The tumour-necrosis-factor-family molecule osteoprotegerin ligand (OPGL; also known as TRANCE, RANKL and ODF) has been identified as a potential osteoclast differentiation factor and regulator of interactions between T cells and dendritic cells in vitro. Mice with a disrupted opgl gene show severe osteopetrosis and a defect in tooth eruption, and completely lack osteoclasts as a result of an inability of osteoblasts to support osteoclastogenesis. Although dendritic cells appear normal, opgl-deficient mice exhibit defects in early differentiation of T and B lymphocytes. Surprisingly, opgl-deficient mice lack all lymph nodes but have normal splenic structure and Peyer's patches. Thus OPGL is a new regulator of lymph-node organogenesis and lymphocyte development and is an essential osteoclast differentiation factor in vivo.

OPGL is a key regulator of osteoclastogenesis, lymphocyte development and lymph-node organogenesis

Src family protein tyrosine kinases are activated following engagement of many different classes of cellular receptors and participate in signaling pathways that control a diverse spectrum of receptor-induced biological activities. While several of these kinases have evolved to play distinct roles in specific receptor pathways, there is considerable redundancy in the functions of these kinases, both with respect to the receptor pathways that activate these kinases and the downstream effectors that mediate their biological activities. This chapter reviews the evidence implicating Src family kinases in specific receptor pathways and describes the mechanisms leading to their activation, the targets that interact with these kinases, and the biological events that they regulate.

Cellular functions regulated by src family kinases

Prostanoids are the cyclooxygenase metabolites of arachidonic acid and include prostaglandin (PG) D2, PGE2, PGF2α, PGI2, and thromboxne A2. They are synthesized and released upon cell stimulation and act on cells in the vicinity of their synthesis to exert their actions. Receptors mediating the actions of prostanoids were recently identified and cloned. They are G protein-coupled receptors with seven transmembrane domains. There are eight types and subtypes of prostanoid receptors that are encoded by different genes but as a whole constitute a subfamily in the superfamily of the rhodopsin-type receptors. Each of the receptors was expressed in cultured cells, and its ligand-binding properties and signal transduction pathways were characterized. Moreover, domains and amino acid residues conferring the specificities of ligand binding and signal transduction are being clarified. Information also is accumulating as to the distribution of these receptors in the body. It is also becoming clear for some types of ...

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Prostanoid Receptors: Structures, Properties, and Functions

In this review most of the various known, suspected, or postulated functions of osteopontin, a secreted highly acidic phosphoprotein, are discussed in terms of what we currently know about the protein. These include 1) binding of OPN both to cells via a GRGDS cell adhesion sequence that recognizes the alpha v beta 3 integrin and to extracellular matrix components via poorly characterized motifs, 2) regulation of the formation and remodeling of mineralized tissue, 3) recruiting and stimulating macrophages and lymphocytes as part of a nonspecific response to microbial infections, 4) multiple interactions with Ca2+ that likely influence OPN protein conformation and may be important in Ca(2+)-mediated or Ca(2+)-dependent processes, 5) inhibiting the growth of calcium oxalate crystals by disruption of the growing crystal lattice, 6) effects on gene expression, Ca2+ regulation, and nitric oxide production, and 7) involvement in cell migration. OPN production is frequently augmented when cell signaling pathways are activated by any of a variety of stimuli, for example in cancer cells.

Osteopontin: a protein with diverse functions.

I. Introduction BONE is a complex tissue in which resorption and formation continue throughout life. This process is called bone remodeling. Osteotropic hormones such as 1α,25-dihydroxyvitamin D3 [1α,25(OH)2D3], PTH, and calcitonin preferentially modulate the process of bone resorption to maintain bone remodeling. The bone tissue contains various types of cells, of which the bone-forming osteoblasts and bone-resorbing osteoclasts are mainly responsible for bone remodeling. Osteoblasts are believed to be derived from undifferentiated mesenchymal cells, which further differentiate into osteocytes and are embedded in calcified tissues. Osteoclasts are multinucleated cells present only in bone. It is believed that osteoclast progenitors are of hemopoietic origin, and they are recruited from hemopoietic tissues such as bone marrow and circulating blood to bone. Osteoclast progenitors then proliferate and differentiate into mononuclear preosteoclasts and fuse with each other to form multinucleated osteoclasts. ...

Modulation of Osteoclast Differentiation

Identification of osteopontin in isolated rabbit osteoclasts.

Generation of osteoclasts from isolated hematopoietic progenitor cells.

The effect of prostaglandin E2 (PGE2) on osteoblastic cell proliferation was investigated using osteoblastic clone MC3T3-E1 cells cultured in serum-free medium. PGE2 at 2 micrograms/ml increased the number of the cells by 2 days after its addition. PGE2 raised the level of DNA synthesis in a dose-related fashion after a constant lag time, the maximal effect being at 2-10 micrograms/ml and the level about fourfold over that of the control at 36 hr after its addition. However, at low doses (below 0.2 microgram/ml), PGE2 rather depressed DNA synthesis. Isobutyl methylxanthine counteracted the stimulation of DNA synthesis by PGE2, and forskolin depressed the synthesis, which was inversely correlated with increasing intracellular cAMP content. These results indicate that an increase in cAMP content inhibits DNA synthesis. In addition, 2',5'-dideoxyadenosine did not negate the stimulatory effect of PGE2 on DNA synthesis, suggesting that PGE2 increases DNA synthesis, probably via a pathway different from the adenylate cyclase/cAMP system. Moreover, at a high dose, PGE2 stimulated both the production and degradation of cAMP; the elevation of cAMP content was rapidly depressed by the stimulated degradation system. Consequently, the stimulatory effect of PGE2 on DNA synthesis would be released from the inhibition by cAMP, resulting in an increase in DNA synthesis. Taken together with data from our previous reports, these results indicate that PGE2 enhances both the proliferation and differentiation of osteoblastic cells in vitro, which are probably mediated by two different second messengers dependent on the concentration of PGE2.

Prostaglandin E2 stimulates DNA synthesis by a cyclic AMP-independent pathway in osteoblastic clone MC3T3-E1 cells.

1,25-Dihydroxyvitamin D3 [1,25-(OH)2D3] was examined for a possible stimulative effect on osteoblastic MC3T3-E1 cells. During the early period of culture, 1,25-(OH)2D3 had a stimulative effect. During the growth phase, however, the steroid had little effect on either the protein or DNA content of the cultures. 1,25-(OH)2D3 increased bone-liverkidney-type alkaline phosphatase activity in a dose-related manner up to a concentration of 5 pg/ml; the increase was 2.2-fold over the control value. Studies on the effect of actinomycin D or cycloheximide treatment indicated that the vitamin may enhance de novo synthesis of ALP. The steroid also stimulated type I collagen production dose dependently via an increase in collagen synthesis rather than by inhibition of collagen degradation. MC3T3-E1 cells have a specific receptor for 1,25-(OH)2D3 which has a dissociation constant of 4.17 × 10−11 m and a sedimentation coefficient of 3.67S. The receptor concentration varied with the period of culture, being higher during...

Effects of 1,25-dihydroxyvitamin D3 on osteoblastic MC3T3-E1 cells.

The effects of prostaglandins (PGs) on the induction of alkaline phosphatase (ALP) were investigated in osteoblastic clone MC3T3-E1 cells cultured in serum-free medium. Prostaglandin E2 (PGE2) stimulated ALP activity in the cells in a dose-dependent fashion with a maximal effect which was about twice that in the control cells at concentrations of 100-500 ng/ml. Actinomycin D and cycloheximide inhibited the stimulative effect of PGE2 on ALP activity in the cells. PGE2-induced and native ALPs in the cells were of the same type as that in adult mouse calvaria, being heat-labile, L-homoarginine- and levamisole-sensitive, and L-phenylalanine-insensitive. Isobutyl methylxanthine (IBMX), a cAMP phosphodiesterase inhibitor, stimulated the inductive effect of PGE2 on ALP activity at 0.1 mM, at which concentration IBMX alone had little effect on the activity. PGE2 also increased the intracellular cAMP content in a dose-dependent fashion with a maximal effect at 100 ng/ml. PGE1, PGF1 alpha, and PGF2 alpha (primary PGs like PGE2) increased the activity. Our present results suggest that PGs stimulate the differentiation of osteoblasts and are involved in bone formation in vivo, as well as in bone resorption.

Noriyoshi Kurihara

Papers

Identification of osteopontin in isolated rabbit osteoclasts.

Generation of osteoclasts from isolated hematopoietic progenitor cells.

Prostaglandin E2 stimulates DNA synthesis by a cyclic AMP-independent pathway in osteoblastic clone MC3T3-E1 cells.

Effects of 1,25-dihydroxyvitamin D3 on osteoblastic MC3T3-E1 cells.

Inductive effects of prostaglandins on alkaline phosphatase in osteoblastic cells, clone MC3T3-E1.