In this study, we assessed the role of lysophosphatidic acid (LPA) in COX-2 induction using synovial fluid of rheumatoid arthritis (RA) patients in fibroblast-like RA synovial cells. We found that synovial fluid from RA patients stimulated COX-2 induction, which was associated with prostaglandin E2 production, in RA synovial cells. The synovial fluid-induced responses were inhibited by pertussis toxin, Gi/0 protein inhibitor, and by Ki16425, antagonist for LPA receptors (LPA1 and LPA3). Indeed, LPA effectively induced COX-2 expression and prostaglandin E2 production. The LPA-induced actions were markedly inhibited by pertussis toxin and Ki16425. Furthermore, LPA synergistically enhanced IL-1-induced COX-2 expression and prostaglandin E2 production in a manner sensitive to pertussis toxin and Ki16425. RA synovial cells abundantly expressed LPA1 receptor compared with other LPA receptor subtypes. The LPA content in RA synovial fluid was just the threshold to induced the action. These results demonstrated that LPA existing in the RA synovial fluid plays a critical role in COX-2 induction in collaboration with inflammatory cytokine, IL-1, in RA synovial cells.

Royal jelly contains a variety of molecules biologically active towards various types of cells. We recently found that an water-soluble extract of royal jelly inhibited COX-2 expression and cellular proliferation in osteosarcoma MG-63 cells. To identify active components in royal jelly that regulate cellular proliferation of MG-63 cells, a water-soluble extract of royal jelly was separated by HPLC. The molecular structure of substances thus isolated was analysed and the structures of 10 substances were determined. AMP N1-oxide and adenosine N1-oxide were found to be active component in royal jelly that inhibits cellular proliferation by inducing G0/G1 cell cycle arrest in MG-63 cells. These results suggest that AMP N1-oxide and adenosine N1-oxide existing in royal jelly might be a useful therapeutic agent for treatment of bone cancer. Further studies on the action mechanism of the compound are now under way.

The complement receptor type 3 (CR3) is a hetero-dimeric molecule consisting of the a and the b subunits. This molecule is involved in leukocyte adhesion reactions through its binding to a variety of ligand molecules. Recently, we have found that guinea pig CR3 can act as an HA binding protein. However, the localization of the ligand binding site on this molecule has not been studied. To solve the problem, we first analyzed the primary structure of the CR3 a subunit and compared it with that of the human CR3 a subunit. We found that the guinea pig CR3 a subunit, but not the human CR3 a subunit, possesses a BX7B motif that is known to be involved in the HA binding activity of the RHAMM molecule. Other HA binding structures such as the Link module that is shared by most HA binding proteins was not found on the guinea pig or human CR3 a subunit. These results suggest the possibility that guinea pig CR3 but not human CR3 may use the BX7B motif of the a subunit in the interaction with the HA molecule. To further clarify the correlation between the function and molecular structure of this molecule, analysis of total primary structure of the b subunit is now in progress.

Signaling of nuclear receptors plays an important role in immune system. We have found that some agonists for the nuclear receptors can regulate homing and differentiation of lymphocytes. Our goal is to develop new treatment for autoimmune diseases.

We found that only GABARAPL1 mRNA among LC3 family members and mitochondria protein Nix mRNA are upregulated by weakly basic agents or mitochondrial membrane potential disrupter, CCCP. These results suggest that GABARAPL1 may play an important role in mitophagy, an autophagy-related pathway specific for mitochondria. Further studies on the mechanism of autophagy are under way.