Study on the Correlation of Thrombomodulin Expression and Cell Proliferation

• Main Authors: Yu-Lan Chou, 周于嵐
• Other Authors: Hua-Lin Wu
• Format: Others
• Language: zh-TW
• Published: 2004
• Online Access: http://ndltd.ncl.edu.tw/handle/a5u4jd

碩士 === 國立成功大學 === 生物化學研究所 === 92 === 　　Thrombomodulin(TM) is widely distributed in a number of different cell types, such as epithelial cells, smooth muscle cells, keratinocytes, and endothelial cells. TM is a glycoprotein receptor and cofactor for thrombin’s activity in a physiologically important natural anticoagulant system. Our recent study has demonstrated that TM may mediate cell-cell adhesion through its lectin-like domain. However, the mechanisms of regulating TM expression are still unclear. In this report, the correlation of TM expression and cell proliferation was investigated. Bovine aortic endothelial cells (BAECs) were incubated for various time intervals and the growth rate and TM expression level were analyzed by MTT assay and Western blot, respectively. Furthermore, flow cytometry was used to determine TM expression levels on cell surface. The results demonstrated that the TM level was increased with cell proliferation and reached maximum at day 3, in which cells grew to confluent monolayer. The TM level was decreased when BAECs were cultured longer than three days. In order to further examine the effect of cell density on TM expression, BAECs with different density were plated onto culture dishes, and TM expression was determined after 15 hr incubation to eliminate the effect of cell proliferation. The data showed that TM expression was not changed significantly under the low cell density condition, whereas TM level was reduced when cell density is over confluent. We also observed that TM expression level was enhanced by adding VEGF to stimulate BAECs proliferation. Furthermore, RT-PCR and Western blot experiment showed that the decrease of TM expression level was due to the downregulation of TM gene transcription, and independent of internalization and degradation of TM. In addition, similar results were observed in HaCaT cells, a human keratinocyte cell line. TM expression in HaCaT cells was also elevated with cell proliferation. These results suggested that TM expression might be increased when cells proliferated, but decreased after cells reached confluence. Taken together, we concluded that TM expression may be regulated by cell proliferation and cell density.