IDENTIFICATION AND IMMUNOLOCALIZATION OF ANNEXIN-V AND ANNEXIN-VI THEMAJOR CARDIAC ANNEXINS, IN RAT-HEART

Objective: The heart contains proteins of the annexin family, a unique group of calcium binding proteins. This study was aimed at identifyin g the major cardiac annexins and determining their distribution in the rat heart. Methods: Annexins were isolated by affinity chromatography and purified by ion exchange high pressure liquid chromatography. Ide ntification of isolated proteins by immunoblotting was confirmed by pa rtial amino acid sequence determination. Antisera raised against the i solated proteins were used for immunohistochemistry by the avidin-biot in-peroxidase technique. Results: Two annexins were isolated and purif ied. Amino acid sequencing confirmed their identities as annexin V and VI. Immunohistochemistry showed that both annexins were present in ca rdiac myocytes and non-myocytes, but a distinct pattern of distributio n was seen for each annexin. Annexin V immunoreactivity was enhanced i n the atria compared with the ventricles, whereas annexin VI was more uniformly distributed. In individual cardiac myocytes annexin V was di stributed throughout the cell by contrast with annexin VI, which local ised to the sarcolemma. Intercalated discs displayed immunoreactivity for both annexins, most prominently for annexin VI. The most striking immunoreactivity for annexin V occurred in vascular endothelial cells, both in the microcirculation and in the major coronary vessels. Immun oreactivity for annexin VI in vascular structures was localised to the nuclei of endothelial and smooth muscle cells. Conclusions: Annexins V and VI are the major cardiac annexins. The localisation of these ann exins to different components of cardiac myocytes will serve to direct the search for their functions in the heart. The striking immunoreact ivity for annexins, particularly annexin V, in the coronary vessels in dicates that the functions of cardiac annexins include a role in the c oronary circulatory system.