EVIDENCE USING IMMUNOELECTRON MICROSCOPY FOR REGULATED AND CONSTITUTIVE PATHWAYS IN THE TRANSPORT AND RELEASE OF ENDOTHELIN

We investigated the distribution of endothelin (ET)-like immunoreactiv ity in the human coronary artery and examined sites linked to the stor age and release of intracellular proteins. Intense ET-like immunoreact ivity was observed at the light-microscope level in luminal coronary a rtery endothelial cells. A low level of staining also was detected in the outer medial smooth-muscle layer and diffusely within the adventit ia. Immunoelectron microscopy was used to determine the ultrastructura l localisation of ET in the endothelium. Positive ET-like immunoreacti ve staining was detected in secretory vesicles at the ultrastructural level. Quantitative immunoelectron microscopy revealed that ET-like im munoreactivity was predominantly localised to the cytoplasmic matrix ( including secretory vesicles) and Weibel-Palade bodies (endothelial ce ll-specific storage granules). Labelling was detected in 44% of Weibel -Palade body profiles positively identified by using antisera to von W illebrand factor and in cytoplasm surrounding these structures. A low level of immunoreactive staining was associated with mitochondria, whe reas the cell nucleus and Golgi complex showed little or no positive s taining. These findings indicate that ET is released from human corona ry artery endothelial cells via two distinct secretory pathways. We pr opose that ET is continuously transported in and released from secreto ry vesicles by the constitutive secretory pathway. ET may also be stor ed in Weibel-Palade bodies and released after an appropriate stimulus by the regulated pathway. Positive immunoreactivity was also observed in plasmalemmal vesicles (50- to 60-nm diameter), indicating a role fo r these structures in endocytosis.