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Biomedical subjects

A Chand

Publications and source records attributed to A Chand.

At least 19 recordsLinked to original sources
PubMed2001

Single incision for implanting a vagal nerve stimulator system (VNSS): technical note.

BACKGROUND: A technique for implanting the vagal nerve stimulator system through a single incision is described. METHOD: A transverse incision is made in the lower part of the neck. Subcutaneous (s.c.) dissection is then done over the clavicle into the infraclavicular area to create a pocket. The vagus nerve is exposed and the electrodes are wrapped around it through the neck incision. The distal ends of the lead are connected to the pulse generator, and latter is then placed in the infraclavicular pocket through the neck incision. RESULTS: Thirty-eight implants were conducted with this technique. The pulse generator could be implanted and anchored to the underlying tissue without any difficulty. Except for wound infections in two patients there was no other complication. CONCLUSION: A single incision is an alternate to the double incision procedure. This procedure can be performed safely.

Adolescent↗
PubMed2000

Human mini-chromosomes with minimal centromeres.

We have introduced a 6.5 Mb human mini-chromosome with a complex centromere structure into DT40 cells and have used sequence targeting and telomere-directed chromosome breakage to dissect the sequence requirements for centromere function. These experiments proved that a vertebrate centromere with two blocks of functional alphoid DNA separated by 2.5 Mb can exist as a stable structure in some but not all vertebrate cells. Further experiments indicated that recovery of chromosomes with less than approximately 100 kb of alphoid DNA is very inefficient, suggesting that a functional centromere requires a minimum of approximately 100 kb of alphoid DNA. Mini-chromosomes with minimal centromeres segregate accurately in some but not all vertebrate cells and should be useful for the detection of sequence-specific factors required for vertebrate centromere maintenance.

Animals↗
PubMed2000

Probing protein-protein interactions in real time.

We have used a prototype small cantilever atomic force microscope to observe, in real time, the interactions between individual protein molecules. In particular, we have observed individual molecules of the chaperonin protein GroES binding to and then dissociating from individual GroEL proteins, which were immobilized on a mica support. This work suggests that the small cantilever atomic force microscope is a useful tool for studying protein dynamics at the single molecule level.

Aluminum Silicates↗
PubMed1998

Glomerular extracellular matrix components and integrins.

It has become apparent that extracellular matrix components and their cellular receptors, the integrins, are important regulators of glomerular development and function. In this rapidly evolving field we studied the production of extracellular matrix components and integrins by rat glomerular visceral epithelial and mesangial cells, using molecular probes and antibodies that have recently become available. Special attention was paid to laminin isoforms and to splice variants of the integrin subunits alpha 3 and alpha 6. Results were compared to the in vivo expression in human fetal, newborn and adult kidneys. The mesangial cells were found to produce laminin-1, nidogen and two as yet unidentified laminin isoforms with putative alpha chains of about 395 (alpha x) and of 375 kDa (alpha y), tentatively described before as bovine kidney laminin. Furthermore, they expressed the integrins alpha 1 beta 1, alpha 2 beta 1, alpha 3A beta 1, alpha 5 beta 1, alpha v beta 3, alpha v beta 5, and small amounts of alpha 6A beta 1 and alpha 6B beta 1. The glomerular visceral epithelial cells produced the two new laminin isoforms mentioned above, laminin-5, but no laminin-1 or nidogen. The integrins alpha 2 beta 1, alpha 3A beta 1, alpha 6A beta 4, alpha 6B beta 4 and the integrin subunit alpha v were found to be expressed. We show that during nephrogenesis, the laminin alpha 1 chain disappears and is replaced by another alpha chain, possibly one of the two as yet unidentified alpha chains mentioned above. The laminin beta 1 chain is replaced by the beta 2 chain somewhat later in glomerular development. In general, the integrins found to be expressed in glomeruli of adult kidney were consistent with those found in cultured glomerular visceral epithelial and mesangial cells. No splice variant switch of the integrin alpha 3 or alpha 6 subunits could be demonstrated during nephrogenesis. Our results suggest an important role for the mesangial cell in providing nidogen as a crucial component of the supramolecular structure of the glomerular basement membrane. Furthermore our results indicate that laminin alpha x beta 2 gamma 1 and alpha y beta 2 gamma 1 isoforms are important in the glomerulus of adult kidney and that the integrin alpha 3A beta 1 is the main integrin receptor for laminin isoforms on glomerular visceral epithelial and mesangial cells, both in vitro and in vivo.

Adult↗
PubMed1996

Mammalian artificial chromosomes.

The development of candidate vectors and of techniques for their manipulation by sequence targeting suggest that a mini-chromosome vector system for the mouse germline may be at hand. Mini-chromosome vectors should allow new sorts of genetic problems to be addressed experimentally and may accelerate the process of gene identification.

Journal Article↗
PubMed1996

Mammalian artificial chromosomes.

The development of candidate vectors and of techniques for their manipulation by sequence targeting suggest that a mini-chromosome vector system for the mouse germline may be at hand. Mini-chromosome vectors should allow new sorts of genetic problems to be addressed experimentally and may accelerate the process of gene identification.

Animals↗
PubMed1995

Long-range organization of reiterated sequences, including the SSX1 cDNA at the OATL1 cluster in Xp11.23.

Analysis of four YACs at the OATL1 locus was undertaken to determine the organization of locally repeated sequences within the OATL1 cluster. A restriction map of ICRFy900C0874, a 600-kb YAC, was constructed using a range of rare-cutting enzymes. Several markers were isolated from the OATL1 cluster, and the YAC map was used for their localization. Markers are shown to be reiterated within the OATL1 cluster, and fine-scale mapping has identified the accurate map position and organization of these repeated sequences within a 275-kb interval. The SSX1 gene involved in synovial sarcoma tumorigenesis and localized at the OATL1 cluster is also shown to be present in a minimum of five copies, all of which map to the aforementioned region. Several markers generated in this study have homologous counterparts at OATL2; they have been utilized to delimit the duplicated region at the OATL clusters. Although instability of one of the OATL1 YACs, ICRFy900F0501, has precluded a precise sizing of this interval, it has been possible to place an upper limit of 520 kb on this region of duplication.

Base Sequence↗
PubMed1995

Construction of two YAC contigs in human Xp11.23-p11.22, one encompassing the loci OATL1, GATA, TFE3, and SYP, the other linking DXS255 to DXS146.

We have constructed two YAC contigs in the Xp11.23-p11.22 interval of the human X chromosome, a region that was previously poorly characterized. One contig, of at least 1.4 Mb, links the pseudogene OATL1 to the genes GATA1, TFE3, and SYP and also contains loci implicated in Wiskott-Aldrich syndrome and synovial sarcoma. A second contig, mapping proximal to the first, is estimated to be over 2.1 Mb and links the hypervariable locus DXS255 to DXS146, and also contains a chloride channel gene that is responsible for hereditary nephrolithiasis. We have used plasmid rescue, inverse PCR, and Alu-PCR to generate 20 novel markers from this region, 1 of which is polymorphic, and have positioned these relative to one another on the basis of YAC analysis. The order of previously known markers within our contigs, Xpter-OATL1-GATA-TFE3-SYP-DXS255146- Xcen, agrees with genomic pulsed-field maps of the region. In addition, we have constructed a rare-cutter restriction map for a 710-kb region of the DXS255-DXS146 contig and have identified three CPG islands. These contigs and new markers will provide a useful resource for more detailed analysis of Xp11.23-p11.22, a region implicated in several genetic diseases.

Animals↗
PubMed1995

Fusion of SYT to two genes, SSX1 and SSX2, encoding proteins with homology to the Kruppel-associated box in human synovial sarcoma.

We demonstrate that the cytogenetically defined translocation t(X;18)(p11.2;q11.2) found in human synovial sarcoma results in the fusion of the chromosome 18 SYT gene to either of two distinct genes, SSX1 or SSX2, at Xp11.2. The SSX1 and SSX2 genes encode closely related proteins (81% identity) of 188 amino acids that are rich in charged amino acids. The N-terminal portion of each SSX protein exhibits homology to the Kruppel-associated box (KRAB), a transcriptional repressor domain previously found only in Kruppel-type zinc finger proteins. PCR analysis demonstrates the presence of SYT-SSX1 or SYT-SSX2 fusion transcripts in 29 of 32 of the synovial sarcomas examined, indicating that the detection of these hybrid transcripts by PCR may represent a very useful diagnostic method. Sequence analysis has demonstrated heterogeneity in the fusion transcripts with the formation of two distinct SYT-SSX1 fusion junctions and two distinct SYT-SSX2 fusion junctions.

Adolescent↗
PubMed1995

Mercuric chloride-induced programmed cell death of a murine T cell hybridoma. II. Opposite effect of interleukin-2 and interleukin-4.

In susceptible animals evidence is accumulating for a primary role for Th2 cells in the course of HgCl2-induced autoimmunity, and for a contribution of Th1 cells in the self-regulated phase of this disease. We have reported that incubation of 2B4.11 T cell hybridoma with HgCl2 induced programmed cell death. This paper shows that recombinant IL-2 significantly diminished HgCl2-induced 2B4.11 cell death. Although no effect was observed upon incubation with exogenous IL-4, we observed a significant protection by adding an anti-IL-4 monoclonal antibody to the culture. Accordingly, by RT-PCR we found the presence of IL-2 receptor-encoding mRNA, and by cytofluorometry, the expression of the protein was detected only after exposure to HgCl2. Moreover, upon HgCl2 treatment, 2B4.11 cells were induced to produce IL-4. Altogether these findings showed that cytokine environment, IL-2, IL-4 otherwise defining the Th1/Th2 dichotomy, in conjunction with a chemical may differentially influence the fate of cell populations, death or survival.

Animals↗
PubMed1995

Modifications of transnasal and transoral stereotactic procedures--technical notes.

Stereotactic transnasal and transoral procedures have been reported by different authors in the past. In this paper, several modifications of these methods are described. The modifications are: transnasal approach to the frontal skull base and suprasellar regions; and transoral approaches to the clivus, the petroclival junction, medial part of the cerebellopontine angle and the lateral mass of C-1. Eleven patients were operated on using these modifications. The procedures were for biopsy and brachytherapy. No complications resulted from the procedures.

Adolescent↗
PubMed1994

Podocyte expression of MHC class I and II and intercellular adhesion molecule-1 (ICAM-1) in experimental pauci-immune crescentic glomerulonephritis.

We examined immunopathological changes of podocytes in vivo which, based on in vitro studies, are thought to be relevant for the pathogenesis of renal diseases. We investigated the alterations of podocytes in local inflammation in a recently developed model of pauci-immune necrotizing crescentic glomerulonephritis (NCGN) in the rat. Frozen and plastic embedded kidney sections at different time points of the disease were incubated with antibodies directed to MHC class I, MHC class II, ICAM-1 and to relevant cytokines. Strong glomerular expression of MHC class I, II and ICAM-1 was found within 4 days, and plastic embedded sections clearly demonstrated increased cell membrane staining of podocytes. Increased glomerular interferon-gamma (IFN-gamma) was detected within 24 h of induction of NCGN, and IL-1 beta and tumour necrosis factor-alpha (TNF-alpha) were found from day 4. The potency of these cytokines to induce adhesion molecules on podocytes was investigated on rat glomerular epithelial cells in vitro. By using FACS analysis and electron microscopical techniques, we found that the in vivo expression of MHC class I, II and ICAM-1 by podocytes could in vitro be simulated by IFN-gamma. IFN-alpha weakly induced MHC class I, while IL-1 beta and TNF-alpha were ineffective. We hypothesize that podocytes in this in vivo model are important to maintain the local inflammatory process in the glomerulus by expression of relevant adhesion molecules and MHC molecules upon stimulation with specific cytokines.

Animals↗
PubMed1991

Subcellular distribution of monoclonal antibody defined epitopes on immunodominant Mycobacterium tuberculosis proteins in the 30-kDa region: identification and localization of 29/33-kDa doublet proteins on mycobacterial cell wall.

Two different groups of monoclonal antibodies (MAbs) directed to different epitopes on 30-kDa region proteins of Mycobacterium tuberculosis were isolated; MAbs 5F9, 5D5 and 5D2 reacted with a single 33-kDa protein band, whereas MAb 3A8 reacted with a distinct 29/33-kDa doublet when analysed by immunoblotting. The present paper describes the distribution of MAbs defined epitopes in the 29-33-kDa region proteins in well-characterized subcellular fractions: cytosol, plasma membrane, cell wall as well as culture filtrate of M. tuberculosis. MAbs 5F9, 5D5 and 5D2 reactive epitopes were found in cytosol, whereas 3A8 epitope is distributed in all cellular compartments of the mycobacterium as well as in the culture filtrate. Localization of these epitopes by indirect immunofluorescence and immunogold-labelling demonstrated that only 3A8 epitope is present on the cell surface of the mycobacterium. Both immunoblotting and ELISA showed that only MAb 3A8, and not 5F9, 5D5 and 5D2, reacted with secreted BCG 85 antigen complex of Mycobacterium bovis BCG. Furthermore, using an MAb 3A8-coupled affinity column, we purified antigen 3A8 from the cytosol fraction of M. tuberculosis. All these MAbs reacted with antigen 3A8 with varying degrees of intensity, thus suggesting that they are directed to a single protein. Absence of 5F9, 5D5 and 5D2 epitopes in the cell wall, culture filtrate and to a single protein. Absence of 5F9, 5D5 and 5D2 epitopes in the cell wall, culture filtrate and BCG-85 complex suggests that these epitopes might have been lost during the processing of the same 33-kDa protein on its way out from cytosol to the cell wall or when the protein is secreted out into the culture filtrate. Our results demonstrate, for the first time, direct evidence of the presence of a 30-kDa region protein not only in secreted antigens but also in the cell wall and on the cell surface of the mycobacterium.

Animals↗
PubMed1990

Gene of type II autosomal dominant retinitis pigmentosa maps on the long arm of chromosome 3.

Linkage analysis has been performed on a large Australian family segregating for the autosomal dominant form of retinitis pigmentosa (ADRP). The majority of patients had no subjective symptoms of night blindness until their second decade and good visual acuity until late in life. The disease in this family has been classified as Type II ADRP according to the subdivisions provided by both Massof and Finkelstein and Fishman and colleagues. Linkage (Omax:0.08 at Zmax:4.78) is here demonstrated between the disease locus and D3S47 (a marker locus on the long arm of chromosome 3), which showed in an earlier study very close linkage without recombination to the disease locus in an Irish pedigree with a clinically more severe and early onset (Type I) ADRP.

Chromosome Aberrations↗
PubMed1990

Exclusion of the autosomal dominant retinitis pigmentosa gene from a substantial region of chromosome 1: study of a large Australian family.

In an attempt to map the gene(s) responsible for autosomal dominant retinitis pigmentosa (ADRP), the technique of reverse genetics was used on a large multigenerational Australian pedigree. The family demonstrated a form of the disease which appears to be less severe than that observed in the Irish pedigree. It was typed for 10 restriction fragment length polymorphism (RFLP) markers on chromosome 1. The data from the linkage study was analysed using the programs LIPED 3; six markers gave informative results. The ADRP gene was excluded from this family from 102 cM using previously prepared chromosome 1 maps. This accounts for 36% of chromosome 1 which is estimated to be the longest human chromosome.

Adolescent↗
PubMed1989

Monoclonal antibodies to 33 KD protein of Mycobacterium tuberculosis with varying epitope specificities.

Five mouse monoclonal antibodies (McAb) 5D2, 5D5, 5F9, 3A8 and 3F2 against Mycobacterium tuberculosis (M. tb) 33 KD protein have been produced. All the McAbs except one were of IgG1, whereas the 3F2 was of IgM isotype. The ELISA and immunoblotting analysis show that 5D2, 5D5 and 5F9 are reactive to only 33 KD protein whereas 3A8 and 3F2 are also cross reactive to 29 KD and 29 KD + 64 KD respectively. ELISA inhibition assays using peroxidase labelled respective McAbs show that 5D2 and 5D5 recognize the same or overlapping epitopes whereas the remaining three McAbs (5F9, 3A8 and 3F2) are reactive to different epitopes of the 33 KD protein. These McAbs are cross reactive to the majority of mycobacterial strains other than M. tb including M. leprae (ML). However such reactivities may involve proteins of differing molecular sizes. Interestingly all these McAbs react to 25 KD and 29 KD of ML. The immunoreactivities of these McAbs to 33 KD appear to be directed to peptide epitopes but not to carbohydrate moiety. Both IgG and IgM antibody activities of lepromatous leprosy (L-lep) sera to 33 KD protein are heterogeneous and directed to more than 4 epitopes. The potential use of these McAbs in studying the pathomechanism of leprosy are currently being investigated.

Antibodies, Bacterial↗
PubMed1989

Concentration dependent functional responsiveness to subcellular antigenic extracts of BCG and different mycobacterial PPDs, of tuberculin (Mtb-PPD) reactive human T-cell lines.

M. tuberculosis purified protein derivative (Mtb-PPD) as tuberculin, other mycobacterial PPD-preparations and subcellular antigenic extracts of BCG showed varying concentration dependent bimodal effects in lymphoproliferative assay (LA) of Mtb-PPD generated human CD4+ T-cell lines (TCLS). Inhibitory effect on LA by high dose of Mtb-PPD is correlated with the inhibition of IL-2 production during the antigen induced stimulation. Consequently, maximal and inhibitory concentrations of different antigens in dose response LA varied for different TCLS. However, inhibitory effect can be overcome by the high concentration of non inhibitory antigen for the particular TCL. These results indicate that i) dose dependent LA is essential for the evaluation of antigen specificity of TCL; ii) some mycobacterial antigen may induce anergy of certain concentration whereas some others exert positive effect to overcome such anergy related to inhibition or production of IL-2.

Antigens, Bacterial↗