Dentinal defects in Hyp mice not caused by hypophosphatemia alone.

The Hyp mouse is a murine homolog of human X-linked hypophosphatemic rickets and displays hypo-mineralization in bone and dentin due to a defect of the phosphate-regulating gene with homology to endopeptidase on the X chromosome (Phex) gene. It has long been considered that the bone and dentin defects in Hyp mice are caused by hypophosphatemia alone, however, several recent studies have indicated the possibility that intrinsic defects are present in Hyp mice osteoblasts. Further, we previously found a hyper-expression of osteocalcin (OC) mRNA in Hyp mouse odontoblasts and suggested the possibility of the presence of intrinsic defects. In the present study, we evaluated morphological features and OC mRNA expression levels in tooth germs of Nor mice with a normal phex gene and a low concentration of serum phosphate, and compared them to those in Hyp and wild-type mice. Nor mice exhibited low serum phosphate levels, however, did not show the characteristic features of dentin defects seen in Hyp mice, such as widened predentin and hyper-expression of OC mRNA. These results suggest that the hypo-mineralization of dentin in Hyp mice is not dependent on serum phosphate level, but rather is affected by intrinsic defects in odontoblasts.

Hyper-expression of osteocalcin mRNA in odontoblasts of Hyp mice.

The Hyp mouse is a murine homologue of human X-linked hypophosphatemia that displays hypo-mineralization in bone and dentin. In this study, we tested the hypothesis that the defect in Hyp mice leads to alterations in the expression of dentin matrix proteins that may be associated with the hypo-mineralization changes in the tissues. Quantitative RT-PCR analyses showed that expression of the osteocalcin gene in Hyp mice tooth germ samples was significantly higher than in wild-type mice, whereas the gene expressions of osteonectin, osteopontin, dentin matrix protein 1, and type I collagen in both types of mice were similar. Further, cultured Hyp mice tooth germ samples exhibited a higher expression of the osteocalcin gene than did those from wild-type mice, which was in accord with the results of our in vivo analysis. These findings suggest that osteocalcin mRNA is highly expressed in Hyp mice odontoblasts and may be associated with dentin hypo-mineralization.

Vascular endothelial growth factor and cellular chemotaxis: a possible autocrine pathway in adult T-cell leukemia cell invasion.

Our previous report (T. Hayashibara et al., Leukemia, 13: 1634-1635, 1999) revealed a possible link between high plasma vascular endothelial growth factor (VEGF) concentration and leukemic cell invasion in adult T-cell leukemia (ATL). However, the biological mechanism of this link has not been elucidated. The purpose of this study was to address that mechanism. Our present observations showed that VEGF mRNA was expressed in ATL cell lines. The corresponding protein was secreted into the extracellular environment, which suggested that the major source of plasma VEGF is ATL cells themselves. More interestingly, all of the cell lines examined were found to express the mRNA and protein for fms-like tyrosine kinase-1 (Flt-1), which is one of the receptors for VEGF. Cytofluorometric analysis demonstrated the VEGF binding potency of these cells. In clinical specimens, expression of VEGF and Flt-1 mRNAs was detected in all (100%) of 11 and 8 (73%) of 11 ATL patients, respectively. Cytofluorometric analysis revealed that VEGF effectively bound only to Flt-1-expressing cells. These findings are highly suggestive of an autocrine pathway involving VEGF operating in ATL. The proliferation of ATL cell lines was not affected by treatment with an anti-VEGF antibody or exogenous VEGF, which indicated that VEGF has no mitogenic effect on ATL cells. In contrast, we made the interesting finding that treatment with exogenous VEGF enhanced the chemotactic activities of some ATL cell lines, which may play a key role in ATL cell invasion. Collectively, these data lead us to propose a possible autocrine mechanism involving VEGF operating by way of Flt-1, in which ATL cells up-regulate their own chemotaxis to facilitate their invasion into various organs.

De novo acute myeloid leukemia in the elderly; a consistent fraction of long-term survivors by standard-dose chemotherapy.

To clarify the characteristics of de novo acute myeloid leukemia (AML) among the elderly, we reviewed 112 patients over 60 years old (median age 72 years) who were treated at hospitals in Nagasaki Prefecture with a population of 1.5 million between 1987 and 1994. Reclassification of morphological diagnosis revealed that the proportion of M3 was lower but that of M6 and the incidence of cases with trilineage dysplasia (TLD), known as poor prognostic features, were higher in the elderly than in patients less than 60 years old. Similarly, chromosomal data showed a lower frequency of favorable karyotypes such as t(8;21) and t(15;17) in the elderly. The overall survival of all 112 patients was 10.3% at 5 years. Multivariate analysis indicated that good performance status (PS), low WBC at diagnosis, standard dose multi-drug chemotherapy and all-trans retinoic acid (ATRA) treatment for M3 patients, and morphological findings without TLD were significantly correlated with longer survival. Most of the long-term survivors were found among those who received standard dose therapy in this series, although no consensus has been established how to treat elderly AML patients. We propose that a prospective controlled trial is necessary to confirm the role of standard dose chemotherapy for elderly patients with de novo AML.

Tooth eruption in a patient with craniometaphyseal dysplasia: case report.

Craniometaphyseal dysplasia (CMD) is a very rare genetic disorder of bone remodeling caused by osteoclast dysfunction. The clinical and radiographical features of oral findings are presented in a sporadic case of CMD in a child (age 10 years, 7 months). An intraoral examination showed severe malocclusions, including anterior crossbite and deep bite. Furthermore, a radiographic examination showed increased radiopacity of the maxilla and mandibular bones due to hyperostosis and sclerosis of the jaw. There was no root resorption of the canines or molars in the primary dentition, although root formation of the permanent teeth was proceeding. Dental age was calculated to be approximately 1 year, 4 months younger than his chronological age. The eruption speed of the permanent lateral incisors after the gingival emergence was shown to be within normal values, and we discuss whether the canines and premolars in the permanent dentition could erupt or not.

Lactacystin activates FLICE (caspase 8) protease and induces apoptosis in Fas-resistant adult T-cell leukemia cell lines.

Lactacystin (LC) is a specific inhibitor of the proteasome, and has recently been shown to induce apoptosis in certain cell lines. In the present study, we established Fas-resistant adult T-cell leukemia (ATL) cell subclones RSO4 and RST1 from their parental Fas-sensitive cell lines SO4 and ST1, and examined whether LC can overcome Fas resistance. LC completely inhibited proteasome function as determined by a peptidyl-MCA substrate (LLVY-MCA and LLE-MCA), and induced apoptosis in these cell lines irrespective of Fas sensitivity at low concentrations (approximately 10 microM). LC induced the activation of caspase 3 (CPP32/Yama) and caspase 6 proteases in an identical manner to Fas-mediated apoptosis. Moreover, LC induced the activation of caspase 8 (FLICE) protease, which is the initiator of the Fas-mediated apoptotic cascade. Synthesized proteasome inhibitory peptide MG-115 (ZLLnV-CHO) also induced apoptosis in these cell lines. These results indicated that proteasome inhibitors overcome Fas-resistance by bypassing the proximal part of the Fas signal. Inhibition of the proteasome function may be a new strategy for the treatment of ATL.

Reproductive and genetic evidence for a reticulate evolutionary history of mass-spawning corals.

Reef-building corals, which reproduce through simultaneous multispecies spawning, are thought to hybridize frequently, and it is hypothesized that they have evolved in repeated rounds of species separation and fusion. We conducted cross-fertilization experiments and molecular analyses with a number of mass-spawning coral species in the genus Acropora. A high rate of interspecific fertilization occurred between some species despite very different morphologies. The hybrid larvae developed normally and contained an allelic sequence transmitted from each parent, suggesting common diploid hybridization. Molecular phylogenetic analyses provided strong evidence for a gene pool shared between the hybridizing species. These reproductive and genetic characteristics are consistent with a species complex formed under the separation/fusion processes predicted for a reticulate evolutionary history.

Fas-resistance in ATL cell lines not associated with HTLV-I or FAP-1 production.

A preventive role for human T-cell leukemia virus type-I (HTLV-I) and Fas-associated phosphatase-1 (FAP-1) in Fas-mediated apoptosis has been reported in HTLV-I-infected cells. In the present study, we examined whether these molecules increased during the acquisition of Fas-resistance in adult T-cell leukemia (ATL) cell lines. SO4, ST1 and KK1 are Fas-sensitive ATL cell lines, and produce small amounts of HTLV-I in vitro. Although their subclones RSO4 and RST1 are completely Fas-resistant, they produced an equivalent amount of HTLV-I to SO4 and ST1. Moreover, FAP-1 mRNA was not detected in these cell lines irrespective of Fas sensitivity. Thus, Fas resistance in ATL cells was not directly associated with the increased production of HTLV-I or FAP-1.

[Hemolytic anemia and thrombocytopenia associated with anti-omeprazole antibody].

An 80-year-old woman was admitted with anemia, jaundice and a bleeding tendency about 5 weeks after starting omeprazole. On admission, the hemoglobin was 6.4 g/dl, platelets 0.1 x 10(4)/microliter, leukocyte count 7,500/microliter, and reticulocyte count 325/1000. The total bilirubin was 1.9 mg/dl, indirect bilirubin 0.6 mg/dl, lactate dehydrogenase 572 IU/l, and haptoglobin < 10 mg/dl. Both the direct and the indirect Coombs' tests were positive. The platelet-associated IgG (PAIgG) was 1,100.0 ng/10(7) cells. A decrease in the complement value was observed. There was an increase in the number of megakaryocytes and erythroblasts in the marrow film. After omeprazole administration was halted, her hemoglobin and platelet levels gradually returned to normal. On the 27th hospital day, the direct Coombs' test was positive but the indirect Coombs' test became negative. The PAIgG value also returned to normal, and she was discharged on the 59th hospital day. The acute phase of the drug-induced lymphocyte stimulation test was negative, however, we detected the IgG antibody to omeprazole. In the recovery phase, the IgG value decreased. Forty days after discharge, the direct Coombs' test had become negative. This is apparently the first report of a patient with acute hemolytic anemia and thrombocytopenia due to omeprazole through an immune complex mechanism.

An Ancient Chemosensory Mechanism Brings New Life to Coral Reefs.

The first scleractinians, progenitors of modern corals, began to appear 240 million years ago; by the late Jurassic (150 Ma) most families of modern corals had evolved and begun forming reefs (1, 2). Mechanisms controlling the recruitment of new corals to sustain these structures are, however, poorly understood (3). Corals, like many marine invertebrates, begin life as soft-bodied larvae that are dispersed in the plankton (3, 4). As the first step in developing a calcified coral colony, the larva must settle out of the plankton onto a suitable substratum and metamorphose to the single calcified polyp stage cemented to the reef (3, 5). Our analyses of the metamorphic requirements of larvae in divergent coral families surprised us by revealing the existence of a common chemosensory mechanism that is required to bring larvae out of the plankton and onto the reef. This mechanism appears to be quite old, predating both the phylogenetic divergence of these coral families and the development of different modes of coral reproduction.

Clinical significance of tissue polypeptide antigen in serum of acute nonlymphocytic leukemia.

Tissue polypeptide antigen (TPA) in serum was measured at diagnosis in 27 patients with acute nonlymphocytic leukemia (ANLL) (1 FAB M0, 1 M1, 10 M2, 7 M3, 5 M4, 1 M5, 1 M6 and 1 MU). Statistical analysis disclosed a close correlation of TPA level with age (P < 0.01), hemoglobin level (P < 0.05), therapeutic response (P < 0.01) and the length of survival after the initial diagnosis (P < 0.02). A significant difference in TPA level was present between patients with complete remission and those with poor response. To our knowledge, this is the first report to prove a correlation of TPA level with therapeutic response and the length of survival in ANLL.

Clinical significance of serum thymidine kinase in adult T-cell leukaemia and acute myeloid leukaemia.

To clarify the clinical and biological significance of serum thymidine kinase (TK) in adult T-cell leukaemia (ATL) associated with human lymphotropic virus type-I (HTLV-I) and in acute myeloid leukaemia (AML), TK was measured in 52 patients with ATL (acute ATL, 35 patients; lymphoma ATL, two patients; chronic ATL, 12 patients; smouldering ATL, three patients), and in 27 patients with AML (one FAB MO, one M1, 10 M2, seven M3, five M4, one M5, one M6, one MU). In ATL patients, statistical analysis disclosed a close correlation between TK level and the leucocyte count (P < 0.01), and absolute number of abnormal lymphocytes (P < 0.01). However, no correlation was observed between serum lactic dehydrogenase (LDH) level and these items. Concerning the therapeutic response, a statistical difference was present in TK between complete remission and no response (P < 0.05), but not in LDH. We also investigated a significant inverse correlation between TK level as well as LDH level and the length of survival after the initial diagnosis (P < 0.01). In AML patients a close correlation of TK level with the count of leucocytes (P < 0.01), percentage of blasts in the blood (P < 0.05), therapeutic response (P < 0.01) and the length of survival after the initial diagnosis (P < 0.05) was present. Therefore the TK level may indicate the aggressiveness of leukaemic cells and predict the response to the chemotherapy and the length of survival in ATL and AML.

A statistical study of calcifying epithelioma, focusing on the sites of origin.

We statistically investigated 396 lesions taken from 355 cases of calcifying epithelioma. The distribution of these lesions did not correlate with the density of the hair follicles, but it was in accord with the distribution of intermediate hairs, such as those in the hair border. This relationship may have etiologic significance.

Isolation and characterization of a mini-collagen gene encoding a nematocyst capsule protein from a reef-building coral, Acropora donei.

Genomic and cDNA clones of a mcol gene encoding mini-collagen (MCOL), a nematocyst capsule protein, have been isolated from a reef-building coral, Acropora donei (Anthozoa). The gene and its flanking regions, comprising 5382 bp and covering three exons and two introns, were sequenced. Exons 2 and 3 together have an open reading frame which can encode a MCOL of 176 amino acids (aa). The coral MCOL has all the characteristic regions present in the four hydra MCOL specified by the four mcol cDNA clones previously isolated from Hydra magnipapillata (Hydrozoa) by Kurz et al. [J. Cell Biol. 115 (1991) 1159-1169], including a central Gly-Xaa-Yaa region and flanking Pro-rich and Cys-repeat regions. This observation suggests that a mcol family is highly conserved in Anthozoa and Hydrozoa, and also that the characteristic regions present in MCOL are essential for the structure and function of these peptides.

Binding of the amino-terminal region of myosin alkali 1 light chain to actin and its effect on actin-myosin interaction.

The role of the amino-terminal region of myosin alkali 1 light chain (A1) in the interaction between actin and myosin subfragment-1 (S-1) was explored. Papain digestion of skeletal myosin filaments produced S-1 whose A1 was found to lose the basic 13 amino-terminal amino acid residues (A1'). We obtained three types of papain S-1 isoenzymes differing in their alkali light chain content: recombined papain S-1 (A1), papain S-1 (A1'), and papain S-1 (A2). Both the maximum turnover rate (Vmax) and the dissociation constant (Km) for actin-activated papain S-1 (A1') ATPase activity were similar to those for papain S-1 (A2) and remarkably larger than those for recombined papain S-1 (A1). The 13 amino-terminal residue peptide of A1 (N-pep) was isolated and characterized. 1H-NMR spectroscopy suggested that the N-pep was relatively immobilized in the presence of actin filaments. A cross-linking study suggested that N-pep binds to actin. The addition of N-pep to acto-S-1 (A1) made Km and Vmax for the actin-activated ATPase activity close to those for S-1 (A2). Removal of the trimethyl group from the N-pep suppressed the above effect on the actin-S-1 interaction. Our findings suggest that the amino-terminal region of A1 binds to the actin molecule to affect the mechanism of actin-activated S-1 ATPase.

Therapeutic and prognostic value of modal number of chromosomes at the blastic phase of Philadelphia-chromosome-positive chronic myeloid leukemia: comparison based on the same criteria between Nagasaki University and Roswell Park Memorial Institute.

In a comparison of 47 patients with Philadelphia-chromosome (Ph)-positive chronic myeloid leukemia (CML) in the Nagasaki University School of Medicine and 64 patients with the same disease in the Roswell Park Memorial Institute, the correlation between the modal number of chromosomes and the therapeutic response and/or survival after the onset of the blastic phase (BP) was evaluated. The patients were divided into four groups on the basis of the modal number of chromosomes of the cells in the bone marrow: those with hypodiploidy (group 1), those with pseudodiploidy carrying a Ph chromosome (group 2), those with 47 chromosomes (group 3), and those with 48 or more chromosomes (group 4). The results revealed similar trends in the two institutes. Namely, the therapeutic response and the survival after the onset of the BP in groups 1 and 4 were more unfavorable and shorter than those in groups 2 and 3, although the former (group 2) had a better prognosis than the latter (group 3). Thus, the statistical analysis revealed that the numerical chromosome findings at the BP are useful parameters for assessing the therapeutic response and survival after the onset of the BP of CML.