Bacopasaponins E and F: two jujubogenin bisdesmosides from Bacopa monniera

Two new dammarane-type jujubogenin bisdesmosides, bacopasaponins E and F of biological interest have been isolated from the reputed Indian medicinal plant Bacopa monniera and defined as 3-O-[beta-D-glucopyranosyl(1 --> 3)[alpha-L-arabinofuranosyl(1 --> 2)]alpha-L-arabinopyranosyl]-20-O-(alpha-L-arabinopyranosyl) jujubogenin and 3-O-[beta-D-glucopyranosyl(1 --> 3)[alpha-L-arabinofuranosyl(1 --> 2)]beta-D-glucopyranosyl]-20-O-alpha-L-arabinopyranosyl) jujubogenin respectively by spectroscopic methods and some chemical transformations.     

Crosstalk between the p190-B RhoGAP and IGF signaling pathways is required for embryonic mammary bud development

P190-B RhoGAP (p190-B, also known as ARHGAP5) has been shown to play an essential role in invasion of the terminal end buds (TEBs) into the surrounding fat pad during mammary gland ductal morphogenesis. Here we report that embryos with a homozygous p190-B gene deletion exhibit major defects in embryonic mammary bud development. Overall, p190-B-deficient buds were smaller in size, contained fewer cells, and displayed characteristics of impaired mesenchymal proliferation and differentiation. Consistent with the reported effects of p190-B deletion on IGF-1R signaling, IGF-1R-deficient embryos also displayed a similar small mammary bud phenotype. However, unlike the p190-B-deficient embryos, the IGF-1R-deficient embryos exhibited decreased epithelial proliferation and did not display mesenchymal defects. Because both IGF and p190-B signaling affect IRS-1/2, we examined IRS-1/2 double knockout embryonic mammary buds. These embryos displayed major defects similar to the p190-B-deficient embryos including smaller bud size. Importantly, like the p190-B-deficient buds, proliferation of the IRS-1/2-deficient mesenchyme was impaired. These results indicate that IGF signaling through p190-B and IRS proteins is critical for mammary bud formation and ensuing epithelial-mesenchymal interactions necessary to sustain mammary bud morphogenesis.     

Synthesis, crystal structure, DNA binding and photo-induced DNA cleavage activity of (S-methyl-L-cysteine)copper(II) complexes of heterocyclic bases

Ternary S-methyl-L-cysteine (SMe-l-cys) copper(II) complexes [Cu(SMe-L-cys)(B)(H(2)O)](X) (1-4), where the heterocyclic base B is 2,2'-bipyridine (bpy, 1), 1,10-phenanthroline (phen, 2), dipyridoquinoxaline (dpq, 3) and dipyridophenazine (dppz, 4), and X is ClO(4)(-) (1-3) or NO(3)(-) (4), are prepared and their DNA binding and cleavage properties studied. Complexes 2 and 4 are structurally characterized by X-ray crystallography. Both the crystal structures show distorted square-pyramidal (4+1) CuN(3)O(2) coordination geometry of the complexes in which the N,O-donor S-methyl-L-cysteine and N,N-donor heterocyclic base bind at the basal plane with a water molecule as the axial ligand. In addition, the dppz structure shows the presence of a 1D-chain formed due to covalent linkage of the carboxylate oxygen atom belonging to another molecule at the elongated axial site. The crystal structures show chemically significant non-covalent interactions like hydrogen bonding involving the axial aqua ligand and pi-pi interactions between dppz ligands. The complexes display a d-d band in the range of 605-654 nm in aqueous dimethylformamide (DMF) solution (9:1 v/v). The redox active complexes show quasireversible cyclic voltammetric response near 0.1 V in DMF assignable to the Cu(II)/Cu(I) couple. The complexes show good binding affinity to calf thymus (CT) DNA giving the order: 4 (dppz)>3 (dpq)>2 (phen)>1 (bpy). The intrinsic binding constants, obtained from UV-visible spectroscopic studies, are 1.3x10(4) and 2.15 x 10(4) M(-1) for 3 and 4, respectively. Control DNA cleavage experiments using pUC19 supercoiled (SC) DNA and minor groove binder distamycin suggest major groove binding propensity for the dppz complex, while the phen and dpq complexes bind at the minor groove of DNA. Complexes 2-4 show DNA cleavage activity in dark in the presence of a reducing agent 3-mercaptopropionic acid (MPA) via a mechanistic pathway involving formation of hydroxyl radical as the reactive species. The complexes also show efficient photo-induced DNA cleavage activity on irradiation with a monochromatic UV light of 365 nm in absence of any external reagent. The cleavage efficiency follows the order: 3>4>2. The complexes exhibit significant DNA cleavage activity on irradiation with visible light of 633 nm. Control experiments show inhibition of cleavage in presence of singlet oxygen quenchers like sodium azide, histidine and enhancement of cleavage in D(2)O, suggesting formation of singlet oxygen as a reactive species in a type-II process. The photosensitizing effect of the thiomethyl group of the amino acid is evidenced from the observation of significant DNA photocleavage activity of the phen complex 2 as the phen ligand itself is not a photosensitizer.     

Co-Carriage of bla KPC-2 and bla NDM-1 in Clinical Isolates of Pseudomonas aeruginosa Associated with Hospital Infections from India

Global spread of KPC poses to be a serious threat complicating treatment options in hospital settings. The present study investigates the genetic environment of bla _KPC-2 among clinical isolates of Pseudomonas aeruginosa from a tertiary referral hospital of India. The study isolates were collected from different wards and clinics of Silchar Medical College and Hospital, India, from 2012–2013. The presence of bla _KPC was confirmed by genotypic characterization followed by sequencing. Cloning of the bla _KPC-2 gene was performed and the genetic environment of this gene was characterized as well. Transferability of the resistance gene was determined by transformation assay and Southern hybridization. Additionally, restriction mapping was also carried out. Two isolates of P. aeruginosa were found to harbor bla _KPC-2, were resistant towards aminoglycosides, quinolone and β-lactam-β-lactamase inhibitor combination. In both the isolates, the resistance determinant was associated with class 1 integron and horizontally transferable. Both the isolates were co-harboring bla _NDM-1. The first detection of this integron mediated bla _KPC-2 coexisting with bla _NDM-1 in P. aeruginosa from India is worrisome, and further investigation is required to track the gene cassette mediated bla _KPC-2 in terms of infection control and to prevent the spread of this gene in hospitals as well as in the community.     

Development of the SAFE Checklist Tool for Assessing Site-Level Threats to Child Protection: Use of Delphi Methods and Application to Two Sites in India

Background

The child protection community is increasingly focused on developing tools to assess threats to child protection and the basic security needs and rights of children and families living in adverse circumstances. Although tremendous advances have been made to improve measurement of individual child health status or household functioning for use in low-resource settings, little attention has been paid to a more diverse array of settings in which many children in adversity spend time and how context contributes to threats to child protection. The SAFE model posits that insecurity in any of the following fundamental domains threatens security in the others: Safety/freedom from harm; Access to basic physiological needs and healthcare; Family and connection to others; Education and economic security. Site-level tools are needed in order to monitor the conditions that can dramatically undermine or support healthy child growth, development and emotional and behavioral health. From refugee camps and orphanages to schools and housing complexes, site-level threats exist that are not well captured by commonly used measures of child health and well-being or assessments of single households (e.g., SDQ, HOME).

Methods

The present study presents a methodology and the development of a scale for assessing site-level child protection threats in various settings of adversity. A modified Delphi panel process was enhanced with two stages of expert review in core content areas as well as review by experts in instrument development, and field pilot testing.

Results

Field testing in two diverse sites in India—a construction site and a railway station—revealed that the resulting SAFE instrument was sensitive to the differences between the sites from the standpoint of core child protection issues.     

Central histaminergic involvement during stress in rats.

Effects of some drugs modulating central histaminergic (HA) transmission were evaluated on restraint stress (RS)-induced gastric ulcerogenesis, plasma corticosterone and immune responses in rats. RS for (i) 6 hr or (ii) 24 hr at room temperature, and (iii) 3 hr at 4 degrees C (CRS) all induced gastric mucosal erosions and elevated plasma corticosterone levels, the effects with the latter two RS procedures being most consistent. Pretreatment of rats with neuronal HA depletor, alpha-FMH (100 mg/kg, ip) attenuated both ulcer severity and corticosterone response, during both 24 hr RS and CRS. Similar effects were also seen with the mast cell degranulator, C-48/80 (10 micrograms/kg, i.c.v.) treatment. Further, the H1-blocker, pheniramine (25 mg/kg, ip) but not the centrally acting H2-blocker, zolantidine (5 mg/kg, ip) produced clearcut attenuations in both stress markers, during the experimental stressors. In rats immunized in SRBC, 24 hr RS (and not CRS) significantly prevented the humoral immune responses to the antigen. alpha-FMH, C 48/80 and pheniramine but not zolantidine, reversed this response during 24 hr RS. The results indicate a central HA ergic involvement in the visceral, endocrinal and immune responses during RS and suggest the probable role of both neuronal as well as extraneuronal (mast cell) HA and activation of H1-receptors in the mediation of these effects.     

2D DIGE based proteomics study of erythrocyte cytosol in sickle cell disease: Altered proteostasis and oxidative stress

Sickle cell disease (SCD) is a hemolytic disorder caused by a mutation in beta‐globin gene and affects millions of people worldwide. Though clinical manifestations of the disease are quite heterogeneous, many of them occur due to erythrocyte sickling at reduced oxygen concentration and vascular occlusion mediated via blood cell adhesion to the vessel wall. We have followed proteomic approach to resolve the differentially regulated proteins of erythrocyte cytosol. The deregulated proteins mainly fall in the group of chaperone proteins such as heat shock protein 70, alpha hemoglobin stabilizing protein, and redox regulators such as aldehyde dehydrogenase and peroxiredoxin‐2 proteoforms. Proteasomal subunits are found to be upregulated and phospho‐catalase level also got altered. Severe oxidative stress inside erythrocyte is evident from the ROS analysis and Oxyblot^TM experiments. Peroxiredoxin‐2 shows significant dimerization in the SCD patients, a hallmark of oxidative stress inside erythrocytes. One interesting fact is that most of the differentially regulated proteins are also common for hemoglobinopathies such as Eβ thalassemia. These could provide important clues in understanding the pathophysiology of SCD and lead us to better patient management in the future.