Mechanical Stimulation of HUVEC: Different Flow Patterns have Differential Effects on TF Expression

Abstract

Human umbilical vein endothelial cells (HUVECs) are known to respond differentially to various mechanical stimuli, and these different responses maybe involved in atherogenesis. Here we investigated the differential effects of different flow patterns, such as laminar flow (LF) and orbital shear stress (OSS), on the expression of tissue factor (TF). HUVECs were seeded on fibronectin coated glass slides and selectively in the periphery of gelatin coated culture dishes. When HUVECs on slides reached confluence, they were exposed to two, four, or six hours of LF (14 dynes/cm2). HUVECs in culture dishes were exposed to the same time course of low OSS (4 dynes/cm2) or high OSS (11 dynes/cm2). OSS was generated by placing dishes on an orbital shaker. TF mRNA levels were quantified with reverse transcription polymerase chain reaction (RT-PCR), with b-actin as the reference gene. Fold changes are expressed as the mean ± SE. Statistical significance between treated groups and controls was evaluated using the Mann-Whitney test. Analysis of variance (ANOVA) and Tamhane's post-hoc analysis were employed for between group analysis and P < .05 was considered significant. Fold changes as compared with controls are summarized in the Table and Fig (P values are in brackets). LF resulted in significant increases in TF expression throughout the six-hour time course when compared with static controls. However no significant differences were noted between the two, four, and six-hour LF groups. Both HOSS and LOSS reduced TF mRNA levels with significance attained at four and six hours respectively. No significant differences were observed between the different OSS groups.TableTF expression fold changes compared with controlsN = 6HOSSLOSSLF2 hrs1.17 ± 0.15 (.80)1.34 ± 0.37 (.21)10.44 ± 6.63 (.002)4 hrs0.67 ± 0.05 (<.001)0.64 ± 0.1 (.08)9.34 ± 3.8 (.002)6 hrs0.73 ± 0.15 (.07)0.69 ± 0.18 (.03)5.00 ± 1.32 (.056)HOSS, High orbital sheer stress; LF, laminar flow; LOSS, low orbital sheer stress; TF, tissue factor. Open table in a new tab HOSS, High orbital sheer stress; LF, laminar flow; LOSS, low orbital sheer stress; TF, tissue factor. LF induced a significant increase of TF mRNA levels. Interestingly both high and low levels of OSS seemed to decrease TF expression after two hours. These data demonstrate that different flow patterns have differential effects on the expression of TF in HUVEC, indicating the involvement of different intracellular signaling pathways. Current efforts are focused on the interaction of flow patterns with chemical stimulation, as a means of elucidating the intracellular signal conduction pathway.