Invasive aspergillosis in a renal transplant recipient successfully treated with interferon-gamma.

Invasive aspergillosis is a serious complication of solid organ transplantation. An early diagnosis is hampered by the lack of reliable serum markers and, even if appropriately diagnosed and treated with current antifungal agents, has a high mortality rate. We report a case of invasive pulmonary and cerebral aspergillosis in a renal transplant patient treated with IFN-γ in conjunction with combination anti-fungal therapy for six weeks in whom complete resolution of the fungal infection was achieved. Renal function remained intact throughout the treatment period. Surveillance CT scans of the chest and head showed resolution of prior disease but revealed a new left upper lobe mass four months after completion of treatment with IFN-γ. Biopsy of the lesion was positive for primary lung adenocarcinoma, for which she underwent left upper lobe resection. The pathology report confirmed clear surgical margins and lymph nodes and no evidence of fungal hyphae. IFN-γ should be considered early in the management of invasive aspergillosis in renal transplant patients. To date, allograft rejection has not been encountered.

The use of drug-eluting stents in the management of transplant renal artery stenosis.

Transplant renal artery stenosis (TRAS) is a common occurrence following kidney transplantation with an incidence rate ranging from 6% to 23%. A single-center retrospective study was conducted to examine the use of drug-eluting stents (DES) in eligible patients with hemodynamically significant TRAS. Between March 2008 and January 2011, 12 patients were diagnosed with TRAS with reference vessel diameter measuring <5 mm and underwent endovascular intervention (EVI) with DES placement. TRAS was detected within the first year posttransplantation in a majority of these patients (83%) and manifested as hypertension (100%), allograft dysfunction (100%) and edema (58%). Procedural success rate was 100%. Patients were followed for a mean period of 16 ± 10 months. Blood pressure improved from a mean of 156/82 to 138/73 mmHg at the end of the follow-up period. In 11/12 patients, serum creatinine improved from 3.1 ± 1.3 mg/dL to 2.3 ± 0.5 mg/dL at the end of the follow-up period. TRAS of early onset is readily amenable to EVI with stent placement resulting in improvement in blood pressure control and allograft function.

Uterine leiomyoma causing urinary obstruction of the transplanted kidney.

Obstruction of the ureter as a cause of acute or chronic kidney injury in the transplanted kidney is unusual beyond the perioperative period. We present a case of ureteric obstruction, infection and septicemia caused by a large uterine leiomyoma in a patient 8 years post transplantation. Initial treatment comprised of intravenous fluid and antibiotics followed by urgent drainage of the collecting system. Subsequent hysterectomy resolved the obstruction with resolution of renal failure. In young female kidney transplant recipients, gynecologic causes, although rare, need to be considered as possible etiologies of urinary obstruction and renal dysfunction.

Sirolimus-associated diffuse alveolar hemorrhage in a renal transplant recipient on long-term anticoagulation.

Sirolimus (rapamycin, rapamune) is an effective immunosuppressant that has been widely used in solid organ transplantation. Recently, two disconcerting side effects, namely pulmonary toxicity, usually in the form of interstitial pneumonitis, and the onset of nephrotic range proteinuria, have been recognized. We report the case of a renal transplant recipient who had been on chronic anticoagulation therapy for a mechanical aortic valve, and who developed pulmonary distress necessitating emergent intubation 18 days after starting sirolimus therapy. Open lung biopsy showed diffuse alveolar hemorrhage with fibrin deposits in the alveolar spaces and small bronchi. Urine protein/creatinine ratio at that time was 16.7. Upon discontinuation of sirolimus, alveolar hemorrhage and nephrotic range proteinuria resolved. We suggest that extra vigilance be paid in individuals who are on chronic anticoagulation and who are started on sirolimus.

Delayed renal allograft dysfunction and cystitis associated with human polyomavirus (BK) infection in a renal transplant recipient: a case report and review of literature.

Human polyomavirus type BK (BKV) associated nephritis (BKVAN) has recently emerged as an important cause of renal allograft dysfunction and failure. Early recognition of this entity as a cause of allograft dysfunction is extremely important since misdiagnosis can accelerate graft loss. We report a case of BKVAN that presented with symptoms related to cystitis, and review the risk factors, the diagnostic tools and the approach to treatment of BK virus associated allograft nephropathy.

Recurrent severe anion gap metabolic acidosis secondary to episodic ethylene glycol intoxication.

Acute ethylene glycol toxicity and its attendant metabolic derangement is a well described clinical entity. Recurrent severe anion gap metabolic acidosis consequent to episodic ingestion of ethylene glycol has not been previously reported. We present a patient who developed severe anion gap metabolic acidosis with no osmolar gap and hypokalemia, consequent to episodic ethylene glycol ingestion. Modest artifactual elevation of the serum lactic acid level and rapid response to intravenous bicarbonate infusion may serve as diagnostic clues. Consideration of these aberrant features should be included in the clinical assessment of severe anion gap metabolic acidosis.

Combined medical surgical therapy for pulmonary mucormycosis in a diabetic renal allograft recipient.

Mucormycosis is a rare opportunistic infection that complicates chronic debilitating diseases and immunosuppressed solid-organ transplant recipients. We present a case of life-threatening pulmonary mucormycosis in a diabetic renal allograft recipient who survived with reasonable renal function. Early recognition of this entity and prompt use of bronchoalveolar lavage (BAL) are critical to the outcome. Antifungal therapy combined with early surgical excision of infected, necrotic tissue appears to be the preferred course of action. Judicious withholding of immunosuppressants until fungemia cleared did not jeopardize allograft function.

Epstein-Barr virus infection of renal proximal tubule cells: possible role in chronic interstitial nephritis.

Chronic interstitial nephritis frequently accompanies renal diseases of different etiologies. Far less common is the entity of primary interstitial nephritis wherein the glomerular and vascular structures of the kidney are not the primary focus of the disease process. Using in situ hybridization and the polymerase chain reaction, we detected DNA from the Epstein-Barr Virus (EBV) exclusively in renal tissue of patients with the idiopathic variety of chronic interstitial nephritis. The EBV genome, but not that of cytomegalovirus or adenovirus, was detected primarily in renal proximal tubule cells. Furthermore, the CD21 antigen, which serves as the receptor for EBV in B lymphocytes, was detected by immunocytochemistry primarily on proximal tubule cells and was markedly upregulated in the EBV-infected tissue. Western blot analysis of primary cultures of normal proximal tubule cells identified a 140-kDa protein, confirming the expression of the CD21 antigen. Colocalization experiments using proximal and distal tubule markers confirmed that EBV DNA and the CD21 antigen are found primarily in proximal tubule cells. EBV infection of renal proximal tubular cells may participate in evoking a cellular immune response that results in a damaged renal interstitium.

Pauci-immune rapidly progressive glomerulonephritis after nephrectomy in a renal donor.

Idiopathic rapidly progressive glomerulonephritis (RPGN) is a clinicopathologic syndrome in which glomerular damage is accompanied by a rapid and progressive decline in renal function, usually resulting in irreversible renal failure in weeks or months. We report the occurrence of pauci-immune RPGN, more specifically microscopic polyarteritis nodosa (PAN), in a 60-year-old woman 15 months after donor nephrectomy, and 3 months after documentation of intact, residual renal function. The transplanted kidney continues to function well in the recipient, 6 years posttransplantation, and 4.5 years beyond destruction of the donor's contralateral kidney by RPGN. The donor underwent cadaveric renal transplantation after 2 years on dialysis, and at the 3-year mark has intact renal function. These intriguing observations strongly argue that host environmental factors, rather than intrarenal factors, play a major causative role in the pathogenesis of RPGN.

Signalling pathways activated by endothelin stimulation of renal cells.

1. Endothelin mediates its effects in a variety of renal cells via a multiplicity of intracellular signalling pathways. 2. Stimulation of phosphatidylinositol-specific phospholipase C (PI-PLC), resulting in the activation of inositol trisphosphate and diacylglycerol, can be detected even at picomolar concentrations of peptide. 3. Endothelin activation of cPLA2 is sensitive to ambient [Ca2+]i, is not contingent upon protein kinase C activation and is independent of PI-PLC stimulation, being coupled to the endothelin receptor in a yet to be determined manner. 4. Activation by endothelin of phosphatidylcholine-specific phospholipase D is under the dual regulation of protein kinase C and [Ca2+]i, with protein kinase C being the major regulator and [Ca2+]i playing a secondary, modulatory role. 5. Phosphatidylcholine-specific phospholipase C (PC-PLC) is stimulated by endothelin and accounts for the prolonged activation of diacylglycerol by this peptide. PC-PLC activity is critically dependent upon [Ca2+]i, whereas protein kinase C plays no role in modulating the activity of this enzyme. 6. Endothelin enhances the phosphorylation of protein tyrosine kinases, with evidence that phosphorylation of pp60 Src may be an important early event.

Endothelin receptor mRNA expression in renal medulla identified by in situ RT-PCR.

Three subtypes of endothelin (ET) receptors have been identified by cDNA cloning, namely ET-RA, ET-RB, and ET-RC. In the current study the precise cellular distribution of the ET receptor subtypes in the renal medulla was explored by detecting the corresponding polymerase chain reaction (PCR)-amplified cDNAs by in situ reverse transcription (RT)-PCR. The PCR-amplified cDNAs were detected either by direct incorporation using digoxigenin-dUTP (dig-dUTP) as a nucleotide substrate in the PCR reaction or by in situ hybridization with the dig-dUTP-labeled probe. ET-RB mRNA was detected exclusively in the epithelial cells of the inner and outer medullary collecting duct. In contrast, ET-RA message was observed primarily in interstitial cells and pericytes of the vasae rectae in the outer and inner medulla. Southern blot analysis of PCR-amplified cDNAs reverse transcribed from extracted RNA of rat renal medulla confirmed the specificity of the RT-PCR products. ET-RC mRNA was not detected. We conclude that ET-RB is the major ET receptor found in rat renal medulla and is expressed exclusively on inner medullary collecting duct cells. The pattern of ET receptor mRNA expression described suggests different physiological actions for ET on the diverse cellular structures of the renal medulla.

Osmolar regulation of endothelin signaling in rat renal medullary interstitial cells.

We tested the hypothesis that endothelin (ET) responsiveness in the renal medulla is modulated by ambient osmolarity. Cultured renal medullary interstitial cells (RMICs) were incubated from 3 to 24 h in isosmolar culture medium (300 mOsm/kg H2O) or media rendered hyperosmolar (600 mOsm/kg H2O) by the addition of urea. Under hyperosmolar conditions, the peak of ET-evoked Ca2+ transient was blunted by 45-58% (P < 0.02) and PGE2 accumulation decreased from 16- to 2-fold above basal values (P < 0.001). To explore whether hyperosmolar conditions blunt intracellular signaling via modulation of receptor number or expression, kinetics of ET binding and Northern blot analysis of ETA receptor mRNA was performed. Under hyperosmolar conditions, ETA receptor density was reduced by 84% versus isosmolar conditions (238 +/- 12 vs. 1450 +/- 184 fmol/mg) (P < 0.01). In contrast to the ligand binding studies, ETA receptor mRNA was increased by 58% (P < 0.05) in cells grown under hyperosmolar versus isosmolar media. These observations indicate that in the hyperosmolar setting, ET-evoked intracellular signaling is blunted in RMICs due to ET receptor downregulation. Since ETA receptor mRNA is increased under hyperosmolar conditions, we conclude that ET receptor downregulation is the consequence of either decreased translation of message, increased degradation of receptor peptide, or increased internalization of specific receptor sites.

cGMP antagonizes angiotensin-mediated phosphatidylcholine hydrolysis and C kinase activation in mesangial cells.

Previous studies from this laboratory have shown that in cultured rat mesangial cells (MC), angiotensin II (ANG II) mediates its effects via activation of phosphatidylinositol-specific phospholipase C (PI-PLC) and phosphatidylcholine-specific phospholipase C (PC-PLC) and phospholipase D (PC-PLD). In addition, guanosine 3',5'-cyclic monophosphate (cGMP)-elevating maneuvers that stimulate particulate and soluble guanylate cyclase [atrial natriuretic factor (ANF) and sodium nitroprusside (SNP), respectively] antagonize ANG II-mediated PI-PLC activation. The current study explored whether cGMP impairs ANG II-mediated PC-PLC and PLD activity. The ANG II-stimulated release of the water-soluble metabolites of PC breakdown (phosphorylcholine and choline) was blocked by ANF and SNP. ANG II-stimulated phosphatidic acid and phosphatidylethanol formation were significantly reduced by ANF and SNP, confirming that cGMP blunted PLD activity. The inhibitory effect of cGMP on PLD could be reversed by N-(2-[methylamino]ethyl)-5-isoquinolinesulfonamide, a blocker of cGMP-dependent protein kinase. In parallel experiments, ANF and SNP abrogated sustained diacylglycerol (DAG) accumulation derived from ANG II stimulation of PC hydrolysis, confirming that cGMP diminished PC-PLC activity. Inhibition of PC-derived DAG accumulation by cGMP was associated with a concomitant decrement in ANG II-mediated translocation of protein kinase C (PKC) activity from the cytosol to the membrane. In summary, in MC, cGMP antagonizes ANG II-mediated PC hydrolysis, DAG formation, and PKC activation. We propose that cGMP-mediated inhibition of phospholipid metabolism and PKC translocation plays an important role in MC vasorelaxation.

Mechanism of endothelin activation of phospholipase A2 in rat renal medullary interstitial cells.

Previous studies from this laboratory have demonstrated that endothelin-1 (ET) stimulates phosphatidylinositol (PI) hydrolysis, activates dihydropyridine-insensitive Ca2+ channels, and promotes prostaglandin E2 (PGE2) accumulation in cultured rat renal medullary interstitial cells (RMIC). The mechanism whereby ET augments PGE2 production was explored in the current study. ET-evoked PGE2 accumulation proceeded independent of large increments in cytosolic free Ca2+ concentration ([Ca2+]i), derived from either extracellular or intracellular sources. Chelation of intracellular Ca2+ with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid eliminated ET-evoked PGE2 production, indicating that eicosanoid production was nonetheless a Ca(2+)-requiring process. Nanomolar concentrations of phorbol 12-myristate 13-acetate (PMA) alone did not stimulate PGE2 production, nor did PMA alter ET-stimulated PGE2 accumulation. Furthermore, downregulation of protein kinase C (PKC) by prolonged exposure of cells to PMA did not mitigate ET-mediated PGE2 production, demonstrating that PKC stimulation was not required for PGE2 production. ET stimulated PGE2 accumulation despite PI-specific phospholipase C (PI-PLC) inhibition by nanomolar concentrations of PMA, indicating that eicosanoid production was not a downstream event of PI hydrolysis. ET stimulated arachidonic acid metabolite release in parallel with a loss of label from membrane phospholipids. Phosphatidylethanolamine was the preferred substrate for ET-mediated activation of phospholipase A2 (PLA2). Immunocytochemical studies including immunostaining, immunoblotting, and immunoprecipitation confirmed the presence of cytosolic PLA2 (cPLA2) in RMIC. In summary, ET stimulation of PGE2 production in RMIC is mediated via agonist activation of cPLA2 independent of activation of PI-PLC, suggesting direct coupling to the ET receptor. Constitutive levels of [Ca2+]i rather than abrupt increments in [Ca2+]i are sufficient for activation of this receptor-effector system, with no obligatory requirement for PKC.

Angiotensin-mediated phosphatidylcholine hydrolysis and protein kinase C activation in mesangial cells.

Angiotensin II (ANG II) in mesangial cells (MC) promotes phosphatidylinositol (PI) hydrolysis resulting in diacylglycerol (DAG)-mediated increases in protein kinase C (PKC) activity. The paucity of MC inositol lipid prompted us to consider whether phosphatidylcholine (PC) could sustain DAG formation. ANG II released choline and increased phosphatidylethanol (PEt) via PC-phospholipase D (PC-PLD). ANG II also stimulated phosphorylcholine consequent to PC-phospholipase C (PC-PLC) activation. ANG II-mediated PC hydrolysis augmented DAG for 30 min. PC breakdown was influenced by extracellular Ca2+, because Ni2+ partially inhibited ANG II-induced PEt and obliterated agonist-mediated DAG formation. The consequence of Ca2+ modulation of PC metabolism was investigated by measuring PKC activity. Ni2+ had no effect on early (PI-associated) activation by ANG II at 90 s but obviated translocation from cytosol to the membrane at 10 min. The pathway responsible for PC-associated DAG was studied in PKC downregulated cells. Whereas downregulation prevented PLD-mediated PEt elevation, ANG II-stimulated DAG formation in myristate-labeled cells was unaltered, indicating PC-PLC activation. In summary, ANG II stimulates PC-PLD and PC-PLC in MC. PC-PLD is tightly regulated by PKC, whereas PC-PLC is stringently controlled by extracellular Ca2+. ANG II mediated PC breakdown principally via PC-PLC provides a mechanism for maintaining elevated DAG levels and PKC activation.

Progesterone-evoked increases in sperm [Ca2+]i correlate with the egg penetrating ability of sperm from fertile but not infertile men.

OBJECTIVES: To investigate the relationship between sperm capacitation and intracellular calcium ([Ca2+]i) and to correlate these findings with routine semen parameters and sperm fertilizing ability. DESIGN: Baseline and P-evoked increases in [Ca2+]i of fresh versus capacitated human sperm were measured for known fertile donors and infertile men and compared with the results of semen analysis and in vitro penetration of zona-free hamster eggs. SETTING: Andrology laboratory in a university hospital. PATIENTS: Infertile men undergoing semen analysis. INTERVENTIONS: Capacitation of spermatozoa and exposure of sperm to P (1 microgram/mL). MAIN OUTCOME MEASURES: [Ca2+]i as measured using fura-2, percent zone-free hamster eggs penetrated, and number of penetrating sperm per egg. RESULTS: Steady state [Ca2+]i increased from 74 +/- 32 nM to 166 +/- 97 nM after capacitation, as did P-evoked peak and plateau [Ca2+]i. Deletion of calcium from the assay buffer with ethylene-bis (oxy-ethylenenitriolo) tetraacetic acid abrogated the P-evoked increments. RU486, a P receptor antagonist; reduced the P-evoked response in a dose-dependent manner. Progesterone-evoked calcium responses of sperm varied between different ejaculates of the same fertile donor and correlated with their egg penetrating ability. Sperm from infertile men with abnormal morphology exhibited lower egg penetrating ability and lower mean peak P-evoked [Ca2+]i than morphologically normal sperm. However, free intracellular calcium parameters correlated only weakly with penetrating ability for individual infertile men. CONCLUSION: Progesterone-evoked increases in [Ca2+]i in motile capacitated spermatozoa cannot be used to discriminate between dysfunctional spermatozoa and those capable of penetrating eggs.

Reversible renal failure in the primary antiphospholipid syndrome--a report of two cases.

The primary antiphospholipid/anticardiolipin syndrome is a recently described entity wherein multiorgan thrombotic events occur in the absence of objective evidence of systemic lupus erythematosus. The spectrum of renal involvement remains poorly described. Two patients with coagulation abnormalities consistent with the primary antiphospholipid/anticardiolipin syndrome who developed profound renal insufficiency are reported. Striking microangiopathic lesions were documented on renal biopsy. Renal function improved concomitant to the institution of steroid therapy. Reversible renal failure should be added to the spectrum of clinical manifestations of this entity. The diagnosis of the primary antiphospholipid/anticardiolipin syndrome should be contemplated in individuals with unexplained acute renal failure.

Endothelin activation of phospholipase D: dual modulation by protein kinase C and Ca2+.

Previous work from this laboratory has identified an endothelin (ET) type A (ETA) receptor on cultured rat renal medullary interstitial cells (RMIC), coupled to phosphatidylinositol-specific phospholipase C (PI-PLC), dihydropyridine-insensitive receptor-operated Ca2+ channels, and phospholipase A2. The current studies explored a role for ET stimulation of phosphatidylcholine-specific phospholipase D (PC-PLD) in intracellular signaling of this cell type. ET stimulated PLD activation, as measured by phosphatidic acid (PA) or phosphatidylethanol (PEt) accumulation, in a time- and concentration-dependent manner. Inhibition of diacylglycerol (DAG) kinase by ethylene glycol dioctanoate or 6-(2)4-[(4-fluorophenyl)-phenylmethylene]-1-piperadinyl]ethy l-7-methyl-5H - thiaxolo-[3,2-alpyrimidin]-5-one (R 59022) failed to blunt PA accumulation, indicating that PLD, and not DAG, was the source of PA. Inhibition of PA phosphohydrolase (PAP) by propranolol increased late accumulation of PA, suggesting that the prevailing metabolic flow was in the direction of PA to DAG. Phorbol 12-myristate 13-acetate (PMA) augmented ET-evoked PEt accumulation, whereas downregulation of protein kinase C (PKC) obviated agonist-induced PEt production. PMA augmentation of PLD activity proceeded independent of cytosolic free Ca2+ concentration. Ca2+ derived from either intracellular or extracellular sources enhanced ET-related PEt accumulation but was without effect in PKC-downregulated cells. Collectively, these observations indicate that ET stimulates PLD production in RMIC. PKC is the major regulator of this process, with Ca2+ playing a secondary, modulatory role. In addition, these data suggest that PC-PLD is coupled to the ETA receptor.