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Isopsoralen Induces Differentiation of Prechondrogenic ATDC5 Cells via Activation of MAP Kinases and BMP-2 Signaling Pathways.

Li L, Eun JS, Nepal M, Ryu JH, Cho HK, Choi BY, Soh Y - Biomol Ther (Seoul) (2012)

Bottom Line: Isopsoralen significantly enhanced the protein expression of BMP-2 in a time-dependent manner.PD98059 and SB 203580, inhibitors of ERK and p38 MAPK, respectively, decreased the number of stained cells treated with isopsoralen.Taken together, these results suggest that isopsoralen mediates a chondromodulating effect by BMP-2 or MAPK signaling pathways, and is therefore a possible therapeutic agent for bone growth disorders.

View Article: PubMed Central - PubMed

Affiliation: Department of Dental Pharmacology, School of Dentistry and Institute of Oral Bioscience, Brain Korea 21 Project, Chonbuk National University, Jeonju 561-756 ; College of Pharmacy, Woosuk University, Wonju 565-701.

ABSTRACT
Endochondral bone formation is the process by which mesenchymal cells condense to become chondrocytes, which ultimately form new bone. The process of chondrogenic differentiation and hypertrophy is critical for bone formation and as such is regulated by many factors. In this study, we aimed to indentify novel factors that regulate chondrogenesis. We investigated the possible role of isopsoralen in induction of chondrogenic differentiation in clonal mouse chondrogenic ATDC5 cells. Isopsoralen treatment stimulated the accumulation of cartilage nodules in a dose-dependent manner. Further, ATDC5 cells treated with isopsoralen were stained more intensely with Alcian blue than control cells, suggesting that isopsoralen increases the synthesis of matrix proteoglycans. Similarly, isopsoralen markedly induced the activation of alkaline phosphatase activity compared with control cells. Isopsoralen enhanced the expressions of chondrogenic marker genes such as collagen II, collagen X, OCN, Smad4 and Sox9 in a time-dependent manner. Furthermore, isopsoralen induced the activation of extracellular signal-regulated kinase (ERK) and p38 MAP kinase, but not that of c-jun N-terminal kinase (JNK). Isopsoralen significantly enhanced the protein expression of BMP-2 in a time-dependent manner. PD98059 and SB 203580, inhibitors of ERK and p38 MAPK, respectively, decreased the number of stained cells treated with isopsoralen. Taken together, these results suggest that isopsoralen mediates a chondromodulating effect by BMP-2 or MAPK signaling pathways, and is therefore a possible therapeutic agent for bone growth disorders.

No MeSH data available.


Related in: MedlinePlus

Effects of Isopsoralen on proliferation and differentiation of ATDC5 cells. Cell Viability was measured with MTT assay (A). Un-treated cells or cells treated with 0.05 μg/ml Isopsoralen or 10 μg/ml insulin were incubated for 14 days, rinsed with phosphate buffered saline twic,e fixed with 95% methanol, and then stained for 16 h with 1% Alcian blue. Cells were washed with 3% acetic acid for 30 s three, times and then photographed (×100) (B). Cells were plated (5×104) cells per well 24-multiwell plates and treated with 0, 0.01, 0.05, 0.5, and 1 μM Isopsoralen for 14 days. Stained cells were dissolved in 10% acetic acid for subsequent quantification of the absorbance at 650 nm (C). The histogram represents the levels of Alcian blue staining compared to the control group (D). The asterisk (*) indicates a significant difference (p<0.05) compared to the control. Each histogram represents the mean ± S.E.M. (n=3).
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Figure 002: Effects of Isopsoralen on proliferation and differentiation of ATDC5 cells. Cell Viability was measured with MTT assay (A). Un-treated cells or cells treated with 0.05 μg/ml Isopsoralen or 10 μg/ml insulin were incubated for 14 days, rinsed with phosphate buffered saline twic,e fixed with 95% methanol, and then stained for 16 h with 1% Alcian blue. Cells were washed with 3% acetic acid for 30 s three, times and then photographed (×100) (B). Cells were plated (5×104) cells per well 24-multiwell plates and treated with 0, 0.01, 0.05, 0.5, and 1 μM Isopsoralen for 14 days. Stained cells were dissolved in 10% acetic acid for subsequent quantification of the absorbance at 650 nm (C). The histogram represents the levels of Alcian blue staining compared to the control group (D). The asterisk (*) indicates a significant difference (p<0.05) compared to the control. Each histogram represents the mean ± S.E.M. (n=3).

Mentions: We evaluated the effect of isopsoralen on ATDC5 proliferation and differentiation in vitro by light microscopy and assessments of matrix proteoglycan formation. In addition, cell growth was evaluated with the treatment of various concentrations of isopsoralen for 24 h, and by MTT assay. Isopsoralen did not affect the growth rates of ATDC5 cells, as shown in Fig. 2A.


Isopsoralen Induces Differentiation of Prechondrogenic ATDC5 Cells via Activation of MAP Kinases and BMP-2 Signaling Pathways.

Li L, Eun JS, Nepal M, Ryu JH, Cho HK, Choi BY, Soh Y - Biomol Ther (Seoul) (2012)

Effects of Isopsoralen on proliferation and differentiation of ATDC5 cells. Cell Viability was measured with MTT assay (A). Un-treated cells or cells treated with 0.05 μg/ml Isopsoralen or 10 μg/ml insulin were incubated for 14 days, rinsed with phosphate buffered saline twic,e fixed with 95% methanol, and then stained for 16 h with 1% Alcian blue. Cells were washed with 3% acetic acid for 30 s three, times and then photographed (×100) (B). Cells were plated (5×104) cells per well 24-multiwell plates and treated with 0, 0.01, 0.05, 0.5, and 1 μM Isopsoralen for 14 days. Stained cells were dissolved in 10% acetic acid for subsequent quantification of the absorbance at 650 nm (C). The histogram represents the levels of Alcian blue staining compared to the control group (D). The asterisk (*) indicates a significant difference (p<0.05) compared to the control. Each histogram represents the mean ± S.E.M. (n=3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3794527&req=5

Figure 002: Effects of Isopsoralen on proliferation and differentiation of ATDC5 cells. Cell Viability was measured with MTT assay (A). Un-treated cells or cells treated with 0.05 μg/ml Isopsoralen or 10 μg/ml insulin were incubated for 14 days, rinsed with phosphate buffered saline twic,e fixed with 95% methanol, and then stained for 16 h with 1% Alcian blue. Cells were washed with 3% acetic acid for 30 s three, times and then photographed (×100) (B). Cells were plated (5×104) cells per well 24-multiwell plates and treated with 0, 0.01, 0.05, 0.5, and 1 μM Isopsoralen for 14 days. Stained cells were dissolved in 10% acetic acid for subsequent quantification of the absorbance at 650 nm (C). The histogram represents the levels of Alcian blue staining compared to the control group (D). The asterisk (*) indicates a significant difference (p<0.05) compared to the control. Each histogram represents the mean ± S.E.M. (n=3).
Mentions: We evaluated the effect of isopsoralen on ATDC5 proliferation and differentiation in vitro by light microscopy and assessments of matrix proteoglycan formation. In addition, cell growth was evaluated with the treatment of various concentrations of isopsoralen for 24 h, and by MTT assay. Isopsoralen did not affect the growth rates of ATDC5 cells, as shown in Fig. 2A.

Bottom Line: Isopsoralen significantly enhanced the protein expression of BMP-2 in a time-dependent manner.PD98059 and SB 203580, inhibitors of ERK and p38 MAPK, respectively, decreased the number of stained cells treated with isopsoralen.Taken together, these results suggest that isopsoralen mediates a chondromodulating effect by BMP-2 or MAPK signaling pathways, and is therefore a possible therapeutic agent for bone growth disorders.

View Article: PubMed Central - PubMed

Affiliation: Department of Dental Pharmacology, School of Dentistry and Institute of Oral Bioscience, Brain Korea 21 Project, Chonbuk National University, Jeonju 561-756 ; College of Pharmacy, Woosuk University, Wonju 565-701.

ABSTRACT
Endochondral bone formation is the process by which mesenchymal cells condense to become chondrocytes, which ultimately form new bone. The process of chondrogenic differentiation and hypertrophy is critical for bone formation and as such is regulated by many factors. In this study, we aimed to indentify novel factors that regulate chondrogenesis. We investigated the possible role of isopsoralen in induction of chondrogenic differentiation in clonal mouse chondrogenic ATDC5 cells. Isopsoralen treatment stimulated the accumulation of cartilage nodules in a dose-dependent manner. Further, ATDC5 cells treated with isopsoralen were stained more intensely with Alcian blue than control cells, suggesting that isopsoralen increases the synthesis of matrix proteoglycans. Similarly, isopsoralen markedly induced the activation of alkaline phosphatase activity compared with control cells. Isopsoralen enhanced the expressions of chondrogenic marker genes such as collagen II, collagen X, OCN, Smad4 and Sox9 in a time-dependent manner. Furthermore, isopsoralen induced the activation of extracellular signal-regulated kinase (ERK) and p38 MAP kinase, but not that of c-jun N-terminal kinase (JNK). Isopsoralen significantly enhanced the protein expression of BMP-2 in a time-dependent manner. PD98059 and SB 203580, inhibitors of ERK and p38 MAPK, respectively, decreased the number of stained cells treated with isopsoralen. Taken together, these results suggest that isopsoralen mediates a chondromodulating effect by BMP-2 or MAPK signaling pathways, and is therefore a possible therapeutic agent for bone growth disorders.

No MeSH data available.


Related in: MedlinePlus