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1.
Brain Dev ; 38(1): 54-60, 2016 Jan.
Article in English | MEDLINE | ID: mdl-26071121

ABSTRACT

BACKGROUND: Glutaric aciduria type I is an autosomal recessive organic acid disorder. The primary defect is the deficiency of Glutaryl-CoA dehydrogenase (EC number 1.3.99.7) enzyme that is involved in the catabolic pathways of the amino acids l-lysine, l-hydroxylysine, and l-tryptophan. It is a treatable neuro-metabolic disorder. Early diagnosis and treatment helps in preventing brain damage. METHODS: The Glutaryl-CoA dehydrogenase gene (GCDH) gene was sequenced to identify disease causing mutations by direct sequencing of all the exons in twelve patients who were biochemically confirmed with GA I. RESULTS: We identified eleven mutations of which nine are homozygous mutations, one heterozygous and two synonymous mutations. Among the eleven mutations, four mutations p.Q162R, p.P286S, p.W225X in two families and p.V410M are novel. A milder clinical presentation is observed in those families who are either heterozygous or with a benign synonymous SNP. Multiple sequence alignment (MSA) of GCDH with its homologues revealed that the observed novel mutations are not tolerated by protein structure and function. CONCLUSIONS: The present study indicates genetic heterogeneity in GCDH gene mutations among South Indian population. Genetic analysis is useful in prenatal diagnosis and prevention. Mutation analysis is a useful tool in the absence of non-availability of enzyme assay in GA I.


Subject(s)
Amino Acid Metabolism, Inborn Errors/genetics , Brain Diseases, Metabolic/genetics , Glutaryl-CoA Dehydrogenase/deficiency , Glutaryl-CoA Dehydrogenase/genetics , Mutation , Amino Acid Metabolism, Inborn Errors/diagnosis , Asian People/genetics , Brain Diseases, Metabolic/diagnosis , DNA Mutational Analysis , Exons , Glutaryl-CoA Dehydrogenase/chemistry , Humans , India , Models, Molecular , Phenotype , Protein Conformation
2.
Exp Mol Pathol ; 90(1): 29-37, 2011 Feb.
Article in English | MEDLINE | ID: mdl-21111730

ABSTRACT

Retinoblastoma (RB), the most common intra-ocular malignancy in children under 5 years of age, has an estimated incidence of about 2000 a year in India, where most cases are in advanced stage at the time of diagnosis. Newer therapeutic approaches would reduce the morbidity of chemotherapy in children with RB. Fatty Acid Synthase (FASN), a lipogenic multi-enzyme complex, is minimally expressed in normal human tissues and over expressed in many cancers, making it an attractive target for cancer therapy. We analyzed RB tissues for FASN protein expression by immunohistochemistry, western blot, and ELISA, and FASN mRNA expression by RT-PCR. FASN expression was correlated with the clinico-pathological characteristics of the tumors. FASN immunostaining was positive in all the 44 RB tissues analyzed (100%). However, FASN expression was heterogeneous within the tumor samples. Tumors with invasion of choroid, optic nerve, orbit and/or retinal pigment epithelium showed significantly higher FASN immunoreactivity than the tumors without invasion (P<0.05), supported by western analysis (P<0.05). FASN expression was significantly high in poorly differentiated retinoblastomas (P<0.05). FASN protein and FASN mRNA estimated by ELISA and RT-PCR respectively showed multi-fold expression over the non-neoplastic muller glial cells that varied quantitatively between tumor tissues. FASN mRNA over-expression was substantially lower than the corresponding FASN protein expression values. The present study reports (i) markedly high expression of FASN protein in poorly differentiated and in invasive retinoblastomas, and (ii) multi-fold over-expression of FASN mRNA and protein in RB tissues, although at varying levels, indicating FASN to be a potential therapeutic target in retinoblastoma management.


Subject(s)
Fatty Acid Synthases/metabolism , Retinal Neoplasms/enzymology , Retinal Neoplasms/pathology , Retinoblastoma/enzymology , Retinoblastoma/pathology , Cell Differentiation , Child, Preschool , Cohort Studies , Female , Humans , India , Infant , Infant, Newborn , Male , Neoplasm Invasiveness , Retrospective Studies
3.
Indian J Biochem Biophys ; 47(3): 166-71, 2010 Jun.
Article in English | MEDLINE | ID: mdl-20653288

ABSTRACT

Triclosan, a broad spectrum antibiotic is currently being evaluated for its anti-cancer property. Though several solvents are available to dissolve lipophilic (hydrophobic) drugs, solubility and toxicity aspects pose a challenge, when combined with the cell culture medium. In this paper, we present a simple approach based on physico-chemical and biologic criteria to choose a suitable solubilizing agent to study the anti-proliferative property of triclosan in breast cancer cell line MCF-7. Triclosan was dissolved in five different solvents viz. DMSO, absolute ethanol, 1 N NaOH, 55% polyethylene glycol + 45% ethanol mixture (PEM) and acetone and diluted with the culture medium (1 mg/ml). Although triclosan dissolved completely in all five solvents, on dilution with culture medium, turbidity was observed in DMSO, 1 N NaOH and ethanol. Cell viability was 95.23% in 10 microl of acetone, when compared with 49.45% at the same volume of PEM. This non-toxic nature of acetone was supported by DNA fragmentation analysis and phase contrast microscopy. A significant decrease in cancer cell proliferation at 100 microg/ml of acetone-solubilized triclosan, compared with 100 microg/ml of PEM-solubilized triclosan (p<0.05) indicated stronger anti-proliferative effect and greater drug-sensitivity of triclosan when solubilized in acetone. Results showed that acetone-solubilized triclosan was suitable for anti-cancer investigations in cultured MCF-7 cells.


Subject(s)
Antineoplastic Agents/pharmacology , Triclosan/pharmacology , Breast Neoplasms/drug therapy , Breast Neoplasms/pathology , Cell Line, Tumor , Cell Proliferation/drug effects , Drug Screening Assays, Antitumor , Female , Humans , Hydrophobic and Hydrophilic Interactions , Solubility , Solvents
4.
J Ocul Biol Dis Infor ; 3(4): 117-28, 2010 Dec.
Article in English | MEDLINE | ID: mdl-23181152

ABSTRACT

Fatty acid biosynthesis is an attractive target for anti-cancer therapeutics. The ocular cancer, retinoblastoma cells were treated with fatty acid synthase (FASN) enzyme inhibitors: cerulenin, triclosan and orlistat. The IC(50) and dose-dependent sensitivity of cancer cells to FASN inhibitors decrease in biologic enzyme activity, and cell morphology alterations were analysed. Molecular interactions of enzyme-inhibitor complexes were studied by molecular modelling and docking simulations. The crystal structures of ketoacyl synthase (PDB ID:3HHD) (cerulenin) and thioesterase (PDB ID:2PX6) (orlistat) domains of human FASN were utilized for docking, while for the non-crystallised human FASN enoyl reductase domain (triclosan), homology model was built and used for docking. All three inhibitors showed significant binding energy indicating stable complex formation with their respective FASN subunits. The predicted Ki value of the FASN inhibitors corroborated well with their corresponding anti-cancer effects.

5.
Indian J Med Res ; 122(2): 165-70, 2005 Aug.
Article in English | MEDLINE | ID: mdl-16177475

ABSTRACT

BACKGROUND & OBJECTIVE: The conventional culture technique for diagnosis of extrapulmonary tuberculosis is time consuming. In order to find a sensitive and rapid technique nested polymerase chain reaction (nPCR) targeting the conserved MPB 64 gene of Mycobacterium tuberculosis was evaluated for detection of M. tuberculosis DNA directly from clinical specimens of extrapulmonary origin. METHODS: A total of 400 clinical specimens from clinically suspected cases of extrapulmonary tuberculosis and 30 control specimens of nontuberculous aetiology were processed by smear and culture and by nPCR technique for detection of M. tuberculosis. The specimens were divided into 3 groups, (group I--280 specimens [104 peritoneal fluid (PF), 120 cerebrospinal fluid (CSF), 44 lymph node biopsies 3 pericardial fluid and 9 other biopsy specimens], group II--120 aqueous humour (AH) from idiopathic granulomatous uveitis cases, and group III--30 control specimens (10 CSF and 20AH). RESULTS: The conventional culture was positive only in 16 of 400 specimens. The overall positivity of nPCR was 35.2 per cent (141/400). Among the 280 specimens from extrapulmonary lesions (group I), 15 were bacteriologically positive, while 115 of 265 bacteriologically negative specimens (43.4%) were positive by nPCR. All the 30 control specimens were negative by nPCR. INTERPRETATION & CONCLUSION: The nPCR using MPB64 gene primers might be a rapid and reliable diagnostic technique for detection of M. tuberculosis genome in clinically suspected extrapulmonary tuberculosis specimens, as compared to the conventional techniques.


Subject(s)
Bacteriological Techniques , DNA Primers/chemistry , Mycobacterium tuberculosis/genetics , Mycobacterium tuberculosis/metabolism , Polymerase Chain Reaction/methods , Tuberculosis/diagnosis , Tuberculosis/microbiology , Aqueous Humor/microbiology , Case-Control Studies , Cerebrospinal Fluid/microbiology , DNA, Bacterial/metabolism , Humans , Lymph Nodes/microbiology , Oligonucleotide Probes/chemistry , Tuberculosis, Ocular/diagnosis , Tuberculosis, Ocular/metabolism , Uveitis/microbiology
7.
Adv Space Res ; 34(6): 1433-7, 2004.
Article in English | MEDLINE | ID: mdl-15881788

ABSTRACT

The existence of significant fluxes of antiparticles in the Earth magnetosphere has been predicted on theoretical considerations in this article. These antiparticles (positrons or antiprotons) at several hundred kilometers of altitudes, we believe are not of direct extraterrestrial origin, but are the natural products of nuclear reactions of the high energy primary cosmic rays (CR) and trapped protons (TP) confined in the terrestrial radiation belt, with the constituents of terrestrial atmosphere. Extraterrestrial positrons and antiprotons born in nuclear reactions of the same CR particles passing through only 5-7 g/cm2 of interstellar matter, exhibit lower fluxes compared to the antiprotons born at hundreds of g/cm2 in the atmosphere, which when confined in the magnetic field of the Earth (in any other planet), get accumulated. We present the results of the computations of the antiproton fluxes at 10 MeV to several GeV energies due to CR particle interactions with the matter in the interstellar space, and also with the residual atmosphere at altitudes of approximately 1000 km over the Earth's surface. The estimates show that the magnetospheric antiproton fluxes are greater by two orders of magnitude compared to the extraterrestrial fluxes measured at energies <1-2 GeV.


Subject(s)
Cosmic Radiation , Electrons , Elementary Particles , Protons , Atmosphere , Elementary Particle Interactions , Extraterrestrial Environment , Models, Theoretical , Nuclear Physics
8.
Indian J Pathol Microbiol ; 44(2): 97-102, 2001 Apr.
Article in English | MEDLINE | ID: mdl-11883143

ABSTRACT

Nucleic acid amplification using IS6110 primers to detect Mycobacterium tuberculosis in clinical specimens has been extensively used as laboratory tool for the diagnosis for tuberculosis. Despite it's dramatic scientific value in practice, it is not as sensitive as expected for the detection of M. tuberculosis. The results of the study suggest that PCR using 123 bp fragment of DNA belonging to IS6110 is specific (95.6%) but only has a sensitivity of 30% to detect M. tuberculosis in clinical specimens.


Subject(s)
Mycobacterium tuberculosis/genetics , Mycobacterium tuberculosis/isolation & purification , Polymerase Chain Reaction/methods , Base Sequence , DNA Primers/genetics , DNA Transposable Elements/genetics , DNA, Bacterial/genetics , Diagnostic Errors , Humans , Polymerase Chain Reaction/statistics & numerical data , Sensitivity and Specificity , Tuberculosis/diagnosis
9.
Invest Ophthalmol Vis Sci ; 41(3): 822-5, 2000 Mar.
Article in English | MEDLINE | ID: mdl-10711699

ABSTRACT

PURPOSE: Tuberculous etiology has been suggested in Eales' disease. Because epiretinal membrane (ERM) is formed on the inner surface of the retina in Eales' disease, it could be the most appropriate intraocular specimen for investigation. Therefore, a nested polymerase chain reaction (nPCR), which detects MPB64 gene of Mycobacterium tuberculosis on the archival specimens of ERM of well-documented Eales' and non-Eales' patients, was applied and the results compared. METHODS: nPCR technique was standardized, and the sensitivity and specificity of the primers were determined. nPCR technique was applied to tissue sections obtained from formalin-fixed and paraffin-embedded tissues of ERM from 23 patients with Eales' disease and 27 noninfective and non-Eales' disease patients as controls. RESULTS: nPCR technique was specific for M. tuberculosis genome and sensitive enough to detect 0.25 fg (corresponding to the presence of a single bacillus). Eleven (47.8%) ERM of 23 Eales' disease and 3 (11.1%) of 27 controls were positive for M. tuberculosis genome. The difference between the two groups was statistically significant (P = 0.001), indicating association of this bacterium with Eales' disease. CONCLUSIONS: The demonstration of the presence of M. tuberculosis DNA by nPCR technique in significant number of ERM of Eales' disease compared with the controls further emphasizes the probable role of this bacterium in the pathogenesis of this enigmatic clinical condition.


Subject(s)
Antigens, Bacterial , Bacterial Proteins/genetics , DNA, Bacterial/analysis , Epiretinal Membrane/microbiology , Mycobacterium tuberculosis/genetics , Phlebitis/microbiology , Retinal Vein , Tuberculosis, Ocular/microbiology , DNA Primers/chemistry , Humans , Polymerase Chain Reaction/methods , Retinal Hemorrhage/microbiology , Sensitivity and Specificity
10.
Indian J Pathol Microbiol ; 43(4): 395-402, 2000 Oct.
Article in English | MEDLINE | ID: mdl-11344601

ABSTRACT

Nucleic acid amplification using IS6110 primers to detect Mycobacterium tuberculosis in clinical specimens has been extensively used as laboratory tool for the diagnosis for tuberculosis. Despite it's dramatic scientific value in practice, it is not as sensitive as expected for the detection of Mycobacterium tuberculosis. The results of the study suggest that PCR using 123 bp fragment of DNA belonging to IS6110 is specific (95.6%) but only has a sensitivity of 30% to detect M. tuberculosis in clinical specimens.


Subject(s)
DNA Transposable Elements/genetics , Mycobacterium tuberculosis/genetics , Mycobacterium tuberculosis/isolation & purification , Polymerase Chain Reaction/methods , Tuberculosis/microbiology , DNA, Bacterial/analysis , Humans , Sensitivity and Specificity
11.
Nature ; 229(5282): 248-50, 1971 Jan 22.
Article in English | MEDLINE | ID: mdl-16059180
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