Angiogenesis is an important biological process not only under physiological conditions but also in a variety of diseases including cancer, diabetic retinopathy and rheumatoid arthritis. Among angiogenic factors, vascular endothelial growth factor-A (VEGF-A) and its receptors appear to mediate the basic signaling for endothelial cell growth and survival, whereas Angiopoietin-Tie system regulates interaction between vascular endothelial cell and smooth muscle cell (pericyte). Furthermore, EphrinB2-EphB4 system determines arterial-venous differentiation. VEGF-A activates two tyrosine kinase receptors, VEGFR-1 (Flt-1) and VEGFR-2 (KDR/Flk-1 in mice). VEGFR-2 is specifically expressed on endothelial cells and is directly involved in angiogenesis. On the other hand, VEGFR-1 is expressed not only on endothelial cells but also on monocyte-macrophages and trophoblasts in placenta, regulating angiogenesis, inflammation and pregnancy. Furthermore, VEGF-C,D and their receptor VEGFR-3 have recently been shown to regulate lymphangiogenesis. Thus, VEGF family and receptor system may be an important target for anti-inflammatory therapy. To regenerate blood vessels in ischemic diseases, VEGF and other angiogenic factors as well as endothelial progenitor cells are thought to be useful, and some are under clinical trial. VEGF-E, novel member of VEGF family, showed an efficient angiogenesis in vivo without side effects such as edema and hemorrhage, suggesting that VEGF-E is also a good candidate for regeneration therapy.

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Rheumatoid arthritis (RA) is a disease raised by synovial proliferation. Overgrown synovium leads to secrete inflammatory cytokines and various kinds of proteinases. Chondrocyte and synovicyte activated by inflammatory cytokines produce extracellular matrix metalloproteinases (MMPs). Among MMPs, matrix metalloproteinase-3 (MMP-3; stromelysin-1) is considered to be the most important proteinases since MMP-3 localizes in erosive cartilage and it has a wide range of substrate specificity. Recently a one-step sandwich enzyme immunoassay (EIA) system for MMP-3 was developed and then several other assay kits have been also used clinically. A lot of clinical data of serum MMP-3 levels measured by these EIA kits have been reported. We review these clinical data of MMP-3 in various diseases and consider that the serum MMP-3 value could be a useful marker for grasping pathology of bone damage and diagnosis for RA. Serum MMP-3 levels in RA patients have been reported to be higher than healthy controls. From these clinical data, MMP-3 may reflect joint destruction and it would be valuable to predict bone damage progression, especially in the early stage of RA. The anti-cytokine therapy such as anti-TNF-α antibody and TNF-α receptor down-regulates serum MMP-3 levels in RA patient. This may suggest that MMP-3 is a useful marker for follow-up of anti-cytokine therapy. Two EIA systems for serum MMP-3, namely, PANACLEAR MMP-3[Plate] and MMP-3[BS], have been approved in Japan as diagnostic tool for RA. The value determined by each kit is different, but correlates each other. To compare with both clinical data, we recommend to convert them with the regression coefficient and to use MMP-3 value for clinical practice.

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As for regenerative medicine in hard tissue field, such as osteoarthritis or bone tumor cases, we have combined both artificial scaffolds and patient's own cells. Osteoarthritis causes the cartilage between the joints to break down, and surgery of total joint replacements using artifitial prosthesis is most common treatment. However, complicatedness which fail in the integrity between the host bone and prosthesis interface are not negligible. Concerning the treatments of bone tumor, synthetic calcium-phosphate ceramics are now available; however, massive bone defects cannot be treated with ceramics themselves. To overcome these problems, we have combined culture-expanded osteogenic cells with synthetic scaffolds. The cell source includes mesenchymal stem cells (MSCs) derived from small amount of patient's bone marrow. We seeded the cells on artificial scaffolds and further cultured to accomplish osteogenic differentiation, resulting in osteoblasts/bone matrix on the scaffolds. We have already applied the hybrid scaffold to more than 30 cases.

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Peptide antibiotics possess potent antimicrobial activities against invading microorganisms and contribute to the innate host defense. We previously revealed that antibacterial cathelicidin CAP11 (cationic antibacterial polypeptide of 11 kDa) exhibits protective actions against endotoxin shock model. During Gram-negative bacterial sepsis, lipopolysaccharide (LPS) activates neutrophils and their apoptosis is suppressed. Prolonged presence of activated neutrophils causes uncontrolled release of toxic metabolites, leading to the systemic tissue injury. In this study, we investigate the action of CAP11 on LPS-induced suppression of neutrophil apoptosis using human neutrophils.
LPS suppressed neutrophil apoptosis, accompanied with the activation of NF-κB, phosphorylation of extracellular signal-related protein kinase, expression of Bcl-XL and inhibition of caspase 3 activation. Interestingly, CAP11 reversed the actions of LPS to trigger these changes, and induced neutrophil apoptosis. Furthermore, LPS activated monocytes to produce anti-apoptotic cytokines (IL-1β, TNF-α and IL-8) and suppressed neutrophil apoptosis. Importantly, CAP11 inhibited the cytokine production from LPS-stimulated monocytes, and induced neutrophil apoptosis. Finally, CAP11 strongly suppressed the LPS-binding to neutrophils and monocytes. These observations indicate that CAP11 can block the LPS-induced prolongation of neutrophil survival via the suppression of anti-apoptotic signaling in neutrophils and anti-apoptotic cytokine production from monocytes by inhibiting the binding of LPS to target cells.

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Vitamin K is widely used for protecting against osteoporosis. Recently, it has been reported that the inhibitory effect of vitamin K₂ (menatetrenone) on bone resorption may be related to its side chain. Geranylgeranylacetone (GGA), known as teprenone, an antiulcer drug, has almost the same chemical structure as that of the side chain of menatetrenone. We hypothesized that GGA also has an inhibitory effect on osteoclastogenesis both in vitro and in vivo. GGA in pharmacological concentrations directly inhibited osteoclastogenesis from human monocytes induced by soluble receptor activator of nuclear factor κB ligand (sRANKL). In addition, GGA induced degradation of actin rings in mature osteoclasts, which was rescued by adding geranylgeranylpyrophosphatase. These results indicate that GGA increases bone mass by maintaining a positive balance of bone turnover through suppression of both the formation and the activity of osteoclasts. Thus, GGA could be used to prevent and improve osteoporosis.

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