transcription factor 6 atf6  (Novus Biologicals)

Journal: Theranostics

Article Title: Hyperglycemia in combination with excess fat intake promotes renal pyroptosis and fibrosis through Gα 12 -dependent endoplasmic reticulum stress

doi: 10.7150/thno.124015

Figure Lengend Snippet: Induction of ER stress in the kidney (A) Heatmap hierarchical correlation analysis of differentially expressed genes (DEGs) (absolute fold-change >1.5 and P <0.05) (left) and GSEA-enrichment plot of the Hallmark category showing the unfolded protein response (UPR) (NES = 1.583, FDR = 0.008) positively correlated with HFD+STZ treatment in kidney tissue using transcriptome data (n = 3 each) (middle). The top 20 genes comprising the enrichment score's leading edge are indicated in the corresponding heatmap (blue, low; red, high). Bar graphs (right) of significantly enriched UPR-related signaling pathways positively correlated with HFD+STZ-treated WT mice in GSEA biological process and molecular function. NES and NOM P -value are shown in the bar graph. (B) Schematic of the induction and treatment protocol. Diabetes was established by feeding mice a 60% HFD for 18 weeks, with the addition of STZ injections (50 mg/kg body weight, i.p three times) at the 12th week. The AZ2 compound (10 mg/kg body weight) was administered orally five times weekly starting at week 15 and continuing until week 18. (C) Immunodetection assays for IRE1α, ATF6, and GRP78 were done using kidney cortex (upper) and medulla (lower) homogenate samples. For C , β-actin was used as normal control for densitometric analysis. The quantification of relative band density is provided for the indicated samples (n = 4 or 5 per group). Results represent the mean ± SEM. One-way ANOVA combined with LSD post-hoc analysis was utilized to calculate significance (*p < 0.05, **p < 0.01, ***p < 0.001). Groups with no significant difference are labeled N.S. (not significant).

Article Snippet: Primary antibodies were sourced from various suppliers as follows: α-SMA (ab7817) was obtained from Abcam (Cambridge, MA, USA); Fibronectin (610078) from BD Biosciences (Franklin Lakes, NJ, USA); and activating transcription factor 6 (ATF6) (NBD1-40256) from Novus Biologicals (Littleton, CO, USA).

Techniques: Protein-Protein interactions, Immunodetection, Control, Labeling

Journal: Theranostics

Article Title: Hyperglycemia in combination with excess fat intake promotes renal pyroptosis and fibrosis through Gα 12 -dependent endoplasmic reticulum stress

doi: 10.7150/thno.124015

Figure Lengend Snippet: Gα 12 family overexpression in the kidney. (A) Immunodetection of Gα 12 , Gα 13 , PGC1α, and PPARα was carried out on the homogenates used in Figure , with band intensity quantification provided for n= 4 or 5 per group. (B) Representative IHC images showing Gα 12 and Gα 13 localization; scale bar represents 100 μm. (n= 4 or 5 per group) (C) Pearsons's correlation were calculated to determine the correlation between Gα 12 or Gα 13 and either IRE1α or ATF6 (n= 4 or 5/group). For A , β-actin was used as normal control for densitometric analysis. Values are represented as mean ± SEM (** p < 0.01, *** p < 0.001, and N.S., not significant). One-way ANOVA in association with an LSD multiple comparison was used for calculating statistical significance ( A ).

Article Snippet: Primary antibodies were sourced from various suppliers as follows: α-SMA (ab7817) was obtained from Abcam (Cambridge, MA, USA); Fibronectin (610078) from BD Biosciences (Franklin Lakes, NJ, USA); and activating transcription factor 6 (ATF6) (NBD1-40256) from Novus Biologicals (Littleton, CO, USA).

Techniques: Over Expression, Immunodetection, Control, Comparison

Journal: Theranostics

Article Title: Hyperglycemia in combination with excess fat intake promotes renal pyroptosis and fibrosis through Gα 12 -dependent endoplasmic reticulum stress

doi: 10.7150/thno.124015

Figure Lengend Snippet: Effects of STZ treatments on ER stress, pyroptosis and fibrosis markers in the kidney. (A) A schematic explanation of an animal experiment showing mice subjected to STZ treatments (50 mg/kg body weight, i.p., 3 times on 1 st week and sacrificed after 15 th week). ( B ) Serum RBS, CRE, and BUN contents (n = 5 or 10/group). ( C ) Immunodetection assays for IRE1α, ATF6 and Gα 12 . ( D ) Immunodetection assays for mature IL-1β, Casp-1 and c-Casp-1. ( E ) Immunodetection assays for Col1A1, α-SMA and fibronectin of the same samples. ( F ) PAS staining of the kidney. Scale bar: 100 μm. ( G ) The bar graph shows the glomerular/bowman's capsule ratio of the kidney (n = 5 or 10/group). Each point represents the mean value of 15 arbitrary visual fields in each kidney sample. For comparison of band intensities within the blots, 5 or 9 samples were chosen for immunoblottings based on blood sugar content. ( H ) Pearsons's correlation assay between Gα 12 and IRE-1α was performed as in Fig. 2C (n = 5 or 9/group). For C, D , and E, β-actin was used as normal control for densitometric analysis. For D and E , β-actin is shared. The quantification of band intensities is given to their corresponding samples (n = 5 or 9/group). Values are represented as mean ± SEM (* p < 0.05, ** p < 0.01, *** p < 0.001, and N.S., not significant). Two-tailed unpaired Student's t test was done for calculating statistical significance ( B-E, G ).

Article Snippet: Primary antibodies were sourced from various suppliers as follows: α-SMA (ab7817) was obtained from Abcam (Cambridge, MA, USA); Fibronectin (610078) from BD Biosciences (Franklin Lakes, NJ, USA); and activating transcription factor 6 (ATF6) (NBD1-40256) from Novus Biologicals (Littleton, CO, USA).

Techniques: Immunodetection, Staining, Comparison, Two-Photon Excitation Fluorescence Cross-Correlation Assay, Control, Two Tailed Test

Journal: Experimental and Therapeutic Medicine

Article Title: ER stress preconditioning ameliorates liver damage after hemorrhagic shock and reperfusion

doi: 10.3892/etm.2021.9679

Figure Lengend Snippet: Topographical changes of UPR signaling. Representative stains of the immunohistochemical proof of pPERK, ATF6, and sXBP1. Vessels of the periportal field (P) and central veins (C) are exemplified. Immunohistochemical staining for detection of (A) pPERK, (B) ATF6 and (C) sXBP1 in the different groups. Scale bar, 300 µm. pPERK, phosphorylated protein kinase RNA-like endoplasmic reticulum kinase; ATF6, activating transcription factor 6; sXBP1, spliced Version of X-box binding protein 1; SC, sham control; HS/R, hemorrhagic shock and reperfusion; DV, drug vehicle; TM, tunicamycin; BC, baseline control; UPR, unfolded protein response.

Article Snippet: The tissue sections were incubated overnight at 4˚C with the rabbit anti-ATF6 antibody (#NBP1-77251, Novus Biologicals Europe, Cambridge, Great Britain), diluted at 1:100.

Techniques: Immunohistochemical staining, Staining, Binding Assay

Journal: Shock (Augusta, Ga.)

Article Title: Modeling Acute ER stress in vivo and in vitro

doi: 10.1097/SHK.0000000000000759

Figure Lengend Snippet: Male Balb/c mice were injected with either control buffer (Con) or thapsigargin (TG) tunicamycin (Tun). After 24 h, the epididymal fat pads were dissected and A) Equal amounts of protein were resolved by SDS-PAGE followed by immunoblotting using antibodies recognizing ATF-6, eIF2α, and IRE1α. B) Densitometry quantification of ER stress proteins expressed over loading control α/β tubulin. Data shown are in mean ± SEM, *p<0.05 vs sham, # p<0.05 vs TG. (n=3-4).

Article Snippet: Proteins were resolved by SDS-PAGE followed by Western blotting using antibodies recognizing GRP78 (BiP), inositol-requiring enzyme (IRE1α), eukaryotic initiation factor (eIF2α), CCAAT-enhancer-binding protein (CHOP), alpha/ beta tubulin, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), isocitrate dehydrogenases (IDH1), and Activating transcription factor 6 (ATF-6) (Novus Biologicals).

Techniques: Injection, Control, SDS Page, Western Blot

Journal: Shock (Augusta, Ga.)

Article Title: Modeling Acute ER stress in vivo and in vitro

doi: 10.1097/SHK.0000000000000759

Figure Lengend Snippet: Male Balb/c mice were injected with either control buffer (Con) or thapsigargin (TG) tunicamycin (Tun). After 24 h, the livers were dissected and A-B) total RNA was extracted and transcribed to cDNA. Relative mRNA levels of the indicated ER stress/UPR genes were quantified using real-time PCR and normalized to 18S rRNA. C-D) Equal amounts of protein were resolved by SDS-PAGE followed by immunoblotting using antibodies recognizing BiP, CHOP, IRE1α, and ATF-6. E) Densitometry quantification of ER stress proteins were either expressed over loading control GAPDH or IDH-1. Data shown are in mean ± SEM, *p<0.05 vs sham, # p<0.05 vs TG. (n=3-4).

Article Snippet: Proteins were resolved by SDS-PAGE followed by Western blotting using antibodies recognizing GRP78 (BiP), inositol-requiring enzyme (IRE1α), eukaryotic initiation factor (eIF2α), CCAAT-enhancer-binding protein (CHOP), alpha/ beta tubulin, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), isocitrate dehydrogenases (IDH1), and Activating transcription factor 6 (ATF-6) (Novus Biologicals).

Techniques: Injection, Control, Real-time Polymerase Chain Reaction, SDS Page, Western Blot