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Well-characterized human glomerular mesangial cells were stimulated in vitro with interferon-gamma (IFN-gamma), interleukin-1, and tumor necrosis factor alpha (TNF-alpha) to determine the influence of these cytokines on the expression of class I and II HLA antigens. IFN-gamma induced a dose-dependent increase in expression of both class I and class II HLA antigens. TNF-alpha induced only class I antigens when used alone, but acted synergistically with IFN-gamma in the expression of class I and II antigens. Interleukin-1 had little effect on HLA antigen expression. These results raise the possibility that stimulated mesangial cells may act as antigen-presenting cells and may mediate local immune reactions in the glomerulus.

A method of assembly of a single-chain Fv fragment is described, whereby asymmetric polymerase chain reactions (APCR) and primer extension were used to join immunoglobulin heavy and light chain variable region genes via a linker sequence. In this procedure heavy and light chain genes, together with a linker gene containing complementary sequences, were amplified by APCR to generate single-stranded products. The single stranded heavy or light chain genes were hybridized to the relevant single-stranded link product, and extended to produce double-stranded heavy-link and link-light genes. These genes then underwent another round of APCR, resulting in two single-stranded genes (heavy-link and link-light) containing extensive overlapping sequences. Hybridization and extension of these two single-stranded products allowed the formation of the complete heavy chain-link-light chain double-stranded product. Using this method, a functional single-chain Fv fragment based on the pan-leukocyte antibody WM65 was expressed and purified from E. coli. This method of immunoglobulin gene assembly by polymerase chain reaction (PCR) offers an alternative to the current methods of the genetic engineering of antibody fragments.

We examined the expression of HLA antigens on post-mortem human uveal tissues and on cultured uveal cells. There was no in vivo expression of class I or class II antigens in these tissues, except for the blood vessel endothelium which expressed class I but not class II antigens. Tissue cultured uveal cells were found to express class I antigens. The effect of interferon-alpha and interferon-gamma on the in vitro expression of these antigens was examined. Class I HLA antigen expression was enhanced by both interferon-alpha and interferon-gamma, while class II HLA antigens were induced on up to 100% of cultured cells, but only by interferon-gamma. These findings are discussed in their relation to immune mechanisms occurring in ocular inflammatory disorders.

The relationship between the nature and severity of inflammatory eye disease was analyzed with respect to HLA antigens and alpha 1-antitrypsin phenotypes. Using standard ophthalmologic criteria, we divided patients with anterior uveitis into acute, chronic (greater than 3-month duration), bilateral, or recurrent disease. There was a significantly increased incidence of alpha 1-antitrypsin-deficient phenotypes in anterior uveitis, especially in those patients with severe (chronic, bilateral, or recurrent) disease. HLA-B27 acts as an independent predisposing factor: it was present in 22% of patients with their first attack of acute uveitis compared with 51% of patients with recurrent disease. Together, these genetic factors are present in 63% of patients with severe anterior uveitis and represent the most significant predisposing and prognostic factors so far detected.

The relationship between HLA B27 anterior uveitis (AU) and evidence of chlamydia trachomatis infection was investigated in 35 consecutive patients attending a uveitis research clinic, using a complement fixation test and lymphocyte transformation test to chlamydia (ornithosis-psittacosis) group antigen. A significant stimulation index (SI greater than 2) was found in 73% (11/15) of the HLA B27 positive patients compared with the HLA B27 negative AU patients and controls (P less than 0.03). There was no difference between AU patients with and without associated rheumatic disease or in terms of antibody response. The results of the present study indicate a significant relationship between the cell-mediated immune response to chlamydia group antigen and the HLA B27 positive subgroup of patients with AU.

The chemiluminescent (CL) associated phagocytic response of peripheral blood monocytes to two serovars of Chlamydia trachomatis, Shigella flexneri and zymosan was assessed in a group of 26 patients with anterior uveitis (AU). HLA-B27+ patients with AU, when compared to HLA-B27- patients with AU and appropriate controls, had a significantly decreased CL response to C. trachomatis but no difference between groups in the response to S. flexneri and zymosan. The decreased chemiluminescent (phagocytic) response of mononuclear phagocytes to Chlamydiae may indicate an important abnormality in the pathogenesis of HLA-B27+ AU.

The eye is a common site for autoimmune inflammatory diseases, many of which are linked to HLA antigens. The role of these HLA phenotypes in the disease process is presently unknown. In view of the importance of HLA antigens to immune responses, a knowledge of the distribution of HLA antigens in ocular tissues may aid in our understanding of the role of these antigens in disease production or susceptibility. Apart from the cornea, the expression of HLA antigens in human ocular tissues has not been thoroughly investigated. In this study we examined the in vivo expression of HLA antigens in postmortem human eyes using immunohistochemical techniques. The majority of ocular cells were found not to express class I or class II HLA antigens, with the exception of the blood vessel endothelium, which was uniformly class I-positive. The significance of these results is discussed in relation to ocular immunity.

Analysis of the immunological features of anterior uveitis (AU) revealed a dichotomy of abnormalities defined in terms of the HLA-B27 status of the patient. HLA-B27-positive AU was characterised by the occurrence of iris autoantibodies and an absolute T cell lymphopenia during active disease which returned to normal with recovery. This phenomenon was not observed in HLA-B27-negative AU or in controls and could not be attributed to antilymphocyte antibodies as these were not detected. Furthermore, there were no changes in T-cell subsets (helper and suppressor T lymphocytes). Compared with HLA-B27-positive AU patients, the HLA-B27-negative group demonstrated elevated IgE levels and increased prevalence of smooth muscle autoantibodies.

HLA antigens are associated with a number of inflammatory eye diseases, most notably HLA B2 with anterior uveitis (AU). This association varies between different populations and ethnic groups. The aim of this study was to investigate the relationship between uveitis and HLA A, B and DR locus antigens in an Australian population. Seventy-two consecutive patients with uveitis were studied (37 males and 35 females) over a 6 month period. Thirty-two percent of the AU patients were HLA B27+, as were 42% of males (19% females) with their first attack of AU compared with 60% of males (23% females) with recurrent AU. The only significant difference in etiology between males and females was the greatly increased incidence of rheumatic diseases in males, in whom 77% (10/13) had radiological evidence of sacroiliitis. Additional findings included a lack of association between the HLA B7 cross reactive group and DR locus antigens in AU as well as the lack of any HLA associations in the 13 patients with posterior uveitis (PU).

We have previously reported the association of Chlamydia trachomatis with HLA B27+ related diseases. To investigate the possibility that chlamydial antibodies serve to localize the immune response in such diseases, we examined the crossreactivity of chlamydial antibodies (rabbit anti-D and anti-L2 serotypes) with peripheral blood mononuclear cells of patients with ankylosing spondylitis (AS) and anterior uveitis (AU) and with human and bovine ocular tissue and cells in culture. Our results indicate a significantly increased percentage binding of chlamydial antibody (D serotype) to the mononuclear cells of HLA B27+ patients with AS when compared with HLA B27- patients with AS (12.9% +/- 2.2 versus 5.4% +/- 2.2), B27+ controls (5.5% +/- 1.5) and B27- controls (6.1% +/- 1.0). There was no significant difference between controls and HLA B27+ patients with AU (6.6% +/- 1.9) and B27- patients with AU (8.7% +/- 1.1). This crossreactivity could not be blocked by monoclonal HLA B27 antibody. Chlamydial antibodies (D and L2) crossreact with human and bovine conjunctiva but not uvea, tissue culture derived iris fibroblasts or smooth muscle cells. Our results provide additional support for the concept of crossreactivity between antibodies to microbial agents and peripheral blood mononuclear cells of patients with HLA B27+ AS.

In previous studies we have shown that normal human uveal cells, with the exception of vascular endothelium, do not express class I or class II HLA antigens in vivo. Class I antigens are induced in vitro by a variety of cytokines, while class II antigens are only induced by gamma interferon. In this study we examine the capacity of cultured uveal cells, rendered class II HLA antigen positive by gamma interferon, to present antigen to T cells. Cultured uveal cells were found to present antigen (tetanus toxoid, PPD, and Candida albicans) to T cells, but only when they were pretreated with gamma interferon. This function of uveal cells was antigen specific and MHC restricted and was blocked by class II-specific monoclonal antibodies, indicating the crucial role of class II HLA antigens in antigen presentation.