REGULATION OF INDUCIBLE RAT SALIVARY CYSTATIN

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Cysteine proteinase inhibitors of the cystatin superfamily are grouped into three families on the basis of their molecular structure. Family 1 cystatins are found mainly intracellularly and are present as constituative components of many rat tissues. Family 2 cystatins are mainly present in extracellular fluids. Kininogens are family 3 cystatins, present mainly in the plasma. Family 1 and 3 cystatins have been fairly well characterized in rat model system but very limited information is available on family 2 cystatins of rat. A family 2 cystatin has been very recently purified and sequenced by us from the saliva and submandibular glands of isoproterenol treated rats. As opposed to family 1 cystatins, which are constituative component of rat tissues, family 2 cystatin is not detected in normal rat tissues. Although family 3 cystatins (kininogens) also act as acute-phase reactants in rat plasma, they are also present in the normal rat plasma. These observations suggest that biosynthesis and secretion of the three cystatins may be different and each may have important physiological role to play. It is speculated that these molecules may be involved as a protective measure against increased concentration of endogenous cysteine proteinases during inflammation. Since family 2 rat cystatin in rat submandibular gland is not present as a constituative component, but is induced following isoproterenol treatment, this offers a novel model system to study the biosynthesis and secretion of this molecule under inflammatory conditions. This pilot study is designed to study the feasibility of induction of family 2 cystatins by different secretogogues and agents, known to produce injury to the salivary glands. The aim of the proposed research is to study the effects of the injury producing reagents on the transcriptional and translational regulation of inducible family 2 cystatin from rat submandibular glands. Immunochemical techniques will be used to study the changes in the salivary and submandibular cystatin in rats following in vivo and in vitro stimulation of submandibular glands. cDNA clones corresponding to family 2 cystatin will be isolated and used to study the transcriptional control. In order to understand precisely the importance of cystatins, it is important to understand their structure- function relationship and to understand the mechanism which control their synthesis and secretion. Based on the information achieved in this pilot study, we would like to extend the study to other models of salivary gland injury of acinar and ductal cells and to understand these control mechanism in greater details. The long term goal of this work is to use the information gained in the rat model as a basis for the study of role of cystatins in normal and pathophysiological conditions.