Ram Sharma - Academia.edu (original) (raw)

Papers by Ram Sharma

Journal of the American Heart Association, Jan 9, 2017

Atrial fibrillation (AF) is the most common cardiac dysrhythmia associated with significant morbi... more Atrial fibrillation (AF) is the most common cardiac dysrhythmia associated with significant morbidity and mortality. Several small studies have reported that low serum total testosterone (TT) levels were associated with a higher incidence of AF. In contrast, it is also reported that anabolic steroid use is associated with an increase in the risk of AF. To date, no study has explored the effect of testosterone normalization on new incidence of AF after testosterone replacement therapy (TRT) in patients with low testosterone. Using data from the Veterans Administrations Corporate Data Warehouse, we identified a national cohort of 76 639 veterans with low TT levels and divided them into 3 groups. Group 1 had TRT resulting in normalization of TT levels (normalized TRT), group 2 had TRT without normalization of TT levels (nonnormalized TRT), and group 3 did not receive TRT (no TRT). Propensity score-weighted stabilized inverse probability of treatment weighting Cox proportional hazard me...

SpringerPlus, 2016

Focal segmental glomerulosclerosis (FSGS) is a major cause of renal failure. Sera of some FSGS pa... more Focal segmental glomerulosclerosis (FSGS) is a major cause of renal failure. Sera of some FSGS patients increase glomerular albumin permeability (Palb) during in vitro testing and cause proteinuria in experimental animals. To determine whether permeability activity of FSGS serum (Palb activity) is associated with rate of progression to renal replacement therapy (RRT). This is an observational study based on medical and demographic information and Palb activity testing. Studies were performed at Medical College of Wisconsin. Serum was submitted by patients' nephrologists for measurement of Palb activity. Each patient had had a biopsy diagnosis of FSGS, had reached ESRD and was on dialysis or had a functioning transplant. Palb activity, clinical characteristics and time between biopsy diagnosis and RRT (T-RRT) were recorded for each patient. Palb activity was measured using established in vitro techniques. Palb and T-RRT were inversely correlated. Neither Palb nor T-RRT varied wit...

American Journal of Physiology-Renal Physiology, 2009

Glomerular capillary filtration barrier characteristics are determined in part by the slit-pore j... more Glomerular capillary filtration barrier characteristics are determined in part by the slit-pore junctions of glomerular podocytes. Protein tyrosine phosphatase receptor-O (PTPro) is a transmembrane protein expressed on the apical surface of podocyte foot processes. Tyrosine phosphorylation of podocyte proteins including nephrin may control the filtration barrier. To determine whether PTPro activity is required to maintain glomerular macromolecular permeability, albumin permeability ( Palb) was studied after incubation of glomeruli from normal animals with a series of monoclonal (mAb) and polyclonal antibodies. Reagents included mAbs to rabbit and rat PTPro and polyclonal rabbit immune IgG to rat PTPro. mAb 4C3, specific to the amino acid core of PTPro, decreased its phosphatase activity and increased Palbof rabbit glomeruli in a time- and concentration-dependent manner. In contrast, mAb P8E7 did not diminish phosphatase activity and did not alter Palb. Preincubation of 4C3 with PTPr...

Nephrology Dialysis Transplantation, 2017

Congenital anomalies of the kidney and urinary tract (CAKUT) including solitary kidney constitute... more Congenital anomalies of the kidney and urinary tract (CAKUT) including solitary kidney constitute the main cause of progressive chronic kidney disease (CKD) in children. Children born with CAKUT develop signs of CKD only during adolescence and do not respond to renin-angiotensin-aldosterone system blockers. Early cellular changes underlying CKD progression to end-stage renal disease by early adulthood are not well understood. The mechanism of maladaptive hyperfiltration that occurs from loss of functional nephrons, including solitary kidney, is not clear. We reexamine the phenomenon of hyperfiltration in the context of biomechanical forces with special reference to glomerular podocytes. Capillary stretch exerts tensile stress on podocytes through the glomerular basement membrane. The flow of ultrafiltrate over the cell surface directly causes fluid flow shear stress (FFSS) on podocytes. FFSS on the podocyte surface increases 1.5-to 2-fold in animal models of solitary kidney and its effect on podocytes is a subject of ongoing research. Podocytes (i) are mechanosensitive to tensile and shear forces, (ii) use prostaglandin E 2 , angiotensin-II or nitric oxide for mechanoperception and (iii) use specific signaling pathways for mechanotransduction. We discuss (i) the nature of and differences in cellular responses to biomechanical forces, (ii) methods to study biomechanical forces and (iii) effects of biomechanical forces on podocytes and glomeruli. Future studies on FFSS will likely identify novel targets for strategies for early intervention to complement and strengthen the current regimen for treating children with CAKUT.

BioMed Research International, 2017

A plasma component is responsible for altered glomerular permeability in patients with focal segm... more A plasma component is responsible for altered glomerular permeability in patients with focal segmental glomerulosclerosis. Evidence includes recurrence after renal transplantation, remission after plasmapheresis, proteinuria in infants of affected mothers, transfer of proteinuria to experimental animals, and impaired glomerular permeability after exposure to patient plasma. Therapy may include decreasing synthesis of the injurious agent, removing or blocking its interaction with cells, or blocking signaling or enhancing cell defenses to restore the permeability barrier and prevent progression. Agents that may prevent the synthesis of the permeability factor include cytotoxic agents or aggressive chemotherapy. Extracorporeal therapies include plasmapheresis, immunoadsorption with protein A or anti-immunoglobulin, or lipopheresis. Oral or intravenous galactose also decreasesPalbactivity. Studies of glomeruli have shown that several strategies prevent the action of FSGS sera. These inc...

American Journal of Physiology-Renal Physiology, 2017

Recently, we and others have found that hyperfiltration-associated increase in biomechanical forc... more Recently, we and others have found that hyperfiltration-associated increase in biomechanical forces, namely, tensile stress and fluid flow shear stress (FFSS), can directly and distinctly alter podocyte structure and function. The ultrafiltrate flow over the major processes and cell body generates FFSS to podocytes. Our previous work suggests that the cyclooxygenase-2 (COX-2)-PGE2-PGE2receptor 2 (EP2) axis plays an important role in mechanoperception of FFSS in podocytes. To address mechanotransduction of the perceived stimulus through EP2, cultured podocytes were exposed to FFSS (2 dyn/cm2) for 2 h. Total RNA from cells at the end of FFSS treatment, 2-h post-FFSS, and 24-h post-FFSS was used for whole exon array analysis. Differentially regulated genes ( P < 0.01) were analyzed using bioinformatics tools Enrichr and Ingenuity Pathway Analysis to predict pathways/molecules. Candidate pathways were validated using Western blot analysis and then further confirmed to be resulting fr...

Journal of investigative medicine : the official publication of the American Federation for Clinical Research, 1999

Primary focal segmental glomerulosclerosis (FSGS) recurs in nearly 30% of patients who progress t... more Primary focal segmental glomerulosclerosis (FSGS) recurs in nearly 30% of patients who progress to end-stage renal disease and then receive a kidney transplant. A circulating plasma factor has been isolated from these patients that increases glomerular permeability to albumin in vitro. Because of the pivotal role of the mesangial cell in the accumulation of extracellular matrix (ECM) material within the glomerulus and the modulation of matrix protein synthesis by nitric oxide (NO), we examined the effect of the FSGS factor on inducible nitric oxide synthase (iNOS) expression and NO production by cultured rat mesangial cells (RMC). RMC were incubated with the supernatant following 70% ammonium sulfate precipitation of serum from patients with recurrent FSGS. Addition of the FSGS factor to cultured RMC led to a significant inhibition of nitrite accumulation, an index of NO synthesis. There was a parallel decline in iNOS gene and protein expression. Sera obtained from control patients ...

Transplantation, 2002

Recurrent focal segmental glomerulosclerosis (FSGS) is heralded by proteinuria that may remit aft... more Recurrent focal segmental glomerulosclerosis (FSGS) is heralded by proteinuria that may remit after treatment with plasmapheresis or immunoadsorption. Study of recurrent FSGS has been hampered by lack of an animal model that exhibits a pattern of proteinuria that mimics human disease. We have obtained a component of FSGS patient plasma (FSGS factor) that increases glomerular albumin permeability (P(alb)) in vitro and causes transient proteinuria in vivo. Plasma fractions containing FSGS factor and comparable plasma fractions from normal donors were injected into normal male Sprague-Dawley rats. Urinary protein, albumin, and creatinine were measured at various time points. Additionally, plasma samples from test animals were collected after injection and tested for FS activity defined by increased P(alb). Finally, glomeruli were isolated from animals after injection and P(alb) of these glomeruli tested. Proteinuria and albuminuria were increased by 24 hr after injection with FSGS factor, and returned to baseline by 48 hr after injection. Injection with the same fraction of normal plasma had no effect on urinary protein. FSGS factor increased urinary protein in a dose-dependent manner. Serum collected from rats 15 or 60 min after injection with FSGS factor increased P(alb) of glomeruli in vitro, whereas serum collected 3 or more hours after injection had no effect. Glomeruli isolated from rats receiving injections with FSGS factor had increased in vitro P(alb) compared with glomeruli from rats injected with a fraction from normal plasma. We have demonstrated that a single injection of FSGS factor increases P(alb) and, causes transient albuminuria and proteinuria in rats. FS activity in the plasma of recipient rats is also transient. This is the first detailed description of the time course and dose-dependence of proteinuria caused by FSGS factor in an animal model.

The Journal of laboratory and clinical medicine, 1995

Sodium nitroprusside (SNP) causes renal vasodilation, increased renal blood flow, and increased r... more Sodium nitroprusside (SNP) causes renal vasodilation, increased renal blood flow, and increased renal sodium excretion. Systemic vascular effects of SNP are dependent on generation of nitric oxide and increase in smooth muscle intracellular cyclic guanosine monophosphate (cGMP). In the current studies isolated glomeruli were used to determine the effects of SNP on glomerular capillary hydraulic conductivity (Lp) and on mesangial tone. The direct effects of the cGMP analogue 8-bromo-cGMP on Lp were studied. Glomeruli were isolated from the superficial renal cortex of adult male Sprague-Dawley rats weighing 143 to 383 gm. Lp was calculated from videomicroscopic images of individual glomeruli during the initial 0.1 second of filtration induced by an albumin oncotic gradient. Incubation of glomeruli in 10(-3) mol/L SNP increased Lp from a control value of 1.38 +/- 0.08 microliter/min.mm Hg.cm2 to 1.91 +/- 0.13 microliter.min.mm Hg.cm2 (p < 0.01). Incubation of glomeruli in 10(-5) mol...

AJP: Renal Physiology, 2009

Chronic kidney disease (CKD) is associated with decreased renal nitric oxide (NO) production and ... more Chronic kidney disease (CKD) is associated with decreased renal nitric oxide (NO) production and increased plasma levels of methylarginines. The naturally occurring guanidino-methylated arginines N-monomethyl-l-arginine (l-NMMA) and asymmetric dimethyl-l-arginine (ADMA) inhibit NO synthase activity. We hypothesized that ADMA and l-NMMA compromise the integrity of the glomerular filtration barrier via NO depletion. We studied the effect of ADMA on albumin permeability (Palb) in isolated glomeruli and examined whether this effect involves NO- and superoxide (O2•−)-dependent mechanisms. ADMA at concentrations found in circulation of patients with CKD decreased cGMP and increased Palb in a dose-dependent manner. A similar increase in Palb was caused by l-NMMA but at a concentration two orders of magnitude higher than that of ADMA. NO donor DETA-NONOate or cGMP analog abrogated the effect of ADMA on Palb. The SOD mimetic tempol or the NAD(P)H oxidase inhibitor apocynin also prevented the...

Journal of Cell Communication and Signaling, 2012

Minimal change disease (MCD), the most common idiopathic nephrotic syndrome in children, is chara... more Minimal change disease (MCD), the most common idiopathic nephrotic syndrome in children, is characterized by proteinuria and loss of glomerular visceral epithelial cell (podocyte) ultrastructure. Lipopolysaccharide (LPS) and puromycin aminonucleoside (PAN) are used to study podocyte injury in models of MCD in vivo and in vitro. We hypothesized that LPS and PAN influence components of the innate immune system in podocytes such as the Toll-Like Receptor (TLRs), TLR adapter molecules, and associated cytokines. Our results show that cultured human podocytes constitutively express TLRs 1-6 and TLR-10, but not TLRs 7-9. LPS (25 μg/ml) or PAN (60 μg/ml) caused comparable derangement of the actin cytoskeleton in podocytes. Quantitative RT-PCR analysis show that LPS differentially up-regulated the expression of genes for TLRs (1>4≥2>3>6>5), the adapter molecule, MyD88, and transcription factor NF-κB within one hour. LPS also caused increased levels of IL-6, IL-8 and MCP1 without exerting any effect on TNF-α, IFN-α or TGF-β1 at 24 h. Immunofluorescence intensity analysis of confocal microscopy images showed that LPS induced a significant increase in nuclear translocation of NF-κB by 6 h. In contrast, PAN-induced only small changes in the expression of TLRs 2-6 that included a persistent increase in TLRs 2 and 5, a transient increase in TLR-4, and a gradual increase in TLRs 3 and 6 between 1 and 6 h. Correspondingly, it did not alter pro-inflammatory cytokine levels in podocytes. However, PAN induced a low but significant increase in NF-κB nuclear translocation within one hour that remained unchanged up to 6 h. In summary, these novel findings show that LPS, a known TLR-4 ligand, induced the gene expression of multiple TLRs with maximum effect on the expression of TLR-1 suggesting a loss of receptor selectivity and induction of receptor interactions in podocytes. A comparable derangement of the podocyte cytoskeleton and significant increase in the nuclear translocation of NF-κB by PAN suggest that disparate but complementary mechanisms may contribute to the development of podocytopathy in MCD.

American Journal of Physiology-Renal Physiology, 1998

Angiotensin II decreases glomerular filtration rate, renal plasma flow, and glomerular capillary ... more Angiotensin II decreases glomerular filtration rate, renal plasma flow, and glomerular capillary hydraulic conductivity. Although angiotensin II receptors have been demonstrated in mesangial cells and proximal tubule cells, the presence of angiotensin II receptors in glomerular epithelial cells has not previously been shown. Previously, we have reported that angiotensin II caused an accumulation of cAMP and a reorganization of the actin cytoskeleton in cultured glomerular epithelial cells. Current studies were conducted to verify the presence of angiotensin II receptors by immunological and non-peptide receptor ligand binding techniques and to ascertain the activation of intracellular signal transduction in glomerular epithelial cells in response to angiotensin II. Confluent monolayer cultures of glomerular epithelial cells were incubated with angiotensin II, with or without losartan and/or PD-123,319 in the medium. Membrane vesicle preparations were obtained by homogenization of wa...

Transplantation, Jan 29, 2018

Kidney donors face a small but definite risk of end-stage renal disease 15-30 years postdonation.... more Kidney donors face a small but definite risk of end-stage renal disease 15-30 years postdonation. The development of proteinuria, hypertension with gradual decrease in kidney function in the donor after surgical resection of 1 kidney has been attributed to hyperfiltration. Genetic variations, physiological adaptations, and co-morbidities exacerbate the hyperfiltration-induced loss of kidney function in the years following donation. A focus on glomerular hemodynamics and capillary pressure has led to the development of drugs that target the renin-angiotensin-aldosterone system (RAAS), but these agents yield mixed results in transplant recipients and donors. Recent work on glomerular biomechanical forces highlights the differential effects of tensile stress and fluid flow shear stress (FFSS) from hyperfiltration. Capillary wall stretch due to glomerular capillary pressure increases tensile stress on podocyte foot processes that cover the capillary. In parallel, increased flow of the u...

Journal of Laboratory and Clinical Medicine, 1996

Matrix metalloproteinases (MMPs) secreted by connective tissue cells are capable of acting on ext... more Matrix metalloproteinases (MMPs) secreted by connective tissue cells are capable of acting on extracellular matrix components of glomerular basement membrane at a slow rate and thus may play a role in the control of protein permeability and in the progression of certain kinds of glomerulonephritis. We have used an in vitro assay to measure the direct effect of three MMPs and human neutrophil elastase on glomerular albumin permeability (Palbumin). Glomeruli were isolated from normal male Sprague-Dawley rats and suspended in isolation medium with or without interstitial collagenase, gelatinase-A, stromelysin-1, or elastase and were incubated at 37 degrees C for up to 4 hours. A tissue-specific inhibitor of matrix metalloproteinases (TIMP-1) and a plasma proteinase inhibitor, alpha2-macroglobulin (alpha2M), were used to block the activity of MMPs. Palbumin was calculated from the change in glomerular volume in response to an applied oncotic gradient. In this study stromelysin-1 (10 microg/ml) and elastase (5 microg/ml) increased Palbumin significantly. Stromelysin-1 increased Palbumin after 4 hours, whereas elastase had an effect after 2 hours. Lower concentrations of stromelysin-1 or shorter incubation time had no effect on Palbumin. Incubation for up to 4 hours with interstitial collagenase (10 microg/ml) or gelatinase-A (10 microg/ml) had no effect on Palbumin. Coincubation with TIMP-1 and alpha2M blocked the stromelysin-1-mediated increase in Palbumin. We conclude that stromelysin-1 is capable of affecting the glomerular filtration barrier directly and that it may play an important role in causing proteinuria in glomerular diseases.

Journal of Laboratory and Clinical Medicine, 2001

lomerular epithelial cells are highly specialized, terminally differentiated cells with interdigi... more lomerular epithelial cells are highly specialized, terminally differentiated cells with interdigitating foot processes. They are in an ideal position to respond to mechanical, endocrine, and paracrine signals through changes in the slit-junctions. 1 GECs are responsible for providing the majority of the resistance (90%) to permeability of water and small molecules. They provide a significant resistance (50%) to permeability of albumin and macromolecules. Calculations with computational fluid dynamics also support the idea that GECs control glomerular capillary Both subtype 1 and 2 receptors of angiotensin II participate in regulation of intracellular calcium in glomerular epithelial cells

Kidney International, 2000

Kidney International, 1994

American Journal of Kidney Diseases, 1998