1 a , 25-Dihydroxyvitamin D 3 UpRegulates Bcl-2 Expression and Protects Normal Human Thyrocytes from Programmed Cell

Apoptosis is thought to play an important role in the pathogenesis of autoimmune thyroid disease. 1a,25-dihydroxyvitamin D3 (VD3) has been shown to suppress several autoimmune diseases. However, the mechanism by which VD3 has these effects is not known. We evaluated the alterations in apoptosis, induced by VD3. Thyrocytes were treated with VD3, and the expression of the Bcl-2 family molecules was studied at both the messenger RNA and protein levels. It was found that VD3 significantly induced the expression of Bcl-2 messenger RNA and protein in thyrocytes but had no effect on the expression of Bcl-xl and Bax. The increase in Bcl-2 expression, mediated by VD3, correlated with protection of thyrocytes against the induction of apoptosis by either staurosporine or UV irradiation. VD3-induced increases in the expression of Bcl-2 could be mimicked by VD3 analogs with high nuclear receptor affinity, but not by analogs only with nongenomic actions. These data indicate a role for Bcl-2 in the regulation of apoptosis in thyrocytes and raise the possibility that VD3 or its agonists may have therapeutic benefit in thyroid disorders. (Endocrinology 140: 1649–1656, 1999) C (HASHIMOTO’S) thyroiditis is an extremely frequent autoimmune disease, affecting approximately 5% of women, and is the most common cause of hypothyroidism in the United States (1). Previous studies indicate that apoptosis plays an important role in the destruction of thyrocytes in thyroiditis (2–4). However, the regulation of apoptosis in thyroid cells has not been fully defined. Several studies have documented that the induction of apoptosis in thyroid cells through different pathways is highly regulated, but the studies differed on the exact manner and effect of this regulation (2–6). Bcl-2 is one member of a family of protooncogenes that regulates apoptosis (7). The strong inhibitory effect of Bcl-2 on programmed cell death is evidenced by the apoptosis of Bcl-2-negative cortical thymocytes during T cell development, whereas positively selected, surviving thymocytes in the medulla show strong staining for Bcl-2 (8). Several members of the Bcl-2 family are expressed in thyroid follicular cells in both normal and pathologic conditions (9). In light of the putative role of apoptotic regulation in the development of autoimmune hypothyroidism, it is of interest to determine whether Bcl-2 has a role in the regulation of apoptosis in thyrocytes. 1a,25-dihydroxyvitamin D3 (VD3), an active form of vitamin D, is a potent secosteroid hormone that produces a wide array of biological effects in its target tissues. In addition to the long recognized role of VD3 in calcium homeostasis, VD3 also has immunomodulatory and antiinflammatory properties, as well as a regulatory role in the growth and differentiation of a variety of cell types and tissues (10, 11). VD3 acts through an intracellular vitamin D receptor (VDR), which belongs to the nuclear hormone receptor gene superfamily (12). The hormone-receptor complex binds to sequence-specific sites in promoters and mediates its effects by altering transcriptional activity of specific genes (13). As might be expected from VD3’s multiple activities, VDRs are present in both classical target tissues, regulating calcium homeostasis, and a wide variety of nonclassical target tissues that include breast cancer cells, lymphocytes, and thyroid follicular cells (11). VD3 has also been suggested to have a variety of nongenomic actions, mediated without the requirement of nuclear receptor binding (11, 13). VD3 has been reported to alter the induction of apoptosis in breast cancer cells (14). Furthermore, VD3 (in the presence of cyclosporin) significantly reduced the incidence of thyroid pathology and the severity of thyroiditis in a mouse model of experimental autoimmune thyroiditis (15). In this report, we demonstrate that VD3 induces the expression of Bcl-2 messenger RNA (mRNA) and protein in human thyroid follicular cells, and this action is associated with protection against apoptosis. These studies indicate a role for Bcl-2 in regulating susceptibility to apoptosis in Hashimoto’s thyroiditis and other thyroid disorders, and they suggest a possible therapeutic role for VD3 in these diseases. Materials and Methods