Expression of Osteoblast–Specific Factor 2 (OSF-2, Periostin) Is Associated with Drug Resistance in Ovarian Cancer Cell Lines.
One of the main obstacles to the effective treatment of ovarian cancer patients continues to be the drug resistance of cancer cells. Osteoblast-Specific Factor 2 (OSF-2, Periostin) is a secreted extracellular matrix protein (ECM) expressed in fibroblasts during bone and teeth development. Expression of OSF-2 has been also related to the progression and drug resistance of different tumors.
The present study investigated the role of OSF-2 by evaluating its expression in the primary serous ovarian cancer cell line, sensitive (W1) and resistant to doxorubicin (DOX) (W1DR) and methotrexate (MTX) (W1MR). The OSF-2 transcript (real-time PCR analysis), protein expression in cell lysates and cell culture medium (western blot), and expression of the OSF-2 protein in cell lines (immunofluorescence) were investigated in this study. Increased expression of OSF-2 mRNA was observed in drug-resistant cells and followed by increased protein expression in cell culture media of drug-resistant cell lines.
A subpopulation of ALDH1A1-positive cells was noted for W1DR and W1MR cell lines; however, no direct co-expression with OSF-2 was demonstrated. Both drugs induced OSF-2 expression after a short period of exposure of the drug-sensitive cell line to DOX and MTX.
The obtained results indicate that OSF-2 expression might be associated with the development of DOX and MTX resistance in the primary serous W1 ovarian cancer cell line.
An analysis of skeletal development in <em>osteoblast</em>-<em>specific</em> and chondrocyte-<em>specific</em> runt-related transcription <em>factor</em>-<em>2</em> (Runx<em>2</em>) knockout mice.
Global gene deletion studies in mice and humans have established the pivotal role of runt related transcription factor-2 (Runx2) in both intramembranous and endochondral ossification processes during skeletogenesis. In this study, we for the first time generated mice carrying a conditional Runx2 allele with exon 4, which encodes the Runt domain, flanked by loxP sites. These mice were crossed with α1(I)-collagen-Cre or α1(II)-collagen-Cre transgenic mice to obtain osteoblast-specific or chondrocyte-specific Runx2 deficient mice, respectively.
As seen in Runx2(-/-) mice, perinatal lethality was observed in α1(II)-Cre;Runx2(flox/flox) mice, but this was not the case in animals in which α1(I)-collagen-Cre was used to delete Runx2. When using double-staining with Alizarin red for mineralized matrix and Alcian blue for cartilaginous matrix, we observed previously that mineralization was totally absent at embryonic day 15.5 (E15.5) throughout the body in Runx2(-/-) mice, but was found in areas undergoing intramembranous ossification such as skull and clavicles in α1(II)-Cre; Runx2(flox/flox) mice. In newborn α1(II)-Cre;Runx2(flox/flox) mice, mineralization impairment was restricted to skeletal areas undergoing endochondral ossification including long bones and vertebrae.
In contrast, no apparent skeletal abnormalities were seen in mutant embryo, newborn, and 3-week-old to 6-week old-mice in which Runx2 had been deleted with the α1(I)-collagen-Cre driver.
These results suggest that Runx2 is absolutely required for endochondral ossification during embryonic and postnatal skeletogenesis, but that disrupting its expression in already committed osteoblasts as achieved here with the α1(I)-collagen-Cre driver does not affect overtly intramembranous and endochondral ossification.
The Runx2 floxed allele established here is undoubtedly useful for investigating the role of Runx2 in particular cells.
<em>Osteoblast</em>-<em>specific</em> <em>factor</em> <em>2</em> expression in prostate cancer-associated stroma: identification through microarray technology.
OBJECTIVE
To better understand the gene expression patterns in tumor-associated stroma, laser-capture-microdissections from clinical specimens were analyzed by genome-wide-expression microarray technology.
The epithelial-stromal interaction plays a critical role in prostate development, reactive changes, and tumorigenesis. Diverse microarray technologies have been used to characterize the molecular changes in prostate cancer.
Even though these gene expression studies are compromised by the heterogeneity of the tumor, as well as by the difficulty associated with collecting appropriate counterparts to represent normal prostate cells, the gene array data from tumors have shown promising results. Currently, little is known about the tumor-associated stromal gene expression profile in prostate cancer.
METHODS
Matching benign and malignant epithelial cell-related stroma cells were subjected to microarray platforms.
RESULTS
The prostatatic stroma expressed several osteogenic molecules. In particular, one of the genes, OSF2, was upregulated in tumor-associated stroma compared with benign epithelial cell associated stroma, which was further validated by immunohistochemical examination.
CONCLUSIONS
These data show that the combination of laser capture dissection with computational enhancement of epithelial and stromal microarray data is a useful tool to assess gene expression changes in prostate cancer stroma.
[The interaction between secreted frizzled-related protein <em>2</em> and <em>osteoblast</em>-<em>specific</em> <em>factor</em> <em>2</em> in keloid].
OBJECTIVE
To verify the interaction between secreted frizzled-related protein 2 (SFRP2) and osteoblast-specific factor 2 (OSF-2).
METHODS
HA-tagged OSF-2 fusion protein recombinant vector pCMV-HA-OSF-2, which could express in mammal cells was constructed, then identified by enzyme-cutting and transfected into human kidney 293 (HK293) cells with or without Myc-SFRP2 recombinant eukaryotic expression vector pCMV-HA-SFRP2.
The interaction between SFRP2 and OSF-2 was detected through coimmunoprecipitation and Western blotting.
RESULTS
In electrophoresis bath, target fragment of SFRP2 coding gene with 800 bp and target gene OSF-2 with 2500 bp could be seen respectively after enzyme-cutting, which showed that pCMV-Myc-SFRP2 and pCMV-HA-OSF-2 were constructed successfully.
No HA-OSF-2 expression was detected after pCMV-Myc-SFRP2 or pCMV-HA-OSF-2 transfection. Whereas, HA-OSF-2 expressed by Myc antibody immunoprecipitation after pCMV-Myc-SFRP2 and pCMV-HA-OSF-2 co-transfection.
CONCLUSIONS
HA-OSF-2 recombinant vector can express in mammal cells. Interaction exists between HA-OSF-2 and SFRP2.
<em>Factors</em> influencing effects of <em>specific</em> COX-<em>2</em> inhibitor NSAIDs on growth and differentiation of mouse <em>osteoblasts</em> on titanium surfaces.
OBJECTIVE
To investigate the influence of exposure time and stages of cell growth on the effects of specific COX-2 inhibitor NSAIDs on growth and differentiation of osteoblasts on smooth titanium surfaces.
METHODS
The study was categorized into 5 groups: group A, 0.1 microM indomethacin; group B, 1.5 microM celecoxib; group C, 3.0 microM celecoxib; group D, 9.0 microM celecoxib; and group E, serum-free culture medium without drug treatment.
A mouse calvarial cell line, MC3T3-E1, was seeded on acid-prickled surface titanium disks. The investigations were performed in 3 experimental phases based on stages of cell growth: static (24 hours after seeding), log (culture day 5), and plateau (culture day 12). In each experimental phase, cells on titanium disks were incubated in a medium treated with drugs according to the groups of study for 1, 3, and 5 days.
RESULTS
Indomethacin and celecoxib in groups A to D inhibited growth of cells on treatment days 3 and 5 in static phase and on treatment day 3 in log phase. Additionally, an inhibitory effect of indomethacin was greater than celecoxib. Effects on alkaline phosphatase (ALP) activity and osteocalcin were not clearly demonstrated. A significant decrease of PGE2 production was found in groups A to D in static and plateau but not log phases.
CONCLUSIONS
A specific COX-2 inhibitor NSAID, celecoxib, inhibited growth of osteoblasts on titanium surfaces and the effects were influenced by exposure time and stages of cell growth. Using a specific COX-2 inhibitor might cause deterioration of osteointegration of dental implants by interfering with osteoblastic cell growth in the proliferative stage.
General transcription <em>factor</em> IIA-gamma increases <em>osteoblast</em>-<em>specific</em> osteocalcin gene expression via activating transcription <em>factor</em> 4 and runt-related transcription <em>factor</em> <em>2</em>.
ATF4 (activating transcription factor 4) is an osteoblast-enriched transcription factor that regulates terminal osteoblast differentiation and bone formation. ATF4 knock-out mice have reduced bone mass (severe osteoporosis) throughout life. Runx2 (runt-related transcription factor 2) is a runt domain-containing transcription factor that is essential for bone formation during embryogenesis and postnatal life.
In this study, we identified general transcription factor IIA gamma (TFIIA gamma) as a Runx2-interacting factor in a yeast two-hybrid screen. Immunoprecipitation assays confirmed that TFIIA gamma interacts with Runx2 in osteoblasts and when coexpressed in COS-7 cells or using purified glutathione S-transferase fusion proteins.
Chromatin immunoprecipitation assay of MC3T3-E1 (clone MC-4) preosteoblast cells showed that in intact cells TFIIA gamma is recruited to the region of the osteocalcin promoter previously shown to bind Runx2 and ATF4.
A small region of Runx2 (amino acids 258-286) was found to be required for TFIIA gamma binding. Although TFIIA gamma interacts with Runx2, it does not activate Runx2. Instead, TFIIA gamma binds to and activates ATF4.
Periostin / Osteoblast Specific Factor 2 (Postn) Antibody |
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| 20-abx114393 | Abbexa |
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Periostin / Osteoblast Specific Factor 2 (Postn) Antibody |
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| abx114393-100l | Abbexa | 100 µl | 612.5 EUR |
Human Periostin (Osteoblast Specific Factor 2) ELISA Kit |
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| MBS286029-10x96StripWells | MyBiosource | 10x96-Strip-Wells | 6255 EUR |
Human Periostin (Osteoblast Specific Factor 2) ELISA Kit |
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| MBS286029-48StripWells | MyBiosource | 48-Strip-Wells | 445 EUR |
Human Periostin (Osteoblast Specific Factor 2) ELISA Kit |
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| MBS286029-5x96StripWells | MyBiosource | 5x96-Strip-Wells | 3290 EUR |
Human Periostin (Osteoblast Specific Factor 2) ELISA Kit |
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| MBS286029-96StripWells | MyBiosource | 96-Strip-Wells | 730 EUR |
Rat periostin/osteoblast specific factor 2 (POSTN) ELISA kit |
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| EKC39533-48T | Biomatik Corporation | 48T | 474.04 EUR |
Rat periostin/osteoblast specific factor 2 (POSTN) ELISA kit |
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| EKC39533-5x96T | Biomatik Corporation | 5x96T | 3216.7 EUR |
Rat periostin/osteoblast specific factor 2 (POSTN) ELISA kit |
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| EKC39533-96T | Biomatik Corporation | 96T | 677.2 EUR |
Rat periostin/osteoblast specific factor 2 (POSTN) ELISA kit |
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| CSB-EL018381RA-24T | Cusabio | 1 plate of 24 wells | 198 EUR |
Rat periostin/osteoblast specific factor 2 (POSTN) ELISA kit |
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| 1-CSB-EL018381RA | Cusabio |
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Rat periostin;osteoblast specific factor 2,POSTN ELISA KIT |
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| E2192Ra-1096T | Jiaxing Korain Biotech Ltd (BT Labs) | 10*96T | 4122 EUR |
Rat periostin;osteoblast specific factor 2,POSTN ELISA KIT |
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| E2192Ra-48wells | Jiaxing Korain Biotech Ltd (BT Labs) | 48 wells | 300 EUR |
Furthermore, TFIIA gamma together with ATF4 and Runx2 stimulates osteocalcin promoter activity and endogenous mRNA expression. Small interfering RNA silencing of TFIIA gamma markedly reduces levels of endogenous ATF4 protein and Ocn mRNA in osteoblastic cells. Overexpression of TFIIA gamma increases levels of ATF4 protein.
Finally, TFIIA gamma significantly prevents ATF4 degradation. This study shows that a general transcription factor, TFIIA gamma, facilitates osteoblast-specific gene expression through interactions with two important bone transcription factors ATF4 and Runx2.