Role of apoptosis signal-regulating kinase 1 in complement-mediated glomerular epithelial cell injury

In the rat passive Heymann nephritis (PHN) model of membranous nephropathy, complement C5b-9 activates protein kinases in glomerular epithelial cells (GEC), and induces sublethal GEC injury and proteinuria. Complement induces production of reactive oxygen species (ROS) via the NAPDH oxidase, and stimulates phosphorylation of c-Jun N-terminal kinase (JNK) and p38 kinase in a ROS-dependent manner. In the present study, we demonstrate that apoptosis signal-regulating kinase 1 (ASK1) was activated in glomeruli of rats with PHN, and that incubation of GEC in culture with antibody and sublytic C5b-9 stimulated ASK1 activity. The latter was, in part, mediated via the NADPH oxidase and ROS. Sublytic complement induced JNK and p38 phosphorylation, which was amplified in GEC that stably overexpress ASK1, as compared with Neo (control) GEC. Complement-induced lysis was enhanced in GEC that overexpress ASK1, as compared with Neo, and was attenuated in GEC that overexpress a dominant negative ASK1 mutant. Inhibition of p38, but not JNK, attenuated complement lysis in GEC that overexpress ASK1, but not in Neo GEC. In Neo GEC, generation of ROS restricted complement-mediated GEC injury but the protective effect of ROS was lost when ASK1 was overexpressed. We propose that the level of ASK1 expression determines the functional effect of p38 activation, i.e. when ASK1 is overexpressed, p38 activation is amplified, and C5b-9 assembly leads to GEC injury via ASK1 and p38. The present study thus defines a novel role for ASK1 as a mediator of C5b-9-dependent cell injury.     

The glomerular complement receptor in immunologically mediated renal glomerular injury.

We examined 25 renal-biopsy specimens to determine whether there is a relation between immunologically mediated renal diseases and the activity of complement receptors that selectively bind antigen-antibody complexes containing activated third component of complement (C3b). These receptors have been termed glomerular complement receptors. Renal lesions associated with in vivo deposition were associated with a loss of receptor sites as demonstrated by reduced or absent in vitro binding of C3b-coated test reagents by glomerular complement receptor. These findings suggest that binding of complement containing immune complexes to glomerular complement receptors in human subjects may participate in the immunopathologic processes of certain immune-complex-mediated renal diseases.     

Leukocytes in glomerular injury

Leukocytes of the innate immune system play a central protective role in immune defense to pathogens but may also mediate injurious inflammatory responses resulting in tissue injury. These leukocytes provide the first rapid cellular defense mechanisms through a limited repertoire of rapid pre-programmed responses, but they are also involved in chronic inflammation and tissue repair. They are directed to sites of pathogen challenge and inflammation by a variety of mechanisms and are activated in response to both exogenous and endogenous stimuli. They do not show the capacity of self–non-self discrimination and memory, which are defining characteristics of the adaptive immune system, although macrophages in particular may show some capacity for differentiation of their effector responses. However, they do play an integral role in adaptive immune responses by their capacity to present antigen, modify T-cell development, and function as effectors of adaptive cell-mediated immunity.     

Complement in glomerular injury

In recent years, research into the role of complement in the immunopathogenesis of renal disease has broadened our understanding of the fragile balance between the protective and harmful functions of the complement system. Interventions into the complement system in various models of immune-mediated renal disease have resulted in both favourable and unfavourable effects and will allow us to precisely define the level of the complement cascade at which a therapeutic intervention will result in an optimal effect. The discovery of mutations of complement regulatory molecules has established a role of complement in the haemolytic uremic syndrome and membranoproliferative glomerulonephritis, and genotyping for mutations of the complement system are already leaving the research laboratory and have entered clinical practice. These clinical discoveries have resulted in the creation of relevant animal models which may provide crucial information for the development of highly specific therapeutic agents. Research into the role of complement in proteinuria has helped to understand pathways of inflammation which ultimately lead to renal failure irrespective of the underlying renal disease and is of major importance for the majority of renal patients. Complement science is a highly exciting area of translational research and hopefully will result in meaningful therapeutic advances in the near future.     

The role of T cells in the mediation of glomerular injury in Heymann's nephritis in the rat.

Heymann's nephritis (HN), a rat model of the membranous glomerulonephritis in man, is thought to be mediated by auto-Ig with subsequent activation of C. Whether T cell mechanisms are involved in the mediation of HN, apart from CD4+ cells providing help for auto-Ig production, was examined by treatment with mAb specific for T cell subsets for 6 weeks after immunization to induce HN. Anti-CD4 mAb therapy totally prevented proteinuria, in that at 6, 8, and 12 week treated rats had less than 15 mg/day of protein compared to controls that all had greater than 260 mg/day. Ig and C deposition in the glomerulus was significantly less and auto-Ig titers in serum were partially suppressed by anti-CD4 therapy. Anti-CD8 mAb therapy markedly reduced proteinuria at all time points, for example at 6 weeks there was 51 +/- 40 mg/day compared to 183 +/- 120 mg/day (P = 0.0003), but had no effect on auto-Ig titers or on Ig and C deposition in the glomerulus. A non-specific effect of high dose mouse mAb therapy was excluded by the findings that a mAb that did not bind to rat cells had no effect on the induction of HN and that serum C was not depleted in any of the mAb treated animals. A role for T effector mechanisms was further supported by the finding that therapy with mAb to T cell receptor alpha/beta chain or with cyclosporine also markedly delayed the onset of proteinuria. Examination of renal biopsies showed a T cell infiltrate in glomeruli and the interstitium of the untreated HN controls that was not present in MRC Ox35 or MRC Ox8 treated groups. This infiltrate included CD4+ and CD8+ T cells and macrophages. These results suggest induction of proteinuria in HN was totally dependent upon CD4+ T cells, and that CD4+ and CD8+ cells may have a direct role in the mediation of glomerular dysfunction in HN.     

Membrane attack complex deposition in experimental glomerular injury.

The complement (C) system is an important mediator of glomerular injury both through its attraction of inflammatory cells and by a cell-independent effect on glomerular capillary wall permeability. We have postulated that the latter effect may be mediated by the terminal components of the C system, the membrane attack complex (MAC). We examined several models of immunologic renal injury in the rat by immunofluorescence for the presence of neoantigens of the MAC. Rats with experimental membranous nephropathy induced by antibody binding to a fixed glomerular antigen (passive Heymann nephritis, PHN) or a planted antigen (autologous phase of PHN) had moderate proteinuria and 1-2+ capillary wall deposits of IgG, rat C3, and MAC. C depletion with cobra venom factor (CVF) significantly decreased proteinuria and prevented deposition of C3 and MAC. Rats with active Heymann nephritis had similar capillary wall deposits of MAC. Rats with anti-glomerular basement membrane nephritis developed severe proteinuria which was not affected by CVF treatment and had no glomerular deposits of MAC. Rats with nonimmunologic proteinuria induced by aminonucleoside of puromycin also had no glomerular deposits of MAC. In rats unilaterally nephrectomized before the induction of PHN segmental glomerular sclerosis developed after 6 months with deposits of MAC in the sclerotic areas. The presence or absence of glomerular deposits of MAC in experimental renal disease correlates well with the pathogenetic role of C in the production of injury. These results support a role for the MAC in the mediation of several types of glomerular injury.     

Losartan逆转钙超负荷及对高血压肾的保护作用

目的:探讨外周血细胞内钙浓度与高血压、血管和肾小球病变之间的关系。方法:动物分自发性高血压(SHR)组和正常对照血压(WKY鼠)组,其中SHR组再分为治疗组(予losartan20mg·kg^-1·d^-1)和非治疗组,分别于3、8月龄测量鼠尾动脉压、外周血淋巴细胞内游离钙浓度及电子显微镜下血管、肾脏有关病理指标。结果:与同龄正常对照组比较,早期高血压鼠外周血淋巴细胞内游离钙浓度已增高,病理表现为血管内皮损伤,管腔表面粗糙变形、内皮细胞增殖肥大,而肾小球内未见明显细胞增生改变,但基底膜有轻度增厚和系膜扩张。8月龄时血管、肾小球细胞均明显肥大,且与钙浓度升高呈密切相关。而经losartan治疗后,SHR钙超负荷得到逆转,血压显著下降,血管、肾小球损伤得到修复。结论:高血压钙超负荷现象不但与血压、血管改变相关,而且与肾小球损害也密切相关,并能通过外周血细胞内钙浓度来反映其变化,losartan能逆转高血压鼠的钙超负荷现象,降压同时,有效地保护血管和肾小球。     

Interaction of antibody with Forssman antigen in guinea pigs. A mechanism of adaptation to antibody- and complement-mediated injury.

Forssman antigen is a glycosphingolipid with antigenic specificity determined by extra-membrane haptenic sugars similar to blood group antigens and antigens that are the main barrier to xenogeneic organ transplantation. Herein, we describe the localization of Forssman antigen in guinea pig lungs and kidneys and the consequences of its interaction with antibodies in vitro and in vivo (Forssman reaction). Exposure of cultured guinea pig aortic endothelial cells to Forssman antibodies induced rapid redistribution of antigen-antibody complexes at the cell surface, followed by shedding that occurred by blebbing of plasma membrane as vesicles or fragments, and was associated with disappearance of antigen from the cell surface (antigenic modulation). Guinea pigs surviving frequent intravenous infections of increasing amounts of antibodies, for a total of 20 to 40 lethal doses, developed a partial or complete adaptation to generalized Forssman reaction, and adaptation was associated with partial or complete modulation of Forssman antigen at the surface of the pulmonary and, in minor degree, renal endothelial and epithelial cells. These findings support the hypothesis that modulation of endothelial carbohydrate antigens contributes to adaptation of highly vascularized organs exposed to tolerable levels of allo- or xenoantibodies.     

系膜增殖性肾炎中血栓素及白三烯对肾小球血液动力学的影响

在抗Ｔｈｙ１．１抗体所致的大鼠系膜增殖性肾炎中，测定了白细胞衍生的血栓素（ＴＸ）及白三烯（ＬＴ）对肾小球血液动力学的作用。在肾炎鼠，肾小球超滤系数（Ｋｆ）、肾小球滤过率（ＧＦＲ）及肾血流量（ＲＢＦ）的急性降低伴有肾小球白细胞计数的升高及肾小球ＴＸＢ２、ＬＴＢ４的合成增多。白细胞除去可抑制ＴＸＢＧ２及ＬＴＢ４的合成并完全防止ＧＦＲ及ＲＢＦ的降低。合用ＴＸ合成酶抑制剂及花生四烯酸５－脂氧化酶抑制剂也防     

Hapten-specific cellular immune response producing glomerular injury.

A new model of experimental glomerulonephritis (GN) in which a cell-mediated (delayed-type) reaction predominated was established. Cationized TNP-BSA conjugates were planted in the kidney of rats by perfusion via the renal artery. Rats which had been sensitized with the TNP hapten 7 days previously revealed marked exudative and proliferative changes in their glomeruli, accompanied by transient proteinuria. A cellular influx was seen without deposition of any autologous antibody. The resulting data are compatible with the idea that cell-mediated immune reactions can participate in tissue damage in glomerular disease.     

Severe anemia: a risk factor for glomerular injury in sickle cell disease.

Approximately 15% to 20% of patients with sickle cell disease have proteinuria. Proteinuria, particularly albuminuria, is the hallmark of glomerular injury. This study examines risk factors for glomerular injury as indicated by urinary albumin excretion (UAE) 30 microgram/minute, directly related to sickle cell disease. A total of seven patients were enrolled between September 1992 and March 1993. Fasting blood chemistries, complete blood cell count, 24-hour urine for protein and creatinine clearance, and glomerular filtration rate determined by 125 I-iothalamate were obtained for each patient. The results indicated that the lower the hematocrit, the higher the UAE rate. Low hematocrits have served as a protective mechanism in sickle cell disease by reducing blood viscosity and thus decreasing the number of vaso-occlusive crises. However, severe anemia appears to have an indirect adverse effect on the kidney in sickle cell disease.     

A possible role for leukotrienes in the regulation of glomerular haemodynamic in the dog

In anaesthetized beagles, saralasin, phentolamine, 1-penicillamine-2-O-methyl-tyrosine-8-arginine-vasopressin and SCH 23390, a DA₁ antagonist, were infused into the left renal artery (i.r.a) and indomethacin and aprotinin intravenously (Group 1). In Groups 2 and 3, i.r.a infusion of two chemically different putative leukotriene (LT) antagonists, FPL 55712 (100 g/kg/min) and LY 171883 (500 g/kg/min), respectively, was superimposed in the fourth period of experiments. In comparison to Group 1, there was an increase (40% in Group 2 and 33% in Group 3) in renal blood flow and a decrease in glomerular filtration rate (16% and 17%, respectively) and filtration fraction (36% and 41%, respectively). Similar changes were observed on the single nephron level. Directly measured glomerular capillary pressure decreased by 10% and 12%, respectively. A decrease in total arteriolar resistance by 34% and 25%, respectively, was caused by a comparatively higher decrease in efferent (44% and 34%, respectively) than afferent (26% and 15%, respectively) arteriolar resistance values. No change in the ultrafiltration coefficient,K _f, was detected. Providing FPL 55712 and LY 171883 are specific LT antagonists, these experiments suggest a possible constrictory role of LT (expressed more on the efferent than afferent arteriole) in anaesthetised mildly surgical stressed dogs.     

Nonimmunologic mechanisms of glomerular injury

From the above discussion it is clear that many factors have been invoked in the pathogenesis of progressive glomerular injury. Those which are most important include increased PGC, coagulation, serum lipid abnormalities, and hypertrophy. Although many hemodynamic alterations have been identified, increased PGC was noted most constantly. Furthermore, the loss of autoregulatory capability which was observed in some models with progressive glomerulosclerosis usually resulted in increased PGC. Increased PGC has been associated with augmented dietary protein and is seen in the Munich-Wistar rat made diabetic. Such an increase in PGC could cause direct mechanical injury to endothelial and epithelial cells, as well as be responsible for increased mesangial traffic of macromolecules with the potential for stimulating cellular proliferation and mesangial matrix increase. Additional support for the importance of increased PGC is provided by the protective effect of decreasing PGC with CEI therapy and anemia, and by the enhanced autoregulatory capability in both the Milan and Okamoto hypertensive rats. The significance of coagulation factors is confirmed by the formation of platelet and fibrin thrombi in the development of the glomerular lesions. The sequence of glomerular injury suggests that endothelial damage occurs with subsequent formation of platelet aggregates as a response to this injury. Formation of platelet aggregates may be associated with the production of substances potentially injurious to the endothelial cells. Although blocking the appearance of such thrombi by administration of heparin or thromboxane synthetase inhibitor prevents glomerular injury, the blood pressure lowering effect of these agents complicates the interpretation of the studies. Serum lipid abnormalities are also important factors in the progression of nonimmunologic glomerular injury. Such abnormalities are observed with increased dietary phosphorus or lipid, in the obese Zucker rat, and in rats with diabetes mellitus. Reduction in serum cholesterol by administration of clofibric acid or mevinolin diminishes glomerular injury independent of alterations in glomerular hemodynamics. The possible link between increased serum lipids and augmentation of glomerular injury is at present indirect. The importance of hypertrophy as a contributing factor to the progression of nonimmunologic glomerular injury is suggested by several lines of evidence. Hypertrophy, with increase in glomerular size and caliber of capillary loops, may amplify the effect of increased PGC by further intensifying the tension and mechanical stress on all elements of the capillary wall.(ABSTRACT TRUNCATED AT 400 WORDS)