12 research outputs found
Studio della modulazione genica di TGF-beta1 da parte di diverse formulazioni di GAGs mediante Real Time PCR e chimica SYBR Green I in cellule mesangiali in vitro.
\u201cReal Time PCR con chimica SYBR Green I e RT/PCR semiquantitativa a confronto per la determinazione dell\u2019espressione genica di fattori di crescita e citochine in differenti tessuti e in sistemi in vivo e in vitro\u201d.
ANALISI DEL PATHWAY MEDIATO DA SMAD DURANTE LA TRANSIZIONE EPITELIO-MESENCHIMALE INDOTTA DA TGFbeta1 IN CELLULE HUTEC IN COLTURA PRIMARIA
Patologia molecolare del gene del canale del cloro CLCN5 in pazienti con nefrolitiasi idiopatica: analisi di mutazione e studio di espressione
STUDIO DELL\u2019ESPRESSIONE GENICA DEL RAS INTRARENALE IN BIOPSIE SERIATE DI PAZIENTI TRAPIANTATI
Precocious activation of genes of the renin-angiotensin system and the fibrogenic cascade in IgA glomerulonephritis
Abstract
BACKGROUND: The renin-angiotensin system (RAS) seems to play a pivotal role in progression of immunoglobulin A (IgA) nephropathy (IgAN). Accordingly, in patients with IgAN a relationship between the RAS and the fibrogenic cascade triggered by transforming growth factor-beta1 (TGF-beta1) should be observed. This study was carried out to obtain deeper insight into the regulation of RAS and the interaction with TGF-beta1 in the diseased kidney.
METHODS: Twenty renal biopsies from IgAN patients and five from renal cancer patients (controls) were analyzed in both microdissected glomerular and tubulointerstitial compartments by reverse transcription-polymerase chain reaction (RT-PCR). All patients had normal renal function. The expression of the following genes was determined: angiotensinogen (Agtg), renin, angiotensin-converting enzyme (ACE), angiotensin II (Ang II) type 1 and type II (AT1 and AT2 receptors), TGF-beta1, collagen IV (Coll IV), alpha-smooth muscle actin (alpha-SMA). Quantitative data were confirmed for TGF-beta1 and ACE genes by real-time PCR. Results. RAS genes were overexpressed in IgAN patients vs. control subjects. There was no difference between glomerular and tubulointerstitial RAS gene expression levels. On the contrary, the overactivation of fibrogenic cascade genes (TGF-beta1, Coll IV, alpha-SMA) in the tubulointerstitium was observed (TGF-beta1, glomerular 0.14 +/- 0.10 SD; tubulointerstial 0.34 +/- 0.20; P = 0.000) (alpha-SMA, glomerular 0.08 +/- 0.07; tubulointerstitial 0.35 +/- 0.19; P = 0.000) (Coll IV, glomerular 0.12 +/- 0.11; tubulointerstitial 0.22 +/- 0.10; P = 0.03). This fibrogenic cascade seems to be triggered by RAS as indicated by statistically significant correlations between the expression of their respective genes. A direct relationship between the putative Ang II activity and the expression of AT receptor genes was found in the tubulointerstitium, whereas in the glomeruli this relationship was negative. In the interstitium, statistically significant positive relationships emerged between interstitial infiltrates and the gene expression of Agtg, AT1 receptor, Coll IV, and TGF-beta1.
CONCLUSION: This study demonstrates that a tight regulation of the intrarenal RAS exists in IgAN and that it follows the general rules disclosed in animal models. Moreover, the RAS seems to be activated early in the diseased kidney and it appears that such activation drives inflammation and a parallel stimulation of the TGF-beta fibrogenic loop, particularly at the tubulointerstitial level
Identification of a novel splice site mutation of CLCN5 gene and characterization of a new alternative 5’ UTR end of ClC-5 mRNA in human renal tissue and leukocytes
Perforin, Granzyme B, and fas ligand for molecular diagnosis of acute renal-allograft rejection: analyses on serial biopsies suggest methodological issues.
BACKGROUND: The Perforin-Granzyme B and Fas/Fas Ligand apoptotic mechanisms are involved in the development of acute renal rejection (AR). We describe our experience of analyzing the expression of cytotoxic T-lymphotoxins (CTL) in biopsies and peripheral blood leukocytes (PBL) for the diagnosis of AR.
METHODS: We analyzed Perforin (P), Granzyme B (GB) and Fas Ligand (FL) expression in 68 renal biopsies and 64 PBL using comparative kinetic RT-PCR and, for GAPDH and FL, we also replicated with real-time RT-PCR. The levels of expression were measured in different groups, such as T0 (biopsies before reperfusion and PBL in recipient before the transplant [Tx]), Td (biopsies and PBL collected for clinical purposes) and Tp (biopsies and PBL two months after Tx).
RESULTS: A higher CTL expression was seen in non-rejecting (NR) biopsies in the first 2 months after Tx. P and FL were significantly more expressed during AR in all biopsies and in Td, while P remained upregulated in Tp. In PBL, there was no significant increase in CTL transcription during AR. A variable expression of CTL emerged in all T0 biopsies.
CONCLUSIONS: Two lytic pathways are activated in biopsies when AR occurs shortly after Tx, whereas the P/GB mechanism prevails if it occurs later on. Only P and FL in biopsies might be able to predict AR diagnosis, but with a considerable variability in each sample, possibly due to the small portion of tissue core, which may be inadequate for molecular diagnosis. CTL expression in PBL does not correlate with histological AR