Yutaka Maruoka

The Novel Gene Encoding a Putative Transmembrane Protein Is Mutated in Gnathodiaphyseal Dysplasia (GDD)

Gnathodiaphyseal dysplasia (GDD) is a rare skeletal syndrome characterized by bone fragility, sclerosis of tubular bones, and cemento-osseous lesions of the jawbone. By linkage analysis of a large Japanese family with GDD, we previously mapped the GDD locus to chromosome 11p14.3-15.1. In the critical region determined by recombination mapping, we identified a novel gene (GDD1) that encodes a 913-amino-acid protein containing eight putative transmembrane-spanning domains. Two missense mutations (C356R and C356G) of GDD1 were identified in the two families with GDD (the original Japanese family and a new African American family), and both missense mutations occur at the cysteine residue at amino acid 356, which is evolutionarily conserved among human, mouse, zebrafish, fruit fly, and mosquito. Cellular localization to the endoplasmic reticulum suggests a role for GDD1 in the regulation of intracellular calcium homeostasis.

Expression of bone morphogenetic protein genes in the human dental pulp cells

Dental pulp has a potential to induce ectopic bone formation, but little is known about its mechanism. We thought that bone morphogenetic proteins (BMPs), members of the transforming growth factor-beta (TGF-beta) superfamily, are involved in the osteoinductive activity of dental pulp. In order to prove this assumption, we constructed a cDNA library from primary culture cells of human dental pulp (HDP cells), and screened the library with previously cloned cDNAs for mouse BMP-2 and -6 as probes. Three distinct cDNA clones encoding human BMP-2, -4 and -6 were isolated. By Northern blot analysis, specific transcripts of the genes of those BMPs were detected in the HDP cells. It was concluded that the BMPs were expressed in a certain population of dental pulp cells and might play some roles in ectopic bone formation by dental pulp.

Deficiency of Chemokine Receptor CCR1 Causes Osteopenia Due to Impaired Functions of Osteoclasts and Osteoblasts

Chemokines are characterized by the homing activity of leukocytes to targeted inflammation sites. Recent research indicates that chemokines play more divergent roles in various phases of pathogenesis as well as immune reactions. The chemokine receptor, CCR1, and its ligands are thought to be involved in inflammatory bone destruction, but their physiological roles in the bone metabolism in vivo have not yet been elucidated. In the present study, we investigated the roles of CCR1 in bone metabolism using CCR1-deficient mice. Ccr1−/− mice have fewer and thinner trabecular bones and low mineral bone density in cancellous bones. The lack of CCR1 affects the differentiation and function of osteoblasts. Runx2, Atf4, Osteopontin, and Osteonectin were significantly up-regulated in Ccr1−/− mice despite sustained expression of Osterix and reduced expression of Osteocalcin, suggesting a lower potential for differentiation into mature osteoblasts. In addition, mineralized nodule formation was markedly disrupted in cultured osteoblastic cells isolated from Ccr1−/− mice. Osteoclastogenesis induced from cultured Ccr1−/− bone marrow cells yielded fewer and smaller osteoclasts due to the abrogated cell-fusion. Ccr1−/− osteoclasts exerted no osteolytic activity concomitant with reduced expressions of Rank and its downstream targets, implying that the defective osteoclastogenesis is involved in the bone phenotype in Ccr1−/− mice. The co-culture of wild-type osteoclast precursors with Ccr1−/− osteoblasts failed to facilitate osteoclastogenesis. This finding is most likely due to a reduction in Rankl expression. These observations suggest that the axis of CCR1 and its ligands are likely to be involved in cross-talk between osteoclasts and osteoblasts by modulating the RANK-RANKL-mediated interaction.

Autosomal Dominant Gnathodiaphyseal Dysplasia Maps to Chromosome 11p14.3-15.1†

Gnathodiaphyseal dysplasia (GDD) is a syndrome characterized by bone fragility, sclerosis of tubular bones, and cemento‐osseous lesions of jawbones. Although some cases of this syndrome exist in families with autosomal dominant inheritance, the underlying gene has never been identified. We analyzed a large four‐generation family with GDD by linkage analysis using genomic DNA from nine affected and six nonaffected family members. A genome‐wide search using a set of highly polymorphic microsatellite markers showed evidence for linkage to chromosome 11p14.3‐15.1. Two‐point linkage analysis of microsatellite markers spanning this locus resulted in a maximum logarithm of odds (LOD) score of 2.70 with a recombination fraction (θ) of 0 at D11S1755, D11S1759, and D11S915, and a maximum LOD score of 3.01 at D11S4114 was obtained in multipoint linkage analysis. Haplotype analysis detected no recombination between GDD and six closely linked markers (D11S928, D11S1755, D11S4114, D11S1759, D11S915, and D11S929) and established the candidate interval of 8.7 cM on chromosome 11p for GDD. Although GDD has been considered to be a variation of osteogenesis imperfecta (MIM 166260), our results indicate that this syndrome is a new and distinct disease entity from other systemic bone diseases. Furthermore, these genetic markers are useful for presymptomatic diagnosis of GDD in some families and for identification of the GDD gene.

Effect of recombinant human fibroblast growth factor‐2 on bone formation in rabbit mandibular distraction models using β‐tricalcium phosphate

This study was designed to evaluate the effect of recombinant human fibroblast growth factor‐2 (rhFGF‐2) on the amount and period of new bone formation in rabbit mandibular distraction models using β‐tricalcium phosphate (β‐TCP) as a bone graft substitute. Sixteen male Japanese White rabbits were divided into the following four experimental groups: 1, distraction alone; 2, distraction with β‐TCP granules; 3, distraction with rhFGF‐2 (25 µg/50 µL) injected into β‐TCP granules; and 4, distraction with rhFGF‐2 (100 µg/50 µL) injected into β‐TCP granules. The bones were harvested at 4 weeks after the operation and examined using soft radiography, micro‐computed tomography (micro‐CT), and peripheral quantitative computed tomography (pQCT). The dissected mandibles were stained using the Villanueva bone staining method, and the amount of new bone formed, bioresorption of β‐TCP, and new blood vessel formation were morphometrically calculated using bone histomorphometry. Radiopaque areas were observed more frequently in the distracted area of groups 3 and 4. Micro‐CT analysis revealed partial new bone formation in the central region of the distracted area in groups 3 and 4. pQCT analysis revealed increased bone mineral density in groups 3 and 4. Histomorphometric analysis revealed increased newly formed bone and blood vessel areas in groups 3 and 4. In group 4, the number of osteoclasts around the β‐TCP granules had significantly increased. The present findings suggested that the combined use of rhFGF‐2 and β‐TCP reduced the treatment period for distraction osteogenesis and accelerated the formation of a new high‐quality bone.

Importance of eliminating potential dental focal infection before the first cycle of chemotherapy in patients with hematologic malignancy

Letter to the Editor, The oral cavity is a potential infection site in patients undergoing immunosuppressive therapy [1]. These patients are more susceptible to systemic infection, particularly those who need chemotherapy for hematologic malignancies [2]. Many dentists find it difficult to select an approach for dental focal infection in these patients [1, 2]. Although there is some literature assessing oral health in patients undergoing chemotherapy or pre-hematopoietic stem cell transplantation [3], these patients had already experienced adverse effects of chemotherapy, making it difficult to interpret the effect of dental treatment. Tooth extraction is a common surgical procedure performed to eliminate dental focal infection, and it carries a risk of perioperative surgical site infection (SSI) [1]. Although the frequency of tooth extraction-associated SSI is <5% in healthy individuals [4], it may be increased in patients with hematologic malignancy. We retrospectively reviewed the records of 160 patients with hematologic malignancy who visited the Department of Oral and Maxillofacial Surgery, Center Hospital, National Center for Global Health and Medicine, Tokyo, Japan, over a 2-year period to clarify the incidence of oral adverse events if dental problems are treated before the first chemotherapy cycle. Seventy-five patients were treated before the first cycle. Dental treatments were performed, and oral health status was assessed after chemotherapy. The observation period was the number of days from the start of the first chemotherapy cycle to just before the start of the second cycle. In cases where the cycle was unclear (e.g., when a single agent was administered repeatedly), the observation period was 30 days from chemotherapy initiation (i.e., the period deemed by hospital management to constitute adequate follow-up). Nine (12.0%) patients experienced oral adverse events after chemotherapy. No significant difference was found between the groups with and without tooth extraction (P = 0.28). The most frequently observed symptom was periodontitis aggravation (n = 3, 33.0%). Although all three patients were deemed to require extraction at the first stomatognathic screening, a decision not to proceed was made in two cases owing to a tight chemotherapy schedule. However, inflammation steadily worsened thereafter. Remarkably, only two of 28 (7.1%) tooth extraction cases had a poor outcome, regardless of the operative stress (surgical or simple extraction). Oral antibiotics (penicillin or cephem) were administered starting just after surgery in all extraction cases and continued at least for 3 days. If oral administration was impossible, the intravenous route was used. The median (min-max) white blood cell * Yasuyuki Shimada yasuyuki.shimada.ys@gmail.com

Bone with a Vascular Flap Induced from Fat Tissue with the Use of rhBMP-2 in Rats

Here we report that successful bone formation with a vascular flap inside a cylindrical mold was induced from fat tissue with the use of recombinant human bone morphogenetic protein-2 in rats. Fat tissue connected to blood vessels was prepared to fit into the mold and implanted intramuscularly into the hind leg in Wistar rats. RhBMP-2 (20 μg) was applied in a collagen sheet previously placed on the inside surface of the mold. Bone formation was confirmed radiologically and morphologically at 2, 4, and 8 weeks after the surgery. In the control group without rhBMP-2 or the group with ligation of the blood vessels before the implantation, bone formation was not observed. Our success in bone formation having a definite size, shape, and blood supply may lead to a therapeutic approach to effective bone reconstitution. The present study is the first report on bone induction from fat tissue by rhBMP-2 in vivo.

Associations between Serum Bone-Specific Alkaline Phosphatase Activity, Biochemical Parameters, and Functional Polymorphisms of the Tissue-nonspecific Alkaline Phosphatase Gene in a Japanese Population

INTRODUCTION We had demonstrated that single nucleotide polymorphism (787T>C) in the tissue-nonspecific ALP (TNSALP) gene was associated with the bone mineral density (BMD). BMD was the lowest among TNSALP 787T homozygotes (TT-type) and highest among TNSALP 787T>C homozygotes (CC-type) in postmenopausal women. In the present study, we investigated the effects of the TNSALP genotype on associations among serum bonespecific alkaline phosphatase (BAP), serum calcium, and phosphorus in healthy young Japanese subjects. METHODS Young healthy adult subjects (n=193) were genotyped for the polymorphism, and we measured the levels of serum BAP, serum calcium, and phosphorus. Dietary nutrient intakes were calculated based on 3-day food records before the day of blood examinations. RESULTS Grouped by the TNSALP genotype, a significant negative correlation between serum BAP and phosphorus was observed in 787T>C homozygotes (CC-type), but not in heterozygotes (TCtype), nor in 787T homozygotes (TT-type). CONCLUSIONS In the present study, we revealed that the single nucleotide polymorphism 787T>C in the TNSALP gene had effects on the correlation between serum BAP and phosphorus in young adult subjects. These results suggest that variation in TNSALP may be an important determinant of phosphate metabolism. Our data may be useful for planning strategies to prevent osteoporosis.

Molecular cloning and sequence of bovine Msx-1 homeobox-containing gene cDNA from a bovine odontoblast library

Screening of a bovine odontoblast cDNA library from developing incisor with murine Msx-1 and Msx-2 cDNA probes led to the isolation of three positive clones. All of them encoded for a sequence of a protein containing 297 amino acids. The responsible gene was designated as bovine Msx-1 (bMsx-1) due to the high homology with the human MSX-1 and mouse Msx-1 sequences.

A case of central odontogenic fibroma of the mandible in a nevoid basal cell carcinoma syndrome patient

Abstract Nevoid basal cell carcinoma syndrome (NBCCS) is an autosomal dominant disorder characterized by tumorigenesis and physical deformities. Although more than 50 clinical manifestations have been described, only two major criteria or one major and two minor criteria are necessary for diagnosis. The most frequently observed manifestation in the oral and the maxillofacial region is an odontogenic keratocyst. In this study, we describe a 14-year-old boy with a diagnosis of NBCCS who presented with a central odontogenic fibroma (COF) in the mandible. This report highlights the importance of precise diagnosis and the choice of surgical method for COF.