Regulation of Human Osteosarcoma Cell Differentiation by Transforming Growth Factor B1 (TGFB and 1a,25- Dihydroxyvitamin D3 : the Mechanism of TGFB1 Potentiation of 1,25D3-induced Alkaline Phosphatase Activity in Human MG63 Osteosarcoma Cell

<p>Transforming growth factor β₁(TGFβ₁) is a pluripotent cytokine that works in conjunction with other cytokines to modulate their effects on cell function. Our previous work demonstrated that the combination of TGFβ₁ and 1α,25-dihydroxyvitamin D₃ had a synergistic effect on alkaline phosphatase induction (500 to 700%) in human osteoblastic osteosarcoma cells (MG63). The current studies focused on the molecular basis of alkaline phosphatase induction, including the regulation of the signal transduction and modification of 1α,25-dihydroxyvitamin D₃ receptors (VDR). We demonstrated that alkaline phosphatase activity was regulated at the transcription level. TGFβ₁ at 1 ng/ml alone had little effect on alkaline phosphatase mRNA induction and 1α,25-dihydroxy vitamin D₃ at 10^-8 M increased alkaline phosphatase mRNA to about 200% over the control value. The combination of TGFβ₁ and 1α,25-dihydroxyvitamin D₃ significantly increased alkaline phosphatase mRNA about 800% over the control. The peak response was at 24 hours. Studies of the signal transduction mechanism provided evidence that the protein kinase C pathway was involved in the alkaline phosphatase stimulation by the combination of TGFβ₁ and 1α,25-dihydroxyvitamin D3; MG63 cells express at least seven protein kinase C isoforms and PKC_y was activated by TGFβ₁ or 1α,25-dihydroxyvitamin D₃ after 30 minute treatment. Studies on the VDR demonstrated that TGFβ₁ or its combination with 1α,25-dihydroxyvitamin D₃ significantly increased VDR mRNA about 300% and 500% respectively over the control by 3 hours. 1α,25-dihydroxyvitamin D₃ increased VDR mRNA about 200% by 24 hour. The VDR protein level was also significantly elevated by the combination of TGFβ₁ with 1α,25-dihydroxyvitamin D3. The peak response at 6 hours was 300% above the control level. Our studies further showed that a 2 hour treatment with TGFβ₁ and 1α,25-dihydroxy vitamin D₃ significantly stimulated VDR phosphorylation to 700% and 300% above the control, respectively. When cells were treated with the combination of TGFβ₁ and 1α,25-dihydroxyvitamin D_3, there was an additive effect on VDR phosphorylation to about 1000% over the control value. Conclusions: (1) PKC_y is involved in the TGFβ and 1α, 25-dihydroxyvitamin D₃ signal transduction pathway. (2) TGFβ potentiates the action of 1α,25-dihydroxyvitamin D₃ by stimulation of VDR mRNA, VDR protein and VDR phosphorylation. These changes further enhance the effect of 1α,25-dihydroxy vitamin D₃ on the cellular levels of alkaline phosphatase mRNA and alkaline phosphatase activity.</p>

Name thesis:degree_name

Doctor of Philosophy (PhD)

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Biochemistry

Year

1996

Author dc:creator

• Song, XinDe

Contributors dc:contributor

• Jon E. Wergedal
• Thomas A. Linkhart
• Lora M. Green
• E. Clifford Herrmann
• Kelvin A. W. Hill

• Biochemistry
• Osteosarcoma; Transforming Growth Factor beta; Calcitriol

Statement dc:rights

• This title appears here courtesy of the author, who has granted Loma Linda University a limited, non-exclusive right to make this publication available to the public. The author retains all other copyrights.

Language dc:language

English

Repository record dc:identifier

https://scholarsrepository.llu.edu/etd/987

OAI identifier oai:identifier

oai:scholarsrepository.llu.edu:etd-2057