PMID- 27411279
OWN - NLM
STAT- MEDLINE
DCOM- 20160930
LR  - 20181202
IS  - 1002-1892 (Print)
IS  - 1002-1892 (Linking)
VI  - 30
IP  - 4
DP  - 2016 Apr
TI  - [EFFECT OF Akt1 GENE TRANSFECTION ON HYPOXIA TOLERANCE OF BONE MARROW MESENCHYMAL 
      STEM CELLS].
PG  - 479-84
AB  - OBJECTIVE: To investigate whether Akt1 gene transfection mediated by recombinant 
      lentivirus (LVs) in the bone marrow mesenchymal stem cells (BMSCs) could enhance 
      the ability of hypoxia tolerance so as to provide a theoretical basis for 
      improving the effectiveness of stem cells transplantation. METHODS: LVs was used 
      as transfection vector, enhanced green fluorescent protein (EGFP) was used as 
      markers to construct the pLVX-EGFP-3FLAG virus vector carrying the Akt1 gene. The 
      3rd generation BMSCs from 3-5 weeks old Sprague Dawley rats were transfected with 
      pLVX-EGFP virus solution as group B and with pLVX-EGFP-3PLAG virus solution as 
      group C; and untransfected BMSCs served as control group (group A). At 2-3 days 
      after transfection, the expression of green fluorescent was observed by 
      fluorescence microscope; and at 48 hours after transfection, Western blot method 
      was used to detect the expression of Akt1 protein in groups B and C. BMSCs of 
      groups B and C were given hypoxia intervention with 94% N(2), 1% O(2), and 5% CO(2) for 
      0, 3, 6, 9, and 12 hours (group B1 and group C1). The flow cytometry was used to 
      analyze the cell apoptosis rate and cell death rate, and the MTT method to 
      analyze the cell proliferation, and Western blot to detect the expression of 
      apoptosis related gene Caspase-3. RESULTS: After transfection, obvious green 
      fluorescence was observed in BMSCs under fluorescence microscopy in groups B and 
      C, the transfection efficiency was about 60%. Akt1 expression of group C was 
      significantly higher than that of group B (t = 17.525, P = 0.013). The apoptosis 
      rate and cell death rate of group B1 increased gradually with time, and 
      difference was significant (P < 0.05). In group C1, the apoptosis rate and cell 
      death rate decreased temporarily at 3 hours after hypoxia intervention, then 
      increased gradually, and difference was significant (P < 0.05). The apoptosis 
      rate and cell death rate of group C1 were significantly lower than those of group 
      B1 at each time point (P < 0.05) except at 0 hour. MTT assay showed tat 
      absorbance (A) values of groups B and C were significantly higher than those of 
      groups B1 and C1 at each time point (P < 0.05); the A value of group B was 
      significantly lower than that of group C at each time point (P < 0.05). The A 
      value of group B1 was significantly lower than that of group Cl at 6, 9, and 12 
      hours after hypoxia intervention (P < 0.05). Western blot results showed that the 
      Caspase-3 expression of group C1 significantly reduced when compared with group 
      B1 at each time point (P < 0.05). CONCLUSION: Akt1 gene transfection mediated by 
      recombinant LVs could significantly improve hypoxia tolerance of BMSCs by 
      inhibiting the apoptosis, which could provide new ideas for improving the 
      effectiveness of stem cells transplantation.
FAU - Yu, Fengxu
AU  - Yu F
FAU - Chen, Yongen
AU  - Chen Y
FAU - Chen, Feng
AU  - Chen F
FAU - Xia, Jiyi
AU  - Xia J
FAU - Liu, Hongduan
AU  - Liu H
FAU - Fu, Yong
AU  - Fu Y
FAU - Li, Miaoling
AU  - Li M
FAU - Liao, Bin
AU  - Liao B
LA  - chi
PT  - Journal Article
PL  - China
TA  - Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi
JT  - Zhongguo xiu fu chong jian wai ke za zhi = Zhongguo xiufu chongjian waike zazhi = 
      Chinese journal of reparative and reconstructive surgery
JID - 9425194
RN  - 0 (Vascular Endothelial Growth Factor A)
RN  - 0 (enhanced green fluorescent protein)
RN  - 147336-22-9 (Green Fluorescent Proteins)
RN  - EC 2.7.11.1 (Akt1 protein, rat)
RN  - EC 2.7.11.1 (Proto-Oncogene Proteins c-akt)
RN  - EC 3.4.22.- (Caspase 3)
SB  - IM
MH  - Animals
MH  - Bone Marrow Cells/cytology
MH  - Caspase 3
MH  - Cell Culture Techniques
MH  - Cell Differentiation
MH  - Cell Proliferation
MH  - Cells, Cultured
MH  - Genetic Vectors
MH  - Green Fluorescent Proteins
MH  - Hematopoietic Stem Cells
MH  - Immune Tolerance
MH  - Lentivirus/*genetics
MH  - Mesenchymal Stem Cells/*cytology/metabolism
MH  - Proto-Oncogene Proteins c-akt/*genetics
MH  - Rats
MH  - Rats, Sprague-Dawley
MH  - Tissue Engineering
MH  - *Transfection
MH  - Vascular Endothelial Growth Factor A/*genetics
EDAT- 2016/07/15 06:00
MHDA- 2016/10/01 06:00
CRDT- 2016/07/15 06:00
PHST- 2016/07/15 06:00 [entrez]
PHST- 2016/07/15 06:00 [pubmed]
PHST- 2016/10/01 06:00 [medline]
PST - ppublish
SO  - Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2016 Apr;30(4):479-84.