Sunday, January 25, 2009


Differential gene expression by Osterix knockdown in mouse chondrogenic ATDC5 cells.

ATDC5 cells are more look chondrocyte progenitor cells than stem cells.
"Osterix (Osx) is a transcription factor required for osteoblast differentiation during intramembranous and endochondral ossification. In an vitro study, which the effects of Osx gene silencing were examined in mouse chondrogenic ATDC5 cells, chondrocyte marker genes were found to be expressionally downregulated and chondrocyte differentiation to be reduced. On the other hand, in vivo studies based on chondrocyte-specific Osx knockouts demonstrated impaired endochondral bone formation with delayed chondrocyte differentiation and reduced cartilage matrix ossification. We investigate global gene expression profile changes caused by Osx knockdown in ATDC5 chondrocytes. The mRNA expressions of 112 genes were significantly modified by Osx knockdown: 68 genes were upregulated and 44 genes downregulated. Functional categories of gene expression classified by gene ontology demonstrated that genes related to cell adhesion, development, and signal transduction were highly affected by Osx knockdown. The expressions of differential genes, such as Sfrp2, Sema3a, Nox4, Rgs4, Zfp521, Has2, Sox6, Scn2a1, Sirpa, and Thbs2, were validated by quantitative real-time PCR."

"Osx gene silencing in mouse chondrogenic ATDC5 cells caused the downregulated expression of chondrocyte marker genes and reduced chondrocyte differentiation"

"Rgs5, Rgs7, and Rgs10 promote chondrocyte differentiation, Rgs4 has an inhibitory effect."

"In chondrocytes, Zfp521 acts on a downstream target gene of PTHR1 signaling and regulates cellular proliferation and differentiation"

"chondrocyte-specific Has2 inactivation causes skeletal deformities and abnormal organization within chondrocytes"

Genes upregulated in Osx knockdown ATDC5 cells also up in LSJL:
Embigin{down as A430106F12Rik}


Haploinsufficiency of Osterix in Chondrocytes Impairs Skeletal Growth in Mice.

"Osteoblast-specific ablation of Osx using Col1α1-Cre resulted in osteopenia, due to impaired osteoblast differentiation in adult mice. Osx is expressed in chondrocytes. We [examine] the skeletal phenotype of mice with conditional disruption of Osx in Col2α1 expressing chondrocytes. Surprisingly, Cre-positive mice that were homozygous for Osx floxed alleles died after birth. Alician blue and alizarin red staining revealed that the lengths of skeleton, femur and vertebrae were reduced by 21, 26 and 14%, respectively, in the knockout (KO) compared with wild type (WT) mice. In order to determine if haploid insufficiency of Osx in chondrocytes influenced postnatal skeletal growth, we compared skeletal phenotype of floxed heterozygous mice that were Cre-positive or Cre-negative. Body length was reduced by 8%, and areal BMD of total body, femur, and tibia was reduced by 5, 7, and 8% respectively in mice with conditional disruption of one allele of Osx in chondrocytes. Micro-CT showed reduced cortical vBMD and trabecular BV/TV in the femurs of Osxflox/+;col2α1-Cre mice. Histological analysis revealed that the impairment of longitudinal growth was associated with disrupted growth plates in the Osxflox/+;col2α1-Cre mice. Primary chondrocytes isolated from KO embryos showed reduced expression of chondral ossification markers, but elevated expression of chondrogenesis markers. Osx expressed in chondrocytes regulates bone growth in part by regulating chondrocyte hypertrophy."

Loss of Osterix downregulated Col10{up} and MMP13.  Loss of Osterix upregulated Fgfr3, Sox9{up}, TGFBr2, and HES1{up in LSJL}.

Osterix regulates corticalization for longitudinal bone growth via integrin β3 expression.
"Corticalization, coalescence of trabecular bone into the metaphyseal cortex, is important for the longitudinal growth of long bones. However, little is known about the molecular mechanisms controlling corticalization. To understand the molecular mechanisms underlying corticalization, we analyzed osteoblast-specific Osterix-knockout mice (Col-OMT). In control mice, corticalization was initiated after 7 postnatal days, and the number of osteoblasts in the peripheral spongiosa was increased compared to the number in the central spongiosa. In contrast, in Col-OMT mice, corticalization was delayed, and the number of osteoblasts in peripheral zones was unchanged compared to the central zone. Furthermore, femoral length was decreased in Col-OMT mice at 1 month. Because Col-OMT mice exhibited impaired matrix coalescence and osteoblast migration, we evaluated integrin signaling in Col-OMT mice. Osterix bound to the Itgb3 promoter and increased transcription of the Itgb3 gene in osteoblast cells. Interestingly, the inner and outer cortical bones were separated in Itgb3-null mice at postnatal day 7. In Itgb3-null mice, the number of osteoblasts in peripheral zones was not changed, and the femoral length was decreased. Taken together, these results indicate that Osterix regulates corticalization for longitudinal bone growth via the control of integrin β3 expression in osteoblasts. Our findings imply that the ability to control osteoblast function during corticalization may help in the treatment of short stature."

"growth plate development, trabecular bone is formed around calcified cartilage in the ossification zone in a process called endochondral bone formation. The newly formed trabecular bone coalesces at the metaphyseal cortex. This event is called “corticalization” and is the process by which longitudinal bone growth is completed. The corticalization process during early growth is essential for the prediction of trabecular and cortical morphology in adulthood"

"During endochondral bone development, the bone collar, which is formed by intramembranous bone, and trabecular bone, which is formed by endochondral bone, are divided into different compartments."

The bone collar is a cuff of periosteal bone that forms around the diaphysis of the hyaline cartilage model in developing long bones

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