title: [Transfection of tumor metastasis suppressor gene nm23-H1 can up-regulate the activity of GSK-3β in human high-metastasis large cell lung cancer cell line L9981].
abstract: BACKGROUND
  To investigate the influence of tumor metastasis suppressor gene nm23-H1 on the activity of glycogen synthase kinase 3β (GSK-3β) in human high-metastasis large cell lung cancer cell line L9981.
  
  
  METHODS
  The levels of GSK-3β expression in cytoplasm and nucleus were determined with anti- GSK-3β antibody in human high-metastasis large cell lung cancer cell line L9981 (cell line with nm23-H1 gene deletion), L9981-nm23-H1 (cell line with nm23-H1 transfected) and L9981-pLXSN (cell line with vector transfected) by Western blot method. The activity of GSK-3β among those three cell lines was detected by immunoprecipitation and analysed by a radioactive isotope scintillation counter before and after treating with 20 mmol/L LiCl.
  
  
  RESULTS
  (1) The expression indensity of GSK-3β of cytoplasm and nucleus was (6 341±541) and (4 356±490) IOD in L9981-nm23-H1, (3 613±383) and (705±75) IOD in L9981-pLXSN, and (3 736±298) and (675±57) IOD in L9981, respectively. A high significance in GSK-3β expressive indensity of both cytoplasm and nucleus existed among L9981-nm23-H1, L9981-pLXSN and L9981 (P < 0.01); Multiple comparison: A highly significant difference was observed when L9981-nm23-H1 was compared with L9981-pLXSN or L9981 (P < 0.01), but no significant difference was observed between L9981-pLXSN and L9981 (P > 0.05). (2) The GSK-3β activity of cytoplasm and nucleus was (28 955±2 509) and (9 247±924) CPM in L9981-nm23-H1, (11 241±1 495) and (1 492±176) CPM in L9981-pLXSN, and (12 505±1 469) and (1 763±125) CPM in L9981, respectively. A highly significant difference in GSK-3β activity of both cytoplasm and nucleus existed among L9981-nm23-H1, L9981-pLXSN and L9981 (P < 0.01); Multiple comparison: the GSK-3β activity in L9981-nm23-H1 was significantly higher than that in L9981-pLXSN and L9981 (P < 0.01), but no significant difference was observed between the L9981-pLXSN and L9981 (P > 0.05). (3) After treatment with 20 mmol/L LiCl, the expressive indensity of GSK-3β of cytoplasm and nucleus was (4 718±549) and (3 823±350) IOD in L9981-nm23-H1, (2 030±155) and (217±15) IOD in L9981-pLXSN, and (2 164±151) and (224±19) IOD in L9981, respectively. No significant difference in GSK-3β expressive indensity existed between before and after treatment with LiCl in L9981-nm23-H1 (P > 0.05). However, the GSK-3β expressive indensity in cytoplasm and nucleus before treatment was remarkably higher than those after treatment in both L9981-pLXSN and L9981 (P < 0.05). (4) After treatment with 20 mmol/L LiCl, the GSK-3β activity in cytoplasm and nucleus was (11 099±1 112) and (3 748±215) CPM in L9981-nm23-H1, (4 447±430) and (1067±159) CPM in L9981, and (4 435±427) and (909±156) CPM in L9981-pLXSN, respectively. The GSK-3β activity both in cytoplasm and nucleus after treatment with LiCl was remarkably lower than that before treatment in L9981-nm23-H1, L9981-pLXSN and L9981 (P < 0.01 or P < 0.05).
  
  
  CONCLUSIONS
  (1) Transfection of nm23-H1 gene can significantly up-regulate the expression level and activity of GSK-3β in human high-metastasis large cell lung cancer cell line L9981; (2) LiCl can remarkably suppress the upregulation effects of nm23-H1 gene on GSK-3β activity in L9981 cell line; (3) The effects of nm23-H1 gene on suppressing the signal transduction of Wnt pathway might be carried out through upregulating GSK-3β expression and activity in human high-metastasis large cell lung cancer cell line L9981.
authors.affiliation: Key Laboratory of Lung Cancer Molecular Biology of Sichuan Province and Department of Thoracic Surgery, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P.R.China.
authors.forename: Junke
authors.initials: J
authors.lastname: Fu
authors.forename: Qinghua
authors.initials: Q
authors.lastname: Zhou
authors.forename: Wen
authors.initials: W
authors.lastname: Zhu
authors.forename: Yanping
authors.initials: Y
authors.lastname: Wang
authors.forename: Xiaohe
authors.initials: X
authors.lastname: Chen
authors.forename: Guowei
authors.initials: G
authors.lastname: Che
authors.forename: Qiang
authors.initials: Q
authors.lastname: Nie
authors.forename: Dingbiao
authors.initials: D
authors.lastname: Li
authors.forename: Lunxu
authors.initials: L
authors.lastname: Liu
authors.forename: Yin
authors.initials: Y
authors.lastname: Li