hsa-miR-485-5p reverses epithelial to mesenchymal transition and promotes cisplatin-induced cell death by targeting PAK1 in oral tongue squamous cell carcinoma.

Oral squamous cell carcinoma (OSCC) is currently a highly prevalent disease worldwide. Cisplatin (CDDP) is widely used for the chemotherapy of OSCC. Yet, the molecular mechanisms responsible for cisplatin resistance have not been fully elucidated. In this study, we showed that overexpression of p21 (RAC1) activated kinase 1 (PAK1) induced epithelial to mesenchymal transition (EMT) and significantly promoted the invasion and migration of oral squamous cell carcinoma SCC25 cells. Emerging evidence indicates a strong link between resistance to therapy and the induction of EMT in cancer. We showed that overexpression of PAK1 induced cisplatin resistance in SCC25 cells. ERCC1 and YAP can promote cisplatin resistance in human OSCC. We showed that ERCC1 and YAP protein were upregulated by PAK1 in SCC25 cells. -We found that miR­485­5p inhibited PAK1 protein expression in the SCC25 cells. Contrary to PAK1, we demonstrated that overexpression of miR­485­5p reversed EMT and significantly inhibited invasion and migration. Moreover, its overexpression sensitized SCC25-CR cells (cisplatin-resistant cells) to cisplatin. Thus, we conclude that miR­485­5p reverses EMT and promotes cisplatin-induced cell death by targeting PAK1 in oral tongue squamous cell carcinoma. This study suggests that PAK1 plays an essential role in the progression of OSCC and it is a potential therapeutic target for OSCC.

[The effect of matrix metalloproteinase-1 on root surface dentin matrix: a scanning electron microscope observation].

OBJECTIVE: To observe the effect of matrix metalloproteinase-1 (MMP-1) from human host on degradation of dentin organic matrix of root dentin. METHODS: The freshly extracted caries-free impacted teeth were selected. Teeth were cut transversely under the enamel-cementum junction into dentin sections with a thickness of about 5 mm. Then all sections with removal of cementum, pulp and predentin were randomly divided into four groups. In the first group, dentin sections were demineralized with acid solution for 21 days, and then incubated with MMP-1 solution for 7 days; the second group were only treated with acid solution for 21 days; the third group were only attacked by MMP-1 solution for 7 days; and the fourth group were untreated as a control. Then all sections were dehydrated in ascending strength of alcohol, critically dried, coated with platinum, and then observed under scanning electron microscope(SEM). RESULTS: The dentin sections of root surface attacked by acid and MMP-1 showed that demineralization of dentin mineral and degradation of dentin matrix fibrae synchronously happened. The dentin matrix fibrae wasn't degradated in the groups treated with acid or MMP-1. CONCLUSION: The proteinases from human host may play an important role in the development of root surface caries. MMP-1 may distinctly degradate the organic matrix of demineralized dentin.

Reconstitution of Src-dependent phospholipase Cgamma phosphorylation and transient calcium release by using membrane rafts and cell-free extracts from Xenopus eggs.

We reported previously that egg membrane rafts serve as a subcellular microdomain for sperm-dependent tyrosine kinase signaling in Xenopus fertilization. Moreover, we demonstrated that raft-associated Src tyrosine kinase was activated by sperm in vitro. Here we show that egg rafts incubated with sperm or hydrogen peroxide (H2O2) can promote Src-dependent phosphorylation of phospholipase Cgamma (PLCgamma) and transient calcium release in the extracts of unfertilized Xenopus eggs. In vivo egg activation by sperm or H2O2 also promotes tyrosine phosphorylation and raft-translocalization of PLCgamma. Immunodepletion of PLCgamma from the egg extracts inhibits the raft-dependent calcium release. Rafts prepared from H2O2-activated eggs also promote Src-dependent dephosphorylation of p42 mitogen-activated protein kinase and cell cycle transition from metaphase II to interphase in egg extracts. PLCgamma phosphorylation and calcium release in egg extracts can be promoted by rafts prepared from COS-7 cells expressing the Xenopus Src gene. These results demonstrate that the signaling events elicited by fertilization in Xenopus eggs can be reconstituted in vitro. The development of such experimental platforms will allow us to dissect the molecular mechanism of sperm-dependent activation of raft-associated Src and subsequent up-regulation of PLCgamma and egg activation machinery in Xenopus eggs.

Porcine sperm factor supports activation and development of bovine nuclear transfer embryos.

A study was undertaken to determine whether injection of porcine sperm factors (pSF), which trigger oscillations in intracellular calcium concentration ([Ca(2+)](i)) in mammalian oocytes, could be used to activate bovine oocytes during nuclear transfer. To date, only combined treatments that induce a monotonic rise in [Ca(2+)](i) and inhibit either phosphorylation or protein synthesis have been utilized in nuclear transfer. Several doses of pSF were tested. Injection of 5 mg/ml pSF triggered [Ca(2+)](i) oscillations that resembled those associated with fertilization with respect to amplitude and periodicity, and as a result, a high percentage of oocytes underwent activation. Furthermore, this concentration of pSF supported in vitro and in vivo development up to 60-90 days of gestation, comparable to development in control nuclear transfer embryos. Nevertheless, neither activation procedure supported development as well as did fertilization. The effectiveness of pSF as an activating agent in bovine oocytes may have been compromised because pSF was unable to support oscillations past 3-5 h postinjection and a second injection was necessary to extend the [Ca(2+)](i) oscillations. Likewise, a single injection of pSF failed to trigger downregulation of the inositol 1,4,5-trisphosphate receptor 1 subtype, whereas a second injection downregulated the receptor in a manner similar to that seen in fertilized oocytes. These results demonstrate that soluble factor(s) from porcine sperm can support early development in bovine nuclear transfer embryos; however, the efficacy may be limited because of the premature cessation of the induced oscillations.