Effects of alendronate and strontium ranelate on cancellous and cortical bone mass in glucocorticoid-treated adult rats.

We studied the effects of alendronate (Aln) and strontium ranelate (SrR) administration on cancellous and cortical bone in glucocorticoid (GC)-treated rats. Thirty-two 3.5-month male Sprague-Dawley rats were randomized into four groups: age-matched normal control (Nrm), methylprednisolone (Met; 5.0 mg/kg/day, sc, for 5 days/week), Met plus Aln orally (1.0 mg/kg/day), and Met plus SrR orally (900 mg/kg/day). The study period was 9 weeks. DXA was used to evaluate the femoral diaphysis and fifth lumbar vertebra (L5). Histomorphometry was performed in the proximal tibial metaphysis and tibial diaphysis. Met significantly decreased body weight and bone mineral density (BMD) compared with Nrm. Aln and SrR significantly increased body weight and BMD compared with Met. SrR resulted in significantly higher BMD than Aln. Met markedly decreased BV/TV, Tb.Th, and Tb.N and increased Tb.Sp compared with Nrm. Aln and SrR showed significantly increased of BV/TV, Tb.Th, and Tb.N and improved bone architecture. Moreover, Met reduced %Ct.Ar, enlarged %Ma.Ar, and decreased bone formation indices in the periosteum as well as increased ES/BS in the endosteum compared with Nrm. Aln significantly decreased endosteal ES/BS compared with Met. SrR significantly increased %Ct.Ar and bone formation indices in the periosteum as well as the endosteum and decreased endosteal ES/BS compared with Met. Furthermore, SrR led to a significantly higher cancellous and endocortical MS/BS and endocortical bone formation compared with Aln. Our findings suggest SrR at a dose of 900 mg/kg has a greater effect than Aln at 1.0 mg/kg, according to BMD and histomorphometric analysis, in preventing GC-induced osteopenia. Therefore, SrR might be applicable as a bone therapeutic agent to treat secondary osteoporosis in the clinic.

C/EBP alpha:AP-1 leucine zipper heterodimers bind novel DNA elements, activate the PU.1 promoter and direct monocyte lineage commitment more potently than C/EBP alpha homodimers or AP-1.

The basic-region leucine zipper (BR-LZ or bZIP) transcription factors dimerize via their LZ domains to position the adjacent BRs for DNA binding. Members of the C/EBP, AP-1 and CREB/ATF bZIP subfamilies form homodimeric or heterodimeric complexes with other members of the same subset and bind-specific DNA motifs. Here we demonstrate that C/EBPalpha also zippers with AP-1 proteins and that this interaction allows contact with novel DNA elements and induction of monocyte lineage commitment in myeloid progenitors. A leucine zipper swap:gel shift assay demonstrates that C/EBPalpha zippers with c-Jun, JunB or c-Fos, but not with c-Maf or MafB. To evaluate activities of specific homodimers or heterodimers we utilized LZs with acid (LZE) or basic (LZK) residues in their salt bridge positions. C/EBPalphaLZE:C/EBPalphaLZK preferentially binds a C/EBP site, c-JunLZE:c-FosLZK an AP-1 site and C/EBPalphaLZE:c-JunLZK a hybrid element identified as TTGCGTCAT by oligonucleotide selection. In murine myeloid progenitors, C/EBPalpha:c-Jun or C/EBPalpha:c-Fos LZE:LZK heterodimers induce monocyte lineage commitment with markedly increased potency compared with C/EBPalpha or c-Jun homodimers or c-Jun:c-Fos heterodimers, demonstrating a positive functional consequence of C/EBP:AP-1 bZIP subfamily interaction. C/EBPalpha:cJun binds and activates the endogenous PU.1 promoter, providing one mechanism for induction of monopoiesis by this complex.

Tissue inhibitor of metalloproteinase-2 (TIMP-2) expression during cardiac neural crest cell migration and its role in proMMP-2 activation.

Matrix metalloproteinases (MMPs) are important mediators of neural crest (NC) cell migration. Here, we examine the distribution of tissue inhibitor of metalloproteinase (TIMP) -2 and TIMP-3 and test whether manipulating TIMP levels alters chicken cardiac NC cell migration. TIMP-2 mRNA is expressed at stage 11 in the neural epithelium and only in migrating cardiac NC cells. TIMP-3 mRNA is expressed only in the notochord at stage 8 and later in the outflow tract myocardium. Exogenous TIMP-2 increases NC motility in vitro at low concentrations but has no effect when concentrations are increased. In vitro, NC cells express membrane type-1 matrix metalloproteinase (MT1-MMP) and TIMP-2 and they secrete and activate proMMP-2. Antisense TIMP-2 oligonucleotides block proMMP-2 activation, decrease NC cell migration from explants, and perturb NC morphogenesis in ovo. Because TIMP-2 is required for activation of proMMP-2 by MT1-MMP, this finding suggests TIMP-2 expression by cardiac NC cells initiates proMMP-2 activation important for their migration.

Synthetic matrix metalloproteinase inhibitor decreases early cardiac neural crest migration in chicken embryos.

During early embryonic development, cardiac neural crest (NC) cells emerge from the forming neural tube, migrate beneath the ectoderm, enter the pharyngeal arches, and subsequently participate in the septation of the heart. Like tumor cells, NC cells penetrate through basement membranes and invade extracellular matrix during their emigration and migration and, therefore, are liable to use similar invasive mechanisms. Matrix metalloproteinases (MMPs) are a family of zinc proteolytic enzymes known to be important in cell migration and invasion of normal and metastatic cells. In an earlier study, we found that the spatial and temporal distribution pattern of MMP-2 positively correlates with cardiac NC migration, suggesting MMP enzymatic activity may be important in mediating cardiac cell NC migration. To test this hypothesis, a synthetic MMP inhibitor, KB8301, was used to block MMP enzymatic activity during in vitro and in vivo cardiac NC cell migration in chick embryos. Injection of KB8301 into the cell-free space adjacent to the neural tube at the level of the second somite before the NC cells emigrated caused major morphologic anomalies in embryos and disrupted cardiac NC morphogenesis. Unilateral injection of KB8301 at lower concentrations, significantly decreased cardiac NC migration on the injected side compared with the noninjected side and compared with that of the injected controls. This decrease correlated with a decrease in MMP activity in the embryos and was not attributable to differences in embryo size or rate of embryonic development after injection. KB8301 also significantly decreased the rate of NC cell motility and distance NC cells migrated from explanted neural tubes and increased cell area and perimeter. These data suggest that MMP enzymatic activity is an important mediator of early cardiac NC migration and that perturbation of endogenous MMP activity may lead to NC-related congenital defects.

Expression of tissue inhibitor of metalloproteinases (TIMPs) during early cardiac development.

Matrix metalloproteinases (MMPs) mediate cell migration and tissue remodeling and are important in cardiac development. We examined the expression patterns of two MMP inhibitors, tissue inhibitor of metalloproteinase (TIMP)-2 and TIMP-3, during critical stages of cardiac development. Both TIMP-2 and TIMP-3 mRNA were expressed in the endocardium prior to and during early cushion cell formation. TIMP-2 was continually expressed within the outflow tract (OT) and atrioventricular (AV) cushion cells at all stages examined, whereas TIMP-3 mRNA was undetectable in the AV cushion cells soon after their formation. Subsequently, TIMP-3 mRNA disappeared in cushion cells of the distal OT and this loss progressed toward the ventricle until eventually all of the OT cushion cells lacked detectable TIMP-3 transcripts. TIMP-3, but not TIMP-2, was also expressed within remodeling myocardium. Immunocytochemistry confirmed these findings. These observations suggest that TIMP-2 and TIMP-3 have important but unique roles in early cardiac development.

MMP-2 expression during early avian cardiac and neural crest morphogenesis.

Matrix metalloproteinase-type 2 (MMP-2) degrades extracellular matrix, mediates cell migration and tissue remodeling, and is implicated in mediating neural crest (NC) and cardiac development. However, there is little information regarding the expression and distribution of MMP-2 during cardiogenesis and NC morphogenesis. To elucidate the role of MMP-2, we performed a comprehensive study on the temporal and spatial distribution of MMP-2 mRNA and protein during critical stages of early avian NC and cardiac development. We found that ectodermally derived NC cells did not express MMP-2 mRNA during their initial formation and early emigration but encountered MMP-2 protein in basement membranes deposited by mesodermal cells. While NC cells did not synthesize MMP-2 mRNA early in migration, MMP-2 expression was seen in NC cells within the cranial paraxial and pharyngeal arch mesenchyme at later stages but was never detected in NC-derived neural structures. This suggested NC MMP-2 expression was temporally and spatially dependent on tissue interactions or differed within the various NC subpopulations. MMP-2 was first expressed within cardiogenic splanchnic mesoderm before and during the formation of the early heart tube, at sites of active pharyngeal arch and cardiac remodeling, and during cardiac cushion cell migration. Collectively, these results support the postulate that MMP-2 has an important functional role in early cardiogenesis, NC cell and cardiac cushion migration, and remodeling of the pharyngeal arches and cardiac heart tube.

[Morphological research on somatoviscera correlation theory].

The experiment was divided into 2 groups. In one group, the animal's nodose ganglion of vagus nerve was taken out; in another group, the animal's half spinal cord was cut off. The results showed that both somatic ascending fibers and vagus nervous fibers all projected in solitary nucleus, commissural nucleus, trigeminal spinal nucleus and in nucleus medulla oblongalae centralis, i.e., these nuclei not only related to somatic nerves, but also related to splanchnic nerves (vagus nerve), which are considered to be foundation of correlation theory of soma and viscera.