T cells and T-cell cytokine transcripts in the synovial membrane in patients with osteoarthritis

Abstract

The synovial membrane in osteoarthritis (OA) often exhibits inflammatory infiltrates, but the role of T cells in these infiltrates is not known. T-cell activation antigens were analyzed by immunohistochemistry, and T-cell cytokine transcripts were measured by competitive PCR in synovial membranes from patients with OA and rheumatoid arthritis (RA). Lymphoid cell aggregates, containing primarily CD3+ T lymphocytes, were found in 65% of patients with OA. Mononuclear cells expressing the activation antigens CD69, CD25, CD38, CD43, CD45RO, and HLA class II were present in both patient groups, although in higher numbers in patients with RA. Interleukin 2 (IL-2) transcripts were found in 10 of 18 patients with OA versus 12 of 13 patients with RA (P = 0.03). Gamma interferon (IFN-gamma) transcripts were detected in 9 of 18 patients with OA versus 10 of 13 patients with RA (not significant), whereas IL-10 transcripts were found in nearly all patients. IL-4 and IL-5 were not detected in any patients. The levels of IFN-gamma and IL-2 transcripts, normalized for T-cell number equivalents, were not statistically different between OA and RA, but the levels of IFN-gamma, normalized for total cell number equivalents, were lower in OA than in RA (P = 0.01). Synovial membranes that expressed IL-2 and IFN-gamma transcripts were more likely to have heavier infiltrations of T cells and cells bearing activation markers than synovial membranes that did not express these cytokines. The presence of activated T cells and TH1 cytokine transcripts in chronic joint lesions of patients with OA suggests that T cells contribute to chronic inflammation in a large proportion of these patients.

FIG. 1

Avidin-biotin complex immunostaining with light hematoxylin counterstaining of the SM from a patient with OA (original magnification, ×400). Serial sections of the same field depicting nodular-perivascular mononuclear cell infiltrate were stained with anti-CD3, anti-CD69, anti-CD25, anti-CD45RO, anti-HLA class II, anti-CD38, and anti-CD43 MAbs and immunoglobulin G1 as a negative control. Immunoreactivity was more widespread with antibodies to late-activation antigens CD45RO and HLA class II than to early-activation antigens CD69 and CD25.

FIG. 2

Degree of SM infiltration with mononuclear cells expressing activation markers in patients with OA and RA. Cell infiltration was graded on a relative scale of 0 to 4, based on the number of positive cells per HPF (×400). CD3 (A) and CD45RO (B) grading: 0, <2 cells; 1, 2 to 50 cells; 2, 51 to 100 cells; 3, 101 to 150 cells; 4, >150 cells. HLA class II (C) grading: 0, <20 cells; 1, 21 to 100 cells; 2, 101 to 200 cells; 3, 201 to 300 cells; 4, >300 cells. CD69 (D), CD25 (E), CD38 (F), and CD43 (G) grading: 0, <1 cell; 1, 1 to 20 cells; 2: 21 to 40 cells; 3, 41 to 60 cells; 4, >60 cells. Bars represent percentages of patients at each grade.

FIG. 3

Cytokine transcript profile in SM specimens from representative patients with OA and RA. The respective MIMIC DNAs or cDNA from phytohemagglutinin-stimulated normal PBMC were used as a positive control (C). A 100-bp DNA ladder was used as a molecular weight marker (M).

FIG. 4

Example of cytokine (IL-10) transcript quantitation by MIMIC PCR. A constant amount of cDNA was amplified along with serial dilutions of a known quantity of IL-10 MIMIC DNA. As the intensity of the MIMIC DNA band was decreasing, the intensity of the IL-10 band was increasing. The lane with 1:1 ratio of MIMIC to IL-10 bands corresponds to 2 × 10⁻⁴ attomol (amoles) of MIMIC DNA input and shows an equimolar amount of IL-10 cDNA.

FIG. 5

IL-2 and IFN-γ transcript levels in OA and RA SM quantitated by MIMIC PCR. Cytokine transcripts were normalized for CD3δ transcripts and therefore for T-cell number equivalents.

FIG. 6

IFN-γ and IL-10 transcript levels in SM from patients with OA and RA normalized for β-actin cDNA and therefore for total cell number equivalents. IFN-γ transcript levels were higher in RA than in OA (P = 0.01). IL-10 transcript levels were 100- to 1,000-fold higher than those of IFN-γ in both patient groups.