Emerging mechanisms for growth and protection of the vasculature by cytochrome P450-derived products of arachidonic acid and other eicosanoids.

Arachidonic acid (AA) is an essential fatty acid that is metabolized by cyclooxygenase (COX), lipoxygenase (LOX) or cytochrome P450 (CYP) enzymes to generate eicosanoids which in turn mediate a number of biological activities including regulation of angiogenesis. While much information on the effects of COX and LOX products is known, the physiological relevance of the CYP-derived products of AA are less well understood. CYP enzymes are highly expressed in the liver and kidney, but have also been detected at lower levels in the brain, heart and vasculature. A number of these enzymes, including members of the CYP 4 family, predominantly catalyze conversion of AA to 20-hydroxyeicosatetraenoic acid (20-HETE) while the CYP epoxygenases generate mainly epoxyeicosatrienoic acids (EETs). This review will focus on the emerging roles of inhibitors of eicosanoid production with emphasis on the CYP pathways, in the regulation of angiogenesis and tumor growth. We also discuss current observations describing the protective effects of EETs for survival of the endothelium.

Chick chorioallantoic membrane as an in vivo model to study vasoreactivity: characterization of development-dependent hyperemia induced by epoxyeicosatrienoic acids (EETs).

Shell-less culture of chick chorioallantoic membrane (CAM) of developing chicken embryos is a useful model to evaluate the effects of vascular agents. We assessed the response of CAM vessels to epoxyeicosatrienoic acids (EETs), derivatives of the essential fatty acid arachidonic acid, that have a number of important biological functions, including dilation of microvessels in the coronary, cerebral, renal, and mesenteric circulations. Three of four regioisomers of EETs, 14,15-, 11,12-, and 8,9-EET, induced a characteristic dose-dependent acute hyperemia within 4 min after application on 10-day-old CAMs. This response was marked in early stages of development (between days 8 and 10), but the frequency and intensity of the response were reduced after 11 days of development. Histological examination demonstrated that the hyperemia was not due to extravasation of erythrocytes. However, many capillaries were distended and contained densely packed erythrocytes as compared to uniformly arranged vessels and erythrocytes in untreated CAMs. Transmission electron microscopy showed the basal laminae surrounding capillaries remained intact, similar to those in vehicle-treated or untreated CAM tissue. The hyperemia was specific to EETs since we did not observe it to be induced by other vasodilators such as nitric oxide or prostacyclin. In conclusion, we report a novel vascular response to EETs using the CAM as an in vivo model. These lipids specifically distend a subset of capillaries in a dose- and development-dependent manner.

Role of JNK in network formation of human lung microvascular endothelial cells.

The signaling mechanisms in vasculogenesis and/or angiogenesis remain poorly understood, limiting the ability to regulate growth of new blood vessels in vitro and in vivo. Cultured human lung microvascular endothelial cells align into tubular networks in the three-dimensional matrix, Matrigel. Overexpression of MAPK phosphatase-1 (MKP-1), an enzyme that inactivates the ERK, JNK, and p38 pathways, inhibited network formation of these cells. Adenoviral-mediated overexpression of recombinant MKP-3 (a dual specificity phosphatase that specifically inactivates the ERK pathway) and dominant negative or constitutively active MEK did not attenuate network formation in Matrigel compared with negative controls. This result suggested that the ERK pathway may not be essential for tube assembly, a conclusion which was supported by the action of specific MEK inhibitor PD 184352, which also did not alter network formation. Inhibition of the JNK pathway using SP-600125 or l-stereoisomer (l-JNKI-1) blocked network formation, whereas the p38 MAPK blocker SB-203580 slightly enhanced it. Inhibition of JNK also attenuated the number of small vessel branches in the developing chick chorioallantoic membrane. Our results demonstrate a specific role for the JNK pathway in network formation of human lung endothelial cells in vitro while confirming that it is essential for the formation of new vessels in vivo.