Effects of recombinate mouse granulocyte-macrophage-colony-stimulating factor along with marrow-derived mesenchymal stem cells on lung injury induced by hyperoxia exposure in newborn rats / 中华急诊医学杂志

ABSTRACT

Objective To investigate the effects of recombinate mouse granulocyte-macrophage-colony-stimulating factor(GM-CSF) alone with mesenchymal stem cells(MSCs) on injured lung of rots after exposure to hyperoxia. Method Mouse MSCs were separated, cultured, amplificated, identified and labeled with 4', 6-diamidino-2-phenylindole(DAPI). Thirty-two 3-day-old Sprague Dawley(SD) rats from four litters were randomly divided(random number) into four groups, namely hypemxia exposured + GM-CSF + MSCs group(group A), hyperoxia exposured + GM-CSF group( group B), hyperoxia exposured + MSCs group( group C) and hypemxia exposured group(group D). All rats were placed in a closed Plexiglas chamber with a minimal flow in and out, providing six to seven exchanges of the chamber volume per hour and maintaining O_2 levels above 95%. Seven days lair,all of them were taken out of the chamber. Rats in group A were treated with 5 x 10~4 MSCs intraperitoneally alone with 9 μg/kg GM-CSF subcutaneously, rats in group B received 9 μg/kg GM-CSF subcutaneously, rats of group C were treated with 5 x 10~4 MSCs intraperitonealiy and rats of group D were administrated with phosphatebuffered solution(PBS). Three days latter, all animals were sacrificed by an injection of 120 mg/kg sodium pentobarbital, perfused with cold 0.9% NaCl, and the left lungs were removed. The upper lobe of them were grinded to make tissue homogenates used for ELASA, and the lower lobes of them were made into frozen sections for fluorescence microscope. The right lungs were fixed in situ for 2 hours by intratracheal instillation of 10% neutral formalin and then were still fixed for additional 24 hours. Sagittal sections of paraffin-embedded middle lobe and upper lobe of left hmg tissue were used for Immunohistochemistry and histological study respectively, Immunohistochemistry was used to detect the expression of telomerase reverse transcrip tase(TERT) grade. Results Among 4groups, there were significantly differences in radical alveolar counts(RAC), TNF-α and IL-β1 in tissue homogenates( P < 0. 01 ). Compared with group D, increase in RAC and the decrease in both TNF-α and IL-β1 were found in other groups, and furthermore there were obvious differences in those changes between group A and group B as well as between group A and group C. There were significant differences in TERT(P<0.01) among four groups. The TERT grade in group A and group B were increased more markedly. DAPI-positive cells were found in group A and group C with significantly differences(6.23 ± 1.88, 5.10 ± 0.91, t = 1.53, P<0.05).Conclusions The protective effects of GM-CSF/MSCs on injuried lungs of new born rats after exposure to hyperoxia may be associated with the increase in the proliferation of stem cells improving the local micro-environment of lung tissue. This protective effects against lung injury induced by hyperoxia exposure may be attributed to the synergism between GM-CSF and MSCs.