EXPRESSION OF THE CYSTEINE PROTEINASE-INHIBITOR CYSTATIN-C GENE IN RAT-HEART - USE OF DIGOXIGENIN-LABELED PROBES GENERATED BY POLYMERASE CHAIN-REACTION DIRECTLY FOR IN-SITU AND NORTHERN BLOT HYBRIDIZATIONS

Cystatins represent a widely distributed superfamily of cysteine proteinase inhibitory proteins. We investigated the expression of the cystatin C gene, belonging to the family 2 of cystatins, in the hearts of female rats. Using a highly sensitive reverse transcriptase-polymerase chain reaction (RT-PCR) we have detected cystatin C mRNA in the ventricle and atrium, as well as in liver add submandibular gland. A digoxigenin-labeled cystatin C probe, generated by PCR, hybridized to a single mRNA species of about 700 nucleotides on Northern blots. Northern blot hybridizations established that neither an acute inflammation produced by injection of turpentine nor administration of the beta-adrenergic agonist isoproterenol had an effect on the level of cystatin C mRNA in the heart. In situ hybridizations with digoxigenin-labeled probe localized the expression of the cystatin C gene to cardiac muscle fibers but not to other cardiac cellular elements. Cystatin C may be released by cardiac muscle fibers under physiological and pathological conditions and may modify inflammatory and necrobiotic processes.