Activated polymorphonuclear leukocytes (PMN) release various types of proteases and express them on the cell surface. The proteases play important roles in PMN-mediated events. In the present study, flow cytometric analysis revealed that CD 14 expression on human gingival fibroblasts (HGF) was markedly reduced by PMA-activated PMN in a coculture system. We found that this reduction was caused by secreted proteases from activated PMN. A protease responsible for the reduction was found to be human leukocyte elastase (HLE) by use of various protease inhibitors. Analysis with purified HLE revealed a timeand dose-dependent reduction of CD 14 on HGF, and complete reduction was observed by 680 nM HLE-treatment for 30-60 min. This reduction of CD 14 resulted from direct proteolysis by HLE on the cell surface, because HLE reduced CD 14 on fixed HGF and also on purified cell membranes. As a result of CD 14 proteolysis, IL-8 production by HGF was suppressed when triggered by 10 ng/ml LPS but not by IL-1 α, indicating that HLE inhibited a CD 14 dependent cell activation. These findings suggested that activated PMNs have a potential negative feedback mechanism for HGF function at the inflammatory site, particularly in periodontal tissues.

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The mechanism of body patterning is complex and includes multiple induction events. Activin, a member of the TGF-β-super family, can induce several kinds of mesodermal and endodermal tissues in the amphibian presumptive ectoderm (animal caps) . The effect of activins on animal caps is distinctly dose dependent, with induction of more dorsal mesodermal tissues and endodermal tissues as the concentration increases. In a recent study of the role of activin in organ formation, we succeeded in raising a beating heart by treating animal caps with a high concentration of activin A. Renal tubules were induced in Xenopus animal caps treated with a combination of activin A and retinoic acid (RA) at a high frequency (100%) accompanying several genes. The renal tubule explants induced by activin and RA in vitro could also function in vivo when the explant was transplanted into the presumptive kidney. Isolated presumptive ectoderm from Xenopus blastula was also treated with activin and retinoic acid to induce differentiation into pancreas. We have already succeeded to make 14 organs and tissues in vitro including sensory organs such as eye and ear vesicle from undifferentiated cells.

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Hepatocyte growth factor (HGF), originally identified and cloned as a long-sought hepatotrophic factor, has mitogenic, motogenic, morphogenic, and anti-apoptotic activities for a wide variety cells. Tissue expression and blood levels of HGF rapidly increases in response to acute injury and endogenously induced HGF plays a role for protection and regeneration/repair of injured tissues. During progression of chronic fibrotic diseases such as liver cirrhosis and chronic renal failure, the expression of HGF decreases in manner reciprocal to the increase in expression of transforming growth factor-β (TGF-β), a key player in tissue fibrosis. Supplement of exogenenous HGF prevents the onset of acute organ failure and chronic fibrotic diseases in laboratory animals. Thus, HGF may be effective for treatment of patients with diseases, by enhancing the intrinsic ability of tissue to regenerate. Our results provide a new therapeutic strategy for practice in regenerative medicine.

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A patient needs the frontier medicine, for example, regenerative medicine, gene therapy and pharmacogenomics. We believe that the regenerative medicine fits for that needs. So, we will discuss about researching and developing a regenerative medicine.
At first, we have to prepare the infrastructure of clinical trial and investment for developing the regenerative medicine. And we will have to build the consortium that contains clinical research center and support center for regenerative medicine. We will call that consortium “regenerative medicine development consortium”.
The clinical research center has a cell processing center, cell and tissue supply center, cell resource development center and cell & tissue bank.
A support center for regenerative medicine has new concept hospital, rehabilitation center, job training center, volunteer organization in support of regenerative medicine and patients.

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Excess of nitric oxide (NO) induces apoptosis in some cell types, including macrophages. Heat shock protein of 70 kDa (hsp 70) has been reported to protect cells from various stresses, including apoptosis-inducing stimuli. Several cytosolic DnaJ homologs, partner chaperones of hsp 70 family members, have been identified in mammals. They include dj 1 (hsp 40/hdj-1) and dj 2 (HSDJ/hdj-2) . A question arose whether these chaperones are required to prevent NO-mediated apoptosis.
Apoptosis occured, when mouse macrophage-like RAW 264.7 cells were treated with an NO donor SNAP. This apoptosis could be prevented by pretreatment of the cells with heat or a low dose of SNAP. With these pretreatment, hsc 70 remained unchanged, whereas hsp 70 and dj 1 showed marked induction and dj 2 moderate induction. In transfection experiments, hsp 70, dj 1 or dj 2 alone was ineffective in preventing NO-mediated apoptosis. In contrast, both dj 1 and dj 2, in combination with hsp 70, prevented the cells from apoptosis. We also found that hsp 70-DnaJ chaperone pairs exert anti-apoptosic effects upstream of caspase 3 activation and also upstream of cytochrome c release from mitochondria.

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The aim of this study was to investigate about endothelial cell injury caused by hypoxia or hypoxia/reoxygenation and the effect of PGE₁ on the injury using human umbilical vein endothelial cells (HUVEC) .
Under hypoxia, HUVEC shrank and peeled from dish, accompanied with reduced thymidine uptake, induced DNA fragmentation, and increased caspase-3 activity. On the other hand, under reoxygenation after hypoxia, HUVEC restarted proliferation as a result of decreasing caspase-3 activity in a time dependent manner.
When PGE₁ was added prior to hypoxia, the number of living cells got fewer and caspase-3 activity was potentiated without changing of cAMP levels. In contrast, the addition of PGE₁ just before reoxygenation, increased the number of living cells and decreased caspase-3 activity with increasing cAMP levels. Under neither hypoxia nor reoxygenation, the gene expression of any prostaglandin EP receptors was not observed. In conclusion, PGE₁ showed opposite effects to be pro-apoptotic under hypoxia, and to be anti-apoptotic under reoxygenation, not through EP receptors.

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To investigate the prevalence of NSAID-related gastrointestinal (GI) complication in patients with rheumatoid arthritis (RA), a cross-sectional analysis was investigated using GI score questionnaire which has been used in the USA as a part of ARAMIS (Arthritis Rheumatism and Aging Medical Information system) . All patients taking NSAID who visited the Institute of Rheumatology, Tokyo Women's Medical University in October, 1999 were requested to answer the questionnaire. The number of patients taking NSAID was 31.4% of all patients who visited the Institute in the month. Analysis of RA patients who has been taking NSAIDs revealed that 31.5% had GI symptoms, 7.2% had histories of having diagnosis of GI ulcer and 1.8% had history of hospitalization due to GI injury. Prevalence of GI ulcer was high in those who were aged or were poor in general condition. These data suggest that the prevalence of NSAID-related GI complication is not few in Japan. In addition, the SCORE point was higher in patients with RA compared with non-RA patients, showing that the RA patients have higher risk for NSAID-related GI complication. However, no difference in the average SCORE point was noted in the class of NSAID. Thus, NSAID use acoording to the risk of each patients were necessary to be instructed.

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High serum cytokine levels may be responsible for various clinical features noted in virus-associated hemophagocytic syndrome (VAHS) . Prognosis of the patients varies widely and the determination of an appropriate therapy for the disease is difficult. Three types of therapy exist: immunochemotherapy which abolishes abnormal cytokine-network: plasma-exchange which reduces serum cytokine levels: and cytotoxic chemotherapy which targets abnormal lymphocyte clones. We successfully treated two of three patients with severe refractory VAHS by a combination of etoposide and cytarabine, both of them have been shown to be cell-cycle phase specific. In two of them, serum IFN-γ level was elevated and Thl dominated: in one, serum IL-4 and IL-6 levels were elevated and Th 2 dominated: and in one, Epstein-Barr virus was a causative agent. We conclude that intensified cytotoxic chemotherapeutic regimens should be considered for the patients who were diagnosed as severe VAHS.

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Cysteinyl leukotrienes (CysLTs) play important roles in the pathogenesis of bronchial asthma, and induce not only the increase of mucus secretion and smooth muscle contraction but also eosinophil recruitment into airways. However, mechanism of eosinophil recruitment by CysLTs has not been thoroughly elucidated. In this study, we examined the LTC₄ on eotaxin production by human lung fibroblasts.
The fibroblasts were cultured with chemical mediators including LTC₄ in the absence or presence of IL-4 for up to 48h. At the end of the culture period, eotaxin in the supernatants was measured by ELISA.
IL-4 clearly enhanced eotaxin production by fibroblasts. LTC₄ alone had no effect on eotaxin production. When fibroblasts were cultured with IL-4 plus LTC₄, eotaxin production was significantly enhanced in comparison with IL-4 alone.
These results suggest that CysLTs may increase eotaxin production by fibroblasts in the presence of IL-4 and that CysLTs may induce eosinophil recruitment into airways through the effect on eotaxin production by fibroblasts in allergic inflammation.

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