Our aim in the present study was to understand the mechanism whereby specific destruction of acinar but not ductal structure occurs in salivary glands in Sjögren's syndrome (SS). Thus, we examined the effects of cytokines including TNF-alpha and IL-1 beta on the proteolytic activity of cultured normal human salivary gland cell clones, because degradation of the basement membrane by proteolytic enzymes leads to the disruption of acinar or ductal structure in salivary glands. Simian virus 40 (SV40)-immortalized normal human salivary gland cell clones with ductal (NS-SV-DC) or acinar (NS-SV-AC) phenotype were treated either with TNF-alpha or IL-1 beta alone or with a combination of both, and then proteolytic activity was examined. Although cytokine-treated NS-SV-AC demonstrated high matrix metalloproteinase-2 (MMP-2) activity at both protein and mRNA levels, no remarkable increase in MMP-2 activity was detected in NS-SV-DC. Expression of tissue inhibitor of metalloproteinase-2 (TIMP-2), a specific inhibitor of MMP-2, was similarly inhibited by cytokines in these cell clones. Thus, the net balance estimated by MMP-2/TIMP-2 suggested enhanced proteolysis in cytokine-treated NS-SV-AC and, to a much lesser extent, in NS-SV-DC. To examine whether enhanced expression of MMP-2 occurred in acinar cells of SS salivary glands, we carried out an immunohistochemical study using SS salivary gland tissues. This study indicated that acinar cells adjacent to the lymphocytic infiltrate exhibited enhanced expression of MMP-2 compared with those distant from infiltrated lymphocytes or with those in normal salivary glands. By Northern blot analysis, NS-SV-DC and NS-SV-AC expressed receptor mRNA for both cytokines. The signal-dependent activation of the transcription factor NF-kappa B was observed only in NS-SV-AC. I kappa B-alpha, a specific inhibitor of NF-kappa B, was down-regulated by treatment with cytokines in NS-SV-AC. However, the expression of I kappa B-alpha protein was not detected in NS-SV-DC at the basal level. Treatment of NS-SV-AC with calpain inhibitor-I restored the expression of I kappa B-alpha protein in cytokine-treated cells, thus leading to the inhibition of NF-kappa B activation. Reverse transcriptase-PCR analysis confirmed that there was a marked reduction in I kappa B-alpha mRNA expression in NS-SV-DC as compared to that in NS-SV-AC. As such, it seems likely that there is a relationship between the activation of MMP-2 and the activity of NF-kappa B, and that the NF-kappa B/I kappa B-alpha complex is not a signal mediator involved in cytokine-induced suppression of TIMP-2. These observations, therefore, indicate that the divergent response to cytokines in each constituent cell of salivary gland may result in the histopathologic manifestation of SS.