Elevated proportion of natural killer T cells in periodontitis lesions: a common feature of chronic inflammatory diseases

Abstract

Although periodontitis is a chronic inflammatory disease caused by a group of so-called periodontopathic bacteria, autoimmune mechanisms have also been implicated in the disease process. Recently, a unique subset of lymphocytes designated natural killer (NK) T cells expressing the Valpha24JalphaQ invariant T cell receptor (TCR) has been reported to have a regulatory role in certain autoimmune diseases. Therefore, we investigated the proportion of the invariant Valpha24JalphaQ TCR within the Valpha24 T cell population in periodontitis lesions and gingivitis lesions using single-strand conformation polymorphism methodology. NK T cells were identified with a specific JalphaQ probe whereas the total Valpha24 TCR was identified using an internal Calpha probe. NK T cells were a significant proportion of the total Valpha24 population both in periodontitis lesions and to a lesser extent in gingivitis lesions but not in the peripheral blood of either periodontitis patients or nondiseased controls. Using immunohistochemistry, some of Valpha24(+) cells in the periodontitis lesions seemed to associate with CD1d(+) cells, which are specific antigen-presenting cells for NK T cells. Although the mechanism underlying the elevation of NK T cells in periodontitis and in gingivitis lesions remains unclear, it can be postulated that NK T cells are recruited to a play regulatory role in the immune response to bacterial infection.

Figure 1.

Demonstration of the Vα24⁺ T cell clonalities and the invariant Vα24JαQ TCR in peripheral blood of controls (A), peripheral blood of periodontitis patients (B), gingival tissue of controls (C), and gingival tissue of periodontitis patients (D). Total RNA was extracted from PBMCs and gingival tissues and then subjected for RT-PCR-SSCP analysis. The top and bottom panels demonstrate the total Vα24⁺ TCR SSCP profiles as detected with a Cα probe and the invariant Vα24JαQ band as detected with the JαQ-specific probe, respectively. The cloned Vα24JαQ DNA fragment as a positive control was applied on the far left lanes. The alphabetical code and the numerical code correspond to each control (A and C) and each patient (B and D), respectively, implying that blood samples and gingival tissues were taken from same subjects and patients. Arrows indicate the position for the invariant Vα24⁺JαQ TCR. The unique bands were indicated by arrowhead. Another unique band appeared in patients 8 and 14 was indicated by open arrowhead.

Figure 2.

Relative proportion of the invariant Vα24JαQ TCR in total Vα24 population. SSCP profiles were analyzed on a computer software (NIH image version 1.62) and relative area and density of Vα24JαQ band to the total area and density of Vα24 TCR was calculated in each lane. The data were expressed as mean ± SD. Relative proportion of the Vα24JαQ TCR in the total Vα24 population was significantly higher in periodontitis tissues than either in gingivitis tissues (P = 0.016) or in peripheral blood of both controls (P = 0.015) and periodontitis patients (P = 0.036).

Figure 3.

Expression of Vα24 and CD1d in periodontitis tissues. Double staining of Vα24 and CD1d was performed by using combined an ABC-PO method and an alkaline-phosphatase anti-alkaline-phosphatase (APAAP) method on cryostat sections. Vα24⁺ cells appeared as brown and CD1d⁺ cells appeared as blue. Anti-CD1d antibody reacted with either cells in the inflammatory infiltrate (A) or cells with spindle shape beneath the oral epithelium (B). Although direct cell-cell contact between Vα24⁺ cells and CD1d⁺ cells was observed in some sections (C), they were also stained separately (D). Original magnification, ×132.