| Objective To investigate the therapeutic potential application of Eeyarestatin I (EerI) and4μ8C, the small-molecule inhibitors of UPR signaling pathway in the cervical cancer.Method The protein expression of GRP78/BiP, p97, Ubiquitin and IRE1α in normal cervical epithelia and cervical cancer were examined by immunohistochemistry. The effect of EerI and4μ8C on HeLa cell proliferation and apoptosis were measured by MTS assay and flow cytometry, respectively. The combined antiproliferative effects of small-molecule inhibitors were carried out in combination of Bortezomib (BTZ) and Cisplatin (CDDP), respectively.Result GRP78/BiP, p97, Ubiquitin and IRE1α were significantly up-regulated in cervical cancer compared with normal cervical epithelia and displayed variability in the cervical cancer of different clinical stages (P<0.05). Eerl and4μ8C inhibited HeLa cell proliferation in a dose-dependent manner with inducing HeLa cell apoptosis in a similar fashion (P<0.05). MTS assays indicated that combination treatment significantly enhanced the antiproliferative effect of single chemotherapy drugs (P<0.05).Conclusion Small-molecule inhibitors targeting the key players of UPR show synergistic effects with other anticancer drugs, suggesting a new therapeutic strategy towards cervical cancer. Objective To investigate the morphological changes, cell repolarization and function of the long-term cultured primary mouse hepatocytes in the sandwich configuration. Moreover, to study the important link between energy metabolism and polarity in hepatocytes.Method Hepatocytes were isolated by a two-step collagenase perfusion and primary mouse hepatocytes were cultured in a sandwich configuration. The morphological changes were observed under an inverted microscope and the canalicular network formation was determined by CDF imaging. By western blot detection of tight junction protein expression changes, using real-time PCR detection of tight junction protein expression levels changes of mRNA.Result Cell viability was80%-90%. Hepatocytes cultured in monolayer exhibited a morphology rather flattener than that of sandwich-cultured counterparts. Sandwich-cultured hepatocytes, but not monolayer-cultured counterparts, displayed extensive canalicular networks. The intensity of CDF accumulation in the bile canaliculi appeared to be more extensive in mouse hepatocytes cultured in sandwich configuration compared to the monolayer configuration. The expression of ZO-1and claudin-2mRNA and their protein were significantly higher in high fat diet than that in normal diet, differences consisted in individuals of one group.Conclusion Primary mouse hepatocytes in sandwich-culture configuration displays extensive canalicular networks. These bile canaliculi-like structures were moreover fully functional since they strongly accumulated the fluorescent dye CF. In addition, the expression changes of tight junction protein were caused by the differences in animal individual, suggesting that hepatocytes polarization parallels changes in energy metabolism. |
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