The effects of atorvastatin on gluten-induced intestinal T cell responses in coeliac disease

Abstract

Various experimental models suggest that the cholesterol-lowering drugs statins may also modulate immune responses. Cellular level studies on human disorders are needed, however, to provide a rational basis for clinical testing of statins as immune therapy. Coeliac disease, a chronic small intestinal inflammation driven by HLA-DQ2 restricted mucosal T cells that are specific for ingested wheat gluten peptides, is in many ways ideal for this purpose. In addition, there is a need for alternative treatment to the gluten-free diet in this disorder. Here we have assessed the effects of atorvastatin on gluten-reactive T cells, dendritic cells and the coeliac mucosa by in vitro culture of biopsies. Atorvastatin inhibited gluten-induced proliferation and specific cytokine production of human intestinal gluten-reactive T cell clones and lines. Dendritic cells exposed to atorvastatin displayed a reduced expression of the costimulatory molecule CD83 upon maturation with lipopolysaccharide. Incubation of intestinal biopsy specimens with atorvastatin in vitro, however, did not influence gluten-induced cytokine release. In conclusion, atorvastatin has specific effects on isolated gluten-reactive T cells and dendritic cells, but does not shut down the gluten-induced production of proinflammatory cytokines in intestinal biopsies.

Fig. 1

Atorvastatin effectively inhibits proliferation of T cell clones, but has less effect on cytokine secretion by the same T cells. (a) Four T cell clones were stimulated by anti-T cell receptor (TCR) α/β antibody coated onto plates in the presence of varying concentrations of atorvastatin. (b) Four T cell clones were stimulated by a B lymphoblastoid cell line (B-LCL) that had been incubated with tissue transglutaminase (TG2)-treated gluten in the presence of varying concentrations of atorvastatin in the presence or absence of mevalonolactone (100 µ M). Mean ± s.e.m. of duplicates. Proliferation and cytokine production was measured in the same experiment after 3 days. AS = atorvastatin (µM), M = mevalonolactone, ND = not done in the presence of both atorvastatin and mevalonolactone.

Fig. 2

Atorvastatin profoundly reduces proliferation and cytokine secretion of gluten-reactive polyclonal T cell lines. (a) Four T cell lines were stimulated by a B lymphoblastoid cell line (B-LCL) that had been incubated with tissue transglutaminase (TG2)-treated gluten in the presence of varying concentrations of atorvastatin (AS) in the presence or absence of mevalonolactone (100 µ M). Mean ± s.e.m. of duplicates.

Fig. 3

Effect of atorvastatin on T cell viability. Apoptosis of T cells was measured after 3 days of stimulation, at the same time as proliferation of the cells. Percentage of viable cells represent cells that were neither stained by annexin V nor by propidium iodide. Mean ± s.e.m. of five experiments. Atorvastatin did not induce significant apoptosis of the T cells (P > 0·9 in every group compared to samples without atorvastatin). Experiments were conducted on TCL437.1.3, TCC387.E9, TCC430.1.135 and TCC412.5.28 with gluten stimulation and on TCC430.1.135 also with anti-T cell receptor (TCR) α/β stimulation.

Fig. 4

Apoptosis and expression of CD83 and CD86 by dendritic cells in the presence of atorvastatin. Monocyte-derived dendritic cells were differentiated and matured by lipopolysaccharide (LPS) in the presence or absence of atorvastatin, assessed by FACS analysis. (a) Rate of apoptosis in immature (white bars) and mature (black bars) dendritic cells differentiated in the presence of varying concentrations of atorvastatin. Mean ± s.e. of three experiments. (b) Histogram of CD83 expression of mature dendritic cells, representative of seven experiments. P = 0·036. (c) Histogram of CD86 expression of mature dendritic cells, representative of four experiments.

Fig. 5

Concentrations of interferon (IFN)-γ, interleukin (IL)-2, tumour necrosis factor (TNF)-α, IL-10, and IL-5 in the supernatants of duodenal biopsies from coeliac patients incubated for 24 h in medium or 2 mg/ml gluten in the presence or absence of 10 µ M atorvastatin. Twelve biopsy specimens of seven patients are shown; individual markers designate specimens of the same patient. The horizontal lines represent the median.