Primary IgA nephropathy, a chronic nephritis with variable prognosis, is characterized by mesangial immunoglobulin A, frequently with codeposition of other immunoglobulin isotypes and complement components accompanying matrix expansion typically preceding glomerular scarring. Glomerular immunoglobulin G, when present, is localized to the mesangial periphery found variably in repeat biopsies. IgG anti-mesangial cell autoantibodies (IgG-MESCA) in sera of patients with IgA nephropathy, specific by F(ab')(2) binding to 48- and 55-kD autoantigen(s) could account for these deposits, but their in vivo localization, and the functional role in promoting scarring is unknown. A specific monoclonal antibody raised previously to these human mesangial cell autoantigen fractions, in this study localized to similar glomerular sites, reinforcing the view that immunoglobulin G deposition in vivo is a result of antibody-autoantigen binding. The propensity for immunoglobulin G more than other isotypes to enhance inflammation prompted study of its functional role in vitro. Using cultured human mesangial cells in a complement-free tritiated glycosaminoglycan synthesis single outcome assay, purified IgG fractions from patient sera increased matrix production in a dose-dependent manner compared with controls. At a constant total IgG concentration, matrix synthesis was proportional to the titre of IgG-MESCA. Autoreactive IgG stimulated matrix synthesis when compared with controls or IgA fractions. These findings are consistent with IgG-MESCA autoantibodies enhancing mesangial matrix synthesis in vitro, which suggests that in IgA nephropathy, similar prosclerotic autoimmune mechanisms might operate. Recombinant TGFbeta(1) also induced matrix synthesis, raising the possibility that both autoimmune mechanisms and those TGFbeta(1)-dependent are functional or inter-related. The pathogenesis of glomerular scarring and loss in IgA nephropathy may include, in part, these mechanisms.

Chromosomal rearrangements involving the telomeres are implied as a significant cause of idiopathic mental retardation. The most frequently used technique to detect these rearrangements was fluorescent in situ hybridization (FISH), an expensive and labor-intensive technique. One of the most promising alternative techniques is multiplex ligation-dependent probe amplification (MLPA). Here, the authors present the evaluation of a double set of probes (the SALSA P036, P019, and P020 human telomere test kits) on a series of 95 patients and 22 normal controls. Overall, 34 patients had been studied by telomeric FISH and MLPA, which was demonstrated to be a reliable method to detect essentially all subtelomeric rearrangements characterized by FISH. In addition, in these 34 patients, 13 dose imbalances were detected by MLPA, but not by FISH analysis. Overall, 12 alterations were observed only with one of the two sets, and they corresponded to polymorphic variants, as they were inherited from healthy parents or also appear in normal controls. The remaining 61 patients were initially studied with SALSA P036, and any putative dose alteration was confirmed with the two other kits and FISH. In the whole series, the authors found 9 dose imbalances evidenced with 2 MLPA kits and confirmed by FISH, representing 10% of patients with subtelomeric rearrangements. On the other hand, one small duplication at 14q11 may be clinically relevant as it appears de novo in one patient. In conclusion, MLPA can be considered a quick, sensitive, cost-effective, and easy method to screen for subtelomeric rearrangements, but any finding based in the testing of one probe should be confirmed by other sources.

Intra- and extracellular lipid accumulation and the production of inflammatory mediators by renal and accessory cells may play an important role in the initiation and progression of these lesions. Platelet activating factor (PAF) is a biologically active phospholipid that is produced by various cells upon activation by different stimuli. It has been suggested that PAF plays a role in atherogenesis, and several studies indicated its participation in the pathogenesis of renal diseases. The aim of this study is to investigate the role of PAF on intracellular lipid accumulation and gene regulation of lipoprotein receptors in human mesangial cells (HMCs). A human mesangial cell line (HMC) was used to study the effects of PAF on foam cell formation by Oil red O staining and on the expression of LDLr, SR-AI, and PAF-R mRNA using RT-PCR. Native LDL caused foam cell formation in HMC in the presence of PAF. PAF enhanced LDLr expression and overrode LDL receptor suppression induced by a high concentration of LDL. Moreover, it enhanced SR-AI expression. PAF also caused increase in PAF-R expression. The above data suggest that PAF enhances its own receptor expression and then increases lipid accumulation by dysregulating LDL receptor regulation and inducing scavenger receptor expression in HMCs. These results suggest that PAF has a potential role in lipid mediated renal injury.

Integrin is the major adhesion molecule for the attachment of podocytes to the glomerular basement membrane, and integrins have been shown to play a major role in the regulation of cell survival. In this study, the authors investigated the apoptosis and its related signal pathways to integrin in cultured rat podocytes. Apoptosis was detected with the terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) technique. Cytochrome c was examined by immunohistochemical stain, and Fas, Fas ligand, Bax, Bcl-2, and ERK activation (p-ERK/ERK) were analyzed by Western blotting analysis. The results demonstrated that the integrin antagonist, Gly-Arg-Gly-Asp (GRGD), increased the percentage of cells with apoptosis (from 0.9+/-0.5% to 27.2+/-9.9%, P < 0.01). Inhibition of protein tyrosine kinase with genistein also caused apoptosis of podocytes (from 0.9+/-0.5% to 26.0+/-8.7%, P < 0.01). In GRGD-treated cells, cytochrome c was found released into cytoplasm by immunohistochemical study and the Bax expression was upregulated, whereas Bcl-2 expression was not changed. Fas was not expressed in both control and GRGD-treated podocytes, although Fas ligand was upregulated in GRGD-treated cells. ERK activation was also found to be increased in GRGD-treated cells. The results indicated that alpha3beta1integrin is necessary for the prevention of the apoptosis of cultured rat podocytes, and that the signaling involves the Bax, Bcl-2, and cytochrome c pathways.

Clear cell carcinoma of the kidney, the most common subtype of renal cell cancer, displays different biological behavior in different patients. This heterogeneity cannot be recognized by light microscopy. In this study, gene expression in 16 clear cell renal cell carcinoma samples and 17 non-malignant tissue types comprising 539 samples was determined using oligonucleotide microarrays representing approximately 40,000 known genes and ESTs. Differences in gene expression were quantified as the fold change in gene expression between the various sets of samples. A set of genes was identified that was overexpressed in the renal cell carcinoma samples compared with the normal kidney samples. Principle component analysis of the set of renal cell carcinomas using this set of genes overexpressed in renal cell cancer revealed the existence of 2 major subgroups among the renal carcinomas. A series of principle component analyses of the set of renal cell carcinomas using different gene sets composed of genes involved in different metabolic pathways also revealed the same 2 major subgroups of the renal cell cancers. Eisen clustering using the same genes also revealed the same 2 major renal cell cancer subsets. Review of the pathology suggested that these 2 subgroups differed in pathologic grade. Genes differentially expressed between the 2 renal cell cancer subsets were identified. Examination of gene expression in each renal cell cancer subset and the pool of renal cell carcinoma samples compared with that in 17 different normal tissues revealed genes specifically overexpressed in renal cell cancer compared with these normal tissues. The authors conclude that gene expression patterns may be useful in helping to further classify subtypes of renal cell carcinoma that may have clinical significance. In addition, the genes identified as overexpressed in each set of clear cell renal cell carcinomas compared with normal tissues may represent useful targets for therapy.

The aim of this study was to compare the number of peripheral blood natural killer (NK) cells, NK cell activity, expression of NK cell activating receptors, and serum cytokine levels in patients with primary Sjögren's syndrome (SS) vs normal controls. The authors found that NK cell number, NK cell killing activity, and the expression of activating receptors CD2 and NKG2D were significantly decreased, and the expression of NKp46, as well as the percentage of apoptotic NK cells, were significantly increased in primary SS patients compared with healthy controls. NK cell killing activity on a per-cell basis was similar in primary SS patients and healthy controls. Moreover, the levels of IL-18 and TNF-alpha, cytokines that have been shown to promote NK cell death, were significantly increased in sera from patients with primary SS compared with controls. These data suggest that reduced NK cell numbers, probably a result of apoptotic death, may contribute to impaired NK cell activity in patients with primary SS.

Proliferation of activated hepatic stellate cells (HSC) is an important event in the development of hepatic fibrosis. Insulin-like growth factor-1 (IGF-1) has been shown to be mitogenic for HSC, but the intracellular signaling pathways involved have not been fully characterized. Thus, the aims of the current study were to examine the roles of the extracellular signal-regulated kinase (ERK), phosphatidylinositol 3-kinase (PI3-K) and p70-S6 kinase (p70-S6-K) signaling pathways in IGF-1- and platelet-derived growth factor (PDGF)-induced mitogenic signaling of HSC and to examine the potential crosstalk between these pathways. Both IGF-1 and PDGF increased ERK, PI3-K and p70-S6-K activity. When evaluating potential crosstalk between these signaling pathways, we observed that PI3-K is required for p70-S6-K activation by IGF-1 and PDGF, and is partially responsible for PDGF-induced ERK activation. PDGF and IGF-1 also increased the levels of cyclin D1 and phospho-glycogen synthase kinase-3beta. Coordinate activation of ERK, PI3-K and p70-S6-K is important for perpetuating the activated state of HSC during fibrogenesis.

Inhibition of erythrocyte (RBC) promotion of platelet reactivity could improve the antiplatelet effect of aspirin (ASA). We tested different ASA regimens for optimal inhibition of platelets and the effects of RBC in patients with a history of vascular diseases. Collagen-induced platelet activation (14C-5HT, TXA2 release) and platelet recruitment (proaggregatory activity of cell-free releasates from activated platelets) were measured in PRP, platelet-RBC (Hct 40%), and whole blood (WB) in 206 patients initially on 200-300-mg ASA/day. Their regimen was modified to biweekly 500 mg (loading dose, L) plus daily or twice-daily low-dose ASA (50 or 100 mg). TXA2 was inhibited with all regimens. Percentage of patients with suboptimal inhibition of platelet recruitment in WB was 200-300 ASA/day (41%), L-50/day (87%), L-100/day (58%), L-50/twice-daily (39%), and L-100/twice-daily (20%; P < 0.05 vs other regimens). 14C-5HT release was inhibited to the greatest extent with L-100/twice-daily in PRP + RBC or WB (P < 0.05 vs other regimens) due to greater inhibition of the RBC prothrombotic effect. Compared with other ASA regimens, L-100 twice-daily (equivalent to 221-mg ASA/day in the 14-day cycle), reduced by >50% the proportion of patients with suboptimal inhibition of platelet recruitment in WB and inhibited 14C-5HT release to the greatest extent.