Elevated urinary excretion of the C5b-9 complex in membranous nephropathy.

In experimental membranous nephropathy, antibody binding to glomerular epithelial cell membrane antigens results in complement activation and formation of complement C5b-9 membrane attack complexes in glomeruli. During active disease, the C5b-9 complexes are shed into the urine. To test the hypothesis that a similar mechanism might be operative in human membranous nephropathy, we measured urinary excretion of C5b-9 and C5 in 146 proteinuric patients with biopsy-proven glomerular diseases or diabetes mellitus. Urinary excretion of C5b-9 relative to C5 excretion was higher in 40 patients with membranous nephropathy than in 106 patients with proteinuria due to non-membranous glomerulonephritis when analyzed by covariance analysis (P less than 0.0002). Urinary C5b-9 excretion was higher in membranous nephropathy than in membranoproliferative glomerulonephritis (N = 13, P less than 0.05), minimal change-focal sclerosis (N = 33, P less than 0.001), mesangial proliferative glomerulonephritis (N = 9, P less than 0.02) and IgA nephropathy (N = 7, P less than 0.025). Urinary C5b-9 excretion was also higher in patients with lupus nephritis (N = 18, P less than 0.02) compared to those with non-membranous glomerulonephritis. The lupus patients with the highest excretion had clinical or pathological features of membranous nephropathy. Nine patients with membranous nephropathy and elevated urinary C5b-9 excretion had a shorter duration of disease (P less than 0.05), lower serum creatinine levels (P less than 0.05) and more proteinuria (P less than 0.02) than the 31 membranous nephropathy patients with normal values.(ABSTRACT TRUNCATED AT 250 WORDS)     

Renal C3 synthesis in idiopathic membranous nephropathy: correlation to urinary C5b-9 excretion

Renal C3 synthesis in idiopathic membranous nephropathy: Correlation to urinary C5b-9 excretion. BACKGROUND: Complement activation plays a central pathogenetic role in idiopathic membranous nephropathy (IMN). Urinary excretion of C5b-9 correlates to the immunologic activity of this disease. Recently, renal cortical C3 gene expression has been described in several nephropathies. METHODS: The aim of this study was to investigate the renal C3 gene expression by in situ hybridization in IMN and to correlate it with histopathologic, pathophysiologic, and immunologic (urinary C5b-9) indices of disease activity. RESULTS: C3 was expressed in 77% of 22 renal biopsies of IMN patients, mainly at the cortical tubular and glomerular parietal epithelial cell levels. C3 protein synthesis by tubular cells was demonstrated by immunofluorescence. The intensity of C3 gene expression by both glomerular and tubulointerstitial compartments correlated with the glomerular stage of disease (P = 0. 0023 and P = 0.0214, respectively). Although no correlation was found with proteinuria, serum creatinine at renal biopsy time was strongly associated with renal C3 expression. IMN patients showed a trend of increased urinary C5b-9 levels, which correlated to C3 at the tubulointerstitial level (P = 0.0143). CONCLUSION: Renal C3 production, mainly at the tubular level, may be induced by urinary excretion of C5b-9 in IMN and may have a pathogenetic role in the tubulointerstitial damage that can be associated with this disease.     

IgA nephropathy in patients with congenital C9 deficiency.

The clinical, histologic, and immunopathological findings of three young Japanese males with congenital C9 deficiency and primary IgA nephropathy are reported. The C9 deficiency was discovered either through mass complement screening, or when low hemolytic activity for CH50 and normal C3 levels were detected in plasma. Hematuria and proteinuria were detected at the age of 8 or 9 years as a result of annual urinary screening tests for school children. Renal biopsy showed focal and segmental mesangial proliferation with small epithelial crescents in one patient, and mild, diffuse mesangial proliferation in two. IgA and C3 were deposited predominantly in the mesangial area, and staining for C9 was negative in these patients. Electron microscopy revealed electron dense deposits predominantly in mesangial and paramesangial zones. Immunohistochemical staining in renal biopsy tissues from two patients showed mesangial staining for C5, C8, and S-protein, but staining for C5b-9 neoantigen was completely negative. These results show that the formation of C5b-9 complex is not essential for the induction of human IgA nephropathy, and also for the proliferation of mesangial and even parietal epithelial cells.     

Urinary excretion of C5b-9 reflects disease activity in passive Heymann nephritis

Passive Heymann nephritis (PHN) is a model of membranous nephropathy in rats in which glomerular injury is mediated by the terminal C5b-9 membrane attack complex of complement. This model has been shown to be associated with markedly elevated urinary excretion of C5b-9, compared to other experimental models of glomerulonephritis To determine if urinary C5b-9 excretion could serve as an index of disease activity by correlating with the formation and quantity of glomerular subepithelial immune deposits in PHN, we measured urinary excretion of C5b-9 in PHN under several experimental conditions. In the heterologous phase a direct correlation was demonstrated between levels of urinary C5b-9 excretion and the amount of anti-Fx1A IgG deposited in glomeruli (r = 0.85). In the autologous phase, C5b-9 excretion correlated with the amount of deposit forming antibody present in the serum and resolved when antibody disappeared, despite persistence of glomerular deposits of antigen, antibody, C5b-9 and heavy proteinuria. Glomerular C3 deposits paralleled urinary C5b-9 excretion. Re-initiation of active deposit formation by a second injection of anti-Fx1A produced new C3 deposits and a marked rise in C5b-9 excretion. Finally, complete abrogation of deposit formation by transplanting PHN kidneys into normal recipients also halted C5b-9 excretion. Our findings demonstrate that urinary excretion of C5b-9 is a sensitive index of on-going immunologic disease activity in the PHN model of membranous nephropathy.     

补体旁路途径过度活化在恶性高血压肾硬化中的作用

目的研究补体旁路途径(alternative complement pathway)过度活化在恶性高血压肾硬化中的作用。方法(1)选取本院经肾穿刺活检证实为恶性高血压肾硬化患者50例为病例组,零点行肾穿刺活检的供肾者25例为正常对照组,采用酶联免疫吸附法(ELISA)检测血浆及尿液中的补体旁路途径活化起始B因子、正向调节P因子、负向调节H因子及补体活化终末产物C3a、C5a水平。(2)免疫组化法检测补体活化终末产物C5b-9、C4d及凝集素途径活化产物甘露糖结合凝集素(MBL)在肾活检组织的沉积;免疫荧光双染检测C5b-9与CD34(内皮细胞标志物)在小动脉内皮及肾小球毛细血管内皮的沉积。结果(1)恶性高血压肾硬化患者血浆及尿液中补体B因子、P因子、C3a及C5a均高于正常对照组(均P<0.05),而H因子则低于正常对照组(P<0.05)。(2)恶性高血压肾硬化患者血浆中补体P因子与24 h尿蛋白量呈正相关(rs=0.465,P=0.001),而补体B因子、H因子、C3a、C5a与血肌酐及24 h尿蛋白量无明显相关性。恶性高血压肾硬化患者尿B因子/尿肌酐、尿P因子/尿肌酐、尿C3a/尿肌酐与血肌酐均呈正相关(rs=0.483,P<0.001;rs=0.352,P=0.012;rs=0.319,P=0.024),尿H因子/尿肌酐与血肌酐及24 h尿蛋白量均呈负相关(rs=-0.299,P=0.035;rs=-0.342,P=0.015),尿C5a/尿肌酐与血肌酐及24 h尿蛋白量均呈正相关(rs=0.525,P<0.001;rs=0.496,P<0.001)。(3)免疫组化显示,恶性高血压肾硬化患者C5b-9沉积于小动脉壁及肾小球毛细血管壁,而正常对照组肾组织中未见沉积。恶性高血压肾硬化患者肾脏C5b-9沉积强度评分与血肌酐及24 h尿蛋白量呈正相关(rs=0.791,P<0.001;rs=0.345,P=0.014)。双重免疫荧光标记法可见C5b-9、CD34沉积于小动脉内皮及肾小球毛细血管内皮。(4)恶性高血压肾硬化患者血浆中B因子与C3a(r=0.331,P=0.022)、P因子与C5b-9评分(rs=0.300,P=0.034)均呈正相关;尿液中补体旁路途径活化B因子与C3a、C5a及C5b-9均呈正相关(rs=0.311,P=0.028;rs=0.465,P=0.001;rs=0.428,P=0.002),P因子与C3a、C5a也均呈正相关(rs=0.307,P=0.030;rs=0.442,P=0.001)。恶性高血压肾硬化患者免疫组化可见C4d沉积于小动脉及肾小球,而未见凝集素途径活化产物MBL沉积。结论补体旁路途径过度活化可能参与恶性高血压肾硬化的发生。恶性高血压肾硬化严重程度与补体旁路途径的活化水平相关。     

Complement activation in IgA nephropathy

Activation of alternative complement pathway is presumed to be important pathogenically in IgA nephropathy since renal biopsies usually exhibit glomerular deposition of C3 and P (properdin). Surprisingly, little is known about plasma complement activation in this disease, and the plasma C3 and C4 concentrations are usually normal or increased. We quantitated C3 activation in 202 plasmas from 81 patients with IgA nephropathy using a sensitive new assay that detects a neoantigen [iC3b-C3d neoantigen) which appears when C3b is inactivated to iC3b, C3dg, or C3d. This assay accurately quantitates small amounts of in vivo C3 activation. The concentration of iC3b-C3d neoantigen in plasma was significantly increased, indicating C3 activation in 37% of the pediatric and 57% of the adult plasmas assayed. When data from serial determinations in the patients were analyzed, 75% of the adult and 57% of the pediatric patients had C3 activation on at least one occasion. Classical pathway activation, quantitated by C4 activation was found in 20% of the adult and 5% of the pediatric plasmas. No association was found between elevated iC3b-C3d neoantigen concentration and history of macroscopic hematuria, chronic renal insufficiency or degree of proteinuria. These studies show that complement activation can frequently be detected in the plasma of IgA nephropathy patients. However, the pathophysiologic significance of this complement activation remains to be determined.     

Immunohistochemical study of the C5b-9 complex of complement in human kidneys

The presence and localization of the C5b-9 neoantigens of the terminal complement sequence, of antigens expressed by cleavage fragments of C3, and of Factor H antigens have been studied by immunohistochemical techniques in morphologically normal adult human kidneys and in biopsy specimens from patients with a wide range of renal diseases with and without immune deposits. In morphologically normal kidneys, C5b-9 neoantigens were observed within all connective matrices (arteriolar media, glomerular basement membrane (GBM), mesangial matrix and tubular basement membrane). The C3d and C3g antigens of the C3dg, and C3bi cleavage fragments of C3 and Factor H antigens were found in similar locations. None of the matrices stained for immunoglobulins. Immunoelectron microscopy demonstrated that C3d, C3g, H antigens and the C5b-9 neoantigens were localized on membranous and vesicular structures embedded in the connective matrices. These structures represent cell membranes shed from adjacent cells as evidenced by their ultrastructural appearance and by the fact that those which were in close vicinity to pedicles within the GBM expressed the C3b receptor antigen, a specific marker for podocyte membranes. Formation of C5b-9 complexes in the shielded environment of connective matrices may explain their persistence over long periods of time in the absence of apparent immunopathological consequences. Biopsies from pathological kidneys were classified into three groups based on the pattern of glomerular staining with anti-C5b-9 antibodies. In the first group, a sparse mesangial labeling was seen, similar to that observed in normal kidneys. In the second group, abundant clusters of C5b-9 were seen in the same location as immune deposits. Activation of the complement system to completion could be documented in the absence of detectable C3 (C3c) antigen in glomeruli. Immunoelectron microscopy demonstrated that C5b-9 neoantigens were present on cell remnants in connective matrices in all specimens that were studied. Labeled cell remnants were present in large amounts in sclerotic matrices. C5b-9 neoantigens were constantly found on old and large immune deposits, and absent or occasionally present on recent and small immune deposits. In membranous nephropathy stage I, proteinuria appeared to be independent of the presence or absence of detectable C5b-9 neoantigens on immune deposits. Thus, the presence of C5b-9 neoantigens in pathological renal tissue does not have an univocal significance, and requires analysis of the localization of the antigens and appropriate controls in order to assess the potential role of C5b-9 in tissue damage.     

Increased urinary excretion of C5b-9 distinguishes passive Heymann nephritis in the rat

Increased urinary excretion of C5b-9 distinguishes passive Heymann nephritis from other forms of experimental glomerulonephritis in the rat. In the passive Heymann nephritis (PHN) model of membranous nephropathy (MN) subepithelial deposits form from anti-Fx1A antibody reacting with antigen expressed on the glomerular epithelial cell membrane followed by membrane patching and shedding of immune complexes. Immune complex deposits are accompanied by deposits of C5b-9 which is required for the mediation of proteinuria. We tested the hypothesis that C5b-9 assembly on the epithelial cell membrane might result in C5b-9 excretion in the urine, which would distinguish this autoimmune mechanism of MN from other processes that result in subepithelial immune complex deposits. Using monoclonal antibodies developed to rat C6 and a rat C5b-9 neoantigen, in a sensitive ELISA assay, elevated urinary excretion of rat C5b-9 was documented in PHN associated with on-going glomerular immune deposit formation. No urinary C5b-9 was detectable in MN induced by an exogenous antigen (cationized IgG) despite equivalent glomerular C5b-9 deposits, or in models of nephrotoxic nephritis, subendothelial immune complex nephritis, anti-mesangial cell membrane antibody-induced nephritis or two non-immune nephropathies. Infusion of preformed C5b-9 in proteinuric animals excluded glomerular filtration of C5b-9 as a contributing mechanism to urinary C5b-9 excretion. We conclude that in the rat, increased urinary excretion of C5b-9 is a marker of MN induced by antibody to a glomerular epithelial cell antigen. Urine C5b-9 excretion reflects active glomerular immune deposit formation and distinguishes MN induced by this mechanism from other forms of MN as well as from other glomerular diseases with equivalent glomerular C5b-9 deposits.     

Antiproteinuric effect of angiotensin-converting enzyme inhibition and C5b-9 urinary excretion in membranous glomerulonephritis

Background: Angiotensin-converting enzyme (ACE) inhibitors have an antiproteinuric effect in membranous glomerulonephritis (MGN). However, no studies hae investigated whether this antiproteinuric effect is influenced by urinary C5b-9 excretion, a marker of immunological activity in this disease. Methods: Eleven patients with biopsy-proven MGN were treated with captopril for 8 weeks. The evolution of several clinical and biochemical parameters, including 24-h urinary protein excretion was evaluated every 4 weeks. Urinary C5b-9 excretion was measured at the onset and at the end of captopril treatment. Results: Patients with MGN had significantly higher C5b-9 excretions than a group of 14 healthy controls (89±23 vs 3.7±1.4 ng/mg UCr; P<0.001). A significant correlation was found between urinary C5b-9 and the magnitude of proteinuria, both at the onset and at the end of treatment. After 8 weeks of captopril treatment, proteinuria had decreased from 8±1.8 to 5.2±1.3 g/day (P<0.05). Four weeks after captopril discontinuation, proteinuria rose to 7.3±1.7 g/day (P<0.05). A marked variability in the antiproteinuric response was observed, ranging from 0 to 85% with respect to baseline values. No correlation between decrease in proteinuria and baseline urinary C5b-9 levels was observed. Several patients with elevated urinary C5b-9 levels had captopril-induced decrease in proteinuria. Conclusions: ACE inhibition induces an antiproteinuric effect in patients with MGN. The urinary C5b-9 excretion does not predict the magnitude of this response.     

Evaluation of urinary factor H excretion in patients with idiopathic membranous nephropathy

BACKGROUND: The complement system plays an important role in renal pathogenesis, and C5b-9, a terminal complement complex, is regarded as the principal mediator of proteinuria in idiopathic membranous nephropathy(MN). Since factor H regulates complement activation at the C3 step and is a crucial factor in complement-mediated tissue injury, the urinary excretion of factor H in patients with idiopathic MN was investigated. METHODS: Seven patients with biopsy-proven idiopathic MN were studied for twenty-four weeks. Urinary factor H levels were measured by ELISA from regularly collected urine samples, and then evaluated and compared with assays of urinary protein and C5b-9 excretion. RESULTS: During the study, five patients were treated with steroid therapy. All seven patients maintained stable renal function and showed a decline in urinary protein excretion. The mean level of urinary factor H was markedly elevated (156.1 +/- 47.1 U/mg U-Cr) before treatment (0 week), and gradually declined to 127.2 +/- 43.5 U/mg U-Cr at 12 weeks, and to 64.7 +/- 26.9 U/mg U-Cr) at 24 weeks. This followed decreases in urinary protein and urinary C5b-9 excretion. Percent change in urinary factor H level significantly decreased 24 weeks after treatment without affecting the plasma factor H level. CONCLUSION: These results suggest that factor H contributes to the regulatory mechanism of in situ complement activation, and thus the study of urinary factor H levels, as well as urinary C5b-9, may be significant in idiopathic MN.     

Activation of complement in IgA nephropathy

Considerable evidence supports a role for the complement system in the pathogenesis of IgA nephropathy (IgAN). The alternative pathway components C3 and properdin (P) and the membrane attack complex (C5b-9) are generally found in the mesangial deposits in IgAN, while the classical pathway components C1q and C4 are usually absent. This pattern of immunofluorescence staining for complement components suggests activation of the alternative and terminal pathways in most patients. Despite normal serum concentrations of C3 and other complement proteins, fragments generated by activation of C3, including iC3b, C3d, and iC3b-C3d neoantigen, and sometimes C4, are often detected in plasma. We found that the severity of the histologic changes in the renal biopsy specimens correlated with plasma iC3b-C3d neoantigen concentrations as measured by an enzyme-linked immunosorbent assay. However, no other clinical feature correlated with the plasma concentrations of this neoantigen.     

Immunohistochemical analysis of C3 cleavage fragments, factor H, and the C5b-9 terminal complex of complement in de novo membranous glomerulonephritis occurring in patients with renal transplant

Fifteen renal biopsies from 13 transplanted patients with de novo membranous nephropathy (DNMN) were investigated by immunofluorescence for the presence of C5b-9 neoantigens of the terminal sequence of complement and for antigens expressed by C3 cleavage fragments. DNMN lesions were classified as stage I, II or III upon light and electron microscopy examination. Seven biopsies were classified as stage I DNMN and 8 stage II-III. All patients were proteinuric. In six biopsies with stage I DNMN, staining for C5b-9 neoantigens was restricted to a fine granular labeling in mesangial areas which was analogous to that seen in normal kidneys in contrast with extensive parietal labeling for IgG, C3d and factor H antigens. In eight biopsies with stage II-III DNMN, the pattern of staining with anti-C5b-9 neoantigens antibodies was similar to that obtained with anti-IgG, anti-C3d and anti-factor H antibodies. These results suggest that in situ activation of the whole complement sequence throughout C5b-9 only occurs on large immune deposits (stage II-III DNMN).     

The temporal relationship between urinary C5b-9 and C3dg and clinical parameters in human membranous nephropathy

We have previously reported that urinary excretion of the complementactivation products C3dg and C5b–9 in human membranousnephropathy (MN) correlated with clinical outcome in a cross-sectionalanalysis. We report here the results of a retrospective longitudinalstudy of the temporal relationship between urinary C3dg andC5b–9 excretion and clinical parameters. A group of 23adult patients with biopsyproven MN were studied over a meantime period per patient of 3.5 years. Freshly voided urine sampleswere collected regularly; C3dg and C5b–9 were measuredby ELISA (mean number of samples per patient=13). During the period of the study, nine patients with decliningrenal function (group A) were treated with a standard steroidregimen. Serum creatinine had improved or stabilized in sevenof these patients at the end of treatment. All nine patientswere excreting C3dg and C5b–9 before treatment. Six otherpatients with declining renal function (group B) were not treatedwith steroids because of clinical contraindications. Serum creatininecontinued to increase during the study in four of these sixpatients. C3dg and C5b–9 were present in the urine samplesof these six patients on the majority of dates tested. Eight patients maintained stable renal function during the study(group C), either normal (6 patients) or impaired (2 patients).Of these patients, six were consistently negative for urinaryC3dg and C5b–9 despite persistent proteinuria, and onepatient who was initially positive became negative within 15months, and remained negative for the rest of the study period.One patient was positive on one of 12 occasions tested. Theseresults suggest that urinary complement activation productsindicate ongoing active glomerular damage and may prove to beimportant determinants for the introduction and monitoring oftherapy.     

Measurement of SC5b-9 in urine in patients with the nephrotic syndrome.

In passive or active Heymann nephritis (HN) in the rat, the immune complexes that form in the glomerular subepithelial space result in complement activation and the urinary (U) excretion of S protein-membrane attack complex (SC5b-9, MAC). Because of the similarities between HN in rats and membranous nephropathy (MN) in humans, it has been suggested that measurement of SC5b-9 in urine (UMAC) could be useful in assessing the immunologic activity of MN in patients. The present study was undertaken in normal individuals and in patients with nephrotic syndrome to determine: 1) the conditions of urine collection and preservation needed for accurate measurement of UMAC for clinical purposes; and 2) whether UMAC levels are a sensitive and/or specific test for MN. In studies conducted on urine specimens from patients with increased UMAC levels, we found that UMAC in freshly voided urine was stable for at least three hours at 37 degrees C, with or without the addition of the enzyme inhibitors that were used to stabilize UMAC levels in the studies of HN in the rat. Urine pH, leukocytes and erythrocytes, over the ranges usually encountered, did not influence UMAC levels. However, freezing urine at -70 degrees C artifactually raised UMAC levels (1500 +/- 550 to 1800 +/- 580 SE ng/ml, P less than 0.001 by paired t-test). Normal urine contained low UMAC levels: 80 +/- 3 ng/mg urinary creatinine (UCr). By contrast, patients with glomerulopathies tended to have elevated UMAC levels: 18 of 38 patients had levels that ranged from 200 to 20,000 ng/mg UCr.(ABSTRACT TRUNCATED AT 250 WORDS)     

Renal ultrasound elastography can reflect clinical-pathological changes in chronic kidney disease patients

Objective To analyze how is the elastography of renal tissue correlated to clinical biochemical indexes and pathological changes in patients with chronic kidney disease (CKD), and to explore the potential of renal elastography to become a new noninvasive method available for the dynamic monitoring of renal disease progression, as well as its efficacy assessment and prognosis evaluation. Methods Patients admitted to the department of nephrology of the First Affiliated Hospital of China Medical University and received renal biopsy from August 2014 to January 2015 were selected. One hundred and thirteen cases of CKD patients, 61 males and 52 females were enrolled, including 23 cases of IgA nephropathy, 39 cases of membranous nephropathy, 15 cases of minimal change nephropathy and 7 cases of focal segmental glomerulosclerosis. The Young modulus of renal cortex and medulla (YMcortex and YMmedulla) were detected by Aix Plorer type full digital color Doppler ultrasound. The correlations between the YMs and clinical biochemical indicators in blood and urine, and the difference of YMs among different pathological changes in patients with CKD were analyzed by statistics. Results The YMcortex and YMmedulla in CKD patients were higher than those in the control group (all P＜0.05); and with the progression of CKD, the YMcortex and YMmedulla gradually increased. The YMcortex in CKD G5 patients was higher than that in CKD G1-3 patients (all P＜0.05). The YMmedulla in CKD G3-5 patients was higher than that in CKD G1-2 patients (all P＜0.05). The YMcortex was correlated with systolic pressure, serum creatinine, cystatin C, serum albumin, serum phosphorus, calcium and phosphorus product, uric acid, intact parathyroid hormone (iPTH), urinary NAG, estimate glomerular filtration rate (eGFR) and hemoglobin (all P＜0.05). The YMmedulla was correlated with systolic pressure, serum creatinine, serum albumin, uric acid, iPTH, urine microalbumin (MA), urinary NAG and hemoglobin (all P＜0.05). Serum cystatin C (β=0.485, P=0.018) and uric acid (β=0.418, P=0.039) were independently correlated with the YMcortex. Serum creatinine (β=0.380, P=0.019), uric acid (β=0.482, P=0.004) and smoking (β=0.337, P=0.009) were independently correlated with YMmedulla. The YMcortex and YMmedulla in different pathological types were statistically significant (P＜0.001, P=0.003). The YMcortex and YMmedulla in patients with membranous nephropathy and IgA nephropathy were higher than those in the patients with minimal change nephropathy (all P＜0.05). The YMmedulla in patients with focal segmental glomerulosclerosis was higher than that in the patients with minimal change nephropathy (P＜0.05). The YMcortex in the patients with phases Ⅳ and Ⅴ based on the Lee grading system of IgA nephropathy was higher than that in the patients with phases Ⅱ and Ⅲ (P＜0.05). According the Oxford classification for IgA nephropathy, the YMcortex and YMmedulla in the T1 and T2 patients were higher than those in the T0 patients (P＜0.05). The YMcortex and YMmedulla showed no statistically significant differences among different stages of membranous nephropathy. Conclusions The YMcortex and YMmedulla are associated with the progress of renal insufficiency, which may become new indicators for determining CKD progression. The renal ultrasound elastography may become a new non-invasive method for early diagnosing CKD, dynamic monitoring disease progression, and assessing efficacy and prognosis.     

Complement (C5b-9) induces glomerular epithelial cell DNA synthesis but not proliferation in vitro.

BACKGROUND: The C5b-9 membrane attack complex of complement is the principal mediator of injury induced experimentally by antibodies directed at glomerular cell membranes. In experimental membranous nephropathy, C5b-9 induced injury to the glomerular visceral epithelial cell (VEC) is associated with DNA synthesis, but not cytokinesis. In the current study we determined if C5b-9 increases DNA synthesis in VEC in vitro, and defined the mechanisms involved. METHODS: Rat VEC in vitro were divided into three groups: (1) sensitized with anti-VEC antibody and exposed to sublytic concentrations of C +/PVG serum (normal complement components); (2) anti-VEC antibody and control C-/PVG serum (C6 deficient); (3) no anti-VEC antibody. DNA synthesis (BrdU staining), mitosis (mitotic figures) and cytokinesis (cell counts) were measured at 24 and 48 hours. To examine the expression of specific S-phase and M-phase cell cycle regulatory proteins and their inhibitors, immunostaining and Western blot analysis was performed for cyclin A, CDK2, p21 and p27, cyclin B and cdc2. RESULTS: In the absence of growth factors, sublytic C5b-9 attack did not increase proliferation. In contrast, sublytic C5b-9 attack (group 1) augmented growth factor induced DNA synthesis by 50% compared to controls (groups 2 and 3; P < 0.001), and was accompanied by increased levels of cyclin A and CDK2, and a decrease in the cyclin kinase inhibitor p27 (but not p21). Sublytic C5b-9 attack reduced the expression of the M phase cell cycle proteins, cyclin B and cdc2, accompanied by reduced mitosis (mitotic figures) and cytokinesis (cell number). CONCLUSIONS: Our results show that the C5b-9 augmented growth factor entry into the S phase in VEC is regulated by changes in specific cell cycle regulatory proteins. However, antibody and complement decreased the M phase cell cycle proteins, and prevented VEC mitosis and cytokinesis, suggesting a delay or arrest at the G2/M phase.     

C6 depletion reduces proteinuria in experimental nephropathy induced by a nonglomerular antigen.

The role of the complement membrane attack complex, C5b-9, in mediating glomerular injury has been well defined in models of membranous nephropathy induced by antibody to endogenous glomerular epithelial cell membrane antigens. The effect of selective C6 depletion (to prevent C5b-9 formation) on morphologic characteristics and proteinuria in a model of in situ subepithelial immune complex nephritis induced by an exogenous cationized antigen (human immunoglobulin G (IgG)) followed by rabbit antibody to human IgG was studied. Selective C6 depletion was achieved by repeated administration of a goat antibody to rat C6. Other groups were treated with cobra venom factor to induce generalized complement depletion and with sublethal irradiation to deplete circulating leukocytes. In C6-depleted rats, C6 levels were reduced to less than 3% of baseline throughout the 2 days of the study compared with over 100% in controls. At 4 h after disease induction, glomerular deposition of antigen and antibody were similar in C6D and control groups by immunofluorescence and by direct measurement of glomerular deposition of radiolabeled antigen and antibody (cationized 131I human IgG, 9.1 +/- 0.1 micrograms/38,000 glomeruli in C6D versus 9.8 +/- 0.9 in controls; P = was not significant; rabbit 125I-labeled anti-human IgG, 104 +/- 10 ng in C6D versus 80 +/- 9 ng in controls; P = was not significant). Circulating C3 levels and glomerular C3 deposition were also similar in C6D and control groups.(ABSTRACT TRUNCATED AT 250 WORDS)     

Polymorphs infiltrate glomeruli in mesangial IgA glomerulonephritis

During episodes of macroscopic hematuria, patients with IgA nephropathy commonly have polymorphonuclear neutrophils (PMNs) as well as fibrin and mononuclear cells in glomerular capillaries. We quantitated PMN and macrophage infiltration in glomeruli of 54 patients with IgA nephropathy in whom renal biopsies were obtained within 30 days of macroscopic hematuria. Control biopsies (N = 22) were from patients with IgA nephropathy and urinary erythrocyte counts below 50,000/ml. PMN monocyte/macrophages were quantitated using the monoclonal antibodies FMC10 and HAM56, respectively. In patients with heavy hematuria, 45.9 +/- 3.4% (mean +/- SE) of glomeruli were positive for PMNs (control 10.5 +/- 2.8%, P = 0.001) with a mean PMN count/glomerulus of 1.10 +/- 0.20 (control 0.13 +/- 0.03, P = less than 0.001). 65.6 +/- 9.7%. Of the glomeruli were positive for monocytes in the heavy hematuria group (control 40.0 +/- 8.4, P less than 0.05) with the mean monocyte count per glomerulus being 1.6 +/- 0.2 (control 0.6 +/- 0.1, P less than 0.01). We conclude that, in the acute phase of mesangial IgA nephropathy, PMN and monocytes are present and presumably participate in glomerular injury.     

Complement activation products in the urine from proteinuric patients

The presence of plasma proteins in the tubular lumen has variety of adverse effects on the tubular cells. Among various plasma proteins filtered through glomerular barrier, complement has been proven as the possible candidate inducing tubulointerstitial injury. To study the role of intratubular complement activation in proteinuric patients, complement activation products (CAP) at C3 level (iC3b and Bb) and C9 level (membrane attack complex) were measured in both plasma and urine of patients with minimal change nephrotic syndrome (MCNS), focal glomerular sclerosis, IgA nephropathy, membranous nephropathy, and diabetic nephropathy. For evaluation of the effect of metabolic acidosis on the intratubular complement activation, urinary CAP were measured before and after sodium bicarbonate administration in patients with renal insufficiency. The following results were obtained: (1) Patients with focal glomerular sclerosis and diabetic nephropathy showed the highest level of urinary CAP excretion rate (unit/creatinine), while MCNS revealed no increase. (2) Patients with membranous nephropathy showed a unique finding, i.e., isolated increase of membrane attack complex excretion. (3) There was no significant correlation between urine and plasma levels of CAP. (4) Except for MCNS patients, the urinary excretion rate of CAP significantly increased when the level of proteinuria exceeded the nephrotic range, and it was significantly correlated with the serum creatinine level. (5) Urinary CAP excretion rate significantly decreased 2 wk after sodium bicarbonate administration without affecting the level of proteinuria or plasma CAP. These results suggest that the degree of intratubular complement activation correlates with the level of proteinuria, type of glomerular disease, impairment of renal function, and metabolic acidosis.     

Immunoglobulin A subclass-containing plasma cells in the jejunum in primary IgA nephropathy and in Henoch-Sch?nlein purpura

We studied the distribution of IgA1- and IgA2-containing plasma cells in the jejunum of 5 patients with primary IgA nephropathy and of 2 patients with Henoch-Sch?nlein purpura. The percentage of IgA-containing cells (62.3 +/- 4.6) was below the level found in normal (79.4 +/- 5.1) (p less than 0.001). The percentage of IgA1-containing plasma cells relative to the total IgA-containing cells was too low in 2, normal in 3 and too high in 2 subjects (mean percentage 68.1 +/- 9.6; normal 69.0 +/- 3.8). Our data do not support an increased production of IgA1 in the jejunal mucosa of patients with primary IgA nephropathy or with Henoch-Sch?nlein purpura.