TY - JOUR
T1 - TGF-β is not involved in early phase growth inhibition of keratinocytes by 1α,25(OH) 2vitamin D 3
AU - Shirakata, Yuji
AU - Ueno, Hikaru
AU - Hanakawa, Yasushi
AU - Kameda, Kenji
AU - Yamasaki, Kenshi
AU - Tokumaru, Sho
AU - Yahata, Yoko
AU - Tohyama, Mikiko
AU - Sayama, Koji
AU - Hashimoto, Koji
PY - 2004/10/1
Y1 - 2004/10/1
N2 - It has been proposed that transforming growth factor-β (TGF-β) is involved in the growth inhibition of normal human epidermal keratinocytes (NHEK) by 1α,25-dihydoxyvitamin D 3 (1α,25(OH) 2D 3), although this is still controversial because of the difficulty in blocking TGF-β activity completely. To determine whether TGF-β is involved in early phase growth inhibition by 1α,25(OH) 2D 3. TGF-β mRNA was detected by ribonuclease protection assay (RPA), and biological active TGF-β was determined by a luciferase reporter assay. To block intrinsic TGF-β activity completely, we constructed an adenovirus vector expressing a truncated TGF-β type II receptor with a dominant negative effect (AdexTβTR) that blocks TGF-β signal transduction. 1α,25(OH) 2D 3 slightly upregulated TGF-β1 and TGF-β2 after 24 h according to an RPA and a luciferase reporter assay, however growth inhibition by 1α,25(OH) 2D 3 occurred at 6 h. The addition of 10 -6 M of 1α,25(OH) 2D 3 to NHEK infected with AdexTβTR or AdexLacZ (control vector) reduced DNA synthesis to 59.3 and 62.2% at 6 h, respectively. There was no significant difference in cell number after a 3-day incubation with AdexTβTR or AdexLacZ-infected cells treated with 1α,25(OH) 2D 3. Since 1α,25(OH) 2D 3 rapidly inhibits NHEK growth regardless of the prevention of TGF-β signal transduction, TGF-β is not involved in early phase growth inhibition by 1α,25(OH) 2D 3.
AB - It has been proposed that transforming growth factor-β (TGF-β) is involved in the growth inhibition of normal human epidermal keratinocytes (NHEK) by 1α,25-dihydoxyvitamin D 3 (1α,25(OH) 2D 3), although this is still controversial because of the difficulty in blocking TGF-β activity completely. To determine whether TGF-β is involved in early phase growth inhibition by 1α,25(OH) 2D 3. TGF-β mRNA was detected by ribonuclease protection assay (RPA), and biological active TGF-β was determined by a luciferase reporter assay. To block intrinsic TGF-β activity completely, we constructed an adenovirus vector expressing a truncated TGF-β type II receptor with a dominant negative effect (AdexTβTR) that blocks TGF-β signal transduction. 1α,25(OH) 2D 3 slightly upregulated TGF-β1 and TGF-β2 after 24 h according to an RPA and a luciferase reporter assay, however growth inhibition by 1α,25(OH) 2D 3 occurred at 6 h. The addition of 10 -6 M of 1α,25(OH) 2D 3 to NHEK infected with AdexTβTR or AdexLacZ (control vector) reduced DNA synthesis to 59.3 and 62.2% at 6 h, respectively. There was no significant difference in cell number after a 3-day incubation with AdexTβTR or AdexLacZ-infected cells treated with 1α,25(OH) 2D 3. Since 1α,25(OH) 2D 3 rapidly inhibits NHEK growth regardless of the prevention of TGF-β signal transduction, TGF-β is not involved in early phase growth inhibition by 1α,25(OH) 2D 3.
KW - 1α,25(OH) D
KW - Adenovirus vector
KW - Dominant negative TGF-β receptor
KW - Growth inhibition
KW - Keratinocyte
KW - Luciferase reporter assay
KW - RNase protection assay
KW - TGF-β
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U2 - 10.1016/j.jdermsci.2004.07.002
DO - 10.1016/j.jdermsci.2004.07.002
M3 - Article
C2 - 15488704
AN - SCOPUS:5644244250
VL - 36
SP - 41
EP - 50
JO - Journal of Dermatological Science
JF - Journal of Dermatological Science
SN - 0923-1811
IS - 1
ER -