Histological study of the nasal septal cartilage in BALB/c-bm/bm mouse which spontaneously induces malocclusion.

OBJECTIVES: The BALB/c-bm/bm mouse is characterized by short limbs and short tail attributed to undersulfated glycosaminoglycans. Anterior transverse crossbite sometimes spontaneously appears in BALB/c-bm/bm mice. The BALB/c-bm/bm mouse shows a short nose and cranium. The reason for hypo-growth of anterior craniofacial structures has not been clarified, although the nasal septal cartilage might be related to the growth of anterior craniofacial structures. Therefore, the purpose of this study was to evaluate histological findings of the nasal septal cartilage at the border region of the ethmoid and sphenoid bone in BALB/c-bm/bm mice. MATERIALS AND METHODS: BALB/c mice (wild type) and BALB/c-bm/bm mice with normal occlusion (bm/bm) were used. Sagittal sections of female mice aged 2, 4, and 8 weeks were stained with hematoxylin and eosin for histological analysis. RESULTS: At the border region between the nasal septal cartilage and the ethmoid bone in bm/bm, the area of proliferative zone was significantly smaller than that in wild type. At the border regions between the nasal septal cartilage and both the ethmoid and sphenoid bones, the number of proliferative chondrocytes was significantly smaller. Normal endochondral ossification was not observed at the border region between the nasal septal cartilage and the sphenoid bone in bm/bm. CONCLUSION: The findings suggest that disorder of endochondral ossification in the nasal septal cartilage contributes to the hypo-growth of anterior craniofacial structures in bm/bm.

Morphological evaluation of cranial and maxillary shape differences of the brachymorphic mouse with spontaneous malocclusion using three-dimensional micro-computed tomography.

OBJECTIVES: The aim of this study was to determine whether significant cranial and maxillary deformity exists in BALB/c-bm/bm (brachymorphism) mouse with spontaneous malocclusion using three-dimensional (3D) images. MATERIALS AND METHODS: Thirty female mice were divided into the following three groups: control group (BALB/c mice, n = 10), Norm group (BALB/c-bm/bm mice with normal occlusion, n = 10), and Mal group (BALB/c-bm/bm mice with malocclusion, n = 10). Nine points in the skull were selected, and transverse and antero-posterior distances were measured using three-dimensional images of micro-computed tomography (CT). Moreover, 3D images were superimposed at the median plane to visualize the skull shape asymmetry. RESULTS: The transverse distances at the posterior cranial and maxillary region and the antero-posterior distances in the Norm and Mal groups were significantly shorter than those in the control group. The nasal septum of the Mal group was significantly shorter than that of the Norm group. Morphological measurements and superimposed 3D images showed that lateral deviation occurred at the anterior cranial and maxillary region in the Mal group. CONCLUSION: The 3D micro-CT images revealed that the antero-posterior length and posterior transverse width at the cranium and maxilla in BALB/c-bm/bm mice were significantly smaller than those in BALB/c mice. It was quantitatively and morphologically clear that BALB/c-bm/bm mice show a spontaneous transverse crossbite owing to lateral deviation of the maxilla and nasal bone.

BubR1 localizes to centrosomes and suppresses centrosome amplification via regulating Plk1 activity in interphase cells.

BubR1 is a critical component of the mitotic checkpoint that delays the onset of anaphase until all chromosomes have established bipolar attachment to the microtubules. We previously reported that mutations of the BUB1B gene (encoding BubR1) caused premature chromatid separation (PCS) syndrome, a condition characterized by constitutional aneuploidy and a high risk of childhood cancer. We here report that the cells from PCS syndrome patients have loss of regulation of the centrosome duplication machinery, resulting in centrosome amplification and multipolar mitosis. PCS syndrome cells show increased activity of Polo-like kinase 1 (Plk1), whose knockdown suppresses centrosome amplification. BubR1 localizes to centrosomes, physically interacts with Plk1 and inhibits Plk1 phosphorylation and its kinase activity during interphase. These results unravel a crucial role of BubR1 in preventing centrosome reduplication through negative regulation of Plk1 in interphase cells.

Two cases of mosaic RhD blood-group phenotypes and paternal isodisomy for chromosome 1.

We encountered a 22-year-old man (case 1) and a 23-year-old woman (case 2), both unrelated and healthy. They were mosaic for the Rh blood group phenotype: one erythrocyte population was D-positive and the other was D-negative. Flow cytometric analysis of density profile of RhD antigen in their erythrocytes, and cytogenetic analysis including in situ hybridization using an RHD/RHCE-containing PAC clone, excluded a deletion of the RHD/RHCE gene complex, but suggested the presence of cells with uniparental disomy for chromosome 1 (UPD1). Microsatellite marker analysis was performed in both probands and their family members. In case 1, the analysis with markers spanning the chromosome 1 revealed both maternal and paternal alleles in his peripheral blood leukocytes (PBL), Epstein-Barr virus-transformed lymphoblastoid cells (EBL), and buccal mucosal cells. However, only paternal alleles were detected in all of 50 individual pieces of his hair or hair-roots and all of five monoclonal cell lines cloned from his established EBL. There was no direct evidence of heterozygous, biparental alleles in these two tissues. The presence of maternal isodisomy 1 was not absolutely ruled out in other tissues examined in case 1. Similar results were obtained in case 2, showing biparental, disomic patterns in her PBL and in 15 of 20 pieces of her hair roots, and showing monoallelic patterns in the remaining five pieces of hair roots. Analysis with markers for other autosomes confirmed their biparental inheritance. These findings indicated that both cases had at least two cell populations, one population having paternal UPD1 (isodisomy 1), and another heterozygous, biparental disomy 1. We emphasize that isodisomy for chromosome 1 is not infrequent and may cause unusual RhD phenotype, as seen in cases we described.

Cancer-prone syndrome of mosaic variegated aneuploidy and total premature chromatid separation: report of five infants.

Five infants (two girls and three boys) from four families all had severe pre- and postnatal growth retardation, profound developmental delay, microcephaly, hypoplasia of the brain with Dandy-Walker complex or other posterior fossa malformations, and developed uncontrollable clonic seizures. Four infants developed Wilms tumors, and one showed cystic lesions in bilateral kidneys. All five infants showed variegated mosaic aneuploidy in cultured lymphocytes. In two infants whose chromosomes were prepared by us, 48.5%-83.2% lymphocytes showed total premature chromatid separation (PCS). Their parents had 3.5%-41.7% of their lymphocytes in total PCS. The remaining three infants and their parents, whose chromosomes were prepared at outside laboratories, tended to show lower frequencies of total PCS. Another five infants reported with the disorder were reviewed together with the five infants we described. Together, their clinical and cytogenetic manifestations were similar enough to suggest a syndrome. Seven of the 10 infants developed proven or probable Wilms tumors. The age at diagnosis of the tumors was younger than usual at 2-16 months. The tumors were bilateral in four infants and unilateral in three infants, and cystic changes were present in six infants. Two infants developed botryoid rhabdomyosarcoma. The carriers of the syndrome are thus liable to tumorigenesis. The possible role of mitotic checkpoint defects, proven in two infants with the syndrome (Matsuura et al. [2000: Am J Hum Genet 69:483-486]), was discussed in connection with tumor development and progression.

Presence of third molar germs in orthodontic patients in Japan.

The purpose of this investigation was to evaluate the existence of third molar germs in orthodontic patients in Japan and to examine the relationship between the existence of third molars and sagittal maxillomandibular jaw relationships. The subjects comprised 306 patients from the orthodontic clinic of Hokkaido University Dental Hospital who were younger than 15 years. The subjects were divided into 2 groups: 1 group included 144 patients who were born between 1966 and 1969 (60s group), and the other group included 162 patients who were born between 1980 and 1987 (80s group). Assessments were made from panoramic radiographs and lateral cephalograms. The following results were obtained: (1) all 4 third molar germs were present in 77% of subjects, (2) mandibular third molars were present significantly more often than maxillary third molars, and (3) the percentage of skeletal Class III subjects who had all 4 third molars was lower than that of skeletal Class II subjects. The chi(2) test was used to determine statistical significance in differences.

Chromosomal instability syndrome of total premature chromatid separation with mosaic variegated aneuploidy is defective in mitotic-spindle checkpoint.

Skin fibroblast cells from two unrelated male infants with a chromosome-instability disorder were analyzed for their response to colcemid-induced mitotic-spindle checkpoint. The infants both had severe growth and developmental retardation, microcephaly, and Dandy-Walker anomaly; developed Wilms tumor; and one died at age 5 mo, the other at age 3 years. Their metaphases had total premature chromatid separation (total PCS) and mosaic variegated aneuploidy. Mitotic-index analysis of their cells showed the absence of mitotic block after the treatment with colcemid, a mitotic-spindle inhibitor. Bromodeoxyuridine-incorporation measurement and microscopic analysis indicated that cells treated with colcemid entered G1 and S phases without sister-chromatid segregation and cytokinesis. Preparations of short-term colcemid-treated cells contained those cells with chromosomes in total PCS and all or clusters of them encapsulated by nuclear membranes. Cell-cycle studies demonstrated the accumulation of cells with a DNA content of 8C. These findings indicate that the infants' cells were insensitive to the colcemid-induced mitotic-spindle checkpoint.

Long-term effects of prostaglandin E2 on the mineralization of a clonal osteoblastic cell line (MC3T3-E1).

Prostaglandin (PG) E2 is thought to be a mediator of the effect of mechanical stress on bone formation, but its effects on osteoblasts have not yet been fully described. Here, the effects of the continuous application of PGE2 and indomethacin, an inhibitor of prostaglandin G/H synthase (cyclo-oxygenase), on the proliferation, differentiation and mineralization of a clonal osteoblastic cell line, MC3T3-E1, were investigated. The cells were cultured in media with either a high (1 microg/ml) or a low (1 ng/ml) concentration of PGE2, with indomethacin (1 microg/ml) and, as a control, with neither agent. The effects of PGE2 and indomethacin were assessed quantitatively. Indomethacin and a high concentration of PGE2 increased the total protein compared to the control and low-PGE2 cultures. 7 days after confluence, alkaline phosphatase (ALP) activity within the cells and extracellular matrices increased. This increase was highest with indomethacin and lowest with a high concentration of PGE2. ALP activity also increased in the medium, but only 21 days after confluence; the effects of the agents were similar to those on the cells and matrices. The accumulation of calcium, inorganic phosphate and hydroxyproline was highest with indomethacin. PGE2 production was at its maximum when the cells were at confluence and was inhibited by indomethacin. Specific [3H]PGE2 binding to the microsomal fraction of the cell was also measured to examine the expression of the PGE2 receptor. The amount of [3H]PGE2 binding per mg of protein was highest at confluence, then decreased and again increased in the mineralizing stage. These results suggest that indomethacin increases ALP activity and the accumulation of mineralized tissue in MC3T3-E1 cells, presumably by inhibiting the production of PGE2. PGE2 could signal the suppression of mineralization as early as confluence.

Mosaic variegated aneuploidy with multiple congenital abnormalities: homozygosity for total premature chromatid separation trait.

Separation of chromatids of all mitotic chromosomes, here called total premature chromatid separation (total PCS), was observed in 67 to 87.5% of repeated cultures of peripheral blood lymphocytes from two unrelated infants. Also noted was a variety of mosaic aneuploidies, especially trisomies, double trisomies, and monosomies, to be called mosaic variegated aneuploidy. The infants both showed severe pre- and postnatal growth retardation, profound developmental retardation, uncontrollable seizures, severe microcephaly, hypoplasia of the brain, Dandy-Walker anomaly, abnormal facial appearance, and bilateral cataract. Patient 1, a girl, in addition had a cleft palate, multiple renal cysts, and Wilms tumor of the left kidney. Whereas patient 2, a boy, had ambiguous external genitalia. They both died within 2 years of age. In the two families of the infants, their parents and three other members showed 2.5 to 47% lymphocytes with total PCS but without mosaic variegated aneuploidy or phenotypic abnormalities. Another 10 relatives studied showed 0 to 1% cells with total PCS and so were judged negative for the total PCS trait. It was deduced that the total PCS trait in the two families was transmitted in an autosomal-dominant fashion, and the two affected infants were homozygous for the trait.

A recurrent translocation, t(16;21)(q24;q22), associated with acute myelogenous leukemia: identification by fluorescence in situ hybridization.

Chromosome fluorescence in situ hybridization (FISH) analyses were performed on bone marrow cells in 3 adult patients with MDS or AML with a (16;21)(q24;q22) translocation. FISH analyses with AML1 probes at 21q22 proved in all 3 patients splitting of the AML1 gene at a region spanning exons 5 and 6 and the translocation of its 5' segment to distal 16q. Chromosome painting FISH analysis in patient 1 proved the translocation of the distal 21q segment to 16q, but it failed to prove the presumed translocation of the distal 16q segment to 21q, most likely because of its small size.

Interstitial deletion of 8p: report of two patients and review of the literature.

Two female infants with de novo interstitial deletions of 8p were studied. One with a deletion from p11.21 to p11.23, and the other patient with a deletion from p11.23 to p21.3 had several clinical manifestations of the terminal 8p- syndrome. Band 8p11.23 was deleted in both patients. The clinical manifestations common to both patients included low birthweight, growth deficiency, congenital heart disease, mental retardation, dolichocephaly, low-set, malformed ears, high-arched palate, thin lips and micrognathia. Since these features may occur in most patients with chromosomal imbalance, and the terminal 8p- syndrome has hitherto been assumed to result from terminal deletions of 8p, ranging from p21.3 to p23, it is likely that these features are simply related to the chromosomal imbalance rather than to band specific imbalance of 8p11.23. The present study suggests that two different types of deletion, interstitial and terminal, are associated with still poorly defined, rather non-specific clinical features.

Congenital heart defects in Aarskog syndrome.

We report on 10 Japanese individuals from 3 families affected with Aarskog syndrome. Pulmonary stenosis and ventricular septal defect with spontaneous closure were detected respectively, in 2 of them as an uncommon finding. A review documented 169 non-Japanese cases (2 with congenital heart defects), while of 30 Japanese individuals reported till now, 4 (including ours) had cardiac anomalies. We propose that this combination is not coincidental and that in all cases of Aarskog syndrome a cardiac evaluation is indicated.

Occipital horn syndrome: report of a patient and review of the literature.

We report an 18-year-old boy with occipital horn syndrome and we review the 20 cases previously published with this syndrome. The distinctive features common to all patients were unusual facial appearance, skeletal abnormalities, chronic diarrhea and genitourinary abnormalities. The skeletal abnormalities included occipital horns, short, broad clavicles, deformed radii, ulnae, and humeri, narrowing of the rib cage, undercalcified long bones with thin cortical walls and coxa valga. Occipital horn syndrome is inherited in an X-linked recessive fashion. Our analysis indicates that occipital horn syndrome is associated with a recognizable characteristic phenotype.

Prenatal diagnosis of fragile X syndrome by direct detection of the dynamic mutation due to an unstable DNA sequence.

The fragile X syndrome is the most common familial form of mental retardation. The mutation causing the syndrome is dynamic mutation due to an unstable DNA (CCG)n repeat localized at Xq27.3. We have previously reported a PCR procedure to prepare a diagnostic probe, pPCRfx1, which can be used to determine the genotype of fragile X mutation individuals by Southern blot analysis. In the present study, pPCRfx1 was applied to the prenatal diagnosis, using chorionic villus cells, of a fetus which was at risk of having fragile X syndrome. In the PstI assay, the Southern blot showed the typical pattern of a female carrier with the full mutation. Analysis of the DNA methylation patterns by EcoRI + EagI assay showed that the EagI restriction site was not methylated on the mutated X chromosome of chorionic villi, but the sites were totally methylated in the brain and other tissues of the fetus. Thus the fetus was diagnosed to be a heterozygous female carrier of the dynamic mutation involved in the fragile X syndrome.

Midkine gene (MDK), a gene for prenatal differentiation and neuroregulation, maps to band 11p11.2 by fluorescence in situ hybridization.

Midkine (MDK) is a retinoic acid-responsive gene concerned with prenatal development and neurite growth. We mapped the gene to band p11.2 of chromosome 11 through fluorescence in situ hybridization analysis and using a 4.5-kb fragment of its human genomic DNA.

Two Japanese cases with microcephalic primordial dwarfism: classical Seckel syndrome and osteodysplastic primordial dwarfism type II.

A male infant with "classical" Seckel syndrome and a girl with osteodysplastic primordial dwarfism type II are described. The boy with classical Seckel syndrome had severe brain dysplasia, a finding hitherto unreported in patients with this syndrome. The patient with osteodysplastic dwarfism type II had skeletal abnormalities including lumbar scoliosis, a small and high pelvis, metaphyseal flaring of the distal radii and ulnae, V-shaped metaphyseal flaring of the distal femorae, and short metacarpals and phalanges. The mother of this girl was short, microcephalic, and had disproportionately short forearms and legs. In view of this, dominant inheritance of the disease was suggested.

Dicentric Y chromosome in azoospermic males.

Two azoospermic, infertile men with a pseudodicentric Y chromosome are reported. The small isodicentric Y chromosomes were composed of duplicated short arm and proximal long arm Y, as proven by fluorescence in situ hybridisation using a Y centromere-specific DNA probe, pDP97, and a short arm probe pY-80. Both lacked germinal cells in the gonads. It was assumed that the azoospermia was caused by deletion or disruption of the azoospermic factor gene located at distal Yq11. Patient 2 measured 147 cm (-4.1 SD) in height and so it was assumed that he had also lost the "statural determinants" gene.