The effect of TAK-778 on gene expression of osteoblastic cells is mediated through estrogen receptor.
This study evaluated the effect of TAK-778 [(2R, 4S)-(-)-N-(4-diethoxyphosphorylmethylphenyl)-1,2,4,5-tetrahydro-4-methyl-7,8-methylenedioxy-5-oxo-3-benzothiepin-2-carboxamide)] on in vitro osteogenic events and on gene expression of osteoblastic cells derived from human alveolar bone and the participation of estrogen receptors (ERs) on such effect. Osteoblastic cells were subcultured, with or without TAK-778 (10(-5) M), to evaluate cell growth and viability, total protein content, and alkaline phosphatase (ALP) activity at 7, 14, and 21 days; bone-like formation at 21 days; and gene expression, using cDNA microarray, at 7 days. Also, osteoblastic cells were exposed to TAK-778 (10(-5) M) combined to ICI182,780, a nonspecific ER antagonist (10(-6) M), and gene expression was evaluated by real-time polymerase chain reaction (PCR) at 7 days. TAK-778 induced a reduction in culture growth and an increase in cell synthesis, ALP activity, and bone-like formation. The cDNA microarray showed genes associated with cell adhesion and differentiation, skeletal development, ossification, and transforming growth factor-beta receptor signaling pathway, with a tendency to be higher expressed in cells exposed to TAK-778. The gene expression of ALP, osteocalcin, Msh homeobox 2, receptor activator of NF-kappa B ligand, and intercellular adhesion molecule 1 was increased by TAK-778 as demonstrated by real-time PCR, and this effect was antagonized by ICI182,780. The present results demonstrated that TAK-778 acts at a transcriptional level to enhance the in vitro osteogenic process and that its effect on gene expression of osteoblastic cells is mediated, at least partially, through ERs. Based on these findings, TAK-778 could be considered in the treatment of bone metabolic disorders.
Bellesini
Larissa S
LS
Cell Culture Laboratory, Molecular Immunogenetics Group, Department of Genetics, School of Dentistry of Ribeirao Preto, University of Sao Paulo, Av do Cafe, s/n, 14040-904-Ribeirao Preto, SP, Brazil.
Beloti
Marcio M
MM
Crippa
Grasiele E
GE
Bombonato-Prado
Karina F
KF
Junta
Cristina M
CM
Marques
Marcia M
MM
Passos
Geraldo A
GA
Rosa
Adalberto L
AL
D001582:Benzothiepins / Q000494:pharmacology*; D001842:Bone and Bones / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis; D010006:Osteoblasts / Q000187:drug effects* / Q000378:metabolism*; D010012:Osteogenesis / Q000187:drug effects; D011960:Receptors, Estrogen / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D001582:Benzothiepins; D011960:Receptors, Estrogen; C120450:TAK 778