Stimulation of whole blood cultures in patients with ankylosing spondylitis by a mitogen derived fromMycoplasma arthritidis (MAS) and other mitogens

Abstract

In this study we compared cytokine production and cell proliferation of immunocompetent cells derived from patients with ankylosing spondylitis (AS) to those from healthy blood donors using a whole blood assay. To this end, blood cell cultures were stimulated with the superantigens MAS (Mycoplasma arthritidis supernatant) and staphylococcal enterotoxin B (SEB) and the plant lectins phytohaemagglutinin (PHA) and concanavalin A (Con A). The number of white blood cells (WBC) and lymphocyte subsets were also determined. Cell proliferation and levels of interferon-gamma (IFN-gamma), interleukin-1 beta (IL-1 beta) and interleukin-6 (IL-6) were measured after stimulation with the different mitogens. An ELISA test was used to analyse supernatant cytokine levels. Individuals with AS showed significantly lower IFN-gamma concentrations and markedly lower cell proliferation rates with all tested mitogens than healthy controls, while there was no significant difference in IL-6 synthesis. IL-1 beta levels were slightly impaired in the patient group, but only blood cell cultures stimulates with MAS showed a statistical significance. Furthermore, there was a significant elevation of leucocytes and lymphocytes in patients with AS resulting in higher numbers of CD4-positive cells, which implies a higher CD4:CD8 cell ratio. CD19- and CD8-positive cells were not significantly distinct compared to healthy controls. This deviation in cytokine levels and cell proliferation points to a suppression of T lymphocytes. A disturbed T-lymphocyte function may play a part in the pathogenesis of AS.

Low T cell production of TNFalpha and IFNgamma in ankylosing spondylitis: its relation to HLA-B27 and influence of the TNF-308 gene polymorphism

“…Our study is in accordance with an earlier study which showed a lower production of IFNγ in patients with AS compared with healthy HLA-B27 negative controls 34. No difference in TNFα levels between HLA-B27 positive patients with ReA and healthy B27 negative controls was found in a Finnish study when lipopolysaccharide stimulated monocytes were used 35.…”

A model of human whole blood lymphokine release for in vitro and ex vivo use

“…For example after 24 h of incubation PHA induced 0.62 ± 0.14 ng/ml TNFa (SEB: 0.57 ± 0.12 ng/ml TNFa). This is in line with previous publications which show a low release of monokines such as IL-6 and IL-110 for stimulation of whole blood with SEB or PHA even after 6~8 days of incubation (Brand et al, 1992(Brand et al, , 1997. However, in those studies, additional SEB-induced lymphokines were not addressed.…”

“…The different types of whole blood lymphokine release assays have been shown to be of value for the evaluation of immune function in a broad range of patient groups, including cancer (Szopinski et al, 1999) , rheumatoid arthritis (Swaak et al, 1997), tuberculosis (van Crevel et al, 1999 ;Hussain et al, 2002), HIV (Hartung et al, 1998 ;Vingerhoets et al, 1998) and ankylosing spondylitis (Brand et al, 1997). However, since very different stimuli and experimental conditions were chosen, these studies are difficult to compare.…”

“…Here, the prerequisites and characteristics of a human whole blood lymphokine release model employing such exotoxins are defmed extending earlier approaches by our own group and others (Brand et al, 1992(Brand et al, , 1997Hartung et al, , 1999Baum et al, 1999 ;Westerholt et al, 2000) . The model appears to be useful in defining the immune characteristics ofdonors, states of disease and the effect of immunomodulatory and/or immunotoxic agents.…”

Cytokines and the immunopathology of the spondyloarthropathies

“…The IFN-g secretion of patients with AS in the supernatant after stimulation with mycoplasma arthritidis was found to be enhanced in one study [63] and diminished in another study [64]. A low IFN-g and a high IL-10 secretion has been correlated with an impaired immune response to Klebsiella pneumoniae [65].…”