Atherosclerotic Pre-Conditioning Affects the Paracrine Role of Circulating Angiogenic Cells Ex-Vivo.

Sara Eslava-Alcon,Jose Angel Alonso-Piñero,Nuria Ibarz,Ismael Sanchez-Gomar,Almudena González-Rovira,Rafael Moreno-Luna,Mª Carmen Durán-Ruiz,Lucía Beltrán-Camacho,Manuel Rodriguez-Piñero,Antonio Rosal-Vela,Mª Jesús Extremera-García,Margarita Jiménez-Palomares,Esther Doiz,Javier Muñoz,Rosario Conejero,Marta Rojas-Torres

doi:10.3390/ijms21155256

Abstract

In atherosclerosis, circulating angiogenic cells (CAC), also known as early endothelial progenitor cells (eEPC), are thought to participate mainly in a paracrine fashion by promoting the recruitment of other cell populations such as late EPC, or endothelial colony-forming cells (ECFC), to the injured areas. There, ECFC replace the damaged endothelium, promoting neovascularization. However, despite their regenerative role, the number and function of EPC are severely affected under pathological conditions, being essential to further understand how these cells react to such environments in order to implement their use in regenerative cell therapies. Herein, we evaluated the effect of direct incubation ex vivo of healthy CAC with the secretome of atherosclerotic arteries. By using a quantitative proteomics approach, 194 altered proteins were identified in the secretome of pre-conditioned CAC, many of them related to inhibition of angiogenesis (e.g., endostatin, thrombospondin-1, fibulins) and cell migration. Functional assays corroborated that healthy CAC released factors enhanced ECFC angiogenesis, but, after atherosclerotic pre-conditioning, the secretome of pre-stimulated CAC negatively affected ECFC migration, as well as their ability to form tubules on a basement membrane matrix assay. Overall, we have shown here, for the first time, the effect of atherosclerotic factors over the paracrine role of CAC ex vivo. The increased release of angiogenic inhibitors by CAC in response to atherosclerotic factors induced an angiogenic switch, by blocking ECFC ability to form tubules in response to pre-conditioned CAC. Thus, we confirmed here that the angiogenic role of CAC is highly affected by the atherosclerotic environment.

Highlights

The potential use of endothelial progenitor cells (EPC) for cell therapy in cardiovascular diseases (CVD) has been largely evaluated due to their assigned role in vascular repairing and tissue revascularization [1,2,3]
Mixed transplantation seems to promote the revascularization effect [13], since these cells participate in a coordinated manner [10]: circulating angiogenic cells (CAC) are recruited to the damaged tissue and secrete paracrine factors that promote the migration of endothelial colony-forming cells (ECFC) to the injured site to restore the integrity of the vascular wall
Our results indicated that after 24 h, angiogenesis was induced in all cases, but significant differences were seen in the angiogenic potential of ECFC depending on Ingenuity Pathway Analysis (IPA) software. the table includes the main categories containing related functions or diseases, predicted activation (↑) or inhibition (↓) according to IPA, numbeIrnInto.t.Jf.J.MMmool.lo.SSclcie.i.2c20u0220l0,e,221s1,xpxFFeOOrRRPcPEaEEtEReRRgREEoVVrIEIyEWW, p-values assigned, and protein names

Summary

Introduction

The potential use of endothelial progenitor cells (EPC) for cell therapy in cardiovascular diseases (CVD) has been largely evaluated due to their assigned role in vascular repairing and tissue revascularization [1,2,3]. Two main sub-populations have been formally defined from the initially named EPC [8,9,10]: early EPC (eEPC), called circulating angiogenic cells (CAC) or myeloid angiogenic cells (MAC), with limited proliferation capacity (lasting 7–10 days after isolation) and angiogenic potential through paracrine mechanisms [11,12]; and late outgrowth cells or endothelial colony-forming cells (ECFC), highly proliferative and with enormous vasculogenic properties, probably by direct engraftment and/or replacement of damaged cells [8,9] Both sets of cells have been used alone or combined in diverse pre-clinical or clinical studies, with certain success in the restoration, among others, of ischemic tissues such as in critical limb ischemia [1,2,13,14,15,16]. The number of circulating EPC is a strong predictor of atherosclerotic plaque recurrence in the common carotid artery, with decreased number of EPC correlating with the presence and progression of preclinical atherosclerosis [18,22,23]

Methods

Results

Discussion

Conclusion