Extracellular Cyclophilin A Inhibitor MM284 Supports Proliferation and Migration of Human Coronary Artery Endothelial Cells

Abstract

IntroductionBiomaterials face an inflammatory environment upon implantation which represents a potential obstacle to their successful application. Therefore, anti‐inflammatory coatings on vascular implants have been extensively studied to improve wound healing and re‐endothelialisation. Cyclophilin A (CyPA) is a mediator of inflammation, which affects cell migration and proliferation. Clinical trials with CyPA‐inhibitors revealed many side effects. However, MM284 is a novel CyPA‐inhibitor targeting only extracellular CyPA and thus potentially offering new means for therapeutic applications. The objective of this study was to characterize the effects of MM284 on human coronary artery endothelial cells (HCAEC) and artery smooth muscle cells (HCASMC).MethodsProliferation was determined in 16‐well E‐plates with MM284 concentrations ranging from 1 nM to 1 μM. The impedance value of each well was continuously monitored for 18 h and expressed as cell index (CI). Viability was determined with resazurin using the same MM284 concentrations. Migration was assessed by live cell imaging under flow (1.5 dyne/cm2) with 50 nM MM284 over 15 h. Images were taken at a frequency of 1 frame/15 min and analyzed by manual tracking and chemotaxis tool using ImageJ (NIH). The results are presented as mean±SD. The relative quantification of specific mRNAs was determined by quantitative real time PCR and analyzed with the 2−ΔΔCT method. ΔΔCT values were normalized to untreated control samples and a t‐test was performed.ResultsMM284 increased HCAEC (p=0.002) but not HCASMC (p=0.648) proliferation in a concentration‐dependent manner compared to untreated controls. Cell viability of HCAEC (p<0.0001), but not HCASMC (p=0.153), was significantly reduced in the presence of MM284. Therefore, further experiments were performed with 50 nM MM284. Shear stress in combination with MM284 treatment increased HCAEC motility regarding the velocity (1.5 dyne/cm²: 0.34±0.01 μm/min; p=0.023), accumulated distance (1.5 dyne/cm²: 301±6 μm; p=0.023) as well as Euclidean distance (1.5 dyne/cm²: 152±7 μm; p=0.048). However, MM284 treatment had no influence on HCASMC migration. Furthermore, MM284 treatment in combination with shear stress reduced gene expression of the inflammatory marker monocyte chemotactic protein‐1 (MCP‐1) in HCAEC (p=0.043) and HCASMC (p=0.022).ConclusionOur results are the first to show that targeting extracellular CyPA via the novel CyPA‐inhibitor MM284, is leading to an enhanced endothelial cell proliferation and migration, while smooth muscle cells are not affected. Therefore, local application of MM284 may positively influence the re‐endothelialisation of cardiovascular implants and suppress the uncontrolled growth of HCASMC as well as an inflammatory reaction. Nevertheless, further in vivo studies should be performed to confirm these results.Support or Funding InformationResearch Project RESPONSE, German Centre for Cardiovascular Research (DZHK)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.