The Role And Mechanism Of Notch1 Regulating Treg Cells In The Pathogenesis Of COPD In The Elderly

Objective: We need to study the expression level of Notch1 gene,HES1 protein and the proportion of Treg cells in peripheral blood if the elderly AECOPD patients are in acute exacerbation period,and to study the influence of immune imbalance that Notch1 signaling pathway in COPD.Methods:We took 40 cases of AECOPD patients between August 2015 and March 2018 in Hunan People's Hospital.The adult AECOPD group(15 years old-64 years old)and the elderly AECOPD group(?65 years old)were 20 cases each.Another 40 healthy subjects were selected as control group,20 of which were adult control group and elderly control group.PBMC was separated by density centrifugation,the proportion of peripheral Treg cells were detected by flow cytometry,the relative expression of Notch1 in PBMC was detected by RT-PCR and the content of HES1 protein in peripheral blood was detected by Western Blot.We collected the peripheral blood from normal human body,and collected Treg cells by immunomagnetic beads,and then foster Treg cells after adding notch inhibitor.The proportion of peripheral Treg cells were detected by flow cytometry.Result:(1)The percentage of Treg cells which were detected by flow cytometry in peripheral blood mononuclear cells(PBMC),in adult AECOPD group [0.38(0.16-8.49)] was significantly higher than that in adult control group[0.06(0.00-0.12)](P<0.05);The percentage of Treg cells in the elderly AECOPD group[4.415(0.39-9.61)] was significantly higher than that in the elderly control group [0.12(0.07-1.32)](P<0.05);The percentage of Treg cells in the aged control group [0.12(0.07-1.32)] was significantly higher than that in the adult control group [0.06(0.00-0.12)](P<0.05);The percentage of Treg cells in the elderly AECOPD[4.415(0.39-9.61)] was significantly higher than that in the adult AECOPD group [ 0.38(0.16-8.49)](P<0.05).(2)The content of HES1 protein(29.49±8.96)in peripheral blood of adult AECOPD group was significantly higher than that of adult control group(9.96 ±6.34)when we used Western blotting method(P< 0.01);The content of HES1 protein in the peripheral blood of the elderly AECOPD group(37.49 ±11.86)was significantly higher than that of the elderly control group(20.47 ±7.28)(P<0.01);The content of HES1 protein in peripheral blood of aged control group(20.47 ±7.28)was significantly higher than that of adult control group(9.96 ±6.34)(P<0.01);The content of HES1 protein in the peripheral blood of the elderly AECOPD group(37.49 ±11.86)was significantly higher than that of the adult AECOPD group(29.49 ±8.96)(P<0.05).(3)The relative expression of Notch1 m RNA in PMBC was detected by RT-PCR,the relative expression of Notch1 in the peripheral blood mononuclear cells of the adult AECOPD group(1.73±0.48)was significantly higher than that in the adult control group(1.03± 0.13)(P<0.01);The relative expression of Notch1 in peripheral blood mononuclear cells in elderly AECOPD group(2.09±0.52)was significantly higher than that in elderly control group(1.15±0.27)(P< 0.01).The relative expression of Notch1 in peripheral blood mononuclear cells of the elderly control group(1.15±0.27)was significantly higher than that of the adult control group(1.03 ±0.13)(P<0.01);The relative expression of Notch1 in peripheral blood mononuclear cells of the elderly AECOPD group(2.09±0.52)was significantly higher than that of the adult AECOPD group(1.73±0.48)(P<0.05).(4)Correlation: There was a positive correlation between age and the content of Hes1 protein in peripheral blood serum(r=0.419,P<0.01).There was a positive correlation between age and the relative expression of Notch1(r=0.226,P<0.05).There was a negative correlation between the percentage of forced expiratory volume in the first second and the level of HES1 protein in peripheral blood(r=-0.710,P<0.01),and has a negative correlation with the relative expression of Notch1(r=-0.657,P<0.01).The ratio of Treg in peripheral blood and the content of HES1 protein in peripheral blood were positively correlated(r=0.635,P<0.01).The ratio of Treg in peripheral blood and the relative expression of notch1 in peripheral blood serum were positively correlated(r=0.686,P < 0.01).(5)Flow cytometry was used to detect the proportion of Treg cells after treatment with different concentrations,Compared with that before treatment,the Treg cells in the Notch1 inhibition group of 1u M concentration were not significantly decreased(P>0.05),and the proportion of Treg cells in other treatment groups with Notch inhibitors decreased significantly(P<0.05),There was no significant difference between 1u M and 2.5u M inhibitor Notch1(P>0.05).There was no significant difference between 2.5u M and 5u M inhibitor Notch1(P> 0.05).But there was a significant difference between the other groups(P < 0.05).Conclusion: Notch1 may affect the proportion of Treg cells through activating Notch signaling pathway,thereby playing a role in regulating AECOPD in the elderly.Inhibition of Notch signaling pathway may affect the immune balance of AECOPD.