Mechanical perturbation of cultured human endothelial cells causes rapid increases of intracellular calcium.
In first-passage human umbilical vein endothelial cells (HUVEC) and bovine aortic endothelial cells (passages 13-16), exposure to gentle mechanical perturbation using a micropipette caused a transient rise in intracellular calcium concentration ([Ca2+]i). The increase in calcium concentration ([Ca2+]) occurred each time the cell was nudged. Three responses were evoked in each of 27 cells using 5 independent HUVEC harvests. Increase in [Ca2+] returned to near baseline levels within approximately 30 s. The stimulus did not cause membrane puncture, as indicated by 1) absence of rapid dye leakage, 2) regulated nature of the [Ca2+] response, 3) absence of membrane blebbing, and 4) repeatable nature of the response in the same cell. As an alternative stimulus, we created very narrow fluid streams (1- to 2-microns diam) from a pressurized pipette that generated shear stresses of approximately 0.001-0.1 dyn/cm2 on the cells. However, these low-shear streams had little effect on [Ca2+]i. The poke-induced change in [Ca2+] was not blocked by lowering extracellular [Ca2+] ([Ca2+]o; 10 microM). In the absence of [Ca2+]o, however, HUVEC did not respond to the first poke, indicating a requirement for some [Ca2+]o as a mediator of signaling. After several poke-induced responses, [Ca2+]i could still be released by caffeine (100 microM), indicating the integrity of the intracellular release mechanism(s). These studies indicate that the response of an endothelial cell to a membrane-deforming event involves a priming step utilizing [Ca2+]o, which facilitates the transient increase of [Ca2+]i.
Sigurdson
W J
WJ
Department of Biophysics, State University of New York, Buffalo 14260.
Sachs
F
F
Diamond
S L
SL
D000814:Aniline Compounds; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D005456:Fluorescent Dyes; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D009994:Osmolar Concentration; D013314:Stress, Mechanical; D013690:Television; D013997:Time Factors; D014966:Xanthenes
D000814:Aniline Compounds; D005456:Fluorescent Dyes; D014966:Xanthenes; C059715:Fluo-3; D002118:Calcium