Effect of Src homology 3 domain domains of intersectin1-S on proliferation and migration of glioma cells
TIAN Chun-ying, LIU Xiao-li, LI Zhi-hui, GU Feng, MA Yong-jie
Published 2013-02-08
Cite as Chin J Exp Surg, 2013,30(02): 293-296. DOI: 10.3760/cma.j.issn.1001-9030.2013.02.033
Abstract
Objective To investigate the effects of Src homology 3 domain (SH3) of intersectin 1-S (ITSN1-S) on proliferation and migration of glioma cell line LN229 and the possible molecular mechanisms.Methods ITSN1-S and fragmental EH1-EH2-CC domain of ITSN1-S were spliced into the linear PCDH-CMV-MCS-EF1-Puro vector which was excised by incision enzyme digestion,and PCDH-CMV-MCS-EF1-Puro vector without treatment served as control group.Each of the three recombinant plasmids was transfected into the LN229 cells by lentivirus,and the stably expressed cell clones were screened.Western blotting was applied to detect the expression of each protein.MTT assay was performed to detect proliferation of LN229 cells persistently cultured for 6 days.Soft agar assay was performed to detect the colony formation ability of glioma cells persistently cultured for 2 weeks.Wound-healing assay was performed to detect the migration of LN229 cells at 0,3,6,9,12,24 h.Results MTT assay and soft agar assay showed that ITSN1-S total-length group proliferated more rapidly than EH1-EH2-CC fragment group and vector control group (P ＜ 0.05),but there was no significant difference between EH1-EH2-CC fragment group and vector control group (P ＞ 0.05).The proliferation rate at 6th day in ITSN1-S total-length group,EH1-EH2-CC fragment group and vector control group was (523.60 ± 32.12)％,(409.54 ± 31.33)％and (353.89 ± 13.98) ％,respectively.The clone formation rate at 14th day in ITSN1-S total-length group,EH1-EH2-CC fragment group and vector control group was (46.49 ± 2.34)％,(23.00 ±1.66) ％ and (17.68 ± 3.47)％,respectively.Wound-healing assay revealed that the cells in ITSN1-S total-length group migrated faster than in EH1-EH2-CC fragment group and vector control group after 12 h (P ＜0.05),but there was no significant difference between EH1-EH2-CC fragment group and vector control group (P ＞0.05).The migration distance at 24th h in ITSN1-S total-length group,EH1-EH2-CC fragment group and vector control group was (0.388 ± 0.023) mm,(0.307 ± 0.021) mm and (0.287 ±0.011) mm,respectively.Conclusion ITSN1-S is involved in regulating the proliferation and migration of glioma cells.The SH3 domains were the crucial functional domain of ITSN1-S to regulate the proliferation and migration of glioma cells.
Key words:
Src homology 3 domain; Glioma; Proliferation; Migration
Contributor Information
TIAN Chun-ying
Research Institute of Tumor Cell Biology Laboratory of Tumour Hospital Affiliated to Tianjin Medical University, Tianjin Municipal Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China
LIU Xiao-li
Research Institute of Tumor Cell Biology Laboratory of Tumour Hospital Affiliated to Tianjin Medical University, Tianjin Municipal Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China
LI Zhi-hui
Research Institute of Tumor Cell Biology Laboratory of Tumour Hospital Affiliated to Tianjin Medical University, Tianjin Municipal Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China
GU Feng
Research Institute of Tumor Cell Biology Laboratory of Tumour Hospital Affiliated to Tianjin Medical University, Tianjin Municipal Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China
MA Yong-jie
Research Institute of Tumor Cell Biology Laboratory of Tumour Hospital Affiliated to Tianjin Medical University, Tianjin Municipal Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China
