Differential Expression of NK Receptors CD94 and NKG2A by T Cells in Rheumatoid Arthritis Patients in Remission Compared to Active Disease

TNF inhibitors (TNFi) have revolutionised the treatment of rheumatoid arthritis (RA). Natural killer (NK) cells and Natural Killer Cell Receptor+ T (NKT) cells comprise important effector lymphocytes whose activity is tightly regulated through surface NK receptors (NKRs). Dysregulation of NKRs in patients with autoimmune diseases has been shown, however little is known regarding NKRs expression in patients with TNFi-induced remission and in those who maintain remission vs disease flare following TNFi withdrawal.Patients with RA were recruited for this study, (i) RA patients in clinical remission following a minimum of one year of TNFi therapy (n = -15); (2) Active RA patients, not currently or ever receiving TNFi (n = 18); and healthy control volunteers (n = 15). Patients in remission were divided into two groups: those who were maintained on TNFi and those who withdrew from TNFi and maintained on DMARDS. All patients underwent full clinical assessment. Peripheral blood mononuclear cells were isolated and NKR (CD94, NKG2A, CD161, CD69, CD57, CD158a, CD158b) expression on T-(CD3+CD56-), NK-(CD3-CD56+) and NKT-(CD3+CD56+) cells was determined by flow cytometry.Following TNFi withdrawal, percentages and numbers of circulating T cells, NK cells or NKT cell populations were unchanged in patients in remission versus active RA or HCs. Expression of the NKRs CD161, CD57, CD94 and NKG2A was significantly increased on CD3+CD56-T cells from patients in remission compared to active RA (p<0.05). CD3+CD56-T cell expression of CD94 and NKG2A was significantly increased in patients who remained in remission compared with patients whose disease flared (p<0.05), with no differences observed for CD161 and CD57. CD3+CD56- cell expression of NKG2A was inversely related to DAS28 (r = -0.612, p<0.005).High CD94/NKG2A expression by T cells was demonstrated in remission patients following TNFi therapy compared to active RA, while low CD94/NKG2A were associated with disease flare following withdrawal of therapy

Expression of NKRs CD94, NKG2A, CD161 and CD57 in patients maintaining or withdrawing therapy.

<p>Graphs illustrate (<b>A</b>) CD94 and NKG2A expression and (<b>B</b>) CD161 and CD57 expression at baseline (▴) and three months (○) in patients maintaining or withdrawing TNF inhibitor therapy. (C) Representative dot plots of two patients withdrawing TNF inhibitor therapy. Patient (I) remained in clinical remission for the study duration and maintained expansion of CD94 and NKG2A; Patient (II) had a flare in disease activity 6 months after withdrawal of therapy with loss of expansion of CD94 and NKG2A at 3 months (i.e. prior to clinical relapse). *p<0.05 significantly different.</p

Clinical, laboratory and functional assessments in patients cohorts maintaining and withdrawing TNF inhibitor therapy.

<p>*p<0.05 vs baseline.</p

Total CD3+, Total CD56+ and CD3+CD56− T cell populations in patients maintaining or withdrawing therapy.

<p>Graphs illustrate total CD3+, Total CD56+ and CD3+CD56− T cell populations (expressed as a percentage of the lymphogate) in patients maintaining or withdrawing therapy at baseline (▴) and three months (○). *p<0.05 significantly different.</p

Baseline characteristics of patients in remission following TNF inhibitor therapy and in patients with active rheumatoid arthritis.

<p>*p<0.05 remission cohort significantly different from active RA cohort.</p

Expression of CD161, HLA-DR and CD57 in active RA, Remission RA and healthy controls.

<p>(A) Representative dot plots of CD161, HLA-DR and CD57 expression on healthy control, patient with active RA and patient in remission following TNF inhibitor therapy. (B) CD161, HLA-DR and CD57 expression on CD3+CD56− cells in healthy controls (▴), patients with active RA (○) and patients in remission following TNF inhibitor therapy (▪). *p<0.05 significantly different.</p

Expression of NKRs CD94 and NKG2A in active RA, Remission RA and healthy controls.

<p>(A) Representative dot plots of CD94 and NKG2A expression on healthy control, patient with active RA and patient in remission following TNF inhibitor therapy. (B) CD94 and NKG2A expression on CD3+CD56− cells in healthy controls (▴), patients with active RA (○) and patients in remission following TNF inhibitor therapy (▪). *p<0.05 significantly different.</p

T cell, NK and NKT cell populations in active RA, Remission RA and healthy controls.

<p>Graph illustrating T cell, NK and NKT cell populations in healthy controls (N = 15) (▴), patients with active RA (n = 18) (○) and patients in remission following TNF inhibitor (n = 15)(▪). *p<0.05 significantly different.</p

Associations between clinical disease activity and NKG2A expression.

<p>(A) Flare in disease activity (Increased DAS28) is associated with reduction in NKG2A expression on CD3+CD56− cells. (B) Negative correlation between DAS28 and T cell expression of *p<0.05 significantly different.</p