| 摘要: | 基質金屬蛋白酶（matrix metalloproteinases，MMPs）能夠降解細胞外基質（extracellular matrix，ECM），且已經被證實在釵h不同腫瘤細胞及基質細胞（stromal cell）中都會分泌這些蛋白酶。我們實驗室先前的研究曾經指出在南台灣的37組口腔腫瘤組織中MMP-3有超過60％的表現率，而且其表現與否和腫瘤期別及淋巴結轉移等有顯著的相關性。MMP-28是基質金屬蛋白酶家族中最新的一種蛋白酶，它在幾種不同腫瘤當中有表現，這型蛋白水解酶於腫瘤生成與發展的過程所扮演的角色至今仍未明瞭。在本研究當中，我們一方面想要了解MMP-28在口腔鱗狀細胞上皮癌（oral squamous cell carcinomas，OSCCs）中的表現情形是否與某些腫瘤特性相關，一方面則擬擴大先前的腫瘤樣本數，觀察MMP-3的表現是否仍與腫瘤期別與頸部淋巴的轉移或其他腫瘤特性具相關性，另一方面則利用antisense strategy來探討這兩種MMPs可能扮演的角色。我們總共偵測99組南台灣口腔腫瘤病人的腫瘤組織，包含7組口腔癌前病變（premalignant lesions，PMLs），及92組OSCCs，範圍從腫瘤第一期至第四期，也包含腫瘤相對應的鄰近正常組織。我們使用反轉錄聚合酶連鎖反應（reverse transcription-polymerase chain reaction，RT-PCR）半定量分析方法及組織免疫化學染色法（immunohistochemical，IHC），分別偵測MMP-3和MMP-28的mRNA的表現情形及其蛋白質表現位置。在IHC染色方法中發現MMP-3和MMP-28蛋白都是偏好表現在OSCCs的腫瘤區域內。而RT-PCR的分析方法中發現99組腫瘤組織當中，有表現MMP-3和MMP-28的樣本分別有73組（74％）和52組（53％）；統計的結果顯示，此兩種MMPs在OSCC樣本中的表現率明顯高於PML的樣本（p<0.01），但是和腫瘤侵犯深度、腫瘤大小、及淋巴結轉移等均無明顯相關性。為要探討此兩種MMPs對OSCC的生長、移動（migration）、或群落形成（colony formation）能力上的重要性，我們利用MMP-3及MMP-28的逆序股寡核苷酸（antisense oligonucleotide，AON）轉染（transfection）OSCC細胞株OECM-1與SCC-9，發現對於細胞的生長影響不大，但是在軟培養基（soft agar）上，有降低OECM-1及另一食道癌細胞株CE81T/VGH群落形成的數目，並且也明顯的抑制了OECM-1與SCC-9的移動（p<0.05）。因此我們的結論為：在七成左右南台灣的口腔腫瘤樣本中有表現MMP-3分子，五成左右有表現MMP-28分子，其表現率雖然不會隨著腫瘤侵犯深度、腫瘤大小、及淋巴結轉移等特性而增加，但很可能從PML進展到OSCC的過程中，此兩種MMPs均扮演了重要角色。另一方面在經MMP-3及MMP-28的AON的轉染後，OECM-1與SCC-9的移動以及OECM-1與CE81T/VGH在軟培養基中群落形成等能力有明顯受到抑制的情況，顯示此兩種MMPs在某些上消化道癌形成的過程中確實扮演了重要的角色。
Matrix metalloproteinases (MMPs) have an ability to degrade virtually all components of the extracellular matrix (ECM), and are abundantly secreted by a variety of tumor cells and stromal cells. Our previous study showed that MMP-3 was expressed in 23/37(62%) oral tumors specimens in southern Taiwan, and its expression was correlated with both tumor stage and lymph node metastasis. MMP-28 is the newest member of the MMP family and MMP-28 is also expressed in several carcinomas. The object of this study was to evaluate the correlation of the expression of MMP-3 and MMP-28 and various tumor parameters of oral squamous cell carcinomas (OSCCs) in southern Taiwan and to explore their possible roles by applying antisense strategy in OSCC cell line. Ninety-nine oral tumor specimens (T) (7 premalignant lesions [PMLs] and 92 OSCCs), as well as their respective neighboring tissues (N) were subjected to MMP-3 and MMP-28 detection. Semi-quantitative reverse transcription- polymerase chain reaction (RT-PCR) and immunohistochemical (IHC) staining were used to detect the mRNA expression and the protein location of MMP-3 and MMP-28, respectively. IHC staining for MMP-3 was detected in the advancing front or marginal portion of the cancerous tissues, and MMP-28 protein molecules was detected in the PML and OSCC tissue. As revealed by RT-PCR, signals of MMP-3 and MMP-28 were positive in 73/99 (74%) and 52/99 (53%) tumors, respectively. Furthermore, the incidence of both MMPs is significantly higher in OSCCs than in premalignant lesions (p<0.01). However, their expressions were not statistically correlated with tumor thickness, tumor size, and lymph node metastasis. To assess the roles of MMP-3 and MMP-28 in tumor growth, migration, and tumorgenicity, transfection of antisense oligonucleotide (AON) was used to block their expressions. Our results have shown that transfection of OECM-1 and SCC-9 with AON of MMP-3 or MMP-28 did not hamper the growth of these cell lines, however, it inhibited the migration of OECM-1 and SCC-9 and the soft-agar-colony-formation of OECM-1 and CE81T/VGH. We conclude that MMP-3 and MMP-28 may be expressed in around 70% and 50% oral tumors in southern Taiwan, respectively. They may play important roles in the progression of oral tumors, especially from the PML to the OSCC. On the other hand, both MMP-3 and MMP-28 may play important roles in the migration and anchorage-independent growth in some aerodigestive carcinomas. |
