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Project

Investigation of molecular basis of osteoarthritis using developmental models.























Musculoskeletal conditions are the leading cause of morbidity
and disability, resulting in enormous healthcare costs and working disability.
About 150diseases, syndromes and disorders are grouped in the musculoskeletal
conditions, each being progressive and associated with pain ((URL:http://www.who.int)http://www.who.int</>). One of them is OA, a chronic
and progressive joint disorder that is caused by degradation of the articular
cartilage and associated defects of the subchondral bone. Although many
research groups have investigated the onset and progressionof this disabling
condition, the underlying cause or mechanism is far from completely understood.


Therefore, our lab analyzed a protein extract from bovine
articular cartilage with in vivo
chondrogenic activity and isolated the SMOC2 protein. In addition, Smoc2 was shown to be expressed in the
joint interzoneof mouse embryonic digits. Recent evidence indicated that
signaling pathways, which are known to regulate the embryonic stages of
development, are reactivated during the onset and progression of multiple
conditions, including musculoskeletal diseases. Therefore, we used
developmental models to analyze the function of SMOC2.

 

To investigate the SMOC2 associated signaling pathway(s), we
used the zebrafish model system and performed loss and gain of function
experiments. 

Our LOF analysis indicated that Smoc2 affected myeloid
development by reducing the number of macrophages and neutrophils during the
primitive wave of zebrafish hematopoiesis. This defect could be traced back to
the reduced expression of the myeloid progenitor specific gene spi1b at earlier stages of development. In
addition, we detected a reduction in Bmp signaling, which was previously shown
to regulate spi1b expression. This
suggests that Smoc2 affects Bmp signaling in the ALPM, potentially by
manipulation of the Bmp gradient.

Our GOF analysis indicated that Smoc2 affects the DV patterning
and CE movements during gastrulation and the subsequent segmentation stages of
zebrafish development.In addition, Smoc2 appeared to mediate the Wnt5b
mediated non-canonical Wnt signaling pathway.

Although Smoc2 could affect both the Bmp and the Wnt pathway
directly, it is as likely that Smoc2 affects just one of them, which
subsequently affectsthe other. Obviously, the interaction and modulation of
these signaling pathways all depends on the specific microenvironment and the
spatio-temporal context of the smoc2
expressing cells.

 

In order to relate the function of SMOC2 to the homeostasis
of the joint, the zebrafish model is less suited. Therefore, we opted for the
chicken limb model, as the joints in these limbs are more equal to human
joints. To overexpress SMOC2 in the
chicken limb bud, we cloned SMOC2
into theRCAS viral vector that allows for infection of the chicken cells.
Localized injection of this RCASBPA-SMOC2
virus resulted in an outgrowth defect along the PD axis. Our preliminary
analysis of the injected limbs indicated a reduced FGF8 and BMP2 expression
in the AER, which are known to regulate the outgrowth of the limb. However, it
remains unclear whether SMOC2 directly affects FGF transcription or affects the
expression through a secondary partner, like BMP.

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As a third animal model, we have used the mouse model to
analyze the phenotype upon loss of SMOC2. Smoc2-/-
mouse embryos presented a lethal phenotype during early stages of
development. As Smoc2 is only
reported to be expressed in the Reicherts membrane at those stages,
potentially a defectin the Reicherts membrane underlies the lethal phenotype.

</> 

</>

In conclusion, our work revealed new insights in the
essentialfunction of SMOC2 during zebrafish, chicken and mouse embryonic development,
which allows further research to determine the role of SMOC2 within the
intricate network of signaling cascades.

























</>

Musculoskeletal conditions are the leading cause of morbidity
and disability, resulting in enormous healthcare costs and working disability.
About 150 diseases, syndromes and disorders are grouped in the musculoskeletal
conditions, each being progressive and associated with pain ((URL:http://www.who.int)http://www.who.int</>). One of them is OA, a chronic
and progressive joint disorder that is caused by degradation of the articular
cartilage and associated defects of the subchondral bone. Although many
research groups have investigated the onset and progression of this disabling
condition, the underlying cause or mechanism is far from completely understood.


</>

Therefore our lab analyzed a protein extract from bovine
articular cartilage with in vivo chondrogenic activity and isolated the SMOC2
protein. In addition, Smoc2 was shown
to be expressed in the joint interzone of mouse embryonic digits. Recent
evidence indicated that signaling pathways, which are known to regulate the
embryonic stages of development, are reactivated during the onset and
progression of multiple conditions, including musculoskeletal diseases.
Therefore, we used developmental models to analyze the function of SMOC2.

</>

 

</>

To investigate the SMOC2 associated signaling pathway(s), we
used the zebrafish model system and performed loss and gain of function
experiments. 

</>

Our loss of function analysis indicated that Smoc2 affected
myeloid development by reducing the number of macrophages and neutrophils
during the primitive wave of zebrafish hematopoiesis. This defectcould be
traced back to the reduced expression of the myeloid progenitor specific gene spi1b at earlier stages of development. In
addition, we detected a reduction in Bmp signaling, which was previously shown
to regulate spi1b expression. This
suggests that Smoc2 affects Bmp signaling in the ALPM, potentially by
manipulation of the Bmp gradient.

</>

Our gain of function analysis indicated that Smoc2 affects
the DV patterning and CE movements during the gastrulation and subsequent segmentation
stages of zebrafish development. In addition, Smoc2 appeared to mediate the
Wnt5b mediated non-canonical Wnt signalingpathway.

</>

Although Smoc2 could affect both the Bmp and the Wnt pathway
directly, it is as likely that Smoc2 affects just one of them, which
subsequently affects the other one. Obviously, the interaction and modulation
of these signaling pathways all depend on the specific microenvironment and the
spatio-temporal context of the smoc2
expressing cells.

</>

 

</>

In order to relate the function of SMOC2 to the homeostasis
of the joint, the zebrafish model is less suited. Therefore, we opted for the
chicken limb model, as the joints in these limbs are more equal to human
joints. To overexpress SMOC2 in the
chicken limb bud, we cloned SMOC2
into the RCAS viral vector thatallows for infection of the chicken cells.
Localized injection of this RCASBPA-SMOC2
virus resulted in an outgrowth defect along the proximo-distal axis. Our
preliminary analysis of the injected limbs indicated a reduced FGF8 expression in the AER, which is
known to regulatethe outgrowth of the limb. However, it remains unclear
whether SMOC2directly affects FGF transcription or affects the expression
througha secondary partner, like BMP.

</>

 

</>

As a third animal model, we have used the mouse model to
analyze the phenotype upon loss of SMOC2. Smoc2-/-
mouse embryos presented a lethal phenotype during early stages of
development. As Smoc2 is only
reported to be expressed in the Reicherts membrane at those stages,
potentially a defect in the Reicherts membrane underlies the lethal phenotype.

</>

 

</>

In conclusion, our work revealed new insights in the
essential function of SMOC2 during zebrafish, chicken and mouse embryonic development,
which allows further research to determine the role of SMOC2 within the
intricate network of signaling cascades.





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Date:1 Oct 2009 →  6 Oct 2014
Keywords:Osteoarthritis
Disciplines:Genetics, Gynaecology and obstetrics, Molecular and cell biology, Morphological sciences, Orthopaedics
Project type:PhD project