Expression Of Calcium-related Proteins In Cystic Ovarian Cancer

As a kind of common gynecologic tumor, the five-year mortality rate of ovarian tumor has reached 60%-70%, it has become the highest fatality rate of gynecologic tumor, and seriously threaten to women’s health and life safety. With the development of Chinese society and changes of life style, there is an increasing trend in the incidence and mortality of ovarian tumor, it will still be one of the major diseases that threaten human health in the next few years.Ovarian tumor has the characteristics of difficult to discover and no typical symptoms in the early stage. Most tumors are diagnosed in its advanced stage which already appeared to widespread peritoneum metastasis. By then, radical surgery has been difficult to implement, which urgently need science researchers to make more indepth reaearch to the early diagnosis of ovarian tumor. It has been reported that the intake of calcium was positively correlated with the occurrence of ovarian tumor. Intracellular Ca2+in and out of cell mainly by a variety of calcium channel proteins distributed on the cell membrane, and Ryanodine receptor calcium release channel(RyR) has been confirmed that plays an important role in maintaining the intracellular calcium homeostasis. This study explored the expression patterns of the second messenger of calcium associated cell interior and exterior protein in ovarian tumor, including series of regulatory proteins of calcium channel protein compIex(RyR), FNs family proteins, cytoskeletal protein and oncogenes. It will provide valuable information for developing earlier effective diagnostic marker.In this study we used ovarian tumors in BALB/C mice model as the research object, the ovarian tumors (OvT) were divided into two parts:solid ovarian tumor (OvT-sol) and liquid ovarian tumor (OvT-liq). We detected the expression changes of all the regulatory proteins in both parts by Western Blot. The measured proteins included the regulatory proteins of channel protein(RyR), such as CAMR1, CAMR2, PP2A A, PP2A B, PP2A B55 and PP2A C; extracellular matrix protein FNs, such as FN-1, FN-2, FN-3, FN-4, FN-5 and N-cadherin; cytoskeletal protein β-tubulin, oncogene proteins P53 and PTEN. We detected the expression of CAMR1 and CAMR2 in normal ovary and ovarian tumor at the level of mRNA by RT-PCR. The histological and pathology changes of ovarian malignant tissue were analyzed by conventional staining methods, such as HE staining, PASM and Masson. Taking N-cadherin as contrast, we detected the expression distribution of CAMR1 and CAMR2 proteins in normal ovary by Immunohistochemistry method, and identified the expression changes in ovarian tumor.The results on the above criteria are as follows:(1) RT-PCR result showed that CAMR1 and CAMR2 were transcripted mRNA both in ovarian tumor and normal ovary at mRNA level.(2) Histologically conventional staining showed that ovarian tumor tissue has obvious change in the structure with follicle obvious atrophy and damaged or even disappeared. The nucleus enlarged, blood vessels, lymphatic vessels and adipose tissue increased. Cancer cells were diffusely distributed, basal membrane structure defect, loose structure, basement membrane lines and collagen fibers increased and deep staining. Tumor fibrosis degree increased.(3) Immunohistochemical results showed that the expression of CAMR1 and CAMR2 in normal ovarian tissue were lower than ovarian tumor tissue, mainly distributed in follicles of different development stages and the supportive tissue of ovary, but the expression in ovarian tumor tissue were up-regulated, the proteins mainly located in cytoplasm and lymphatic vessels. The high expression of N-cadherin in normal ovarian tissue, mainly distributed in the membranes of each follicle cell and the cell periphery of ovary connective tissue, but the expression of N-cadherin in ovarian tumor tissue was reduced or even disappeared.(4) The Western Blot results showed that, as the regulatory protein of RyR, CAMR1’s expression was significantly up-regulated in both OvT and OvT-sol, and was no significant change in the liquid tissue; the expression amount of CAMR2 in ovarian tumor tissues was significantly higher than normal ovarian tissue, especially the highest expression in OvT-sol. All above results were supported by immunohistochemical staining. The expression of PP2A A and PP2A B in ovarian tumor tissues were all significantly higher than normal ovarian tissues; the expression of PP2A B55 was also significantly higher than normal ovarian tissue, but the expression of PP2A B55 in OvT-liq was low, significantly lower than OvT and OvT-sol; the expression of PP2A C was all significantly down-regulated in ovarian tumor tissues, especially the lowest expression in OvT-liq, significantly lower than OvT and OvT-sol.The expression of ECM proteins FN-1 and FN-2 were all significantly up-regulated in ovarian tumor tissues, and the degree of up-regulated expression in OvT-liq was significantly lower than OvT and OvT-sol; the expression amount of FN-3 in OvT and OvT-liq were lower than normal ovarian tissue, but there were no significant difference, expression of FN-3 in OvT-sol was significantly lower than normal ovarian tissue; there was no significant difference of the FN-4 expression between ovarian tumor tissue and normal ovarian tissue; the expression of FN-5 in OvT was significantly higher than normal ovarian tissue, but there were no significant change in OvT-sol and OvT-liq.Compared with normal ovarian tissue, the expression amount of cytoskeleton protein β-tubulin in ovarian tumor tissues were increased, but there were no significant difference; the expression of N-cadherin in ovarian tumor tissues was significantly lower than normal ovarian tissue; the expression of P53 and PTEN in ovarian tumor tissues were all significantly higher than normal ovarian tissues, but the degree of up-regulated expression in OvT-liq was significantly lower than OvT.In conclusion, we systematically studied the expression level of calcium related proteins in mice ovarian tumor first time, and have a certain practical significance of the follow-up study of ovarian tumor at the cellular and molecular level. According to the detection of IHC, RT-PCR and Western Blot, we found that the expression of CAMR1 and CAMR2 were all significantly up-regulated in ovarian tumor, these proteins may be involved in the regulation of ovarian tumor occurrence and development process, probably become effective biomarkers for early diagnosis of ovarian tumor. The up-regulations of PP2A A, PP2AB, PP2A B55, FN-1, FN-2, FN-5, P53 and PTEN, and the down-regulations of PP2A C and N-cadherin in ovarian tumor indicate that calcium is closely associated with the tumor occurrence, and these proteins may be potential biomarkers for early diagnosis of ovarian tumor. Therefore, this study can provide valuable information and ways for the research of early diagnosis and treatment of ovarian tumor.