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Biomedical subjects

J Chandra

Publications and source records attributed to J Chandra.

At least 19 recordsLinked to original sources
PubMed2003

Uses and limitations of the XTT assay in studies of Candida growth and metabolism.

Colorimetric tetrazolium assays are used increasingly in studies of fungi, often in the absence of standardization or correlation with other methods. We examined species- and strain-related tetrazolium metabolism in Candida albicans and Candida parapsilosis by using XTT [2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-2H-tetrazolium hydroxide] and WST-8 [2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulphonyl)-2H-tetrazolium] and found marked variations. Also, significant signal was often missed in the absence of dimethyl sulfoxide extraction.

Candida↗
PubMed2002

Antifungal susceptibility of Candida biofilms: unique efficacy of amphotericin B lipid formulations and echinocandins.

Biofilms, likely the predominant mode of device-related microbial infection, exhibit resistance to antimicrobial agents. Evidence suggests that Candida biofilms have dramatically reduced susceptibility to antifungal drugs. We examined antifungal susceptibilities of Candida albicans and Candida parapsilosis biofilms grown on a bioprosthetic model. In addition to conventional agents, we determined if new antifungal agents (triazoles, amphotericin B lipid formulations, and echinocandins) have activities against Candida biofilms. We also explored effects of preincubation of C. albicans cells with subinhibitory concentrations (sub-MICs) of drugs to see if they could modify subsequent biofilm formation. Finally, we used confocal scanning laser microscopy (CSLM) to image planktonic- and biofilm-exposed blastospores to examine drug effects on cell structure. Candida biofilms were formed on silicone elastomer and quantified by tetrazolium and dry weight (DW) assays. Susceptibility testing of fluconazole, nystatin, chlorhexidine, terbenafine, amphotericin B (AMB), and the triazoles voriconazole (VRC) and ravuconazole revealed resistance in all Candida isolates examined when grown as biofilms, compared to planktonic forms. In contrast, lipid formulations of AMB (liposomal AMB and AMB lipid complex [ABLC]) and echinocandins (caspofungin [Casp] and micafungin) showed activity against Candida biofilms. Preincubation of C. albicans cells with sub-MIC levels of antifungals decreased the ability of cells to subsequently form biofilm (measured by DW; P < 0.0005). CSLM analysis of planktonic and biofilm-associated blastospores showed treatment with VRC, Casp, and ABLC resulted in morphological alterations, which differed with each agent. In conclusion, our data show that Candida biofilms show unique susceptibilities to echinocandins and AMB lipid formulations.

Amphotericin B↗
PubMed2002

Comparison of biofilms formed by Candida albicans and Candida parapsilosis on bioprosthetic surfaces.

Little is known about fungal biofilms, which may cause infection and antibiotic resistance. In this study, biofilm formation by different Candida species, particularly Candida albicans and C. parapsilosis, was evaluated by using a clinically relevant model of Candida biofilm on medical devices. Candida biofilms were allowed to form on silicone elastomer and were quantified by tetrazolium (XTT) and dry weight (DW) assays. Formed biofilm was visualized by using fluorescence microscopy and confocal scanning laser microscopy with Calcofluor White (Sigma Chemical Co., St. Louis, Mo.), concanavalin A-Alexafluor 488 (Molecular Probes, Eugene, Oreg.), and FUN-1 (Molecular Probes) dyes. Although minimal variations in biofilm production among invasive C. albicans isolates were seen, significant differences between invasive and noninvasive isolates (P < 0.001) were noted. C. albicans isolates produced more biofilm than C. parapsilosis, C. glabrata, and C. tropicalis isolates, as determined by DW assays (P was <0.001 for all comparisons) and microscopy. Interestingly, noninvasive isolates demonstrated a higher level of XTT activity than invasive isolates. On microscopy, C. albicans biofilms had a morphology different from that of other species, consisting of a basal blastospore layer with a dense overlying matrix composed of exopolysaccharides and hyphae. In contrast, C. parapsilosis biofilms had less volume than C. albicans biofilms and were comprised exclusively of clumped blastospores. Unlike planktonically grown cells, Candida biofilms rapidly (within 6 h) developed fluconazole resistance (MIC, >128 microg/ml). Importantly, XTT and FUN-1 activity showed biofilm cells to be metabolically active. In conclusion, our data show that C. albicans produces quantitatively larger and qualitatively more complex biofilms than other species, in particular, C. parapsilosis.

Antifungal Agents↗
PubMed2001

Cloning and expression of a novel human glutaredoxin (Grx2) with mitochondrial and nuclear isoforms.

Glutaredoxin (Grx) is a glutathione-dependent hydrogen donor for ribonucleotide reductase. Today glutaredoxins are known as a multifunctional family of GSH-disulfide-oxidoreductases belonging to the thioredoxin fold superfamily. In contrast to Escherichia coli and yeast, a single human glutaredoxin is known. We have identified and cloned a novel 18-kDa human dithiol glutaredoxin, named glutaredoxin-2 (Grx2), which is 34% identical to the previously known cytosolic 12-kDa human Grx1. The human Grx2 sequence contains three characteristic regions of the glutaredoxin family: the dithiol/disulfide active site, CSYC, the GSH binding site, and a hydrophobic surface area. The human Grx2 gene, located at chromosome 1q31.2--31.3, consisted of five exons that were transcribed to a 0.9-kilobase human Grx2 mRNA ubiquitously expressed in several tissues. Two alternatively spliced Grx2 mRNA isoforms that differed in their 5' region were identified. These corresponded to alternative proteins with a common 125-residue C-terminal Grx domain but with different N-terminal extensions of 39 and 40 residues, respectively. The 125-residue Grx domain and the two full-length variants were expressed in E. coli and exhibited GSH-dependent hydroxyethyl disulfide and dehydroascorbate reducing activities. Western blot analysis of subcellular fractions from Jurkat cells with a specific anti-Grx2 antibody showed that human Grx2 was predominantly located in the nucleus but also present in the mitochondria. We further showed that one of the mRNA isoforms corresponding to Grx2a encoded a functional N-terminal mitochondrial translocation signal.

Amino Acid Sequence↗
PubMed2001

Pulmonary function tests in beta thalassemia.

To study pulmonary function tests (PFT) in multiple transfusion recipient thalassemics, PFTs were done for 30 thalassemics and 20 matched controls. Confirmed cases of thalassemia on regular transfusion therapy were the subject of study. Apart from history and physical examination of the thalassemics, serum ferritin estimation and spirometry were done. Parameters studied included lung volumes--functional residual capacity (FRC), forced vital capacity (FVC), residual volume (RV) and total lung capacity (TLC); and flow rates--forced expiratory volume in one second (FEV1), forced expiratory volume in 1 second/forced vital capacity (FEV1/FVC), peak expiratory flow 25-75 (PEF 25-75%) and peak expiratory flow rate (PEFR). Single breath carbon monoxide diffusing capacity (DLco) and arterial blood gas (ABG) were also analysed. The mean height and weight of thalassemics were below that of age matched controls. A restrictive abnormality in PFT was found in 86.6% cases. These patients were found to have a decrease in all the lung volumes namely FVC, FRC, RV and TLC with a proportional decrease in the flow rates, FEV1, PEF 25-75% and PEF with a normal (> 0.75) FEV1/FVC ratio. DLco was decreased in all the patients with restrictive lung disease and fall in DLco showed a good correlation (r = 0.7, P < .001) with the severity of restrictive disease suggesting that some intrapulmonary pathology is likely to be responsible for the restrictive pattern. None of the cases had an obstructive or mixed pattern of pulmonary dysfunction. No correlation was found between severity of restrictive disease and the serum ferritin levels. A negative correlation with degree of hepatosplenomegaly was found. No correlation was found between severity of the defect and age, number of blood transfusions received and hemoglobin at the time of doing the test. To conclude, restrictive lung disease is the predominant abnormality in multi-transfused thalassemics, which is probably due to pulmonary parenchymal pathology. The abnormality of PFTs is not directly related to iron overload.

Adolescent↗
PubMed2001

Pharmacological basis for cladribine resistance in a human acute T lymphoblastic leukaemia cell line selected for resistance to etoposide.

Cross-resistance between different classes of anti-neoplastic agents can jeopardize successful combination cancer chemotherapy. In this study, we observed an unexpected cross-resistance between the podophyllotoxine derivative etoposide (VP) and the nucleoside analogue cladribine (CdA) in CCRF-CEM cells developed for resistance to VP. The resistant cells also displayed 14- and twofold resistance to cytarabine (ara-C) and gemcitabine respectively. Closer analysis of these cells showed that they contained lower amounts of topoisomerase (topo) IIalpha (P < 0.001) and beta protein (P < 0.026), formed substantially lower amounts of the topo II-DNA complex, and had a markedly decreased level of Fas (CD95/APO-1)-ligand mRNA expression. Interestingly, Fas expression in the resistant cells did not differ from that in the parental cell line. No differences were observed in the accumulation/efflux of daunorubicin or in the gene expressions of P-glycoprotein, multidrug resistance-associated protein and the lung resistance-related protein. The activity of deoxycytidine kinase (dCK), responsible for activation of CdA and ara-C, was the same for resistant and wild-type cells. However, there was an increase in the activity of the cytosolic 5'-nucleotidases (5'-NT), responsible for deactivation of nucleotides, amounting to 206% (P < 0.001) for the high Km and 134% (P < 0.331) for the low Km 5'-NT in resistant cells. The high Km 5'-NT is probably responsible for the decreased amount of the active metabolite CdA 5'-triphosphate [40% decreased (P < 0.045)], as well as for other purine ribonucleosides and deoxyribonucleosides triphosphates in the resistant cells. In contrast, a significantly higher deoxycytidine triphosphate (dCTP) level (167%, P < 0.001) was observed in the resistant cells. Thus, this study suggests that the major cause of resistance to the nucleoside analogues CdA and ara-C in cells selected for resistance to VP is a result of metabolic alterations producing increased activity of 5'-NT and higher dCTP levels. Furthermore, these results indicate that there is a common factor in the regulation of nucleotide-degrading enzymes and DNA topoisomerases, which may be altered in cross-resistant cells.

5'-Nucleotidase↗
PubMed2001

Endophthalmitis in the western Sydney region: a case-control study.

BACKGROUND: A retrospective case-control study was conducted to investigate risk factors for endophthalmitis following routine intraocular surgery. METHODS: A review was performed of consecutive cases of endophthalmitis from three teaching hospitals in the western Sydney region and matched controls from the same institutions between 1996 and 1998. RESULTS: There were 31 cases and 66 controls. Eighty procedures were phacoemulsification, 15 conventional extracapsular cataract extraction, and two were penetrating keratoplasties. Of the 80 patients who had phacoemulsification surgery, 50 had a clear corneal incision, and 26 had a scleral incision (four were unknown). Logistic regression showed an increased risk of endophthalmitis with surgical complications (P = 0.002) and clear cornea temporal incisions (P = 0.007). Risk of endophthalmitis was reduced with use of subconjunctival injections (P = 0.008). The yield for the Gram stain was 47% and for culture was 67%. Anterior chamber tap in addition to vitreous biopsy alone did not increase the yield for microorganism (P = 0.78). Mean visual acuity on presentation was hand movement with 13 patients (50%) showing visual improvement following intravitreal injections of antibiotics (P = 0.003). Visual prognosis did not correlate with presenting visual acuity but appeared to be better in those who grew Staphylococcus epidermidis or were culture negative. CONCLUSIONS: Although this study is unable to draw definite conclusions regarding risk of endophthalmitis in clear corneal temporal cataract surgery, sufficient data suggest the importance of incision type and location. Surgical complication is an important risk factor for endophthalmitis. Use of subconjunctival antibiotic injections at the conclusion of the procedure is recommended.

Aged↗
PubMed2001

Apoptosis and the response to anticancer therapy.

Apoptosis is a distinctive form of cell death that reflects cleavage of a subset of intracellular polypeptides by proteases known as caspases. Two major intracellular caspase cascades, one activated predominantly by death receptor ligands and the other triggered by various cellular stresses, including DNA damage and microtubule disruption, have been delineated. Activation of these protease cascades is tightly regulated by a number of polypeptides, including Bcl-2 family members, inhibitor of apoptosis proteins, and several protein kinases. The demonstration that many antineoplastic agents induce apoptosis in susceptible cells raises the possibility that factors affecting caspase activation and activity might be important determinants of anticancer drug sensitivity. Here, we review recent studies describing the regulation of apoptotic pathways and identify potential implications of these findings for resistance to antineoplastic agents.

Antineoplastic Agents↗
PubMed2001

Biofilm formation by the fungal pathogen Candida albicans: development, architecture, and drug resistance.

Biofilms are a protected niche for microorganisms, where they are safe from antibiotic treatment and can create a source of persistent infection. Using two clinically relevant Candida albicans biofilm models formed on bioprosthetic materials, we demonstrated that biofilm formation proceeds through three distinct developmental phases. These growth phases transform adherent blastospores to well-defined cellular communities encased in a polysaccharide matrix. Fluorescence and confocal scanning laser microscopy revealed that C. albicans biofilms have a highly heterogeneous architecture composed of cellular and noncellular elements. In both models, antifungal resistance of biofilm-grown cells increased in conjunction with biofilm formation. The expression of agglutinin-like (ALS) genes, which encode a family of proteins implicated in adhesion to host surfaces, was differentially regulated between planktonic and biofilm-grown cells. The ability of C. albicans to form biofilms contrasts sharply with that of Saccharomyces cerevisiae, which adhered to bioprosthetic surfaces but failed to form a mature biofilm. The studies described here form the basis for investigations into the molecular mechanisms of Candida biofilm biology and antifungal resistance and provide the means to design novel therapies for biofilm-based infections.

Antifungal Agents↗
PubMed2001

Antifungal resistance of candidal biofilms formed on denture acrylic in vitro.

Denture biofilms represent a protective reservoir for oral microbes. The study of the biology of Candida in these biofilms requires a reliable model. A reproducible model of C. albicans denture biofilm was developed and used to determine the susceptibility of two clinically relevant C. albicans isolates against 4 antifungals. C. albicans, growing as a biofilm, exhibited resistance to amphotericin B, nystatin, chlorhexidine, and fluconazole, with 50% reduction in metabolic activity (50% RMA) at concentrations of 8, 16, 128, and > 64 microg/mL, respectively. In contrast, planktonically cultured C. albicans were susceptible (50% RMA for the same antifungals was obtained at 0.25, 1.0, 4.0, and 0.5 microg/mL, respectively). In conclusion, results obtained by means of our biofilm model show that biofilm-associated C. albicans cells, compared with cells grown in planktonic form, are resistant to antifungals used to treat denture stomatitis.

Acrylic Resins↗
PubMed2001

Role of apoptosis in pancreatic beta-cell death in diabetes.

Apoptosis is a physiological form of cell death that occurs during normal development, and critical mediators of this process include caspases, reactive oxygen species, and Ca2+. Excessive apoptosis of the pancreatic beta-cell has been associated with diabetes. Consequently, apoptosis research has focused on how infiltrating macrophages or cytotoxic T-cells might kill pancreatic beta-cells using cytokines or death receptor triggering. Meanwhile, the intracellular events in the target beta-cell have been largely ignored. Elucidation of such targets might help develop improved treatment strategies for diabetes. This article will outline recent developments in apoptosis research, with emphasis on mechanisms that may be relevant to beta-cell death in type 1 and type 2 diabetes. Several of the models proposed in beta-cell killing converge on Ca2+ signaling, indicating that the pancreatic beta-cell may be an ideal system in which to carefully dissect the role of Ca2+ during apoptosis.

Animals↗
PubMed2001

Effects of serum from patients with type 1 diabetes on primary cerebellar granule cells.

Type 1 diabetes is an autoimmune disease of unknown etiology. Our previous work has shown that a factor present in serum from type 1 diabetic patients causes increased Ca2+ channel activity and apoptotic DNA fragmentation in pancreatic beta-cells. Here we examined the effects of type 1 diabetic serum on primary cerebellar granule cells (CGCs). In CGCs, exposure to type 1 diabetic serum did not cause increased apoptosis or changes in Ca2+ channel activity. However, patient serum did cause modulation of Ca2+ signals in a cell type with triangular soma that exhibited low voltage-gated Ca2+ currents. This cell was present primarily in cultures exposed to type 1 diabetic serum. The presence of low voltage-gated Ca2+ currents and long neuronal dendrites indicated that this unique cell was of neuronal origin and not of glial origin.

Animals↗
PubMed2001

Neuroglial cyst: a rare occurence in abdomen and pelvis.

We hereby report a case of a child who presented with abdominal distension and urinary retention. Radiological investigations and exploratory laparotomy revealed a cystic mass in the sacral region pushing the urinary bladder anteriorly and upwards. Histopathological examination revealed neuroglial elements. Neuroglial cysts are uncommon outside the central nervous system and this is the first report of its occurrence in abdomen and pelvis.

Abdomen↗
PubMed2000

Idiopathic pulmonary hemosiderosis.

Two cases of idiopathic pulmonary hemosiderosis in children are reported. Both cases presented with a combination of acute/recurrent respiratory symptoms along with iron deficiency anemia. In one case diagnosis was delayed for 18 months after onset of symptoms. After initial stabilisation with corticosteroid therapy, both cases were put on chloroquine therapy and showed improvement. Various other forms of therapy and outcome are discussed. It is suggested that in cases with recurrent respiratory symptoms, idiopathic pulmonary hemosiderosis should also be considered after excluding infectious etiology.

Adrenal Cortex Hormones↗
PubMed2000

Triggering and modulation of apoptosis by oxidative stress.

Cell survival requires multiple factors, including appropriate proportions of molecular oxygen and various antioxidants. Although most oxidative insults can be overcome by the cell's natural defenses, sustained perturbation of this balance may result in either apoptotic or necrotic cell death. Numerous, recent studies have shown that the mode of cell death that occurs depends on the severity of the insult. Oxidants and antioxidants can not only determine cell fate, but can also modulate the mode of cell death. Effects of oxidative stress on components of the apoptotic machinery may mediate this modulation. This review will address some of the current paradigms for oxidative stress and apoptosis, and discuss the potential mechanisms by which oxidants can modulate the apoptotic pathway.

Animals↗