Heparin-binding EGF-like growth factor (HB-EGF) is a member of the EGF family that binds to and activates the EGF receptor, and is expressed in a variety of tissues, predominantly lung, heart, brain and skeletal muscle. HB-EGF is known to induce VSMC proliferation, and this involves PI3K-Akt and MAPK pathway. Our previous data showed that JAK-STAT pathway was also involved in HB-EGF induced VSMC proliferation. Interestingly, HB-EGF (10 ng/ml) induced a biphasic activation of STAT3 (5 min and 60-120 min). Therefore, we tried to elucidate the underlying mechanism of this delayed STAT3 activation by HB-EGF and its effect in VSMCs. First, we examined the effect of HB-EGF on IL-6 gene expression in VSMCs, since IL-6 has been implicated in the regulation of STAT3 activation. According to our data, HB-EGF increased transcription of IL-6 gene (30 - 60 min) and subsequent secretion of IL-6 as evidenced by ELISA assay. Furthermore, HB-EGF-mediated stimulation of IL-6 gene expression and IL-6 secretion was inhibited by NF-觀B inhibitor Bay 117082 (2.5 關M) treatment suggesting it involves NF-觀B pathway. Again, the delayed activation of STAT3 by HB-EGF was abolished by both Bay 117082 and IL-6 neutralizing antibody (1 關g/ml) indicating IL-6 was a key molecule in delayed activation of STAT3. In addition, STAT3 activation induced by HB-EGF stimulated conditioned media was also inhibited by IL-6 neutralizing antibody, and HB-EGF induced VSMC proliferation was suppressed by IL-6 neutralizing antibody. In conclusion, IL-6 plays an important role in the delayed activation of STAT3 and VSMC proliferation induced by HB-EGF.