Osteoclastogenesis inhibitory factor exhibits hypocalcemic effects in normal mice and in hypercalcemic nude mice carrying tumors associated with humoral hypercalcemia of malignancy

Osteoclastogenesis inhibitory factor (OCIF) is a novel secreted protein that inhibits osteoclastogenesis both in vitro and in vivo. In this study, we examined the effects of OCIF on serum calcium (Ca) concentrations in normal mice and in hypercalcemic nude mice carrying tumors associated with humoral hypercalcemia of malignancy. In normal mice, a single intraperitoneal injection of OCIF reduced serum Ca levels in a dose-dependent manner. Significant decrease in serum Ca (by 1.6 ± 0.3 mg/dL, n = 5) was observed 2 h after the injection of OCIF at 20 mg/kg and the hypocalcemic effect continued for up to 12 h. Serum phosphate (Pi) concentrations also decreased in response to OCIF. Urinary excretion of Ca, Pi, and creatinine did not change significantly after injection of OCIF or vehicle. In hypercalcemic, tumor-bearing nude mice, a single intraperitoneal injection of OCIF at 20 mg/kg resulted in a dramatic decrease in serum Ca (maximal decrease 2.8 ± 0.37 mg/dL, n = 11), which continued for up to 24 h. The results suggest that OCIF decreased serum Ca through its inhibitory effect on bone resorption. Furthermore, it is suggested that OCIF has therapeutic potential for the treatment of hypercalcemic conditions such as malignancy-associated hypercalcemia.     

Differential Expression of Subspecies of Polyomavirus and Murine Leukemia Virus Enhancer Core Binding Protein, PEBP2, in Various Hematopoietic Cells

The core sequence of the enhancer of murine leukemia virus (MuLV) long terminal repeat is highly conserved in a large number of MuLV strains and appears to play an essential role when SL3-3 or Moloney strains induce T cell lymphoma in mice. We found by using the electrophoretic mobility shift assay that a polyomavirus enhancer core-binding protein, PEBP2, bound to this core motif of MuLV. We also noted that PEBP2 in several hematopoietic cell lines derived from B lymphocyte, macrophage and myelocyte lineages migrated significantly faster than the authentic PEBP2 detected in NIH3T3 (ibroblasts. Interestingly, PEBP2 detected in the cell lines of T lymphocyte lineage appeared to contain both types, which were indistinguishable in electrophoretic mobility from those of NIH3T3 and of B lymphocyte, macrophage and myelocyte lineages. The treatment of the nuclear extract containing PEBP2 with phosphatase generated PEBP3, which is a subcomponent of PEBP2 and retained the same DNA-binding specificity as PEBP2. The altered mobility of hematopoietic cell-derived or T lymphocyte-derived PEBP2 was found to be due to the alteration of the mobility of PEBP3. Based on the distinct mobility of PEBP2/3 of T lymphocytes from those of other hematopoietic cells, we discuss the implication of PEBP2 in MuLV-induced T cell leukemia and T cell-specific gene expression.     

Mechanism of atrial natriuretic polypeptide and sodium nitroprusside-induced relaxation in guinea-pig tracheal smooth muscle

Atrial natriuretic polypeptide (ANP) is composed of a family of peptides isolated from rat and human atria. In the present study, the relaxant effects of ANP, sodium nitroprusside and 8-bromo-cyclic guanosine monophosphate (GMP) were investigated in guinea-pig tracheal smooth muscle, and the tissue cyclic GMP and cyclic adenosine monophosphate (AMP) concentrations were measured. ANP, sodium nitroprusside and 8-bromo-cyclic GMP showed relaxant effects on the spontaneous tone in normal Krebs solution (5.9 mmol/l K(+)-2.4 mmol/l Ca++ solution). They diminished relaxant effects on 40 mmol/l K(+)-0.1 mmol/l Ca++ induced contraction, which was approximately the same tension as the spontaneous tone. Sodium nitroprusside and 8-bromo-cyclic GMP diminished less relaxant effects on 40 mmol/l K(+)-2.4 mmol/l Ca++ induced contraction, but ANP showed no relaxation. The tissue cyclic GMP levels following administration of ANP and sodium nitroprusside in normal Krebs solution, in 40 mmol/l K(+)-2.4 mmol/l Ca++ solution, and in 40 mmol/l K(+)-0.1 mmol/l Ca++ solution increased dose-dependently without regard to external Ca++ concentrations, while the tissue cyclic AMP levels did not change. These results suggest that ANP might be a novel potent relaxant in airway smooth muscle and the relaxant effect may be, at least in part, mediated by cyclic GMP. There was a difference in relaxant effects on tracheal smooth muscle between ANP and sodium nitroprusside.     

Differential inhibitory effects of nitroglycerin on contractile responses to the alpha-adrenoceptor agonists, methoxamine and clonidine, in rabbit aorta

The vasoinhibitory action of nitroglycerin was examined on contractile responses to methoxamine and clonidine in isolated rabbit aorta. Nitroglycerin at 10(-5) M, but not 10(-6)-10(-8) M, shifted the concentration response curve for methoxamine to the right. Nitroglycerin (10(-8)-10(-5) M), however, noncompetitively inhibited responses to clonidine in a concentration dependent manner. Nitroglycerin (10(-5) M) had no effect on responses to potassium (10-70 mM), but slightly inhibited responses to Ca2+ (0.1-5 mM) in a Ca2+-free medium containing potassium. Nifedipine (10(-6) and 10(-5) M), however, almost abolished responses to both potassium and Ca2+ but had no effect on responses to either methoxamine or clonidine. Agonist-antagonist interactions using prazosin and yohimbine revealed that responses to both methoxamine and clonidine were due to activation of alpha 1-adrenoceptors. Results with phenoxybenzamine suggested that the aorta has more receptor reserve for methoxamine than for clonidine. Furthermore, in tissues pretreated with phenoxybenzamine, nitroglycerin (10(-5) M) inhibited the maximal contractile response to methoxamine (3 x 10(-4) M). The maximal response to clonidine in tissues pretreated with phenoxybenzamine was not affected by nitroglycerin (10(-8) M). Nitroglycerin (10(-9)-10(-4) M) had greater inhibitory effect on residual responses to clonidine (10(-5) M) than that to methoxamine (10(-5) M) in a Ca2+-free medium containing EGTA. The contractile responses to Ca2+ (2 mM) in a Ca2+-free medium containing EGTA, nifedipine, and either methoxamine (5 x 10(-7) M) or clonidine (3 x 10(-7) M) were inhibited by nitroglycerin (10(-9) - 10(-5) M). The effect of nitroglycerin was greater on responses in the presence of clonidine than methoxamine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mechanism of barium-induced contraction in the vascular smooth muscle of rabbit aorta.

In a solution containing 1.5 mM Ca2+, cumulative application of 0.3-10.0 mM Ba2+ induced a concentration-dependent contraction of the rabbit aorta. This contraction was reduced by the Ca2+ channel inhibitors, verapamil (10(-6) M), nifedipine (10(-7) M) and lanthanum (2.0 mM), and was potentiated by the Ca2+ channel facilitator, Bay K8644 (10(-7) M). In a Ca2+-free solution containing EGTA (1.0 mM), cumulative application of Ba2+ still induced a concentration-dependent contraction, the maximum contractile tension of which was comparable to that in the presence of 1.5 mM Ca2+. The Ba2+-induced contraction which was not dependent on the external Ca2+ was also inhibited by verapamil, nifedipine and lanthanum and was potentiated by Bay K8644. A high concentration (65.4 mM) of K+ potentiated this Ba2+-induced contraction whereas noradrenaline (10(-6) M) did not have such an effect. In order to deplete the releasable Ca2+ store in the cell, the muscle strip was treated with noradrenaline (10(-6) M) and/or caffeine (20.0 mM) in a Ca2+-free solution. In such a Ca2+-depleted muscle, Ba2+ still induced a contraction of a similar magnitude to that without such treatment. Further, the second application of Ba2+ in a Ca2+-free solution induced a similar contraction to that induced by the first application of Ba2+. These results suggest that Ba2+ depolarizes the cell membrane and opens the voltage-dependent Ca2+ channels resulting in a Ca2+ influx in the presence of Ca2+. In the absence of external Ca2+, Ba2+ may enter the cell through the voltage-dependent Ca2+ channels and induce contraction without mobilizing the Ca2+ store which is sensitive to noradrenaline and caffeine.     

Presence of ghost cells and the Wnt signaling pathway in odontomas

BACKGROUND: Although it has been reported that ghost cells are present in odontomas, the generation mechanism of these cells is unclear. To evaluate the presence of ghost cells and involvement of the Wnt signaling pathway, we examined the expression of hard keratins, beta-catenin and Lef-1 in odontomas. METHODS: Sixty-nine cases of odontoma were examined immunohistochemically with the use of antibodies against human hair proteins, beta-catenin and Lef-1. RESULTS: Expression of hard keratins was found only in the cytoplasm of ghost cells in 46 (66.7%) of the 69 odontomas. Compound odontomas (78.8%) showed a higher incidence of ghost cells than complex odontomas (29.4%). Histopathologically, ghost cells were found within odontogenic epithelium adjacent to immature enamel and in the centre of Liesegang-ring-like calcified materials. Expression of beta-catenin and Lef-1 was observed in the cytoplasm and nucleus of odontogenic epithelial cells adjacent to the ghost cells in immature odontomas. CONCLUSION: These findings suggest that odontoma is a hard keratin-expressing tumor-like lesion, and that the Wnt signaling pathway may be involved in the formation of ghost cells in odontomas.     

Diversity in DNA rearrangements and in RNA expressions of immunoglobulin gene on common variable immunodeficiency.

Six heterogeneous common variable immunodeficiency (CVID) patients were analysed for germ-line DNA, DNA rearrangements, and RNA expressions of immunoglobulin (Ig) gene by Southern or northern blotting using appropriate probes. We detected no polymorphism in neutrophil DNA hybridized to a C mu and a C gamma probe. In three patients, both serum Ig and Ig-bearing cells were scarcely detected, and by northern hybridization methods, neither mu mRNA, gamma mRNA, alpha mRNA nor kappa mRNA was detected. However, one Epstein-Barr virus-transformed B lymphoblastoid cell line (LCL) of these three patients was different from the germ line in the region of JH, C gamma, and C kappa, and expressed mu mRNA at a higher level. The B cell defects of these three patients lay on the B cell maturation stage similar to X-linked agammaglobulinaemia (XLA). In two others among the six CVID patients, serum IgM and IgM-bearing cells were detected to a certain degree, and by northern hybridization, mu mRNA was detected at a lower level, but neither mu mRNA, alpha mRNA, nor kappa mRNA was detected. One LCL of these two patients could express mu mRNA at the normal level. In the last patient, the serum IgM was normal, serum IgG and IgA were somewhat low, Ig-bearing cells were normal, mu mRNA and kappa mRNA were detected at the normal level, and gamma mRNA and alpha mRNA were detected at a lower level. The defect of this patient affected the class switch stage. These results showed that primary B cell defects in CVID occurred at several B cell differentiation stages which could be classified by expression of the Ig gene, and at the degree of clonal diversity in the B cell repertoire. Furthermore, this study provides support for the idea that the CVID defect is related to a more generalized cellular function, such as regulating the proliferation and/or clonal expansion of cells of the B lymphoid lineage.