Oleanolic acid inhibits oxidative damage of HUVECs via PPARγ

doi:10.3877/cma.j.issn.2095-1221.2017.06.003

Abstract

Abstract:

Objective

To investigate the effects of oleanolic acid (OA) on oxidative damage in human vascular endothelial cells (HUVECs) and its underlying mechanism.

Methods

The groups are as followings: the control group, ox-LDL alone group, ox-LDL+OA (10μmol/L,20μmol/L, 40μmol/L) group, ox-LDL+OA (10μmol/L, 20μmol/L, 40μmol/L) +GW9662 group, and GW9662 alone group. The cell viability was measured using MTT assay. The levels of SOD, GSH as well as MDA were detected by enzyme linked immunosorbent assay (Elisa) . The level of ROS was detected using reactive oxygen species detection kit. The expression level of PPARγwas measured by Western blot. All indexed are performed biology-repeatedly. The statistical analysis was used with Analysis of Variance and Kruskal-WallisHtest.

Results

OA (10μmol/L, 20μmol/L,40μmol/L) significantly attenuated the cell viability of HUVECs inhibited by ox-LDL (100μg/ml) in a dose-dependent manner and each cell viability is (68.51±1.16) ﹪, (82.64±0.73) ﹪, (92.37?±?0.13) ﹪ (t = 24.35, 26.18, 35.17; P = 0.034, 0.027, 0.008) . OA reversed oxidative damage was induced by ox-LDL including decreases in the levels of SOD and GSH, and increases in the level of MDA and ROS in HUVECs. The levels of SOD, GSH, ROS and MDA in ox-LDL group are (16.12±0.06) μmol/g, (132.16±2.11) μmol/g, (2.63±0.02) kU/g, (158.12?±0.39) ﹪; the levels of SOD, GSH, ROS and MDA in ox-LDL and OA (10μmol/L) group are (13.28?±0.09) μmol/g, (129.58±0.09) μmol/g, (2.26±0.15) kU/g, (126.43±1.51) ﹪respectively; the levels of SOD, GSH, ROS and MDA in ox-LDL and OA (20μmol/L) group is (13.28±0.09) μmol/g, (129.58?±?0.09) μmol/g, (2.26±0.15) kU/g, (126.43±1.51) ﹪respectively; the levels of SOD, GSH, ROS and MDA in ox-LDL and OA (40μmol/L) group are (14.86±0.16) μmol/g, (131.47±0.76) μmol/g, (2.14?±0.08) kU/g, (112.39±1.07) ﹪respectively (F = 26.38, 31.27, 56.82, 41.16; P = 0.005, 0.004, 0.002, 0.003) . Western blot result showed that OA attenuated downregulation of PPARγprotein induced by ox-LDL in HUVECs. Our result also showed that PPARγinhibitor GW9662 reversed the attenuation of OA on ox-LDL-caused inhibition of cell viability: the cell viability in ox-LDL+OA (20μmol/L) +GW9662 group (t = 16.47,P = 0.035) . In addition, we found that PPARγinhibitor GW9662 reversed the improvement of OA on ox-LDL-caused oxidative damage: the levels of SOD, GSH, MDA and ROS in ox-LDL+OA (10μmol/L) +GW9662 group are (8.42±0.05) μmol/g, (88.38±0.48) μmol/g, (2.83±0.01) kU/g, (154.41±1.04) ﹪ (t = 38.47, 39.25, 43.69, 41.27; P = 0.008, 0.008, 0.006, 0.006) respectively; the levels of SOD, GSH, MDA and ROS in ox-LDL+OA (20μmol/L) +GW9662 group are (10.59±0.12) μmol/g, (106.42±0.1) μmol/g, (2.61?±?0.07) kU/g, (138.12±1.15) ﹪ (t?= 46.08, 38.11, 49.35, 35.59,P =0.005, 0.008, 0.004, 0.009) respectively; the levels of SOD, GSH, MDA and ROS in ox-LDL+OA (40μmol/L) +GW9662 group was (13.65.±0.03) μmol/g, (124.61±1.27) ?μmol/g, (2.49±0.04) ?kU/g, (126.51±0.73) ﹪ (t = 48.04, 38.62, 45.14, 50.13;P = 0.004, 0.008, 0.005, 0.002) respectively. Furthermore, we also found that after inhibiting PPAR gamma, OA still dose dependently alleviated ox-LDL-caused oxidative damage of HUVECs cells.

Conclusion

OA inhibits oxidative damage induced by ox-LDL and PPARγplays a regulatory role in the protection of OA against ox-LDL-induced oxidative damage of HUVECs.

Key words: Oleanolic acid, Ox-LDL, Oxidative damage, PPARγ, Human umbilical vein endothelial cells

Baohua Zhu, Chunjing Gu, Fenglian Liu, Lu Liu, Yanhong Ren. Oleanolic acid inhibits oxidative damage of HUVECs via PPARγ[J]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2017, 07(06): 327-333.

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References 20

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分组	样本数	SOD（μmol/g）	GSH（μmol/g）	MDA（kU/g）	ROS相对荧光强度（﹪）
对照组	3	16.12±0.06	132.16±2.11	2.03±0.04	100.04±0.45
ox-LDL	3	5.43±0.08^a	49.72±1.28^a	2.63±0.02^a	158.12±0.39^a
ox-LDL+OA（10 μmol/L）	3	10.60±0.14^b	108.36±2.05^b	2.41±0.21^b	136.18±1.24^b
ox-LDL+OA（20 μmol/L）	3	13.28±0.09^b	129.58±0.09^b	2.26±0.15^b	126.43±1.51^b
ox-LDL+OA（40 μmol/L）	3	14.86±0.16^b	131.47±0.76^b	2.14±0.08^b	112.39±1.07^b
F值	?	26.38	31.27	56.82	41.16
P值	?	0.005	0.004	0.002	0.003

分组	样本数	SOD（μmol/g）	GSH（μmol/g）	MDA（kU/g）	ROS相对荧光强度（﹪）
对照组	3	16.12±0.06	132.16±2.11	2.03±0.04	100.04±0.45
ox-LDL	3	5.43±0.08^a	49.72±1.28^a	2.63±0.02^a	158.12±0.39^a
ox-LDL+OA（10 μmol/L）	3	10.60±0.14^b	108.36±2.05^b	2.41±0.21^b	136.18±1.24^b
ox-LDL+OA（20 μmol/L）	3	13.28±0.09^b	129.58±0.09^b	2.26±0.15^b	126.43±1.51^b
ox-LDL+OA（40 μmol/L）	3	14.86±0.16^b	131.47±0.76^b	2.14±0.08^b	112.39±1.07^b
F值	?	26.38	31.27	56.82	41.16
P值	?	0.005	0.004	0.002	0.003

分组	样本数	SOD（μmol/g）	GSH（μmol/g）	MDA（kU/g）	ROS相对荧光强度（﹪）
对照组	3	17.05±0.12	135.28±2.14	2.16±0.15	100.53±1.08
ox-LDL	3	6.41±0.04^a	50.38±0.13^a	2.71±0.12^a	160.05±0.46^a
ox-LDL+OA（10 μmol/L）	3	10.58±0.13^b	102.46±0.06^b	2.42±0.08^b	144.38±2.02^b
ox-LDL+OA（20 μmol/L）	3	13.25±0.05^b	122.59±0.33^b	2.23±0.16^b	123.94±0.15^b
ox-LDL+OA（40 μmol/L）	3	15.88±0.14^b	140.26±1.05^b	2.02±0.13^b	187.52±0.68^b
oxLDL+OA(10 μmol/L)+GW9662	3	8.42.±0.05^c	88.38±0.48^c	2.83±0.01^c	154.41±1.04^c
oxLDL+OA(20 μmol/L)+GW9662	3	10.59.±0.12^c	106.42±0.15^c	2.61±0.07^c	138.12±1.15^c
oxLDL+OA(40 μmol/L)+GW9662	3	13.65.±0.03^c	124.61±1.27^c	2.49±0.04^c	126.51±0.73^c
GW9662	3	16.02±0.13	130.14±2.04	1.98±0.29	103.15±0.29
F值	?	35.08	42.31	26.84	33.73
P值	?	0.003	0.001	0.004	0.003

分组	样本数	SOD（μmol/g）	GSH（μmol/g）	MDA（kU/g）	ROS相对荧光强度（﹪）
对照组	3	17.05±0.12	135.28±2.14	2.16±0.15	100.53±1.08
ox-LDL	3	6.41±0.04^a	50.38±0.13^a	2.71±0.12^a	160.05±0.46^a
ox-LDL+OA（10 μmol/L）	3	10.58±0.13^b	102.46±0.06^b	2.42±0.08^b	144.38±2.02^b
ox-LDL+OA（20 μmol/L）	3	13.25±0.05^b	122.59±0.33^b	2.23±0.16^b	123.94±0.15^b
ox-LDL+OA（40 μmol/L）	3	15.88±0.14^b	140.26±1.05^b	2.02±0.13^b	187.52±0.68^b
oxLDL+OA(10 μmol/L)+GW9662	3	8.42.±0.05^c	88.38±0.48^c	2.83±0.01^c	154.41±1.04^c
oxLDL+OA(20 μmol/L)+GW9662	3	10.59.±0.12^c	106.42±0.15^c	2.61±0.07^c	138.12±1.15^c
oxLDL+OA(40 μmol/L)+GW9662	3	13.65.±0.03^c	124.61±1.27^c	2.49±0.04^c	126.51±0.73^c
GW9662	3	16.02±0.13	130.14±2.04	1.98±0.29	103.15±0.29
F值	?	35.08	42.31	26.84	33.73
P值	?	0.003	0.001	0.004	0.003

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