Journal: Current Issues in Molecular Biology

Article Title: GPX4 Inhibition Enhances the Pro-Oxidant and ER Stress Effects of Tempol in Colon and Gastric Cancer Cell Lines

doi: 10.3390/cimb47100856

Figure Lengend Snippet: Data are presented as observed vs. Bliss-expected normalized protein concentrations (mean ± SEM). Statistical significance of Δ values was assessed by one-sample t -tests (see ). ATF6 showed strong synergy in both cell lines. IRE1α exhibited slight, non-significant synergy in both CRL-1739 and HT29. GRP78 responses were cell line–dependent, with strong synergy in CRL-1739 but no significant difference (moderate antagonism) in HT29 ( p ≥ 0.05).

Article Snippet: Specifically, IRE1α (catalog no. NBP3–49699), GRP78 (catalog no. NBP2–62145), and ATF6 (catalog no. NBP2–69885) ELISA kits (Novus Biologicals, Centennial, CO, USA) were employed according to the manufacturer’s protocols.

Techniques:

Journal: Molecular cancer

Article Title: MYC regulates the unfolded protein response and glucose and glutamine uptake in endocrine resistant breast cancer.

doi: 10.1186/1476-4598-13-239

Figure Lengend Snippet: Figure 8 Glutamine in glucose-deprived conditions activates the UPR. A, Cells were plated at 70% confluence. 24 hr later, media was changed to 0, 2, or 4 mM glutamine alone or in presence of 12 mM glucose. Western blot analysis showed Increased levels of GRP78, IRE1a, phospho-JNK, CHOP and decreased levels of BCL2 were present in LCC1 (right) and LCC9 (left) cells in glutamine-only conditions. MYC protein levels were highest when both glucose and glutamine are present; MYC is undetectable when these metabolites are absent in the media. MYC expression in the presence of glutamine-only, but not in presence of glucose-only, conditions correlated with increased expression of UPR proteins. B, Knockdown of MYC for 24 h was followed by media change to either glucose + glutamine, glucose-only, glutamine-only or no glucose + no glutamine conditions for another 48 h. Western blots analysis showed that a decrease in MYC protein levels correlated with an increase in the UPR proteins IRE1α and phospho-JNK(Thr183/ Ty4185), and the autophagosome formation marker LC3II, and the autophagosome cargo degradation marker p62/SQSTM1. GRP78 was also increased in glucose + glutamine, glucose-only and no glucose + no glutamine conditions but robust expression of GRP78 in glutamine-only conditions was not affected by MYC siRNA. Total levels of JNK did not change.

Article Snippet: 5 μM MYC siRNA (SMARTpool: ON-TARGETplus set of four MYC siRNA Dharmacon, Lafayette, CO), 10 GLS1, GRP78 (HSPA5), IRE1a or XBP1 (10 nM of 3 unique 27mer siRNA duplexes; Origene, Rockville, MD) or their respective control siRNA, were transfected using the TransIT-siQUEST (Mirus, Madison, WI) transfection reagent.

Techniques: Western Blot, Expressing, Knockdown, Marker

Journal: Molecular cancer

Article Title: MYC regulates the unfolded protein response and glucose and glutamine uptake in endocrine resistant breast cancer.

doi: 10.1186/1476-4598-13-239

Figure Lengend Snippet: Figure 9 UPR in glutamine-only conditions can lead to both pro-survival and pro-death outcomes. Effect of transfection of siRNA targeting GRP78, IRE1α, XBP1(s), and MYC for 24 h; or JNK inhibition with a small molecule inhibitor (SP600125) on growth in either glucose + glutamine or glutamine-alone media. Western blot (48 h); A-C, GRP78. D-F, IRE1α. G-I, JNK. J-L, XBP1. M-O, MYC. Inhibition of GRP78 did not significantly further affect cell numbers in glutamine-only conditions in both LCC1 and LCC9 cell lines, A. Western blot analysis of total GRP78 protein are shown in both cell lines in different conditions, B-C. Knockdown of IRE1α, D-F and XBP1, J-L, significantly increased inhibition of cell growth in glutamine-only conditions in both cell lines. However, inhibition of JNK with SP600125 significantly decreased the inhibition of cell growth in glutamine-only conditions, G-I. Also, knockdown of MYC, M-O, significantly decreased inhibition of cell growth in glutamine-only conditions. Overall, MYC may have facilitate an IRE1α-XBP1 pathway to promote cell survival during glutamine-only conditions, and an IRE1α-phospho-JNK pathway to promote cell death in this condition. ANOVA, p ≤0.001; *p < 0.05 for respective cell lines transfected with indicated siRNA (or treated with SP600125, for JNK) compared with control siRNA (or vehicle alone, for JNK) in glutamine-only conditions.

Article Snippet: 5 μM MYC siRNA (SMARTpool: ON-TARGETplus set of four MYC siRNA Dharmacon, Lafayette, CO), 10 GLS1, GRP78 (HSPA5), IRE1a or XBP1 (10 nM of 3 unique 27mer siRNA duplexes; Origene, Rockville, MD) or their respective control siRNA, were transfected using the TransIT-siQUEST (Mirus, Madison, WI) transfection reagent.

Techniques: Transfection, Inhibition, Western Blot, Knockdown, Control

Journal: Oncotarget

Article Title: Unfolded protein response signaling impacts macrophage polarity to modulate breast cancer cell clearance and melanoma immune checkpoint therapy responsiveness

doi: 10.18632/oncotarget.19849

Figure Lengend Snippet: ( A ) RAW 264.7 cells were pretreated with vehicle, 1 mM DTT, or 1 μg/mL tunicamycin (Tn) for 4 hours before treatment with 1 μg/mL LPS for 24 hours. GRP78, IRE1, and PERK were measured by Western blot hybridization. Protein loading was normalized to β-actin. n = 3; * p < 0.05. ( B ) RAW 264.7 cells were pretreated with vehicle, 1 mM DTT, or 1 μg/mL tunicamycin (Tn) for 4 hours before treated with 1 μg/mL LPS for 24 hours. iNOS and Arg-1 were measured by Western blot hybridization. Protein loading was normalized to β-actin. n = 3; * p < 0.05. ( C ) RT-PCR of iNOS, ARG-1, IL-6, IL-10, and IL-12 in RAW 264.7 cells pretreated with vehicle, 1 mM DTT, or 1 μg/mL tunicamycin (Tn) for 4 hours before treatment with 1 μg/mL LPS for 24 hours. Gene expression was normalized to the 18s housekeeping gene. n = 5; * p < 0.05. ( D ) 4T1 breast cancer cells were plated in an ACEA E-plate for 24 hours; RAW 264.7 macrophages pretreated with vehicle, 1 mM DTT, or 1 μg/mL Tn for 24 hours were then added to the E-plate. Each well was treated with 1 μg/mL LPS and the cell index was measured at 8 hours by electrical impedance. n = 3; * p < 0.05.

Article Snippet: The following materials were obtained as indicated: Mouse specific GRP78 ( HSPA5 ), IRE1 ( ERN1 ), and PERK ( EIF2AK3 ) siRNA (Origene, Rockville, MD). ePlates were purchased from ACEA.

Techniques: Western Blot, Hybridization, Reverse Transcription Polymerase Chain Reaction, Gene Expression

Journal: Oncotarget

Article Title: Unfolded protein response signaling impacts macrophage polarity to modulate breast cancer cell clearance and melanoma immune checkpoint therapy responsiveness

doi: 10.18632/oncotarget.19849

Figure Lengend Snippet: ( A ) UPR signaling components IRE1, PERK, and GRP78 in RAW 264.7 cells were inhibited by RNAi treatment for 24 hours followed by treatment with 1 μg/mL LPS for 24 hours. iNOS, Arg-1, PERK, IRE1, and GRP78 were measured by Western blot hybridization. Protein loading was normalized to β-actin. ( B ) UPR signaling components IRE1, PERK, and GRP78 were inhibited by RNAi for 24 hours and then plated in an ACEA E-plate. Each well was treated with 1 μg/mL LPS and the cell index was measured every 12 hours by electrical impedance. n = 3; * p < 0.05. ( C ) 4T1B breast cancer cells were plated in an ACEA E-plate for 24 hours; then 5 × 10 4 control, IRE1, PERK, or GRP78 siRNA transfected RAW 264.7 macrophages were added to the E-plate. Each well was treated with 1 μg/mL LPS and the cell index was measured every 4 hours by electrical impedance. n = 3; * p < 0.05. ( D ) RT-PCR analysis of iNOS, ARG-1, IL-6, IL-10, and IL-12 gene expression in control, IRE1, PERK, or GRP78 siRNA transfected RAW 264.7 cells treated with 1 μg/mL LPS for 24 hours. Gene expression was normalized to 18S housekeeping gene. n = 4; * p < 0.05.

Article Snippet: The following materials were obtained as indicated: Mouse specific GRP78 ( HSPA5 ), IRE1 ( ERN1 ), and PERK ( EIF2AK3 ) siRNA (Origene, Rockville, MD). ePlates were purchased from ACEA.

Techniques: Western Blot, Hybridization, Control, Transfection, Reverse Transcription Polymerase Chain Reaction, Gene Expression

Journal: Oncotarget

Article Title: Unfolded protein response signaling impacts macrophage polarity to modulate breast cancer cell clearance and melanoma immune checkpoint therapy responsiveness

doi: 10.18632/oncotarget.19849

Figure Lengend Snippet: ( A ) RAW 264.7 macrophage cells were transfected with control, GRP78, IRE1, or PERK siRNA for 24 hours, and then treated with or without LPS. Intracellular lipids were stained for oil-red-o and representative images obtained at 40×. ( B ) UPR signaling components IRE1, PERK, and GRP78 in RAW 264.7 cells were inhibited by RNAi treatment for 24 hours followed by treatment with 1 μg/mL LPS for 24 hours. ATGL was measured by Western blot hybridization. Protein loading was normalized to β-actin. ( C ) Glucose uptake in RAW 264.7 cells transfected with scrambled control or UPR targeting siRNA for 24 hours. n = 3; * p < 0.05. ( D ) Mitochondrial metabolomics was determined in transfected RAW 264.7 macrophages using a Seahorse Bioanalyzer. ( E ) Basal ECAR rates in transfected RAW 264.7 macrophage cells. n = 3–4; * p < 0.05.

Article Snippet: The following materials were obtained as indicated: Mouse specific GRP78 ( HSPA5 ), IRE1 ( ERN1 ), and PERK ( EIF2AK3 ) siRNA (Origene, Rockville, MD). ePlates were purchased from ACEA.

Techniques: Transfection, Control, Staining, Western Blot, Hybridization

Journal: Oncotarget

Article Title: Unfolded protein response signaling impacts macrophage polarity to modulate breast cancer cell clearance and melanoma immune checkpoint therapy responsiveness

doi: 10.18632/oncotarget.19849

Figure Lengend Snippet: ( A ) Protein lysates from IFNγ-treated bone marrow cells from wild-type and GRP78 heterozygous mice were analyzed for iNOS, ARG-1, and β-actin by Western blot hybridization. n = 3; * p < 0.05. ( B ) Protein lysates from IFNγ-treated bone marrow cells from wild-type and GRP78 heterozygous mice were analyzed for PERK, GRP78, IRE1, and β-actin by Western blot hybridization. n = 3; * p < 0.05. ( C ) 4T1B breast cancer cells were plated in an ACEA E-plate for 24 hours and then wild-type or GRP78 heterozygous CD11b+ cells that were pre-treated with IFNγ for 48 hours + LPS were added to the E-plate. Cell index was measured every 6 hours by electrical impedance. n = 3; * p < 0.05. ( D ) Signaling schematic representing how UPR targeting differentially regulates cellular bioenergetics to control macrophage polarity.

Article Snippet: The following materials were obtained as indicated: Mouse specific GRP78 ( HSPA5 ), IRE1 ( ERN1 ), and PERK ( EIF2AK3 ) siRNA (Origene, Rockville, MD). ePlates were purchased from ACEA.

Techniques: Western Blot, Hybridization, Control

Journal: Oncotarget

Article Title: Unfolded protein response signaling impacts macrophage polarity to modulate breast cancer cell clearance and melanoma immune checkpoint therapy responsiveness

doi: 10.18632/oncotarget.19849

Figure Lengend Snippet: ( A ) Control, IRE1, PERK, or GRP78 transfected 4T1B breast cancer cells were plated in an ACEA E-plate and RAW 264.7 macrophages were added to the E-plate. Each well was treated with 1 μg/mL LPS and the cell index was measured by electrical impedance. n = 3; * p < 0.05. ( B ) Conditioned media from control or GRP78 siRNA transfected 4T1B or ZR-75-1 breast cancer cells were used to treat RAW 264.7 cells for 24 hours. iNOS and Arg-1 were measured by Western blot hybridization. Protein loading was normalized to β-actin. n = 3; * p < 0.05. ( C ) Vehicle treated, LPS treated, ZR-75-1 conditioned media (CM), or GRP78-silenced ZR-75-1 conditioned media were used to treat RAW 264.7 macrophage cells for 24 hours. Macrophages were stained for CD68-FITC (green), CD80-Cy5 (red), or CD206-cy7 (pink) and counterstained with DAPI. The M1/M2-like macrophage population was determined by immunocytochemistry. ( D ) Vehicle treated, LPS treated, ZR-75-1 conditioned media, or GRP78-silenced ZR-75-1 conditioned media were used to treat RAW 264.7 macrophage cells for 24 hours. Macrophages were stained for IL12-Cy3 (yellow) or IL10-cy7 (pink) and counterstained with DAPI. The M1/M2-like macrophage population was determined by immunocytochemistry.

Article Snippet: The following materials were obtained as indicated: Mouse specific GRP78 ( HSPA5 ), IRE1 ( ERN1 ), and PERK ( EIF2AK3 ) siRNA (Origene, Rockville, MD). ePlates were purchased from ACEA.

Techniques: Control, Transfection, Western Blot, Hybridization, Staining, Immunocytochemistry

Journal: Oncotarget

Article Title: Unfolded protein response signaling impacts macrophage polarity to modulate breast cancer cell clearance and melanoma immune checkpoint therapy responsiveness

doi: 10.18632/oncotarget.19849

Figure Lengend Snippet: ( A ) IL-12p70, IL-1β, IL-6, TARC, Eotaxin, and RANTES were measured by ELISA from serum of WT and GRP78 morpholino treated mice. n = 4; * p < 0.05. ( B ) DMBA-induced mammary tumors from WT and GRP78 heterozygous mice (untreated or treated with tamoxifen) were stained with fluorescently labelled CD68 (green) or CD80 (red). Tumor sections were counterstained with DAPI. ( C ) Control, IRE1, PERK, or GRP78 transfected 4T1B breast cancer cells were plated in an ACEA E-plate and then control, IRE1, PERK, or GRP78 transfected RAW 264.7 macrophages were added to the E-plate. Each well was treated with 1 μg/mL LPS and the cell index was measured every 12 hours by electrical impedance. n = 3; * p < 0.05.

Article Snippet: The following materials were obtained as indicated: Mouse specific GRP78 ( HSPA5 ), IRE1 ( ERN1 ), and PERK ( EIF2AK3 ) siRNA (Origene, Rockville, MD). ePlates were purchased from ACEA.

Techniques: Enzyme-linked Immunosorbent Assay, Staining, Control, Transfection

Journal: Oncotarget

Article Title: Unfolded protein response signaling impacts macrophage polarity to modulate breast cancer cell clearance and melanoma immune checkpoint therapy responsiveness

doi: 10.18632/oncotarget.19849

Figure Lengend Snippet: ( A ) Western blot analysis of Arg-1 and iNOS protein expression from matched human PBMC isolated from melanoma patients before Ipilimumab treatment or after progression of disease (Ipilimumab resistance). n = 5 * p = 0.02. ( B ) Western blot analysis of PERK, GRP78, and IRE1 protein expression from matched human PBMC isolated from melanoma patients before Ipilimumab treatment or after progression of disease (Ipilimumab resistance). n = 6; * p = 0.003 ( C ) Adherent PBMC from matched human PBMC isolated from melanoma patients before treatment and after ipilimumab therapy resistance were stained with CD206 M2-like macrophage marker and PERK antibodies. Cells were counterstained with DAPI.

Article Snippet: The following materials were obtained as indicated: Mouse specific GRP78 ( HSPA5 ), IRE1 ( ERN1 ), and PERK ( EIF2AK3 ) siRNA (Origene, Rockville, MD). ePlates were purchased from ACEA.

Techniques: Western Blot, Expressing, Isolation, Staining, Marker

Journal: Journal of clinical immunology

Article Title: Anti-citrullinated protein antibodies activated ERK1/2 and JNK mitogen-activated protein kinases via binding to surface-expressed citrullinated GRP78 on mononuclear cells.

doi: 10.1007/s10875-012-9841-6

Figure Lengend Snippet: Fig. 2 Effect of ACPAs and different protein molecules on the ratio of p-ERK1/2/ERK1/2, p-JNK/JNK and p-p38/p38 MAPKs in U937 cells. The concentration of different proteins were: non-specific human IgG (6 μg/ml), ACPAs (100 IU/ml), ACPA-depleted [ACPA(−) RA] sera (6 μg/ml), ACPAs (100 IU/ml)+anti-Toll-like receptor 4 blocking antibody (anti-TLR4, 10 μg/ml), ACPAs (100 IU/ml)+cit-GRP78 (10 μg/ml) or ACPAs (100 IU/ml)+GRP78 (10 μg/ml) for 24 h. The cells were lysed and both phosphorylated and native forms of three MAPKs were analyzed by Western blotting. a pERK1/2/ERK1/2 ratio, b p-JNK/JNK ratio, c p-p38/p38 ratio, and d A representative example to show the expression of phosphorylated and native forms of MAPKs after incubation with different proteins by Western blot; * denote p<0.05 compared to the non-specific human IgG

Article Snippet: Recombinant GRP78 (Novus Biologicals, Littleton, CO, USA) was successfully citrullinated (cit-GRP78) after incubation of the molecules with 10 mM CaCl2 and peptidylarginine deiminase (Sigma-Aldrich) at a concentration of 20 IU per 1 mg GRP78 protein for 2 h.

Techniques: Concentration Assay, Blocking Assay, Western Blot, Expressing, Incubation

Journal: Journal of clinical immunology

Article Title: Anti-citrullinated protein antibodies activated ERK1/2 and JNK mitogen-activated protein kinases via binding to surface-expressed citrullinated GRP78 on mononuclear cells.

doi: 10.1007/s10875-012-9841-6

Figure Lengend Snippet: Fig. 4 Effect of ACPAs and different protein molecules on p-Akt/Akt and p-IKK-α/IKK-α ratio of U937 cells. U937 cells (1×106 cells/ml) were incubat- ed with non-specific human IgG (5 μg/ml), ACPAs (80 IU/ ml), ACPA(−) RA sera (5 μg/ ml), ACPAs (80 IU/ml)+anti- TLR4 (10 μg/ml), ACPAs (80 IU/ml)+cit-GRP78 (10 μg/ ml) for 24 h. The cells were lysed and both phosphorylated and native form of a Akt and b IKK-α were analyzed by West- ern blotting. * denote p<0.05 compared to the non-specific human IgG

Article Snippet: Recombinant GRP78 (Novus Biologicals, Littleton, CO, USA) was successfully citrullinated (cit-GRP78) after incubation of the molecules with 10 mM CaCl2 and peptidylarginine deiminase (Sigma-Aldrich) at a concentration of 20 IU per 1 mg GRP78 protein for 2 h.

Techniques:

Journal: Journal of clinical immunology

Article Title: Anti-citrullinated protein antibodies activated ERK1/2 and JNK mitogen-activated protein kinases via binding to surface-expressed citrullinated GRP78 on mononuclear cells.

doi: 10.1007/s10875-012-9841-6

Figure Lengend Snippet: Fig. 5 Comparison of citrullinated (cit-GRP78) and native forms (GRP78) GRP78 molecules on the surface mem- brane extracts (150 μg) among U937 cells, normal human PBMCs, and RA PBMCs by Western blotting. The expres- sion of transferrin receptor (TfR) was used as an internal control. a The expression ratio of cit-GRP78/TfR among the three groups b The expression of GRP78/TfR among three groups c The expression ratio of cit-GRP78/GRP78 among three groups. d A representative example is shown

Article Snippet: Recombinant GRP78 (Novus Biologicals, Littleton, CO, USA) was successfully citrullinated (cit-GRP78) after incubation of the molecules with 10 mM CaCl2 and peptidylarginine deiminase (Sigma-Aldrich) at a concentration of 20 IU per 1 mg GRP78 protein for 2 h.

Techniques: Comparison, Western Blot, Control, Expressing

Journal: Journal of clinical immunology

Article Title: Anti-citrullinated protein antibodies activated ERK1/2 and JNK mitogen-activated protein kinases via binding to surface-expressed citrullinated GRP78 on mononuclear cells.

doi: 10.1007/s10875-012-9841-6

Figure Lengend Snippet: Fig. 6 A schematic diagram to indicate that the MAPK signaling pathways are transduced by ACPAs after binding to surface- expressed cit-GRP78 on RA PBMCs that leads to TNF-α production

Article Snippet: Recombinant GRP78 (Novus Biologicals, Littleton, CO, USA) was successfully citrullinated (cit-GRP78) after incubation of the molecules with 10 mM CaCl2 and peptidylarginine deiminase (Sigma-Aldrich) at a concentration of 20 IU per 1 mg GRP78 protein for 2 h.

Techniques: Protein-Protein interactions, Binding Assay

Journal: Cell reports

Article Title: TAILS N-Terminomics and Proteomics Show Protein Degradation Dominates over Proteolytic Processing by Cathepsins in Pancreatic Tumors.

doi: 10.1016/j.celrep.2016.06.086

Figure Lengend Snippet: Figure 6. Structural Modeling of GRP78 Processing by Cathepsins (A and B) Two distinct neo-N termini were identified using TAILS; whereas cleavage in the hinge region (L418YD419; depicted in red, A) results in separation of the ATPase domain of the protein interaction domain, the second cleavage site (N249YG250; depicted in blue, B) is located centrally in the ATPase domain, dismantling the nucleotide binding site and thus inactivating the enzyme. ADP is shown in yellow as a ball and stick model, and calcium is depicted as a red sphere.

Article Snippet: Proteins (30 mg) were resolved on a 10% Bis-Tris mini protein gel (Invitrogen) and transfer membranes probed with polyclonal anti-GRP78 goat antibody (R&D systems) and were revealed by Donkey anti-goat-HRP secondary (Jackson Immunoresearch) with Enhanced Chemiluminescent substrate (ThermoFisher).

Techniques: Binding Assay

Journal: Frontiers in Immunology

Article Title: Secreted glucose regulated protein78 ameliorates DSS-induced mouse colitis

doi: 10.3389/fimmu.2023.986175

Figure Lengend Snippet: GRP78 is decreased in the intestinal mucosa of UC patients. (A) Representative immunofluorescent staining of GRP78 in human colonic tissues (left). Scare bar: 50 μm. The quantitative results about the percentage of GRP78 immunoreactive cells per field of view was shown in right. sGRP78 (B) and TNF-α (C) levels in the supernatants of mucosa punches collected from UC (n=8) and non-colitic control (n=6) patients. The levels were normalized to colonic surface area. Results are expressed as mean ± standard error of mean (SEM). ns, not significant. * p < 0.05, ** p < 0.01.

Article Snippet: Female C57BL/6 mice (HFK Bioscience, Beijing, China) were randomly divided into following groups: non-colitic control group; colitic group receiving saline (200 μl); colitic group receiving sGRP78 (10 mg/kg) or selective GRP78 serine proteinase subA (20 μg/mouse) intraperitoneally.

Techniques: Staining

Journal: Frontiers in Immunology

Article Title: Secreted glucose regulated protein78 ameliorates DSS-induced mouse colitis

doi: 10.3389/fimmu.2023.986175

Figure Lengend Snippet: sGRP78 expression in experimental models of colitis. (A) Scheme of colitis induction. (B) Body weight was measured every two days and expressed as the percentage from day 0. At indicated time points, the colonic tissues were removed from sacrificed mice for following measurements: (C) Colon length. (D) Histological scores. Levels of TNF-α (E) and sGRP78 (F) in colonic punches supernatants. (G) Correlation analysis between sGRP78 and TNF-α (n=39). Pearson correlation coefficients (r) and p-values were calculated and shown. Relationship curve of sGRP78 with weight loss (H) , colon shortening (I) , and histological changes (J) in colitis. (K) Representative photomicrographs of colon sections (left) stained with HE (upper panel) or GRP78 (lower panel). (Day 0: black circle; Day2: red square; Day4: blue regular triangle; Day6: Purple inverted triangle; Day8: orange rhombus; Day10: black pentagram; Day12: brown error). Scale bar: 100 μm (upper panel) and 50 μm (lower panel). Numbers of GRP78-positive cells were calculated and shown in the right. Data represent mean ± SEM (n=5–6). ns, not significant. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Female C57BL/6 mice (HFK Bioscience, Beijing, China) were randomly divided into following groups: non-colitic control group; colitic group receiving saline (200 μl); colitic group receiving sGRP78 (10 mg/kg) or selective GRP78 serine proteinase subA (20 μg/mouse) intraperitoneally.

Techniques: Expressing, Staining

Journal: Frontiers in Immunology

Article Title: Secreted glucose regulated protein78 ameliorates DSS-induced mouse colitis

doi: 10.3389/fimmu.2023.986175

Figure Lengend Snippet: sGRP78 induced macrophage M2 polarization in colitis. Representative immunofluorescent images of GRP78 (green) and CD68 (red) stained human (A) and mouse (B) colonic mucosa. Scale bar: 50 μm. (C) Frequencies of F4/80 + cells in lamina propria (LP) and their CD80 + or CD206 + expressions. Upper: gating strategy followed to distinguish the LP macrophages. Middle: representative FCM dot plots. Lower: FCM plots based quantitative analysis for F4/80 + CD11b + macrophages. (D) Frequencies of indicated cell populations in spleen. (E) Frequencies of indicated cell populations in MLN. (F) mRNA expression of Arg1 , Fizz1 , Ym1 , Mgl1 , and Inos in mice colonic tissues (Ctrl group: black circle; DSS+NS group: red square; DSS+GRP78 group: blue regular triangle). Data represent mean ± SEM (n=5). ns, not significant. * p < 0.05, ** p < 0.01, **** p < 0.0001.

Article Snippet: Female C57BL/6 mice (HFK Bioscience, Beijing, China) were randomly divided into following groups: non-colitic control group; colitic group receiving saline (200 μl); colitic group receiving sGRP78 (10 mg/kg) or selective GRP78 serine proteinase subA (20 μg/mouse) intraperitoneally.

Techniques: Staining, Expressing

Journal: Frontiers in Immunology

Article Title: Secreted glucose regulated protein78 ameliorates DSS-induced mouse colitis

doi: 10.3389/fimmu.2023.986175

Figure Lengend Snippet: Adoptive transfer of sGRP78-conditioned BMDMs ameliorated DSS-induced colitis. (A) Timetable scheme showing DSS and BMDMs administration in mice. Effect of sGRP78-induced BMDMs on (B) weight loss, (C) DAI score (n=5) in DSS exposed mice. (D) Representative gross picture of colons (left) and quantification of colon length (right). (E) Representative HE-staining (left) and respective colitis scores of indicated groups (right). Scale bar: 100 μm. (F) mRNA expression of Arg1, Fizz1,Il10, Inos, Il6, Tnfα, Occludin in colonic tissues. (G) FITC-dextran concentrations in serum of mice. (Ctrl group: black circle; DSS+NS group: green square; DSS+BMDMs group: red regular triangle; DSS+BMDMs (GRP78) group: Purple inverted triangle). Data represent mean ± SEM (n=4-5). ns, not significant. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Female C57BL/6 mice (HFK Bioscience, Beijing, China) were randomly divided into following groups: non-colitic control group; colitic group receiving saline (200 μl); colitic group receiving sGRP78 (10 mg/kg) or selective GRP78 serine proteinase subA (20 μg/mouse) intraperitoneally.

Techniques: Adoptive Transfer Assay, Staining, Expressing