| Objective: To observe the effects and possible role of bone marrow mesenchymal stem cells genetically modified hepatocyte growth factor in a rat with acute lung injury induced by LPS.Methods: Sixty male F344 inbred line rats, 7 weeks old, weighting 200~250g, were divided into 3 groups randomly: Control group, EGFP-MSCs group(MSCs were transduced with empty vector), h HGF-MSCs group(MSCs were transduced with lentiviral vector modified with human HGF). All rats with acute lung injury(ALI) was induced by instillation of LPS(7 mg/kg) intratracheally. Immediately after LPS administration, rats were given either EGFP-MSCs(5×105 EGFP-MSCs in 1ml of DMEM, EGFP-MSCs treatment group) or h HGF-MSCs(5×105 h HGF-MSCs in 1ml of DMEM, h HGF-MSC treatment group) or 1ml of DMEM(Control treatment group)through caudal vein injection. The distribution of MSCs was traced by labelling the cells with 4?, 6-diamidino-2-phenylindole dihydrochloride(DAPI) by the laser scanning confocal microscope at days 7 after cells administration. Sixty rats experiment specimens were harvested at the day 1, day 3 and day 7 from every group after LPS administration: wet-to-dry ratio(W/D)of lung tissue and total protein and total cells and neutrophil count of BALF and severity scorings of lung injury were assessed; myeloperoxidase activity was measured by means of chemical colorimetry; total protein of BALF was measured by means of BCA protein determination method; the content of rat HGF(r HGF), h HGF and in serum and TNF-?, IL-10, IL-6 in lung tissue and BALF were measured by means of ELASA; the expression of TNF-? m RNA and IL-10 m RNA and IL-6 m RNA were measured by means of RT-q PCR. Apoptosis index of alveolar epithelial cells was measured by means of TUNEL. Thirty-six rats with ALI were divided into 3 groups randomly according to the grouping method above(n=12) and the survival rates within 90 days of rats among the 3 groups were observed after cells were transduced.Results:(1) A large number of green fluorescent cells were observed in the pulmonary interstitial among the EGFP-MSCs treatment group and the h HGF-MSCs treatment group while there were no green fluorescent cells observed in the Control group.(2) The h HGF was detected in h HGF-MSCs group, while it was not detected in other groups. Level of r HGF in serum was significantly reduced in the h HGF-MSCs treatment group compared with the EGFP-MSCs- and Controlgroups at day 7( P < 0.05).(3) Severity scoring of lung injury was significantly reduced in the EGFP-MSCs- and h HGF-MSCs- treatment groups compared with the Control group at the three time-points( P < 0.05). Compared with Control group, level of r HGF in serum was significantly increased in the EGFP-MSCs treatment group at day 3( P < 0.05) and increased in the h HGF-MSCs treatment group at day 1 and day 3( P < 0.05, P < 0.01). Compared with the EGFP-MSCs group, level of r HGF in serum was significantly increased in the h HGF-MSCs treatment group at day 3( P < 0.05).(4) W/D of lung was significantly reduced in the EGFP-MSCs- and h HGF-MSCs- treatment groups compared with the Control group at the day 1 and day 7( P < 0.01).(5) BALF total protein was significantly reduced in the EGFP-MSCs- and h HGF-MSCs- treatment groups compared with the Control group at at all the time-points( P < 0.05).(6) Total cells and neutrophil count of BALF were significantly reduced in the h HGF-MSCs treatment group compared with the Control group at the day 3 and day 7( P < 0.05, P < 0.01), compared with the EGFP-MSCs treatment group at the day 1( P < 0.01). Total cells and neutrophil count of BALF were significantly reduced in the EGFP-MSCs treatment group compared with the Control group at the day 7( P < 0.05).(7) There was not significantly different of Lung MPO activity between EGFP-MSCs treatment group and Control group at all the time-points. Lung MPO activity was significantly reduced in the h HGF-MSCs treatment groups compared with the Control group at the day 7( P < 0.05).(8) Compared with the Control group, level of TNF-? and IL-6 in BALF was significantly reduced in the EGFP-MSCs- and h HGF-MSCs- treatment groups at the day 1 and day 7( P < 0.05); level of IL-10 in BALF was significantly increased in the h HGF-MSCs treatment group at all the time point( P < 0.05, P < 0.01), level of IL-10 was significantly increased in the EGFP-MSCs treatment group at the day 1 and day 7( P < 0.01). Compared with the EGFP-MSCs treatment group, level of IL-6 in BALF was reduced at the day 7( P < 0.01), level of IL-10 was significantly increase at the day 3 and day 7( P < 0.05) in h HGF-MSCs treatment group.(9) Compared with the Control group, level of TNF-? and IL-6 in lung tissue was significantly reduced in the EGFP-MSCs- and h HGF-MSCs- treatment groups( P < 0.01); level of IL-10 in lung tissue was increased in h HGF-MSCs treatment group at the three time-points( P < 0.01)and in EGFP-MSCs treatment group at the day 1( P < 0.05). Compared with the EGFP-MSCs treatment group, level of TNF-? in lung tissue was significantly reduced at the day 1( P < 0.01) and level of IL-6 in lung tissue was significantly reduced at the day 3( P < 0.01) and level of IL-10 in lung tissue was significantly increased at the day 3 and day 7( P < 0.05, P < 0.01) in h HGF-MSCs treatment group.(10) Compared with the Control group, levels of TNF-? and IL-6 in m RNA was significantly reduced at the three time-points( P < 0.01) and IL-10 in m RNA was significantly increased at the three time-points( P < 0.01) in the h HGF-MSCs treatment group; levels of TNF-? and IL-6 in m RNA was significantly reduced at the three time-points( P < 0.05, P < 0.01) and IL-10 in m RNA was significantly increased at the day 3 and day 7( P < 0.05) in the EGFP-MSCs treatment group. Compared with the EGFP-MSCs treatment group, level of TNF-? in m RNA was significantly reduced at the day 1( P < 0.05) and IL-10 in m RNA was significantly increased at the three time-points( P < 0.05, P < 0.01) in the h HGF-MSCs treatment group.(11) The AI of alveolar epithelial cells was significantly reduced in the h HGF-MSCs treatment group compared with the Control group at the day 3 and day 7( P < 0.05).(12) The survive rate was significantly increased in the EGFP-MSCs- and h HGF-MSCs- treatment groups compared with the Control group( P < 0.05).Conclusion: The treatments of EGFP-MSCs or h HGF-MSCs all could reduce inflammation and improve survive time,while the effect of h HGF-MSCs was more better. |
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