ABSTRACT
Aim To explore the effect of Buyang Huanwu Decoction on cerebral ischemia-reperfusion injury in rats by regulating autophagy through PI3K/AKT pathway. Methods The rats were randomly divided into five groups(n=10): sham operation group(Sham), model group(Model), Buyang Huanwu Decoction group(BYHWD), PI3K inhibitor group(LY294002)and Vehicle group(Vehicle). Except Sham group, the other groups were treated with 2h ischemia and 72 h reperfusion for modeling. The Zea Longa score was used to assess the neurological defects, HE was used to observe brain injury in the ischemic penumbra(IP), immunofluorescence was employed to detect LC3, and Western blot was used to detect pathway and autophagy marker proteins. Results Compared BYHWD group with model group, the neurological score of rats decreased, cerebral infarction volume decreased, the pathological lesions of brain IP were relieved, PI3K and p-AKT/AKT expression increased, and LC3Ⅱ/ decreased and p62 increased(P<0.05). The regulatory effect of BYHWD was weakened by LY294002(P<0.05). Conclusion Buyang Huanwu Decoction alleviates cerebral ischemia-reperfusion injury in rats by activating PI3K/AKT pathway to inhibit autophagy.
ABSTRACT
BACKGROUND: Superparamagnetic iron oxide (SPIO) labeling technology is a classic noninvasive tracing method, which has been widely used in the stem cell transplantation. Induced pluripotent stem cells (iPSCs) are currently one of the most promising seed cells for cell transplantation. Whether SPIO labeling can also be used to noninvasively trace induced pluripotent stem cells is rarely reported, and concern has been raised about whether SPIO markedly impacts the differentiation of iPSCs. OBJECTIVE:To investigate the effects of SPIO labeling on the differentiation of iPSCs in vitro. METHODS: Rat fibroblasts were isolated and cultured. Efficient recombinant vector and plasmids that were packaged by virus and contained target genes (Oct4, Sox2, Klf4 and c-Myc) were transfected into 293T cells for virus packaging and production. The packaging lentiviral vectors that contained target genes infected rat fibroblasts to obtain iPSCs. SPIO-labeled (experimental) or unlabeled (control) iPSCs were subjected to neural induction and differentiation. Prussian blue staining and transmission electron microscope observation were performed for SPIO-labeled iPSCs. Immunohistochemical method was used to detect neuron-specific enolase expression after induced differentiation. Flow cytometry was used to detect the proportion of neurons and glial cells differentiated from iPSCs. RESULTS AND CONCLUSION: There were dense iron particles in the cytoplasm of SPIO-labeled iPSCs shown by Prussian staining and under transmission electron microscope. Differentiated iPSCs were positive for neuron-specific enolase. In addition, the proportion of neurons and glial cells showed no difference between the experimental and control groups. To conclude, SPIO labeling has no obvious effect on the capacity of iPSCs differentiating into neurons. Reasonable application of this new cell labeling technique will promote the development of seed cells in regenerative medicine.