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Biomedical subjects

Rajiv Kumar

Publications and source records attributed to Rajiv Kumar.

At least 109 records · Page 6Linked to original sources
PubMed2003

Differences in transient outward current properties between neonatal and adult human atrial myocytes.

UNLABELLED: Knowledge of postnatal modulation of I(to) in human atrial myocytes is quite limited. The present study investigated the differences in I(to) properties between neonatal and adult human atrial myocytes. METHODS: Atrial myocytes were dissociated enzymatically from biopsies of human right atrial appendage. I(to) and action potentials were recorded by whole-cell patch-clamp technique. The expressed protein levels of Kv4.3 and KChIP2 in atrial tissue were detected by western blot technique. RESULTS: I(to) was present in all atrial cells (n = 37) from 10 neonatal patients (2.5-7 months). The mean value of I(to) density in neonatal atrial cells was significantly larger than in adult atrial cells. The time constants for I(to) current decay were significantly faster for neonatal cells, compared to adult cells. I(to) recovery from inactivation at holding potential of - 80 mV was significantly slower for neonatal atrial cells than for adult atrial cells. There was no difference in the voltage dependence of I(to) activation between neonatal and adult cells. The voltage-dependent inactivation slope factor was smaller for neonatal compared to adult atrial cells. A more significant frequency-dependent suppression of I(to) peak current and a more significant lengthening of APD(30) were observed in neonatal atrial cells compared to adult atrial cells. Western blots showed both Kv4.3 and KChIP2 are expressed in neonatal atria, but with significantly higher level of Kv4.3 and lower level of KChIP2 protein compared to adult. CONCLUSION: There are significant differences in the properties of I(to) between neonatal and adult human atrial cells, including a larger current density, faster inactivation and slower recovery from inactivation in the neonatal atrial cells. The physiological differences of I(to) are consistent with the different expression protein levels of Kv4.3 and KChIP2.

Action Potentials↗
PubMed2003

Comparison of biometric measurements using partial coherence interferometry and applanation ultrasound.

PURPOSE: To compare and contrast axial length (AL) measurements assessed by ultrasound biometry and optical biometry. SETTING: Department of Ophthalmology, Johannes Gutenberg-University, Mainz, Germany. METHODS: Optical biometry and ultrasound biometry were performed in 360 eyes to measure AL. RESULTS: In eyes with a clear-appearing crystalline lens, there was a statistically significant median difference in AL measurements between the 2 methods in eyes with a normal or long length. In eyes with cataract, there was a statistically significant median difference in AL measurements in eyes with a normal or short length. In these cases, optical biometry produced larger readings than ultrasound biometry. In eyes with a posterior chamber intraocular lens, there was no statistically significant difference between the 2 methods in eyes of all lengths. In all eyes, the 2 methods produced reliable measurements. CONCLUSIONS: These results show that optical biometry and ultrasound applanation biometry give statistically significant differences in AL measurement in patients with cataract and normal lenses. In these cases, optical biometry provided clinically relevant larger values than ultrasound applanation. Optical biometry provides an alternative to ultrasound biometry. However, ultrasound biometry may be indicated in certain complex cases.

Biometry↗
PubMed2003

Expression of protein phosphatases during postnatal development of rabbit heart.

Protein phosphatases play a major role in the regulation of L-type calcium current (I(Ca)) in heart cells. We previously showed developmental differences in the effects of inhibitors of protein phosphatases (PP's) on the modulation of I(Ca), with greater stimulatory effects on I(Ca) observed in newborn than in adult ventricular cells. We hypothesized that this developmental difference might be due to greater expression and levels of PP 1 and PP 2A in newborn than in adult ventricular cells. We thus determined the mRNA expression of alpha and beta subunits of PP 1 and the a subunit of PP 2A in adult and newborn rabbit ventricles and levels of PP 1 and PP 2A in total homogenates, particulate membranes, and in soluble fraction prepared from isolated ventricular myocytes from adult and newborn rabbits. RT-PCR analysis demonstrated the presence of mRNA of these subunits of PP's in both newborn and adult ventricles. Northern blot analysis using 32P labeled cDNA probes specific for PP 1alpha, PP 1beta and PP 2Aalpha showed that the expression of steady state mRNA levels for PP 1alpha, PP 1beta and PP 2Aalpha were much higher in newborn compared to adult rabbit ventricles. mRNA for glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and for sarcoplasmic reticulum Ca(2+)-ATPase (SERCA) in rabbit ventricles were measured as controls. GAPDH did not show significant developmental changes while mRNA for SERCA was higher in adult compared to newborns. Western blot analysis showed that PP 1 and PP 2A protein levels were also much higher in newborn compared to adult rabbit ventricular cells. Immunoblot analysis in particulate membranes and soluble fraction showed that PP1 was mainly membrane bound while PP 2 was present only in soluble fraction. These findings suggest that the two major protein phosphatases (PP 1 and PP 2A) in heart are expressed at much higher levels in newborn and decline to lower levels in adult ventricular myocytes. The presence of high levels of PP's and particularly PP 1 in newborn cells may be responsible for the greater dependence of newborn cells on the inhibition of PP as a mechanism of action of beta-agonist isoproterenol on I(Ca).

Aging↗
PubMed2003

Simultaneous detection of the exon 10 polymorphism and a novel intronic single base insertion polymorphism in the XPD gene using single strand conformation polymorphism.

We developed a new method based on the single strand conformation polymorphism (SSCP) technique for the detection of a G23591A (Asp312Asn) polymorphism in exon 10 of the XPD gene. In the process we also identified a novel polymorphism 23623C-ins (IVS10+17C-ins) in intron 10 of the same gene. With this newly developed SSCP-based method of genotyping we could detect both polymorphisms in the same assay and thus consequently determine the haplotype. In order to determine the population frequency of the novel polymorphism and the haplotype frequency, 302 healthy individuals were genotyped. The allelic frequency of the 23623C-ins intronic polymorphism was 0.16, whereas the frequency of the variant allele for the G23591A polymorphism was 0.39. Forty-three individuals (14%) were heterozygous for both polymorphisms but none carried polymorphic variants for both G23591A and 23623C-ins on the same allele. The effect of the novel intronic insertion polymorphism, which is located 16 nt downstream of the 3'-end of exon 10 of the XPD gene and involves a mononucleotide C repeat sequence, on expression remains to be determined.

Adult↗
PubMed2003

White Star technology.

White Star micropulse technology is a software modification that allows extremely short bursts of ultrasound energy. Studies have shown that this decreases wound heat build-up with the retained efficiency of continuous ultrasound. Decreased energy utilization with improved corneal function and improved nuclear fragment followability appear to be additional benefits.

Hot Temperature↗
PubMed2003

Secreted frizzled-related protein 4 is a potent tumor-derived phosphaturic agent.

Tumors associated with osteomalacia elaborate the novel factor(s), phosphatonin(s), which causes phosphaturia and hypophosphatemia by cAMP-independent pathways. We show that secreted frizzled-related protein-4 (sFRP-4), a protein highly expressed in such tumors, is a circulating phosphaturic factor that antagonizes renal Wnt-signaling. In cultured opossum renal epithelial cells, sFRP-4 specifically inhibited sodium-dependent phosphate transport. Infusions of sFRP-4 in normal rats over 2 hours specifically increased renal fractional excretion of inorganic phosphate (FEPi) from 14% +/- 2% to 34% +/- 5% (mean +/- SEM, P < 0.01). Urinary cAMP and calcium excretion were unchanged. In thyro-parathyroidectomized rats, sFRP-4 increased FEPi from 0.7% +/- 0.2% to 3.8% +/- 1.2% (P < 0.05), demonstrating that sFRP-4 inhibits renal inorganic phosphate reabsorption by PTH-independent mechanisms. Administration of sFRP-4 to intact rats over 8 hours increased FEPi, decreased serum phosphate (1.95 +/- 0.1 to 1.53 +/- 0.09 mmol/l, P < 0.05) but did not alter serum 1alpha, 25-dihydroxyvitamin D, renal 25-hydroxyvitamin D 1alpha-hydroxylase cytochrome P450, and sodium-phosphate cotransporter mRNA concentrations. Infusion of sFRP-4 antagonizes Wnt action as demonstrated by reduced renal beta-catenin and increased phosphorylated beta-catenin concentrations. The sFRP-4 is detectable in normal human serum and in the serum of a patient with tumor-induced osteomalacia. Thus, sFRP-4 displays phosphatonin-like properties, because it is a circulating protein that promotes phosphaturia and hypophosphatemia and blunts compensatory increases in 1alpha, 25-dihydroxyvitamin D.

25-Hydroxyvitamin D3 1-alpha-Hydroxylase↗
PubMed2003

Fibroblast growth factor 23 concentrations in humoral hypercalcemia of malignancy and hyperparathyroidism.

OBJECTIVE: To determine whether fibroblast growth factor 23 (FGF 23) concentrations are increased in the serum of patients with humoral hypercalcemia of malignancy and in those with hyperparathyroidism. PATIENTS AND METHODS: Serum samples from patients with hypercalcemia and documented elevations of parathyroid hormone-related protein or parathyroid hormone levels were examined for FGF 23 concentrations. Specimens from healthy age-matched controls were evaluated to establish a reference range for FGF 23. RESULTS: Mean +/- SEM concentrations of FGF 23 were elevated in the 7 patients with humoral hypercalcemia of malignancy (385 +/- 134 relative unit [RU]/mL) compared with 11 healthy controls (43.9 +/- 5.8 RU/mL; P = .005). In the 11 patients with hyperparathyroidism, FGF 23 concentrations were increased (mean +/- SEM 86.6 +/- 228 RU/mL), but the increases were not statistically significant. The increases in FGF 23 in these disorders did not correlate with changes in serum phosphate or 1alpha,25-dihydroxyvitamin D levels. CONCLUSION: Levels of FGF 23 are elevated in patients with tumors associated with humoral hypercalcemia of malignancy. The precise cause of such elevations requires further investigation.

Case-Control Studies↗
PubMed2003

Detecting homozygous deletions in the CDKN2A(p16(INK4a))/ARF(p14(ARF)) gene in urinary bladder cancer using real-time quantitative PCR.

PURPOSE: 9p21 is a major target in the pathogenesis of human urinary bladder cancer. The locus harbors the CDKN2A/ARF tumor suppressor gene, which encodes two cell cycle regulatory proteins cyclin dependent kinase 2A (p16(INK4a)) and alternate reading frame (p14(ARF)). We have designed a real-time quantitative PCR (QPCR) application to study homozygous deletion (HD) of CDKN2A/ARF in 186 urinary bladder cancer patients. EXPERIMENTAL DESIGN: Real-time QPCR, based on simultaneous amplification of ARF and a reference gene, GAPDH, was developed and evaluated in three melanoma cell lines with HDs at the CDKN2A/ARF locus (IGR-1, SK-MEL-5, and WM-266-4). In addition, loss of heterozygosity was analyzed at the D9S942, D9S1748, and D12S99 markers. Mutation analysis of the CDKN2A/ARF gene was performed using single-strand conformational polymorphism and sequencing. Results from the present investigation were combined with previous p53 analysis of the same urinary bladder neoplasms. RESULTS: Real-time QPCR analysis showed 26 (14%) HDs, 22 (12%) hemizygous deletions, and 3 (2%) multiple duplications. Loss of heterozygosity was determined in 30 (22%) cases at the D9S942 locus, which is located between E1alpha and E1beta of the CDKN2A/ARF gene. No association was established between occurrence of genetic aberrations at 9p21 and tumor stage or grade, supporting previous suggestions that CDKN2A/ARF inactivation is an early event in bladder carcinogenesis. CONCLUSIONS: We have established a fast and efficient method for detection of HDs. Our data support the notion that inactivation, including HDs, of CDKN2A/ARF is an early event in transitional cell carcinoma. We observed separate and specific targeting of the CDKN2A and ARF genes, respectively, and that simultaneous inactivation of ARF and p53 occurs.

Carcinoma, Transitional Cell↗
PubMed2002

Enzyme mediated extracellular synthesis of CdS nanoparticles by the fungus, Fusarium oxysporum.

The biosynthesis of Q-state CdS nanoparticles by reaction of aqueous CdSO4 solution with the fungus, Fusarium oxysporum, is demonstrated. Nanoparticle formation proceeds by release of sulfate reductase enzymes by the fungus, conversion of sulfate ions to sulfide ions that subsequently react with aqueous Cd2+ ions to yield highly stable CdS nanoparticles. Elucidation of an enzymatic pathway using fungi opens up the exciting possibility of developing a rational, biosynthesis strategy for nanomaterials over a range of chemical compositions which is currently not possible.

Cadmium↗
PubMed2002

Distinctive dendritic cell modulation by vitamin D(3) and glucocorticoid pathways.

Dendritic cell (DC) maturation plays a central role in regulating immunity. We show that glucocorticoid and 1alpha,25(OH)(2)D(3) agonists modulate DCs via distinct and additive signaling pathways. Phenotypic and functional indices were examined in DCs treated with dexamethasone (DEX) and/or a 1alpha,25(OH)(2)D(3) analog (D(3) analog). DEX potently attenuated pro-inflammatory cytokines and chemokines but had modest, reversible effects on T-cell stimulatory capacity. D(3) analog produced significantly greater inhibition of T-cell stimulation in vitro and in vivo and, unlike DEX, increased expression of the chemokines MCP-1 and MIP-1alpha. Both DEX and D(3) analog were associated with reduced expression of the NF-kappaB proteins c-Rel and Rel B but not Rel A. Combined DEX and D(3) analog treatment of DCs resulted in significant additive inhibition of pro-inflammatory cytokines, T-cell stimulation, chemokines, chemokine receptors, and NF-kappaB components. Additive inhibition was most striking for RANTES, CCR5, CCR7, and Rel B. The combined effects of the two hormonal pathways on DCs have unique immunomodulatory potential.

Animals↗
PubMed2002

Characterization of a novel hexameric repeat DNA sequence in the promoter of the immediate early gene, IEX-1, that mediates 1alpha,25-dihydroxyvitamin D(3)-associated IEX-1 gene repression.

1alpha,25-Dihydroxyvitamin D(3)(1alpha,25(OH)(2)D(3)), the active metabolite of vitamin D(3), mediates anti-proliferative effects in cells by regulating the expression of 1alpha,25(OH)(2)D(3)-responsive genes. The expression of the proliferation-promoting Immediate Early gene X-1 (IEX-1) is reduced by 1alpha,25(OH)(2)D(3) through unknown mechanisms. Here we report the presence of a novel inhibitory hexameric repeat DNA response element in the promoter of the human IEX-1 gene that mediates 1alpha,25(OH)(2)D(3)-associated IEX-1 gene repression. To localize a vitamin D sensitive DNA response element we transfected the keratinocyte-like cell line, HaCaT, (referred as HaCaT) with a series of plasmids containing full-length and truncated IEX-1 promoter elements fused to the luciferase reporter gene in the absence or presence of 1alpha,25(OH)(2)D(3), and we performed electrophoretic gel mobility assays in the presence of receptors for 1alpha,25(OH)(2)D(3) (vitamin D receptor, VDR) and 9-cis-retinoic acid (RXRalpha). We mapped a negative response element between nt -405 and -391(15 bp) of theIEX-1 promoter (5'-TGAACC AGG GAGTCA-3') that mediates transcriptional inhibition in response to 1alpha,25(OH)(2)D(3) and which requires expression of both nuclear receptors for 1alpha,25(OH)(2)D(3) and 9-cis-retinoic acid. Our data indicate that the physiological repression of IEX-1 gene expression by 1alpha,25(OH)(2)D(3) is directly mediated by nuclear VDR/RXRalpha heterodimers through a specific transcriptional element.

Apoptosis Regulatory Proteins↗
PubMed2002

Divergent regulation of the growth-promoting gene IEX-1 by the p53 tumor suppressor and Sp1.

IEX-1, a recently discovered early response gene, regulates cell growth and apoptosis. IEX-1 gene expression is regulated by a variety of factors such as x-irradiation, ultraviolet radiation, steroids, growth factors, and inflammatory stimuli. By systematic examination of the IEX-1 promoter, we show that IEX-1 gene expression is controlled by multiple conserved gene regulatory elements and that IEX-1 is a downstream target of the p53 tumor suppressor and Sp1. In addition, p300, Sox, nuclear factor-kappaB, and AP4 appear to be modulators of IEX-1 gene expression to a lesser degree. We found that there is at least one Sp1 element that functions as an activator and contributes to high basal transcriptional levels of the IEX-1 gene. We demonstrate the presence of a p53 response element that represses IEX-1 promoter activity in HaCaT keratinocytes, indicating that Sp1 and p53 have opposite effects on IEX-1 gene expression. We conclude that IEX-1 expression in cells is regulated by the p53 tumor suppressor and Sp1, thus providing a direct mechanism for control of cell proliferation.

Apoptosis Regulatory Proteins↗
PubMed2002

1 alpha,25-dihydroxyvitamin D3 increases TGF beta 1 binding to human osteoblasts.

1alpha,25-Dihydroxyvitamin D3 (1alpha,25(OH)2D3) increases the binding of transforming growth factor beta1(TGF beta1) via TGF beta receptors to the surface of human osteoblasts. The increase in TGF beta receptors induced by 1alpha,25(OH)2D3 is dependent on increases in TGF beta secretion induced by 1alpha,25(OH)2D3, since antibodies directed against TGF beta block the increase in TGF beta1 binding. The increase in TGF beta type I and II receptors on cell surfaces following 1alpha,25(OH)2D3 treatment is associated with increases in receptor mRNA concentrations. Increases in receptor mRNA concentrations following 1alpha,25(OH)2D3 treatment are not due to changes in receptor gene transcription. The role of TGF beta receptors, in mediating the growth responses to 1alpha,25(OH)2D3 is demonstrated by showing that osteoblasts which express dominant negative, kinase-deficient TGF beta type II receptors, fail to respond to the growth-inhibitory effects of 1alpha,25(OH)2D3. An increase in TGF beta receptor expression is important in mediating 1alpha,25(OH)2D3-associated changes in the growth rate of osteoblasts.

Activin Receptors, Type I↗
PubMed2002

The metal-binding properties of DREAM: evidence for calcium-mediated changes in DREAM structure.

DREAM, an EF-hand protein, associates with and modulates the activity of presenilins and Kv4 potassium channels in neural and cardiac tissues and represses prodynorphin and c-fos gene expression by binding to DNA response elements in these genes. Information concerning the metal-binding properties of DREAM and the consequences of metal binding on protein structure are important in understanding how this protein functions in cells. We now show that DREAM binds 1 mol of calcium/mol of protein with relatively high affinity and another 3 mol of calcium with lower affinity. DREAM binds 1 mol of magnesium/mol of protein. DREAM, pre-loaded with 1 mol of calcium, binds 1 mol of magnesium, thus demonstrating that the magnesium-binding site is distinct from the high affinity calcium-binding site. Analysis of metal binding to mutant DREAM protein constructs localizes the high affinity calcium-binding site and the magnesium-binding site to EF-hands 3 or 4. Binding of calcium but not magnesium changes the conformation, stability, and alpha-helical content of DREAM. Calcium, but not magnesium, reduces the affinity of apo-DREAM for specific DNA response elements in the prodynorphin and c-fos genes. We conclude that DREAM binds calcium and magnesium and that calcium, but not magnesium, modulates DREAM structure and function.

Amino Acid Sequence↗