The Migration Of Human Umbilical Vein Endothelial Cells Is Collectively Regulated By S1P/S1P1 And SR-BI Through PI3K-Akt Signaling Pathway

【 objective 】 Sphingosine-1-phosphate(S1P) is a signal sphingomyelin, most of them combine with HDL in the plasma, and they mediate HDL’s many functions through combining with its membrane receptors(S1PR1- S1PR5). Scavenger receptor class B type I(SR-B1) is the natural high affinity physiological receptor of the HDL, and it also can mediate HDL’s many functions through combining with apo A-I. It was reported that S1P/S1PR1 can promote the migration of endothelial cells through PI3K-Akt signaling pathway. Besides, some reports said that SR-BI can mediate HDL’s many biological functions through PI3K-Akt signaling pathway. But it has not been reported that HDL/SR-BI can mediate the migration of Human Umbilical Vein Endothelial Cell(HUVEC) through PI3K-Akt signaling pathway and HDL/SR-BI and HDL-S1P/S1P1 collectively regulate the migration of HUVECs through PI3K-Akt signaling pathway. Besides, it’s unclear whether SR-BI can affect the regulation of HDL/SR-BI to the migration of human umbilical vein endothelial cells. Therefore, we mainly study that HDL/SR-BI and HDL-S1P/S1P1 collectively regulate the migration of HUVECs through PI3K-Akt signaling pathway and SR-BI affects the HDL-S1 P /S1P1’s regulation function of HUVECs migration. So that it will provide the new theoretical basis for more all-sided cognition of HDL’s protection function of the endothelium.【Methods】1. Transfected with recombined expression plasmid, pc DNA3.1(-)-h SR-B1, for 48 hours or incubated with BLT-1 later, then HUVECs were stimulated with S1 P, HDL and apo A-I for 24 hours and 8hours, respectively. The migration of HUVECs was tested respectively by wound healing assay and Transwell. The protein expression levels of phosphorylated PI3 K and Akt were tested by Western Blot. 2. Incubated with S1P1 agonist SEW2871 or antagonist W146 later, then HUVECs were stimulated with S1 P, HDL and apo A-I for 24 hours and 8hours, respectively. The migration of HUVECs was tested respectively by wound healing assay and Transwell. The protein expression levels of phosphorylated PI3 K and Akt were tested by Western Blot. 3. Incubated with PI3 K inhibitor LY294002 or Akt inhibitor GSK690693 later, then HUVECs were stimulated with S1 P, HDL and apo A-I for 24 hours and 8hours, respectively. The migration of HUVECs was tested respectively by wound healing assay and Transwell. 4. Transfected with recombined expression plasmid, pc DNA3.1(-)-h SR-B1, for 48 hours and incubated with BLT-1 later, then HUVECs were stimulated with HDL for 0hours, 2hours and 6hours, respectively. The protein expression level of S1P1 was tested by Western Blot.【Results】 S1 P, HDL and apo A-I can promote the migration of HUVECs. The migration of HUVECs treated with HDL is the most obvious. SR-B1 overexpression, compared with HDL group in the control, the migration of HUVECs treated with HDL increased by about 20%. SR-B1 inhibition, compared with HDL group in the control, the migration of HUVECs treated with HDL reduced by about 25%. S1P1 activation, compared with HDL group in the control, the migration of HUVECs treated with HDL increased by about 10%. S1P1 inhibition, compared with HDL group in the control, the migration of HUVECs treated with HDL reduced by about 10%. S1 P, HDL and apo A-I can increase the protein expression levels of phosphorylated PI3 K and Akt. It is the most obvious that the increase of the protein expression levels of phosphorylated PI3 K and Akt in HDL group. For the protein expression levels of phosphorylated PI3 K and Akt, it is more obvious that the magnitude of the change in HDL group which was treated with overexpressed SR-B1. However, it is not obvious that the magnitude of the change in HDL group which was treated with S1P1 agonist SEW2871 or antagonist W146. Incubated with PI3 K inhibitor LY294002 or Akt inhibitor GSK690693 later, compared with the control, the migration of HUVECs obviously reduced in HDL, S1 P and apo A-I group. S1P1 degradation was mediated by HDL.Transfected with recombined expression plasmid, pc DNA3.1(-)-h SR-B1, for 48 hours later, S1P1 degradation had slow down mediated by HDL.【Conclusion】1. As same as S1 P and HDL, apo A-I belongs to HDL also can promote the migration of HUVECs. 2. SR-BI can affect the regulative function of HDL-S1 P /S1P1 to HUVECs. 3. apo A-I /SR-BI and S1P/S1P1 collectively regulate the migration of HUVECs through PI3K-Akt signaling pathway.