Leopoldo Ardiles

Gremlin: A Novel Mediator of Epithelial Mesenchymal Transition and Fibrosis in Chronic Allograft Nephropathy

BACKGROUND Chronic allograft nephropathy (CAN) is the most frequent cause of chronic dysfunction and late loss of renal allografts. Epithelial mesenchymal transition (EMT) has been identified as responsible for the presence of activated interstitial fibroblasts (myofibroblasts) and transforming growth factor beta (TGF-beta)/Smad is the key signaling mediator. It has been proposed that the bone morphogenetic protein 7 (BMP-7) antagonist, Gremlin, could participate in EMT, as a downstream mediator of TGF-beta. METHODS We evaluated 33 renal allograft biopsies, 16 of which showed CAN, versus 17 controls. By in situ hybridization we studied the expression of TGF-beta and Gremlin mRNA. Gremlin, BMP-7, E-cadherin, and alpha-smooth muscle actin (alpha-SMA) proteins were evaluated by immunohistochemistry and Smad3 activation by Southwestern. In cultured human tubuloepithelial cells (HK2 cell line), Gremlin induction by TGF-beta was studied by confocal microscopy. RESULTS Among renal biopsies of transplanted patients with CAN, we detected up-regulation of TGF-beta in colocalization with Gremlin (RNA and protein), mainly in areas of tubulointerstitial fibrosis. In the same tubules, we observed decreased expression of E-cadherin and induction of vimentin and alpha-SMA. BMP-7 was significantly decreased in the CAN biopsies. In addition, HK2 stimulated with TGF-beta (1 ng/mL) induced Gremlin production at 72 hours. CONCLUSION We postulated that Gremlin is a downstream mediator of TGF-beta, suggesting a role for Gremlin in EMT observed in CAN.

Glomerular Localization of Platelet Factor 4 in Streptococcal Nephritis

Since platelet factor 4 (PF4), a cationic (pI 7.6) platelet secretory protein, binds avidly to glomerular polyanions both in vitro and in vivo, and is implicated in neutrophil chemotaxis, we studied by indirect immunofluorescence microscopy the presence of PF4 deposits in glomeruli of patients with poststreptococcal nephritis (APSGN). Goat antihuman PF4 serum was used as primary antibody and fluorescein-conjugated IgG fraction of rabbit antigoat IgG as second antibody. Controls consisted of nonimmune goat serum or anti-PF4 serum preabsorbed with human PF4, as primary antibodies. Glomerular deposits of PF4 were demonstrated in renal tissues obtained by biopsy in 14 of 20 patients studied; the deposits were particularly intense in 9 patients. PF4 was bound to the mesangium and to the capillary walls. There was a significant positive correlation between intraglomerular deposits of PF4 and the levels of proteinuria (p = 0.024). These findings provide further evidence for a role of platelets in the pathogenesis of APSGN and suggest that PF4 may contribute to alter the glomerular permeability in this disease.

Study of neoral kinetics in adult renal transplantation treated with diltiazem

THE INTERACTION between cyclosporine A (CsA) and diltiazem that results in increased CsA blood concentration has been established previously. Both drugs are cytochrome P450 3A4 substrates, and their interaction explained by competitive inhibition of CsA metabolism in the liver. Calcium channels blockers (CCB) are useful in preventing CsA nephrotoxicity. Cotreatment with CCB has been postulated to improve early graft function, reduce acute rejection, and decrease the incidence of delayed graft function. The effect of cotreatment with the CCB diltiazem on the pharmacokinetic of the new microemulsion formulation of CsA (Neoral) and its metabolites has not been specifically examined in adults.

Antineutrophil-Cytoplasmic-Autoantibodies in Poststreptococcal Nephritis

Sera from 210 patients with APSGN, were tested for the presence of ANCA (IgG-isotype). Indirect immunofluorescence (IF) on ethanol fixed human PMNs was used, and for those positive sera, ELISA kits for PR3 (Proteinase 3) and MPO (Myeloperoxidase) was performed. ANCA were detected in 9% (18 out of 210 cases) in a predominantly diffuse cytoplasmic staining pattern in 14 cases (77%), and in a perinuclear pattern in the remaining 4 cases (22%). Anti-MPO was found in 4 cases (C-ANCA 3; P-ANCA 1) and anti-PR3 was always negative. The presence of ANCA was significantly associated with a more severe glomerular disease as assessed by the serum creatinine value and the crescents formation. Longitudinal studies performed in 11 cases have shown that raised levels of these autoantibodies may persist for at least six months, without relationship with disease activity. Further studies are required to dilucidate the specificity of these autoantibodies, and if its presence is either an epiphenomenon of the heterogeneous humoral immune response in streptococcal infection, or they play some pathogenic role in APSGN.

Decreased Platelet Counts and Decreased Platelet Serotonin in Poststreptococcal Nephritis

Mean platelet survival time in patients with acute poststreptococcal glomerulonephritis (APSGN) is reduced to 50-60% of the control values, and glomerular deposits of platelet factor 4 are found in these patients. In order to investigate further systemic platelet changes of pathogenic, clinical or prognostic significance, we measured the platelet serotonin (5-HT) content and the blood platelet counts during the 1st week of the disease in 27 patients with APSGN. Platelet 5-HT was significantly reduced in patients with APSGN as compared with patients with impetigo without glomerular involvement (785 +/- 54 vs. 1,329 +/- 94 ng 5-HT/10(9) platelets; p < 0.001). Similarly, the mean blood platelet count was reduced to 247 +/- 16 x 10(3) as compared with 303 +/- 14 x 10(3) in the controls (p < 0.05). Thirteen (48%) of these patients had individual values of platelet 5-HT lower than the 95% confidence interval calculated in the control group. No significant correlation was observed between the concentration of 5-HT and either the severity of the disease judged by the amount of urinary protein excretion and the serum creatinine value or the presence of circulating immune complexes. Significant correction of the platelet 5-HT content (to 1,180 +/- 111 ng/10(9) platelets; p < 0.01) and of the platelet counts (to 309 +/- 21 x 10(3); p < 0.01) were observed in the longitudinal study at least 2 weeks later. Platelet activation, with secretion of granular content and increased consumption, may explain these findings. Additionally, the reduced mean age of the circulating platelets could contribute to their decreased 5-HT levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Presence of Circulating Immune Complexes in the Classic Form of Hemolytic Uremic Syndrome: A Constant Finding

In an attempt to further study the possible contribution of circulating immune complexes (CIC) in the pathogenesis of the classic form of hemolytic uremic syndrome, 9 patients were studied during the acute phase of the diseases. C1q solid-phase ELISA and conglutinin solid-phase ELISA, were used to measure the levels of immune complexes. All 9 were positive in one or both assays. No correlation was found between the levels of CIC and the clinical severity of the disease. The constant finding of positive CIC in these patients might represent an epiphenomenon, point out the postinfectious nature of this disease but also suggest a possible pathogenic role.

Age Influence on Mononuclear Phagocyte System Fc-Receptor Function in Poststreptococcal Nephritis

The Fc-receptor function of the mononuclear phagocyte system (MPS) was examined in 41 children and adult patients, by measuring the clearance of IgG-sensitized, 51Cr-labeled erythrocytes. The Fc-receptor-mediated clearance observed in patients (mean +/- SE) was not significantly different as compared to the control group of similar age distribution. However, the immune clearance time was significantly age-correlated in both groups (acute poststreptococcal glomerulonephritis, APSGN, r = 0.39, p less than 0.05; control r = 0.63, p less than 0.01). The magnitude of the Fc-specific immune clearance and the serum creatinine were also significantly correlated (r = 0.59; p less than 0.01). Circulating immune complexes (as measured by the C1q and conglutinin ELISA) did not correlate with immune clearance, which remained stable in longitudinal studies. Age-related changes in MPS Fc-receptor function could explain, at least in part, prognostic differences between children and adult patients with APSGN.