Expression of ADAMTS1 in endothelial cells is induced by shear stress and suppressed in sprouting capillaries

Abstract

ADAMTS1 inhibits capillary sprouting, and since capillary sprouts do not experience the shear stress caused by blood flow, this study undertook to clarify the relationship between shear stress and ADAMTS1. It was found that endothelial cells exposed to shear stress displayed a strong upregulation of ADAMTS1, dependent upon both the magnitude and duration of their exposure. Investigation of the underlying pathways demonstrated involvement of phospholipase C, phosphoinositide 3-kinase, and nitric oxide. Forkhead box protein O1 was identified as a likely inhibitor of the system, as its knockdown was followed by a slight increase in ADAMTS1 expression. In silico prediction displayed a transcriptional binding site for Forkhead box protein O1 in the promotor region of the ADAMTS1 gene, as well as sites for nuclear factor 1, SP1, and AP-1. The anti-angiogenic effects of ADAMTS1 were attributed to its cleavage of thrombospondin 1 into a 70-kDa fragment, and a significant enhancement of this fragment was indeed demonstrated by immunoblotting shear stress-treated cells. Accordingly, scratch wound closure displayed a slowdown in conditioned medium from shear stress-treated endothelial cells, an effect that could be completely blocked by a knockdown of thrombospondin 1 and partially blocked by a knockdown of ADAMTS1. Non-perfused capillary sprouts in rat mesenteries stained negative for ADAMTS1, while vessels in the microcirculation that had already experienced blood flow yielded the opposite results. The shear stress-dependent expression of ADAMTS1 in vitro was therefore also demonstrated in vivo and thereby confirmed as a mechanism connecting blood flow with the regulation of angiogenesis.

Pulsatile Atheroprone Shear Stress Affects the Expression of Transient Receptor Potential Channels in Human Endothelial Cells

“…16 We showed that a shear stress profile taken from an atheroprone site increases TRPC6 and TRPV1 mRNA expression in endothelial cells. Although atheroprone shear stress increased TRPC6 and TRPV1 transcripts, protein levels were not significantly affected within 24 hours.…”

“…Third, static, that is, no-flow conditions, represent nonperfused vessels and occur in vivo only in sprouting capillaries and after vessel occlusion. 16,[20][21][22] Morphological changes represent a late response of endothelial cells on flow exposure. 23 The experiments performed confirmed typical morphological changes of human endothelial cells under the influence of atheroprotective shear stress 24 versus atheroprone 25 and static …”

“…Using the Transcriptor first-strand cDNA synthesis kit (Roche Diagnostics, Mannheim, Germany), cDNA was synthesized from 2 g of total RNA using oligo dT [12][13][14][15][16][17][18] and 5 U of Avian Myeloblastosis Virus reverse transcriptase at 50°C for 60 minutes, followed by heating to 85°C for 5 minutes.…”

“…We analyzed the expression of TRP channel mRNA in an established in vitro dynamic flow system, 4 in which HUVECs are exposed to 2 well-defined types of mechanical stimulation, considered to have direct pathophysiological relevance to atherosclerotic disease in vivo. Specifically, we compared the effect of an atheroprotective flow profile with shear stress ranging from 0 to 40 dyne/cm 2 atheroprone bidirectional flow profile with a mean shear stress near 0 dyne/cm 24,16 on TRP channel mRNA expression. TRPC6 and TRPV1 mRNA expressions were significantly increased after 24 hours of exposure to an atheroprone flow profile compared with an atheroprotective flow profile (normalized ratio, 185Ϯ21 versus 100Ϯ26 arbitrary units; Pϭ0.04; each nϭ6 for TRPC6; normalized ratio, 260Ϯ48 versus 100Ϯ35 arbitrary units; Pϭ0.02; each nϭ6 for TRPV1).…”

Synergistic Regulation of Angiogenic Sprouting by Biochemical Factors and Wall Shear Stress

“…Recombinant, soluble ADAMTS1 inhibited angiogenesis in corneal pocket and chick chorioallantoic angiogenic assays (107). In rat mesentery, Hohberg et al (48) demonstrated positive immunostaining for ADAMTS1 in microvessels experiencing flow, while there was a lack of ADAMTS1 staining in unperfused sprouts. Hohberg et al also reported that ADAMTS1 expression and liberation of TSP fragments in HUVECs increased monotonically with WSS in the range of 1 to 20 dyn/cm 2 (28).…”