The effects of ERK1/2 inhibitor on the chondrogenesis of bone marrow- and adipose tissue-derived multipotent mesenchymal stromal cells.
This study tested the hypothesis that mitogen-activated protein kinase inhibitors suppress hypertrophy and enhance chondrogenesis during chondrogenesis of multipotent mesenchymal stromal cells (MSCs). The effects of PD98059 (an extracellular signal-regulated kinase-1/2 inhibitor) and SB203580 (a p38 inhibitor) were tested on bone marrow-derived MSCs (BMMSCs) and adipose tissue-derived MSCs (ATMSCs). In vitro pellet cultures were carried out using 2.5 x 10(5) MSCs in a chondrogenic medium containing 5 ng/mL of transforming growth factor-beta(2) (TGF-beta(2)) for BMMSCs, and 5 ng/mL of TGF-beta(2) and 100 ng/mL of bone morphogenetic protein-7 (BMP-7) for ATMSCs. From the 14th day of culture, the pellets were additionally treated with PD98059 or SB203580. After 14 more days of in vitro culture, pellets were harvested for analysis. PD98059 increased DNA content and glycosaminoglycan amount in BMMSCs and ATMSCs, whereas SB203580 had little effect. Collagen type I (COL1A1) mRNA decreased to almost a quarter in BMMSCs treated with PD98059. The mRNA levels of collagen type II (COL2A1) and SRY (sex determining region Y)-box 9 (SOX-9) increased several fold in both cells after PD98059 treatment, whereas SB203580 had only a slight effect. The gene expression of collagen type X (COL10A1) and runt-related transcription factor 2 (Runx-2) decreased by half after PD98059 treatment in BMMSCs, and decreased further in ATMSCs. SB203580 elevated COL10A1 and Runx-2 gene expression in both cell types. Safranin-O staining and immunohistochemistry generally mirrored findings from real-time PCR except for diminished expression of type I collagen in ATMSCS, and more pronounced decrease in type X collagen and Runx-2 in BMMSCs after PD98059 treatment. Our study demonstrated that PD98059 suppressed hypertrophy and promoted chondrogenesis of MSCs, and provides a ground for using them in cartilage tissue engineering.
Kim
Hye-Joung
HJ
Department of Orthopaedics, Dongguk University Ilsan Hospital, Goyang, Republic of Korea.
Im
Gun-Il
GI
D000273:Adipose Tissue / Q000166:cytology*; D000328:Adult; D001854:Bone Marrow Cells / Q000166:cytology*; D002452:Cell Count; D020219:Chondrogenesis / Q000187:drug effects*; D003094:Collagen / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors*; D005419:Flavonoids / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007150:Immunohistochemistry; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D008875:Middle Aged; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors; D039902:Multipotent Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011509:Proteoglycans / Q000096:biosynthesis; D011725:Pyridines / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017154:Stromal Cells / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D005419:Flavonoids; D007093:Imidazoles; D047428:Protein Kinase Inhibitors; D011509:Proteoglycans; D011725:Pyridines; D014157:Transcription Factors; D003094:Collagen; D004247:DNA; D048049:Extracellular Signal-Regulated MAP Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; C093642:SB 203580; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one