Journal: iScience

Article Title: Advanced oxidation protein products induce Paneth cells defects by endoplasmic reticulum stress in Crohn's disease

doi: 10.1016/j.isci.2023.107312

Figure Lengend Snippet: Chronic administration of AOPP induced PC defects in intestinal crypts of mice (A and B) Representative images of HE staining for PC number quantification and immunohistochemical staining for Lysozyme in intestine sections of mice with or without AOPP treatment. These images were representative of results from each experimental group, each with five mice. Images showed that AOPP challenge reduced the PC number and lysozyme expression in intestinal crypts compared with vehicle-treated mice. Scale bar, 20 μm. (C) Representative Western blotting and densitometric quantification showed the protein level of Lysozyme in isolated intestinal crypts of mice with or without AOPP administration. (D and E) Bar graphs representing qPCR analysis showed the mRNA levels of Lysozyme and cryptdins in isolated intestinal crypts of mice with or without AOPP administration. Relative protein and mRNA levels were normalized by GAPDH, respectively. Data were represented by median with range, n = 5 mice; error bars indicating range. Student’s t test, ∗∗p < 0.01 versus vehicles. Vehicle, PBS treatment for 28 days; AOPP, AOPP treatment for 28 days; qPCR, quantitative real-time polymerase chain reaction; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: Anti-mouse GAPDH, anti-rabbit GRP75, anti-rabbit VDAC1, anti-rabbit Mfn1/2 and anti-rabbit PDI were all purchased from Proteintech (Wuhan, China).

Techniques: Staining, Immunohistochemical staining, Expressing, Western Blot, Isolation, Real-time Polymerase Chain Reaction

Journal: iScience

Article Title: Advanced oxidation protein products induce Paneth cells defects by endoplasmic reticulum stress in Crohn's disease

doi: 10.1016/j.isci.2023.107312

Figure Lengend Snippet: Chronic challenge of AOPP induced ER stress in intestinal crypts of mice (A and B) Representative images of immunohistochemical staining for GRP78 and CHOP in mice with or without AOPP challenge. These images were representative of results from each experimental group, each with five mice. Images showed that AOPP treatment elevated GRP78 and CHOP levels in intestinal crypts compared with vehicle-treated mice. Scale bar, 20 μm. (C) Representative Western blotting and densitometric quantification showed the protein levels of GRP78, CHOP and IRE1α in isolated intestinal crypts of mice with or without AOPP challenge. (D and E) Bar graphs representing qPCR analysis showed the levels of GRP78 and CHOP mRNA in isolated intestinal crypts of mice with or without AOPP challenge. Relative protein and mRNA levels were normalized by GAPDH, respectively. Data were represented by median with range, n = 5 mice; error bars indicating range. Student’s t test, ∗p < 0.05 and ∗∗p < 0.01 versus vehicles.

Article Snippet: Anti-mouse GAPDH, anti-rabbit GRP75, anti-rabbit VDAC1, anti-rabbit Mfn1/2 and anti-rabbit PDI were all purchased from Proteintech (Wuhan, China).

Techniques: Immunohistochemical staining, Staining, Western Blot, Isolation

Journal: iScience

Article Title: Advanced oxidation protein products induce Paneth cells defects by endoplasmic reticulum stress in Crohn's disease

doi: 10.1016/j.isci.2023.107312

Figure Lengend Snippet: AOPP-mediated PC defects depended on ER stress in intestinal crypts of mice (A and B) Representative images of HE staining for PC number quantification and immunohistochemical staining for Lysozyme in intestine sections of mice that underwent intraperitoneal administration of AOPP with or without TUDCA. These images were representative of results from each experimental group, each with five mice. Images reveled that treatment with TUDCA alleviated AOPP-mediated decreasing PC number and lysozyme expression in intestinal crypts compared with AOPP-treated mice. Scale bar, 20 μm. (C) Representative Western blotting and densitometric quantification revealed the protein level of Lysozyme in isolated intestinal crypts of mice that underwent intraperitoneal administration of AOPP with or without TUDCA. (D and E) Bar graphs representing qPCR analysis revealed the levels of mRNA of Lysozyme and cryptdins in isolated intestinal crypts of mice that underwent intraperitoneal administration of AOPP with or without TUDCA. Relative protein and mRNA levels were normalized by GAPDH, respectively. Data were represented by median with range, n = 5 mice; error bars indicating range. ANOVA, ∗∗p < 0.01 versus vehicles; # p < 0.05 and ## p < 0.01 versus AOPP-treated mice.

Article Snippet: Anti-mouse GAPDH, anti-rabbit GRP75, anti-rabbit VDAC1, anti-rabbit Mfn1/2 and anti-rabbit PDI were all purchased from Proteintech (Wuhan, China).

Techniques: Staining, Immunohistochemical staining, Expressing, Western Blot, Isolation

Journal: iScience

Article Title: Advanced oxidation protein products induce Paneth cells defects by endoplasmic reticulum stress in Crohn's disease

doi: 10.1016/j.isci.2023.107312

Figure Lengend Snippet: TUDCA inhibited AOPP-resulted ER stress in intestinal crypts of mice (A and B) Representative images of immunohistochemical staining for GRP78 and CHOP in mice that underwent intraperitoneal administration of AOPP with or without TUDCA. These images were representative of results from each experimental group, each with five mice. Images revealed that treatment with TUDCA attenuated AOPP-induced increased GRP78 and CHOP levels in intestinal crypts compared with AOPP-treated mice. Scale bar, 20 μm. (C) Representative Western blotting and densitometric quantification revealed the protein levels of GRP78, CHOP and IRE1α in isolated intestinal crypts of mice that underwent intraperitoneal administration of AOPP with or without TUDCA. (D and E) Bar graphs representing qPCR analysis revealed the levels of GRP78 and CHOP mRNA in isolated intestinal crypts of mice that underwent intraperitoneal administration of AOPP with or without TUDCA. Relative protein and mRNA levels were normalized by GAPDH, respectively. Data were represented by median with range, n = 5 mice; error bars indicating range. ANOVA, ∗∗p < 0.01 versus vehicle; # p < 0.05 and ## p < 0.01 versus AOPP-treated mice. AOPP+TUDCA, AOPP treatment plus daily intraperitoneal injection of TUDCA; TUDCA, tauroursodeoxycholic acid, an inhibitor of ER stress.

Article Snippet: Anti-mouse GAPDH, anti-rabbit GRP75, anti-rabbit VDAC1, anti-rabbit Mfn1/2 and anti-rabbit PDI were all purchased from Proteintech (Wuhan, China).

Techniques: Immunohistochemical staining, Staining, Western Blot, Isolation, Injection

Journal: iScience

Article Title: Advanced oxidation protein products induce Paneth cells defects by endoplasmic reticulum stress in Crohn's disease

doi: 10.1016/j.isci.2023.107312

Figure Lengend Snippet:

Article Snippet: Anti-mouse GAPDH, anti-rabbit GRP75, anti-rabbit VDAC1, anti-rabbit Mfn1/2 and anti-rabbit PDI were all purchased from Proteintech (Wuhan, China).

Techniques: Recombinant, Lysis, Protein Quantitation, Isolation, ATP Assay, Software

Journal: iScience

Article Title: Advanced oxidation protein products induce Paneth cells defects by endoplasmic reticulum stress in Crohn's disease

doi: 10.1016/j.isci.2023.107312

Figure Lengend Snippet: Primer sequences

Article Snippet: Anti-mouse GAPDH, anti-rabbit GRP75, anti-rabbit VDAC1, anti-rabbit Mfn1/2 and anti-rabbit PDI were all purchased from Proteintech (Wuhan, China).

Techniques:

Journal: American Journal of Physiology - Renal Physiology

Article Title: Circulating extracellular vesicles of patients with steroid-sensitive nephrotic syndrome have higher RAC1 and induce recapitulation of nephrotic syndrome phenotype in podocytes

doi: 10.1152/ajprenal.00097.2021

Figure Lengend Snippet: Characterization of extracellular vesicles (EVs) obtained from plasma by serial ultracentrifugation. A: transmission electron microscopy images showing the typical bilayer membrane of EVs (arrowheads). Scale bars = 50 nm. B: Western blot images of expression of EV markers [ALG-2-interacting protein X (ALIX), tumor susceptibility gene 101 (TSG-101), and flotillin] on EVs from six relapse patients with nephrotic syndrome (NS) and podocyte cell lysates. Glucose-regulated protein 94 (GRP94) was used as a negative EV marker. C: circulating EVs of healthy controls (n = 10) and relapse patients with NS (n = 20) were stained with the corresponding cell type-specific markers. Data are presented as means ± SD (in %). An unpaired t test was used. ****P < 0.0001; ***P < 0.001; **P < 0.01. ns, nonsignificant. D: podocalyxin expression represented as mean fold induction (FI) on the matched isotype control on NS relapse EVs (n = 20) was significantly higher than in healthy controls (n = 10, P < 0.001). Data are presented as means ± SD. An unpaired Student’s t test was performed.

Article Snippet: Twenty-five micrograms of protein were loaded per well and separated by 10% or 12% SDS-PAGE, transferred to a PVDF membrane, and probed with 1:1,000 diluted primary antibodies against anti-phospho-p38 (p-p38; clone E-1, Santa Cruz Biotechnology), anti-tumor susceptibility gene 101 (TSG-101) antibody [clone EPR7130(B), Abcam, Cambridge, UK)], anti-α-tubulin (clone B-7, Santa Cruz Biotechnology), anti-synaptopodin (clone D-9, Santa Cruz Biotechnology), anti-flotillin (Cat. No. 3252, Cell Signaling Technology, Frankfurt am Main, Germany), anti-glucose-regulated protein 94 (GRP94; clone D6X2Q, Cell Signaling Technology), and anti-ALG-2-interacting protein X (ALIX; clone 3A9, Cell Signaling Technology).

Techniques: Transmission Assay, Electron Microscopy, Western Blot, Expressing, Marker, Staining

Journal: Molecular medicine reports

Article Title: Kaempferol induces apoptosis in HepG2 cells via activation of the endoplasmic reticulum stress pathway.

doi: 10.3892/mmr.2016.4845

Figure Lengend Snippet: Figure 3. Dose‑ and time‑dependent effects of kaempferol on protein and mRNA levels of endoplasmic reticulum stress markers. (A and B) For dose‑dependent effects, HepG2 cells were treated in the presence or absence of different concentrations of kaempferol for 24 h. Western blotting and RT‑qPCR were performed with GRP78, GRP94, PERK, partial ATF‑6, IRE1α, caspase‑4 and CHOP antibodies. (C and D) For time‑dependent effects, HepG2 cells were exposed to 100 µM kaempferol for different time periods. Western blotting and RT‑qPCR were performed with GRP78, GRP94, PERK, partial ATF‑6, IRE1α, caspase‑4 and CHOP antibodies. Data are expressed as the mean ± standard deviation of at least three independent experiments. GRP78, glucose regulated protein 78; GRP94, glucose regulated protein 94; PERK, protein kinase R‑like endoplasmic reticulum kinase; ATF‑6, activation transcription factor 6; IRE1α, inositol‑requiring enzyme 1α; CHOP, C/EBP homologous protein; RT‑qPCR, reverse transcription‑quantitative polymerase chain reaction; HPRT, hypoxan thine phosphoribosyl transferase.

Article Snippet: The membranes were subsequently incubated with 5% skimmed milk in Tris‐buffered saline with Tween‐20 (Beyotime Institute of Biotechnology) for 1 h to block nonspecific binding and then overnight with the following antibodies: Monoclonal rabbit anti‐human GRP78 (1:1000; cat. no. 3177; Cell Signaling Technology, Inc., Danvers, MA, USA), monoclonal rabbit anti‐human GRP94 (1:1,000; cat. no. 20292; Cell Signaling Technology, Inc.), monoclonal rabbit anti‐human PERK (1:1,000; cat. no. 5683; Cell Signaling Technology, Inc.), monoclonal mouse anti‐human partial ATF‐6 (1:1,000; cat. no. IMG‐273; Imgenex Corporation, San Diego, CA, USA), monoclonal rabbit anti‐human IRE1α (1:1,000; cat. no. 3294; Cell Signaling Technology, Inc.), monoclonal rabbit anti‐human caspase‐4 (1:1,000; cat. no. 4450; Cell Signaling Technology, Inc.), monoclonal mouse anti‐human CHOP (1:1,000; cat. no. 2895; Cell Signaling Technology, Inc.), monoclonal rabbit anti‐human cleaved caspase‐3 (1:1,000; cat. no. 9664; Cell Signaling Technology, Inc.) and monoclonal rabbit anti‐human β‐actin (1:1,000; cat. no. 4970; Cell Signaling Technology, Inc.) at 4 ̊C, followed by incubation with monoclonal goat anti‐rabbit IgG (1:2,000; cat. no. 14708; Cell Signaling Technology, Inc.) secondary antibody for 1 h at room temperature.

Techniques: Western Blot, Standard Deviation, Activation Assay, Polymerase Chain Reaction

Journal: Molecular & Cellular Proteomics : MCP

Article Title: Interactome Analysis Identifies the Role of BZW2 in Promoting Endoplasmic Reticulum-Mitochondria Contact and Mitochondrial Metabolism

doi: 10.1016/j.mcpro.2023.100709

Figure Lengend Snippet:

Article Snippet: Antibody , Anti-GRP75 (rabbit polyclonal) , Abclonal , A0558 , WB (1:1000).

Techniques: Cloning, Transfection, Transduction, Construct, Protease Inhibitor, Software, Calcium Assay, Bioassay, Isolation, Viability Assay, Sequencing, Modification, Western Blot, Bicinchoninic Acid Protein Assay