IGF-1-mediated osteoblastic niche expansion enhances long-term hematopoietic stem cell engraftment after murine bone marrow transplantation

The efficiency of hematopoietic stem cell (HSC) engraftment after bone marrow (BM) transplantation depends largely on the capacity of the marrow microenvironment to accept the transplanted cells. While radioablation of BM damages osteoblastic stem cell niches, little is known about their restoration and mechanisms governing their receptivity to engraft transplanted HSCs. We previously reported rapid restoration and profound expansion of the marrow endosteal microenvironment in response to marrow radioablation. Here, we show that this reorganization represents proliferation of mature endosteal osteoblasts which seem to arise from a small subset of high-proliferative, relatively radio-resistant endosteal cells. Multiple layers of osteoblasts form along the endosteal surface within 48 hours after total-body irradiation, concomitant with a peak in marrow cytokine expression. This niche reorganization fosters homing of the transplanted hematopoietic cells to the host marrow space and engraftment of long-term (LT)-HSC. Inhibition of insulin-like growth factor (IGF)-1-receptor tyrosine kinase signaling abrogates endosteal osteoblast proliferation and donor HSC engraftment, suggesting that the cytokine IGF-1 is a crucial mediator of endosteal niche reorganization and consequently donor HSC engraftment. Further understanding of this novel mechanism of IGF-1-dependent osteoblastic niche expansion and HSC engraftment may yield clinical applications for improving engraftment efficiency after clinical HSC transplantation.The efficiency of hematopoietic stem cell (HSC) engraft-ment after bone marrow (BM) transplantation depends largely on the capacity of the marrow microenvironment to accept the transplanted cells. While radioablation of BM damages osteoblastic stem cell niches, little is known about their restoration and mechanisms governing their receptivity to engraft transplanted HSCs. We previously reported rapid restoration and profound expansion of the marrow endosteal microenvironment in response to marrow radioablation. Here, we show that this reorganization represents proliferation of mature endosteal osteoblasts which seem to arise from a small subset of high-proliferative, relatively radio-resistant endosteal cells. Multiple layers of osteoblasts form along the endosteal surface within 48 hours after total body irradiation, concomitant with a peak in marrow cytokine expression. This niche reorganization fosters homing of the transplanted hematopoietic cells to the host marrow space and engraft-ment of long-term-HSC. Inhibition of insulin-like growth factor (IGF)-1-receptor tyrosine kinase signaling abrogates endosteal osteoblast proliferation and donor HSC engraft-ment, suggesting that the cytokine IGF-1 is a crucial mediator of endosteal niche reorganization and consequently donor HSC engraftment. Further understanding of this novel mechanism of IGF-1-dependent osteoblastic niche expansion and HSC engraftment may yield clinical applications for improving engraftment efficiency after clinical HSC transplantation. © AlphaMed Press

The Roles of Indian Hedgehog Signaling in TMJ Formation

The temporomandibular joint (TMJ) is an intricate structure composed of the mandibular condyle, articular disc, and glenoid fossa in the temporal bone. Apical condylar cartilage is classified as a secondary cartilage, is fibrocartilaginous in nature, and is structurally distinct from growth plate and articular cartilage in long bones. Condylar cartilage is organized in distinct cellular layers that include a superficial layer that produces lubricants, a polymorphic/progenitor layer that contains stem/progenitor cells, and underlying layers of flattened and hypertrophic chondrocytes. Uniquely, progenitor cells reside near the articular surface, proliferate, undergo chondrogenesis, and mature into hypertrophic chondrocytes. During the past decades, there has been a growing interest in the molecular mechanisms by which the TMJ develops and acquires its unique structural and functional features. Indian hedgehog (Ihh), which regulates skeletal development including synovial joint formation, also plays pivotal roles in TMJ development and postnatal maintenance. This review provides a description of the many important recent advances in Hedgehog (Hh) signaling in TMJ biology. These include studies that used conventional approaches and those that analyzed the phenotype of tissue-specific mouse mutants lacking Ihh or associated molecules. The recent advances in understanding the molecular mechanism regulating TMJ development are impressive and these findings will have major implications for future translational medicine tools to repair and regenerate TMJ congenital anomalies and acquired diseases, such as degenerative damage in TMJ osteoarthritic conditions

Novel ANKH mutation in a patient with sporadic craniometaphyseal dysplasia

Craniometaphyseal dysplasia is caused by mutations in ANKH (ankylosis, progressive homolog [mouse]) in the majority of cases, and all of the reported mutations are single amino acid changes. Genomic DNA from an affected patient, his biological parents, and a sibling was amplified and ANKH was sequenced. The affected patient had a complex heterozygous mutation in exon 7 (c.936T>C, c.938C>G, c.942-953delTGGTTGACGGAA), predicting p.Try290Gln and p.Trp292-Glu295del. We studied the effect of the predicted mutation on the subcellular distribution of ANKH protein. Immunofluorescent labeling of COS-7 cells transduced with normal or mutant Ank (murine progressive ankylosis), showed that normal Ank localized to both the plasma membrane and cytoplasm, whereas mutant Ank was detected only in the cytoplasmic compartment. We propose that this craniometaphyseal dysplasia mutationcauses a loss of ANKH protein expression and activity in the plasma membrane as a result of aberrant intracellular protein trafficking.This study was supported in part by NIH/NIAMS grant AR50627 to H.D.N

NovelANKHmutation in a patient with sporadic craniometaphyseal dysplasia

Spojeni mlaz je fenomen turbulentnog strujanja koji u sustavima ventilacije nastaje meñusobnom interakcijom dvaju nasuprotnih zračnih mlazova iz susjednih istrujnih otvora te mu je, iako predstavlja važan čimbenik toplinske ugodnosti u zoni boravka, u dosadašnjim istraživanjima u ventilaciji posvećeno vrlo malo pozornosti. Cilj istraživanja je temeljem teorije samosličnosti strujanja u razvijenoj zoni turbulentnog mlaza razviti bezdimenzijski matematički model koji bi omogućio procjenu brzine i intenziteta turbulencije zraka kao veličina o kojima ovisi osjećaj propuha u zoni spojenog mlaza, a koji bi u primjeni bio znatno jeftiniji i jednostavniji od složenih CFD proračuna koji još uvijek ne daju pouzdane rezultate kod ovakvih strujanja. Razvoj modela temelji se na eksperimentalnim mjerenjima brzine i intenziteta turbulencije CTA anemometrom te na teorijskoj analizi strujanja uz primjenu analitičkih i jednostavnih numeričkih metoda. Obzirom na oskudna istraživanja vrtložnog ventilacijskog difuzora za proizvodnju mlaza, odabran je vrtložni radijalni stropni difuzor. Mjerenja su vršena u zoni izotermnog priljubljenog stropnog radijalnog mlaza i u zoni izotermnog spojenog mlaza koji nastaje interakcijom te koji se takoñer, na temelju dokaza drugih autora, smatra radijalnim mlazom te se opisuje modelom strujanja slobodnog radijalnog mlaza usmjerenog prema zoni boravka. \Nalaženje rješenja osrednjenog strujanja radijalnog mlaza, kao unaprjeñenje dosadašnjih rješenja, temelji se na pretpostavci o razdiobi brzine u glavnom smjeru strujanja u obliku Gaussove funkcije, te uzimajući u obzir da turbulentna viskoznost u zoni mlaza nije konstantna. S obzirom da eksperimentalni rezultati govore da spojeni mlaz ima više nego dvostruko veći koeficijent širenja od klasičnog radijalnog mlaza, provedena je analiza primjenjivosti Prandtlovih pretpostavki graničnog sloja te su rješenja osrednjenog strujanja donesena uz primjenu i bez primjene pojednostavljenja graničnog sloja. Uz primjenu ovih pretpostavki došlo se do analitičkog rješenja za razdiobu turbulentne viskoznosti, srednje brzine i turbulentnog naprezanja dok je bez primjene ovih pretpostavki bilo potrebno primijeniti jednostavne numeričke metode. Kinetička energija turbulencije odnosno intenzitet turbulencije dobiven je numeričkim rješenjem transportne jednadžbe kinetičke energije turbulencije pri čemu su, kao modeli turbulencije, korištena rješenja \Navier-Stokesove jednadžbe odnosno turbulentne viskoznosti dobivene uz i bez primjene pretpostavki graničnog sloja. Usporedba razvijenog modela s eksperimentalnim rezultatima govori o dobrom slaganju rezultata srednje brzine, turbulentnog naprezanja i kinetičke energije turbulencije u usporedbi s rezultatima klasičnog mlaza drugih autora. Slaganje teorijskog modela s eksperimentalnim rezultatima intenziteta turbulencije u spojenom mlazu je bolje u slučaju bez primjene pretpostavki graničnog sloja nego u slučaju primjene ovih pretpostavki.Merged jet is a turbulent flow phenomenon which is formed by interaction of two opposed air jets from adjacent ventilation diffusers. Regarding its significant influence on thermal comfort conditions in occupied spaces it received very little attention in former studies. The research in this work aims at developing non-dimensional mathematical model for the assessment of air velocity and turbulence intensity as the parameters of sensation of draught in merged jet. This model is intended to be cheaper and simpler than more complex CFD calculations that still do not provide good results for this kind of flow. Development of the model is based on CTA (constant temperature anemometry) measurements of velocity (speed) and turbulence intensity and on theoretical analysis of the flow using analytical and simple numerical methods. Regarding limited research on vortex diffusers found in literature ceiling vortex ventilation diffuser is used for the production of the jet. Measurements are performed in the zone of isothermal attached ceiling radial jet and in the zone of isothermal merged jet that is formed by the interaction of ceiling jets. Merged jet is considered to be a free radial jet flowing towards the occupied zone. Integral solution for the mean flow profiles of the turbulent radial jet, as the improvement of former solutions, is found with the assumption of Gaussian functional form for the streamwise velocity distribution and the assumption of non-constant turbulent viscosity distribution in the zone of the jet. As the experimental data shows that coefficient of spread for the merged jet is more than twice higher than the coefficient for the classical free radial jet the analysis of applicability of Prandtl shear flow approximations is performed and solutions of the mean flow were derived with and without these approximations. Using these approximations enables derivation of algebraic equations for turbulent viscosity, mean velocity and turbulent shear stress. Solutions without these approximations are numerical and demand use of simple numerical methods. Turbulence kinetic energy and turbulence intensity are obtained by numerical solution of transport equation for turbulent kinetic energy by using solutions of Navier-Stokes equation, that are turbulence viscosity solutions with and without shear flow approximations, as the turbulence models. Agreement of mean velocity, turbulent shear stress and turbulent kinetic energy data of the developed theoretical model with the experimental data of classical jet found in literature is good. Agreement of the developed theoretical model with experimental data for mean velocity of the merged jet is good and agreement with experimental data for turbulence intensity is better in the case of not using approximations of shear flow than in the case of using them

Runx2 regulates FGF2-induced Bmp2 expression during cranial bone development

Calvarial bone is formed by the intramembranous bone-forming process, which involves many signaling molecules. The constitutive activation of the fibroblast growth factor (FGF) signaling pathway accelerates osteoblast differentiation and results in premature cranial suture closure. Bone morphogenetic protein (BMP) signaling pathways, which involve the downstream transcription factors Dlx5 and Msx2, are also involved in the bone-forming processes. However, the relationships between these two main signaling cascades are still unclear. We found that FGF2 treatment of developing bone fronts stimulated Bmp2 gene expression but that BMP2 treatment could not induce Fgf2 expression. Moreover, the disruption of the Runx2 gene completely eliminated the expression of Bmp2 and its downstream genes Dlx5 and Msx2 in the developing primordium of bone, while the expression of Fgf2 was maintained. In addition, cultured Runx2-/- cells expressed very low baseline levels of Bmp2 that were up-regulated by transfection with a Runx2-expressing plasmid. These levels in turn were markedly elevated by FGF2 treatment. FGF2 treatment also strongly enhanced the Bmp2 expression in MC3T3-E1 cells, whose endogenous Runx2 gene is intact and which express Bmp2 at low baseline levels as well. These results indicate that Runx2 is an important mediator of the expression of Bmp2 in response to FGF stimulation in cranial bone development. Developmental Dynamics 233:115-121, 2005. © 2005 Wiley-Liss, Inc

Cleft lip and cleft palate in Esrp1 knockout mice is associated with alterations in epithelial-mesenchymal crosstalk

Cleft lip is one of the most common human birth defects. However, there remain a limited number of mouse models of cleft lip that can be leveraged to characterize the genes and mechanisms that cause this disorder. Crosstalk between epithelial and mesenchymal cells underlies formation of the face and palate, but the basic molecular events mediating this crosstalk remain poorly understood. We previously demonstrated that mice lacking the epithelial-specific splicing factor Esrp1 have fully penetrant bilateral cleft lip and palate. In this study, we further investigated the mechanisms leading to cleft lip as well as cleft palate in both existing and new Esrp1 mutant mouse models. These studies included a detailed transcriptomic analysis of changes in ectoderm and mesenchyme in Esrp1-/- embryos during face formation. We identified altered expression of genes previously implicated in cleft lip and/or palate, including components of multiple signaling pathways. These findings provide the foundation for detailed investigations using Esrp1 mutant disease models to examine gene regulatory networks and pathways that are essential for normal face and palate development - the disruption of which leads to orofacial clefting in human patients