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Biomedical subjects

N Mani

Publications and source records attributed to N Mani.

At least 19 recordsLinked to original sources
PubMed2002

Angiogenic and astroglial responses to vascular endothelial growth factor administration in adult rat brain.

The effects of exogenous vascular endothelial growth factor (VEGF) on angiogenesis, blood-brain barrier permeability and astroglial proliferation in the adult rat CNS in situ were investigated. Recombinant human VEGF(165) (25 or 50 ng/ml) was delivered for up to 1 week using either intracerebral osmotic minipumps or less traumatic subdural gelatin sponge placement. By 3 days, VEGF delivery caused significantly increased cerebral angiogenesis (25 ng/ml was most effective) in both experimental models when compared to saline controls; VEGF infusion resulted in a 100% increase in an index of vascular proliferation, and gelatin sponge delivery produced a 65% increase. The blood-brain barrier hallmark endothelial glucose transporter-1 was not present in nascent vascular sprouts. Infusion of VEGF produced extensive protein leakage that persisted after saline-induced permeability was mostly resolved, while gelatin sponge administration caused milder barrier dysfunction. Administration of the angiogenic factor had unexpected proliferative effects on astroglia in both models, resulting in an 80-85% increase in mitotically active astroglia when compared to controls. Immunohistochemical results and semi-quantitative reverse transcriptase-polymerase chain reaction indicated that the VEGF receptors flk-1 and flt-1 were up-regulated in response to the infusion trauma; flt-1 was localized to reactive astroglia, while flk-1 was expressed in vascular endothelium but predominantly in neuronal somata and processes adjacent to the delivery site. mRNA for the VEGF(121), VEGF(165) and VEGF(188) isoforms was also increased after delivery of the recombinant protein. These data show that VEGF application has substantial proliferative effects on CNS endothelium and astroglia and causes up-regulation of its own message. Flt-1 and flk-1 receptor mRNAs and proteins are up-regulated in both vascular and non-vascular cell types following infusion trauma. From these results we suggest that administered VEGF has heretofore unanticipated pleiotrophic effects in the adult CNS.

Animals↗
PubMed2001

Preparation and characterization of two-phase melt systems of lidocaine.

The melting point of lidocaine was significantly lowered when mixed with thymol and/or aqueous ethanol. Mixtures of lidocaine and thymol at ratios within the range of 30:70-70:30 (w:w) became homogeneous oils at 25 degrees C. In a pH 9.2 carbonate buffer containing 25% ethanol, lidocaine (5%, w:w) also liquefied at 25 degrees C. The studies led to the development of novel two-phase melt systems of lidocaine (TMS) which consisted of a highly concentrated oil phase of lidocaine and an alcoholic aqueous phase. A compositional phase diagram showed that in aqueous dispersions of lidocaine, concurrent use of thymol and ethanol depressed the melting point of lidocaine more effectively than when they were used individually. Both thymol and aqueous ethanol were necessary as melting point depressing agents to achieve the highest possible lidocaine concentration of 87% (w:w) in the oil phase of a TMS at 25 degrees C. Containing an internal oil phase and an external aqueous phase at ambient temperature, such a TMS can be readily formulated into topical O/W cream after addition of proper surfactants and thickening agents. In an anesthetic activity test using mouse tail-flick model, a 5% lidocaine cream prepared was highly effective as shown by the prolonged latency time of the mice to a heat stimulus as compared with a placebo (P<0.05).

Analgesia↗
PubMed2001

Regulation of toxin synthesis in Clostridium difficile by an alternative RNA polymerase sigma factor.

Clostridium difficile, a causative agent of antibiotic-associated diarrhea and its potentially lethal form, pseudomembranous colitis, produces two large protein toxins that are responsible for the cellular damage associated with the disease. The level of toxin production appears to be critical for determining the severity of the disease, but the mechanism by which toxin synthesis is regulated is unknown. The product of a gene, txeR, that lies just upstream of the tox gene cluster was shown to be needed for tox gene expression in vivo and to activate promoter-specific transcription of the tox genes in vitro in conjunction with RNA polymerases from C. difficile, Bacillus subtilis, or Escherichia coli. TxeR was shown to function as an alternative sigma factor for RNA polymerase. Because homologs of TxeR regulate synthesis of toxins and a bacteriocin in other Clostridium species, TxeR appears to be a prototype for a novel mode of regulation of toxin genes.

Bacterial Toxins↗
PubMed2000

CcpC, a novel regulator of the LysR family required for glucose repression of the citB gene in Bacillus subtilis.

Synergistic carbon catabolite repression of the Bacillus subtilis aconitase (citB) gene by glucose and a source of 2-ketoglutarate is dependent on DNA sequences located upstream of the gene. Mutations in a dyad symmetry element centered at position -66 and in a repeat of the downstream arm of the dyad symmetry at position -27 cause derepressed citB expression. In this work, a protein able to bind to a DNA fragment containing these elements was purified and identified. This protein, named CcpC (Catabolite control protein C), shares sequence similarity with members of the LysR family of transcriptional regulators. In addition to binding to the citB promoter, CcpC bound to the promoter of the citZ gene, which encodes the cell's major citrate synthase and is subject to carbon catabolite repression. In a ccpC null mutant, expression of both citB and citZ was derepressed in glucose-glutamine minimal medium, indicating that CcpC is a negative regulator of citB and citZ gene expression. DNase I footprinting experiments showed that CcpC binds to two sites within the citB promoter region, corresponding to the dyad symmetry and -27 elements. In the presence of citrate, a putative inducer, only the dyad symmetry element was fully protected by CcpC. When the dyad symmetry element was mutated, CcpC was no longer able to bind to either the dyad symmetry or -27 elements. Repression of citB and citZ gene expression during anaerobiosis also proved to be mediated by CcpC.

Aconitate Hydratase↗
PubMed1999

Vascular, glial and neuronal effects of vascular endothelial growth factor in mesencephalic explant cultures.

Vascular endothelial growth factor is a highly conserved, heparin-binding protein which mediates a number of critical developmental processes in both vertebrates and invertebrates, including angiogenesis, vasculogenesis and hematopoiesis. We employed an organotypic rat explant model (produced from embryonic day 17 fetuses) to assess the effects of vascular endothelial growth factor on brain microvasculature in general and the ventral midbrain specifically. Immunohistochemistry using antisera to rat endothelial cell antigen and laminin demonstrated a robust, dose-dependent effect of vascular endothelial growth factor, resulting in increased vessel neogenesis, branching and lumen size by three days in vitro. This effect was blocked by addition of an anti-vascular endothelial growth factor antibody. At higher doses of vascular endothelial growth factor, the effect was attenuated, though a statistically significant increase in both astrocyte, and neuronal density was observed using antisera to glial and neuronal markers. Tyrosine hydroxylase-immunoreactive (i.e. dopaminergic) neurons, particularly, exhibited increased survival in response to vascular endothelial growth factor application. Vascular endothelial growth factor had a mitogenic effect on endothelial cells and astrocytes, but not dopaminergic neurons, as demonstrated by the addition of [3H]thymidine to the cultures 2 h after the cultures were established. Similarly, results of a radioreceptor assay indicated that specific vascular endothelial growth factor binding sites were present on blood vessels and astrocytes, and were up-regulated by exposure to vascular endothelial growth factor. We conclude that, in explants of the ventral mesencephalon, exogenously applied vascular endothelial growth factor is mitogenic for endothelial cells and astrocytes, and promotes growth/survival of neurons in general and dopaminergic neurons in particular.

Animals↗
PubMed1998

Patterns of brain angiogenesis after vascular endothelial growth factor administration in vitro and in vivo.

Vascular endothelial growth factor (VEGF) is a secreted endothelial cell mitogen that has been shown to induce vasculogenesis and angiogenesis in many organ systems and tumors. Considering the importance of VEGF to embryonic vascularization and survival, the effects of administered VEGF on developing or adult cerebrovasculature are unknown: can VEGF alter brain angiogenesis or mature cerebrovascular patterns? To examine these questions we exposed fetal, newborn, and adult rat cortical slice explants to graduated doses of recombinant VEGF. The effects of another known angiogenic factor, basic fibroblast growth factor (bFGF), were evaluated in a comparable manner. In addition, we infused VEGF via minipump into the adult cortex. Significant angiogenic effects were found in all VEGF experiments in a dose-responsive manner that were abolished by the addition of VEGF neutralizing antibody. Fetal and newborn explants had a highly complex network of branched vessels that immunoexpressed the flt-1 VEGF receptor, and flk-1 VEGF receptor expression was determined by reverse transcription-PCR. Adult explants had enlarged, dilated vessels that appeared to be an expansion of the existing network. All bFGF-treated explants had substantially fewer vascular profiles. VEGF infusions produced both a remarkable localized neovascularization and, unexpectedly, the expression of flt-1 on reactive astrocytes but not on endothelial cells. The preponderance of neovascularization in vitro and in vivo, however, lacked the blood-brain barrier (BBB) phenotype marker, GLUT-1, suggesting that in brain the angiogenic role of VEGF may differ from a potential BBB functional role, i.e., transport and permeability. VEGF may serve an important capacity in neovascularization or BBB alterations after brain injury.

Animals↗
PubMed1998

Screening systems for detecting inhibitors of cell wall transglycosylation in Enterococcus. Cell wall transglycosylation inhibitors in Enterococcus.

We devised two screening systems to detect cell wall transglycosylation inhibitors. One screen utilizes a mutant of Enterococcus faecalis strain A256 that is dependent on vancomycin or moenomycin for growth. In the absence of transglycosylation inhibitors the strain fails to grow, while in the presence of inhibitors, cells are rescued. A second screening organism E. faecalis strain MDD212 utilizes a translational fusion of the lacZ gene to the vanH promoter in a derivative of E. faecalis that contains a vancomycin resistance determinant. Induction of beta-galactosidase occurs when cells are exposed to inhibitors of transglycosylation. Our natural products drug source of fungal fermentations was tested with these screens. Several cultures that produced the same family of compounds, called the thielavins, were detected. Thielavin B inhibited the formation of peptidoglycan in an in vitro assay, suggesting that these screening systems can detect compounds that interfere with cell wall transglycosylation.

Base Sequence↗
PubMed1997

Molecular analysis of lytic genes of bacteriophage 80 alpha of Staphylococcus aureus.

Nucleotide sequencing of a 3779-bp fragment of the Staphylococcus aureus bacteriophage 80 alpha revealed two open reading frames: ORF1, designated as lytA, which encodes a polypeptide of 481 amino acids with an apparent M(r) of 53.81 kDa; and ORF2, designated as holin, which encodes for a hydrophobic polypeptide of 145 amino acids with an apparent M(r) of 15.58 kDa and exhibits two putative transmembrane helices. Both genes showed 100% sequence homology to that of the peptidoglycan hydrolase and holin genes of the S. aureus phage phi 11 reported earlier. In addition, the downstream sequences of the lytA gene were homologous to the phage attachment site (attP) of the phage phi 11. Based on our data we propose that the lytic system of the phage 80 alpha evolved from that of phage phi 11.

Amino Acid Sequence↗
PubMed1994

Effect of physiological conditions on the autolysis of Staphylococcus aureus strains.

The effect of physiological conditions on autolysis and autolytic activity in various strains of Staphylococcus aureus was determined. The rate of whole cell autolysis of S. aureus was growth phase dependent and a maximum rate was observed in early stationary phase cultures. However, the autolysins extracted by the freeze-thaw method (cell-wall bound autolytic activity) did not show any significant increase in activity. The addition of NaCl to the growth medium enhanced the rate of autolysis with the highest rate being displayed by cultures grown in 1.5 M NaCl. However, lower autolytic activity was found in the freeze-thaw extracts of cultures grown at higher concentrations of NaCl. The rate of autolysis of cultures grown at 30 degrees C was higher than cultures grown at 37 or 43 degrees C. Thus, the rate of autolysis seems to be independent of the bacterial growth rate. Cultures grown in slightly acidic conditions showed a faster rate of autolysis compared to cultures grown under alkaline conditions. SDS-polyacrylamide gel containing 0.2% crude cell-wall of S. aureus did not show any obvious correlation with the appearance of any particular lytic band in the zymogram to autolytic activity or rate of autolysis of cultures grown under various environmental conditions. A nonhemolytic phenotype, mutations in the accessory gene regulator, and lysogeny (phages phi 11, phi 12, phi 13) had no obvious effect either on the rate of autolysis or on the pattern of lytic bands in the zymograms.

Autolysis↗
PubMed1994

Autolysis-defective mutant of Staphylococcus aureus: pathological considerations, genetic mapping, and electron microscopic studies.

In an earlier report, we had described the isolation and characterization of autolysis-defective mutants of Staphylococcus aureus (N. Mani, P. Tobin, and R.K. Jayaswal, J. Bacteriol. 175:1493-1499, 1993). In the study reported here, an autolysis-defective mutant showed attenuated virulence in a rat model of experimental endocarditis, supporting the role of autolysins in pathogenicity. Transmission electron micrographs of the mutant cells revealed a rough outermost surface as compared with the parent strain, ISP2018. Treatment of mutant cells with lysozyme, proteases, and lipase failed to alter this rough appearance. Physical and genetic data locate the site of mutation between the omega 1100 and ilv loci on the S. aureus chromosome.

Animals↗
PubMed1993

Isolation and characterization of autolysis-defective mutants of Staphylococcus aureus created by Tn917-lacZ mutagenesis.

Two autolysis-defective mutants (Lyt-1 and Lyt-2) of Staphylococcus aureus have been isolated by transposon Tn917-lacZ mutagenesis. The mutants exhibited normal growth rate, cell division, cell size, and adaptive responses to environmental changes. No autolytic activities were detected in a crude autolytic enzyme preparation from the Lyt- mutants. The rate of autolysis of whole cells and cell walls in the mutants were negligible, but mutant cell wall preparations were degraded by crude enzyme preparations from the wild-type strain. Zymographic analyses of enzyme extracts from the mutants showed a single autolytic enzyme band, compared with more than 10 autolytic enzyme bands from the parent strain. Analyses of intracellular and exoprotein fractions gave results similar to those in experiments with total-cell extracts. Southern blot analysis indicated the insertion of a single copy of the transposon into the chromosome of Lyt mutants. Isogenic Lyt mutants constructed by phage phi 11 transduction showed similar phenotypes. Because both Lyt- mutants had Tn917-lacZ inserted in the appropriate orientation, it was possible to determine gene activity under various conditions by measuring beta-galactosidase activity. The gene activity was found to be induced by low pH, low temperature, and high sucrose and high sodium chloride concentrations. From these data, we propose that the mutation lies in either a master regulatory gene or a structural gene which is responsible for the synthesis or processing of a majority of the autolytic enzyme bands.

Electrophoresis, Polyacrylamide Gel↗
PubMed1992

Analysis of a peptidoglycan hydrolase gene from Staphylococcus aureus NCTC 8325.

We have investigated the expression of the peptidoglycan hydrolase gene (lytA) of Staphylococcus aureus NCTC 8325. Results from in vitro transcription-translation analysis, maxicell experiments, and Northern (RNA) blot analysis suggest that the lytA gene encodes a polypeptide of M(r) approximately 50,000. Physical mapping data indicate that the lytA gene originated from prophage 11 in the NCTC 8325 strain.

Chromosome Mapping↗
PubMed1991

Treatment of pure red-cell aplasia following major ABO-mismatched T-cell-depleted bone marrow transplantation. Two case reports with successful response to plasmapheresis.

Major ABO-mismatched bone marrow transplantation (BMT) may be accompanied by red-cell haemolysis, but pure red-cell aplasia following BMT is a rare complication. Two cases of transient pure red-cell aplasia following T-lymphocyte-depleted BMT for a period of greater than 20 weeks are described, both of which responded to one cycle of plasmapheresis. The prompt response of the two patients described with red-cell aplasia with no evidence of haemolysis suggests that plasmapheresis may be considered in such clinical situations as a first treatment of choice before attempting more complex modes of therapy.

ABO Blood-Group System↗
PubMed1991

[Not Available].

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General Surgery↗
PubMed1990

Management of traumatic rupture of the globe in aphakic patients.

We report the successful treatment of three cases of traumatic ruptures of the globe complicated by massive choroidal hemorrhage, uveal prolapse and retinal detachment. All three of the eyes were aphakic prior to injury and all patients were age 64 or older. The presenting visual acuity in all patients was light perception. The blunt injury in each case caused a wound dehiscence at the site of previous cataract extraction. All injuries were associated with uveal prolapse. Secondary surgical intervention was performed when the hemorrhagic choroidal detachments had decreased as demonstrated by echography in the suprachoroidal space, occurring at an average of 14 days after injury. The management consisted of surgical drainage of the choroidal hemorrhage combined with vitrectomy and silicone oil injection. Successful reattachment of the retina was achieved in all cases. Postoperative epiretinal membranes formed in two cases but all were anatomically successful at six months. Final visual acuities varied from 20/70 to 1/200, visual acuity being a function of secondary contusive damage to the retina and choroid. We believe that in eyes sustaining severe blunt injuries resulting in rupture of the globe complicated by massive choroidal hemorrhage and retinal detachment, properly timed external drainage of the choroidal hemorrhage combined with pars plana vitrectomy and silicone oil injection is a useful approach.

Aged↗
PubMed1983

[Not Available].

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Epidemiology↗