Journal: Neural regeneration research

Article Title: Electroacupuncture preconditioning attenuates ischemic brain injury by activation of the adenosine monophosphate-activated protein kinase signaling pathway.

doi: 10.4103/1673-5374.160095

Figure Lengend Snippet: Figure 3 Western blot analysis of protein expression of AMPKα (A) and p-AMPKα (B) in the hippocampal CA1 region at 72 hours after BCCAO. Data are presented as the mean ± SD (n = 6 mice in each group), and were analyzed by one-way analysis of variance followed by post hoc Stu- dent-Newman-Keuls test. Values represent the optical density ratio of target protein to GAPDH. Experiments were performed in triplicate. *P < 0.05, vs. BCCAO group; #P < 0.05, vs. EA + BCCAO group. AMPKα: Adenosine monophosphate-activated protein kinase α; p-AMPKα: phosphorylated adenosine monophosphate-activated protein kinase α; EA: electroacupuncture; BCCAO: bilateral common carotid artery occlusion; CC: com- pound C (an adenosine monophosphate-activated protein kinase antagonist).

Article Snippet: Membranes were blocked in 5% fat-free milk prepared in Tris-buffered saline/Tween-20 buffer for 1 hour, and incubated with primary monoclonal antibodies for rabbit anti-mouse AMPKα (1:1,000; Abcam, Cambridge, UK), rabbit anti-mouse phosphorylated-AMPKα (p-AMPKα) (1:1,000; Cell Signaling Technology, Danvers, MA, USA), and GAPDH (1:500; rabbit anti-mouse; Zhongshan Goldenbridge Biotechnology, Beijing, China) overnight at 4°C.

Techniques: Western Blot, Expressing

Journal: Human Molecular Genetics

Article Title: Superoxide dismutating molecules rescue the toxic effects of PINK1 and parkin loss

doi: 10.1093/hmg/ddy069

Figure Lengend Snippet: PINK1 - and Parkin -editing in HeLa and HEK-293T cells. ( A ) Schematic representation of the gRNA target sequences in PINK1 gene. Arrows indicate Cas9 DNA cutting sites. ( B ) Western blot analysis of PINK1 in HeLa cells transiently expressing scramble, CRISPR-PINK1-A or CRISPR-PINK1-B plasmids. GAPDH signal was used as loading control. ( C ) Densitometric analysis of PINK1 levels normalized to GAPDH expressed in percentage. Data are expressed as mean ± SEM of three independent experiments. ( D ) Schematic representation of the gRNA target sequences in parkin gene. Arrows indicate Cas9 DNA cutting sites. ( E ) Western blot analysis of Parkin in HEK-293T cells transiently expressing scramble, CRISPR-Parkin-A or CRISPR-Parkin-B plasmids. GAPDH signal was used as loading control. ( F ) Densitometric analysis of Parkin levels normalized to GAPDH expressed in percentage. Data are expressed as mean ± SEM of three independent experiments.

Article Snippet: The following antibodies were used: rabbit monoclonal anti-PINK1 antibody (D8G3, Cell Signalling Technology), mouse monoclonal anti-parkin antibody (sc-32282, Santa Cruz Biotechnology), rabbit Polyclonal anti-GAPDH antibody (Origene), anti-mouse and anti-rabbit IgG peroxidase antibodies (Sigma), Alexa Fluor ® 488 anti-human CD4 antibody (clone OKT4, BioLegend) and Alexa Fluor ® 647 anti-human CD4 antibody (clone OKT4, BioLegend).

Techniques: Western Blot, Expressing, CRISPR, Control

Journal: Bioscience Reports

Article Title: Insulin down-regulates the expression of ubiquitin E3 ligases partially by inhibiting the activity and expression of AMP-activated protein kinase in L6 myotubes

doi: 10.1042/bsr20150017

Figure Lengend Snippet: Figure 1 Effects of AICAR and compound C on p-AMPK, p-ACC, MAFbx and MuRF1 Effects of AICAR and compound C (CC) on p-AMPK, p-ACC (A) and mRNA expression of MAFbx and MuRF1 (B) in L6 myotubes. L6 myotubes were pre-treated with 20 μM CC or without CC for 1 h, followed by treatment with AICAR (1 mM) for 30 min (A) or 24 h (B). (A) After stimulation with AICAR for 30 min, p-AMPK at Thr172 and p-ACC at Ser79 were measured by western blotting, with GAPDH as the internal standard. Shown are representative blots from three independent experiments. (B) At 24 h after AICAR treatment, extracted RNA from L6 myotubes were assayed with qRT-PCR. Data were normalized to GAPDH and the values for the control group set at 1.0. Data are expressed as means +−S.D. (n = 3). *P < 0.05 compared with control. #P < 0.05 compared with AICAR treatment group.

Article Snippet: Equal protein loading was determined by stripping each membrane and re-probing for GAPDH (glyceraldehyde-3phosphate dehydrogenase, Zhong Shan Golden Bridge Biotechnology).

Techniques: Expressing, Western Blot, Quantitative RT-PCR, Control

Journal: Bioscience Reports

Article Title: Insulin down-regulates the expression of ubiquitin E3 ligases partially by inhibiting the activity and expression of AMP-activated protein kinase in L6 myotubes

doi: 10.1042/bsr20150017

Figure Lengend Snippet: Figure 2 Effects of insulin on p-AMPK, p-ACC, MAFbx and MuRF1 Effects of insulin on p-AMPK, p-ACC (A) and mRNA expression of MAFbx and MuRF1 (B) in L6 myotubes. L6 myotubes were pre-treated with AICAR (1 mM), compound C (CC; 20 μM) or not with AICAR and CC for 1 h, followed by addition of insulin (100 nM) for 30 min (A) or 24 h (B). (A) At 30 min, p-AMPK and p-ACC were measured by western blotting, with GAPDH as the internal standard. Shown are representative blots from three independent experiments. (B) At 24 h, extracted RNA from L6 myotubes were assayed with qRT-PCR. Data were normalized to GAPDH and the values for the control group set at 1.0. Data are expressed as means +−S.D. (n = 3). *P < 0.05 compared with control. #P < 0.05 compared with insulin only treatment group.

Article Snippet: Equal protein loading was determined by stripping each membrane and re-probing for GAPDH (glyceraldehyde-3phosphate dehydrogenase, Zhong Shan Golden Bridge Biotechnology).

Techniques: Expressing, Western Blot, Quantitative RT-PCR, Control

Journal: Bioscience Reports

Article Title: Insulin down-regulates the expression of ubiquitin E3 ligases partially by inhibiting the activity and expression of AMP-activated protein kinase in L6 myotubes

doi: 10.1042/bsr20150017

Figure Lengend Snippet: Figure 3 Effect of Akt inhibitor on inhibition of p-AMPK and p-ACC by insulin The effect of Akt inhibitor (Akt inhibitor IV) on inhibition of p-AMPK and p-ACC by insulin in L6 myotubes. L6 myotubes were pre-treated with Akt inhibitor IV (1 μM) for 1 h, followed by addition of insulin (100 nM) or without insulin for 30 min. p-AMPK, p-ACC and phosphor-Akt (p-Akt) were measured by western blotting, with GAPDH as the internal standard. Shown are representative blots from three independent experiments.

Article Snippet: Equal protein loading was determined by stripping each membrane and re-probing for GAPDH (glyceraldehyde-3phosphate dehydrogenase, Zhong Shan Golden Bridge Biotechnology).

Techniques: Inhibition, Western Blot

Journal: Bioscience Reports

Article Title: Insulin down-regulates the expression of ubiquitin E3 ligases partially by inhibiting the activity and expression of AMP-activated protein kinase in L6 myotubes

doi: 10.1042/bsr20150017

Figure Lengend Snippet: Figure 4 Effect of insulin on AMPK α1 and α2 Effects of insulin on mRNA expression of AMPK α1 and α2. L6 myotubes were either untreated (control) or pre-treated with 1 mM AICAR or 20 μM CC or not with AICAR and CC for 1 h, followed by stimulation with 100 nM insulin for 24 h. Extracted RNA from L6 myotubes were assayed with qRT-PCR. Data were normalized to GAPDH and the values for the control group set at 1.0. Values are expressed as means +−S.D. (n = 3). *P < 0.05 compared with control. #P < 0.05 compared with insulin only group.

Article Snippet: Equal protein loading was determined by stripping each membrane and re-probing for GAPDH (glyceraldehyde-3phosphate dehydrogenase, Zhong Shan Golden Bridge Biotechnology).

Techniques: Expressing, Control, Quantitative RT-PCR