Cell proliferation and colony formation assays revealed that overexpression of miR-148a diminished the proliferation of these cell lines , whereas miR-148a inhibition enhanced the proliferation of those cell lines . Overexpression of HPIP reversed the impact of miR-148a on HepG2 cell proliferation . Soft agar assay showed that miR-148a inhibited anchorage-independent HepG2 cell proliferation . Again, introduction of HPIP reversed the effect of miR-148a on anchorage-independent HepG2 cell proliferation . These effects suggest that miR-148a inhibits hepatoma cell proliferation by targeting HPIP. Subsequent, we examined the effects of miR-148a on migration and invasive capability of hepatoma cells. miR-148a overexpression suppressed cell migration in HepG2, SMMC-7721, and BEL- 7402 cells utilizing a wound-healing assay . Matrigel invasion assays demonstrated that miR- 148a overexpression decreased the quantity of invaded cells in these cell lines .
Conversely, miR-148a inhibition had opposite results . HPIP reexpression in miR-148a-HepG2 cells reversed the effects of miR-148a on cell migration and invasion . Importantly, equivalent benefits have been observed in HBx-expressing MHCC97-H cells . Therefore, we examined direct results of miR-148a on HBx-mediated selleck chemicals you can find out more development and migration of hepatocytes. As anticipated, HBx greater LO2 cell development and migration . Intriguingly, these effects had been rescued by miR-148a reexpression. Equivalent effects have been observed in HepG2 cells . These information suggest that HBx enhances liver cell development and migration as a result of inhibition of miR-148a. miR-148a inhibits EMT through inhibition of HPIP expression. Since EMT is properly regarded to become involved in invasion and metastasis of cancer cells , we tested the effects of miR-148a on EMT in MHCC97-H cells.
miR-148a overexpression inhibited morphologic adjustments from a polarized epithelial phenotype, which brought about an elongated fibroblastoid phenotype , suggesting that b-AP15 concentration miR-148a suppresses EMT. Furthermore, miR-148a greater expression in the epithelial marker E-cadherin and decreased that of your E-cadherin repressor Snail also as N-cadherin and Vimentin, two mesenchymal markers, accompanied through the inhibition of mTOR signaling . The observed miR-148a¨Cmediated phenotype was rescued by HPIP overexpression. Also, miR-148a reversed HBx-mediated effects on EMT and mTOR signaling . miR-148a also inhibited EMT in HepG2 cells . These success suggest that miR-148a might possibly control HCC progression and metastasis via regulation of EMT. miR-148a inhibits tumor growth and metastasis of HCC in nude mice.
To confirm the in vitro phenotype of miR-148a expression, we to begin with examined the effect of miR-148a on HepG2 cell growth in nude mice. miR-148a markedly suppressed tumor growth . As expected, the tumors in mice inoculated with miR-148a- HepG2 cell lines had diminished amounts of HPIP and phosphorylation of mTOR, S6K1, and 4E-BP1 as well as the mTOR effectors c-myc and cyclin D1 .