C5a-blockade improves burn-induced cardiac dysfunction

We previously reported that generation of the anaphylatoxin C5a is linked to the development of cardiac dysfunction in sepsis due to C5a interaction with its receptor (C5aR) on cardiomyocytes. Burn injury involves inflammatory mechanisms that can lead to C5a generation as well. In this study, we investigated the effects of C5a blockade on burn-induced cardiac dysfunction. Using a standardized rat model of full thickness scald injury, left ventricular pressures were recorded in vivo followed by in vitro assessment of sarcomere contraction of single cardiomyocytes. Left ventricular pressures in vivo and cardiomyocyte sarcomere contractility in vitro were significantly reduced following burn injury. In the presence of anti-C5a Ab, these defects were greatly attenuated 1, 6, and 12 h after burn injury and completely abolished 24 h after burn. In vitro incubation of cardiomyocytes with bacterial LPS accentuated the impaired contractility, which was partially prevented in cardiomyocytes from burned rats that had received an anti-C5a Ab. Based on Western blot analyses, real-time PCR, and immunostaining of left ventricular heart tissue, there was a significant increase in cardiomyocyte expression of C5aR after burn injury. In conclusion, an in vivo blockade of C5a attenuates burn-induced cardiac dysfunction. Further deterioration of contractility due to the exposure of cardiomyocytes to LPS was partially prevented by C5a-blockade. These results suggest a linkage between C5a and burn-induced cardiac dysfunction and a possible contribution of LPS to these events.     

Recognition and cooperation between the ATP-dependent RNA helicase RhlB and ribonuclease RNase E

The Escherichia coli protein RhlB is an ATP-dependent motor that unfolds structured RNA for destruction by partner ribonucleases. In E. coli, and probably many other related gamma-proteobacteria, RhlB associates with the essential endoribonuclease RNase E as part of the multi-enzyme RNA degradosome assembly. The interaction with RNase E boosts RhlB's ATPase activity by an order of magnitude. Here, we examine the origins and implications of this effect. The location of the interaction sites on both RNase E and RhlB are refined and analysed using limited protease digestion, domain cross-linking and homology modelling. These data indicate that RhlB's carboxy-terminal RecA-like domain engages a segment of RNase E that is no greater than 64 residues. The interaction between RhlB and RNase E has two important consequences: first, the interaction itself stimulates the unwinding and ATPase activities of RhlB; second, RhlB gains proximity to two RNA-binding sites on RNase E, with which it cooperates to unwind RNA. Our homology model identifies a pattern of residues in RhlB that may be key for recognition of RNase E and which may communicate the activating effects. Our data also suggest that the association with RNase E may partially repress the RNA-binding activity of RhlB. This repression may in fact permit the interplay of the helicase and adjacent RNA binding segments as part of a process that steers substrates to either processing or destruction, depending on context, within the RNA degradosome assembly.     

Evaluation of hepatoprotective activity of Clerodendrum serratum L

The ethanol extract of C. serratum roots and ursolic acid isolated from it were evaluated for hepatoprotective activity against carbon tetrachloride induced toxicity in male Wistar strain rats. The parameters studied were estimation of liver function serum markers such as serum total bilirubin, total protein, alanine transaminase, aspartate transaminase and alkaline phosphatase activities. The ursolic acid showed more significant hepatoprotective activity than crude extract. The histological profile of the liver tissue of the root extract and ursolic acid treated animal showed the presence of normal hepatic cords, absence of necrosis and fatty infiltration as similar to the controls. The results when compared with the standard drug silymarin, revealed that the hepatoprotective activity of the constituent ursolic acid is significant as similar to the standard drug.     

Metabolic flux screening of Saccharomyces cerevisiae single knockout strains on glucose and galactose supports elucidation of gene function

New methods for an extended physiological characterization of yeast at a microtiter plate scale were applied to 27 deletion mutants of Saccharomyces cerevisiae cultivated on glucose and galactose as sole carbon sources. In this way, specific growth rates, specific rates of glucose consumption and ethanol production were determined. Flux distribution, particularly concerning branching into the pentose phosphate pathway was determined using a new (13)C-labelling method using MALDI-ToF-mass spectrometry. On glucose, the growth was predominantly fermentative whereas on galactose respiration was more active with correspondingly lower ethanol production. Some deletion strains showed unexpected behavior providing very informative data about the function of the corresponding gene. Deletion of malic enzyme gene, MAE1, did not show any significant phenotype when grown on glucose but a drastically increased branching from glucose 6-phosphate into the pentose phosphate pathway when grown on galactose. This allows the conclusion that MAE1 is important for the supply of NADPH during aerobic growth on galactose.     

High iNOS mRNA and protein levels during early third trimester suggest a role for NO in prelabor cervical ripening in the bovine

Nitric oxide (NO) plays a key role in the processes leading to cervical softening prior to labor. Inducible nitric oxide synthase (iNOS) contributes most to the increased production of NO during labor, as demonstrated in the rat cervix, or at term pregnancy in women. Changes in expression of iNOS during late gestation have not yet been studied longitudinally in any species, because repeatedly taking biopsies could not be performed. iNOS mRNA (n = 6) and protein expression (n = 3) in serial cervical biopsies of pregnant pluriparous cows taken around days 225, 250, and 275 of pregnancy and within 1.5 hr after calving (d225, d250, d275 and parturition biopsies, respectively) were measured using quantitative RT-PCR and Western blotting. iNOS mRNA expression decreased from the d225 biopsy onwards, differences being significant between the d250 and d275 (P < 0.05) and between the d275 and parturition biopsies (P < 0.05). iNOS protein expression decreased from d225 to d250 onwards. Immunohistochemical analysis of biopsies showed, besides positive staining in endothelium and epithelium, which remained unchanged at different time points, that iNOS expressing cells in the connective tissue cells of early biopsies were predominantly spindle shaped (mostly smooth muscle cells and some fibroblasts). In the parturition biopsies, iNOS reactivity was mainly found in mononuclear leucocytes. These results lead us to suggest that iNOS from spindle shaped cells is involved in prepartum cervical ripening, while iNOS in mononuclear inflammatory cells may be important for local tissue repair mechanisms during postpartum cervical involution.     

Stem anatomy of the dwarf subshrub Cressa cretica L. (Convolvulaceae)

Stem anatomy and development of medullary phloem are studied in the dwarf subshrub Cressa cretica L. (Convolvulaceae). The family Convolvulaceae is dominated by vines or woody climbers, which are characterized by the presence of successive cambia, medullary- and included phloem, internal cambium and presence of fibriform vessels. The main stems of the not winding C. cretica shows presence of medullary (internal) phloem, internal cambium and fibriform vessels, whereas successive cambia and included phloem are lacking. However, presence of fibriform vessels is an unique feature which so far has been reported only in climbing members of the family. Medullary phloem develops from peri-medullary cells after the initiation of secondary growth and completely occupies the pith region in fully grown mature plants. In young stems, the cortex is wide and formed of radial files of tightly packed small and large cells without intercellular air spaces. In thick stems, cortical cells become compressed due to the pressure developed by the radial expansion of secondary xylem, a feature actually common to halophytes. The stem diameter increases by the activity of a single ring of vascular cambium. The secondary xylem is composed of vessels (both wide and fibriform), fibres, axial parenchyma cells and uni-seriate rays. The secondary phloem consists of sieve elements, companion cells, axial and ray parenchyma cells. In consequence, Cressa shares anatomical characteristics of both climbing and non-climbing members. The structure of the secondary xylem is correlated with the habit and comparable with that of other climbing members of Convolvulaceae.     

MTHFR C677T Predisposes to POAG but Not to PACG in a North Indian Population: A Case Control Study

Hyperhomocysteinemia induced by the C677T genetic variant in MTHFR (methylenetetrahydrofolate reductase) has been implicated in neuronal cell death of retinal ganglion cells (RGC), which is a characteristic feature of glaucoma. However, association of MTHFR C677T with glaucoma has been controversial because of inconsistent results across association studies. Association between MTHFR C677T and glaucoma has not been reported in Indian population. Therefore, with a focus on neurodegenerative death of RGC in glaucoma, the current study aimed to investigate association of MTHFR C677T with Primary Open Angle Glaucoma (POAG) and Primary Angle Closure Glaucoma (PACG) in a North Indian population. A total of 404 participants (231 patients and 173 controls) were included in this study. Genotyping was performed by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism. A few random samples were also tested by direct sequencing. Genotypic and allelic distributions of the POAG and PACG cohorts were compared to that of controls by chi-square test and odds ratios were reported with 95% confidence intervals. Genotypic and allelic distributions between POAG cases and controls were significantly different (p = 0.03 and p = 0.01 respectively). Unlike POAG, we did not find significant difference in the genotypic and allelic distributions of C677T between PACG cases and controls (p>0.05). We also observed a higher proportion of TT associated POAG in females than that in males. However, this is a preliminary indication of gender specific risk of C677T that needs to be replicated in a larger cohort of males and females. The present investigation on MTHFR C677T and glaucoma reveals that the TT genotype and T allele of this polymorphism are significant risk factors for POAG but not for PACG in North Indian population. Ours is the first report demonstrating association of MTHFR C677T with POAG but not PACG in individuals from North India.     

Novel coumarin derivatives of heterocyclic compounds as lipid-lowering agents

Coumarin derivatives of different heterocycles (5,7a-i, 10 and 11) were designed based on cyclisation of 2-ethoxy-3-phenylpropanoic acid and 2-benzylmalonic acid as novel lipid-lowering agents and their preliminary in vivo screening indicates 7c has moderate triglyceride-lowering activity.     

Prolonged treatment with prostaglandin E1 increases the rate of lipolysis in rat adipocytes

Prolonged treatment of adipocytes with certain inhibitors of lipolysis, including N(6)-phenylisopropyl adenosine (PIA) and prostaglandin E(1) (PGE(1)) leads to down-regulation of G(i). Prolonged treatment with PIA increases the rate of lipolysis, and we have reported that tumor necrosis factor-alpha (TNF alpha) stimulates lipolysis by down-regulating G(i). To determine the relationship between G(i) concentration and lipolysis, we have investigated the effect of two other acute inhibitors of lipolysis; PGE(1), which down-regulates G(i), and nicotinic acid (NA), which does not down-regulate G(i). Rat adipocytes were incubated with PIA (300 nM), PGE(1) (3 microM) or nicotinic acid (1 mM) for 24 h. The rate of lipolysis (glycerol release) was increased approximately 2 to 3-fold in PIA-treated cells, and in PGE(1)-treated cells. Conversely, the rate of lipolysis was not altered by the prolonged nicotinic acid treatment. These findings support the hypothesis that the rate of lipolysis in adipocytes is determined, at least partly, by the cellular concentration of G(i) proteins.