Pathologic high shear stress induces apoptosis events in human platelets.

Recently, it has been discovered that apoptosis of anucleate platelets can be induced by chemical agonists. Other studies demonstrated that mechanical forces (shear stresses) stimulate platelet activation and signaling in the absence of exogenous chemical stimuli. We analyzed whether shear stresses can trigger platelet apoptosis, a question that has not yet been studied. Using a cone-and-plate viscometer, we exposed human platelet-rich plasma to different shear stresses, ranging from physiologic arterial and arteriole levels (10-44 dyn/cm2) to pathologic high levels (117-388 dyn/cm2) occurring in stenotic vessels. We found that pathologic shear stresses induce not only platelet activation (P-selectin upregulation and GPIbalpha downregulation) but also trigger apoptosis events, including mitochondrial transmembrane potential depolarization, caspase 3 activation, phosphatidylserine exposure, and platelet shrinkage and fragmentation, whereas physiological shear stresses are not effective. This novel finding suggests that shear-induced platelet apoptosis can be mediated by mechanoreceptors, does not require nuclear participation, and may affect platelet clearance.

Smooth muscle cell growth in monolayer and aortic organ culture is promoted by a nonheparin binding endothelial cell-derived soluble factor/s.

OBJECTIVE: To characterize endothelial derived soluble factor(s) that regulate neointimal formation in porcine aortic organ cultures. METHODS AND RESULTS: Endothelial cell (EC) conditioned medium, collected in preconfluent EC cultures at 4 days after plating, stimulates vascular smooth muscle cell (SMC) growth in cell culture and in the intima of porcine aortic organ cultures. EC conditioned medium was fractionated consecutively by salt precipitation, ion exchange chromatography and affinity chromatography on a heparin column. Heparin column nonbound fraction (HNBF) contains an endothelial cell-derived soluble factor/s (ECDSF) that promotes neointimal formation by increasing intimal SMC (iSMC) proliferation, as detected by BrdU labeling and inhibiting iSMC apoptosis, as shown by TUNEL. Trypsin digestion of HNBF resulted in loss of mitogenic activity. HNBF show a prominent 70-kDa band in SDS-NuPAGE. CONCLUSIONS: Endothelial-derived soluble factor(s) has a molecular weight higher than other growth factors, does not have affinity to heparin, is a protein, at least in the active part of the molecule and increases iSMC number due to increased proliferation and suppression of apoptosis leading to neointimal formation.