xbp1 antibodies sc-7160  (Santa Cruz Biotechnology)

Journal: PLoS ONE

Article Title: L-lysine protects C2C12 myotubes and 3T3-L1 adipocytes against high glucose damages and stresses

doi: 10.1371/journal.pone.0225912

Figure Lengend Snippet: The gel electrophoresis pattern of the spliced and unspliced (XBP-1s and XBP-1u, respectively) amplicons as studied by RT-PCR in (A) C2C12 myoblasts and (B) 3T3-L1 adipocytes with no treatment (control), in the presence of HG or HG+Lys after 2, 4 and 6 h of incubation. The semiquantitative analysis of the XBP1s and XBP1u mRNA values, obtained by ImageJ analysis, and the ratio of XBP1s/XBP1u mRNA are shown in figures (C) C2C12 myoblasts and (D) 3T3L1 adipocytes. The data represent as mean ± SD of three independent experiments. The data of the named four independent groups at different time intervals were analyzed by repeated-measures ANOVA. The significant differences (p-value) in the XBP1 mRNA splicing within the groups at different time intervals are shown in the figures. The statistical differences between the groups are shown by stars in the figures and defined as follows: * indicates the differences between control and HG groups ( p = 0.000). ** indicates the differences between HG and HG+ Lys groups ( p = 0.000).

Article Snippet: The antibody against XBP1s (sc-7160) was from Santa Cruz, USA; and the antibodies against p-eIF2α (phospho-Ser51) (ab32157), eIF2α (ab5369), β-actin (ab227387), and LC3 (48394) were bought from Abcam Inc., Cambridge, MA.

Techniques: Nucleic Acid Electrophoresis, Reverse Transcription Polymerase Chain Reaction, Control, Incubation

Journal: PLoS ONE

Article Title: L-lysine protects C2C12 myotubes and 3T3-L1 adipocytes against high glucose damages and stresses

doi: 10.1371/journal.pone.0225912

Figure Lengend Snippet: (A) The Western blot data of whole-cell lysates of C2C12 myoblasts subjected to Lys (1 mM) and/ or HG treatments at different times of incubation that exposed to different antibodies. The β-actin served as a loading control. (B and C) Show the ratio of the spliced XBP1 (XBP1s)/β-actin and p -eIF2α/eIF2α, respectively, as determined by semiquantitative analysis of the bands in (A). All data are represented as means ± SD of three independent experiments. The data of the named four independent groups at different time intervals were analyzed by repeated-measures ANOVA. The significant differences (p-value) in the XBP1s/β-Actin and p -eIF2α/eIF2α ratios within the groups at different time intervals are shown in the figures. The statistical differences between the groups are shown by stars in the figures and defined as follows: * indicates the differences between control and HG groups ( p = 0.000). ** indicates the differences between HG and HG+ Lys groups ( p = 0.000). *** indicates the differences between control and Lys groups ( p = 0.032).

Article Snippet: The antibody against XBP1s (sc-7160) was from Santa Cruz, USA; and the antibodies against p-eIF2α (phospho-Ser51) (ab32157), eIF2α (ab5369), β-actin (ab227387), and LC3 (48394) were bought from Abcam Inc., Cambridge, MA.

Techniques: Western Blot, Incubation, Control

Journal: PLoS ONE

Article Title: L-lysine protects C2C12 myotubes and 3T3-L1 adipocytes against high glucose damages and stresses

doi: 10.1371/journal.pone.0225912

Figure Lengend Snippet: (A) The Western blot data of whole-cell lysates of 3T3-L1 adipocytes subjected to Lys (1 mM) and/ or HG treatments at different times of incubation that exposed to different antibodies. The β-actin served as a loading control. (B) and (C) Show the ratio of the spliced XBP1 (XBP1s)/ β-actin and p -eIF2α/eIF2α, respectively, as determined by semiquantitative analysis of the bands in (A). All data are represented as means ± SD of three independent experiments. The data of the named four independent groups at different time intervals were analyzed by repeated-measures ANOVA. The significant differences (p-value) in the XBP1s/β-Actin and p -eIF2α/eIF2α ratios within the groups at different time intervals are shown in the figures. The statistical differences between the groups are shown by stars in the figures and defined as follows: * indicates the differences between control and HG groups ( p = 0.000). ** indicates the differences between HG and HG+ Lys groups ( p = 0.000 for XBP1/β-Actin and p = 0.003 for p-eIF2α/eIF2α ratios).

Article Snippet: The antibody against XBP1s (sc-7160) was from Santa Cruz, USA; and the antibodies against p-eIF2α (phospho-Ser51) (ab32157), eIF2α (ab5369), β-actin (ab227387), and LC3 (48394) were bought from Abcam Inc., Cambridge, MA.

Techniques: Western Blot, Incubation, Control

Journal: Circulation Research

Article Title: Unspliced XBP1 Confers VSMC Homeostasis and Prevents Aortic Aneurysm Formation via FoxO4 Interaction

doi: 10.1161/circresaha.117.311450

Figure Lengend Snippet: Figure 3. Unspliced XBP1 maintains the contractile phenotype of VSMCs. A, Representative western blot and quantification of SMC contractile markers and XBP1 in primary VSMCs isolated from thoracic aortas (TA) and abdominal aortas (AA) of XBP1WT and XBP1SMKO mice (8- to 12-week-old, male). Five aortas were pooled in each sample. The experiments were performed four times independently. *P<0.05. NS, no significance. B, Western blot analysis of the expression of SMC contractile markers and XBP1 in treated A7r5 cells. The data were presented as means ± SEM from 7 independent experiments in duplicate. *P<0.05. NS, no significance. C, Representative western blot and quantification of SMC contractile markers and XBP1 in primary VSMCs isolated from XBP1SMKO mice (8- to 12-week-old, four aortas were pooled in each sample) after transfection of Flag-vector or Flag-XBP1u for 48 h. The experiments were performed four times independently. *P<0.05.

Article Snippet: Antibodies against XBP1 (sc-7160) used for western blot to detect both isoforms and co-immunoprecipitation to detect FoxO4 interaction with XBP1u, myocardin (sc-21561), GST-tag (sc-138) and eIF-5 (sc-282) were from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Western Blot, Isolation, Expressing, Transfection, Plasmid Preparation

Journal: Circulation Research

Article Title: Unspliced XBP1 Confers VSMC Homeostasis and Prevents Aortic Aneurysm Formation via FoxO4 Interaction

doi: 10.1161/circresaha.117.311450

Figure Lengend Snippet: Figure 4. Unspliced XBP1 modulates VSMC phenotype via FoxO4. A, Luciferase reporter analysis in COS-7 cells. SMA promoter-luc reporter- or SM22α promoter-luc reporter-overexpressing COS-7 cells were transfected with Flag-myocardin, Flag-XBP1u or Flag-XBP1s plasmids. The luciferase activity normalized to β-galactosidase activity was presented as the means ± SEM from three independent experiments performed in triplicate. *P<0.05. NS, no significance. B, Chromatin immunoprecipitation

Article Snippet: Antibodies against XBP1 (sc-7160) used for western blot to detect both isoforms and co-immunoprecipitation to detect FoxO4 interaction with XBP1u, myocardin (sc-21561), GST-tag (sc-138) and eIF-5 (sc-282) were from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Luciferase, Transfection, Activity Assay, Chromatin Immunoprecipitation

Journal: Circulation Research

Article Title: Unspliced XBP1 Confers VSMC Homeostasis and Prevents Aortic Aneurysm Formation via FoxO4 Interaction

doi: 10.1161/circresaha.117.311450

Figure Lengend Snippet: Figure 6. Unspliced XBP1 but not spliced inhibites myocardin interaction with FOXO4. A, Co- immunoprecipitation analysis of FoxO4 and myocardin. Flag-Myo (Flag-myocardin) and His-FOXO4- overexpressing HEK293A cells, transfected with pcDNA3.1, pcDNA3.1-XBP1u or pcDNA3.1-XBP1s, were harvested for co-immunoprecipitation with an anti-Flag antibody and immunoblotting with an anti- His antibody, sequentially. The quantitative data of his after immunoprecipitation with flag antibody were from three independent experiments. *P<0.05. NS, no significance. B, Co-immunoprecipitation analysis of FoxO4 and myocardin in serum-starved rat primary VSMCs infected with Ad-FoxO4-DN or Ad-EGFP. C, Western blot analysis of SMC contractile markers in serum-starved A7r5 cells transfected with Flag-vector or Flag-FoxO4-DN. Five independent experiments were performed in duplicate. *P<0.05. D, Quantitative real-time PCR analysis of mRNA levels of inflammatory cytokines and MMP-9 in serum-starved A7r5 cells transfected with Flag-vector or Flag-FoxO4-DN. The data were expressed as the means ± SEM from 5 independent experiments performed in duplicate. *P<0.05. E-G, 16-week-old ApoE-/- mice were infected

Article Snippet: Antibodies against XBP1 (sc-7160) used for western blot to detect both isoforms and co-immunoprecipitation to detect FoxO4 interaction with XBP1u, myocardin (sc-21561), GST-tag (sc-138) and eIF-5 (sc-282) were from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Immunoprecipitation, Transfection, Western Blot, Infection, Plasmid Preparation, Real-time Polymerase Chain Reaction