NK, T and NK T-cells in ageing, coeliac disease and inflammatory bowel disease. (2008)

This thesis investigated the number and function of natural killer T-cells (NK\nT-cells) as a function of age, in coeliac disease, Crohn’s disease and ulcerative\ncolitis.\nNK T-cells are a newly appreciated class of immune cells that are able to\nregulate the activity of the broader T-cell population. NK T-cells have been\nimplicated in animal models of autoimmune disease and in human autoimmune\ndisease. A subset of NK cells express the T-cell receptor (TCR) and are termed\nNK T-cells. In humans a further small subset of NK T-cells express an invariant\nTCR α chain (Vα24Jα18) and contain the immunoregulatory cell population\nthat is distinguished from classical T-cells by promptly producing interleukin-4\n(IL-4). Invariant NK T-cells (iNK T-cells) have the surface phenotype of\nVα24+ Vβ11+ T-cells and express CD161+ NK markers. They are CD4+\n(single positive; SP) or CD4- (double negative; DN), CD1d restricted and are\nα-galactosylceramide (α-GalCer) reactive.\nNKT cells have been implicated in numerous autoimmune disorders. Early\nwork showed a major deficiency of NKT cell numbers in nonobese diabetic\n(NOD) mice, a well-established model of spontaneous, autoimmune T-cell\nmediated insulin-dependent diabetes. Both the number of NKT cells and\nfunction, as assessed by IL-4 release following TCR ligation, are dramatically\nreduced in NOD mice. NK T-cells have been implicated in other models of\nautoimmunity such as, experimental allergic encephalomyelitis (EAE). They\nhave since been investigated and shown to be deficient in a number of human\nautoimmune diseases including, systemic sclerosis (SSc), and systemic lupus\nerythematosus (SLE), multiple sclerosis, atopic asthma, atopic dermatitis,\nrheumatoid arthritis, type 1 diabetes mellitus and scleroderma. The basis of the\nwork presented within this thesis originated from the deficiency of NK T-cells\nin models of autoimmune diseases and human autoimmune diseases.\nThe initial aim of this thesis was to investigate the phenotype and function of\nVα24+ NK T-cells in normal healthy control subjects and with respect to age.\nThe original aim was to investigate whether NK cells, T-cells, NK T-like cells\nand invariant NK T-cells (iNK T-cells) are deficient in coeliac disease, Crohn’s\ndisease and/or ulcerative colitis.\nBlood was collected for flow cytometry from normal control subjects, subjects\nwith coeliac disease, Crohn’s disease and ulcerative colitis. The number of\ncirculating NK cells, T-cells, NK T-like cells and iNK T-cells was assessed by\nthree-colour flow cytometry. Intracellular cytokine production was measured\nafter in vitro anti-CD3/ anti-CD28 antibodies, gluten fraction 3 and\nPMA:ionomycin stimulation. Vα24+ T-cells were quantified in ileocolonic\nbiopsies by immunofluorescence and as mRNA by relative and real-time PCR\n(RT-PCR).\nThe number of circulating Vα24+ T-cells and iNK T-cells decrease with age in\nnormal healthy control subjects. Cytokine production was also affected by age.\nThe work of this thesis has identified a subpopulation of otherwise normal\nhealthy individuals whom have normal numbers of circulating Vα24+ T-cells,\nreduced numbers of circulating Vα24+ Vβ11+ T-cells and consequently iNK Tcells.\nCirculating CD161+ NK cells, Vα24+ T-cells and the SP subset of Vα24+ Tcells\nwere reduced in coeliac disease. The low numbers of circulating Vα24+\nT-cells was independent of diet. The number of circulating Vα24+ Vβ11+ Tcells\nwere reduced in coeliac disease, and as a consequence, the number of\ncirculating Vα24+ Vβ11+ α-GalCer/CD1d tetramer+ and Vα24+ 6B11+ iNK\nT-cells were reduced. The deficiency of Vα24+ T-cells was not confined to the\nblood, but observed within the intestinal mucosa. Intestinal Vα24 mRNA\nexpression from subjects with coeliac disease was reduced compared to levels\nin normal subjects as assessed by relative and RT-PCR. Thus, Vα24+ T-cells\nwere deficient in coeliac disease both systemically and mucosally. Cytokine production by Vα24+ T-cells, 6B11+ and Vα24+ α-GalCer/CD1d tetramer+\niNK T-cells after 4 h in vitro anti-CD3 stimulation was also impaired in\nsubjects with coeliac disease.\nCirculating CD56+, CD57+, CD94+, CD161+ NK cells were reduced in\nCrohn’s disease and ulcerative colitis. Vα24+ T-cells and the SP subset of\nVα24+ T-cells were reduced in Crohn’s disease but not in ulcerative colitis.\nCirculating Vα24+ Vβ11+ T-cells, Vα24+ Vβ11+ α-GalCer/CD1d tetramer+\nand Vα24+ 6B11+ iNK T-cells were deficient in both Cohn’s disease and\nulcerative colitis. The deficiency of Vα24+ T-cells was also observed within\nthe intestinal mucosa. Intestinal Vα24 mRNA expression from Crohn’s disease\nand ulcerative colitis was reduced compared to levels in normal subjects as\nassessed by relative and RT-PCR. Cytokine production by Vα24+ T-cells,\n6B11+ and Vα24+ α-GalCer/CD1d tetramer+ iNK T-cells after 4 h in vitro\nanti-CD3 stimulation was impaired for subjects with Crohn’s disease and\nulcerative colitis.\nIn summary, Vα24+ T-cell number and function were affected by age. Further\ninvestigations are warranted to see if deficiency of this immunoregulatory\npopulation is associated with disease. The decrease and dysfunction in\nimmunoregulatory cells, Vα24 T-cells and iNK T-cells could contribute to the\npathogenesis of coeliac disease, Crohn’s disease and ulcerative colitis. Coeliac\ndisease, Crohn’s disease and ulcerative colitis are polygenetic diseases in\nwhich environmental factors play a significant role in disease development and\nstate. The reduced numbers of iNK T-cell along with their impaired function\nmay only be two factors. Presumably, other factors are involved. Nevertheless,\niNK T-cells offer a potential target for the therapeutic intervention of coeliac\ndisease, ulcerative colitis and Crohn’s disease.