Introduction: In 80’s Professor Barker proposed that the perinatal period plays a major role on the development of health and disease at adulthood. Individuals born after an intrauterine growth restriction (IUGR) are more prone to develop cardiovascular diseases, especially arterial hypertension at adulthood. Several mechanisms are involved in the development of hypertension. Endothelial dysfunction is one of them. Endothelial progenitors cells (EPCs) are the circulating components of the endothelium, contributing to endothelial repair. EPCs can be classified as early EPCs and later EPCs, also called endothelial colony-forming cells (ECFCs). ECFCs are the only EPCs to have the capacity of endothelial repair. They can proliferate, have auto-renewal capacities, and can improve vascular proliferation and neovascularization. Oxidative stress (OS), and senescence are also playing major roles in endothelial dysfunction and arterial hypertension development by reducing angiogenesis and vascular repair.
The aim of this study was to explored whether individuals born after IUGR displayed a reduced ECFCs proportion and functionality and to identify mechanisms contributing to these dysfunction like OS and senescence.
Methods: We used an IUGR rat model induced by a maternal low protein diet during pregnancy, with 9% casein in the IUGR group compared to 23% casein in control group. The arterial pressure has been measured at the age of 6 months. Males ECFCs have been isolated from bone marrow using a ficoll gradient. We evaluated the ECFCs proportion by flow cytometry using CD45-/CD3+/CD 146+ and with the expression of CD34 by immunofluorescence. We measured the production of NO with fluorescent DAF-2DA, and the expression of eNOS by immunofluorescence and western blot. The expression of angiogenic and antiangiogenic proteins like angiopoietin, angiomotin, vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor 2 (VEGFR2) and thrombospondin have been evaluated by immunofluorescence. In addition, we have explored the presence of oxidative stress by measurement of superoxide anions and antioxidant defenses (Cu/Zn superoxide dismutase, catalase). We explored the cellular senescence by measurement of beta- galactosidase activity and the related factors associated to senescence proteins like p21WAFand p16INK4a expression with western blots.
Results: We found a decreased proportion of ECFCs in the IUGR group. They had less capacities to organize in tubular structure and to form new vessels. We also observed a reduction of NO production, associated with decreased eNOS expression. We found a decreased expression of angiogenic proteins (VEGF-A, VEGFR-2 angiopoietin, angiomotin, and an increased production of antiangiopoietic proteins (thrombospondin). Superoxide anion production is increased with a reduction of antioxidant defenses and some senescence markers (beta-galactosidase, p16INK4a ) are also increased.
Conclusion: We showed that IUGR is contributing to development of arterial hypertension associated with a decreased proportion and impaired function of ECFCs, altered angiogenic profile and increased OS and senescence.