[Show abstract][Hide abstract] ABSTRACT: Mucopolysaccharidosis IV A (Morquio syndrome A, MPS IVA) is a lysosomal storage disease caused by the deficiency of N-acetylgalactosamine-6-sulfatase (GALNS). The mutation spectrum in this condition is yet to be determined in Indians. We aimed to analyze the mutations in the GALNS gene in Asian Indians with MPS IVA. All the exons and the adjacent intronic regions of the gene were amplified and sequenced in sixty-eight unrelated Indian families. We identified 136 mutant alleles comprising of 40 different mutations. We report twenty-two novel mutations that comprise of seventeen missense (p.Asn32Thr, p.Leu36Arg, p.Pro52Leu, p.Pro77Ser, p.Cys79Arg, p.His142Pro, p.Tyr191Asp, p.Asn204Thr, p.Gly188Ser, p.Phe216Ser, p.Trp230Cys, p.Ala291Ser, p.Gly317Arg, p.His329Pro, p.Arg386Ser, p.Glu450Gly, p.Cys501Ser), three splice-site variants (c.120 + 1G > C, c.1003-3C > G, c.1139 + 1G > A), one nonsense mutation (p.Gln414*) and one frameshift mutation (p.Pro420Leufs*440). Eighteen mutations have been reported earlier. Among these p.Ser287Leu (8.82%), p.Phe216Ser (7.35%), p.Asn32Thr (6.61%) and p.Ala291Ser (5.88%) were the most frequent mutations in Indian patients but were rare in the mutational profiles reported in other populations. These results indicate that the Indian patients may have a distinct mutation spectrum compared to those of other populations. Mutant alleles in exon 1, 7 and 8 accounted for 44.8% of the mutations, and sequencing of these exons initially may be a cost-effective approach in Asian Indian patients. This is the largest study on molecular analysis of patients with MPS IVA reported in the literature, and the first report from India
American Journal of Medical Genetics Part A 09/2014; · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Constitutional type of an individual or prakriti is the basic clinical denominator in Ayurveda, which defines physical, physiological, and psychological traits of an individual and is the template for individualized diet, lifestyle counseling, and treatment. The large number of phenotype description by prakriti determination is based on the knowledge and experience of the assessor, and hence subject to inherent variations and interpretations.
Journal of Ayurveda and integrative medicine 09/2014; 5(3):167-175.
[Show abstract][Hide abstract] ABSTRACT: ABSTRACT Acute lymphoblastic leukemia (ALL) arises due to several genetic alterations in progenitor cells and methotrexate is frequently used as part of the treatment regimen. Although there are evidences for the effect of MTHFR C677T and A1298C variations on the drug response in ALL, its risk association for ALL is still unresolved. In a case-control study of total 203 ALL patients and 246 controls and meta-analysis on Indian population, we show an insignificant association of MTHFR C677T and A1298C genotypes with childhood and adult ALL. A comprehensive insilico characterization of non-synonymous single nucleotide polymorphisms (nsSNPs) and SNPs of 3'UTR region revealed 9 nsSNPs as deleterious and 3SNPs in 3'UTR region could possibly alter the binding of miRNA. The study revealed that several overlooked SNPs may contribute to risk of ALL susceptibility and further studies of these SNPs with functional characterization in large sample size is required to understand the significant role of MTHFR gene in ALL development.
[Show abstract][Hide abstract] ABSTRACT: While virulence factors and the biofilm forming capabilities of microbes are the key regulators of wound healing process, the host immune response may also contribute in the events following wound closure or exacerbation of non-closure. We examined samples from diabetic and non-diabetic foot ulcers/wounds for microbial association and tested the microbes for their antibiotic susceptibility and ability to stimulate biofilm formation. A total of 1074 bacterial strains were obtained with staphylococci, Pseudomonas, Citrobacter and enterococci as major colonisers in diabetic samples. Though non-diabetic samples had a similar assemblage, the frequency of occurrence of different groups of bacteria was different. Gram negative bacteria were found to be more prevalent in the diabetic wound environment while, Gram positive bacteria were predominant in non-diabetic ulcers. A higher frequency of monomicrobial infection was observed in samples from non-diabetic individuals when compared to samples from diabetic patients. The prevalence of different groups of bacteria varied when the samples were stratified according to age and sex of the individuals. Several multidrug resistant strains were observed among the samples tested and most of these strains had the ability to elicit moderate to high levels of biofilm. The weakened immune response in diabetic individuals and synergism among pathogenic microorganisms may be critical factors that may determine the delicate balance of wound healing process.
Journal of Medical Microbiology 07/2014; · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Objectives: Biofilms, secretions and excretions from bacteria such as Pseudomonas aeruginosa infecting the wound exacerbate or inhibit various stages of wound repair (such as the inflammatory and proliferative phases) resulting in delayed healing or in total inability of the wound to completely heal. Therefore we set out to study not only the antibiotic susceptibility and minimum inhibitory concentrations of bacteria forming biofilms but also the effect of their secretions and excretions on various cell types involved in wound healing such as fibroblasts and keratinocytes. Methods: The biofilm forming ability of 27 strains of Pseudomonas aeruginosa were assessed using the micro-titer plate method. Three strains were selected on the basis of their biofilm forming ability and further analyzed by checking their antibiotic susceptibility profiles by disk diffusion method. Pulsed field gel electrophoresis was then performed to type the strains. WST-1 and Wound Scratch assays were performed to study the viability, migration and proliferation of fibroblast and keratinocyte cells in the presence of lyophilized metabolites and toxins of the bacterial strains. Results: The Pseudomonas aeruginosa strains screened had high, mid-range and low biofilm formers. The three strains selected for further analysis were Multi Drug Resistant (MDR). A positive correlation between the biofilm forming ability of a strain and the inhibition of migration and proliferation of human fibroblasts and keratinocytes was observed.
24th European Congress of Clinical Microbiology and Infectious Diseases, Barcelona, Spain; 05/2014
[Show abstract][Hide abstract] ABSTRACT: Double C2 like Domain Beta (DOC2B) gene encodes for a calcium binding protein which is involved in neurotransmitter release, sorting and exocytosis. We have identified promoter region of DOC2B gene as hypermethylated in premalignant, malignant cervical tissues and cervical cancer cell lines by methylation sensitive dimethyl sulfoxide polymerase chain reaction (MS DMSO PCR) and bisulfite genome sequencing (BGS); while, it was unmethylated in normal cervical tissues (P<0.05). The promoter hypermethylation was inversely associated with mRNA expression in SiHa, CaSki and HeLa cells and treatment with demethylating agent 5 aza 2 deoxycytidine restored DOC2B expression. The region -630 to +25bp of DOC2B gene showed robust promoter activity by luciferase reporter assay and was inhibited by in-vitro artificial methylation with Ss1 methylase prior to transient transfections. The overexpression of DOC2B gene in SiHa cells when compared to controls, showed significantly reduced colony formation, cell proliferation, induced cell cycle arrest and repressed cell migration and invasion (P<0.05). Ectopic expression of DOC2B resulted in anoikis mediated cell death and repressed tumor growth in a nude mice xenograft model (P<0.05). DOC2B expressing cells showed a significant increase in intracellular calcium level (P<0.05), impaired AKT 1 and ERK1/2 signaling and induced actin cytoskeleton remodeling. Our result show that promoter hypermethylation and silencing of DOC2B gene is an early and frequent event during cervical carcinogenesis and whose reduced expression due to DNA promoter methylation may lead to selective cervical tumor growth.
Journal of Biological Chemistry 02/2014; · 4.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Aim:MTHFR mediates the one carbon metabolism pathway. Two common genetic variants, C677T and A1298C, of MTHFR are associated with number of human diseases, including cancer, as well as being involved in the modulation of therapy outcome to antifolate drugs. To understand the distribution pattern of SNPs among different tissues of an individual, we examined MTHFR polymorphisms in normal and colon cancer tissues and compared the genotype frequencies in peripheral blood samples. Materials & methods: DNA was isolated from tumor tissue and matched normal tissues from 155 colon cancer patients. These samples as well as DNA from blood samples of the control group (n = 294) were analyzed for MTHFR polymorphisms by PCR-RFLP and confirmed by a direct DNA sequencing method. Results: Our data suggest that the allele and genotype frequencies of C677T and A1298C were significantly different between tumor tissues and both types of normal tissues. We have established that MTHFR variants that exist in tumor and matched normal tissues of colon cancer patients differ suggesting somatic variation in MTHFR polymorphisms among different tissues of an individual. The MTHFR A1298C polymorphism was associated with risk of colon cancer. Conclusion: Different MTHFR variants may exist in different tissues to maintain physiological functions and may have implications for disease susceptibility and pharmacogenomics based therapies. Original submitted 21 January 2013; Revision submitted 3 January 2014.
[Show abstract][Hide abstract] ABSTRACT: Autofluorescence characteristics of human serum albumin (HSA) are highly sensitive to its local environment. Identification and characterization of the proteins in normal and disease conditions may have great clinical implications. Aim of the present study was to understand how autofluorescence properties of HSA varies with denaturation under urea (3.0M, 6.0M, 9.0M) and guanidine hydrochloride (GnHCl) (2.0M, 4.0M, 6.0M) as well as digestion with trypsin. Towards this, we have recorded the corresponding autofluorescence spectra of HSA at 281nm laser excitation and compared the outcomes. Although, HSA contains 1 tryptophan and 17 tyrosine residues, it has shown intense autofluorescence due to tryptophan as compared to the tyrosine in native form, which may be due to the fluorescence resonance energy transfer (FRET) from tyrosine to tryptophan. As the unfolding progresses in denatured and digested forms of the protein, a clear increase in tyrosine fluorescence as compared to tryptophan was observed, which may be due to the increase of tryptophan - tyrosine separation disturbing the FRET between them resulting in differences in the overall autofluorescence properties. The decrease in tryptophan fluorescence of around 17% in urea denatured, 32% in GnHCl denatured and 96% in tryptic digested HSA was observed as compared to its native form. The obtained results show a clear decrease in FRET between tyrosine and tryptophan residues with the progression of unfolding and urea seems to be less efficient than GnHCl in unfolding of HSA. These results demonstrate the potential of autofluorescence in characterizing proteins in general and HSA in particular.