Malignant Phenotype And Molecular Mechanism Of Cancer Cell Lamellipodium

The malignant degree of cancer is closely correlated with the malignant phenotype of cancer. There are three malignant phenotypes in cancer cell: immortality, mobility and loss of contact inhibition. Certainly, there are many others physiological, biochemical and morphological characteristics in cancer cell like heteroploid, metabolized abnormality, drug resistance and so on. And mobility of cancer cell is one of the most common phenotypes, which directly determining the malignant degree. The mobility of cancer cell involved in reduction of cell adhesion, rearrangement of cell skeleton, degradation of extracellar matrix and formation of pseudopodium and cell protruding. Pseudopodium formation, especially formation is the key point of cancer cell mobility. Pseudopodium is divide to filopodium, lamellipodium, invadopodia and podosome. As a special structure of cell membrane. They have various functions their own: filopodium are in charge of cell adhesion, lamellipodium play a important role in cell mobility. And HAb18G/CD147 as a tumor marker expressed on many kinds of cancer with a high level. There are many references reports that HAb18G/CD147 makes a important influence in cancer metastasis and invasion. However, the influence mechanism needs a further clarification.Hence, in order to understand the relationship between HAb18G/CD147 and malignant phenotypes, especially the metastasis and invasion of cancer, and how the HAb18G/CD147 regulate cell mobility by alteration of pseudopodium, we used a series of approaches of cell biology and molecular biology. Finally, we proved that HAb18G/CD147 activated Cortactin through phosphorylation of Fak and Src, and activated Cortactin as a support protein stabilized the Arp2/3 complex, which induce Actin forming dendritic framework which namely lamellipodium. Eventually, lamellipodium promote mobility of cancer cell. The whole research elucidated the biology mechanism that HAb18G/CD147 promotes metastasis and invasion at last. Part 1 HAb18G/CD147 mediates cancer metastasis through promoting the formation of lamellipodium of cancer cellThere are many references reports that HAb18G/CD147 makes a important influence in cancer metastasis and invasion. Through whether sealing the function of HAb18G/CD147 from antibody or intervening the expression of HAb18G/CD147, it will be inhibited the ability of cancer metastasis and invasion; But, how the HAb18G/CD147 promotes cancer metastasis and invasion is still need a reasonable interpretation.For this reason, we found that the low expression of HAb18G/CD147 can significantly inhibit the mobility of cancer cell through transwell experiments with some cancer cells which HAb18G/CD147 gene been knock down or knock out, and we validated that the low expression of HAb18G/CD147 can significantly inhibit the velocity of cell movement through cell tracking experiments; all above experiments point out that the change of the expression of HAb18G/CD147 can significantly influence the ability of cancer metastasis. After that, we observed the process of cell movement using wound-healing assay by high-resolution confocal and found that less formation of lamellipodium in HAb18G/CD147 low expression group but more formation of lamellipodium in leading edge of cancer cell in HAb18G/CD147 high expression group. And we also validated that HAb18G/CD147 expression influence the formation of lamellipodium using immunofluorescent staining assay. We reach a conclusion that the change of the low expression of HAb18G/CD147 can significantly inhibit the ability of cancer metastasis and the inhibition mainly be directed against lamellipodium formation. Part 2 The molecular mechanism of lamellipodium formation which impact by HAb18G/CD147So, in order to reveal the specific molecular mechanism of lamellipodium formation which impact by HAb18G/CD147, we used the cancer cells which the first part mentioned to do some western blot experiments to test the expression of some common molecules related with lamellipodium formation, and we found that HAb18G/CD147 can not change the expression of these molecules directly. So, next we test the phosphorylation level of these molecules by their phosphorylate antibodies with western blot. And we found that the low expression of HAb18G/CD147 can change the phosphorylation level of these common molecules related with lamellipodium formation like Fak, Src and Cortactin. After that, we test interactions between these molecules by FRET experiments. Next we validated that the sequence of interactions by western blot. At last, we validated that the phosphorylation of Cortactin is the most important point of lamellipodium formation by live cell observation and transwell experiments after tyrosine point mutation in Cortactin.Through above outcomes We reach a conclusion that HAb18G/CD147 promote lamellipodium formation was through the sequence phosphorylation from HAb18G/CD147, Fak, Src to Cortactin. Part 3 In vivo research about that HAb18G/CD147 promote cancer metastasisThe objective of this part is validating that HAb18G/CD147 promote cancer metastasis by a series of animal experiments and analysing the relationship between the expression of HAb18G/CD147 and TNM stages by case collection.We used the methods that construct metastasis model in nude mice via tail intravenous administration cancer cells. At endpoint, we euthanized and anatomize the nude mice, analysed the lung and liver by HE staining. And we found that HAb18G/CD147 significantly influence the liver metastasis. This outcomes also been verified by two-photon Microscope observation. All the in vivo experiments point out that HAb18G/CD147 promote cancer metastasis.On the other hand, we analysed the relationship between the expression of HAb18G/CD147 and TNM stages by case collection. And we found that there are correlations between the expression of HAb18G/CD147 and TNM stages. All these clinical evidences also proved that HAb18G/CD147 promote cancer progression.All in all, HAb18G/CD147 can significantly promote cancer metastasis and the influence malignant phenotypes via promote lamellipodium formation. And the promoting is through a signal pathway which involve a series of sequence phosphorylation from HAb18G/CD147, Fak, Src to Cortactin. In vivo and clinical research also validated that HAb18G/CD147 promote cancer progression.