| Despite decades of research, synthetic materials elicit undesirable responses (such as thrombosis, inflammation) in blood-contacting materials. However, the human body has created a material which, for the most part, works well for decades without recruiting such undesirable responses---this material is the blood vessel wall. Studying how endothelial cells (ECs) sense the properties of this matrix, convert those sensations into intracellular signals, and then further convert those signals into desirable behaviors can provide a roadmap for reverse-engineering the properties of such natural materials. Here human umbilical vein endothelial cells (HUVECs) were cultured on ten different surfaces---9 adsorbed extracellular matrix (ECM) proteins and tissue culture plastic (PS). Activities of four kinases (namely Extracellular regulated kinase (ERK), c-Jun N-Terminal Kinase (JNK), I-Kappa-B kinase (IKK) and Protein kinase B (PKB/Akt)) are measured at several time points by rate of transfer of a P (phosphorous) 32-labeled phosphate. Endothelial cell behaviors such as cell adhesion (using toluidene blue staining), beta1 integrin expression (using Flow cytometry) and thrombotic behavior (Prostacyclin production (using a 6-keto-PGF1alpha competition assay (Cayman Chemicals)) and Coagulation time of blood (using Thromboelastograph)) are quantified on the ten different surfaces. Data are inserted into matrices and then analyzed by Principal component analysis and Partial Least Squares Regression Analysis (Eigenvector, Inc.). From this study, Principal component analyses indicated that with modulation of activity of only four kinases, the ECMs were uniquely identified by ECs. Partial least squares correlated cell behavior with intracellular cell signaling. Cell adhesion and beta1-integrin expression are correlated to high Mitogen activated protein kinases (MAPK) namely ERK and JNK activity whereas the thrombotic behaviors are correlated to high ERK, JNK and Akt activities. These correlations were validated by chemical inhibition of the kinase activity hyphothesized to link the biomaterial property (ECM) to a particular behavior of the HUVECs. This provides insight into how biomaterials can be reverse engineered to achieve the desired cell behavior and also provides a generalizable method for reverse engineering other cell behaviors. |
