-2Cl⁻/NaCl symport, were ineffective in reducing blood to brain sodium transport. Acetazolamide (0.25 m M ), an inhibitor of carbonic anhydrase, did not change sodium transport at the BBB. Finally, a perfusate pH of 7.0 or 7.8 or a perfusate P co ₂ of 86 mm Hg failed to change sodium transport. These results indicate that 50% of transcellular transport of sodium from blood to brain occurs through Na⁺/H⁺ exchange and a sodium channel in the luminal membrane of the BBB. We propose that the sodium transport systems at the luminal membrane of the BBB, in conjunction with Cl⁻/HCO₃⁻ exchange, lead to net NaCl secretion and obligate water transport into the brain.     

Picroside III,an Active Ingredient of Picrorhiza scrophulariiflora,Ameliorates Experimental Colitis by Protecting Intestinal Barrier Integrity

This study aims to explore the protective effects of Picroside III, an active ingredient of Picrorhiza scrophulariiflora, on the intestinal epithelial barrier in tumor necrosis factor-α (TNF-α) induced Caco-2 cells and dextran sulfate sodium (DSS) induced colitis in mice. Results show that Picroside III significantly alleviated clinical signs of colitis including body weight loss, disease activity index increase, colon shortening, and colon tissue damage. It also increased claudin-3, ZO-1 and occludin expressions and decreased claudin-2 expression in the colon tissues of mice with colitis. In vitro, Picroside III also significantly promoted wound healing, decreased the permeability of cell monolayer, upregulated the expressions of claudin-3, ZO-1 and occludin and downregulated the expression of claudin-2 in TNF-α treated Caco-2 cells. Mechanism studies show that Picroside III significantly promoted AMP-activated protein kinase (AMPK) phosphorylation in vitro and in vivo, and blockade with AMPK could significantly attenuate the upregulation of Picroside III in ZO-1 and occludin expressions and the downregulation of claudin-2 expression in TNF-α treated Caco-2 cells. In conclusion, this study demonstrates that Picroside III attenuated DSS-induced colitis by promoting colonic mucosal wound healing and epithelial barrier function recovery via the activation of AMPK.     

Successful synthesis of a glial-specific blood–brain barrier shuttle peptide following a fragment condensation approach on a solid-phase resin

Successful manual synthesis of the TD2.2 peptide acting as a blood–brain barrier shuttle was achieved. TD2.2 was successfully synthesised by sequential condensation of four protected peptide fragments on solid-phase settings, after several unsuccessful attempts using the stepwise approach. These fragments were chosen to minimise the number of demanding amino acids (in terms of coupling, Fmoc removal) in each fragment that are expected to hamper the overall synthetic process. Thus, the hydrophobic amino acids as well as Arg(Pbf) were strategically spread over multiple fragments rather than having them congested in one fragment. This study shows how a peptide that shows big challenges in the synthesis using the common stepwise elongation methodology can be synthesised with an acceptable purity. It also emphasises that choosing the right fragment with certain amino acid constituents is key for a successful synthesis. It is worth highlighting that lower amounts of reagents were required to synthesise the final peptide with an identical purity to that obtained by the automatic synthesiser.     

Heat shock proteins: A dual carrier-adjuvant for an anti-drug vaccine against heroin

Heroin is a highly abused opioid that has reached epidemic status within the United States. Yet, existing therapies to treat addiction are inadequate and frequently result into rates of high recidivism. Vaccination against heroin offers a promising alternative therapeutic option but requires further development to enhance the vaccine’s performance. Hsp70 is a conserved protein with known immunomodulatory properties and is considered an excellent immunodominant antigen. Within an antidrug vaccine context, we envisioned Hsp70 as a potential dual carrier-adjuvant, wherein immunogenicity would be increased by co-localization of adjuvant and antigenic drug hapten. Recombinant Mycobacterium tuberculosis Hsp70 was appended with heroin haptens and the resulting immunoconjugate granted anti-heroin antibody production and blunted heroin-induced antinociception. Moreover, Hsp70 as a carrier protein surpassed our benchmark Her-KLH cocktail through antibody-mediated blockade of 6-acetylmorphine, the main mediator of heroin’s psychoactivity. The work presents a new avenue for exploration in the use of hapten-Hsp70 conjugates to elicit anti-drug immune responses.     

Membrane mediated development of the vertebrate blood‐gas‐barrier

During embryonic lung development, establishment of the gas‐exchanging units is guided by epithelial tubes lined by columnar cells. Ultimately, a thin blood‐gas barrier (BGB) is established and forms the interface for efficient gas exchange. This thin BGB is achieved through processes, which entail lowering of tight junctions, stretching, and thinning in mammals. In birds the processes are termed peremerecytosis, if they involve cell squeezing and constriction, or secarecytosis, if they entail cutting cells to size. In peremerecytosis, cells constrict at a point below the protruding apical part, resulting in fusion of the opposing membranes and discharge of the aposome, or the cell may be squeezed by the more endowed cognate neighbors. Secarecytosis may entail formation of double membranes below the aposome, subsequent unzipping and discharge of the aposome, or vesicles form below the aposome, fuse in a bilateral manner, and release the aposome. These processes occur within limited developmental windows, and are mediated through cell membranes that appear to be of intracellular in origin. In addition, basement membranes (BM) play pivotal roles in differentiation of the epithelial and endothelial layers of the BGB. Laminins found in the BM are particularly important in the signaling pathways that result in formation of squamous pneumocytes and pulmonary capillaries, the two major components of the BGB. Some information exists on the contribution by BM to BGB formation, but little is known regarding the molecules that drive peremerecytosis, or even the origins and composition of the double and vesicular membranes involved in secarecytosis. Birth Defects Research (Part C) 108:85–97, 2016. © 2016 Wiley Periodicals, Inc.     

Quantification of the permeability of the blood-CSF barrier to alpha-MSH in the rat

The ability of alpha-MSH to cross the blood-CSF barrier of the rat was assessed by measurement of the rate of appearance of immunoreactive alpha-MSH in a cerebrospinal fluid (CSF) perfusate following intravenous injection of peptide. Comparisons were made with the rate of appearance of a simultaneously administered dose of 14C-inulin which is poorly permeable at the blood-CSF barrier. Concentrations of drugs measured in plasma were fitted to two-compartment pharmacokinetic models, and those measured in the CSF perfusate to one-compartment open systems receiving an input from the plasma compartment. The rate constant for entry of alpha-MSH into CSF was 0.00087 min-1, which was not significantly different from that for inulin of 0.00055 min-1. As alpha-MSH penetrated into CSF at a rate comparable to inulin, it was concluded that the limited entry of peptide was by aqueous diffusion along with other water-soluble macromolecules.