Moore D H, Allison B, Look K Y, Sutton G P, Bigsby R M
Department of Obstetrics and Gynecology, Indiana University Medical Center, Indianapolis 46202, USA.
Gynecol Oncol. 1997 Apr;65(1):78-82. doi: 10.1006/gyno.1997.4628.
There are several distinct forms of the matrix metalloproteinases (MMP) that degrade collagen in the extracellular matrix. Most involved in the metastatic process are collagenases exhibiting specificity for type IV collagen, the collagen that makes up the backbone of the basement membrane. We chose to examine the expression of one type IV collagenase, MMP-9, in several ovarian cancer cell lines.
Under the influence of phorbol esters such as TPA, the fibrosarcoma cell line HT1080 is known to express MMP-9. We examined the effect of TPA on MMP-9 expression with Northern analysis, and on MMP-9 collagenase activity with polyacrylamide gel electrophoresis-zymography, in HT1080 cells (positive control) and in six ovarian cancer cell lines: Hey, HeyA8, CAOV3, PA-1, SKOV3, and OVCAR3. Cells were grown to 70% confluence and the growth medium was changed to a serum-free medium for 24 hr. Cells were then treated with 12 ng/mL TPA or its vehicle for an additional 24 hr. Medium was collected for PAGE-zymographic analysis. The cells were lysed and their RNA collected for Northern analysis.
PA-1, a teratocarcinoma, expressed more than 10-fold the amount of RNA for MMP-9 than the other lines following TPA treatment. Both Northern analysis and zymography showed that four of the six ovarian cell lines responded to TPA with increased collagenase expression and activity. The other two cell lines showed no MMP-9 activity either before or after TPA treatment.
These results suggest that ovarian cancer cell lines express collagenase and that this expression may be regulated by phorbol esters which activate the protein kinase C pathway.
基质金属蛋白酶（MMP）有几种不同形式，可降解细胞外基质中的胶原蛋白。参与转移过程的大多数是对IV型胶原具有特异性的胶原酶，IV型胶原构成基底膜的骨架。我们选择检测几种卵巢癌细胞系中一种IV型胶原酶MMP - 9的表达。
在佛波酯如TPA的影响下，已知纤维肉瘤细胞系HT1080表达MMP - 9。我们用Northern分析检测TPA对HT1080细胞（阳性对照）和六种卵巢癌细胞系（Hey、HeyA8、CAOV3、PA - 1、SKOV3和OVCAR3）中MMP - 9表达的影响，并用聚丙烯酰胺凝胶电泳酶谱法检测TPA对MMP - 9胶原酶活性的影响。细胞生长至70%汇合，将生长培养基换成无血清培养基24小时。然后用12 ng/mL TPA或其溶剂再处理细胞24小时。收集培养基进行PAGE - 酶谱分析。裂解细胞并收集其RNA进行Northern分析。
畸胎瘤细胞系PA - 1在TPA处理后，MMP - 9的RNA表达量比其他细胞系高出10倍以上。Northern分析和酶谱分析均显示，六个卵巢细胞系中有四个对TPA有反应，胶原酶表达和活性增加。另外两个细胞系在TPA处理前后均未显示MMP - 9活性。
这些结果表明卵巢癌细胞系表达胶原酶，且这种表达可能受激活蛋白激酶C途径的佛波酯调控。
