1.Regulation of Transforming Growth Factor -beta 1/Smad3 Signaling Pathway of Asthmatic Rat Airway Remodeling Model by Back Three-acupoint Electroacupuncture
Zheng LI ; Pengdong JI ; Yayun WANG ; Xiaojie LIN ; Zhenyi CHEN ; Xiaozhe LI ; Xinsheng LAI
Journal of Guangzhou University of Traditional Chinese Medicine 2015;(1):71-75
Objective To observe the effect of electroacupuncture on back three-acupoints ( namely bilateral Dazhu, Fengmen, Feishu) on the protein and mRNA expression of transforming growth factor-beta 1 (TGF-β1) and Smad3 in asthmatic rat airway remodeling model, and to explore its therapeutic efficacy and molecular mechanism. Methods Rat asthma model was established by inhalation of ovalbumin. After sensitization for 6 weeks, rats were killed. And then the airway morphological parameters of rats were measured by image analysis. The protein and mRNA expression of TGF-β1 and Smad3 in lung tissues were detected by immunohistochemistry and real-time quantitative polymerase chain reaction (qRT-PCR) respectively. Results Compared with the blank group, the standardized values of muscle cross-sectional area including airway smooth muscle area (WAm) /perimeter of the basement membrane (Pbm), and bronchial inner wall area (WAi)/Pbm were increased in the model group. The protein and mRNA expression levels of TGF-β1 and Smad3 were also increased in the model group. In electroacupuncture group, the above observation indexes were decreased, and the differences were statistically significant ( P<0.05 compared with the model group). Conclusion The experimental results indicated that back three-acupoint electroacupuncture has an effect on remodeling airway, and one of the mechanisms is probably associated with the regulation of TGF-β1/Smad3 signaling pathway.
2.Role of GPx4 in somatic cells reprogramming into stem cells
Yang YANG ; Xiahua LIN ; Junyi YANG ; Yin ZHENG ; Pengdong LI ; Zizhang OUYANG ; Guojun ZHAO ; Baojian LIAO
Chongqing Medicine 2024;53(19):2898-2906
Objective To investigate the effect of the glutathione peroxidase 4(GPx4)on mouse so-matic cell reprogramming.Methods To compare the expressions of GPx4 in OG2 mouse embryonic fibroblast(OG2-MEF)cells(MEFs group)and mouse embryonic stem cells(mESC,mESCs group),the expression lev-el of intracellular GPx4 was determined by transcriptome sequencing technique and Western blot.To verify the effect of GPx4 on the efficiency of the somatic cells reprogramming,the complete open reading frame se-quence of GPx4 gene and its selenocysteine insertion sequence(SECIS)were connected to the retroviral vector pMXs for constructing the overexpressed plasmid pMXs-GPx4.Gpx4-targeting short hairpin RNA(shRNA)was synthesized and connected to pSUPER vector,GPx4 shRNA1 and GPx4 shRNA2 were constructed to knockdown GPx4 expression.The above plasmids were co-transfected with pMXs-Sox2,pMXs-Klf4 and pMXs-Oct4 into MEF cells for reprogramming induction to obtain the pMXs no-load control group(pMXs NC),pMXs GPx4 group,pSUPER no-load control group(pSUPER NC),GPx4 shRNA1 group and GPx4 shRNA2 group.The expressions of GPx4 gene and multifunctional marker genes Rex1,Sox2,Dappa3,Sall4,Oct4 and Nanog were detected by real-time fluorescence quantitative PCR.The induced pluripotent stem cells(iPSC)were detected by immunofluorescence staining;the number of iPSC clones generation was detected by alkaline phosphatase staining of pluripotent stem cells;the GPx4 protein expression was detected by Western blot.Results The mRNA and protein expression of GPx4 in the mESCs group was higher than that in the MEFs group;compared with the pMXs NC group,the expression level of GPx4 mRNA in the pMXs GPx4 group was significantly increased;compared with the pSUPER NC group,the GPx4 mRNA and protein levels in the GPx4 shRNA1 group and GPx4 shRNA2 group were decreased(P<0.05);the iPSC clone number in the pMXs GPx4 group was higher than that in the pMXs NC group,but the difference was not statistically significant(P>0.05).The number of iPSC clones in the GPx4 shRNA1 group and GPx4 shRNA2 group was significantly lower than that in the pSUPER NC group,and the difference was statistically significant(P<0.05).After completing the reprogramming,compared with the original MEF cells,the expression levels of various pluripotent marker genes Rex1,Sox2,Dappa3,Sall4,Oct4 and Nanog in the generated iPSC of each group were increased.Conclusion GPx4 knockdown could inhibit the efficiency of somatic cell reprogram-ming,its generated induced pluripotent stem cells have the normal pluripotent gene expression ability.