Search

Skeletal muscle is a major tissue of glucose consumption and plays an important role in glucose homeostasis. Prenylflavonoids, a component of Macaranga tanarius fruits, have been reported to have antioxidant, antibacterial, and anticancer effects. However, the effects of these compounds on skeletal muscle glucose metabolism are unclear. Here, we isolated five prenylflavonoids from M. tanarius fruits, and investigated the mechanism of action of these compounds on skeletal muscle cells using L6 myotubes. We found that isonymphaeol B and 3′-geranyl naringenin increased glucose uptake in a dose-dependent manner. Furthermore, both isonymphaeol B and 3′-geranyl naringenin increased AMPK phosphorylation but did not affect PI3K-Akt phosphorylation. Isonymphaeol B and 3′-geranyl naringenin also increased Glut1 mRNA expression and plasma membrane GLUT1 protein levels. These results suggest that isonymphaeol B and 3′-geranyl naringenin have beneficial effects on glucose metabolism through AMPK and GLUT1 pathway. Isonymphaeol B and 3′-geranyl naringenin may be potential lead candidates for antidiabetic drug development. [ABSTRACT FROM AUTHOR]

Bone mass is regulated by osteoblast-mediated bone formation and osteoclast-mediated bone resorption. Osteoporosis is a bone metabolism disorder in which bone mass decreases due to increased bone resorption rather than bone formation. We focused on the traditional plant Alpinia zerumbet in Okinawa, Japan, and searched for promising compounds for the prevention and treatment of osteoporosis. Pinocembrin isolated from the leaves of A. zerumbet showed enhanced alkaline phosphatase (ALP) activity and mineralization and increased mRNA expression of osteoblast-related genes Alp and Osteocalcin (Ocn) in MC3T3-E1 cells. Pinocembrin increased the mRNA expression of Runx2 and Osterix, which are important transcription factors in osteoblast differentiation, and the mRNA expression of Dlx5 and Msx2, which are enhancers of these transcription factors. The bone morphogenetic protein (BMP) antagonist noggin, its receptor kinase inhibitor LDN-193189 and p38 MAPK inhibitor SB203580 attenuated pinocembrin-promoted ALP activity. Pinocembrin increased the mRNA of Bmp-2 and its target gene Id1. In addition, the estrogen receptor (ER) inhibitor ICI182780 suppressed pinocembrin-stimulated ALP activity. Pinocembrin may increase BMP-2 expression via ER. Then, the BMP-2 promotes osteoblast specific genes expression and mineralization through both Smad-dependent and independent pathway following Runx2 and Osterix induction. Our findings suggest that pinocembrin has bone anabolic effects and may be useful for the prevention and treatment of bone metabolic diseases such as osteoporosis. [ABSTRACT FROM AUTHOR]

Alcohol consumption causes the accumulation of reactive oxygen species in liver, which leads to alcoholic fatty liver and hepatocyte injury. In this study, we investigated the effects of an ethanolic Citrus depressa extract and those of its main components on alcohol-induced liver damage using a mouse model. Four polymethoxylated flavonoids, namely, nobiletin, tangeretin, 5-O-demethylnobiletin, and sinensetin, were isolated from C. depressa extract. Treatment of ethanol fed mice with C. depressa extract, nobiletin, tangeretin, or 5-O-demethylnobiletin at 300 mg/kg for 8 weeks by oral administration alleviated the accumulation of lipid droplets in liver and significantly decreased the serum levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) (markers of liver damage). Also, in mice treated with ethanol plus nobiletin, tangeretin, or 5-O-demethylnobiletin, liver level of glutathione (an antioxidant) increased whereas levels of tumor necrosis factor-alpha (TNF-α), hepatic malondialdehyde, and hepatic cytochrome P450 2E1 (CYP2E1) mRNA decreased as compared with ethanol fed controls. These findings suggest C. depressa extract and polymethoxylated flavonoids had a protective effect on alcohol-induced liver injury, and that the mechanism involved is related to the regulation of hepatic CYP2E1-mediated oxidative stress. [ABSTRACT FROM AUTHOR]

Acinetobacter baumannii is a nosocomial pathogen that can survive unfavorable conditions, such as, desiccation, nutrient starvation, and antimicrobial treatment, and this is primarily due to its ability to form biofilms on biotic and abiotic surfaces like tissues and medical devices. In this study, honokiol and magnolol were investigated for antibiofilm activity against A. baumannii ATCC 17978. Both were found to inhibit biofilm formation dose-dependently and to disperse matured biofilms. Honokiol and magnolol were found to inhibit biofilm formation by five and four of eight additional clinical A. baumannii isolates, respectively. Furthermore, honokiol and magnolol effectively suppressed pellicle formation and the surface motilities of the A. baumannii and prolonged the survival of infected nematode Caenorhabditis elegans. These results demonstrate that honokiol and magnolol may be useful for controlling A. baumannii infections. [ABSTRACT FROM AUTHOR]

Nepodin, found in the roots of Rumex japonicus Houtt. (Polygonaceae), inhibits osteoclast differentiation and has an antidiabetic effect. We propose nepodin as an ingredient of new functional foods or as a drug candidate for reducing the risk of reduced locomotion resulting from diseases such as osteoporosis. Although there are no previous reports of R. obtusifolius L., which is found throughout Japan, having roots containing nepodin, we found nepodin in the roots of this species. Therefore, R. obtusifolius as well as R. japonicus was considered a candidate raw material for nepodin extraction. We also discuss the suitability of R. japonicus and R. obtusifolius as sources of raw nepodin for cultivation on the Ryukyu Islands. In this study, all specimens on the Ryukyu Islands were identified as R. japonicus. Conversely, all specimens on mainland Japan were R. obtusifolius. The DNA sequence of the chloroplast trnL- trnF intergenic spacer region and partial nuclear internal transcribed spacer was consistent with the identification of R. japonicus and R. obtusifolius by morphological characteristics of the perianth segments. Therefore, to avoid erroneous identification and misuse of the plant species used for extraction of raw materials, it is preferable to develop DNA markers for these two regions. The content of nepodin varied from undetectable to 0.34% of the fresh weight (%FW) in R. japonicus and from undetectable to 0.21%FW in R. obtusifolius. From a pharmacological perspective, as plants that might be suitable as raw materials for nepodin extraction, it became clear that both R. japonicus and R. obtusifolius can be used with the same expected extraction efficiency. Based on our findings, R. obtusifolius could not be confirmed as inhabiting the Ryukyu Islands. For this reason, to conserve the endemic genetic characteristics of the Ryukyu Islands and to prevent genetic pollution by R. obtusifolius, only R. japonicus should be cultivated on the Ryukyu Islands. [ABSTRACT FROM AUTHOR]

Honokiol and magnolol are neolignans contained in the Chinese medicinal herb Magnolia officinalis that exert anti-tumor and anti-inflammatory effects. Both compounds have been reported to enhance glucose uptake. Little is known about effects when used in combination. The effects of honokiol, magnolol, and a combination of both compounds on lipid and glucose metabolism in highfat diet-induced obese mice were investigated, and underlying mechanisms were examined. All 3 treatments significantly ( p<0.05) reduced plasma total cholesterol and glucose levels, and improved glucose tolerance, compared with controls. In addition, treatments increased mRNA expression of the peroxisome proliferator-activated receptor (PPAR)-γ, glucose transporter (GLUT)-4, and adiponectin genes in white adipose tissue (WAT). Both compounds individually and in combination significantly ( p<0.05) increased Akt phosphorylation and GLUT4 protein expression in WAT compared to the control group. Honokiol and magnolol improve dyslipidemia and hyperglycemia and act synergistically when used in combination. [ABSTRACT FROM AUTHOR]

Using B16 melanoma cells for screening, we found that a marine sponge extract has a potent anti-pigmenting effect and identified arenarol as its major active compound. In normal human melanocytes (NHMs), arenarol significantly abrogated the endothelin 1 (EDN1) stimulated expression of tyrosinase, tyrosinase-related protein 1 and dopachrome tautomerase at the transcriptional, translational and enzymatic activity (only for tyrosinase) levels. That effect was accompanied by the attenuation of the increased expression level of microphthalmia-associated transcription factor (MITF) protein at the transcriptional and translational levels. Analysis of EDN1 signaling demonstrated that arenarol significantly suppressed the EDN1-induced phosphorylation of MEK, ERK, MITF and CREB but not of Raf-1s. In contrast, the forskolin-induced phosphorylation of CREB was not down-regulated by arenarol. As for the mode of action of the suppressed phosphorylation of MEK, Raf-1 activity was not directly inhibited by arenarol in vitro and pretreatment with the protein phosphatase inhibitor okadaic acid did not affect the down-regulated phosphorylation of MEK that was induced by arenarol in NHMs. The sum of these findings suggests that arenarol abrogates the EDN1-stimulated expression of melanocyte-specific proteins by interrupting MEK phosphorylation in an as yet unknown Raf-1 inactivation mechanism. [ABSTRACT FROM AUTHOR]

Flos Magnoliae (FM) is a commonly used Chinese medicinal herb for symptomatic relief of allergic rhinitis, sinusitis and headache. Although several FM species have been used as substitutes or adulterants for clinical use, possible differences in their pharmacological actions have not been reported. To confirm the effects of FM on skeletal muscle glucose metabolism, we tested the effects of several compounds isolated from FM on glucose uptake by L6 myotubes. We found that fargesin, a component of FM, dose-dependently stimulated glucose consumption in L6 myotubes, which was accompanied by enhanced glucose transporter (GLUT)-4 translocation to the cell surface. Fargesin-stimulated glucose uptake was blocked by wortmannin, a phosphatidylinositol-3 kinase (PI3 K) inhibitor. Fargesin stimulated Akt phosphorylation, a key component in the insulin signaling pathway, which was completely inhibited by wortmannin. Here, we demonstrated that fargesin, a bioactive component of Flos Magnoliae, increases basal glucose uptake and GLUT4 translocation in L6 myotubes by activating the PI3 K-Akt pathway. [ABSTRACT FROM AUTHOR]

Leaves develop as flat lateral organs from the indeterminate shoot apical meristem. The establishment of polarity along three-dimensional axes, proximal-distal, medial-lateral, and adaxial-abaxial axes, is crucial for the growth of normal leaves. The mutations of ASYMMETRIC LEAVES1 ( AS1) and AS2 of Arabidopsis thaliana cause defects in repression of the indeterminate state and the establishment of axis formation in leaves. Although many mutations have been identified that enhance the adaxial-abaxial polarity defects of as1 and as2 mutants, the roles of the causative genes in leaf development are still unknown. In this study, we found that wild-type plants treated with berberine produced pointed leaves, which are often observed in the single mutants that enhance phenotypes of as1 and as2 mutants. The berberine-treated as1 and as2 mutants formed abaxialized filamentous leaves. Berberine, an isoquinoline alkaloid compound naturally produced in various plant sources, has a growth inhibitory effect on plants that do not produce berberine. We further showed that transcript levels of meristem-specific class 1 KNOX homeobox genes and abaxial determinant genes were increased in berberine-treated as1 and as2. Berberine treated plants carrying double mutations of AS2 and the large subunit ribosomal protein gene RPL5B showed more severe defects in polarity than did the as2 single mutant plants. We suggest that berberine inhibits (a) factor(s) that might be required for leaf adaxial cell differentiation through a pathway independent of AS1 and AS2. Multiple pathways might play important roles in the formation of flat symmetric leaves. [ABSTRACT FROM AUTHOR]

The dried flower buds of Magnolia sp. are widely used as herbal medicines because of their anti-inflammatory, anti-malarial and anti-platelet activities. Here, we found that veraguensin and galgravin, lignan compounds derived from Magnolia sp., dose-dependently inhibited osteoclast formation in co-cultures of bone marrow cells and osteoblastic cells. These compounds also inhibited receptor activator of nuclear factor κB ligand (RANKL)-induced osteoclast differentiation in RAW264.7 cells and bone marrow macrophages. In the RANKL-induced signaling pathway, veraguensin and galgravin reduced p38 phosphorylation and suppressed the expression of c-Fos, a key transcription factor for osteoclastogenesis. Veraguensin and galgravin also inhibited osteoclastic pit formation, which was accompanied by decreased mature osteoclast viability. In conclusion, these results indicate that veraguensin and galgravin can inhibit bone resorption and may offer novel compounds for the development of drugs to treat bone-destructive diseases such as osteoporosis. [ABSTRACT FROM AUTHOR]

Excessive receptor activator of NF-κB ligand (RANKL) signaling causes enhanced osteoclast formation and bone resorption. The downregulation of RANKL expression and its downstream signals may be an effective therapeutic approach to the treatment of bone loss diseases such as osteoporosis. Here, we found that coptisine, one of the isoquinoline alkaloids from Coptidis Rhizoma, exhibited inhibitory effects on osteoclastogenesis in vitro. Although coptisine has been studied for its antipyretic, antiphotooxidative, dampness dispelling, antidote, antinociceptive, and anti-inflammatory activities in vitro and in vivo, its effects on osteoclastogenesis have not been investigated. Therefore, we evaluated the effects of coptisine on osteoblastic cells as well as osteoclast precursors for osteoclastogenesis in vitro. The addition of coptisine to cocultures of mouse bone marrow cells and primary osteoblastic cells with 10 M 1α,25(OH)D caused significant inhibition of osteoclast formation in a dose-dependent manner. Reverse transcriptase polymerase chain reaction (RT-PCR) analyses revealed that coptisine inhibited RANKL gene expression and stimulated the osteoprotegerin gene expression induced by 1α,25(OH)D in osteoblastic cells. Coptisine strongly inhibited RANKL-induced osteoclast formation when added during the early stage of bone marrow macrophage (BMM) cultures, suggesting that it acts on osteoclast precursors to inhibit RANKL/RANK signaling. Among the RANK signaling pathways, coptisine inhibited NF-κB p65 phosphorylations, which are regulated in response to RANKL in BMMs. Coptisine also inhibited the RANKL-induced expression of NFATc1, which is a key transcription factor. In addition, 10 μM coptisine significantly inhibited both the survival of mature osteoclasts and their pit-forming activity in cocultures. Thus, coptisine has potential for the treatment or prevention of several bone diseases characterized by excessive bone destruction. [ABSTRACT FROM AUTHOR]

dipose tissue plays an essential role in energy homeostasis as a metabolic and endocrine organ. Accordingly, adipocytes are emerging as a major drug target for obesity and obesity-mediated metabolic syndrome. Dysfunction of enlarged adipocytes in obesity is involved in obesity-mediated metabolic syndrome. Adipocytokines, such as adiponectin released from small adipocytes, are able to prevent these disorders. In this study, we found that honokiol, an ingredient of Magnolia officinalis used in traditional Chinese and Japanese medicines, enhanced adipocyte differentiation in 3T3-L1 preadipocytes. Oil Red O staining showed that treatment with honokiol in the presence of insulin dose-dependently increased lipid accumulation in 3T3-L1 preadipoyctes although its activity was weak compared with rosiglitazone. During adipocyte differentiation, the expression of peroxisome proliferator-activated receptor γ2 (PPARγ2) mRNA and PPARγ target genes such as adipocyte protein 2 (aP2), adiponectin, and GLUT4 was induced by treatment with 10 μM honokiol. However, honokiol failed to show direct binding to the PPARγ ligand-binding domain in vitro. In preadipocytes, treatment with honokiol in the presence of insulin increased the phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 protein and Akt protein, early insulin signaling pathways related to adipocyte differentiation, compared with insulin-only treatment. Taken together, our results suggest that honokiol promotes adipocyte differentiation through increased expression of PPARγ2 mRNA and potentiation of insulin signaling pathways such as the Ras/ERK1/2 and phosphoinositide-3-kinase (PI3K)/Akt signaling pathways. [ABSTRACT FROM AUTHOR]

To effectively treat degenerative joint diseases including osteoarthritis (OA), small chemical compounds need to be developed that can potently induce chondrogenic differentiation without promoting terminal differentiation. For this purpose, we screened natural and synthetic compound libraries using a Col2GFP-ATDC5 system and identified oxytetracycline (Oxy) as a chondrogenic compound. Oxy induced cartilaginous matrix synthesis and mRNA expressions of chondrocyte markers in ATDC5 cells. In addition, Oxy suppressed mineralization and mRNA expressions of terminal chondrocyte differentiation markers in ATDC5 cells, primary chondrocytes, and cultured metatarsal bones. Oxy's induction of Col2 mRNA expression was decreased by the addition of Noggin and was increased by the addition of BMP2. Furthermore, Oxy increased mRNA expression of Id1, Bmp2, Bmp4, and Bmp6. These data suggest that Oxy induces chondrogenic differentiation in a BMP-dependent manner and suppresses terminal differentiation. Oxy may be useful for treatment of OA and also for regeneration of cartilage tissue. [ABSTRACT FROM AUTHOR]

The present study examined the anti-obesity effect and mechanism of action of Korean white ginseng extracts (KGE) using high-fat diet (HFD)-induced obese mice. Mice were fed a low-fat diet (LFD), HFD or HFD containing 0.8 and 1.6% (w/w) KGE diet (HFD + 0.8KGE and HFD + 1.6KGE) for 8 weeks. We also examined the effects of KGE on plasma triglyceride (TG) elevation in mice administrated with oral lipid emulsion. Body weight gain and white adipose tissue (WAT) weight were significantly decreased in the HFD + 1.6KGE group, compared with the HFD group. The plasma TG levels were also significantly reduced in both HFD + 0.8KGE and HFD + 1.6KGE groups, while leptin levels were significantly decreased in only the HFD + 1.6KGE group, compared with the HFD group. The HFD + 1.6KGE group showed significantly lower mRNA levels of lipogenesis-related genes, including peroxisome proliferator-activated receptorγ2 (PPARγ2), sterol regulatory element binding protein-1c (SREBP-1c), lipoprotein lipase (LPL), fatty acid synthase (FAS) and diacylglycerol acyltransferase 1 (DGAT1), compared with the HFD group. In addition, a dose of 1000 mg/kg KGE inhibited the elevation of plasma TG levels compared with mice given the lipid emulsion alone. These results suggest that the anti-obesity effects of KGE may be elicited by regulating expression of lipogenesis-related genes in WAT and by delaying intestinal fat absorption. [ABSTRACT FROM AUTHOR]

Anoectochilus formosanus, a plant native to Taiwan, is used as a folk medicine. It was found that oral administration of A. formosanus extract (AFE) (500 mg/kg) for 4 weeks suppressed bone weight loss and trabecular bone loss in ovariectomized mice, an experimental model of osteoporosis. Although AFE at 12.5 and 25 mug/ml inhibited osteoclast formation in co-culture of osteoblasts and bone marrow cells, AFE did not inhibit the formation of osteoclast progenitor cells and preosteoclast cells in bone marrow cells and RAW264 cells. However, AFE (at 12.5 and 25 microg/ml) decreased RANKL expression. These results suggested that AFE might suppress the bone loss caused by estrogen deficiency through suppression of RANKL expression required for osteoclast formation. [ABSTRACT FROM AUTHOR]

Although the mouse bone marrow stromal cell line ST2 has been known to be differentiated into osteoblasts, the differentiation characteristics of the cell into adipocyte and the concerned relationship between its adipogenesis and osteogenesis remains unknown. The adipogenic induction medium which is made up of insulin, dexamethasone (DEX) and 3-isobutyl-1-methylxanthine(IBMX), stimulated the expression of n early adipogenic marker PPAR γ and a late marker GPDH in ST2 cells. The triglyceride accumulation and lipid stain level generated by the induction medium in ST2 cells was inhibited by RA with IC50 at about 1 nM. The induction medium up-regulated expression of PPARγ and GPDH was also inhibited by RA whereas RA (30 nM) exterted no effect on the cell growth. Interestingly, treatment of the cells with induction medium in the presense of RA caused a 3- or 10-fold higher in ALP activity respectively as compared to those treated with RA or the induction medium alone. RT-PCR analysis showed that such a synergistic effect of RA and the induction medium paralleled the process of inhibition on adipogenesis. Additional experiments showed that IBMX played a key role in increasing the effect of RA and ALP activity. Our results suggested that the relationship between adipogenesis and osteogenesis in ST2 cells was reciprocally interrelated and the process of adipogenesis could be potentially reversed into an osteoblastogenic tendency. This is the first report demonstrating that RA transforms adipogenic potential into an osteoblastic tendency. [ABSTRACT FROM AUTHOR]

The osteoclasts are bone-resorbing multinucleatedcells formed by the fusion of mononuclearpreosteoclasts (pOCs) of hematopoietic origin.Although receptor activator of NF-κBligand (RANKL) has been shown to regulate osteoclastdifferentiation and function, its effect on the fusionof pOCs into multinucleated osteoclast-like cells(OCLs) has not been known. Using our fusion assaysystem, that is not contaminated with multinucleatedcells (MNCs) and osteoblastic cells, we determined theeffect of RANKL on the fusion of pOCs into MNCs. WhenpOCs were cultured on the plates, most of pOCs diedand disappeared from the plates within 24 h in theabsence of additives, but pOCs were fused to MNCswithin 6 h in the presence of RANKL. RANKL-inducedMNCs showed typical properties of OCL such astartrate-resistant acid phosphatase (TRAP) activity,actin ring formation, and bone-resorbing activity. Thefusion of pOCs into OCLs induced by osteoblastic cellsor RANKL was inhibited by OPG/OCIF, but that inducedby IL-1β was not. Both RANKL- andIL-1β-induced OCL formation from pOCs wasinhibited by ZLLL-H, a peptide inhibitor ofproteasome. These findings indicate that RANKLsupports the survival of pOCs and induces the fusionof pOCs into OCLs and suggest that NF-κBactivation is involved in these processes induced byRANKL and IL-1β. [ABSTRACT FROM AUTHOR]

Fusion and activation of osteoclasts are the final two events in osteoclastic bone resorption. To investigate the regulatory mechanism of these events, mononuclear osteoclasts (preosteoclasts, pOCs) were isolated from co-cultures of mouse osteoblastic cells and bone marrow cells. Most of the pOCs cultured without any additives died within 12 h. Survival of pOCs was supported by addition of either osteoblastic cells or macrophage-colony-stimulating factor (M-CSF). pOCs began to fuse with each other after culture for 12 h in the presence of osteoblastic cells or M-CSF. However, the properties of multinucleated osteoclast-like cells (OCLs) induced by osteoblastic cells were considerably different from those induced by M-CSF. Fusion of pOCs induced by osteoblastic cells was retarded after culture for 24 h. In contrast, M-CSF-induced fusion of pOCs continued throughout the 48-h culture period, which was not inhibited by addition of calcitonin. When pOCs together with osteoblastic cells were cultured for 48 h on dentine slices, many resorption pits were formed on the slices. Calcitonin completely inhibited the fusion and pit-forming activity of pOCs treated with osteoblastic cells. Resorption pits were hardly detected on dentine slices in pOC cultures treated with M-CSF. Osteoblastic cells prepared from osteopetrotic ( op/op) mice, which cannot produce functional M-CSF, stimulated the fusion and pit-forming activity of pOCs. Recombinant RANKL (receptor activator of NF-κB ligand), a cytokine which is produced by osteoblastic cells and is responsible for osteoclast differentiation, induced the fusion and pit-forming activity of pOCs. These results suggested that osteoblastic cells are involved in fusion and activation of osteoclasts through a mechanism independent of M-CSF production. RANKL appears to be responsible for fusion and activation of osteoclasts induced by osteoblastic cells. [ABSTRACT FROM AUTHOR]

In the course of screening for immunomodulators, we found a significant blastogenic activity specific for splenic B cells in the extracts of safflower (Carthamus tinctorius L.). Active fractions termed SF1 and SF2 were purified from dried petals of safflower by boiling water extraction, ethanol precipitation and Sepharose CL-2B column chromatography. The elution profiles of the gel filtration indicated that the molecular weight of SF1 and SF2 was estimated to be more than 100 kD. Major components of SF1 and SF2 seem to be polysaccharides, and structural analysis of alditol acetate derivatives by GC-MS revealed some differences between SF1 and SF2 in the sugar component. Biological activities of SF1 and SF2 on B cells and macrophages were examined in comparison with lipopolysaccharides (LPS). SF1 and SF2 induced both the proliferation and the IgM production of B cells to the equivalent level as those induced by LPS. In macrophages, SF1 and SF2 effectively stimulated the production of NO. However, SF1 stimulated the production of IL-1, IL-6, and TNF as much as LPS, while SF2 induced them only weakly or not at all. Thus, these results suggest that SF1 and SF2 activate B cells and macrophages in different mechanisms. [ABSTRACT FROM AUTHOR]

The osteoclasts, which play an essential role in bone resorption, are multinucleated cells (MNCs). They are formed by the fusion of mononuclear preosteoclasts (pOCs). However, because of the difficulty of isolating the pOCs, the process of fusion of pOCs has not been elucidated. To establish a fusion assay system, we succeeded in isolating pOCs that are not contaminated with MNCs and osteoblastic cells from the coculture of mouse bone marrow cells and osteoblastic cells. When pOCs were cultured in the presence of osteoblastic cells, the fusion of pOCs into MNCs took place within 24 h. No MNCs were formed in the absence of osteoblastic cells. The number of MNCs formed by the fusion of pOCs was dependent on the number of both pOCs and osteoblastic cells in the culture. Osteoblastic cells were also required for bone resorption by the osteoclasts that were formed by the fusion of pOCs. Osteotropic hormones, such as 1α,25-dihydroxy vitamin D3 [1α,25(OH)2D3] and parathyroid hormone (PTH), did not affect the fusion and pit formation of pOCs in the absence of osteoblastic cells. Eel calcitonin (eCT), however, significantly inhibited the fusion of pOCs induced by osteoblastic cells. These results suggest that the fusion of pOCs was induced by the direct contact between pOCs and osteoblastic cells.. [ABSTRACT FROM AUTHOR]