Down-regulation of PTH/PTHrP receptor in the kidney of patients with renal impairment.
(25/2477)
OBJECTIVE: To investigate whether the down-regulation of renal parathyroid hormone/parathyroid hormone related protein (PTH/PTHrP) receptor messenger ribonucleic acid (mRNA) expression is a general phenomenon in patients with different stages of renal disease, besides chronic renal failure. METHODS: Twenty-five patients were divided into the following groups: (1) chronic glomerulonephritis with normal renal function (GNN) (2) chronic glomerulonephritis with moderate renal insufficiency (GNI) (3) severe chronic renal failure undergoing maintenance dialysis (CRF) (4) acute renal failure during oliguric phase (ARF) (5) normal control without renal suffering (NC). Using relatively quantitative reverse transcription/polymerase chain reaction (RT/PCR) method, we investigated PTH/PTHrP receptor mRNA expression in renal specimens obtained through biopsy or operation. The levels of plasma C-terminal PTH, serum phosphorus and calcium were also observed at the same time. RESULTS: Plasma C-terminal PTH levels in GNI, CRF and ARF patients were 1.90, 9.73 and 8.63 times higher than those in NC. However, the difference between GNN and NC was insignificant. CRF and ARF patients also presented obviously elevated serum phosphorus (1.61, 1.86 vs 1.14 mmol/L) and reduced serum calcium (1.82, 1.71 vs 2.26 mmol/L) compared with that in the control. These two parameters for GNN and GNI patients were normal. The levels of PTH/PTHrP receptor mRNA (corrected by beta-actin mRNA) in the kidney of GNN, GNI, CRF and ARF patients was markedly decreased by up to 35.7%, 68.5%, 77.9% and 92.2%, respectively. CONCLUSIONS: The down-regulation of renal PTH/PTHrP receptor mRNA occurs much earlier than the changes of renal function, plasma PTH, serum phosphorus and calcium in the course of human renal disease. (+info)
Glomerular deposition of mannose-binding lectin in human glomerulonephritis.
(26/2477)
BACKGROUND: Mannose-binding lectin (MBL), a member of the collectin family, binds to various oligosaccharides and activates the classical pathway of complement independent from C1q. At present it is unknown whether this so-called lectin pathway of complement activation plays a role in the pathogenesis of human glomerulonephritis. METHODS: Direct immunofluorescence of 84 renal biopsies using an MBL-specific monoclonal antibody and antibodies directed against IgG, IgA, IgM, C1q, C3, and terminal complement complex (TCC) was performed. Serum MBL levels of 50 patients were determined by enzyme-linked immunosorbent assay. RESULTS: MBL was detected in the glomeruli of patients with lupus nephropathy (15 of 16), membranous nephropathy (10/15), membranoproliferative glomerulonephritis type I (5/6) and anti-GBM nephritis (2/4). MBL deposition paralleled that of immunoglobulins, C1q, C3, and TCC but was less intense as compared to C1q. Focal segmental deposits of MBL were present in focal segmental glomerulosclerosis (4/6), IgA nephropathy (3/11), amyloidosis AL (1/4), and advanced renal fibrosis (2/2). Here MBL staining was identical to IgM and C3 and considered an unspecific entrapment of MBL in sclerotic lesions in these cases. No significant difference in MBL serum levels was observed between normal controls and patients with lupus nephritis, membranous nephropathy, membranoproliferative glomerulonephritis, focal segmental sclerosis, minimal change disease or IgA nephropathy. In patients suffering from membranous nephropathy with (n=10) or without (n=5) glomerular MBL deposits serum creatinine, C3, C4, serum protein, and proteinuria were not statistically different. CONCLUSION: MBL is present in the glomeruli of patients with glomerulonephritis involving deposition of IgG and activation of the classical pathway of complement. We propose that MBL binds to agalactosyl oligosaccharides of IgG that terminate in N-acetylglucosamine. The extent to which the lectin pathway of complement contributes to overall complement activation in the glomeruli remains unknown, but is likely to be marginal. (+info)
Increased plasma malondialdehyde levels in glomerular disease as determined by a fully validated HPLC method.
(27/2477)
BACKGROUND: Reactive oxygen species and particularly free radical induced lipid peroxidative tissue damage have been implicated in the pathogenesis of various renal diseases. Lipid peroxidation is assessed indirectly by the measurement of secondary products, such as malondialdehyde (MDA), using the widely employed thiobarbituric acid reactive substances (TBARS) method. However, this method lacks sensitivity and specificity. We have therefore developed and validated an HPLC (high-performance liquid chromatography) method for measurement of MDA and applied this to a variety of plasma samples in renal patients. METHODS: The optimized method involves antioxidant treatment of the plasma sample, followed by a protein precipitation step using trichloroacetic acid, acid hydrolysis and formation of an MDA thiobarbituric acid complex. The MDA-(TBA)2 adduct is separated from other interfering compounds by C18 reverse-phase HPLC techniques, with visible detection at 532 nm. RESULTS: The assay was linear over the ranges 0.25-1.0 microM MDA and the detection limit was 0.06 microM MDA. Within-run precision was <4.5% and between-run precision was <10.0%. MDA plasma concentrations (mean+/-SD) were higher in ESRF diabetic patients (0.32 +/- 0.14 microM, n=20), non-diabetic ESRF patients (0.32 +/- 0.09 microM, n=20), and CRF patients (0.14 +/- 0.06 microM, n=40) compared to healthy controls (0.11 +/- 0.03 microM, n=40), (P < 0.001, P < 0.001 and P = 0.008). Levels were similar in healthy controls with normal renal function and transplanted patients (0.12 +/- 0.03 microM MDA, n=40), (P=NS). No correlation was observed between MDA and creatinine levels (r2 = 0.05, n=80), which suggests that MDA does not correlate with the degree of renal impairment. We matched CRF patients with glomerular and non-glomerular causes of renal failure for creatinine levels and found that MDA levels were higher in patients with glomerulonephritis (0.16 +/- 0.06 microM) than in those with CRF from non-glomerular causes (0.12 +/- 0.04 microM, P = 0.002). CONCLUSIONS: We have introduced a reliable and sensitive HPLC technique to enhance the specificity of MDA-(TBA)2 measurement, with a significant improvement in HPLC column life. Using this method, picomole quantities of MDA can be detected in plasma. We have shown that MDA levels are significantly raised in patients with CRF due to glomerulonephritis, regardless of serum creatinine, which suggests that there is oxidative injury independent of any possible MDA retention due to renal impairment. (+info)
L-Arginine supplementation increases mesangial cell injury and subsequent tissue fibrosis in experimental glomerulonephritis.
(28/2477)
BACKGROUND: Mesangial cell lysis in the antithymocyte serum (ATS)-induced model of glomerulonephritis is dependent on the generation of cytotoxic nitric oxide (NO) through transient induction of NO synthase (iNOS). We hypothesized that increased availability of L-arginine (L-Arg) during mesangial cell lysis might provide iNOS with increased substrate leading to increased lysis, and that this increased lysis would be reflected in more severe fibrotic disease at day 6. METHODS: To ensure whole body equilibration with high L-Arg at the time of injury, rats were pretreated with 1% L-Arg in drinking water for one week prior to the administration of ATS. Animals were sacrificed six hours after ATS injection when previous experiments had indicated iNOS induction had occurred and at six days. At six hours, plasma was obtained for L-Arg levels and nitrite/nitrate (NOx) content. Renal tissues were taken for histological evaluation of glomerular cell counts, macrophage infiltration (ED-1), and iNOS expression. Glomeruli were isolated for detection of iNOS mRNA and placed in culture to study the dependence of NO production on L-Arg concentration. In rats sacrificed at six days, L-Arg supplementation was stopped 16 hours after ATS injection. Fibrotic disease was evaluated by urinary protein excretion, histological assessment of glomerular cell number, matrix accumulation, and production of transforming growth factor-beta1 and matrix components fibronectin and plasminogen activator inhibitor type-1 (PAI-1) by isolated glomeruli in culture. RESULTS: At six hours, the glomerular cell number was significantly reduced by ATS injection (P < 0.01) and further significantly (P < 0. 05) reduced by L-Arg feeding [normal control (NC) = 64.2 +/- 1, ATS = 53.4 +/- 0.7, ATS + L-Arg = 50.8 +/- 0.7]. Disease increased macrophage infiltration and iNOS protein and iNOS mRNA levels markedly (P < 0.01), whereas L-Arg feeding did not further increase these variables. Plasma L-Arg levels (nmol/ml) were reduced by disease (NC = 121 +/- 9, ATS = 84 +/- 13, P < 0.01) and elevated by L-Arg feeding (ATS + L-Arg = 166 +/- 12, P < 0.01). Plasma NOx was significantly increased by ATS and further increased by ATS + L-Arg (P < 0.05). Production of NOx by cultured glomeruli showed striking L-Arg concentration dependence in six hours but not in normal glomeruli. In the group sacrificed at day 6, day 2 proteinuria was higher in the ATS + L-Arg group compared with the ATS alone group (P < 0.05). Measures of fibrotic disease at day 6 all showed large increases over control with ATS alone (P < 0.01), and further small, but significant increases when L-Arg was combined with ATS (P < 0.05). CONCLUSIONS: The results indicate that if given during disease induction, L-Arg supplementation can enhance iNOS-dependent tissue injury by providing increased substrate. Although the increase in injury with L-Arg supplementation was small, it led to increased fibrosis at day 6. These data predict that in diseases with repeated iNOS-dependent tissue injury, L-Arg supplementation may produce cumulative increases in tissue fibrosis. (+info)
Heparin-binding epidermal growth factor-like growth factor is expressed in the adhesive lesions of experimental focal glomerular sclerosis.
(29/2477)
BACKGROUND: In this study, we attempted to determine whether heparin-binding epidermal growth factor-like growth factor (HB-EGF) was up-regulated in two chronic models of proteinuria. METHODS: Chronic passive Heymann nephritis (PHN) and puromycin aminonucleoside (PAN) models were induced in Sprague-Dawley rats. HB-EGF expression was studied by Northern blotting, in situ hybridization, and immunohistochemistry. RESULTS: The chronic PAN model was associated with the development of glomerular lesions of focal glomerular sclerosis (FGS), severe interstitial fibrosis, and renal failure. Lesions of FGS were seen in approximately 80% of glomeruli at all time points, with a slight increase in the number of glomeruli showing extensive adhesion between 40 and 90 days. Northern blots of whole kidney tissue showed a 3- to 5.8-fold increased expression of HB-EGF mRNA in the chronic PAN group. Increased mRNA and protein were localized by in situ hybridization and immunohistochemistry to tubules, glomerular epithelial cells (GECs), and cells of Bowman's capsule. HB-EGF mRNA and protein were strongly expressed by epithelial cells involved in the formation of the lesions of FGS. By contrast, in chronic PHN, there was a small increase in HB-EGF, and the extensive lesions of FGS did not develop despite continued, heavy proteinuria. CONCLUSIONS: These data suggest that HB-EGF may contribute to formation of the lesions of FGS, perhaps through stimulation of abortive mitogenesis in GECs or an adhesive interaction between transmembrane HB-EGF and the exposed glomerular basement membrane. (+info)
The cyclin kinase inhibitor p21CIP1/WAF1 limits glomerular epithelial cell proliferation in experimental glomerulonephritis.
(30/2477)
BACKGROUND: During glomerulogenesis, visceral glomerular epithelial cells (VECs) exit the cell cycle and become terminally differentiated and quiescent. In contrast to other resident glomerular cells, VECs undergo little if any proliferation in response to injury. However, the mechanisms for this remain unclear. Cell proliferation is controlled by cell-cycle regulatory proteins where the cyclin-dependent kinase inhibitor p21Cip1,WAF1 (p21) inhibits cell proliferation and is required for differentiation of many nonrenal cell types. METHODS: To test the hypothesis that p21 is required to maintain a differentiated and quiescent VEC phenotype, experimental glomerulonephritis was induced in p21 knockout (-/-) and p21 wild-type (+/+) mice with antiglomerular antibody. DNA synthesis (proliferating cell nuclear antigen, bromodeoxyuridine staining), VEC proliferation (multilayers of cells in Bowman's space), matrix accumulation (periodic acid-Schiff, silver staining), apoptosis (TUNEL), and renal function (serum urea nitrogen) were studied on days 5 and 14 (N = 6 per time point). VECs were identified by location, morphology, ezrin staining, and electron microscopy. VEC differentiation was measured by staining for Wilms' tumor-1 gene. RESULTS: Kidneys from unmanipulated p21-/- mice were histologically normal and did not have increased DNA synthesis, suggesting that p21 was not required for the induction of VEC terminal differentiation. Proliferating cell nuclear antigen and bromodeoxyuridine staining was increased 4.3- and 3.3-fold, respectively, in p21-/- mice with glomerulonephritis (P < 0.0001 vs. p21+/+ mice). At each time point, VEC proliferation was also increased in nephritic p21-/- mice (P < 0.0001 vs. p21+/+ mice). VEC re-entry into the cell cycle was associated with the loss of Wilms' tumor-1 gene staining. Nephritic p21-/- mice had increased extracellular matrix protein accumulation and apoptosis and decreased renal function (serum urea nitrogen) compared with p21+/+ mice (P < 0.001). CONCLUSION: These results show that the cyclin kinase inhibitor p21 is not required by VECs to attain a terminally differentiated VEC phenotype. However, the loss of p21, in disease states, is associated with VEC re-entry into the cell cycle and the development of a dedifferentiated proliferative phenotype. (+info)
Genetic dissection of lupus pathogenesis: a recipe for nephrophilic autoantibodies.
(31/2477)
Sle1 and Sle3 are 2 loci that confer susceptibility to lupus nephritis in the NZM2410 strain of mice. Our previous work has shown that B6.NZMc1 mice, congenic for Sle1, exhibit loss of tolerance to chromatin but do not develop any pathogenic autoantibodies or disease. B6.NZMc7 mice, congenic for Sle3, exhibit low-grade polyclonal B- and T-cell activation, elevated CD4/CD8 ratios, and mildly penetrant glomerulonephritis. In contrast to these monocongenics, the present study reveals that B6.NZMc1|c7 mice, bicongenic for Sle1 and Sle3, exhibit splenomegaly, significantly expanded populations of activated B and CD4(+) T cells, and a robust, variegated IgG autoantibody response targeting multiple components of chromatin (including double-stranded DNA), intact glomeruli, and basement membrane matrix antigens. As one might predict, these mice, particularly the females, exhibit highly penetrant glomerulonephritis. These findings lend strong support to a two-step epistatic model for the formation of pathogenic, nephrophilic autoantibodies in lupus. Whereas loci such as Sle1 may serve to breach tolerance to chromatin, full-blown pathogenic maturation of the autoantibody response appears to require additional input from other loci (such as Sle3) and gender-based factors. (+info)
Urine IgG2/IgG4-ratio indicates the significance of the charge selective properties of the glomerular capillary wall for the macromolecular transport in glomerular diseases.
(32/2477)
BACKGROUND: Alterations of the charge-selective properties of the glomerular capillary wall are important constituents of the pathogenesis of many glomerular diseases. Thus, differences in the degree of such changes could be of help in understanding the mechanisms governing the transport of macromolecules across the glomerular capillary wall. METHODS: The ratio between urine concentrations of neutral IgG2 and negatively charged IgG4 (IgG2/IgG4-ratio) was measured in 150 proteinuric patients and 21 healthy controls. The patients were subdivided into seven biopsy verified diagnostic groups. RESULTS: The study revealed decreased IgG2/ IgG4-ratio in membranous glomerulonephritis (0.57) compared to healthy controls (2.09) and to all other diagnosis groups; crescentic necrotizing glomerulonephritis (1.28), diffuse proliferative glomerulonephritis (1.10), IgA nephropathy (1.11), mesangial proliferative glomerulonephritis (1.55), minimal change nephropathy (1.00), and nephrosclerosis secondary to hypertension (1.06). Although not statistically significant, there was a tendency towards lower IgG2/IgG4-ratio values in all the studied glomerular diseases compared to healthy controls. CONCLUSIONS: Since IgG is transported entirely through the large pores of the glomerular basement membrane decreased IgG2/IgG4-ratio implies that this pathway is strongly influenced by the charge-selective properties of the glomerular capillary wall. The conclusion that could be drawn from that is that the large pore radius must be discrete, in the order of 80-90 A, and thus not non-discriminatory to macromolecules as previously thought. (+info)