Taurine protects against carbon tetrachloride toxicity in the cultured neurons and in vivo

Carbon tetrachloride (CCl4) has been found to induce cellular damage by generating oxygen free radicals. A study was carried out to investigate the effects of taurine (extracted from Pegasus laternarius Cuvier) on CCl4 intoxicated cultured neurons. CCl4 application (0.4 mmol x l(-1), 0.8 mmol x l( -1), 1.2 mmol x l (-1) and 1.6 mmol x l(-1 )) increased the lipid peroxidation product and decreased glutathione peroxidase (GPx) activity significantly in a concentration dependent manner. Pretreatment of cultures with taurine (10 micromol x l(-1), 30 micromol x l(-1) and 60 micromol x l(-1)) prevented the loss of GPx activity and lipid peroxidation. The effects of three different dosages of taurine (10 mg/kg body wt., 20 mg/kg body wt. and 40 mg/ kg body wt.) for 45 days on the activities of superoxide dismutase and glutathione peroxidase were examined in the cerebrum, cerebellum and medulla of normal and CCl4 treated mice. Treatment of mice with taurine provided protection against CCl4 toxicity as was evident by lipid peroxide status. Taurine was not so successful at inducing the activity of SOD in normal animals except in the medulla where it could increase the activity of SOD (p < 0.05). Taurine induces the GPx activity in a dose dependent manner in all regions of the brain studied. Also in the CCl4 poisoned mice taurine could augment the status of GPx activity in a dose dependent manner. Hence it is concluded that taurine can protects neurons from the oxidative stress induced by CCl4.     

Maharishi Amrit Kalash,an ayurvedic medicinal preparation,enhances cholinergic enzymes in aged guinea pig brain

The effect of orally fed Maharishi Amrit Kalash was examined on the activities of cholinergic enzymes in the guinea pig brain. The activity of the cholinergic enzymes viz. choline-acetyltransferase and acetylcholinesterase enzymes was found to be reduced significantly (P<0.05) in the various regions of CNS of the aged guinea pigs. Oral administration of MAK(500 mg/kg body weight daily) for 2 months significantly increased (P<0.05) the activity of choline acetyltransferase and acetylcholinesterase in the older animals. The present study indicates that this food supplement can be helpful in alleviating the cholinergic deficits in the old age.     

Simultaneous recognition of two chiral centers in α,β-diastereomeric amino acids by an N-carbamoyl-L-α-amino acid amidohydrolase from Alcaligenes xylosoxidans

Cells of Alcaligenes xylosoxidans containing N-carbamoyl-L--amino acid amidohydrolase strictly distinguished the configuration of not only the -carbon but also the -carbon of N-carbamoyl--methylphenylalanine, and produced threo-l--methylphenylalanine specifically from a mixture of the four stereoisomers.     

Flow cytometric analysis of DNA indices, expression of p53 and multidrug resistance genes in multiple myeloma patients

OBJECTIVE: To perform a quantitative analysis of DNA ploidy, S-phase fraction % (SPF%), p53 and multidrug resistance (MDR) gene expression as independent prognostic parameters and to compare these parameters with stage of the disease in multiple myeloma (MM) patients. STUDY DESIGN: Peripheral blood bone marrow samples were analyzed for DNA ploidy and SPF% using a FACScan flow cytometer (Becton Dickinson). Detection of p53 and MDR gene expression was done using immunocytochemistry. RESULTS: Aneuploidy was found in 5/48 (10.42%) total myeloma patients, all of whom revealed hyperdiploidy. High SPF% was noted in 18/37 (48.65%) newly diagnosed MM patients and 5/11 (45.45%) follow-up cases of myeloma. p53 Gene product was noted in 8/48 (16.66%) myeloma patients, 6 newly diagnosed and 2 on follow-up. MDR gene expression was detected in 4/27 (10.81%) newly diagnosed patients and in 1/11 (9.09%)follow-up patients. CONCLUSION: All myeloma patients with aneuploidy revealed hyperdiploidy. The majority of cases with high SPF% were at advanced stages, indicating the prognostic significance of SPF%. Although, there was no statistical significance of DNA ploidy, SPF%, p53 and MDR gene product expression, they are important prognostic parameters. Our results can provide baseline data for comparison with future studies since these parameters have not been reported earlier from the Indian subcontinent.     

Genetic admixture and population structure analysis of Indian water buffaloes (Bubalus bubalis) using STR markers

Molecular Biology Reports - India has a vast riverine and swamp buffalo diversity adapted to various agro-ecological conditions. In the present study, genetic diversity data for 10 different...     

Functional refolding of a recombinant C-type lectin-like domain containing intramolecular disulfide bonds

The lectin-like oxidized low-density lipoprotein scavenger receptor (LOX-1) is a pro-inflammatory marker and Type II membrane protein expressed on vascular cells and tissues. The LOX-1 extracellular domain mediates recognition of oxidized low-density lipoprotein (oxLDL) particles that are implicated in the development of atherosclerotic plaques. To study the molecular basis for LOX-1-mediated ligand recognition, we have expressed, purified and refolded a recombinant LOX-1 protein and assayed for its biological activity using a novel fluorescence-based assay to monitor binding to lipid particles. Overexpression of a hexahistidine-tagged cysteine-rich LOX-1 extracellular domain in bacteria leads to the formation of aggregates that accumulated in bacterial inclusion bodies. The hexahistidine-tagged LOX-1 molecule was purified by affinity chromatography from solubilized inclusion bodies. A sequential dialysis procedure was used to refold the purified but inactive and denatured LOX-1 protein into a functionally active form that mediated recognition of oxLDL particles. This approach allowed slow LOX-1 refolding and assembly of correct intrachain disulfide bonds. Circular dichroism analysis of the refolded LOX-1 molecule demonstrated a folded state with substantial alpha-helical content. Using immobilized recombinant, refolded LOX-1 we demonstrated a 70-fold preferential recognition for oxLDL over native LDL particles. Thus, a protein domain containing intrachain disulfide bonds can be reconstituted into a functionally active state using a relatively simple dialysis-based technique.     

Protective efficacy and immunogenicity of Vi-porin conjugate against Salmonella typhi.

A conjugate vaccine against Salmonella typhi was prepared by covalently binding capsular polysaccharide (Vi) with porin, both isolated from S. typhi. First, Vi and porins were extracted. The Vi was purified from S. typhi Ty2. The purified Vi conformed to the requirements of the World Health Organization. Porins were purified from S. typhi 0901. The Vi was bound to the porins by a heterobifunctional cross-linking reagent, N-succinimidyl-3-(2-pyridyl dithio)-propionate (SPDP). After preparing the Vi-porin conjugate, its protective ability and immunogenicity were studied in mice following systemic immunization. The results showed that the conjugate is 6.5-fold more protective than Vi alone against S. typhi. The mice immunized with conjugate elicited higher anti-Vi antibody (IgG) levels (P < 0.01) than the mice immunized with Vi alone. Anti-porin antibodies were also induced by the conjugate. To study the mucosal immune responses, secretory IgA (sIgA) in the intestinal fluid was measured. Conjugate-immunized mice showed the induction of sIgA as compared to Vi alone. The results showed that when Vi is bound to porins, both isolated from same organism, the resultant conjugate induced both systemic and mucosal immune responses and provided better protection against S. typhi than Vi alone.     

Methodological Considerations for the Use of Stable Isotope Probing in Microbial Ecology

Stable isotope probing (SIP) is a method used for labeling uncultivated microorganisms in environmental samples or directly in field studies using substrate enriched with stable isotope (e.g., ¹³C). After consumption of the substrate, the cells of microorganisms that consumed the substrate become enriched in the isotope. Labeled biomarkers, such as phospholipid-derived fatty acid (PLFA), ribosomal RNA, and DNA can be analyzed with a range of molecular and analytical techniques, and used to identify and characterize the organisms that incorporated the substrate. The advantages and disadvantages of PLFA-SIP, RNA-SIP, and DNA-SIP are presented. Using examples from our laboratory and from the literature, we discuss important methodological considerations for a successful SIP experiment.     

Antiangiogenic drugs synergize with a membrane macrophage colony-stimulating factor-based tumor vaccine to therapeutically treat rats with an established malignant intracranial glioma

Combining a T9/9L glioma vaccine, expressing the membrane form of M-CSF, with a systemic antiangiogenic drug-based therapy theoretically targeted toward growth factor receptors within the tumor's vasculature successfully treated >90% of the rats bearing 7-day-old intracranial T9/9L gliomas. The antiangiogenic drugs included (Z)-3-[4-(dimethylamino)benzylidenyl]indolin-2-one (a platelet-derived growth factor receptor beta and a fibroblast growth factor receptor 1 kinase inhibitor) and oxindole (a vascular endothelial growth factor receptor 2 kinase inhibitor). A total of 20-40% of the animals treated with the antiangiogenic drugs alone survived, while all nontreated controls and tumor vaccine-treated rats died within 40 days. In vitro, these drugs inhibited endothelial cells from proliferating in response to the angiogenic factors produced by T9/9L glioma cells and prevented endothelial cell tubulogenesis. FITC-labeled tomato lectin staining demonstrated fewer and constricted blood vessels within the intracranial tumor after drug therapy. Magnetic resonance imaging demonstrated that the intracranial T9 glioma grew much slower in the presence of these antiangiogenic drugs. These drugs did not affect in vitro glioma cell growth nor T cell mitogenesis. Histological analysis revealed that the tumor destruction occurred at the margins of the tumor, where there was a heavy lymphocytic infiltrate. Real-time PCR showed more IL-2-specific mRNA was present within the gliomas in the vaccinated rats treated with the drugs. Animals that rejected the established T9/9L glioma by the combination therapy proved immune against an intracranial rechallenge by T9/9L glioma, but showed no resistance to an unrelated MADB106 breast cancer.     

Identifying subset errors in multiple sequence alignments

Multiple sequence alignment (MSA) accuracy is important, but there is no widely accepted method of judging the accuracy that different alignment algorithms give. We present a simple approach to detecting two types of error, namely block shifts and the misplacement of residues within a gap. Given a MSA, subsets of very similar sequences are generated through the use of a redundancy filter, typically using a 70–90% sequence identity cut-off. Subsets thus produced are typically small and degenerate, and errors can be easily detected even by manual examination. The errors, albeit minor, are inevitably associated with gaps in the alignment, and so the procedure is particularly relevant to homology modelling of protein loop regions. The usefulness of the approach is illustrated in the context of the universal but little known [K/R]KLH motif that occurs in intracellular loop 1 of G protein coupled receptors (GPCR); other issues relevant to GPCR modelling are also discussed.