Two distinct types of crescentic glomerulonephritis.

For a characterization of the clinical course of crescentic glomerulonephritis (Cres. GN), reciprocals of serum creatinine concentration (1/Cr) as a function of time were studied in 24 patients. The patients fulfilled the following criteria; 1) crescents were observed in more than 50% of glomeruli, and 2) the increment of serum creatinine could be determined sequentially on three or more occasions in the phase of progression of renal impairment. In all patients 1/Cr declined linearly with time with correlation coefficients between 0.881 and 0.993. According to the slope, the patients were divided into two groups; an acute group (13 patients) with slopes of -1.0 x 10(-2) dl/mg/day or more steep, i.e. with increments of serum creatinine from 1.0 to 10.0 mg/dl within 90 days, and a subacute group (11 patients) with less steep slopes. Histologically, there was no difference in the percentage of glomeruli with crescents between the two groups. However, in the acute group a negative correlation was found between the period from onset to histological examination and the percentage of glomeruli with cellular crescents in all glomeruli with various crescents (r = -0.872, p less than 0.001), while in the subacute group the percentage was nearly constant regardless of the time of histological examination. These mean drastic but transient activation of the disease in the former in spite of a persistence of indolent activity in the latter. Clinically, in the acute group an improvement of serum creatinine was observed in 8 (61.5%) vs. only 2 (18.2%) in the subacute group (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Pentoxifylline attenuates experimental mesangial proliferative glomerulonephritis.

BACKGROUND: Accumulation of glomerular macrophages, proliferation of mesangial cells (MCs), and deposition of extracellular matrix proteins are pathobiological hallmarks of glomerulonephritis. We previously reported that a clinically available nonselective inhibitor of cyclic 3',5'-nucleotide phosphodiesterase, pentoxifylline (PTX), inhibits proliferation of cultured rat MCs, as well as collagen production by these cells. In this study, we investigated the in vivo effects of PTX on rat anti-Thy1 disease, a model of mesangial proliferative nephritis. METHODS: Anti-Thy1 nephritis was induced in Sprague-Dawley rats by injecting mouse anti-rat Thy1 antibodies intravenously. Nephritic rats were randomly assigned to receive PTX (0.1 g/kg/day) or vehicle (phosphate-buffered saline) and were sacrificed at various time points. Paraffin kidney sections were stained with hematoxylin and periodic acid-Schiff reagents for glomerular histology. Frozen kidney sections were stained by monoclonal antibodies against proliferating cell nuclear antigen, ED-1, and alpha-smooth muscle actin and were visualized by color development from a horseradish peroxidase reaction. Monocyte chemoattractant protein-1 (MCP-1), intercellular adhesion molecule-1 (ICAM-1), and various extracellular matrix mRNAs were analyzed by Northern blotting. Urine protein concentrations were determined by Lowry's method. RESULTS: Nephritic rats treated with PTX excreted less urinary protein on day 5 of nephritis than vehicle-treated nephritic rats. In periodic acid-Schiff-stained kidneys from PTX-treated nephritic rats, there was attenuation of both glomerular cellularity and glomerular sclerosis compared with vehicle-treated nephritic rats. PTX decreased the augmented glomerular mRNA levels of MCP-1 and ICAM-1 at two hours and on day 1 of nephritis. Immunoreactive staining showed that PTX reduced the number of proliferating glomerular macrophages on days 1, 2, and 3, but not at two hours of nephritis, compared with vehicle-treated nephritic rats. On day 5, PTX decreased the number of activated proliferating MCs and attenuated the glomerular mRNA levels of type I (alpha1), type III (alpha1), and type IV (alpha1) collagen and fibronectin compared with vehicle-treated nephritic rats. CONCLUSION: The administration of PTX to rats with anti-Thy1 disease reduces accumulation and proliferation of glomerular macrophages, attenuates proteinuria, suppresses activation and proliferation of MCs, and ameliorates glomerular sclerosis. These results suggest that PTX may have a suppressive effect in acute phases or relapses of mesangial proliferative glomerulonephritis.     

The use of heparin in the treatment of idiopathic rapidly progressive glomerulonephritis.

14 cases of idiopathic rapidly progressive glomerulonephritis (I. R. P. G. N.) treated by heparin between 1968 and 1974 were collected by the authors. Extra-capillary crescents (E. C. C.) occurred in 75 to 100% of glomeruli in ten patients and in 50 to 75% in four. Gross proteinuria, hematuria and renal failure were always present. 9 patients were admitted with primary oligo-anuric renal failure. 11 patients were treated by repeated hemodialysis before and during anticoagulant treatment. Heparin was given by intra-venous injection every 3 hours for one to two months with Howell times range from 150 to 200% of control. Heparin was the only treatment in 6 cases, and was given with dipyridamole in 4, with prednisone in 3 and with azathioprine in one case. 5 severe or fatal hemorragic complications were observed. The clinical course was usually unfavorable with 5 early deaths, 3 provisional steady-states with 2 late deaths. Six patients were treated by periodic hemodialysis. Repeat kidney biopsies were obtained in 8 patients. The findings suggest that heparin affects mainly the E. C. C. and fibrinoid deposits but not glomerular sclerosis. The inefficiency of all current treatments of primary oligo-anuric IRPGN is stressed. In patients with better initial renal function choice between anticoagulant and/or immuno-depressive drugs must be scrutinized in individual cases bearing in mind potential iatrogenic complications. In equivocal cases, patients should be referred to the chronic hemodialysis and/or transplantation program.     

蕲蛇酶联合肝素治疗兔急性下腔静脉血栓的实验研究

目的 探讨蕲蛇酶联合肝素治疗兔急性下腔静脉血栓的有效性及安全性.方法 新西兰大白兔72只建立急性下腔静脉血栓模型后随机分为肝素组(A组)、尿激酶联合肝素组(B组)、蕲蛇酶联合肝素组(C组),每组24只.各组在血栓形成3 d后给药,分别在用药后的3、7、10 d检测各组凝血指标评估其安全性,通过静脉彩超评估其有效性.结果 C组的凝血酶原时间(prothrombin time,PT)延长时间较B组短(P<0.05),纤维蛋白原(fibrinogen,FBG)值较B组小(P<0.05),B组和C组PT延长时间较A组长(P<0.01),FBG值较A组大(P<0.01),B组和C组的D-二聚体值(D-dimer,D-D)均逐渐降至正常,差异无统计学意义(P>0.05).B组和C组的溶栓效果均较A组效果好,B组与A组差异有统计学意义(P<0.01),第10天时,C组与A组差异有统计学意义(P<0.01).B组和C组溶栓效果差异无统计学意义(P>0.05).结论 蕲蛇酶联合肝素治疗兔急性下腔静脉血栓是有效的,并且具有较高的安伞性.     

Platelet-derived growth factor in experimental glomerulonephritis

Growth factors and in particular platelet-derived growth factor(PDGF) have been implicated in the pathogenesis of glomerulonephritisand glomerulosclerosis. We have studied the distribution ofimmunoreactive PDGF (iPDGF) within serial kidney biopsies (days7, 15, 30, 90 and 120) of eight rats with an accelerated formof nephrotoxic serum nephritis (NTN). The course of NTN wasmild in five rats and severe in three. Two patterns of immunostainfor PDGF were noted. The first consisted of iPDGF-B chain ina glomerular segmental distribution similar to that of infiltratingmonocytes (OX6+cells). At all stages of NTN the distributionof iPDGF-B chain correlated closely with the immunostain formonocytes. The second pattern of immunostain showed iPDGF-Achain in a diffuse distribution along the glomerular capillarylining and to a lesser extent in some mesangial cells. In severelyaffected rats the magnitude of the iPDGF-A increased along withglomerulosclerosis but disappeared later from areas of segmentaland global glomerular obsolescence. By contrast, in rats withmilder NTN glomerular iPDGF-A chain peaked early and decreasedsubsequently. During the acute phase of NTN, on day 7, iPDGFA chain correlated with the severity of proteinuria (r=0.848,P<0.05) and that of glomerular cellular proliferation (r=0.91,P<0.02). On day 30 iPDGF-B correlated positively with proteinuria(r=0.998, P<0.01, n=5) and with serum creatinine (r=0.949,P<0.02, n=5). During the late stages of NTN, both iPDGF-A(day 90, r=0.884, P<0.05) and iPDGF-B (day 120, r=0.941,P<0.01) correlated closely with the extent of glomerulosclerosis.Both PDGF chains (A and B) are detectable through out the courseof nephrotoxic nephritis in rats. PDGF may play a role in theearly proliferative and late sclerotic changes of experimentalglomerulonephritis.     

Interferon-beta reduces proteinuria in experimental glomerulonephritis

Interferon-beta (IFN-beta) is a multifunctional cytokine with immunomodulatory properties. We examined the effect of IFN-beta in three separate rat models of glomerular injury and in cultured human glomerular endothelial cells and podocytes. In nephrotoxic nephritis in WKY rats, recombinant rat IFN-beta started either at induction or after establishment of disease significantly reduced 24-h proteinuria by up to 73% and 51%, respectively, but did not affect serum creatinine. There was a slight reduction in numbers of glomerular macrophages, but no difference in glomerular or tubulointerstitial scarring. In Thy-1 nephritis in Lewis rats, IFN-beta started at induction of disease reduced proteinuria by up to 66% with no effect on numbers of glomerular macrophages, but a reduced number of proliferating cells. In puromycin nephropathy in Wistar rats, IFN-beta started at induction of disease reduced proteinuria by up to 93%, but had no effect on glomerular histology. In cultured cells, human IFN-beta-1a had a dramatic effect on barrier properties, increasing electrical resistance across monolayers of either glomerular endothelial cells or podocytes and decreasing trans-monolayer passage of albumin. In conclusion, these results show that IFN-beta reduces proteinuria in three different rat models of glomerular injury and that its anti-proteinuric action may result from direct effects on cells that comprise the glomerular filtration barrier. These data indicate that IFN-beta may have potential as a therapeutic agent in proteinuric renal disease.     

Low-dose heparin in experimental peritonitis

The aim of this investigation was to study the beneficial role of low-dose heparin in peritonitis. Peritonitis was induced in 20 dogs by the technique of Rosato and co-authors, 10 of which received low-dose heparin subcutaneously for 5 days in a dose of 100 units/kg body weight. Decreased mortality as well as decrease in the formation of intra-abdominal abscesses and adhesions were observed in the heparin-treated dogs. Some of the possible underlying mechanisms for beneficial effects of heparin in peritonitis are discussed.     

The protective effect of heparin in experimental bladder infection.

The surface mucopolysaccharide layer of the urinary bladder has been shown to interfere with bacterial attachment, and we therefore consider it to be the primary mechanism of combatting bacterial colonization and subsequent infection. Experimentally, hydrochloric acid alters this layer in such a way as to permit massive bacterial adsorption to the vesical mucosa. Heparin, an acid mucopolysaccharide, was instilled into rabbit bladders that had their naturally occurring mucopolysaccharide altered by hydrochloric acid. Heparin was shown to inhibit the attachment of inoculated Escherichia coli to these acid-treated bladder mucosas, and the degree of protection afforded was similar to that of the mucopolysaccharide layer of the intact urothelium.     

Dissociation of mesangial cell migration and proliferation in experimental glomerulonephritis.

BACKGROUND: Recently, we documented that following in vivo mesangial cell (MC) ablation in the Thy1 model, reconstitution of the mesangium occurs by a coordinated proliferation and migration of Thy1 (OX-7)-positive cells originating from the hilus and extraglomerular mesangium. We investigated the role of basic fibroblast growth factor (bFGF) in the mediation of these events. METHODS: Rats were injected with antithymocyte serum and 48 hours later were pulsed with 3H-thymidine to label proliferating cells. Ninety minutes later, a baseline renal biopsy was obtained, and rats were injected with neutralizing anti-bFGF antibodies or control IgG. Sacrificial biopsies were obtained at 96 hours of disease. Using computer image analysis, biopsies from both time points were quantitated for the number of radiolabeled MC (proliferation) and their mean distance from the hilus (migration). The effect of bFGF on the migration of MCs in culture was examined using a chemotactic assay. RESULTS: At sacrifice, autoradiographs of rats receiving anti-bFGF had significantly fewer radiolabeled MCs as compared with rats receiving control IgG (8.7+/-1.9 vs. 14.7+/-3.5, P = 0.0001), yielding an overall 40% reduction in proliferation. There was no difference, however, in the final distance of radiolabeled MCs from the glomerular hilus in the two groups, indicating that the administration of anti-bFGF did not effect MC migration in this model. In an in vitro chemotactic assay, MCs migrated in response to platelet-derived growth factor (PDGF) BB (20 ng/ml), but did not migrate in response to bFGF at a wide range of concentrations (0.5 to 50 ng/ml). CONCLUSIONS: These studies demonstrate that bFGF is an important mediator of MC proliferation but that it does not significantly influence MC migration. This is the first demonstration showing that the mediators effecting proliferation can be dissociated from those mediating migration in renal injury.     

Initiation and evolution of interstitial leukocytic infiltration in experimental glomerulonephritis.

Most forms of glomerulonephritis have a significant interstitial leukocytic infiltrate which is associated with disease progression. However, there is little data concerning the timing, initial location, and development of this interstitial component. Therefore, we have addressed these issues in a study of passive accelerated anti-GBM glomerulonephritis in the rat. In this model, interstitial leukocytic infiltration was an early event in the disease process with a significant infiltrate apparent at 12 hours after administration of nephrotoxic serum (NTS). This initial infiltrate was restricted to a perivascular sheath surrounding the hilar arterioles. The sheath infiltrate then spread to include the whole hilar area by day 1, the entire periglomerular area by day 3, and became widespread throughout the cortical tubulointerstitium by day 7. The early sheath infiltrate was composed of macrophages and T cells. Both cell types continued to increase as the infiltrate expanded, and a significant accumulation of activated cells (IL-2R+) was evident from day 7 onwards. There was a highly significant correlation between interstitial macrophage infiltration and renal function impairment, proteinuria, and histologic damage. Interstitial T cell infiltration correlated with proteinuria and histologic damage, while the appearance of immune-activated mononuclear cells (IL-2R+) exhibited a highly significant correlation with all disease parameters. This study demonstrates the importance of the glomerular hilar arteriolar region as a focus for mononuclear leucocytic migration and accumulation which not only affects the structure and function of the glomerulus but subsequently the entire tubulointerstitium.     

Contribution of superficial nephron segments to sodium excretion in experimental glomerulonephritis

The present study examined the contribution of individual superficial nephron segments to sodium excretion in antiglomerular basement membrane nephritis in the rat by sampling the same nephron successively from the end and beginning of the distal tubule and end of the proximal tubule. Whole kidney GFR in glomerulonephritic rats was reduced by approximately 40% from controls; absolute sodium excretion was about 25% of normal. Metabolic balance studies in the awake state had suggested that the animals were in sodium balance. Plasma renin levels before and during micropuncture were similar to controls. These findings suggest that the defect in sodium handling is intrinsic to the kidney. Glomerulotubular balance was maintained along the proximal tubule. Sodium reabsorption in the loop of Henle was reduced in absolute terms but was proportional to the load delivered. Due to the decreased absolute sodium reabsorption in the preceding segments, sodium delivery to the beginning of the distal tubule was comparable in the two groups of animals. Along the distal tubule sodium reabsorption was comparable to control animals. Therefore, the avid urinary sodium retention seen during micropuncture was due to increased sodium reabsorption by segments past the superficial proximal tubule and/or by deep nephrons.     

Contribution of the distal tubule to potassium excretion in experimental glomerulonephritis

The renal adaptations that maintain potassium homeostasis in diffuse forms of glomerular disease are not well defined. Thus, handling of potassium by superficial nephron segments was examined in a rat model of antiglomerular basement membrane nephritis. Sampling the same nephron successively from the end and beginning of the distal tubule and the end of the proximal tubule allowed a segmental analysis. Despite a 40% reduction in GFR, potassium excretion in the glomerulonephritis animals was normal due to an increase in FEK. The proximal tubule and loop segment did not contribute to the enhanced FEK seen in these animals. In contrast, potassium entry along the distal tubule was significantly greater in the experimental group averaging 13.7 +/- 4.3 pmol/min compared to 1.2 +/- 1.7 pmol/min in controls (P less than 0.01). Multiple linear regression analysis showed that distal tubule potassium entry at any level of flow was enhanced in glomerulonephritis compared to controls (P less than 0.0001). Plasma aldosterone levels were similar in both groups of animals. Thus, the adaptation to potassium excretion seen in glomerulonephritis is partly achieved by the distal tubule through flow-rate independent mechanisms and appears to be independent of plasma aldosterone levels.     

Glomerular expression of C-C chemokines in different types of human crescentic glomerulonephritis.

BACKGROUND: Crescentic glomerulonephritis (CGN) presents a rapidly progressive glomerulonephritis clinically, in which macrophages play a crucial role in the pathogenesis. However, the precise molecular mechanism of macrophage recruitment and activation has not been fully elucidated. C-C chemokines, monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1alpha and beta (MIP-1alpha and MIP-1beta), are major chemoattractants for macrophages. We attempted to study the expression of C-C chemokines and their correlation with CD68-positive macrophages in crescentic glomeruli to investigate further their possible roles in crescent formation and progression to fibrosis in different types of human CGN. METHODS: The expression of MCP-1, MIP-1alpha, MIP-1beta and CD68 was detected in glomeruli with different forms of crescents (cellular, fibrocellular and fibrous crescents) by immunohistochemistry in serial sections of renal biopsies taken from 32 patients with biopsy-proven CGN including eight patients with anti-glomerular basement membrane (GBM) disease (type I CGN), 12 patients with immune complex-mediated CGN (type II CGN) and another 12 patients with pauci-immune CGN (type III CGN) enrolled in this study. Eight normal human kidneys were obtained from cadaveric renal transplant donors whose kidneys were technically unsuitable for transplantation, serving as controls. RESULTS: MCP-1, MIP-1alpha, MIP-1beta and CD68 were undetectable in glomeruli of normal kidney. In crescentic biopsies, MCP-1, MIP-1alpha, MIP-1beta and CD68 were detected in fibrocellular crescents and were even more prominent in cellular crescents, but were undetectable in fibrous crescents. Using consecutive sections for staining, it was demonstrated that a high proportion of infiltrating CD68-positive macrophages, mainly localized to the area of the expression of chemokines, were MCP-1, MIP-1alpha and MIP-1beta positive in crescents. Chemokines were expressed mainly by CD68-positive macrophages and parietal epithelial cells in crescents. The number of MCP-1- and MIP-1alpha-positive cells in glomeruli with cellular crescents was positively correlated with the number of CD68-positive cells (r = 0.568 and 0.749, respectively, both P < 0.01). The number of MCP-1- and MIP-1alpha-positive cells and the incidence of Bowman's capsule rupture in glomeruli of patients with type I CGN were higher than those of type II and type III CGN. CONCLUSIONS: These observations suggest that the expressed C-C chemokines, MCP-1, MIP-1alpha and MIP-1beta, may mediate the inflammatory process of crescent formation and progression to fibrosis. The strong correlation of MCP-1 and MIP-1alpha with infiltrating macrophages within glomeruli with cellular crescents suggested that these chemokines might be of particular importance for macrophage recruitment to this site. MCP-1 and MIP-1alpha were correlated to type I CGN with its more severe inflammatory course and worse prognosis. The variance of glomerular expression of C-C chemokines may contribute to the difference in histopathological features and prognosis in these three types of CGN.     

The expression of matrix metalloproteinase-11 protein in various types of glomerulonephritis.

BACKGROUND: Matrix metalloproteinases (MMPs) have been implicated to play important roles in a number of pathological processes such as inflammation. In human glomeruli, the mesangial matrix turnover is controlled by a dynamic equilibrium between synthesis and degradation to which metalloproteinases are known to contribute. Metalloproteinase-11 (MMP-11) was originally discovered as a gene whose expression was associated with tissue remodelling. The aim of this study was to investigate whether MMP-11 protein is expressed in various types of glomerulonephritis and to elucidate the role of this expression. METHODS: Using standard immunohistochemistry, we analysed MMP-11 expression in renal biopsies from 95 patients with primary glomerulonephritis (n = 44) and secondary, either lupus-associated glomerulonephritis (n = 22) or pauci-immune, ANCA-associated glomerulonephritis due to small vessel vasculitis (n = 23) or Wegener's granulomatosis (n = 6). The examined cases were divided into two groups (proliferative and non-proliferative). Anti-Ki67 and -CD68 immunostaining was also performed in order to estimate cell proliferation and number of macrophages, respectively. RESULTS: MMP-11 immunopositivity was detected in the glomeruli of the majority of pathological samples. The highest incidence of MMP-11 immunopositivity (26.3%) was noticed in glomerulonephritides associated with microscopic polyangiitis and Wegener's granulomatosis. Generally, MMP-11 was often expressed in segmental areas of sclerosis, microadhesions, cellular and fibrocellular crescents. Fibrotic crescents and fibrotic glomeruli were constantly MMP-11-immunonegative. In MMP-11 immunoreactive glomeruli, increased numbers of macrophages were often detected in the mesangium (P = 0.001), while no such observation could be made with regard to proliferating cells (P = 0.170). CONCLUSIONS: MMP-11, like an inflammatory mediator, may exert a chemotactic influence on macrophages which aggregate in the mesangium; MMP-11 is not likely to have a parallel mitogenic or antifibrotic effect in diseased glomeruli.     

Surgical technique of experimental lung transplantation in rabbits.

BACKGROUND: Although rabbits have often been used as an experimental model for the analysis of lung preservation, there are no reports of long-term survival after rabbit lung allotransplantation. The purpose of this study was to establish a lung allotransplantation model for the evaluation of acute lung rejection in the rabbit and to investigate the transplantation techniques in the rabbit. METHODS: Left unilateral lung allotransplantations were performed in 10 pairs of Japanese white male rabbits, weighing from 2.8 to 3.7 kg. Rabbits were divided into two groups. Group A rabbits (n=5) received Cyclosporine A (CsA) (20 mg/kg/day) orally for 5 days postoperatively, while Group B rabbits (n=5) received no CsA. All rabbits were sacrificed at the fifth postoperative day for histological examinations. RESULTS: Anastomoses of the pulmonary vein were achieved by using the atrial cuff technique. Satisfied blood flow was obtained in all arterial and venous anastomosis sites. Bronchial anastomosis was also well healed and all rabbits could maintain adequate spontaneous ventilation. In Group A, histopathology revealed that three cases were grade A0, one was grade A1 and the other was A2. In Group B, three cases lived for five days, and histopathology showed two cases were grade A2 and one case was grade A3. CONCLUSION: We established a left unilateral lung allotransplantation model in the rabbit and observed suppression of acute rejection of the transplanted lung by CsA. This study suggests that the rabbit is also an experimental model suited for the analysis of lung preservation as well as lung allotransplant rejection.     

Strain susceptibility to active induction and passive transfer of experimental autoimmune glomerulonephritis in the rat.

BACKGROUND: Previous studies have shown that different inbred rat strains vary in their susceptibility to experimental autoimmune glomerulonephritis (EAG). The Wistar Kyoto (WKY) rat is highly susceptible and develops crescentic glomerulonephritis, while the Lewis (LEW) rat is resistant. When immunized with collagenase-solubilized rat glomerular basement membrane (GBM), both strains produce circulating autoantibodies reactive with rat GBM by enzyme-linked immunosorbent assay, but only the WKY rat shows strong linear deposits of IgG on the GBM. METHODS: We investigated the hypothesis that differences in the characteristics of the anti-GBM antibodies produced, or in the inflammatory response to antibody deposition, could account for susceptibility. RESULTS: We found that circulating anti-GBM antibodies from WKY rats immunized with GBM were present at a higher concentration than those from LEW rats. Antibodies from WKY rats also recognized the rat alpha3 chain of type IV collagen [alpha3(IV)NC1], whereas those from LEW rats did not. Antibody eluted from the kidneys of WKY rats with EAG induced by GBM showed a higher affinity for GBM and recombinant rat alpha3(IV)NC1 than circulating antibody. This eluted antibody bound strongly to normal kidney sections from both WKY and LEW rats. Passive transfer of eluted anti-GBM antibodies from WKY rats with EAG resulted in similar binding of IgG to the GBM of WKY and LEW rats at 24 h. However, only the WKY recipients went on to develop crescentic glomerulonephritis by 28 days. CONCLUSIONS: This study demonstrates that the characteristics of the anti-GBM antibodies induced in WKY rats contribute to their susceptibility to EAG. However, the passive transfer experiments reveal that factors related to the inflammatory response to antibody deposition are also important in determining susceptibility. A combination of these genetic influences could explain the variation in severity of human anti-GBM disease.