Study on the SHAP-hyaluronoan (HA) complex as a functional entity of HA in the process of inflammation.

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We have shown that HA in the inflammatory tissues with influx of blood usually contain covalently bound proteins that we named SHAP and fount to correspond to the heavy chain subunits of inter a-trypsin inhibitor (ITI) circulating in blood. Therefore, we hypothesized that the SHAP-HA complex is a functional entity of HA involved in inflammation. Here, we demonstrate evidences obtained from the following experiments shown below, and our final goal is to find some ways to regulate inflammations through the study on the regulation of the SNAP-HA complex formation.1) Electron microscopic observation and biochemical analysis of the complex purified from the synovial fluid obtained from rheumatoid arthritic patients have revealed that the HA of-Mr 1,000,000 in the patient fluid bound about 5 SHAPs and is polymerized via SNAP.2) SHAP-HA complex-knockout mice obtained by homologous recombination of the genes for molecules involved in the ITI synthesis show not only a marked decrease in the ovu … More lation of the cumulus-oocyte complex (COC) which is caused by the defective formation of the COC, due to the absence of the SNAP-HA complex in the COC matrix, but also the impaired fertilization which is likely due to the abnormality of the zona pellucida judging from the IVF (in vitro fertilization) and ISCI (intra-cytoplasmic spermatozoa injection) experiments.3) Comparison of cell adhesion and activation of CD44-positive inflammatory lymphocyte cells, HUT78 to the SHAP-HA complex substrata with those to HA alone has revealed that the cell adhesion to the complex is about 100 times more efficient than that to HA and SHAP bound to HA mediates such a high capacity of the HA-CD44 interaction.4) We have developed the highly sensitive assay method for the activity of the SHAP-HA complex formation and found the presence of the activity in human sera and conditioned media of some liver cells. In collaboration with the Kaketsu Institute Corporation we successfully purified the fraction with the high activity and, based upon the informations of the peptide analysis, have obtained the cDNA of the candidate molecule. We are now on the way to the definitive answer.5) In order to clarify in vivo role of the SNAP-HA complex in inflammation, we induced type II collagen-induced arthritis and dextran sulfate-induced colitis in the knockout mice, and have found that the disease conditions tented to be significantly reduced, compared to those in wild mice. However, this seemed not the case for ConA-induced hepatitis, which suggests some involvement of the deficient formation of ITI itself in the hepatitis. Less