Increased excretions of beta2-microglobulin, IL-6, and IL-8 and decreased excretion of Tamm-Horsfall glycoprotein in urine of patients with active lupus nephritis

Tubulointerstitial nephritis is a less frequently recognized but important complication of systemic lupus erythematosus. We have investigated the cytokine beta2-microglobulin (beta2M) and Tamm-Horsfall glycoprotein (THG) excretions in the urine of systemic lupus erythematosus patients to identify indices for evaluation of tubulointerstitial inflammation in lupus nephritis (LN). Daily urine was collected from 15 patients with active LN, from 12 patients with inactive LN, and from 17 normal subjects. The amounts of soluble interleukin (IL) 2 receptor, IL-6, IL-8, beta2M, and THG in urine were measured. Beta2M and THG were regarded as indicators of proximal and distal renal tubule function, respectively. The urinary excretions of IL-6 and IL-8 were significantly higher in patients with active LN than in those with inactive LN and in normal individuals. The excretion of soluble IL-2 receptor in all three groups of subjects was not significantly different. On the other hand, the excretion of beta2M in patients with LN was significantly higher than that in normal individuals. The excretion of beta2M in patients with active or inactive LN was not significantly different. The THG excretion was lower in patients with active LN and tubulointerstitial inflammation as compared with patients with inactive LN or normal individuals. Six patients underwent pulse cyclophosphamide therapy during the course of experiments. Five of them showed a decrease in IL-8 and IL-6 excretions in urine after the treatment. The excretions of beta2M and THG in urine, in addition to IL-6 and IL-8, can reflect the renal inflammatory activity in patients with lupus tubulointerstitial nephritis as well as in those having lupus glomerulonephritis.     

白细胞介素6在狼疮性肾炎患者小管间质的表达及其意义

探讨白细胞介素6(IL-6)在狼疮性小管间质病变中的作用。方法 采用ELISA方法与原位分子杂交技术(后者结合IBAS计算机图像分析系统),分别检测42例活动期狼疮性肾炎(LN)患者尿IL-6浓度与其中的15例肾小管间质IL-6mRNA水平。结果 42例活动期LN患者有36例尿IL-6>5pg/mg·cr,其增高程度与尿β_2-m及NAG活性水平呈显著正相关;其中15例肾组织切片中,肾小管间质均有IL-6mRNA表达,并且小管间质病变愈严重,其表达量愈高,而健康肾组织小管间质几无IL-6mRNA表达。结论 LN患者活动期尿IL-6浓度异常增高与肾小管间质IL-6mRNA异常表达有关,提示IL-6在狼疮性小管间质损害过程中可能具有重要作用。     

Interaction between proximal tubular epithelial cells and infiltrating monocytes/T cells in the proteinuric state

We hypothesize an interaction between T cells/monocytes and the tubules in the development of tubulointerstitial injury in chronic proteinuric nephropathy. We established in vitro co-culture systems of proximal tubular epithelial cells (PTEC) and T cells/monocytes to study the contribution of soluble factors and cell-to-cell contact in the development of tubulointerstitial injury. The release of monocyte chemoattractant protein-1 (MCP1 or CCL2), Regulated upon Activation, normal T cell Expressed and Secreted (RANTES or CCL5), soluble intracellular adhesion molecules-1 (sICAM-1), or interleukin-6 (IL-6) was increased in PTEC following apical exposure to human serum albumin (HSA). The release of CCL2, CCL5, or sICAM-1 from PTEC was enhanced by contact of monocytes/T cells on the basolateral surface. Tumor necrosis factor-alpha (TNF-alpha) and IL-1beta are important soluble factors as suggested by the blocking effect of antibodies (Abs) against TNF-alpha or IL-1beta but not against other cytokines. The percentage of CD4+ T cells expressing both chemokine receptors, CCR2 and CCR5, was increased after culturing with supernatant from the apical or basolateral surface of PTEC following apical exposure to HSA. However, only CCR2 was upregulated in CD8+ T cells, whereas CCR5 expression was increased in monocytes. The chemotaxis of CD4+ or CD8+ T cells to supernatant from PTEC upon apical exposure to HSA was reduced with neutralizing Abs against CCL5 and/or CCL2, whereas the chemotaxis of monocytes was only reduced by anti-CCL5 but not by anti-CCL2. In summary, chemokines released by HSA-activated PTEC are amplified by monocytes/T cells. Mediators released by HSA-activated PTEC can differentially modulate the expression of chemokine receptors in monocytes/T cells and hence, alter their chemotaxis towards activated PTEC. These interactions are pivotal in the development of tubulointerstitial injury.     

Activation of tubular epithelial cells by mesangial-derived TNF-alpha: glomerulotubular communication in IgA nephropathy

BACKGROUND: IgA nephropathy (IgAN), characterized by mesangial IgA deposition, runs a variable clinical course with tubulointerstitial damage and renal failure in no less than 30% of patients. Histologically, IgA is rarely detected in renal tubules. The direct toxicity by IgA on renal tubules remains uncertain. We hypothesize that mediators released from human mesangial cells (HMC) triggered by IgA deposition may lead to activation of proximal tubular epithelial cells (PTEC). METHODS: The binding of IgA to PTEC or HMC was assessed by flow cytometry. IgA-HMC medium was prepared by collecting the spent medium in which growth arrested HMC were incubated with IgA isolated from patients with IgAN, healthy control subjects, or other nephritic control patients. PTEC was cultured with the IgA-HMC medium in the presence or absence of neutralizing antibodies to TNF-alpha, IL-1beta, TGF-beta, or PDGF. Gene expression and protein synthesis of TNF-alpha, MIF, or ICAM-1 by PTEC were determined by RT-PCR and ELISA, respectively. RESULTS: The binding of IgA isolated from patients with IgAN to PTEC was increased when compared to binding of IgA from healthy control subjects (P < 0.005). However, the binding to PTEC was less than one tenth that of HMC in IgAN. The binding to PTEC was not mediated through known IgA receptors, as shown by competitive binding assays and gene expression of the receptors. Despite the in vitro binding, PTEC cultured with isolated IgA exhibited no increased cell proliferation or enhanced synthesis of TNF-alpha, MIF, or sICAM-1. However, when PTEC were cultured with IgA-HMC medium prepared from IgAN patients, there was enhanced proliferation of PTEC (P < 0.001) and increased synthesis of TNF-alpha, MIF, and sICAM-1 when compared with PTEC cultured with IgA-HMC medium from control subjects (P < 0.001). The synthesis of MIF and sICAM-1 by PTEC cultured with IgA-HMC medium was reduced by neutralizing antibodies to TNF-alpha (P < 0.001) but not by neutralizing antibodies to IL-1beta, TGF-beta, or PDGF. CONCLUSION: Our finding implicates that TNF-alpha released from the mesangium after IgA deposition activates renal tubular cells. The glomerulotubular communication could play an important role in the pathogenesis of tubulointerstitial damage in IgAN.     

Interferon-gamma is required for lupus nephritis in mice treated with the hydrocarbon oil pristane.

BACKGROUND: Although the precise mechanisms leading to lupus nephritis remain obscure, both TH1 and TH2 cytokines have been implicated. The present study examined the roles of interleukin (IL)-4 and interferon-gamma (IFN-gamma) in a novel inducible form of lupus that develops in non-autoimmune mice treated with the hydrocarbon oil pristane. METHODS: BALB/c IL-4 or IFN-gamma deficient mice (IL-4 -/-, IFNgamma -/-) and wild type controls (+/+) received either pristane or phosphate-buffered saline (PBS) IP. Serial sera were analyzed for anti-DNA/chromatin, anti-RNP/Sm, and total immunoglobulin levels. Proteinuria was measured and kidneys were examined by direct immunofluorescence and light microscopy. RESULTS: Renal disease did not develop in pristane-treated IFN-gamma -/- mice, as assessed by the absence of capillary immune deposits, glomerular pathology and proteinuria whereas IL-4 -/- mice developed renal disease similar to +/+ mice. Production of IgG anti-single stranded DNA and anti-chromatin antibodies was abrogated in IFN-gamma -/- mice. In contrast, these autoantibodies were produced at similar or higher frequencies and levels by IL-4 -/- versus wild-type mice. The frequency of anti-nRNP/Sm was markedly reduced in IFN-gamma -/- mice. IL-4 deficiency had little effect on the production of anti-DNA/chromatin and anti-nRNP/Sm. CONCLUSIONS: IFN-gamma is essential for the induction of nephritis and anti-DNA/chromatin following pristane exposure in BALB/c mice, suggesting that genetic or environmental factors influencing TH1-TH2 balance could be an important determinant of renal disease in lupus.     

Serum levels of IL-1 beta,TNF-alpha,IL-8, and acute phase proteins in seronegative spondyloarthropathies

OBJECTIVES: Some immunological abnormalities have been described in seronegative spondyloarthropathies (SpA). The aim of this study is to determine the serum levels of IL-1beta, TNF-alpha and IL-8, which are proinflammatory cytokines in active and inactive patients with SpA, to compare the results with those of controls and to investigate a relationship with clinical activity and acute phase proteins. METHODS: Forty-two patients (34 males and eight females) and 22 healthy controls (17 M and 5 F) were included in the study. All patients fulfilled Amor criteria for the classification of SpA. Among patients 23 had active and 19 had inactive disease. IL-1beta, TNF-alpha and IL-8 were determined by enzyme-linked immunosorbent assay ( ELISA), acute phase proteins were measured by nephelometric assay. RESULTS: There was no statistically significant difference between mean IL-1beta levels of patient groups and controls. Serum mean TNF-a levels in active and inactive patients were significantly increased as compared to that in the controls (P < 0.05, P < 0.05, respectively). Serum mean IL-8 levels in active patients was significantly increased as compared to that in the controls and in inactive patients (P < 0.01, P < 0.01, respectively). High serum IL-8 levels correlated well with C-reactive protein and haptoglobulin, but there was no correlation between IL-1beta or TNF-alpha levels and acute phase proteins such as C-reactive protein, alpha-1 acid glycoprotein, alpha-1 antitrypsin and haptoglobulin. CONCLUSIONS: These results suggest that serum IL-8 may reflect clinical activity of the disease and may be helpful for monitoring patients with SpA.     

Inflammation-Induced IL-6 Functions as a Natural Brake on Macrophages and Limits GN

IL-6 can mediate proinflammatory effects, and IL-6 receptor (IL-6R) blockade as a treatment for inflammatory diseases has entered clinical practice. However, opposing effects of IL-6 have been observed in models of GN. Although IL-6 is proinflammatory in murine lupus nephritis, protective effects have been observed for IL-6 in the nephrotoxic nephritis (NTN) model of acute crescentic GN. In light of the potential dangers of IL-6–directed treatment, we studied the mechanisms underlying the contradictory findings in GN. IL-6 can signal through the membrane-bound IL-6R, which is expressed only on hepatocytes and certain leukocytes (classic), or through the soluble IL-6R, which binds the ubiquitously expressed gp130 (alternative). Preemptive treatment of mice with anti-IL-6R or anti-IL-6 worsened NTN, whereas selective blockade of alternative IL-6 signaling by the fusion protein sgp130Fc did not. FACS analysis of mouse spleen cells revealed proinflammatory macrophages express the highest levels of IL-6Rα, and in vitro treatment with IL-6 blocked macrophage proliferation. Furthermore, proinflammatory macrophages were expanded during inflammation in IL-6^−/− mice. Late application of anti-IL-6 after establishment of adaptive nephritogenic immunity was sufficient to aggravate NTN within 2.5 days, a period when macrophages are active. Finally, NTN was aggravated in mice with macrophage-specific impairment of IL-6 classic signaling, coincident with enhanced macrophage proliferation and accumulation in the kidney. Our data thus reveal a novel mechanism in which IL-6–mediated dampening of macrophage activation protects tissues from overshooting immune responses. This finding has important implications for potential IL-6–directed therapies and supports the careful choice of recipient patients and timing.     

Heterogeneity of immune complex-derived anti-DNA antibodies associated with lupus nephritis

The mechanisms responsible for the tissue injuries associated with lupus nephritis have not yet been well explained. We have investigated the characteristics of anti-DNA antibodies in circulating immune complexes (CIC) and in the deposits of renal glomeruli in patients with active lupus nephritis. The CIC-derived antibodies expressed anti-DNA idiotypes (Id) designated as 0-81 Id and NE-1 Id, and bound mainly to single-stranded DNA but never to glomerular basement membrane (GBM) antigens. On the other hand, the immunoglobulins (Ig) eluted from renal glomeruli of lupus patients reacted not only with DNA but also with GBM, proteoglycan, and heparan sulfate. The binding of glomeruli-deposited Ig was markedly low when GBM antigens were used after treatment with heparitinase, suggesting that some anti-DNA antibodies may bind directly to GBM antigens associated with heparan sulfate, and form in situ IC in renal glomeruli. It was also revealed that the renal eluates obtained after passing through GBM antigen-coupled Sepharose lost the binding ability with GBM but still retained DNA-binding and 0-81 Id activity, showing the participation of circulating IC-derived anti-DNA antibodies in the glomerular deposits. Theoretically there may be two mechanisms in the pathogenesis of lupus nephritis through the deposition of circulating IC and through in situ formation of anti-DNA IC in renal glomeruli. The diversity of histological features in lupus kidneys may be attributed to the heterogeneity of the mechanisms.     

Mesangial cell-binding anti-DNA antibodies in patients with systemic lupus erythematosus

The mechanisms by which anti-DNA antibodies contribute to the pathogenesis of lupus nephritis (LN) remain to be elucidated. This study investigates the binding of polyclonal anti-DNA immunoglobulins from patients with systemic lupus erythematosus (SLE) to human mesangial cells (HMC) in vitro. Testing of cross-sectional serum samples from 280 LN patients (108 during active disease; 172 during remission), 35 SLE patients without renal involvement, 72 patients with non-lupus primary glomerular diseases, and 37 healthy subjects with a cellular enzyme-linked immunosorbent assay showed significant IgG mesangial cell-binding activity in patients with SLE, particularly those with active LN (P < 0.0001). Significant HMC-binding activity was demonstrated in 83.9%, 42.8%, and 47.1% of patients with active LN, inactive LN, and non-renal SLE, respectively. This was predominantly attributed to binding by anti-DNA antibodies, and immune complex binding accounted for 4.6%, 3.5%, and 2.8% of seropositive samples in the respective groups. Longitudinal studies in 27 LN patients demonstrated correlation between serial levels of anti-DNA antibodies, serum HMC-binding activity, and disease activity in 18 patients (66.7%). Affinity-purified polyclonal IgG anti-DNA antibodies from sera with HMC-binding activity showed significant binding to cultured HMC, and to a lesser extent glomerular and proximal tubular epithelial cells and human umbilical vein endothelial cells, but not tumor cell lines, peritoneal mesothelial cells, bronchial epithelial cells, or fibroblasts. The binding of anti-DNA antibodies to HMC was increased 1.47-fold (P = 0.0059) after the removal of Ig-associated DNA by DNase treatment, but it was unaffected by DNase treatment of HMC membrane. Controlled trypsinization of membrane proteins in HMC resulted in a 1.26-fold (P = 0.0025) increase in their binding by anti-DNA antibodies. In conclusion, subsets of anti-DNA antibodies from patients with SLE are capable of binding to HMC. The association of such binding with renal involvement and disease activity and its modulation by DNA concentration suggest that Ig binding to HMC can be a potential marker for disease activity in selected patients and that the binding of anti-DNA antibodies to HMC may be a pathogenetic mechanism in LN.     

Wound-healing factors secreted by epidermal keratinocytes and dermal fibroblasts in skin substitutes

Full-skin substitutes, epidermal substitutes, and dermal substitutes are currently being used to heal deep burns and chronic ulcers. In this study, we investigated which wound-healing mediators are released from these constructs and whether keratinocyte-fibroblast interactions are involved. Autologous skin substitutes were constructed from human keratinocytes, fibroblasts, and acellular donor dermis. Full-thickness skin was used to represent an autograft. Secretion of wound-healing mediators was investigated by means of protein array, enzyme-linked immunosorbent assay, neutralizing antibodies, and conditioned culture supernatants. Full-skin substitutes and autografts produce high amounts of inflammatory/angiogenic mediators (IL-6, CCL2, CXCL1, CXCL8, and sST2). Epidermal and dermal substitutes produced less of these proteins. Epidermal-derived proinflammatory cytokines interleukin-1alpha (IL-1alpha) and tumor necrosis factor-alpha (TNF-alpha) were found to mediate synergistically the secretion of these wound-healing mediators (with the exception of sST2) from fibroblasts in dermal substitutes. The secretion of proinflammatory cytokines (IL-1alpha, TNF-alpha), chemokine/mitogen (CCL5) and angiogenic factor (vascular endothelial growth factor) by epidermal substitutes and tissue remodeling factors (tissue inhibitor of metalloproteinase-2, hepatocyte growth factor) by dermal substitutes was not influenced by keratinocyte-fibroblast interactions. The full-skin substitute has a greater potential to stimulate wound healing than epidermal or dermal substitutes. Both epidermal-derived IL-1alpha and TNF-alpha are required to trigger the release of dermal-derived inflammatory/angiogenic mediators from skin substitutes.     

Viral double-stranded RNA aggravates lupus nephritis through Toll-like receptor 3 on glomerular mesangial cells and antigen-presenting cells

How viral infections trigger autoimmunity is poorly understood. A role for Toll-like receptor 3 (TLR3) was hypothesized in this context as viral double-stranded RNA (dsRNA) activates dendritic cells to secrete type I interferons and cytokines that are known to be associated with the disease activity in systemic lupus erythematosus (SLE). Immunostaining of nephritic kidney sections of autoimmune MRL(lpr/lpr) mice revealed TLR3 expression in infiltrating antigen-presenting cells as well as in glomerular mesangial cells. TLR3-positive cultured mesangial cells that were exposed to synthetic polyinosinic-cytidylic acid (pI:C) RNA in vitro produced CCL2 and IL-6. pI:C RNA activated macrophages and dendritic cells, both isolated from MRL(lpr/lpr) mice, to secrete multiple proinflammatory factors. In vivo, a single injection of pI:C RNA increased serum IL-12p70, IL-6, and IFN-alpha levels. A course of 50 microg of pI:C RNA given every other day from weeks 16 to 18 of age aggravated lupus nephritis in pI:C-treated MRL(lpr/lpr) mice. Serum DNA autoantibody levels were unaltered upon systemic exposure to pI:C RNA in MRL(lpr/lpr) mice, as pI:C RNA, in contrast to CpG-DNA, failed to induce B cell activation. It therefore was concluded that viral dsRNA triggers disease activity of lupus nephritis by mechanisms that are different from those of bacterial DNA. In contrast to CpG-DNA/TLR9 interaction, pI:C RNA/TLR3-mediated disease activity is B cell independent, but activated intrinsic renal cells, e.g., glomerular mesangial cells, to produce cytokines and chemokines, factors that can aggravate autoimmune tissue injury, e.g., lupus nephritis.     

抗核小体抗体与狼疮肾炎活动性关系的研究

目的探讨抗核小体抗体与狼疮肾炎活动性的关系。方法对2007年4月至2009年12月在我院住院的75例经肾活检确诊为狼疮肾炎患者的临床资料进行回顾性研究。结果抗核小体抗体在活动性增殖性狼疮肾炎中的阳性率（59．3％）较非活动性增殖性狼疮肾炎（27．3％）和非增殖性狼疮肾炎（20．0％）高（P〈0．05）。活动性狼疮肾炎组的抗核小体抗体阳性率（53．8％）较非活动性狼疮‘肾炎组阳性率（23．8％）高（P〈0．05）。抗核小体抗体阳性组的病理活动性指数（active index,AI）较阴性组高（P〈0．05）,两组慢性指数（chronic index.CI）和狼疮肾炎的临床活动性评分（renalactivity score,RAS）无显著差异（P〉0．05）。结论抗核小体抗体与狼疮肾炎的病理活动性相关,在活动性增殖性狼疮肾炎中出现较为普遍,是提示活动性增殖性狼疮肾炎的一项重要的指标。     

Anti-DNA autoantibodies initiate experimental lupus nephritis by binding directly to the glomerular basement membrane in mice

The strongest serological correlate for lupus nephritis is antibody to double-stranded DNA, although the mechanism by which anti-DNA antibodies initiate lupus nephritis is unresolved. Most recent reports indicate that anti-DNA must bind chromatin in the glomerular basement membrane or mesangial matrix to form glomerular deposits. Here we determined whether direct binding of anti-DNA antibody to glomerular basement membrane is critical to initiate glomerular binding of anti-DNA in experimental lupus nephritis. Mice were co-injected with IgG monoclonal antibodies or hybridomas with similar specificity for DNA and chromatin but different IgG subclass and different relative affinity for basement membrane. Only anti-DNA antibodies that bound basement membrane bound to glomeruli, activated complement, and induced proteinuria whether injected alone or co-injected with a non-basement-membrane-binding anti-DNA antibody. Basement membrane-binding anti-DNA antibodies co-localized with heparan sulfate proteoglycan in glomerular basement membrane and mesangial matrix but not with chromatin. Thus, direct binding of anti-DNA antibody to antigens in the glomerular basement membrane or mesangial matrix may be critical to initiate glomerular inflammation. This may accelerate and exacerbate glomerular immune complex formation in human and murine lupus nephritis.     

Mechanisms of tubulointerstitial injury in IgA nephropathy

BACKGROUND: IgA nephropathy (IgAN) runs a highly variable clinical course, with frequent involvement of tubulointerstitial damage. A subgroup of IgAN with severe tubulointerstitial damage is often associated with the most rapid progression to end-stage renal failure. In IgAN, mesangial sclerosis and tubulointerstitial damage were found to be correlated with the increase in pore size of the glomerular barrier. METHODS: The direct toxicity of proximal tubular epithelial cells (PTEC) by IgA in IgAN is still unresolved. Activation of PTEC by mediators released from infiltrating cells or resident kidney cells that induce tubular inflammation is the common final pathway in most chronic renal diseases. We hypothesize that mediators released from human mesangial cells (HMC) triggered by IgA deposition may lead to PTEC activation. RESULTS: We found that IgA binding to PTEC was less than one tenth that of HMC. The binding was nonspecific and exhibited no increased cell proliferation or enhanced synthesis of cytokines or adhesion molecules. However, when PTEC were cultured with IgA-HMC spent medium prepared from IgAN patients, there was enhanced proliferation of PTEC and increased synthesis of cytokines and adhesion molecules. CONCLUSION: These findings implicate a glomerulotubular cross-talk with mediators released from the mesangium, contributing to the pathogenesis of tubulointerstitial damage in IgAN. There are preliminary data to suggest that the expression of angiotensin II subtype-1 receptor and angiotensin II subtype-2 receptor in PTEC differs from that of HMC. These novel findings may provide clinicians new therapeutic approach for selective blockade of the tubulointerstitial injury in IgAN.     

Predominant tubulointerstitial nephritis in a patient with systemic lupus nephritis

In most cases of systemic lupus erythematosus (SLE), glomerular lesions are the main renal complication. Although tubulointerstitial lesions are often associated with severe glomerular lesions, predominant or isolated tubulointerstitial injury in the presence of minimal glomerular abnormalities with SLE, so-called predominant tubulointerstitial lupus nephritis, is rare. Only ten cases are reported in the English literature. Herein, we describe the case of a 64-year-old man with SLE who presented with acute renal deterioration attributable to acute tubulointerstitial nephritis. Renal biopsy showed diffuse infiltration of inflammatory mononuclear cells in the interstitium and tubulitis without significant glomerular lesions. Immunofluorescence study revealed positive staining for IgG, C3, and C1q along the renal tubular basement membrane (TBM). Electron microscopy also showed electron-dense deposits in the TBM. Other causes of tubulointerstitial injury, such as drug use and infection, were ruled out. Taking these findings together with the presence of antitubular basement membrane antibody, predominant tubulointerstitial lupus nephritis was diagnosed. Treatment with oral corticosteroids for 6 weeks improved renal function. Even after tapering of the corticosteroid, renal function and serological markers of SLE activity have remained stable in this patient for more than 12 months.     

Angiotensin II regulates the synthesis of proinflammatory cytokines and chemokines in the kidney

BACKGROUND: Emerging evidence suggests that angiotensin II (Ang II) is not only a vasoactive peptide, but also a true cytokine that regulates cell growth, inflammation and fibrosis. Many studies have demonstrated that this peptide plays an active role in the progression of renal injury. Some of Ang II-induced effects are mediated by the production of a large array of growth factors. The aim of this study was to investigate whether Ang II could regulate the expression of cytokines and chemokines in the kidney and its correlation with the Ang II-induced renal damage. METHODS: The model of Ang II-induced renal damage was done by systemic Ang II infusion into normal rats (50 ng/kg/min; subcutaneous osmotic minipumps). In addition, the implication of Ang II was investigated in a model of immune complex nephritis in rats treated with the angiotensin converting enzyme (ACE) inhibitor quinapril. The mRNA expression was analyzed by RT-PCR and/or Northern blot, and protein levels by Western blot and/or immunohistochemistry. RESULTS: Rats infused with Ang II for 3 days caused elevated renal expression of tumor necrosis factor-alpha (TNF-alpha; gene and protein levels). TNF-alpha positive cells were observed in glomeruli (mainly in endothelial cells), tubules and vessels. In rats with immune complex nephritis, the renal overexpression of TNF-alpha was diminished by the ACE inhibitor quinapril. Systemic infusion of Ang II also increased renal synthesis of cytokines (interleukin-6, IL-6) and chemokines (monocyte chemoattractant protein-1; MCP-1) that were associated with elevated tissue levels of activated nuclear factor-kappaB (NF-kappaB) and the presence of inflammatory cell infiltration. CONCLUSIONS: Ang II in vivo increases TNF-alpha production in the kidney. Ang II also up-regulates other proinflammatory mediators, including IL-6, MCP-1 and NF-kappaB, coincidentally associated to the presence of glomerular and interstitial inflammatory cells in the kidney. All these data further strengthen the idea that Ang II plays an active role in the inflammatory response in renal diseases.