Suppression of Oxygen Evolving System in Spinach Chloroplast Membranes due to Release of Manganese on Exposure to Osmotic Stress in vitro in Presence of Magnesium

When spinach chloroplast membranes were exposed to osmotic stress in vitro, by incubation in 1.0 M sorbitol + 10 mM MgCl₂ their oxygen evolving system was suppressed. The possible reasons for such inactivation of PS II mediated oxygen evolution were examined. There were conformational changes in the chloroplast membranes, as indicated by their absorption spectra. The pattern of sensitivity to DCMU was not altered. The sensitivity of PS II to water stress remained, even after a pre-wash treatment with NaCI (which removed 18 and 24 kD proteins) but not when the thylakoids were pretreated with NH₂0H or CaCl₂ (removed manganese and 33 kD). The manganese content of thylakoid membranes was markedly reduced under osmotic stress in presence of magnesium. We suggest that exposure of chloroplasts to 1.0 M sorbitol in presence of Mg₂₊ released manganese from thylakoid membranes, thereby leading to a suppression in oxygen evolution.     

Hydrophobic surfaces in oligosaccharides: linear dextrins are amphiphilic chains.

Polysaccharide chains are usually considered to be highly hydrophilic, since they have no obvious nonpolar moieties in them. Yet, it is possible to realise conformations in these chains wherein all the hydroxy groups are disposed in one side or face of the chain and the hydrogens disposed in the other. We experimentally demonstrate that such an amphiphilic surface is present in linear oligomeric dextrins, i.e., alpha-1,4-linked D-glucosides, but not in alpha-1,6-D-glucosides (dextrans) or in beta-1,4-D-glucosides (cellulose). This amphiphilicity is generated as a consequence of the stereochemical constraints, which vary with the structure of the sugar and with the type of linkage. Oligosaccharide chains that can adopt incipient helical structures might display amphiphilicity. This property might be relevant to intermolecular recognition on cell surfaces, lectin-sugar binding, antigen-antibody interactions and the like, and might be manifested more in heteromolecular recognition process than as homomolecular self-aggregation.     

A possible cause of non-disjunction of additional chromosome 21 in Down syndrome

Summary A possible cause of non-disjunction of chromosome 21 in Down Syndromes has been cytogenetically evaluated by examining the parents by Ag-staining technique. In all the cases studied so far, the contributing parents have active ribosomal cistrons on both chromosomes 21 i.e. both chromosomes are stained positively by silver staining. These results show that the active NORs might play an essential role in meiotic non-disjunction. Furthermore, the preliminary results demonstrate that the acrocentric associations of homologous and non-homologous nature involving chromosome 21 are the most frequent in the contributing parent which may further indicate the role of multiple cellular factors affecting the associations in promoting the non-disjunction in addition to active NORs. The possible mechanisms regarding the non-disjunction of chromosome 21 have been described.Presented at the 34th Annual Meeting of the American Society of Human Genetics, Norfolk, VA, USA     

Search for α3β2/3γ2 subtype selective ligands that are stable on human liver microsomes

Selective modulation of specific benzodiazepine receptor (BzR) gamma amino butyric acid-A (GABA_A) receptor ion channels has been identified as an important method for separating out the variety of pharmacological effects elicited by BzR-related drugs. Importantly, it has been demonstrated that both α2β_(2/3)γ2 (α2BzR) and α3BzR (and/or α2/α3) BzR subtype selective ligands exhibit anxiolytic effects with little or no sedation. Previously we have identified several such ligands; however, three of our parent ligands exhibited significant metabolic liability in rodents in the form of a labile ester group. Here eight analogs are reported which were designed to circumvent this liability by utilizing a rational replacement of the ester moiety based on medicinal chemistry precedents. In a metabolic stability study using human liver microsomes, four compounds were found to undergo slower metabolic transformation, as compared to their corresponding ester analogs. These compounds were also evaluated in in vitro efficacy assays. Additionally, bioisostere 11 was evaluated in a rodent model of anxiety. It exhibited anxiolytic activity at doses of 10 and 100 mg/kg and was devoid of sedative properties.     

Rosiglitazone Treatment of Type 2 Diabetic db/db Mice Attenuates Urinary Albumin and Angiotensin Converting Enzyme 2 Excretion

Alterations within the renal renin angiotensin system play a pivotal role in the development and progression of cardiovascular and renal disease. Angiotensin converting enzyme 2 (ACE2) is highly expressed in renal tubules and has been shown to be renoprotective in diabetes. The protease, a disintegrin and metalloprotease (ADAM) 17, is involved in the ectodomain shedding of several transmembrane proteins including ACE2. Renal ACE2 and ADAM17 were significantly increased in db/db mice compared to controls. We investigated the effect of the insulin sensitizer, rosiglitazone, on albuminuria, renal ADAM17 protein expression and ACE2 shedding in db/db diabetic mice. Rosiglitazone treatment of db/db mice normalized hyperglycemia, attenuated renal injury and decreased urinary ACE2 and renal ADAM17 protein expression. Urinary excreted ACE2 is enzymatically active. Western blot analysis of urinary ACE2 demonstrated two prominent immunoreactive bands at approximately 70 & 90 kDa. The predominant immunoreactive band is approximately 20 kDa shorter than the one demonstrated for kidney lysate, indicating possible ectodomain shedding of active renal ACE2 in the urine. Therefore, it is tempting to speculate that renoprotection of rosiglitazone could be partially mediated via downregulation of renal ADAM17 and ACE2 shedding. In addition, there was a positive correlation between blood glucose, urinary albumin, plasma glucagon, and triglyceride levels with urinary ACE2 excretion. In conclusion, urinary ACE2 could be used as a sensitive biomarker of diabetic nephropathy and for monitoring the effectiveness of renoprotective medication.     

Freeze-drying vegetative mycelium of Laccaria fraterna and its subsequent regeneration

Laccaria fraterna, a basidiomycetous, filamentous, non-sporulating (in vitro) fungus, was subjected to freeze-drying and tested for viability after rehydration. The optimization of the freeze-drying protocol included selection of the candidate fungus; optimizing physiological growth conditions and age; standardizing the protectant type and concentration; optimizing the pre-freezing method, freeze-drying run and extent of drying; choice of the rehydrant and the extent of rehydration. The culture retained its viability after lyophilization and when subjected to quality assurance tests gave consistent results similar to the non-lyophilized culture, indicating the stability of the product and the applicability of the developed process to freeze-dry vegetative mycelium of filamentous non-sporulating fungi.     

A Src inhibitor regulates the cell cycle of human pluripotent stem cells and improves directed differentiation

Driving human pluripotent stem cells (hPSCs) into specific lineages is an inefficient and challenging process. We show that a potent Src inhibitor, PP1, regulates expression of genes involved in the G1 to S phase transition of the cell cycle, activates proteins in the retinoblastoma family, and subsequently increases the differentiation propensities of hPSCs into all three germ layers. We further demonstrate that genetic suppression of Src regulates the activity of the retinoblastoma protein and enhances the differentiation potential of hPSCs across all germ layers. These positive effects extend beyond the initial germ layer specification and enable efficient differentiation at subsequent stages of differentiation.     

Xanthophylls are preferentially taken up compared with beta-carotene by retinal cells via a SRBI-dependent mechanism

The purpose of this study was to investigate the mechanisms by which carotenoids [xanthophylls vs. beta-carotene(beta-C)] are taken up by retinal pigment epithelial (RPE) cells. The human RPE cell line, ARPE-19, was used. When ARPE-19 cells were fully differentiated (7-9 weeks), the xanthophylls lutein (LUT) and zeaxanthin (ZEA) were taken up by cells to an extent 2-fold higher than beta-C (P < 0.05). At 9 weeks, cellular uptakes were 1.6, 2.5, and 3.2%, respectively, for beta-C, LUT, and ZEA. Similar extents were observed when carotenoids were delivered in either Tween 40 or "chylomicrons" produced by Caco-2 cells. Differentiated ARPE-19 cells did not exhibit any detectable beta-C 15,15'-oxygenase activity or convert exogenous beta-C into vitamin A. When using specific antibodies against the lipid transporters cluster determinant 36 (CD36) and scavenger receptor class B type I (SR-BI), cellular uptake of beta-C and ZEA were significantly decreased (40-60%) with anti-SR-BI but not with anti-CD36. Small interfering RNA transfection for SR-BI led to marked knockdown of SR-BI protein expression (approximately 90%), which resulted in decreased beta-C and ZEA uptakes by 51% and 87%, respectively. Thus, the present data show that RPE cells preferentially take up xanthophylls versus the carotene by a process that appears to be entirely SR-BI-dependent for ZEA and partly so for beta-C. This mechanism may explain, in part, the preferential accumulation of xanthophylls in the macula of the retina.