Haematinic effect of Hygrophila spinosa T. Anderson on experimental rodents

Ethanolic extract of the aerial parts of H. spinosa a semiwoody herb was examined on male albino rats for certain haematological changes. The extract (100 & 200 mg/kg, po) significantly increased the haemoglobin, haematocrit, RBC and total WBC, as compared with vehicle treated control rat haemogram. In anemic male albino rats, the extract significantly increased haemoglobin, haematocrit and RBC count. Serum iron and serum total iron binding capacity were significantly decreased in H. spinosa extract treated anemic rats as compared with those in the vehicle treated anemic control rats. These findings demonstrated the haematinic effect of H. spinosa extract on experimental animals.     

Evaluation of orange-fleshed sweet potato (<Emphasis Type="Italic">Ipomoea batatas</Emphasis> L.) genotypes for salt tolerance through shoot apex culture under in vitro NaCl mediated salinity stress conditions

Fifteen genotypes of sweet potato were evaluated for salinity stress tolerance under in vitro NaCl mediated salinity stress conditions (MS, MS + 0.5% and MS + 1.0% NaCl). The growth parameters such as number of leaves, number of shoots, number of roots, length of plantlets and length of roots decreased significantly among the genotypes with increase in level of salinity. Of the 15 genotypes tested, six genotypes (108X1, 90/606, 90/696, CIP 8, S-30X15 and SP-61) were unable to sprout even at 0.5% NaCl and were characterized as susceptible to salt stress, three genotypes (CIP 6, 90/774 and CIP 3) which could tolerate 0.5% NaCl as moderately tolerant and six genotypes (CIP 12, CIP 13, JO 14, JP 13, SB-198/115 and Gouri) as tolerant to salinity at 1.0% NaCl. Amongst the six genotypes showing tolerance to 1.0% NaCl, the exotic genotypes––JP 13, CIP 12 and indigenous one SB-198/115 continued to exhibit significant higher values for growth parameters over the susceptible one. Based on the performance under NaCl mediated salinity stress (1.0%), the pattern of salinity tolerance in the genotypes through shoot apex culture was JP 13 > SB-198/115 > JO 14 > Gouri > CIP 12 > CIP 13. The effect of salt stress on the activity of antioxidative enzymes was studied in leaves of 8-week-old plantlets of those six genotypes, which responded at higher NaCl stress along with a susceptible genotype 90/606. In leaves of salt stressed plants, superoxide dismutase (SOD), guaiacol peroxidase (GPX) and catalase (CAT) activities increased when compared with the stress free control. The increase was more pronounced in the tolerant genotypes than that in the susceptible one. These results indicate that oxidative stress may play an important role in salt stressed sweet potato plants and that the greater protection of tolerant plants from salt induced oxidative damage results, at least in part, through the increase in the activity of antioxidant enzymes.     

Nampt/PBEF/Visfatin regulates insulin secretion in beta cells as a systemic NAD biosynthetic enzyme

Intracellular nicotinamide phosphoribosyltransferase (iNampt) is an essential enzyme in the NAD biosynthetic pathway. An extracellular form of this protein (eNampt) has been reported to act as a cytokine named PBEF or an insulin-mimetic hormone named visfatin, but its physiological relevance remains controversial. Here we show that eNampt does not exert insulin-mimetic effects in vitro or in vivo but rather exhibits robust NAD biosynthetic activity. Haplodeficiency and chemical inhibition of Nampt cause defects in NAD biosynthesis and glucose-stimulated insulin secretion in pancreatic islets in vivo and in vitro. These defects are corrected by administration of nicotinamide mononucleotide (NMN), a product of the Nampt reaction. A high concentration of NMN is present in mouse plasma, and plasma eNampt and NMN levels are reduced in Nampt heterozygous females. Our results demonstrate that Nampt-mediated systemic NAD biosynthesis is critical for beta cell function, suggesting a vital framework for the regulation of glucose homeostasis.     

Differential interactions of antitumor antibiotics chromomycin A(3) and mithramycin with d(TATGCATA)(2) in presence of Mg(2+)

The antitumor antibiotics chromomycin A(3) (CHR) and mithramycin (MTR) are known to inhibit macromolecular biosynthesis by reversibly binding to double stranded DNA with a GC base specificity via the minor groove in the presence of a divalent cation such as Mg(2+). Earlier reports from our laboratory showed that the antibiotics form two types of complexes with Mg(2+): complex I with 1:1 stoichiometry and complex II with 2:1 stoichiometry in terms of the antibiotic and Mg(2+). The binding potential of an octanucleotide, d(TATGCATA)(2), which contains one potential site of association with the above complexes of the two antibiotics, was examined using spectroscopic techniques such as absorption, fluorescence, and circular dichroism. We also evaluated thermodynamic parameters for the interaction. In spite of the presence of two structural moieties of the antibiotic in complex II, a major characteristic feature was the association of a single ligand molecule per molecule of octameric duplex in all cases. This indicated that the modes of association for the two types of complexes with the oligomeric DNA were different. The association was dependent on the nature of the antibiotics. Spectroscopic characterization along with analysis of binding and thermodynamic parameters showed that differences in the mode of recognition by complexes I and II of the antibiotics with polymeric DNA existed at the oligomeric level. Analysis of the thermodynamic parameters led us to propose a partial accommodation of the ligand in the groove without the displacement of bound water molecules and supported earlier results on the DNA structural transition from B --> A type geometry as an obligatory requirement for the accommodation of the bulkier complex II of the two drugs. The role of the carbohydrate moieties of the antibiotics in the DNA recognition process was indicated when we compared the DNA binding properties with the same type of Mg(2+) complex for the two antibiotics.     

Isolation,purification and partial chemical characterization of a lethal factor from common indian toad (Bufo melanostictus,Schneider) skin extract

Indian toad (Bufo melanostictus, Schneider) skin extract (TSE) is pharmacologically potent and probably contains several bioactive compounds [Das et. al., Indian J Pharmacol, 28 (1996) 72]. A lethal factor was isolated and purified by neutral alumina column chromatography followed by HPLC. Spectroscopic (UV, IR, FAB-MASS) study indicated that the lethal factor (TSE-LF) was a 254 Da long chain compound with carbonyl, hydroxyl and ester as functional groups. LD50 of TSE-LF was found to be 3.5 mg/kg (iv). Biological study showed that TSE-LF possesses hypotensive, cardiotoxic, neurotoxic activity and produced death by apnoea in experimental animal. Cyproheptadine antagonised TSE-LF induced contraction of isolated smooth muscle indicating involvement of histamine/serotonin receptors. TSE-LF induced neurotoxic action on chick biventer cervices was mediated through Ca2+ ion.     

RecA-mediated rescue of Escherichia coli strains with replication forks arrested at the terminus

The recombinational rescue of chromosome replication was investigated in Escherichia coli strains with the unidirectional origin oriR1, from the plasmid R1, integrated within oriC in clockwise (intR1(CW)) or counterclockwise (intR1(CC)) orientations. Only the intR1(CC) strain, with replication forks arrested at the terminus, required RecA for survival. Unlike the strains with RecA-dependent replication known so far, the intR1(CC) strain did not require RecBCD, RecF, RecG, RecJ, RuvAB, or SOS activation for viability. The overall levels of degradation of replicating chromosomes caused by inactivation of RecA were similar in oriC and intR1(CC) strains. In the intR1(CC) strain, RecA was also needed to maintain the integrity of the chromosome when the unidirectional replication forks were blocked at the terminus. This was consistent with suppression of the RecA dependence of the intR1(CC) strain by inactivating Tus, the protein needed to block replication forks at Ter sites. Thus, RecA is essential during asymmetric chromosome replication for the stable maintenance of the forks arrested at the terminus and for their eventual passage across the termination barrier(s) independently of the SOS and some of the major recombination pathways.     

Polymerized liposome assemblies: bifunctional macromolecular selectin inhibitors mimicking physiological selectin ligands

Monomeric sialyl Lewis(X) (sLe(x)) and sLe(x)-like oligosaccharides are minimal structures capable of supporting selectin binding in vitro. However, their weak binding interactions do not correlate with the high-affinity binding interactions witnessed in vivo. The polyvalent display of carbohydrate groups found on cell surface glycoprotein structures may contribute to the enhanced binding strength of selectin-mediated adhesion. Detailed biochemical analyses of physiological selectin ligands have revealed a complicated composition of molecules that bind to the selectins in vivo and suggest that there are other requirements for tight binding beyond simple carbohydrate multimerization. In an effort to mimic the high-affinity binding, polyvalent scaffolds that contain multicomponent displays of selectin-binding ligands have been synthesized. Here, we demonstrate that the presentation of additional anionic functional groups in the form of sulfate esters, on a polymerized liposome surface containing a multimeric array of sLe(x)-like oligosaccharides, generates a highly potent, bifunctional macromolecular assembly. This assembly inhibits L-, E-, and P-selectin binding to GlyCAM-1, a physiological ligand better than sLe(x)-like liposomes without additional anionic charge. These multivalent arrays are 4 orders of magnitude better than the monovalent carbohydrate. Liposomes displaying 3'-sulfo Lewis(X)-like oligosaccharides, on the other hand, show slight loss of binding with introduction of additional anionic functional groups for E- and P-selectin and negligible change for L-selectin. The ability to rapidly and systematically vary the composition of these assemblies is a distinguishing feature of this methodology and may be applied to the study of other systems where composite binding determinants are important for high-affinity binding.     

Characterization of herpes simplex virus-containing organelles by subcellular fractionation: role for organelle acidification in assembly of infectious particles

The cytoplasmic compartments occupied by exocytosing herpes simplex virus (HSV) are poorly defined. It is unclear which organelles contain the majority of trafficking virions and which are occupied by virions on a productive rather than defective assembly pathway. These problems are compounded by the fact that HSV-infected cells produce virus continuously over many hours. All stages in viral assembly and export therefore coexist, making it impossible to determine the sequence of events and their kinetics. To address these problems, we have established assays to monitor the presence of capsids and enveloped virions in cell extracts and prepared HSV-containing organelles from normally infected cells and from cells undergoing a single synchronized wave of viral egress. We find that, in both cases, HSV particles exit the nucleus and accumulate in organelles which cofractionate with the trans-Golgi network (TGN) and endosomes. In addition to carrying enveloped infectious virions in their lumen, HSV-bearing organelles also displayed nonenveloped capsids attached to their cytoplasmic surface. Neutralization of organellar pH by chloroquine or bafilomycin A resulted in the accumulation of noninfectious enveloped particles. We conclude that the organelles of the TGN/endocytic network play a key role in the assembly and trafficking of infectious HSV.     

Lack of Apparent Neurological Abnormalities in Rabbits Sensitized by Gangliosides

Two very high titer polyclonal antibodies against two ganglioside antigens, GM1 and GD1a, have been raised in New Zealand white rabbits using a homogeneous suspension of the highly purified antigens in Keyhole Limpet Hemocyanin and Freunds adjuvant. The antisera were prepared over a period of 6 months with repeated injections of the ganglioside suspension, followed by an intravenous injection of the purified ganglioside solution, and collecting the serum (approximately 50 ml) at defined time intervals. The GM1-antibody, thus prepared, showed a cross reactivity toward GD1b and asialo-GM1 (GA1), while the GD1a-antibody reacted with GD1a, GM1 and GA1 and GD1b as determined by immuno-overlay and ELISA methods. The titer for GM1 antiserum, determined by ELISA, was greater than 1/10,000 dilution while the titer for GD1a antibody was greater than 1/5,000 dilution. No neurological or behavioral abnormality was observed during the period of antiserum production. To evaluate any likely pathological damage caused by such a high titer ganglioside-antibody, autopsy of CNS as well PNS tissues from the rabbits were carried out after the final bleeding. No obvious pathological changes, including demyelination, were noted in any of the four rabbits. These observations cast doubt as to the direct effect of anti-ganglioside antibody induced neurological and pathological disorders.Special issue dedicated to Lawrence F. Eng.     

CoPS: Comprehensive Peptide Signature database

We present the development of a Comprehensive database of 12 076 invariant Peptide Signatures (CoPS) derived from 52 bacterial genomes with a minimum occurrence in at least seven organisms. These peptides were observed in functionally similar proteins and are distributed over nearly 1250 different functional proteins. The database provides function, structure and occurrence in biochemical pathways of the proteins containing these signature peptides. It houses additional information on the signature peptides, such as identical match in other motif/pattern (e.g. PROSITE, BLOCKS, PRINTS and Pfam) databases and the database of interacting proteins, human proteome and mutation effect on these signature peptides. There is a wide applicability of this database in the identification of critical functional residues in proteins. The database also facilitates the identification of folding nucleus/structural determinants in proteins and functional assignment to yet unknown proteins. We demonstrate functional assignment to 2605 hypothetical proteins in bacterial genomes and 112 unknown proteins in human using this database. AVAILABILITY: The database can be freely accessed through the following URL: http://203.195.151.46/copsv2/index.html or http://203.90.127.70/copsv2/index.html