Runx2 haploinsufficiency ameliorates the development of ossification of the posterior longitudinal ligament

Ossification of the Posterior Longitudinal Ligament (OPLL) is a disease that is characterized by the ectopic calcification of the ligament; however, the pathogenesis of OPLL remains to be investigated. We attempted to identify the in vivo role of Runx2, a master regulator of osteoblast differentiation and skeletal mineralization, in the pathogenesis of OPLL. The expression of Runx2 in the ligament was examined using in situ hybridization and immunohistochemistry and by monitoring the activity of a LacZ gene that was inserted into the Runx2 gene locus. To investigate the functional role of Runx2, we studied ENPP1(ttw/ttw) mice, a mouse model of OPLL, that were crossed with heterozygous Runx2 mice to decrease the expression of Runx2, and we performed histological and quantitative radiological analyses using 3D-micro CT. Runx2 was expressed in the ligament of wild-type mice. The induction of Runx2 expression preceded the development of ectopic calcification in the OPLL-like region of the ENPP1(ttw/ttw) mice. Runx2 haploinsufficiency ameliorated the development of ectopic calcification in the ENPP1(ttw/ttw) mice. Collectively, this study demonstrated that Runx2 is expressed in an OPLL-like region, and its elevation is a prerequisite for developing the complete OPLL-like phenotype in a mouse model of OPLL.     

Characteristic differences among osteogenic cell populations of rat bone marrow stromal cells isolated from untreated, hemolyzed or Ficoll-treated marrow

Background aimsAlthough bone marrow (BM) stromal cells (SC; BMSC) isolated from adherent cultures of untreated BM are known to contain both committed and uncommitted osteogenic cells, it remains unknown whether BMSC isolated either by hemolysis or Ficoll centrifugation also contain both of these populations.MethodsDifferences in the osteogenic cell populations of rat BMSC isolated from untreated, hemolyzed or Ficoll-treated BM were analyzed by in vivo transplantation, flow cytometry, alkaline phosphatase (ALP) assay, real-time polymerase chain reaction (PCR) and alizarin red staining.ResultsTransplantation of non-cultured samples indicated that the Ficolled BMSC contained the lowest number of committed osteogenic cells. Flow cytometric analysis of cultured, non-induced samples showed that the percentage of ALP-positive cells was significantly lower in Ficolled BMSC. Quantitative ALP assays confirmed that the lowest ALP activity was in the Ficolled BMSC. Hemolyzed BMSC also contained lower numbers of committed osteogenic cells than untreated BMSC, but still more than Ficolled BMSC. Interestingly, the Ficolled BMSC showed the greatest levels of osteogenic ability when cultured in osteogenic induction medium.ConclusionsThese findings suggest that, although Ficolled BMSC rarely contain committed osteogenic cells, they are able to show comparable or even greater levels of osteogenic ability after induction, possibly because they contain a greater proportion of uncommitted stem cells. In contrast, induction is optional but recommended for both untreated and hemolyzed BMSC before use, because both these groups contain both committed and uncommitted osteogenic cells. These findings are of significant importance when isolating BMSC for use in bone tissue engineering.     

Effects of an Epidermal Growth Factor on Growth of Zea Primary Roots and Mesocotyls

Sections of the elongation zones of primary roots and mesocotylsof Zea mays L. were incubated with various concentrations ofhuman epidermal growth factor (EGF). The growth rates of rootsections incubated with 1,000, 100 and 10 µg liter^–1EGF were higher than that of control by 15%, 26% and 14%, respectively.The rates of mesocotyl sections incubated with 100, 10, 1.0and 0.1µg liter^–1 EGF were 23%, 31%, 24% and 22%higher than that of control. (Received August 8, 1994; Accepted November 1, 1994)     

Polymerase chain reaction detection of two novel human N-acetylgalactosamine-6-sulfate sulfatase gene polymorphisms by single-strand conformation polymorphism analysis or by StyI and StuI cleavages

A novel polymorphism (6376 G/T) in intron 7 (I7) of the human PROC gene has been identified by direct DNA sequencing. Restriction analysis with the use of mutagenic primers indicate that the allele frequencies are 0.17 (allele T) and 0.83 (allele G), with a calculated heterozygosity of 28%.     

Akazara scallop myosins hybridized with DTNB-light chains of skeletal myosin and with regulatory light chains of gizzard myosin

Two different hybrid myosins were obtained by combining "desensitized" myosin (DM) of Akazara scallop striated adductor with rabbit skeletal DTNB-light chains (DTNB-LC) and with chicken gizzard regulatory light chains (GR-LC). Using the two hybrid myosins, the following were found: (a) DTNB-LC has an inhibitory effect on the Mg-ATPase activities of Akazara DM and acto-DM both in the absence of calcium and in its presence. (b) DTNB-LC also has an enhancing effect on the superprecipitation activity of acto-DM. (c) The Mg-ATPase activities of DM and acto-DM are made sensitive to calcium by GR-LC, regardless of whether GR-LC is phosphorylated or unphosphorylated. (d) However, the Mg-ATPase activity of acto-myosin hybridized with phosphorylated GR-LC is definitely higher than that of acto-myosin hybridized with unphosphorylated GR-LC.     

Replication asynchrony between homologs 15q11.2: Cytogenetic evidence for genomic imprinting

Summary Replication kinetics of the Prader-Willi syndrome critical region (15q11.2) was investigated in seven normal healthy adult females using RBG replication bands. Replication asynchrony between homologs 15q11.2 was identified consistently in about 40% of cells in all individuals. It was limited to the stages in which Xp22, Xp11, Xq13 and Xq24/26 were visible in the late-replicating X chromosome. This asynchrony suggested that replication timing overlapped between 15q11.2 and the early replicating R-bands of the late X chromosome in some cells, and that the difference in replication timing between homologs was probably related to genomic imprinting; the latter has been suggested as a pathogenetic basis of Prader-Willi syndrome. As a result of an analysis of the proportions of asynchronous and synchronous cells in each replication stage, two types of cells were deduced providing 11 methylation mosaicism of genomic imprinting was assumed. The first type was composed of cells with normal replication in one homolog and delayed replication in the other. The second type was composed of cells with normal replication in both homologs. Our results provide cytogenetic evidence of methylation mosaicism for mammalian genomic imprinting.     

Caespitenone,a new cyclopropanoid pseudoguaiane and ent-sesquiterpenes from Porella species

Six Porella species and one Macvicaria species have been investigated and a new cyclopropane pseudoguaiane was isolated and its structure elucidated by chemical and spectral evidence. Macvicaria ulophylla and the Porella species, except P. caespitans ssp. setigera, contain the diterpene dialdehyde, perrottetianal A. (+)-Aristolone, (?)-α-eudesmol, and related sesquiterpene hydrocarbons and alcohols, enantiomeric to those found in higher plant, have been isolated from the Porella species.     

Sesquiterpenoids of fourteen Plagiochila species

The distribution of ent-2,3-secoaromadendrane-, ent-aromadendrane-, ent-bicyclogermacrane- and ent-maaliane-type sesquiterpenoids in fourteen Plagiochila species is described. These sesquiterpenes are the significant chemosystematic markers of Plagiochila. The intense pungent substance of some Plagiochila species is due to an ent-2,3-secoaromadendrane-type sesquiterpene hemiacetal, plagiochiline A.     

Dehydropinguisanin,dehydropinguisenol and pinguisenal,three pinguisane-type sesquiterpenes from Trocholejeunea sandvicensis

Three new pinguisane-type sesquiterpenes, dehydropinguisanin, dehydropinguisenol and pinguisenal together with the previously known deoxopinguisone, pinguisanin, pinguisanolide and (—)-bicyclogermacrene, have been isolated from the liverwort, Trocholejeunea sandvicensis, and their structures have been established from spectral evidence.