Human fibroblasts ubiquitously express glutamic acid decarboxylase 65 (GAD 65): possible effects of connective tissue inflammation on GAD antibody titer.

BACKGROUND

Type 1 diabetes is caused by a destruction of pancreatic beta cells due to autoimmunity. Autoantibody against glutamic acid decarboxylase (GAD) 65 expressed in pancreatic beta cells is widely used as a predictive marker for pancreatic destruction. In this study, we hypothesized that if certain cells in periodontal tissues could express GAD, then it may influence GAD antibody titer.

METHODS

We used: 1) reverse transcription-polymerase chain reaction (PCR) analysis to detect GAD 65 mRNA in various cells; 2) nucleotide sequencing analysis to confirm that amplified PCR product is the gene encoding GAD; and 3) Western blotting to determine the expression of GAD 65 protein in human gingival fibroblasts. Immunohistochemical staining of GAD 65 protein in normal and inflamed gingiva was performed to examine the potential influence of periodontal inflammation on GAD 65 expression. GAD antibody titer in sera of periodontal patients as well as healthy subjects was measured to determine if periodontal patients could develop autoantibody against GAD 65.

RESULTS

Cultured human gingival, periodontal, and dermal fibroblasts and mesangial cells expressed GAD mRNA. Nucleotide sequencing analyses confirmed the amplified PCR product as GAD 65. Western immunoblotting analyses and immunohistochemical staining revealed that the GAD 65 protein was expressed in vitro and in vivo. The expression of GAD 65 in inflamed tissue was higher than that in normal tissues. Two of 62 periodontal patients without diabetes showed an increased antibody titer against GAD 65, while none of the systemically healthy subjects showed an increased antibody titer against this antigen.

CONCLUSIONS

We concluded that periodontal inflammation may result in higher levels of GAD and influence GAD antibody titer, and, hence, affect diabetic diagnosis based upon GAD antibody production.