Ya Huan Lou

CD4+ T cells specific to a glomerular basement membrane antigen mediate glomerulonephritis

Ab-mediated mechanisms have been considered the major causes of glomerulonephritis (GN). However, recent studies suggest that T cells may be more important in mediating GN. To investigate the effects of antigen-specific CD4(+) T cells, we generated Th1 cell lines specific for this antigen from rats that had been immunized with a recombinant form of the glomerular basement membrane (GBM) antigen, Col4alpha3NC1. Upon the transfer of in vitro-activated T cell lines to pertussis toxin-primed, naive syngeneic rats, the recipients developed severe proteinuria/albuminuria, which plateaued after approximately 35 days. Although no IgG binding to GBM or C3 deposition could be detected by immunofluorescence, five out of eleven rats exhibited severe GN, as judged by the formation of characteristic crescent-shaped lesions in the glomeruli, whereas the others exhibited modest GN. Thus Col4alpha3NC1-specific T cells directly initiated glomerular injury in the recipients. One notable difference from GN induced by active immunization was a T cell infiltration in the renal interstitium, which affected some tubules. We therefore injected fluorescence-labeled Col4alpha3NC1-specific into naive rats, and we found that they were enriched 4.5-fold in the kidney cortex relative to nonspecific control T cells 24 hours later. Many of the T cells were located in the Bowmans space and had a flattened shape, suggesting that the primary target for the T cells was in or adjacent to the Bowmans capsule.

Glomerulonephritis Induced by Recombinant Collagen IVα3 Chain Noncollagen Domain 1 Is Not Associated with Glomerular Basement Membrane Antibody: A Potential T Cell-Mediated Mechanism

Glomerulonephritis is believed to result commonly from Ab-mediated glomerular injury. However, Ab-associated mechanisms alone cannot explain many cases of human glomerulonephritis. We developed a rat model of human anti-glomerular basement membrane (GBM) disease to investigate T cell and Ab response, and their associations with the disease. A single immunization of highly denatured recombinant mouse collagen IVα3 chain noncollagen domain 1 (rCol4α3NC1) induced severe glomerulonephritis in 100% of Wistar Kyoto rats, 33% of which died of this disease around day 35 postimmunization. The renal pathology demonstrated widespread glomerular damage and a mononuclear cell infiltration within the interstitial tissue. T cells from immunized rats responded not only to rCol4α3NC1, but also to isolated rat GBM. Sera Abs to rCol4α3NC1 were detectable in 100% of the rats, but only 20% of the rats had low levels of Ab to isolated rat GBM by Western blot, and none by immunofluorescence. Furthermore, IgG/M binding to or C3 deposition on endogenous GBM in immunized rats were not detected in most of the experimental rats, and showed no statistical correlation with disease severity. Additionally, no electronic dense deposition in the glomeruli was detected in all rats. Those data revealed a disassociation between the disease and anti-GBM Ab. T cell-mediated mechanisms, which are currently under our investigation, may be responsible for the glomerular disease.

Anti-Inflammatory and Renal Protective Actions of Stanniocalcin-1 in a Model of Anti-Glomerular Basement Membrane Glomerulonephritis

We have previously shown that stanniocalcin-1 (STC1) inhibits the transendothelial migration of macrophages and T cells, suppresses superoxide generation in macrophages, and attenuates macrophage responses to chemoattractants. To study the effects of STC1 on inflammation, in this study we induced a macrophage- and T-cell-mediated model of anti-glomerular basement membrane disease in STC1 transgenic mice, which display elevated serum STC1 levels and preferentially express STC1 in both endothelial cells and macrophages. We examined the following parameters both at baseline and after anti-glomerular basement membrane antibody treatment: blood pressure; C(3a) levels; urine output; proteinuria; blood urea nitrogen; and kidney C(3) deposition, fibrosis, histological changes, cytokine expression, and number of T cells and macrophages. Compared with wild-type mice, after anti-glomerular basement membrane treatment STC1 transgenic mice exhibited: i) diminished infiltration of inflammatory macrophages in the glomeruli; ii) marked reduction in crescent formation and sclerotic glomeruli; iii) decreased interstitial fibrosis; iv) preservation of kidney function and lower blood pressure; v) diminished C(3) deposition in the glomeruli; and vi) reduced expression of macrophage inhibitory protein-2 and transforming growth factor-beta2 in the kidney. Compared with baseline, wild-type mice, but not STC1 transgenic mice, had higher proteinuria and a marked reduction in urine output. STC1 had minimal effects, however, on both T-cell number in the glomeruli and interstitium and on cytokine expression characteristic of either TH1 or TH2 activation. These data suggest that STC1 is a potent anti-inflammatory and renal protective protein.

T Cell Epitope Mimicry in Antiglomerular Basement Membrane Disease

Antiglomerular basement membrane (GBM) disease or Goodpasture’s syndrome is among the earliest recognized human autoimmune diseases. Although collagen 4α3 NC1 (Col4α3NC1) has been identified as the responsible autoantigen, it remains unknown how autoimmunity to this autoantigen is provoked. We have demonstrated in our rat model that a single nephritogenic T cell epitope pCol28–40 of Col4α3NC1 induces glomerulonephritis. We hypothesized that microbial peptides that mimic this T cell epitope could induce the disease. Based on the critical residue motif (xxtTxNPsxx) of pCol28–40, seven peptides derived from human infection-related microbes were chosen through GenBank search and synthesized. All peptides showed cross-reactivity with pCol28–40-specific T cells at various levels. Only four peptides induced transient proteinuria and minor glomerular injury. However, the other three peptides induced severe proteinuria and modest to severe glomerulonephritis in 16–25% of the immunized rats. Unexpectedly, the most nephritogenic peptide, pCB, derived from Clostridium botulinum, also induced modest (25%) to severe (25%) pulmonary hemorrhage, another important feature of anti-GBM disease; this was not correlated with the severity of glomerulonephritis. This finding suggests that subtle variations in T cell epitope specificity may lead to different clinical manifestations of anti-GBM disease. In summary, our study raises the possibility that a single T cell epitope mimicry by microbial Ag may be sufficient to induce the anti-GBM disease.

Regulatory T-cell, endogenous antigen and neonatal environment in the prevention and induction of autoimmune disease

Summary: Recent studies on autoimmune ovarian disease (AOD) induced by thymectomy on d3 (d3tx), and AOD induced by immunization with the ovary‐specific zona pellucida 3 peptide (pZP3), have yielded the following results. First, female tolerance to pZP3 depends on the persistence of endogenous antigen (Ag). Second, following regulatory T‐cell depletion, endogenous Ag in prepubertal d3tx mice triggers AOD and drives disease progression. Third, endogenous ZP3 from ovaries without AOD stimulates a diversified IgG autoantibody (autoAb) response that rapidly follows pZP3 T epitope immunization. Fourth, induction of AOD and autoimmune memory in neonatal female mice by pZP3 in incomplete Freunds adjuvant depends on endogenous Ag stimulation within the neonatal week. Fifth, in a rodent pinworm‐positive environment, neonatal but not adult female mice injected with pZP3 in water develop Th2‐mediated AOD and Th2 memory. Sixth, neonatal T cells transfer AOD to syngeneic athymic recipients, whereas adult T cells are non‐pathogenic and in fact suppress AOD conferred by neonatal T cells. Therefore: 1) the continuous presence of physiologically‐expressed autoAg is critical for both tolerance maintenance and autoimmune disease pathogenesis; the outcome is determined by the integrity of regulatory T cells; and 2) the neonatal mice, deficient in the regulatory T‐cell function, are more responsive than adults to Ag and environmental stimuli that promote autoimmune disease and memory.

Retargeting T Cell-Mediated Inflammation: A New Perspective on Autoantibody Action

To understand the pathogenesis of organ-specific autoimmune disease requires an appreciation of how the T cell-mediated inflammation is targeted, and how the organ function is compromised. In this study, autoantibody was documented to influence both of these parameters by modulating the distribution of T cell-mediated inflammation. The murine autoimmune ovarian disease is induced by immunization with the ZP3330–342 peptide of the ovarian zona pellucida 3 glycoprotein, ZP3. Passively transferred or actively induced Ab to ZP3335–342 bound to the zona pellucida in the functional and degenerative ovarian follicles, and the ovaries remained histologically normal. Transfer of ZP3330–342 peptide-specific T cells targeted the degenerative follicles and spared the functional follicles, and the resultant interstitial oophoritis was associated with unimpaired ovarian function. Unexpectedly, the coexistence of ZP3330–342 peptide-specific T cells and zona-bound autoantibody led to a dramatic translocation of the ovarian inflammation to the growing and mature ovarian follicles, with destruction of the ovarian functional unit. Ab retargeted both Th1-induced mononuclear inflammation and Th2-induced eosinophilic inflammation, and retargeting was induced by murine and rat polyclonal Abs to multiple distinct native B cell determinants of the zona pellucida. Therefore, by reacting with the native determinants in tissue Ag, Ab alters the distribution of T cell-mediated inflammation, and results in destruction of the functional units of the target organ. We propose that this is a clinically important and previously unappreciated element of Ab action in autoimmune disease.

A Self T Cell Epitope Induces Autoantibody Response: Mechanism for Production of Antibodies to Diverse Glomerular Basement Membrane Antigens

The anti-glomerular basement membrane (GBM) Ab has been regarded as a prototypical example of pathogenic autoantibodies. However, the mechanism for elicitation of this Ab remains unknown. In the present paper, we report that the Ab to diverse GBM Ags was induced by a single nephritogenic T cell epitope in a rat model. The T cell epitope pCol28–40 of noncollagen domain 1 of collagen type IV α3 chain not only uniformly induced severe glomerulonephritis but also elicited anti-GBM Ab in 76% of the immunized rats after prominent glomerular injury. Furthermore, we demonstrated that the anti-GBM Ab was not related to the peptidic B cell epitope nested in pCol28–40; that is, 1) elimination of the B cell epitope, either by substitution of the critical residues of the B cell epitope or by truncation, failed to abrogate anti-GBM Ab production, and 2) the anti-GBM Ab, eluted from the diseased kidneys, reacted only with native GBM, but not with pCol28–40. Confocal microscopy and immunoprecipitation further demonstrated that the eluted anti-GBM Ab recognized conformational B cell epitope(s) of multiple native GBM proteins. We conclude that autoantibody response to diverse native GBM Ags was induced by a single nephritogenic T cell epitope. Thus, anti-GBM Ab may actually be a consequence of T cell-mediated glomerulonephritis.

Autoimmune ovarian disease: mechanism of disease induction and prevention.

Investigation of the versatile models for autoimmunity of the ovary and other selected organs has contributed to our understanding of the following aspects of autoimmunity: the mechanism of T cell molecular mimicry; T-->B epitope spreading, as a basis for autoantibody diversification, and as a link between organ-specific and systemic autoimmunity; the localization of genetic loci potentially influencing multiple autoimmune diseases; and the elucidation of regulatory T cells as a component of physiological self tolerance.

Repression of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) but not its receptors during oral cancer progression

BackgroundTRAIL plays an important role in host immunosurveillance against tumor progression, as it induces apoptosis of tumor cells but not normal cells, and thus has great therapeutic potential for cancer treatment. TRAIL binds to two cell-death-inducing (DR4 and DR5) and two decoy (DcR1, and DcR2) receptors. Here, we compare the expression levels of TRAIL and its receptors in normal oral mucosa (NOM), oral premalignancies (OPM), and primary and metastatic oral squamous cell carcinomas (OSCC) in order to characterize the changes in their expression patterns during OSCC initiation and progression.MethodsDNA microarray, immunoblotting and immunohistochemical analyses were used to examine the expression levels of TRAIL and its receptors in oral epithelial cell lines and in archival tissues of NOM, OPM, primary and metastatic OSCC. Apoptotic rates of tumor cells and tumor-infiltrating lymphocytes (TIL) in OSCC specimens were determined by cleaved caspase 3 immunohistochemistry.ResultsNormal oral epithelia constitutively expressed TRAIL, but expression was progressively lost in OPM and OSCC. Reduction in DcR2 expression levels was noted frequently in OPM and OSCC compared to respective patient-matched uninvolved oral mucosa. OSCC frequently expressed DR4, DR5 and DcR1 but less frequently DcR2. Expression levels of DR4, DR5 and DcR1 receptors were not significantly altered in OPM, primary OSCC and metastatic OSCC compared to patient-matched normal oral mucosa. Expression of proapoptotic TRAIL-receptors DR4 and DR5 in OSCC seemed to depend, at least in part, on whether or not these receptors were expressed in their parental oral epithelia. High DR5 expression in primary OSCC correlated significantly with larger tumor size. There was no significant association between TRAIL-R expression and OSSC histology grade, nodal status or apoptosis rates of tumor cells and TIL.ConclusionLoss of TRAIL expression is an early event during oral carcinogenesis and may be involved in dysregulation of apoptosis and contribute to the molecular carcinogenesis of OSCC. Differential expressions of TRAIL receptors in OSCC do not appear to play a crucial role in their apoptotic rate or metastatic progression.

Transient Expression of CC Chemokine TECK in the Ovary during Ovulation: Its Potential Role in Ovulation

Problem:  Chemokine thymus‐expressed chemokine (TECK), which is expressed exclusively in the thymus and small intestine, plays a critical role in T‐cell development. Our previous study revealed its expression in the ovary also. This study investigated its ovarian expression during ovulatory process.