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Hypoxia Induces Expression and Activation of AMPK in Rat Dental Pulp Cells

¹ Department of Endodontics and
² Department of Biochemistry, Tokyo Dental College, 1-2-2, Masago, Mihama-ku, Chiba City, 261-8502, Japan

View larger version (8K): [in this window] [in a new window]   Figure 1. Effects of hypoxia on cell proliferation. RPC-C2A cells were incubated under normoxic (20% O2) or hypoxic (2% O2) conditions for times indicated. Cell proliferation was analyzed by WST-1 reduction assay, and is expressed as percentage of control. Data represent means ± SD of three independent experiments. *p < 0.05.

View larger version (17K): [in this window] [in a new window]   Figure 2. Expression of AMPK subunit isoforms mRNA. mRNA expression of AMPK subunit isoforms in RPC-C2A was analyzed by qRT-PCR. mRNA level was normalized to internal control, GAPDH, and relative mRNA level to that of 1, which was set at 1.0. Data represent means ± SD of three independent experiments.

View larger version (34K): [in this window] [in a new window]   Figure 3. Effects of hypoxia on expression of major AMPK subunit isoforms and HIF-1. RPC-C2A cells were incubated under hypoxic conditions for the times indicated. Protein expressions of AMPK1, β1 and 1 (A) and HIF-1 (B) were analyzed by Western blotting. Each protein level was normalized to expression of β-actin, and relative protein levels to that at 0 hr are shown. Blot values represent means ± SD of three independent experiments. *p<0.05 vs. 0 hr.

View larger version (20K): [in this window] [in a new window]   Figure 4. Effects of AMPK1 silencing on cell proliferation and AMPK1 and HIF-1 expression. RPC-C2A cells were transfected with control siRNA or siRNA against AMPK1, and incubated for 48 hrs. (A) Expression of AMPK1 protein was analyzed by Western blotting. (B) Cells were then incubated under normoxic or hypoxic conditions for an additional 24 hrs. Cell proliferation was analyzed by WST-1 reduction assay, and is expressed as percentage of control. Data represent means ± SD of three independent experiments. *p < 0.05. (C) After transfection, cells were incubated under normoxic or hypoxic conditions for an additional 4 hrs. Expression of HIF-1 protein was analyzed by Western blotting. N, normoxic conditions; H, hypoxic conditions.

Journal of Dental Research, Vol. 86, No. 9, 903-907 (2007)
DOI: 10.1177/154405910708600919


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