Ana Konvalinka

Randomized Controlled Trial of Chlorhexidine Gluconate for Washing, Intranasal Mupirocin, and Rifampin and Doxycycline Versus No Treatment for the Eradication of Methicillin-Resistant Staphylococcus aureus Colonization

BACKGROUND Eradication of methicillin-resistant Staphylococcus aureus (MRSA) carriage may reduce the risk of MRSA infection and prevent transmission of the organism to other patients. METHODS To determine the efficacy of decolonization therapy, patients colonized with MRSA were randomized (3:1 allocation) to receive treatment (2% chlorhexidine gluconate washes and 2% mupirocin ointment intranasally, with oral rifampin and doxycycline for 7 days), or no treatment. Follow-up samples for MRSA culture were obtained from the nares, perineum, skin lesions, and catheter exit sites monthly for up to 8 months. The primary outcome measure was detection of MRSA at 3 months of follow-up. Univariate and multivariable analyses were performed to identify variables associated with treatment failure. RESULTS Of 146 patients enrolled in the study, 112 patients (87 treated; 25 not treated) were followed up for at least 3 months. At 3 months of follow-up, 64 (74%) of those treated had culture results negative for MRSA, compared with 8 (32%) of those not treated (P=.0001). This difference remained significant at 8 months of follow-up, at which time, 54% of those treated had culture results negative for MRSA (chi2=64.4; P<.0001, by log-rank test). The results of the multivariable analysis indicated that having a mupirocin-resistant isolate at baseline was associated with treatment failure (relative risk, 9.4; 95% confidence interval, 2.8-31.9; P=.0003), whereas decolonization therapy was protective (relative risk, 0.1; 95% confidence interval, 0.04-0.4; P=.0002). Mupirocin resistance emerged in only 5% of follow-up isolates. CONCLUSIONS Treatment with topical mupirocin, chlorhexidine gluconate washes, oral rifampin, and doxycycline for 7 days was safe and effective in eradicating MRSA colonization in hospitalized patients for at least 3 months.

Utility of HLA Antibody Testing in Kidney Transplantation

HLA antigens are polymorphic proteins expressed on donor kidney allograft endothelium and are critical targets for recipient immune recognition. HLA antibodies are risk factors for acute and chronic rejection and allograft loss. Solid-phase immunoassays for HLA antibody detection represent a major advance in sensitivity and specificity over cell-based methods and are widely used in organ allocation and pretransplant risk assessment. Post-transplant, development of de novo donor-specific HLA antibodies and/or increase in donor-specific antibodies from pretransplant levels are associated with adverse outcomes. Although single antigen bead assays have allowed sensitive detection of recipient HLA antibodies and their specificities, a number of interpretive considerations must be appreciated to understand test results in clinical and research contexts. This review, which is especially relevant for clinicians caring for transplant patients, discusses the technical aspects of single antigen bead assays, emphasizes their quantitative limitations, and explores the utility of HLA antibody testing in identifying and managing important pre- and post-transplant clinical outcomes.

Effect of protein kinase Cβ inhibition on renal hemodynamic function and urinary biomarkers in humans with type 1 diabetes: a pilot study.

OBJECTIVE—The aim of this study was to examine the effect of protein kinase Cβ inhibition with ruboxistaurin on renal hemodynamic function and urinary biomarkers (monocyte chemoattractant protein-1 [MCP-1] and epidermal growth factor) in renin angiotensin system blockade-treated type 1 diabetic subjects. RESEARCH DESIGN AND METHODS—Albuminuric subjects were randomized (2:1) to ruboxistaurin (32 mg daily; n = 13) or placebo (n = 7) for 8 weeks. Renal hemodynamic function was measured during clamped euglycemia or hyperglycemia and before and after ruboxistaurin or placebo. RESULTS—Ruboxistaurin was not associated with between-group differences during clamped euglycemia or hyperglycemia. In a post hoc analysis comparing hyperfilterers with normofilterers during euglycemia, glomerular filtration rate and MCP-1 decreased, whereas the epidermal growth factor–to–MCP-1 ratio increased in hyperfilterers versus normofilterers (all P < 0.05). CONCLUSIONS—The effect of ruboxistaurin is modest and dependent, at least in part, on the level of ambient glycemia and baseline glomerular filtration rate.

Searching for New Biomarkers of Renal Diseases through Proteomics

BACKGROUND Technological advances have resulted in a renaissance of proteomic studies directed at finding markers of disease progression, diagnosis, or responsiveness to therapy. Renal diseases are ideally suited for such research, given that urine is an easily accessible biofluid and its protein content is derived mainly from the kidney. Current renal prognostic markers have limited value, and renal biopsy remains the sole method for establishing a diagnosis. Mass spectrometry instruments, which can detect thousands of proteins at nanomolar (or even femtomolar) concentrations, may be expected to allow the discovery of improved markers of progression, diagnosis, or treatment responsiveness. CONTENT In this review we describe the strengths and limitations of proteomic methods and the drawbacks of existing biomarkers, and provide an overview of opportunities in the field. We also highlight several proteomic studies of biomarkers of renal diseases selected from the plethora of studies performed. SUMMARY It is clear that the field of proteomics has not yet fulfilled its promise. However, ongoing efforts to standardize sample collection and preparation, improve study designs, perform multicenter validations, and create joint industry-regulatory bodies offer promise for the recognition of novel molecules that could change clinical nephrology forever.

Determinants of Long-Term Graft Outcome in Transplant Glomerulopathy

Background. Transplant glomerulopathy (TG) is a renal allograft disease defined by glomerular basement membrane duplication with peritubular capillary basement membrane multilayering (PTCML), and associated with anti-human leukocyte antigen antibodies and C4d. Outcome in TG is poor but variable, and prognostic factors, particularly those affecting long-term outcome, are not well known. We investigated several potentially prognostic clinical and pathologic factors in TG and evaluated estimated glomerular filtration rate (eGFR) slopes to assess graft function and early decline. Methods. We examined all cases of TG from 2001 to 2005 with at least 4-year follow-up after biopsy, excluding those with a second confounding diagnosis. Results. Among 36 cases of pure TG, mean graft age at biopsy was 8.8±6 years. C4d stain was positive in 11 (33%) cases. Clinical characteristics at biopsy were not different based on C4d. C4d was associated with greater PTCML (P=0.03), peritubular capillaritis (P=0.04), and glomerulitis (P=0.03). Death-censored graft survival was significantly associated with interstitial fibrosis (P=0.001), PTCML (P=0.001), and arteriolar hyalinosis (P=0.007), and it showed a trend with proteinuria (P=0.07) and C4d positivity (P=0.08). C4d-positive cases also showed a trend toward rapid graft loss. Analysis of eGFR slopes showed a pattern of preserved, slightly negative slope from transplant until approximately 1 year before biopsy, at which point the slope became significantly more negative (P<0.001). Conclusion. Interstitial fibrosis, PTCML, and arteriolar hyalinosis were significant predictors of graft survival in TG. C4d positivity was associated with a more rapid rate of function decline. eGFR slope data showed significant deterioration in graft function well before the diagnostic biopsy.

Loss of ACE2 exacerbates murine renal ischemia-reperfusion injury.

Ischemia-reperfusion (I/R) is a model of acute kidney injury (AKI) that is characterized by vasoconstriction, oxidative stress, apoptosis and inflammation. Previous studies have shown that activation of the renin-angiotensin system (RAS) may contribute to these processes. Angiotensin converting enzyme 2 (ACE2) metabolizes angiotensin II (Ang II) to angiotensin-(1–7), and recent studies support a beneficial role for ACE2 in models of chronic kidney disease. However, the role of ACE2 in models of AKI has not been fully elucidated. In order to test the hypothesis that ACE2 plays a protective role in AKI we assessed I/R injury in wild-type (WT) mice and ACE2 knock-out (ACE2 KO) mice. ACE2 KO and WT mice exhibited similar histologic injury scores and measures of kidney function at 48 hours after reperfusion. Loss of ACE2 was associated with increased neutrophil, macrophage, and T cell infiltration in the kidney. mRNA levels for pro-inflammatory cytokines, interleukin-1β, interleukin-6 and tumour necrosis factor-α, as well as chemokines macrophage inflammatory protein 2 and monocyte chemoattractant protein-1, were increased in ACE2 KO mice compared to WT mice. Changes in inflammatory cell infiltrates and cytokine expression were also associated with greater apoptosis and oxidative stress in ACE2 KO mice compared to WT mice. These data demonstrate a protective effect of ACE2 in I/R AKI.

Insights into Diabetic Kidney Disease Using Urinary Proteomics and Bioinformatics

A number of proteomic and peptidomic analyses of urine from diabetic subjects have been published in the quest for a biomarker that predicts progression of nephropathy. Less attention has been paid to the relationships between urinary proteins and the underlying biological processes revealed by the analyses. In this review, we focus on the biological processes identified by studying urinary proteins and protein-protein interactions at each stage of diabetic nephropathy to provide an overview of the events underlying progression of kidney disease reflected in the urine. In uncomplicated diabetes, proteomic/peptidomic analyses indicate that early activation of fibrotic pathways in the kidney occurs before the onset of microalbuminuria. In incipient nephropathy, when albumin excretion rates are abnormal, proteomic/peptidomic analyses suggest that changes in glomerular permselectivity and tubular reabsorption account, at least in part, for the proteins and peptides that appear in the urine. Finally, overt nephropathy is characterized by proteins involved in wound healing, ongoing fibrosis, and inflammation. These findings suggest that there is a spectrum of biological processes in the diabetic kidney and that assessing protein networks may be more informative than individual markers with respect to the stage of disease and the risk of progression.

Determination of an Angiotensin II-regulated Proteome in Primary Human Kidney Cells by Stable Isotope Labeling of Amino Acids in Cell Culture (SILAC)

Background: Specific markers of kidney angiotensin-II (AngII) activity are needed. Results: Eighteen AngII-regulated proteins were discovered and confirmed by proteomics in human kidney cells, whereas heme oxygenase-1 was validated in a mouse. Conclusion: Heme oxygenase-1 and other AngII-regulated proteins represent novel markers of AngII activity. Significance: AngII-regulated proteins may represent kidney-specific AngII activity markers in patients and in experimental models. Angiotensin II (AngII), the major effector of the renin-angiotensin system, mediates kidney disease progression by signaling through the AT-1 receptor (AT-1R), but there are no specific measures of renal AngII activity. Accordingly, we sought to define an AngII-regulated proteome in primary human proximal tubular cells (PTEC) to identify potential AngII activity markers in the kidney. We utilized stable isotope labeling with amino acids (SILAC) in PTECs to compare proteomes of AngII-treated and control cells. Of the 4618 quantified proteins, 83 were differentially regulated. SILAC ratios for 18 candidates were confirmed by a different mass spectrometry technique called selected reaction monitoring. Both SILAC and selected reaction monitoring revealed heme oxygenase-1 (HO-1) as the most significantly up-regulated protein in response to AngII stimulation. AngII-dependent regulation of the HO-1 gene and protein was further verified in PTECs. To extend these in vitro observations, we overlaid a network of significantly enriched gene ontology terms from our AngII-regulated proteins with a dataset of differentially expressed kidney genes from AngII-treated wild type mice and AT-1R knock-out mice. Five gene ontology terms were enriched in both datasets and included HO-1. Furthermore, HO-1 kidney expression and urinary excretion were reduced in AngII-treated mice with PTEC-specific AT-1R deletion compared with AngII-treated wild-type mice, thus confirming AT-1R-mediated regulation of HO-1. Our in vitro approach identified novel molecular markers of AngII activity, and the animal studies demonstrated that these markers are relevant in vivo. These interesting proteins hold promise as specific markers of renal AngII activity in patients and in experimental models.

Characterization of the Intrarenal Renin-Angiotensin System in Experimental Alport Syndrome

Blockade of the renin-angiotensin system attenuates the progression of experimental and clinical Alport syndrome (AS); however, the underlying mechanism(s) remains largely unknown. We evaluated the renin-angiotensin system in 4- and 7-week-old homozygous for collagen, type IV, α3 gene (Col4A3(-/-)) and wild-type mice, a model of AS characterized by proteinuria and progressive renal injury. Renal angiotensin (Ang) II levels increased, whereas renal Ang-(1-7) levels decreased in 7-week-old Col4a3(-/-) mice compared with age-matched controls; these changes were partially reversed by recombinant angiotensin-converting enzyme 2 (ACE2) treatment. The expression of both the angiotensinogen and renin protein increased in Col4a3(-/-) compared with wild-type mice. Consistent with the Ang-(1-7) levels, the expression and activity of kidney ACE2 decreased in 7-week-old Col4a3(-/-) mice. The urinary excretion rate of ACE2 paralleled the decline in tissue expression. Expression of an Ang II-induced gene, heme oxygenase-1, was up-regulated in the kidneys of 7-week-old Col4a3(-/-) mice compared with wild-type mice by microarray analysis. Heme oxygenase-1 (HO-1) protein expression was increased in kidneys of Col4a3(-/-) mice and normalized by treatment with ACE inhibitor. Urinary HO-1 excretion paralleled renal HO-1 expression. In conclusion, progressive kidney injury in AS is associated with changes in expression of intrarenal renin Ang system components and Ang peptides. HO-1 and ACE2 may represent novel markers of AS-associated kidney injury, whereas administration of recombinant ACE2 and/or Ang-(1-7) may represent novel therapeutic approaches in AS.

Urine proteomics for acute kidney injury prognosis: another player and the long road ahead

Urine represents a mine for proteomic markers of renal diseases, but its analysis is hindered by unresolved technical issues. Aregger et al. used an unbiased proteomics approach to analyze urine of critically ill patients. They discovered insulin-like growth factor-binding protein 7 (IGFBP-7) and linked it to acute kidney injury (AKI) outcome. Future efforts should address the biology of IGFBP-7 and related proteins to determine their role in AKI.