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Versican G3 domain enhanced tumor cell survival in serum free medium by up-regulating pERK.

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posted on 2011-11-09, 01:21 authored by William Weidong Du, Burton B. Yang, Bing L. Yang, Zhaoqun Deng, Ling Fang, Sze Wan Shan, Zina Jeyapalan, Yaou Zhang, Arun Seth, Albert J. Yee

a) G3- and vector-transfected 66c14 (2×105) 2×105 were cultured in 10% FBS/DMEM medium in culture dishes for 12 hours. After cell attachment, we changed the medium to serum free DMEM and cultured them for 6 days. Cell lysates were prepared and subjected to immunoblotting with antibodies to ERK2, pERK, GSK-3β (S9P), GSK-3β, CDK2, CDK6, CDC2P34, and β-actin. b) G3-transfected and vector-transfected 66c14 cells (1×104) were inoculated and cultured in 10% FBS/DMEM medium in 96 well culture dishes for 12 hours. After cell attachment, we changed the medium to serum free DMEM, with EGF (20 ng/ml), selective EGFR inhibitor AG 1478 (2.0 µM), selective MEK (ERK kinase) inhibitor PD 98059 (50 µM), or selective JNK inhibitor SP 600125 (100 nM) cultured for 6 days with medium changed every 2 days. WST-1 Cell Survival Assays were used to analyze cell viability. Compared with vector control group, n = 6, * p<0.05, **p<0.01, analyzed with t-test. c) After culture in serum free medium for 6 days, cell lysates were prepared and subjected to immunoblotting with antibodies to ERK2, pERK, GSK-3β (S9P), GSK-3β, and β-actin.

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