BACKGROUND/AIMS: Over the past decade, heat shock protein 90 (Hsp90) has emerged as a potential therapeutic target for cancer. However, the molecular mechanisms of down-regulation Hsp90 expression in osteosarcoma are incompletely understood. To develop potential therapy targeting Heat shock protein 90B1 (Hsp90B1), we studied the roles of miR- 223 in the proliferation and apoptosis of human osteosarcoma. METHODS: pcDNA3.1(+)- miR-223 plasmid vectors were constructed and transfected into MG63 cells. Co-transfection of miR-223 expression vector with pMIR-Hsp90B1 (The recombined vector of pMIR-GLOTM luciferase vector containing Hsp90B1-3'UTR) led to the reduced activity of luciferase in a dual-luciferase reporter gene assay, suggesting that Hsp90B1 is a target gene of miR-223. Expression of HSP90B1 was detected by RT-PCR and western blotting analysis. Cell proliferation was determined using the MTT assay. Cell-cycle distribution and apoptosis were examined by flow cytometry. PI3K, p-Akt, Akt, mTOR, Bcl-2 and Bid were also detected by western blotting analysis. After a mouse xenograft model of human MG63 tumors was constructed, tumor growth, microvessel density and proliferation in each group was determined. RESULTS: The pcDNA3.1(+)-miR-223 vector efficiently suppressed the expression of HSP90B1, while silencing miR-223 increased expression of Hsp90B1. Furthermore, overexpression of miR-223 results in significant inhibition of cell growth on culture plates. Moreover, cancer cells showed significant G0/G1 arrest and increased apoptosis due to gene silencing. Protein levels of PI3k, p-Akt, mTOR, and Bcl-2 were decreased, whereas Bid levels were increased. Microvessel density as assessed by CD34 levels and cell growth by PCNA levels decreased according to immunohistochemical analysis. CONCLUSION: Hsp90B1 is a direct target of miR-223 and miR- 223 may have a tumor suppressor function in osteosarcoma through the PI3K/Akt/mTOR pathway and could be used in anticancer therapies in osteosarcoma.
BACKGROUND/AIMS: Over the past decade, heat shock protein 90 (Hsp90) has emerged as a potential therapeutic target for cancer. However, the molecular mechanisms of down-regulation Hsp90 expression in osteosarcoma are incompletely understood. To develop potential therapy targeting Heat shock protein 90B1 (Hsp90B1), we studied the roles of miR- 223 in the proliferation and apoptosis of humanosteosarcoma. METHODS: pcDNA3.1(+)- miR-223 plasmid vectors were constructed and transfected into MG63 cells. Co-transfection of miR-223 expression vector with pMIR-Hsp90B1 (The recombined vector of pMIR-GLOTM luciferase vector containing Hsp90B1-3'UTR) led to the reduced activity of luciferase in a dual-luciferase reporter gene assay, suggesting that Hsp90B1 is a target gene of miR-223. Expression of HSP90B1 was detected by RT-PCR and western blotting analysis. Cell proliferation was determined using the MTT assay. Cell-cycle distribution and apoptosis were examined by flow cytometry. PI3K, p-Akt, Akt, mTOR, Bcl-2 and Bid were also detected by western blotting analysis. After a mouse xenograft model of human MG63 tumors was constructed, tumor growth, microvessel density and proliferation in each group was determined. RESULTS: The pcDNA3.1(+)-miR-223 vector efficiently suppressed the expression of HSP90B1, while silencing miR-223 increased expression of Hsp90B1. Furthermore, overexpression of miR-223 results in significant inhibition of cell growth on culture plates. Moreover, cancer cells showed significant G0/G1 arrest and increased apoptosis due to gene silencing. Protein levels of PI3k, p-Akt, mTOR, and Bcl-2 were decreased, whereas Bid levels were increased. Microvessel density as assessed by CD34 levels and cell growth by PCNA levels decreased according to immunohistochemical analysis. CONCLUSION:Hsp90B1 is a direct target of miR-223 and miR- 223 may have a tumor suppressor function in osteosarcoma through the PI3K/Akt/mTOR pathway and could be used in anticancer therapies in osteosarcoma.
Authors: G M Viera; K B Salomao; G R de Sousa; M Baroni; L E A Delsin; J A Pezuk; M S Brassesco Journal: Clin Transl Oncol Date: 2019-04-04 Impact factor: 3.405
Authors: Mirte Mayke Streppel; Shweta Pai; Nathaniel R Campbell; Chaoxin Hu; Shinichi Yabuuchi; Marcia Irene Canto; Jean S Wang; Elizabeth A Montgomery; Anirban Maitra Journal: Clin Cancer Res Date: 2013-06-11 Impact factor: 12.531