Bilirubin reversibly affects cell death and odontogenic capacity in stem cells from human exfoliated deciduous teeth.

OBJECTIVE: Hyperbilirubinemia in patients with biliary atresia causes deciduous tooth injuries such as green pigmentation and dentin hypoplasia. In patients with biliary atresia who received liver transplantation, tooth structure appears to be recovered radiographically. Nevertheless, little is known about cellular mechanisms underlying bilirubin-induced damage and suppression of deciduous tooth formation. In this study, we examined the effects of bilirubin in stem cells from human exfoliated deciduous teeth (SHED) in vitro. MATERIALS AND METHODS: SHED were cultured under exposure to excess of bilirubin and then interruption of bilirubin stimulation. RESULTS: Bilirubin induced cell death and inhibited the odontogenic capacity of SHED by suppressing AKT and extracellular signal-regulated kinase 1 and 2 (ERK1/2) pathways and enhancing nuclear factor kappa B p65 (NF-κB p65) pathway. The interruption of bilirubin stimulation reduced cell death and recovered the inhibited odontogenic capacity of bilirubin-damaged SHED. The bilirubin interruption also normalized the impaired AKT, ERK1/2, and NF-κB p65 signaling pathways. CONCLUSION: These findings suggest that tooth hypodontia in patients with hyperbilirubinemia might be due to bilirubin-induced cell death and dentinogenic dysfunction of odontogenic stem cells via AKT, ERK1/2, and NF-κB pathways and also suggested that bilirubin-induced impairments in odontogenic stem cells were reversible when bilirubin stimulation is interrupted.

Immune therapeutic potential of stem cells from human supernumerary teeth.

Discoveries of immunomodulatory functions in mesenchymal stem cells (MSCs) have suggested that they might have therapeutic utility in treating immune diseases. Recently, a novel MSC population was identified from dental pulp of human supernumerary teeth, and its multipotency characterized. Herein, we first examined the in vitro and in vivo immunomodulatory functions of human supernumerary tooth-derived stem cells (SNTSCs). SNTSCs suppressed not only the viability of T-cells, but also the differentiation of interleukin 17 (IL-17)-secreting helper T (Th17)-cells in in vitro co-culture experiments. In addition, systemic SNTSC transplantation ameliorated the shortened lifespan and elevated serum autoantibodies and nephritis-like renal dysfunction in systemic lupus erythematosus (SLE) model MRL/lpr mice. SNTSC transplantation also suppressed in vivo increased levels of peripheral Th17 cells and IL-17, as well as ex vivo differentiation of Th17 cells in MRL/lpr mice. Adoptive transfer experiments demonstrated that SNTSC-transplanted MRL/lpr mouse-derived T-cell-adopted immunocompromised mice showed a longer lifespan in comparison with non-transplanted MRL/lpr mouse-derived T-cell-adopted immunocompromised mice, indicating that SNTSC transplantation suppresses the hyper-immune condition of MRL/lpr mice through suppressing T-cells. Analysis of these data suggests that SNTSCs are a promising MSC source for cell-based therapy for immune diseases such as SLE.

Platelet-rich plasma suppresses osteoclastogenesis by promoting the secretion of osteoprotegerin.

BACKGROUND AND OBJECTIVE: Platelet-rich plasma is characterized by containing fundamental protein growth factors. Although many in vitro studies have documented the capability of platelet-rich plasma to induce the growth of osteoblasts or osteoblast-like cells, the effect of platelet-rich plasma on osteoclastogenesis has not yet been studied. The aim of the present study was to evaluate the effects of platelet-rich plasma and platelet-poor plasma on osteoclastogenesis with rat bone marrow cell culture. MATERIAL AND METHODS: Platelet-rich plasma and platelet-poor plasma were produced from the whole blood of rat. For cell culture, rat bone marrow cells were isolated from rat tibiae and then treated with 1,25alpha dihydroxy vitamin D(3) and with different concentrations of platelet-rich plasma or platelet-poor plasma. After 4 d of culture, rat bone marrow cells were stained with tartrate-resistant acid phosphatase (TRAP), and TRAP-positive cells that had more than three nuclei (TRAP-positive multinucleated cells) were counted as osteoclast-like cells. Osteoprotegerin, known as an osteoclastogenesis-related factor, cells was quantified using an enzyme-linked immunosorbent assay (ELISA). RESULTS: Although platelet-poor plasma had no effect on the formation of TRAP-positive multinucleated cells, platelet-rich plasma decreased the number of TRAP-positive multinucleated cells in a dose-dependent manner. The amount of osteoprotegerin produced from rat bone marrow cells and from MC3T3-E1 cells was enhanced in platelet-rich plasma-treated groups. CONCLUSION: Under our experimental conditions, platelet-rich plasma decreased the formation of TRAP-positive multinucleated cells and increased the secretion of osteoprotegerin. This study suggests that platelet-rich plasma suppresses osteoclastogenesis, therefore inhibiting bone resorption. In addition we also demonstrated that platelet-rich plasma increased the secretion of osteoprotegerin, an inhibitor for osteoclast formation, thus suggesting that the enhancement of osteoprotegerin secretion induces this inhibitory effect.

Differential-phase-shift quantum secret sharing.

A quantum secret sharing (QSS) protocol based on a differential-phase-shift scheme is proposed, which quantum mechanically provides a full secret key to one party and partial keys to two other parties. A weak coherent pulse train is utilized instead of individual photons as in conventional schemes. Compared with previous QSS protocols, the present one features a simple setup, is suitable for fiber transmission, and offers the possibility for a high key creation rate. An experiment is also carried out to demonstrate the operation.

Fibroblasts from the inner granulation tissue of the pseudocapsule in hips at revision arthroplasty induce osteoclast differentiation, as do stromal cells.

BACKGROUND: It has previously been shown that many osteoclast precursors are included in the granulation tissue within the pseudocapsule obtained at revision arthroplasty from hips with osteolysis. In vitro culture of only cells isolated from the granulation tissue has been previously shown to generate many mature osteoclasts. OBJECTIVE: To investigate the presence or otherwise of supporting cells, similar to stromal cells, which differentiate osteoclasts within the granulation tissue. METHODS: Cells isolated from the granulation tissue were cultured alone, and after four weeks fibroblast-like cells (granulation fibroblasts) remained. Rat non-adherent bone marrow cells (NA-BMCs) were co-cultured with the granulation fibroblasts with or without 1alpha,25(OH)2D3 (10(-8) M) or heat treated ROS 17/2.8 cell conditioned medium (ht ROSCM), or both. Multinucleated cells (MNCs), which formed, were assessed by biochemical and functional characterisation of osteoclasts. Receptor activator of NFkappaB ligand (RANKL) was investigated by immunohistochemistry. RESULTS: Co-culture of NA-BMCs and granulation fibroblasts caused the formation of tartrate resistant acid phosphatase (TRAP) positive MNCs, which had the calcitonin receptor (CTR), the Kat-1 antigen, which is specific to the surface of rat osteoclasts, and the ability to form pits in the presence of both 1alpha,25(OH)2D3 and ht ROSCM or in the presence of just ht ROSCM. RANKL was detected in fibroblast-like cells in the granulation tissue. CONCLUSION: These data suggest that granulation fibroblasts support osteoclast differentiation, as do osteoblasts/stromal cells, and may play a part in aseptic loosening.

Adult T-cell leukemia with anti-HTLV-I antibody but no HTLV-I DNA in tumor cells.

Adult T-cell leukemia (ATL) is usually defined as a malignant disease of T cells infected by human T-lymphotropic virus type I (HTLV-I). In the present study, we describe a 49-year-old woman with an acute type ATL, whose leukemic cells do not contain the HTLV-I genome. Laboratory tests revealed an increase in abnormal lymphocytes with convoluted nuclei, elevated serum lactate dehydrogenase levels, increased thymidine kinase activity and soluble interleukin-2 receptor-alpha levels. Serum examination demonstrated positive anti-HTLV-I antibody, but Southern blot analysis using the whole HTLV-I genome as a probe did not detect any integration of the viral genome. In contrast, PCR detected the HTLV-I pX region in the same DNA samples as used for Southern blot analysis. These findings suggest two possibilities. One possibility is that ATL in this patient is generated by other pathogens than HTLV-I virus. She is also an HTLV-I carrier. The other possibility is that her leukemic T cell clone derived its malignant phenotype from HTLV-I infection, and once this malignant phenotype was obtained, partial deletions of viral genome repeated until the whole viral genome was deleted. Although there is no direct evidence, the former possibility is more likely in the present case.

Osteoclast differentiation antigen, distinct from receptor activator of nuclear factor kappa B, is involved in osteoclastogenesis under calcitonin-regulated conditions.

Although calcitonin has been clinically utilized as a primary treatment for several metabolic bone diseases, its inhibitory effects against osteoclastic function diminish after several days owing to the calcitonin 'escape phenomenon'. We have previously found a unique cell-surface antigen (Kat1-antigen) expressed on rat osteoclasts. Here we show evidence that, in the presence of calcitonin, the Kat1-antigen is involved in osteoclastogenesis. Treatment of bone marrow cultures for forming osteoclast-like cells with anti-Kat1-antigen monoclonal antibody (mAb Kat1) provoked a marked stimulation of osteoclast-like cell formation only in the presence of calcitonin but not in its absence. Osteoclastogenesis stimulated by the receptor activator of nuclear factor kappa B (NF-kappaB) ligand/osteoclast differentiation factor was further augmented by mAb Kat1 in the presence of calcitonin. Furthermore, even in the presence of the osteoprotegerin/osteoclast inhibitory factor, mAb Kat1 induced osteoclast-like cell formation. Our current data suggest that the Kat1-antigen is a molecule that is distinct from receptor activator of NF-kappaB. The presence of the unique Kat1-antigen on cells in the osteoclast lineage appears to contribute to the fine regulation of osteoclastogenesis in vivo. Expression of this cell-surface molecule in cells in the osteoclast lineage may partly explain the mechanism responsible for the escape phenomenon.

Kat1-antigen--a reliable immunological marker for identifying osteoclast precursors of rats: detection of subpopulations among precursors and initiation of osteoclastogenesis.

Previously we found a unique cell surface antigen [Kat1-antigen (Kat1-Ag)] expressed on rat osteoclasts. In the present study, we focused on the expression of this antigen in preosteoclasts, mononuclear precursors of osteoclasts. Immunohistochemical and immunoelectron microscopic observations of the Kat1-Ag expressed in vivo showed the antigen to be present on mononuclear cells having the morphological features of preosteoclasts. The relationship between Kat1-Ag expression and calcitonin receptor (CTR) expression was examined in detail by a double-detection technique for CTR and Kat1-Ag by use of autoradiography and immunocytochemistry, respectively. In a culture system for forming mononuclear preosteoclast-like cells, almost 100% of the mononuclear cells expressing Kat1-Ag also expressed CTR, demonstrating that Kat1-Ag is a reliable immunological marker for identifying preosteoclasts. Interestingly, a significant number of the CTR-positive mononuclear cells did not express the Kat1-Ag. Detection of these cells expressing CTR but not Kat1-Ag strongly suggests the presence of subpopulations in preosteoclasts. We also obtained evidence suggesting that expression of the Kat1-Ag is initiated during the postmitotic stage of the osteoclast progenitors.

Tumor necrosis factor-alpha cooperates with receptor activator of nuclear factor kappaB ligand in generation of osteoclasts in stromal cell-depleted rat bone marrow cell culture.

A member of the tumor necrosis factor (TNF) family, receptor activator of nuclear factor kappaB ligand (RANKL; also known as ODF, OPGL, and TRANCE), plays critical roles in osteoclast differentiation and activation in the presence of macrophage colony-stimulating factor (M-CSF). Recently, TNF-alpha has also been shown to induce the formation of multinucleated osteoclast-like cells (MNCs) in the presence of M-CSF from mouse macrophages. We demonstrated that mononuclear preosteoclast-like cells (POCs) were formed in the presence of conditioned medium of osteoblastic cells in a rat bone marrow culture depleted of stromal cells. Using this culture system, in this study we examined whether TNF-alpha affects differentiation into POCs from hematopoietic progenitor cells. Human TNF-alpha (hTNF-alpha) markedly stimulated the formation of POCs. Moreover, a concentration as low as 0.005 ng/mL of hTNF-alpha increased the level of mRNA for calcitonin receptor (CTR) and cathepsin-K of POCs. The POCs induced by hTNF-alpha formed MNCs, which showed dentine-resorbing activity after coculture with primary osteoblasts. Stimulation was observed after 24 h of treatment with hTNF-alpha only on day 1 or day 2 of the culture. After 24 h of hTNF-alpha treatment, expression of the receptor activator of nuclear factor kappaB (RANK) mRNA was markedly increased. The addition of soluble RANKL (sRANKL) to the preformed POCs efficiently induced MNCs. Interestingly, treatment of bone marrow cells with hTNF-alpha and sRANKL synergistically augmented the formation of MNCs. This formation was abolished by the addition of human osteoprotegerin (hOPG). These results suggest that cooperation of TNF-alpha and RANKL is important for osteoclastogenesis.

The significance of cytomegalovirus infection over the clinical course of adult T-cell leukemia/lymphoma.

The significance of cytomegalovirus (CMV) infections developed over the clinical course of adult T-cell leukemia/lymphoma (ATLL) were evaluated in relation to the patient survival rate, ATL activity and immunocompetent cells. ATLL patients with CMV infections on admission exhibited a poor survival rate, while patients with CMV infections at any time after admission survived longer than those not infected with this virus. ATLL patients who exhibited a numbers of CMV infection on admission showed higher ATL activity and had lower numbers of CD8-positive and CD56-positive cells than those who developed CMV infections at any time after admission. Therefore, it appears likely that patients with CMV infections on admission were in an immunosuppressive state due to aggressive ATL activity.

Granulocyte colony-stimulating factor production by adult T-cell leukaemia cells.

We previously demonstrated that, in about 30% of primary adult T-cell leukaemia (ATL) cases, the leukaemic cells proliferated in response to granulocyte colony-stimulating factor (G-CSF). In the present report, we describe five patients with the acute leukaemia type of ATL who showed marked neutrophilia and elevated serum G-CSF concentrations in the absence of infection. We further examined two of these patients for detailed clinical features and cellular characteristics of the tumour cells. The white blood cell counts of both patients were 62 x 10(9)/l, consisting of approximately 90% neutrophils and 10% ATL cells. Serum concentrations of G-CSF in the two patients were 138 pg/ml and 93 pg/ml. The G-CSF concentrations in supernatants of short-term cultures of the patients' peripheral blood T-cells were 2 5 pg/ml and 13 pg/ml respectively. Immunostaining with anti-G-CSF antibody demonstrated G-CSF production by primary ATL cells in both cases. The neutrophil count fluctuated simultaneously with activity of ATL. Primary ATL cells from one patient were shown to proliferate in response to G-CSF in vitro. These results suggest autocrine growth stimulation of primary ATL cells in a subgroup of patients.

Cytomegalovirus infection is not necessarily a poor prognostic factor in adult T-cell leukemia/lymphoma.

The relationship between cytomegalovirus (CMV) antigenemia and the clinical course was examined in 57 patients with adult T-cell leukemia/lymphoma (ATLL). All patients included had the acute/lymphoma type of ATL according to the criteria of the Japan Lymphoma Study Group (LSG). CMV antigenemia was assessed on admission and at the time when the patients had fever higher than 37. 5 degrees C, which did not respond to antibiotics for longer than 3 days. The incidence of CMV antigenemia was 44%. Approximately 90% of patients with CMV antigenemia died of infections with viruses, bacteria, and/or fungi, while approximately 40% of patients without CMV antigenemia died of deterioration of ATLL (P<0.0001). In this study, the patients with CMV antigenemia tended to survive longer than those negative for it (321.4 days vs. 266.2 days), although there was no statistical significance (P=0.09). Kaplan-Meier analysis revealed that CMV antigenemia was not a poor prognostic factor. When the disease status of ATLL was evaluated by thymidine kinase (TK) and soluble interleukin 2 receptor (sIL-2R), both had lower titers during CMV antigenemia (TK: P=0.01, sIL-2R: P=0.03, respectively). Therefore, CMV infections in ATLL patients seemed to have bimodal meanings; CMV infection at the end of clinical course were life-threatening, but infection during the first half of clinical course seemed to suppress ATLL activity and to contribute to the longer survival of the patients.

Long-term maintenance combination chemotherapy with OPEC/MPEC (vincristine or methotrexate, prednisolone, etoposide and cyclophosphamide) or with daily oral etoposide and prednisolone can improve survival and quality of life in adult T-cell leukemia/lymphoma.

Acute leukemia and lymphoma varieties of adult T-cell leukemia/lymphoma (ATL) usually carry a poor prognosis. While etoposide is generally useful for treating ATL, especially as a daily oral maintenance regimen, etoposide has not proven effective in severe types of ATL efficient in some patients. Of 87 ATL patients whom we have treated, 51 had acute leukemia, 22 lymphoma and 14 progressive chronic leukemia. Seventy-nine patients were treated with a long term maintenance combination protocol, OPEC/MPEC (weekly doses of vincristine, 0.7 mg/m2 or methotrexate, 14 mg/m2; prednisolone, 20 mg/m2; etoposide, 70 mg/m2 and cyclophosphamide, 200 mg/m2). The other 8 patients, 3 with acute leukemia, 2 with lymphoma and 3 with progressive chronic leukemia, were treated with daily oral administration of 25 mg of etoposide and 10 mg of prednisolone (DOEP). The dose administered was modified in individual cases to maintain the granulocyte count and reduce the number of ATL cells. Considering both protocols, a complete response and a partial response were achieved in 31.0% and 58.6% patients, respectively. Median survival times (MST) of all patients and, acute leukemia, lymphoma and progressive chronic leukemia types were 7.5, 6.7, 9.6 and 12.4 months, respectively. Respective MST of patients treated with OPEC/MPEC or DOEP protocols were 7.1 and 18.0 months. Relatively normal WBC counts, lower lactate dehydrogenase concentration and normal calcium concentration, limited numbers of anatomic sites involved, good performance status and good response to chemotherapy were significantly associated with long survival time. Drug toxicity was not apparent, and about half of patients were treated in an outpatient setting.

Osteoclast-derived zinc finger (OCZF) protein with POZ domain, a possible transcriptional repressor, is involved in osteoclastogenesis.

The differentiation of osteoclasts is regulated by transcription factors expressed in cells of osteoclast lineage. We isolated here a potential transcription factor from a cDNA library of an enriched population of preosteoclasts and osteoclasts. The cDNA encodes a protein with N-terminal POZ domain and C-terminal Krüppel-like zinc fingers. We designate this protein as osteoclast-derived zinc finger (OCZF). OCZF was found to be rat homologue of mouse leukemia/lymphoma-related factor (LRF). Northern blot and in situ hybridization analysis showed OCZF mRNA at a high level in osteoclasts and kidney cells. OCZF had a nuclear targeting sequence and was localized in the nucleus of transfected cells. In addition, OCZF specifically bound to the guanine-rich consensus sequences of Egr-1 and c-Krox. Transient transfection assays indicate that OCZF can repress transcription activity like other POZ domain proteins. Furthermore, antisense but not sense phosphorothioate oligodeoxynucleotides (ODNs) for OCZF cDNA suppressed the formation of osteoclast-like multinucleated cells (MNCs) in bone marrow culture, whereas the same ODNs did not significantly affect the formation of macrophage polykaryons and mononuclear preosteoclast-like cells (POCs). These results suggest that OCZF is a unique transcription factor that plays an important role in the late stage of osteoclastogenesis.

NF-kappaB involvement in the activation of primary adult T-cell leukemia cells and its clinical implications.

The HTLV-I provirus-encoded Tax protein induces NF-kappaB in Tax-transfected Jurkat T cells or HTLVL-I- infected T cells in vitro. Tax induction of NF-kappaB is presumed to be involved in proliferation and activation of primary leukemia cells in vivo. Recent studies have demonstrated that NF-kappaB activities in human T cells are mediated by at least four c-Rel-related DNA binding proteins - p50, p55, p75 and p85. We examined the significance of NF-kappaB induction in primary adult T cell leukemia cells and the induction kinetics of each of the four NF-kappaB species. Marked NF-kappaB activity was detected using an electrophoretic mobility shift assay (EMSA) in the primary cells of patients with acute disease, but little activity was noted in the cells of chronic patients. NF-kappaB activity was enhanced in a time-dependent manner in acute type cells cultured with mitogen-free medium; there was no induction of activity in chronic type cells. UV crosslinking demonstrated all four species of NFkappaB complex - high levels of p50 and lower levels of p55 and p75, in acute type cells; chronic type cells showed only the p50. As a control, normal resting T cells similarly showed only p50; control cells showed little change in activity when cultured without mitogenic stimulation, analogous to chronic type ATL. Northern blotting revealed enhancement of c-rel (encoding p85) and KBFI (encoding p50 and p55) expression in acute type cells during culture, while there was no significant enhancement of mRNAs in chronic type ATL cells or unstimulated normal T cells. Northern blotting also revealed that Tax is upregulated at the mRNA level in acute- but not chronic-type cells during culture. Expression of c-rel and KBF1 mRNAs in acute type cells appeared to be related to Tax mRNA expression. These results suggest that Tax is capable of inducing nuclear expression of all four NF-kappaB species in primary ATL cells of acute type patients, with marked effects on p55, p75, and p85. Tax induction of NF-kappaB species is regulated, at least in part, at a pretranslational level involving increases in c-rel and KBF1 mRNA.

Spontaneous regression associated with apoptosis in a patient with acute-type adult T-cell leukemia.

We describe a 76-year-old man with acute-type adult T-cell leukemia, who demonstrated a spontaneous decrease in leukemic cell number, apparently coincident with apoptotic cell death. On admission the patient's white blood cell count was 38.9 x 10(9)/l with 77% abnormal lymphocytes. He also had hypoproteinemia (4.3 g/dl) from protein losing enteropathy. After admission the leukemic cell count decreased without chemotherapy, reaching 5.9 x 10(9)/l after 2 months. Studies of peripheral lymphocytes demonstrated appearance of the apoptotic cells and DNA ladder formation from the beginning of regression. Same truncated proviral DNA was recognized in primary ATL cells through the whole clinical course. The hypoproteinemia improved with intravenous nutrition, followed by increase of the leukemic cells. This case is the first report that demonstrates tumor-cell apoptosis induced clinical regression in adult T-cell leukemia. Further, we speculate that the hypoproteinemia may have been involved in the leukemic cell apoptosis.

A novel role of IL-15 in the development of osteoclasts: inability to replace its activity with IL-2.

IL-15 shares many activities with IL-2 on stimulating lymphocytes, hematopoietic progenitor cells, and macrophages. However, the role of IL-15 in osteoclastogenesis has not been elucidated. The recent finding of abundant IL-15 in rheumatoid arthritis synovial fluids suggested a possible role for this cytokine in the pathological destruction of bone and prompted us to determine whether IL-15 stimulates osteoclast formation. IL-15 stimulated the formation of multinucleated osteoclast-like cells in rat bone marrow cultures. In stroma-free cultures, IL-15 increased the number of mononuclear preosteoclast-like cells in the early stage of osteoclast formation. The stimulation was observed even after treatment with IL-15 for only 24 or 48 h of culture. Moreover, low IL-15 concentration (0.1 ng/ml) strongly increased the level of calcitonin receptor mRNA of mononuclear preosteoclast-like cells. Although IL-15 is known as a potent stimulator of TNF-alpha, its activity was not abolished by addition of anti-TNF-alpha Ab. Interestingly, IL-2 and IL-7, which utilize some IL-15R components, had no effect on osteoclast differentiation, but pretreatment with IL-2 or IL-7 of bone marrow cells before the addition of IL-15 inhibited the enhancing activity of IL-15. In summary, IL-15 has a novel activity to stimulate the differentiation of osteoclast progenitors into preosteoclasts, which cannot be replaced by IL-2 but may use components in common with IL-2R to mediate its effects.

Fluid shear stress increases interleukin-11 expression in human osteoblast-like cells: its role in osteoclast induction.

It is unclear how mechanical stress influences bone cells. Mechanical stress causes fluid shear stress (FSS) in the bone. Osteoblast lineage cells are thought to sense FSS and regulate bone remodeling. We therefore investigated the effects of FSS on human osteoblast-like osteosarcoma cells: SaOS-2 cells in vitro. The conditioned medium of the SaOS-2 cells after 24 h of FSS (24 h-FSS CM) showed such osteoclastic phenotype inductions as significantly increasing the number of tartrate-resistant acid phosphatase (TRAP) positive multinuclear cells in rat bone marrow cells and TRAP-positive cells in human preosteoclastic cells: FLG 29.1 cells. An enzyme-linked immunosorbent assay showed interleukin-11 (IL-11) protein to increase 7-fold in the 24 h-FSS CM. A Northern analysis showed that IL-11 mRNA increased 4-fold in the SaOS-2 cells after 6 h-FSS; however, no IL-6 mRNA expression was detected. Furthermore, the anti-human IL-11 antibody significantly neutralized the osteoclastic phenotype induction of the 24 h-FSS CM. The IL-11 mRNA up-regulation in SaOS-2 cells by the 6 h-FSS was not inhibited by the anti-human transforming growth factor-beta1 antibody, but it was significantly inhibited by indomethacin. An enzymeimmunoassay showed prostaglandin E2 to increase 7-fold in the 1 h-FSS CM. These findings thus suggest that FSS induces osteoblasts to produce IL-11 (mediated by prostaglandins) and thus stimulates bone remodeling.

Localization of a novel inositol 1,4,5-trisphosphate binding protein, p130 in rat brain.

We have isolated a novel inositol 1,4,5-trisphosphate binding protein with molecular mass of 130 kDa (p130), homologous to phospholipase C-delta1 in amino acid sequence but with no catalytic activity. Here we report the expression and localization of p130 at the mRNA level in rat brain. Northern blotting showed that gene expression encoding p130 was most abundant in brain. Brain localization of p130-mRNA using an in situ hybridization technique revealed that in the cerebellum, the mRNA was detected in the granular cell and Purkinje cell layers, and cerebellar nuclei. In the cerebrum, the mRNA was localized in hippocampal pyramidal cells, dentate granule cells and pyramidal and/or granule cells of the cerebral cortex. The brain localization of p130-mRNA was similar to that of the beta-subtype of phospholipase C, indicating that p130 may be mainly involved in phospholipase Cbeta-mediated signaling.