hspa5 recombined protein  (MedChemExpress)

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: Pro-inflammatory diet inhibits endoplasmic reticulum stress signaling involving ATF4/HSPA5 pathway. (A) RNA-seq analysis on principal component analysis of all genes to evaluate the differences among CN group (in black, n = 4) and PI group (in red, n = 5). (B) Number of differentially expressed colonic genes between two groups. (C) Heatmap of gene set variation analysis showing several downregulated genes in PI group belonging to heat shock protein 70 family. (D) Quantitative RT–PCR analysis showing the down-regulation of Hspa1b , Hspa1a , Hspa5 and Hsph1 after pro-inflammatory diet treatment. (E) Comparisons of the relative mRNA level of Perk , Atf4 , Atf6 and Chop between mice treated with or without pro-inflammatory diet. (F) Heatmap on the expression of Perk , Atf4 , Hspa5 and GPX4 between ND group and PD group. (G-I) Western blotting and immunohistochemical results demonstrating decreased expression of ATF4 and HSPA5 in PD group. Corresponding animal experiments had been repeated for 3 or more times. Data are represented as the mean ± SD. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: RNA Sequencing, Quantitative RT-PCR, Expressing, Western Blot, Immunohistochemical staining

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: The involvement of ferroptosis in intestinal inflammation of CD patients with pro-inflammatory diet pattern. (A-C) Results from Dataset Accession number GSE6731 showing the downregulation of GPX4 and the upregulation of HSPA5 and ATF4 in the disease affected colon from patients with IBD. CD-un, disease unaffected tissues from CD patients; CD-aff, disease affected tissues from CD patients; UC-un, disease unaffected tissues from UC patients; UC-aff, disease affected tissues from UC patients. (D-F) Relative expression of PERK , ATF4 , HSPA5 and GPX4 in colon from CD patients of our hospital in mRNA and protein level. HC, health control group; CD, patients with CD group; (G) The DII scores of 32CD patients included in this study were compared between High-DII group (patients with higher DII group) and Low-DII group (patients with lower DII group). (H) Representative immunohistochemical staining of GPX4 and HSPA5 in CD colon tissues from High-DII group and Low-DII group. (I-J) Quantification of GPX4 and HSPA5 expression using integrated optical density/specimen area (IOD/area). Data are represented as the mean ± SD. ns, no significance.

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: Expressing, Control, Immunohistochemical staining, Staining

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: Bacteroides uniformis ameliorates DSS-induced acute colitis. (A) Experimental design for antibiotics and DSS treatment, as well as the microbiota transplantation. BU group (oral administration of B. uniformis , n = 5), UA group (oral S. mutans , n = 5), and PBS group ( n = 5). (B) Culture of the fecal bacteria before and after antibiotic cocktails treatment using the blood agar plate. (C-D) Measurement of body weight lost and colon length at sacrifice of three groups. (E-F) H&E staining of colon section and the pathological score for each group (Scale bars: 625um). (G) Comparison of colonic MPO activity among three groups. (H-I) Detection of the relative mRNA level of Mucin-2 , tight juncture proteins ( Zo-1 and Cldn8 ) and inflammatory cytokines ( Tnfα , Il17a , Il6 , Il1β , Il12 , Il1α , Il10 and Tgfβ1 ) in colon. (J-L) Higher expression of GPX4, HSPA5 and ATF4 in both RNA and protein level after administrated with B. uniformis. Corresponding animal experiments had been repeated for 3 or more times. Data are represented as the mean ± SD.

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: Transplantation Assay, Bacteria, Staining, Comparison, Activity Assay, Expressing

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: Bacteroides uniformis ameliorates Pro-inflammatory diet-induced colitis in DSS model by reversing ferroptosis. (A) Experimental design for dietary and DSS treatment. Mice fed on the pro-inflammatory diet were divided into three groups: BU group (oral B. uniformis , n = 5), UA group (oral S. mutans , n = 5), PBS group ( n = 5). (B-D) Comparisons of body weight lost during DSS treatment, the colon length and the colonic MPO activity at sacrifice among three groups. (E-F) Quantitative RT–PCR detection of inflammatory cytokines ( Tnfα , Il17a , Il6 , Il1β , Il12 Il1α , Il10 and Tgfβ1 ), tight junction proteins ( Cldn1 , Cldn2 , Jam and Cldn14 ) and Mucin-2 expression in colon. (G-H) Representative images with H&E staining and the pathological score to estimate intestinal inflammation (Scale bars: 625um). (I-K) Measurement of Fe, GSH and MDA content in colon. (L) Relative mRNA expression of ferroptosis related genes ( Acsl4 , Tfr1 , Nox1 , Cox1 , Gpx4 , Fpn , Fth1 , Ftl , Tf , Ireb2 , Slc7a11 and Slc3a2 ). (M) Representative images of transmission electron microscopy for distal colon. Arrows head to mitochondria. (N-O) The recovery of GPX4, HSPA5, ATF4 and PERK expression in both RNA and protein level after administrated with B. uniformis. Corresponding animal experiments had been repeated for 3 or more times. Data are represented as the mean ± SD.

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: Activity Assay, Quantitative RT-PCR, Expressing, Staining, Transmission Assay, Electron Microscopy

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: Co-cultured with Bacteroides uniformis inhibits ER stress pathway-mediated ferroptosis in vitro . (A-B) Improvement of cell viability in a concentration- and time-dependent pattern when co-culture with B. uniformis . (C-D) Decreased level of LDH level in cell coculture supernatant. Relative expression of PERK, ATF4, HSPA5 and GPX4 in RNA level (E, F) and in protein level (G H) after treated cells with B. uniformis . A, C, E and G showed the experimental results in Caco-2 cell. B, D, F and H showed the experimental results in NCM460 cell. Experiments had been repeated for 3 or more times. Data are represented as the mean ± SD.

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: Cell Culture, In Vitro, Concentration Assay, Co-Culture Assay, Expressing

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: Pro-inflammatory diet inhibits endoplasmic reticulum stress signaling involving ATF4/HSPA5 pathway. (A) RNA-seq analysis on principal component analysis of all genes to evaluate the differences among CN group (in black, n = 4) and PI group (in red, n = 5). (B) Number of differentially expressed colonic genes between two groups. (C) Heatmap of gene set variation analysis showing several downregulated genes in PI group belonging to heat shock protein 70 family. (D) Quantitative RT–PCR analysis showing the down-regulation of Hspa1b , Hspa1a , Hspa5 and Hsph1 after pro-inflammatory diet treatment. (E) Comparisons of the relative mRNA level of Perk , Atf4 , Atf6 and Chop between mice treated with or without pro-inflammatory diet. (F) Heatmap on the expression of Perk , Atf4 , Hspa5 and GPX4 between ND group and PD group. (G-I) Western blotting and immunohistochemical results demonstrating decreased expression of ATF4 and HSPA5 in PD group. Corresponding animal experiments had been repeated for 3 or more times. Data are represented as the mean ± SD. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: RNA Sequencing, Quantitative RT-PCR, Expressing, Western Blot, Immunohistochemical staining

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: The involvement of ferroptosis in intestinal inflammation of CD patients with pro-inflammatory diet pattern. (A-C) Results from Dataset Accession number GSE6731 showing the downregulation of GPX4 and the upregulation of HSPA5 and ATF4 in the disease affected colon from patients with IBD. CD-un, disease unaffected tissues from CD patients; CD-aff, disease affected tissues from CD patients; UC-un, disease unaffected tissues from UC patients; UC-aff, disease affected tissues from UC patients. (D-F) Relative expression of PERK , ATF4 , HSPA5 and GPX4 in colon from CD patients of our hospital in mRNA and protein level. HC, health control group; CD, patients with CD group; (G) The DII scores of 32CD patients included in this study were compared between High-DII group (patients with higher DII group) and Low-DII group (patients with lower DII group). (H) Representative immunohistochemical staining of GPX4 and HSPA5 in CD colon tissues from High-DII group and Low-DII group. (I-J) Quantification of GPX4 and HSPA5 expression using integrated optical density/specimen area (IOD/area). Data are represented as the mean ± SD. ns, no significance.

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: Expressing, Control, Immunohistochemical staining, Staining

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: Bacteroides uniformis ameliorates DSS-induced acute colitis. (A) Experimental design for antibiotics and DSS treatment, as well as the microbiota transplantation. BU group (oral administration of B. uniformis , n = 5), UA group (oral S. mutans , n = 5), and PBS group ( n = 5). (B) Culture of the fecal bacteria before and after antibiotic cocktails treatment using the blood agar plate. (C-D) Measurement of body weight lost and colon length at sacrifice of three groups. (E-F) H&E staining of colon section and the pathological score for each group (Scale bars: 625um). (G) Comparison of colonic MPO activity among three groups. (H-I) Detection of the relative mRNA level of Mucin-2 , tight juncture proteins ( Zo-1 and Cldn8 ) and inflammatory cytokines ( Tnfα , Il17a , Il6 , Il1β , Il12 , Il1α , Il10 and Tgfβ1 ) in colon. (J-L) Higher expression of GPX4, HSPA5 and ATF4 in both RNA and protein level after administrated with B. uniformis. Corresponding animal experiments had been repeated for 3 or more times. Data are represented as the mean ± SD.

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: Transplantation Assay, Bacteria, Staining, Comparison, Activity Assay, Expressing

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: Bacteroides uniformis ameliorates Pro-inflammatory diet-induced colitis in DSS model by reversing ferroptosis. (A) Experimental design for dietary and DSS treatment. Mice fed on the pro-inflammatory diet were divided into three groups: BU group (oral B. uniformis , n = 5), UA group (oral S. mutans , n = 5), PBS group ( n = 5). (B-D) Comparisons of body weight lost during DSS treatment, the colon length and the colonic MPO activity at sacrifice among three groups. (E-F) Quantitative RT–PCR detection of inflammatory cytokines ( Tnfα , Il17a , Il6 , Il1β , Il12 Il1α , Il10 and Tgfβ1 ), tight junction proteins ( Cldn1 , Cldn2 , Jam and Cldn14 ) and Mucin-2 expression in colon. (G-H) Representative images with H&E staining and the pathological score to estimate intestinal inflammation (Scale bars: 625um). (I-K) Measurement of Fe, GSH and MDA content in colon. (L) Relative mRNA expression of ferroptosis related genes ( Acsl4 , Tfr1 , Nox1 , Cox1 , Gpx4 , Fpn , Fth1 , Ftl , Tf , Ireb2 , Slc7a11 and Slc3a2 ). (M) Representative images of transmission electron microscopy for distal colon. Arrows head to mitochondria. (N-O) The recovery of GPX4, HSPA5, ATF4 and PERK expression in both RNA and protein level after administrated with B. uniformis. Corresponding animal experiments had been repeated for 3 or more times. Data are represented as the mean ± SD.

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: Activity Assay, Quantitative RT-PCR, Expressing, Staining, Transmission Assay, Electron Microscopy

Journal: Journal of Advanced Research

Article Title: Bacteroides uniformis ameliorates pro-inflammatory diet-exacerbated colitis by targeting endoplasmic reticulum stress-mediated ferroptosis

doi: 10.1016/j.jare.2024.11.025

Figure Lengend Snippet: Co-cultured with Bacteroides uniformis inhibits ER stress pathway-mediated ferroptosis in vitro . (A-B) Improvement of cell viability in a concentration- and time-dependent pattern when co-culture with B. uniformis . (C-D) Decreased level of LDH level in cell coculture supernatant. Relative expression of PERK, ATF4, HSPA5 and GPX4 in RNA level (E, F) and in protein level (G H) after treated cells with B. uniformis . A, C, E and G showed the experimental results in Caco-2 cell. B, D, F and H showed the experimental results in NCM460 cell. Experiments had been repeated for 3 or more times. Data are represented as the mean ± SD.

Article Snippet: Upon reaching 70–80 % confluence, the cell were treated with HSPA5 inhibitor (1 uM, HY-100437, MCE, Shanghai, China), HSPA5 recombined protein (10 ng / mL, HY-P71742, MCE, Shanghai, China) or PERK inhibitor (1uM or 5uM, GSK2606414, MCE, Shanghai, China) for two days and were then harvested for RT-qPCR and Western blot analysis.

Techniques: Cell Culture, In Vitro, Concentration Assay, Co-Culture Assay, Expressing

Journal: Disease Markers

Article Title: HSPA5 Inhibitor Meliorate DSS-Induced Colitis through HSPA1A/CHIP

doi: 10.1155/2022/7115181

Figure Lengend Snippet: Primer sequences used for RT-PCR.

Article Snippet: The HSPA5 inhibitor HA15 (batch no. 1609402-14-3) was purchased from the MedChemExpress (New Jersey, USA).

Techniques:

Journal: Disease Markers

Article Title: HSPA5 Inhibitor Meliorate DSS-Induced Colitis through HSPA1A/CHIP

doi: 10.1155/2022/7115181

Figure Lengend Snippet: Expressions of HSPA5 in colonic tissue and the effects of pharmacotherapies on DSS-induced colitis in mice. (a) The colonic protein levels of HSPA5 measured by Western blotting. (b) Representative protein levels of HSPA5/GAPDH. (c) Levels of HSPA5 mRNA in colonic tissue. Data are shown as means ± SD ( n = 3). (d) The disease activity index (DAI) score of each group was monitored daily. (e) Typical colonic appearance of each group. (f) Colon length of each group. (g) Histological analysis (×50, ×200) about pathological changes of colonic tissue under a microscope. (h) Histological score of each group. Data are shown as means ± SD ( n = 8). ∗∗ P < 0.01, ∗∗∗ P < 0.001, vs. DSS group.

Article Snippet: The HSPA5 inhibitor HA15 (batch no. 1609402-14-3) was purchased from the MedChemExpress (New Jersey, USA).

Techniques: Western Blot, Activity Assay, Microscopy