title: [Inhibitory effect and significance of rapamycin on the mammalian target of rapamycin signaling pathway in osteosarcoma stem cells and osteosarcoma cells].
abstract: OBJECTIVE
  To observe the effect of rapamycin on the MG-63 osteosarcoma cells (OC), osteosarcoma stem cells (OSC) and on mTOR signaling pathway, and explore the feasibility of rapamycin as a novel therapeutic measure in osteosarcoma chemotherapy regimens.
  
  
  METHODS
  OC and OSC were cultured in vitro. Immunofluorescence assay was used to detect the expression of Nanog and Oct4 in OC and OSC. OC and OSC were treated with rapamycin in concentrations of 0, 20, 50 and 100 nmol/L. Semi-quantitative PCR and RT-PCR were used to detect the mTOR mRNA and CCK-8 assay was used to detect cell proliferation, and the cell morphology was observed under an inverted microscope.
  
  
  RESULTS
  The cores of MG-63 cellular spheres exhibited embryonic stem cell characteristics such as Nanog and Oct4 expession. The mTOR pathway was activated in the OSC and the expression of mTOR mRNA was higher in OSC (0.761 ± 0.080) than that in OS (0.406 ± 0.090, P < 0.05) by semi-quantitative PCR. RT-PCR showed that the expression of mTOR mRNA was lower in OSCs treated with 100 nmol/L rapamycin (0.961 ± 0.060) than that with 0 nmol/L rapamycin (1.654 ± 0.246, P < 0.05). Cell counting kit-8 (CCK-8) assay showed that the proliferation of OC treated with 20, 50 and 100 nmol/L rapamycin was significantly inhibited, compared with that with 0 nmol/L rapamycin (P < 0.05). Compared with 0 nmol/L rapamycin, the proliferation of OSC treated with 20 and 50 nmol/L rapamycin was not significantly inhibited (P > 0.05), but that with 100 nmol/L rapamycin was significantly inhibited (P < 0.05). The invert microscopic observation revealed that rapamycin inhibited the formation of OSC spheres.
  
  
  CONCLUSIONS
  Rapamycin can effectively inhibit cell proliferation and the ability of sphere formation of OSCs. It will provide a basis for a novel therapeutic approach in osteosarcoma chemotherapy regimens.
authors.affiliation: Shanghai Institute of Orthopedics and Traumatology, Shanghai Key Laboratory for Prevention and Treatment of Bone and Joint Diseases with Integrated Chinese-Western Medicine, Shanghai 200025, China.
authors.forename: Pei-yi
authors.initials: PY
authors.lastname: Liu
authors.forename: Wei-bin
authors.initials: WB
authors.lastname: Zhang
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
authors.forename: Yu-hui
authors.initials: YH
authors.lastname: Shen
authors.forename: Yi-yong
authors.initials: YY
authors.lastname: Wei
mesh_terms: D000903:Antibiotics, Antineoplastic / Q000008:administration & dosage / Q000494:pharmacology; D001859:Bone Neoplasms / Q000378:metabolism / Q000473:pathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D000071317:Nanog Homeobox Protein; D014411:Neoplastic Stem Cells / Q000378:metabolism / Q000473:pathology*; D050814:Octamer Transcription Factor-3 / Q000378:metabolism; D012516:Osteosarcoma / Q000378:metabolism / Q000473:pathology*; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction; D020123:Sirolimus / Q000008:administration & dosage / Q000494:pharmacology*; D058570:TOR Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*
chemical_list: D000903:Antibiotics, Antineoplastic; D018398:Homeodomain Proteins; C486780:NANOG protein, human; D000071317:Nanog Homeobox Protein; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; D012333:RNA, Messenger; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus