Gabriel Joelsons

Noninvasive Tim-3 messenger RNA evaluation in renal transplant recipients with graft dysfunction.

Background. Renal biopsies are usually needed to elucidate graft dysfunction. In this study, T-cell immunoglobulin domain, mucin domain mRNA expression in the peripheral blood leukocytes (PBL) and urinary cells (UC) were studied as a noninvasive method for the diagnosis of acute rejection (AR) of kidney transplant patients with dysfunction. Methods. One hundred sixty biopsies were obtained from 115 patients. Blood and urine samples were collected immediately before the biopsies. Histopathologic diagnoses were acute tubular necrosis with superimposed AR or acute tubular necrosis in patients with delayed graft function (DGF), and (AR), or calcineurin inhibitor nephrotoxicity (CIN), or interstitial fibrosis and tubular atrophy in patients with acute graft dysfunction (AGD). Fifteen protocol biopsies of stable grafts were used as controls. mRNA relative quantification was performed by real-time polymerase chain reaction. Results. Gene expression in tissue, PBL, and UC was always higher in patients with AR than in patients with the other causes of graft dysfunction (P<0.001). Significant correlations of gene expression in different compartments were observed (P<0.001). The obtained diagnostic parameters were 100% accurate in the DGF group and, respectively, for blood and urine: sensitivity (87% and 84%); specificity (95% and 96%); positive predictive value (87% and 89%); negative predictive value (93% and 94%); and accuracy (91% and 93%) for the group of patients with AGD. Conclusion. T-cell immunoglobulin domain, mucin domain mRNA quantification by real-time polymerase chain reaction in PBL and UC of renal transplant patients undergoing DGF or AGD may become a useful tool for an accurate noninvasive diagnosis of AR.

Messenger RNA levels of podocyte-associated proteins in subjects with different degrees of glucose tolerance with or without nephropathy

BackgroundTo investigate gene expression of podocyte-specific proteins in urine of diabetes and prediabetes subjects and the association of these proteins with albuminuria.MethodsFifteen controls, 19 prediabetes, and 67 diabetes subjects were included. Messenger RNA of nephrin, podocin, podocalyxin, synaptopodin, TRPC6, alpha-actinin-4, and TGF-β1 were measured using RT-PCR. Podocyte marker expression was correlated with albuminuria, glycemic control, and renal function. The diagnostic performance of the genes used to detect increased albuminuria was assessed using ROC curves and Poisson regressions.ResultsPodocyte marker expression was significantly higher in diabetic subjects. Urinary nephrin was correlated with increasing levels of albuminuria; risk of albuminuria increased by 20% for every one-unit increase in the log10 of nephrin mRNA. Nephrinuria was found in 53%, 71%, and 90% of normo-, micro-, and macroalbuminuric diabetes subjects, respectively (p = 0.023). Urinary nephrin, podocalyxin, TRPC6, podocin, and alpha actinin-4 were correlated with glycemic control and albuminuria but not with renal function.ConclusionsDiabetes subjects had higher urinary mRNA levels of podocyte proteins than nondiabetic subjects, even the normoalbuminuric patients. Nephrinuria was correlated with diabetic nephrophathy stage and predicted pathological albuminuria. Urinary mRNA levels of podocyte markers of prediabetic subjects did not differ from controls.

Analysis of FOXP3 gene and protein expressions in renal allograft biopsies and their association with graft outcomes

Background: The transcription factor FOXP3 is increased in acute renal rejection, but its influence on graft outcomes is unclear. This study correlated FOXP3 with dendritic cells and graft outcomes. Methods: We assessed 96 kidney transplants undergoing allograft biopsy for cause. FOXP3 mRNA was analyzed by real-time polymerase chain reaction (PCR) and FOXP3 protein and DCsCD83+ by immunohistochemistry. Graft function and survival were assessed at 5 years post-transplantation, as well as by independent predictors of graft loss. Results: Intragraft FOXP3 gene and protein expression were significantly correlated (r = 0.541, p < 0.001). Both FOXP3 mRNA and protein were increased in patients with acute rejection (AR). High expression of FOXP3 mRNA or protein in biopsies did not correlate with clinical variables, but there was a trend to higher positive variation in the glomerular filtration rate (GFR) from biopsy to last follow-up. Patients with FOXP3-mRNAhigh had more DCsCD83+ in biopsy, but these cells did not associate with AR. Five-year graft survival was not influenced by either FOXP3 mRNA or protein expressions. Conclusions: FOXP3 mRNA and protein had a good correlation in archival renal graft tissue. Increased FOXP3 expression was found in AR and FOXP3 associated with high numbers of DCs. However, both FOXP3 mRNA and protein was not associated with better allograft outcomes.