| Cervical carcinoma is one of the most common tumor of women. There are lots of clinical and experimental studies about the causeing of this disease, but the molecule mechanism is not very clearly. Several studies have demonstrated that Cx26 and Cx 43 express in normal cervix. But in malignant cell lines and tumours, disruption of gap junctional intercellular communication and/or connexins (gap junction proteins) is frequently reported The expression of connexin is also associated with cervical tumour progression. Lack of some gap junction protein and interruption of GJIC is one of the mechanism that tumor cells escape from cell differentiation monitor.Gap junction, a kind of intercellular connection form, commonly existed in animal tissues, and constructed of a multigene family of integral membrane proteins, the connexins (Cx), with over 20 connexin isoforms extending from 25 to 62 kDa found in human genomes. Gap junction channels span two plasma membranes and result from the docking of two half channels, or connexons, which arehexameric torus of connexins around an aqueous pore, providing a direct route permit the intercytoplasmic exchange of small metabolites and ions between two adjacent cells. This exchange of substance and information is called gap junction intercellular communication(GJIC).Gap junction regulates the process of metabolism, proliferation and differentiation. It appears to be important event in metabolism balance and growth of organism. Abnormal functional correlates closely with carcinogenesis. Compare with normal cells, most tumor cells absent of connexin and lack of gap junctions. It was reported that Cx inhibits the growth of tumor cells by GJIC or other ways.In our trial, we investigated the expression and signal transduction regulation of gap junction protein gene in cervical malignant cell lines, and its direct effects to the GJIC, attempted to enucleate the etiological mechanism of the disease, and to discuss the . possible gene therapy methods.1 we applied RT-PCR, Western blot and flow cytometry (FCM) to analyze the expression of Cx26, Cx43 gene in normal cervix tissue and the human cervical carcinoma cell line HeLa, And found that they are expressed in cervix tissue but not in HeLa cell.2 By gene recombinant technology, we constructed Cx26, Cx43 eukaryotic expression vector and transfected into HeLa cells. By Immunohistochemical staining and LSCM, we detect the expression of Cx., observed the effects of Cx transfection to cells. We found that the two Cxco-expreesed in the transfection cells and cell growth and proliferation was declined.3 We observed the effect of retinoic acid(RA) stimulate cells in time-dose dependant fashion. Lucefer yellow scrape loading and dye transfer(SLDT) showed that the the transfection cells resume function gap junction channel, and RA enhance the GJIC.4 After Fluo-3 AM loading ,laser scanning confocal microscope(LSCM) examined intercelluar to analyze the change of free [Ca2+]i. We found the concentration of [Ca2+]i reduced in the transformed cell and after RA stimulate it became markedly. Furthermore, Western blot was used to detect the tyrosine phosphorylation of Cx43 in transfected cell and the expression of activated Raf-K ERK in order to find out the possible signal transduction pathway.From all above, we can draw the following conclusion:1. Cx26, Cx43 gene and protein can be detected in the normal cervix tissue and can not in cervical carcinoma cell line HeLa.2. Cx26, Cx43 gene transfection recovered the expression of Cx26, Cx43 protein in HeLa cells, induced the functional GJIC, inhibited the growth of the cells.3. Stimulation of RA can augment the growth inhibition of transfected cells in time-dose dependent fashion.4. Stimulation of RA can augment the communicate function, reduce the concentration of free [Ca2+], RA enhance Cx43 phosphorylation and activate ERK...
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