Generation of a novel decay accelerating factor (DAF) knock-out rat model using clustered regularly-interspaced short palindromic repeats, (CRISPR)/associated protein 9 (Cas9), genome editing.

Decay accelerating factor (DAF), a key complement activation control protein, is a 70 kDa membrane bound glycoprotein which controls extent of formation of the C3 and C5 convertases by accelerating their decay. Using clustered regularly-interspaced short palindromic repeats, (CRISPR)/associated protein 9 (Cas9) genome editing we generated a novel DAF deficient (Daf-/-) rat model. The present study describes the renal and extrarenal phenotype of this model and assesses renal response to complement-dependent injury induced by administration of a complement-fixing antibody (anti-Fx1A) against the glomerular epithelial cell (podocyte). Rats generated were healthy, viable and able to reproduce normally. Complete absence of DAF was documented in renal as well as extra-renal tissues at both protein and mRNA level compared to Daf+/+ rats. Renal histology in Daf-/- rats showed no differences regarding glomerular or tubulointerstitial pathology compared to Daf+/+ rats. Moreover, there was no difference in urine protein excretion (ratio of urine albumin to creatinine) or in serum creatinine and urea levels. In Daf-/- rats, proteinuria was significantly increased following binding of anti-Fx1A antibody to podocytes while increased C3b deposition was observed. The DAF knock-out rat model developed validates the role of this complement cascade regulator in immune-mediated podocyte injury. Given the increasing role of dysregulated complement activation in various forms of kidney disease and the fact that the rat is the preferred animal for renal pathophysiology studies, the rat DAF deficient model may serve as a useful tool to study the role of this complement activation regulator in complement-dependent forms of kidney injury.

Curcumin Improves the Renal Autophagy in Rat Experimental Membranous Nephropathy via Regulating the PI3K/AKT/mTOR and Nrf2/HO-1 Signaling Pathways.

Membranous nephropathy (MN, also known as membranous glomerulopathy) is one of the many glomerular diseases causing nephrotic syndrome. The literature indicates that autophagy is associated with the homeostasis of podocytes in glomeruli. Curcumin, the main active component in turmeric, has drawn attention for its effective bioactivities against chronic kidney disease. The current study was aimed at assessing the effects of curcumin and exploring the underlying mechanism that mediates autophagy in an animal model of passive Heymann nephritis (PHN) in rats. Passive Heymann nephritis (PHN) was induced in male SD rats by intraperitoneal injection of anti-Fx1A serum. The rats were divided into 3 groups: control (n = 10, normal diet), model group (n = 10, 0.5% sodium carboxymethylcellulose), and curcumin (n = 10, 300 mg/kg/d). The kidney function and oxidative stress indicators were measured using commercial diagnostic kits, and the histomorphology of renal tissues was observed. The number of podocytes was measured by immunohistochemistry. Meanwhile, the autophagosomes in podocyte were analyzed by transmission electron microscopy and the immunofluorescence assay pointing to p62, an autophagic marker. Western blot analyzed the levels of apoptosis, autophagy, PI3K/AKT/mTOR, and Nrf2/HO-1 pathway-associated proteins. The total cholesterol (TC), triglycerides (TG), creatinine (Scr), blood urea nitrogen (BUN), urine volume, and urine albumin of PHN rats were significantly reduced by the administration of curcumin and attenuated renal histomorphological changes in model rats. Meanwhile, curcumin improved the oxidative stress response by decreasing MDA and increasing SOD, GSH, and CAT levels in the kidney of PHN rats. Furthermore, curcumin significantly ameliorated the podocyte loss, along with the fusion, and increased the autophagic vacuoles compared to the PHN control rats. In addition, curcumin downregulated the expression of Bax, Caspase-3, p62, PI3K, p-AKT, and p-mTOR proteins and upregulated the Bcl-2, beclin1, LC3, Nrf2, and HO-1 levels in this animal model. The results provide a scientific basis that curcumin could significantly alleviate the development of MN by inducing autophagy and alleviating renal oxidative stress through the PI3K/AKT/mTOR and Nrf2/HO-1 pathways.

A plasmid-encoded peptide from Staphylococcus aureus induces anti-myeloperoxidase nephritogenic autoimmunity.

Autoreactivity to myeloperoxidase (MPO) causes anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV), with rapidly progressive glomerulonephritis. Here, we show that a Staphylococcus aureus peptide, homologous to an immunodominant MPO T-cell epitope (MPO409-428), can induce anti-MPO autoimmunity. The peptide (6PGD391-410) is part of a plasmid-encoded 6-phosphogluconate dehydrogenase found in some S. aureus strains. It induces anti-MPO T-cell autoimmunity and MPO-ANCA in mice, whereas related sequences do not. Mice immunized with 6PGD391-410, or with S. aureus containing a plasmid expressing 6PGD391-410, develop glomerulonephritis when MPO is deposited in glomeruli. The peptide induces anti-MPO autoreactivity in the context of three MHC class II allomorphs. Furthermore, we show that 6PGD391-410 is immunogenic in humans, as healthy human and AAV patient sera contain anti-6PGD and anti-6PGD391-410 antibodies. Therefore, our results support the idea that bacterial plasmids might have a function in autoimmune disease.

Betulinic acid, isolated from the leaves of Syzygium cumini (L.) Skeels, ameliorates the proteinuria in experimental membranous nephropathy through regulating Nrf2/NF-κB pathways.

Membranous nephropathy (MN) is associated with increased oxidative stress and inflammatory markers in the kidney. Betulinic acid (BA) is a potent antioxidant and anti-inflammatory compound isolated from the leaves of Syzygium cumini (L.) Skeels. In the present study, we investigated the effects of BA on experimental MN in rats and explored the mechanisms by which it enhances antioxidant activities and resolves inflammatory condition in experimental MN. Passive Heymann nephritis (PHN) was induced in Sprague-Dawley rats by a single tail vein injection of anti- Fx1A antiserum. The rats were orally administered BA (25 and 50 mg kg -1 d -1) or dexamethasone (DEX; 0.07 mg kg-1, reference compound) for 4 weeks after the induction of PHN. Blood, urine, and kidney tissue were collected for analysis at the end of the study. Treatment of PHN rats with BA or DEX significantly attenuated renal dysfunction, histopathological alterations and reduced immune complex deposition in the kidneys. Furthermore, BA ameliorated mRNA and protein expression of NF-κB, iNOS, TNF-α, Nrf2, HO-1 and NQO1 in the kidney. BA also restored malondialdehyde level and antioxidant enzyme activities in the kidney. In a nutshell, the protective effect of BA can be explained by its anti-inflammatory and anti-oxidant activities, which in turn is due to downregulation of NF-κB pathway and activation of Nrf2. The results indicated that BA can effectively suppress experimental PHN in rats by regulating Nrf2/NF-κB pathways.

Identification of Promising Urinary MicroRNA Biomarkers in Two Rat Models of Glomerular Injury.

MicroRNAs (miRNAs) are small, noncoding RNAs that regulate protein levels posttranscriptionally. miRNAs play important regulatory roles in many cellular processes and have been implicated in several diseases. Recent studies have reported significant levels of miRNAs in a variety of body fluids, raising the possibility that miRNAs could serve as useful biomarkers. Here, changes in miRNA expression patterns are described in 2 different rodent models of glomerular injury (acute puromycin aminonucleoside nephropathy and passive Heymann nephritis). By employing 2 different modes of glomerular insult, oxidative stress and immune-mediated toxicity, miRNA changes in both isolated glomeruli as well as urine specimens allow for identification of urinary miRNA biomarkers that are suggestive of drug-induced injury specifically to the glomerulus. Subsets of glomerular urinary miRNAs associated with these different modes of glomerular toxicity seem to be dependent on the mechanism of the induced injury, while 9 miRNAs that changed early in both glomerular and urine specimens were common to both studies. We further show that the miRNAs identified as mechanism-specific early glomerular injury biomarkers target key pathways and transcripts relevant to the type of insult, while the insult-independent changes might serve as ideal glomerular injury biomarkers.

Heymann nephritis in Lewis rats.

Human membranous nephritis is a major cause of end-stage kidney disease. Active Heymann nephritis (HN) is an auto-immune model of membranous nephritis induced in Lewis rats by immunization with a crude renal tubular antigen (Fx1A) or megalin (gp330). The pathogenesis of HN is through the binding of anti-Fx1A autoantibodies to the auto-antigen expressed on glomerular epithelial cells, resulting in severe glomerular injury and proteinuria. The pathological features of HN include immune deposits in glomeruli and infiltration of glomeruli and the tubulointerstitium by macrophages and T cells. This unit describes the method of the preparation of Fx1A and the induction of HN in Lewis rats by immunization with Fx1A.

Autophagy can repair endoplasmic reticulum stress damage of the passive Heymann nephritis model as revealed by proteomics analysis.

Membranous nephropathy is a common cause of nephrotic syndrome in adults. Although many mechanisms have been proposed, whole proteomic research is still lacking. We analyzed the passive Heymann nephritis animal model using label-free quantitative proteome technology. Results showed 160 differential proteins between control and PHN model groups at days 14 and 21. The expression level of endoplasmic reticulum stress (ERS)-associated protein GRP78 and GRP94 protein was up-regulated on day 14 or 21, which was confirmed by Western blotting. The results also showed that the autophagy marker LC3 was up-regulated in the models. Furthermore, we used tunicamycin to induce ERS of podocytes in vitro to investigate the mechanism. Results of Western blotting revealed that the expression of GRP78, GRP94, and LC3 was up-regulated, while that of the cytoskeletal protein tubulin-ß was down-regulated, and immunofluorescence displayed disordered distribution of tubulin-ß. These suggest that ERS plays an important role in podocyte damage. Autophagy can repair the cytoskeleton damage caused by ERS as a protective mechanism. This provides an important basis for a thorough understanding of the mechanism of podocyte damage and the pathogenesis of membranous nephropathy.

Production of heterologous IgG antibody against Heymann nephritis antigen by injections of immune complexes.

Heterologous IgG antibody (ab) can be produced against Heymann nephritis (HN) antigen (ag) in rabbits by administering it in Freund's complete adjuvant. The developing abs reacted at high titre with rat kidney brush border (BB) regions of the renal proximal tubules in an indirect fluorescence ab test. A single IV injection of the heterologous ab into a susceptible strain of rat resulted in the localization of IgG ab to glomerular fixed ags, producing immune complex glomerular nephritis. The injected ab also reacted with the BB region of the renal proximal tubules. The aim of this experiment was to find out whether heterologous IgG ab against the HN ag can also be produced in recipient rabbits by injecting immune complexes (ICs) composed of a rat kidney tubular preparation [rat kidney fraction 3 (rKF3)] and donor rabbit-derived rabbit anti-rKF3 IgG ab. We found that anti-rKF3 IgG ab--against the BB region of the renal proximal tubules--could be induced in rabbits injected with ICs, and the resulting ab was able to initiate passive HN in rats. This was the first time a pathogenic IgG ab was produced against HN ag in rabbits without the use of adjuvant. Ab responses in recipient rabbits were achieved by ab information transfer. Recipient rabbits injected with the IC produced the same class of immunoglobulin with the same specificity against the target ag rKF3, as was present in the innoculum, namely rabbit anti-rKF3 IgG ab.

Cytochrome P450 2B1 mediates complement-dependent sublytic injury in a model of membranous nephropathy.

Membranous nephropathy is a disease that affects the filtering units of the kidney, the glomeruli, and results in proteinuria accompanied by loss of kidney function. Passive Heymann nephritis is an experimental model that mimics membranous nephropathy in humans, wherein the glomerular epithelial cell (GEC) injury induced by complement C5b-9 leads to proteinuria. We examined the role of cytochrome P450 2B1 (CYP2B1) in this complement-mediated sublytic injury. Overexpression of CYP2B1 in GECs significantly increased the formation of reactive oxygen species, cytotoxicity, and collapse of the actin cytoskeleton following treatment with anti-tubular brush-border antiserum (anti-Fx1A). In contrast, silencing of CYP2B1 markedly attenuated anti-Fx1A-induced reactive oxygen species generation and cytotoxicity with preservation of the actin cytoskeleton. Gelsolin, which maintains an organized actin cytoskeleton, was significantly decreased by complement C5b-9-mediated injury but was preserved in CYP2B1-silenced cells. In rats injected with anti-Fx1A, the cytochrome P450 inhibitor cimetidine blocked an increase in catalytic iron and ROS generation, reduced the formation of malondialdehyde adducts, maintained a normal distribution of nephrin in the glomeruli, and provided significant protection at the onset of proteinuria. Thus, GEC CYP2B1 contributes to complement C5b-9-mediated injury and plays an important role in the pathogenesis of passive Heymann nephritis.

Triptolide reduces proteinuria in experimental membranous nephropathy and protects against C5b-9-induced podocyte injury in vitro.

Membranous nephropathy is a major cause of nephrotic syndrome in adults where podocyte injuries were found to mediate the development of proteinuria. Triptolide, a major active component of Tripterygium wilfordii Hook F, has potent immunosuppressive, anti-inflammatory and antiproteinuric effects. To study its antiproteinuric properties, we established an experimental rat model of passive Heymann nephritis and a C5b-9 injury model of podocytes in vitro. Treatment or pretreatment with triptolide markedly reduced established proteinuria as well as the titer of circulating rat anti-rabbit IgG antibodies in these nephritic rats, accompanied by a reduction in glomerular C5b-9 deposits. Expression of desmin, a marker of podocyte injury, diminished after triptolide treatment, whereas quantitative analysis of mean foot process width showed that effacement of foot processes was substantially reversed. In in vitro studies we found that triptolide deactivated NADPH oxidase, suppressed reactive oxygen species generation and p38 mitogen-activated protein kinase, and restored RhoA signaling activity. Triptolide did not interfere with the formation of C5b-9 on the membrane of podocytes. Thus, triptolide reduces established heavy proteinuria and podocyte injuries in rats with passive Heymann nephritis, and protects podocytes from C5b-9-mediated injury.

De novo expression of podocyte proteins in parietal epithelial cells during experimental glomerular disease.

Studies have shown that certain cells of the glomerular tuft begin to express proteins considered unique to other cell types upon injury. Little is known about the response of parietal epithelial cells (PEC) to injury. To determine whether PECs change their phenotype upon injury to also express proteins traditionally considered podocyte specific, the following four models of glomerular disease were studied: the transforming growth factor (TGF)-beta1 transgenic mouse model of global glomerulosclerosis, the adriamycin model of focal segmental glomerulosclerosis (FSGS), the anti-glomerular basement membrane (GBM) model of crescentic glomerulonephritis, and the passive Heymann nephritis model of membranous nephropathy. Double immunostaining was performed with antibodies to podocyte-specific proteins (synaptopodin and Wilms' tumor 1) and antibodies to PEC specific proteins (paired box gene 8 and claudin-1). No double staining was detected in normal mice. In contrast, the results showed a statistical increase in the number of cells attached to Bowman basement membrane that were double-positive for both podocyte/PEC proteins in TGF-beta1 transgenic, anti-GBM, and membranous animals. Double-positive cells for both podocyte and PEC proteins were also statistically increased in the glomerular tuft in TGF-beta1 transgenic, anti-GBM, and FSGS mice. These results are consistent with glomerular cells coexpressing podocyte and PEC proteins in experimental glomerular disease, but not under normal circumstances.

Acute poststreptococcal glomerulonephritis: an update.

PURPOSE OF REVIEW: Acute poststreptococcal glomerulonephritis, the most common form of acute glomerulonephritis in children, continues to be a major concern worldwide. This review summarizes the recent advances in the pathogenesis, host susceptibility factors, diverse clinical presentations, and treatment of the condition. RECENT FINDINGS: Several recent advances have been made in identifying streptococcal antigens that may play a pathogenic role in acute poststreptococcal glomerulonephritis. Nephritis-associated streptococcal plasmin receptor and streptococcal pyrogenic exotoxin B are currently considered major putative nephritogens. Host susceptibility factors including HLA-DRB1*03011 have been found at a higher frequency in acute poststreptococcal glomerulonephritis patients than in healthy controls. Reversible posterior leukoencephalopathy and autoimmune hemolytic anemia are newly reported clinical associations with the disease. Studies from developing countries question whether the outcome is always benign. Treatment remains mostly conservative; however, controversy exists over the use of aggressive therapy with poor prognostic factors. SUMMARY: Severe group A streptococcal disease including acute poststreptococcal glomerulonephritis remains a cause of morbidity and mortality in developing countries and among impoverished populations. Various reports on the diverse clinical manifestations that can be associated with the condition will aid physicians in prompt diagnosis and intervention, while studies focusing on better understanding of immunopathogenesis may facilitate vaccine development and prevention.

Differential capacity of anti-RAP and anti-megalin antibodies to produce progressive passive Heymann nephritis - implications for the pathogenesis of idiopathic human membranous glomerulonephritis.

Passive Heymann nephritis (PHN) induced with heterologous antisera has been described according to various criteria, which may or may not include induction of chronic disease and proteinuria. Characteristics of the glomerular immune deposits determined by the antigenic specificities of the antisera presumably account for differences in disease outcome. In this study, the clinical and immunohistological features in the model produced with monospecific antisera were compared against megalin or receptor associated protein (RAP), two proteins that have been implicated as target antigens in PHN. Rats injected with either anti-megalin or anti-RAP antiserum developed typical glomerular immune deposits of PHN when examined after 7 days. Although the deposits stained for complement, none of the animals had abnormal proteinuria in this time frame. Over a longer time course (7-16 weeks), immune deposits persisted and proteinuria increased to pathological levels in all animals injected with anti-megalin serum. By contrast, immune deposits had cleared from the kidneys of rats injected with anti-RAP antiserum when examined at 7-8 weeks post-injection and the proteinuria levels observed up to 13 weeks remained in the normal range. Additional doses of anti-RAP antiserum given 4 and 17 days after the first injection did not prolong the duration of glomerular immune deposits. These results demonstrate a clear divergence in pathogenic potential of antisera generated against the two renal antigens, which suggest differences in the immune deposits linked to a soluble antigen that is non-covalently bound to the podocyte membrane versus those linked to an integral membrane antigen. These observations could provide clues to the nature of the unknown glomerular autoantigen of idiopathic membranous glomerulonephritis in humans.

Effect of rat kidney fraction 3 (rKF3) antigen and specific IgM antibody against rKF3 on the progression of slowly progressive Heymann nephritis.

The aim of the present study was to find out if specific IgM (M) antibody (directed against rat kidney fraction 3 (rKF3)) or rKF3 antigen were able to influence disease progression in an experimental autoimmune kidney disease called slowly progressive Heymann nephritis (SPHN). The level of circulating autoantibodies (aabs) and the morphological and functional changes to the kidney were studied in six groups of rats. All of the treatment components (except post-treatment with M) used in the SPHN pre- and post-treated rats and post-treated-only rats had measurable beneficial effects (even during restimulation with the chemically modified renal antigen, 22 weeks after the induction of the disease) as demonstrated by diminished pathogenic IgG aab production, less severe kidney lesions, and proteinuria reductions. The injected rKF3 minimized progression best in this experiment, especially when administered in a pre- and post-treatment regimen. It is thought that the effect of rKF3 in the reduced progression of SPHN was due to increased production of specific IgM aabs, which in turn limited pathogenic aab production and continuous buildup of immune complexes in the glomeruli by facilitating removal or blockage of nephritogenic autoantigens from the circulation.

Downregulation of a pathogenic autoantibody response by IgM autoantibodies directed against the nephritogenic antigen in slowly progressive Heymann nephritis.

The purpose of the study was to find out if a new modified vaccination technique would be effective in downregulating immunopathological events during the course of an experimental autoimmune kidney disease (which is morphologically and functionally similar to Heymann nephritis) called 'slowly progressive Heymann nephritis' (SPHN). We have shown that the pathogenic IgG autoantibody (aab)-induced experimental autoimmune kidney disease process can be downregulated early on as well as during the chronic progressive phase, when rats were restimulated. The IgM aab, resulting from stimulation by immune complexes made up of rat kidney fraction 3 (rKF3) antigen and rat anti-rKF3 IgM antibody in antigen excess (MIC), can greatly diminish pathogenic aab production by removing or blocking nephritogenic antigens. Reduced IgG aab production limits the formation of damaging immune complexes (IC) in the glomeruli and development of proteinuria. At the end of the experiment 60% and 80% of the MIC-treated groups had no pathogenic IgG aab in their circulation, while all the untreated SPHN rats had high levels of IgG aab associated with disease progression manifesting in increased proteinuria and severe immune complex glomerulonephritis.

Fucoidan inhibits the development of proteinuria in active Heymann nephritis.

Fucoidan, the sulphated polysaccharide extracted from brown seaweed, has various biological activities. The effect of fucoidan on the formation of proteinuria and renal functions in active Heymann nephritis was investigated in this study. Active Heymann nephritis was induced by administering brush border protein of rat proximal uriniferous tubules (FX1A). Fucoidan was administered by oral intubation to Heymann nephritis rats at three doses (50, 100 and 200 mg/kg) once daily for 4 weeks. The elevated urinary protein excretion and plasma creatinine due to the induction of Heymann nephritis were significantly reduced by fucoidan at doses of 100 and 200 mg/kg. The results indicated that fucoidan has a renoprotective effect on active Heymann nephritis and is a promising therapeutic agent for nephritis.

[The effect of splenic lymphocytes from oral immune tolerance rats on activity of NF-kappaB p65 in glomerular mesangial cells].

AIM: To explore the effect of splenic lymphocytes from oral immune tolerance rats on NF-kappaB p65 activity in glomerular mesangial cells. METHODS: 10 female Wistar rats with 6-8 weeks of age were divided randomly into two groups, 5 rats for each group. The rats of one group were given through intragastric feeding with 2 mL Fx1A antigen to induce oral immune tolerance, and the rats of the other group only with PBS (as control group). Two weeks later, the rats were sacrificed, and then their spleens were excised and lymphocytes were separated routinely for antigen-specific lymphocytes reaction. The culture supernatants of splenic lymphocytes from oral immune tolerance rats were prepared. The IL-10 level in the supernatants was detected by sandwich ELISA. After the cultured mesangial cells were stimulated with high dose of insulin, the culture supernatants of rat's splenic lymphocytes were added to insulin-treated mesangial cells. Twenty-four hours later, the NF-kappaB p65 activity in mesangial cells was detected by immunohistochemical staining and sandwich ELISA. RESULTS: As compared with control group, the proliferation reaction of splenic lymphocytes from oral immune tolerance rats was inhibited obviously, and IL-10 level in splenic lymphocyte culture supernatants from the tolerance rats rose markedly (P<0.01). NF-kappaB p65 activity induced by insulin in mesangial cells was inhibited by culture superatants of splenic lymphocytes from oral immune tolerance rats (P<0.05). CONCLUSION: NF-kappaB p65 activity in mesangial cells may be inhibited by IL-10 secreted by splenic lymphocytes from the oral immune tolerance rats.

Production of Heymann nephritis by a chemically modified renal antigen.

An autoimmune kidney disease morphologically and functionally similar to Heymann nephritis (HN) was induced in mature male Sprague Dawley rats by repeated weekly IP injections of a chemically modified azo sonicated ultracentrifuged (u/c) rat kidney fraction 3 (rKF3) antigen in an aqueous medium. The experiment was terminated 15 weeks after the first injection of the chemically altered antigen. Serum samples collected and analysed by an indirect fluorescent antibody test on normal rat kidney sections during the course of the experiment showed a gradual rise in circulating pathogenic autoantibodies directed against the proximal tubular brush border regions. Proteinuria was present and significantly increased in the urine of two of eight rats. The arising immune-complex glomerulonephritis (ICGN) revealed typical HN kidney disease lesions in 70% of the rats in histological, direct fluorescent antibody and electron-microscopical examinations. Control rats injected similarly with the an unmodified version of the same antigen did not develop the HN-characteristic morphological and functional changes. To our knowledge, this is the first time that the autoimmune kidney disease designated as an active HN has been produced by the administration of a chemically altered renal antigen in an aqueous solution and not by the usual presentation of the nephritogenic renal antigen in an adjuvant.

DNA vaccination against specific pathogenic TCRs reduces proteinuria in active Heymann nephritis by inducing specific autoantibodies.

We have previously identified potential pathogenic T cells within glomeruli that use TCR encoding Vbeta5, Vbeta7, and Vbeta13 in combination with Jbeta2.6 in Heymann nephritis (HN), a rat autoimmune disease model of human membranous nephritis. Vaccination of Lewis rats with naked DNA encoding these pathogenic TCRs significantly protected against HN. Proteinuria was reduced at 6, 8, 10, and 12 wk after immunization with Fx1A (p < 0.001). Glomerular infiltrates of macrophages and CD8(+) T cells (p < 0.005) and glomerular IFN-gamma mRNA expression (p < 0.01) were also significantly decreased. DNA vaccination (DV) causes a loss of clonality of T cells in the HN glomeruli. T lymphocytes with surface binding of Abs were found in DNA vaccinated rats. These CD3(+)/IgG(+) T cells expressed Vbeta5 and Vbeta13 that the DV encoded. Furthermore, FACS shows that these CD3(+)/IgG(+) cells were CD8(+) T cells. Analysis of cytokine mRNA expression showed that IL-10 and IFN-gamma mRNA were not detected in these CD3(+)/IgG(+) T cells. These results suggest that TCR DNA vaccination produces specific autoantibodies bound to the TCRs encoded by the vaccine, resulting in blocking activation of the specific T cells. In this study, we have shown that treatment with TCR-based DV, targeting previously identified pathogenic Vbeta families, protects against HN, and that the mechanism may involve the production of specific anti-TCR Abs.