Journal: World Journal of Gastroenterology : WJG

Article Title: Caspase-12 mediates carbon tetrachloride-induced hepatocyte apoptosis in mice

doi: 10.3748/wjg.v20.i48.18189

Figure Lengend Snippet: Carbon tetrachloride induced endoplasmic reticulum stress in the liver. The representative Western blots show that there was an increase in endoplasmic reticulum stress response in CCl4-treated mice. Glucose-regulated protein 78 (GRP78), CCAAT/-enhancer-binding protein homologous protein (CHOP) and serine/threonine-protein kinase/endoribonuclease (IRE-1) were upregulated in the livers of both caspase-12+/+ and -12-/- mice treated with CCl4.

Article Snippet: The antibodies used were as follows: rabbit anti-caspase-12 (AB3612, Lot 22101182; Chemicon of EMD Millipore, Billerica, MA, United States) and mouse anti-actin (MAB1501; Chemicon); rabbit anti-caspase-3 (sc-7148), caspase-8 (sc-7890), caspase-9 (sc-8355), and rabbit anti-cytochrome C (sc-7159) from Santa Cruz Biotechnology (Dallas, TX, United States); anti-glucose-regulated protein 78 (GRP78) (NBP1-06274; Novus Biologicals, Littleton, CO, United States); anti-CCAAT/-enhancer-binding protein homologous protein (CHOP) (#2895; Cell Signaling Technology, Danvers, MA); anti-serine/threonine-protein kinase/endoribonuclease (IRE-1) (ab37073; Abcam, Cambridge, United Kingdom).

Techniques: Western Blot, Binding Assay

Journal: BMC pulmonary medicine

Article Title: IL-32 induces epithelial-mesenchymal transition by triggering endoplasmic reticulum stress in A549 cells.

doi: 10.1186/s12890-020-01319-z

Figure Lengend Snippet: Fig. 4 GRP78 expression characteristics induced by IL-32 or 4-PBA. a GRP78 mRNA expression levels. *P < 0.05 compared with the control group, **P < 0.05 compared with the rhIL-32 group. b and c GRP78 protein expression levels, measured by western blotting. *P < 0.05 compared with the control group, **P < 0.05 compared with the rhIL-32 group

Article Snippet: A sample containing 20 μg of protein was separated on a 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel (Solarbio), transferred to a polyvinylidene fluoride membrane, and incubated with rabbit anti-mouse N-cadherin, GRP78, and α-SMA primary antibodies (Proteintech, Wuhan, China), or βactin antibody (Bioss, Beijing, China) as a loading control, at room temperature for 2 h. Subsequently, the membrane was washed, incubated with the horseradish peroxidase-conjugated goat anti-rabbit secondary antibody (Proteintech, Wuhan, China) for 1 h at room temperature, and exposed using the ECL kit.

Techniques: Expressing, Control, Western Blot

Journal: Experimental and therapeutic medicine

Article Title: Glucose-regulated protein 78 is an antiviral against hepatitis A virus replication.

doi: 10.3892/etm.2017.4407

Figure Lengend Snippet: Figure 1. Effects of GRP78 knockdown on HAV replication in Huh7 cells. Cells were transfected with si‑GRP78 or si‑C 1 day prior to HAV infection. Cellular RNA was extracted and subjected to reverse transcription‑quantita tive polymerase chain reaction 96 h after HAV HA‑11‑1299 genotype IIIA strain infection at a multiplicity of infection of 0.1. Data are expressed as the mean ± standard deviation. *P<0.05 vs. si‑C‑transfected controls using Student's t‑test. All experiments were performed in triplicate. GRP78, glucose regulated protein 78; HAV, hepatitis A virus; si‑GRP78, siRNA against GRP78; si‑C, control siRNA.

Article Snippet: The membranes were probed with primary antibodies against GRP78 (#3177), ATF4 (#11815; all 1:1,000; all from Cell Signaling Technology, Danvers, MA, USA), ATF6 (sc‐166659), XBP1 (sc‐80154) or GAPDH (sc‐25778; all 1:1,000; all from Santa Cruz Biotechnology, Inc.) at 4 ̊C for 16 h. Anti‐mouse IgG HRP‐linked antibody (NA931; dilution, 1:7,000; GE Healthcare Life Sciences, Chalfont, UK) or anti‐rabbit IgG HRP‐linked antibody (#7074; dilution, 1:3,500; Cell Signaling Technology) were used as secondary antibodies, incubated at room temperature for 1 h. Proteins were visualized using an enhanced chemiluminescent ECL Western blot substrate (Amersham ECL Prime Western Blotting Detection Reagent; GE Healthcare Life Sciences) and scanned with an image analyzer LAS‐4000 (Fujifilm Corporation, Tokyo, Japan) and Image Gauge (version 3.1; Fujifilm) (4).

Techniques: Knockdown, Transfection, Infection, Polymerase Chain Reaction, Standard Deviation, Virus, Control

Journal: Experimental and therapeutic medicine

Article Title: Glucose-regulated protein 78 is an antiviral against hepatitis A virus replication.

doi: 10.3892/etm.2017.4407

Figure Lengend Snippet: Figure 2. Effects of GRP78 KO on HAV replication in Huh7 cells. (A) Effect of KO of GRP78 on HAV replication in control Huh7 cells and GRP78 KO cells. Cellular RNA was extracted and subjected to reverse transcription‑quantitative polymerase chain reaction 96 h after HAV HA‑11‑1299 genotype IIIA strain infection at a multiplicity of infection of 0.1. Data are expressed as mean ± standard deviation. *P<0.05 vs. si‑C‑transfected controls using Student's t‑test. All experiments were performed in triplicate. (B) Effects of knockout of GRP78 on endoplasmic reticulum stress molecules. Western blot analysis was performed using specific primary antibodies. Clone #5 was subjected to HAV infection. GRP78, glucose regulated protein 78; HAV, hepatitis A virus; KO, knockout; si‑C, control siRNA; ATF, activating transcription factor; XBP, X‑box‑binding protein.

Article Snippet: The membranes were probed with primary antibodies against GRP78 (#3177), ATF4 (#11815; all 1:1,000; all from Cell Signaling Technology, Danvers, MA, USA), ATF6 (sc‐166659), XBP1 (sc‐80154) or GAPDH (sc‐25778; all 1:1,000; all from Santa Cruz Biotechnology, Inc.) at 4 ̊C for 16 h. Anti‐mouse IgG HRP‐linked antibody (NA931; dilution, 1:7,000; GE Healthcare Life Sciences, Chalfont, UK) or anti‐rabbit IgG HRP‐linked antibody (#7074; dilution, 1:3,500; Cell Signaling Technology) were used as secondary antibodies, incubated at room temperature for 1 h. Proteins were visualized using an enhanced chemiluminescent ECL Western blot substrate (Amersham ECL Prime Western Blotting Detection Reagent; GE Healthcare Life Sciences) and scanned with an image analyzer LAS‐4000 (Fujifilm Corporation, Tokyo, Japan) and Image Gauge (version 3.1; Fujifilm) (4).

Techniques: Control, Polymerase Chain Reaction, Infection, Standard Deviation, Knock-Out, Western Blot, Virus

Journal: Investigative Ophthalmology & Visual Science

Article Title: In Search of the Identity of the XAP-1 Antigen: A Protein Localized to Cone Outer Segments

doi: 10.1167/iovs.09-4286

Figure Lengend Snippet: LC-MS/MS results obtained from the protein band corresponding to the XAP-1 antigen. (A) The MS/MS spectrum of a peptide from the LC-MS/MS analysis. It contains product ions of the b- and y- series that matched peptide sequence ITPSYVAFTPEGER from Grp78 from Mus musculus. (B) The m/z ratios for theoretical product ions of the y- and b- series. Product ions that were observed in the MS/MS spectrum are shown in bold.

Article Snippet: Blot A was incubated in rabbit anti-human Grp78 (1:1000 dilution; Anaspec) overnight at 4°C.

Techniques: Liquid Chromatography with Mass Spectroscopy, Tandem Mass Spectroscopy, Sequencing

Journal: Investigative Ophthalmology & Visual Science

Article Title: In Search of the Identity of the XAP-1 Antigen: A Protein Localized to Cone Outer Segments

doi: 10.1167/iovs.09-4286

Figure Lengend Snippet: Protein sequence of Mus musculus Grp78. The peptide from our MS/MS analysis is shown in gray text and represents 2.14% coverage.

Article Snippet: Blot A was incubated in rabbit anti-human Grp78 (1:1000 dilution; Anaspec) overnight at 4°C.

Techniques: Sequencing, Tandem Mass Spectroscopy

Journal: Investigative Ophthalmology & Visual Science

Article Title: In Search of the Identity of the XAP-1 Antigen: A Protein Localized to Cone Outer Segments

doi: 10.1167/iovs.09-4286

Figure Lengend Snippet: Depletion of IgM and Grp78 from the outer segment–enriched preparation. IgM and Grp78 were depleted from outer segment (OS) cell extracts. IgM and the XAP-1 antigen are shown in red, whereas Grp78 is shown in green. Overlap of the signals (E, F, yellow). A small amount of IgM was detected after one round of depletion, but none remained after a second round (A). Both the XAP-1 antigen (B) and Grp78 (C, D) remained in the sample after all IgM was removed. The XAP-1 antigen and Grp78 overlap at an identical relative molecular mass (F). Depletion of Grp78 (C, D) abolished the signal obtained with the XAP-1 antibody (B).

Article Snippet: Blot A was incubated in rabbit anti-human Grp78 (1:1000 dilution; Anaspec) overnight at 4°C.

Techniques:

Journal: Investigative Ophthalmology & Visual Science

Article Title: In Search of the Identity of the XAP-1 Antigen: A Protein Localized to Cone Outer Segments

doi: 10.1167/iovs.09-4286

Figure Lengend Snippet: Immunohistochemical localization of Grp78 in the retinas of mouse and frog. After antigen retrieval, retinas from C57BL6/J mice were immunostained using anti-Grp78 (A, green), PNA (B, red), and iodide (C, blue). (C) Composite image. Yellow (arrows) indicates areas of overlap of the Grp78 and PNA, indicating that in the outer retina, Grp78 is localized to cone photoreceptors in a pattern identical with that of the XAP-1 antigen. (C, inset) Higher magnification image of the area within the inset in C. In the mouse, Grp78 is also found in all cell layers of the retina in a location corresponding to ER. (D) Examination of the frog retina revealed a different pattern. The peripheries of both rod and cone photoreceptor outer segments and the inner segments are immunolabeled in this species. Scale bar, 10 μm. OS, outer segment; IS, inner segment; ONL, outer nuclear layer; OPL, outer plexiform layer; INL, inner nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer.

Article Snippet: Blot A was incubated in rabbit anti-human Grp78 (1:1000 dilution; Anaspec) overnight at 4°C.

Techniques: Immunohistochemical staining, Immunolabeling