Objective To investigate effects of neuromuscular electrical stimulation (NMES) on pulmonary arterial hypertension induced by chronic hypoxic hypercapnia in rats.Methods Eighteen male Sprague-Dawley rats were randomly divided into a normal control group (the control group),a hypoxic hypercapnia group (the model group),and a hypoxic hypercapnia + NMES group (the NMES group),each of 6.The rats in both the model and NMES groups were placed in an isobaric cabin with an O2 concentration of 9％ to 11％ and a CO2 concentration of 5％ to 6％ for 8 hours a day for 4 weeks.After leaving the cabin,NMES was performed on the NMES group's bilateral calf muscles for 30 minutes every day.The heart was removed,and the right ventricle (RV) and the left ventricle plus the septum (LV+S) were dissected.An index of right ventricular hypertrophy was calculated as RVHI=RV/(LV+S).Any changes in the pulmonary vasculature were observed using an optical microscope.WT％ and WA％ were calculated.The expression of hypoxia-inducible factor-1α (HIF-1α),PDH-E1α and PDK1 in the lung tissue were determined using western blotting.The LDH activity and the concentration of PDH in the lung tissue homogenate were measured was measured by spectrophotometric method using the LDH assay kit and ELISA,respectively.Results Compared with the control group,the average RVHI,WT％ and WA％,the protein expression of HIF-lα and PDK1,and LDH activity had all increased significantly in the NMES group,while the average expression of PDH-1Eα had decreased significantly.Compared with the model group,significant decrease was observed in the average RVHI,WT％,WA％,protein expression of HIF-1α and PDK1,and LDH activity in the NMES group,but the average expression of PDH-1Eα increased significantly.No significant differences in PDH concentration were detected among the 3 groups.Conclusions NMES may alleviate pulmonary artery hypertension induced by chronic hypoxic hypercapnia,at least in rats.The mechanism may be attributed to inhibiting the expression of HIF-1α protein,which may inhibit the activity of PDH-E1α and LDH,then the aerobic metabolism into glycolysis,finally improving the remodeling of the pulmonary vascular structure.

Objective:1. To detect any change in the PTEN/Akt/FoxO1 signaling pathway in the muscles of rats with chronic hypoxia-hypercapnia treated using neuromuscular electrical stimulation (NMES), and 2. To document the role of chronic hypoxia-hypercapnia in inducing muscle atrophy.Methods:Thirty-two male Sprague-Dawley rats were randomly divided into a control group, a model group, a mock stimulation group, and an NMES group, each of eight. All of the rats in the model group, the mock stimulation group and the NMES group were placed in a hypoxia-hypercapnia chamber with a 9-11% O 2 and 5.5-6.5% CO 2 atmosphere for 8h per day and 7d per week, lasting 4 weeks. The control group were placed in a similar chamber with normal air. In the last 2 weeks, after the 8h in the chamber, the NMES group were given 30min of electrical stimulation at 100Hz to the calf muscles of their bound lower limbs. The mock stimulation group were only bound without any electrical stimulation. After the 4-week intervention, the gastrocnemius muscles were resected and their cross-sectional areas (CSAs) were observed using hematoxylin-eosin staining. Immunohistochemistry and western blotting were employed to detect the protein expression of phosphatase and tensin (PTEN), p-Akt, Akt and FoxO1. Results:Compared with the control group, a significant decrease was observed in the average CSA and in the expression of p-Akt and Akt in the model group, while a significant increase was found in the average protein expression of PTEN and FoxO1. Compared with the model group, there was a significant increase in the average CSA, as well as the average expression of p-Akt and Akt in the NMES group, but a significant decrease in the average expression of PTEN and FoxO1.Conclusion:Neuromuscular electrical stimulation can relieve muscle atrophy from chronic hypoxia-hypercapnia by inducing skeletal muscle protein synthesis through regulating the PTEN/Akt/FoxO1 signaling pathway, at least in rats.

Objective:To discuss the effect of N-myc downstream-regulated gene 1(NDRG1)on the enzalutamide(ENZA)resistance in the castration-resistant prostate cancer(CRPC),and to clarify its mechanism.Methods:The human CRPC C4-2 cells and ENZA-resistant strain C4-2/ENZA cells were cultured in vitro.The expression levels of NDRG1 mRNA in the C4-2/ENZA cells and their parental C4-2 cells were detected by real-time fluorescence quantitative PCR(RT-qPCR)method.The expression levels of NDRG1,androgen receptor(AR),and prostate-specific antigen(PSA)proteins in the cells were detected by Western blotting method to verify the transfection efficiency of the cells.The C4-2/ENZA cells were divided into blank group(normally cultured without treatment),negative control lentivirus(Lv-NC)group(transfected with Lv-NC),Lv-NDRG1 group(transfected with Lv-NDRG1),Lv-NC+ENZA group(transfected with Lv-NC followed by ENZA treatment),Lv-NDRG1+ENZA group(transfected with Lv-NDRG1 followed by ENZA treatment),Lv-NDRG1+epidermal growth factor(EGF)group(transfected with Lv-NDRG1 followed by EGF treatment),and Lv-NDRG1+EGF+ENZA group(transfected with Lv-NDRG1 followed by EGF and ENZA treatment).The half-maximal inhibitory concentration(IC50),resistance index(RI),and proliferation activity of the cells were detected by MTT assay;the apoptotic rates of the cells in various groups were detected by flow cytometry;RT-qPCR method was used to detect the expression levels of NDRG1 mRNA in the cells in various groups;Western blotting method was used to detect the expression levels of NDRG1,AR,phosphorylated AR at serine213(p-ARSer213),phosphorylated AR at serine81(p-ARSer81),and PSA proteins in the cells in various groups.Results:Compared with C4-2 cells,the expression levels of NDRG1 mRNA and protein in the C4-2/ENZA cells were significantly decreased(P<0.01)and the expression levels of AR and PSA proteins were increased(P<0.01),indicating low expression of NDRG1 in the ENZA-resistant C4-2/ENZA strain.Compared with Lv-NC group,the expression levels of NDRG1 mRNA and protein in the cells in Lv-NDRG1 group were significantly increased(P<0.01),indicating the successful construction of an NDRG1 gene over-expression strain of C4-2/ENZA resistant cells.The MTT assay results showed that compared with the C4-2 cells,the IC50 of the C4-2/ENZA cells was increased(P<0.01)and the RI was 17.78;compared with Lv-NC group,the IC50 of the C4-2/ENZA cells in Lv-NDRG1 group was decreased(P<0.01).After 24 h of EGF treatment,compared with Lv-NC group,the IC50 of the C4-2/ENZA cells in Lv-NC+EGF group was significantly increased(P<0.01);compared with Lv-NDRG1 group,the IC50 of the C4-2/ENZA cells in Lv-NDRG1+EGF group was increased(P<0.01).Compared with before ENZA treatment,after 24 h of ENZA treatment,the proliferation activities of C4-2 and C4-2/ENZA cells were gradually decreased(F=223.80,P<0.01;F=81.46,P<0.01).Compared with Lv-NC group,the proliferation activity in the C4-2/ENZA cells in Lv-NDRG1 group after 24 h of ENZA treatment was significantly decreased(P<0.01).After 24 h of EGF treatment,compared with Lv-NC group,the proliferation activity of the C4-2/ENZA cells in Lv-NC+EGF group was significantly increased(P<0.01),while the the proliferation activity of the C4-2/ENZA cells in Lv-NDRG1+EGF group was significantly decreased(P<0.01).The chosen concentration and treatment duration for further testing were 10 000 μmol·L-1 ENZA and the intervention time was 24 h.The flow cytometry results showed that after 24 h of ENZA treatment,compared with Lv-NC group,the apoptotic rate of the cells in Lv-NDRG1 group was significantly increased(P<0.01);compared with Lv-NC+ENZA group,the apoptotic rate of the cells in Lv-NDRG1+ENZA group was significantly increased(P<0.01).After 24 h of EGF treatment,compared with Lv-NDRG1 group,the apoptotic rate of the cells in Lv-NDRG1+EGF group was significantly decreased(P<0.01),while the apoptotic rate of the cells in Lv-NDRG1+ENZA group was significantly increased(P<0.01);compared with Lv-NDRG1+ENZA group,the apoptotic rate of the cells in Lv-NDRG1+EGF+ENZA group was significantly decreased(P<0.01).The Western blotting results showed that after 24 h of ENZA treatment,compared with Lv-NC group,the expression levels of AR and PSA proteins and the ratio of p-ARSer213/AR and p-ARSer81/AR in the cells in Lv-NDRG1 group were significantly decreased(P<0.05 or P<0.01).After 24 h of EGF treatment,compared with Lv-NC group,the expression levels of AR and PSA proteins and the ratio of p-ARSer213/AR and p-ARSer81/AR in the cells in Lv-NC+EGF group were significantly increased(P<0.05 or P<0.01);compared with Lv-NDRG1 group,the expression levels of AR and PSA proteins and the ratio of p-ARSer213/AR and p-ARSer81/AR in the cells in Lv-NDRG1+EGF group were significantly increased(P<0.01).Conclusion:Over-expression of NDRG1 can reduce the resistance of CRPC to ENZA,and its mechanism may be related to the inhibition of AR signaling.