Journal: Journal of Advanced Research

Article Title: Novel function of macrophage migration inhibitory factor in regulating post-infarct inflammation and the therapeutic significance

doi: 10.1016/j.jare.2025.05.030

Figure Lengend Snippet: Macrophage migration inhibitory factor (MIF) promotes inflammatory cell migration and infiltration through C – C motif chemokine receptor 2 (CCR2) and C-X-C motif chemokine receptor (CXCR) 4 and promotes splenic monocyte mobilization via interaction with angiotensin Ⅱ type 1 receptor (AT-1R) after myocardial infarction (MI). A, peripheral blood mononuclear cells (PBMCs) chemotaxis in response to homogenized sham or infarct heart tissue from wild type (WT) mice at day 3 post-MI, or MI homogenate together with inhibition of C–C motif chemokine receptor 2 (CCR2), C-X-C motif chemokine receptor (CXCR) 2 and CXCR4, respectively. Cells/HPF, cells/high power field. PBMCs harvested from WT mice at 3 days post-MI. CCR2i, CCR2 inhibition. CXCR2i, CXCR2 inhibition. CXCR4i, CXCR4 inhibition. n = 4–8 per group. *** P < 0.001 vs. sham . B, PBMCs chemotaxis in response to recombinant human MIF (rMIF) or rMIF with inhibition of CCR2, CXCR2 and CXCR4, respectively. PBMCs harvested from WT mice at 3 days post-MI. n = 4–8 per group. *** P < 0.001 vs. control . C, Representative immunoblotting images for monocyte chemoattractant protein-1 (MCP-1), CXCR4 and internal reference protein (heat shock protein 60, HSP-60) in hearts from WT and global MIF deficient (MIFKO) mice with sham-operation or 72 h MI. D-E, Quantitative analysis of MCP-1 and CXCR4 expression. n = 6–9 per group. ** P < 0.01 vs. sham in the same genotype. *** P < 0.001 vs. sham in the same genotype. F, Splenic monocyte (from normal WT mice) chemotaxis in response to plasma from normal mice or mice with 1.5 h MI, or MI plasma together with losartan. Lor, losartan, AT-1R inhibitor. n = 6–8 per group. *** P < 0.001 vs. normal. G, Splenic monocytes (from normal WT mice) chemotaxis in response to rMIF and angiotensin II (Ang II), or together with losartan addition, respectively. n = 6–8 per group. ** P < 0.01 vs. Control. *** P < 0.001 vs. Control. H, Representative immunoblotting images for AT-1R and internal reference protein (HSP-60) in spleens from WT and global MIF deficient (MIFKO) mice with sham-operation and 1.5 h MI. I, Quantitative analysis of AT-1R expression. n = 8 per group. *** P < 0.001 vs. sham in the same genotype . J. Co-immunoprecipitation (Co-IP) assay using an anti-AT-1R antibody or IgG demonstrated the interaction between AT-1R and MIF, as well as between AT-1R and CD74 in splenic monocytes and spleen tissue. Input, input sample. IgG, negative control sample. IP, Co-IP sample.

Article Snippet: Primary antibodies against MCP-1 (1:1000), CCR2 (1:1000), CXCR2 (1:1000), CXCR4 (1:1000), tissue inhibitor of metalloproteinases-4 (Timp-4, 1:1000), AT-1R (AGTR-1, 1:1000), heat shock protein 60 (HSP-60, 1:1000) and HSP-90 (1:1000) were purchased from ProteinTech.

Techniques: Migration, Chemotaxis Assay, Inhibition, Recombinant, Control, Western Blot, Expressing, Clinical Proteomics, Co-Immunoprecipitation Assay, Negative Control

Journal: medRxiv

Article Title: Anomalous epithelial variations and ectopic inflammatory response in chronic obstructive pulmonary disease

doi: 10.1101/2020.12.03.20242412

Figure Lengend Snippet: A. Representative images immunostained with CCL2, TNC, and CXCL8 with AT2 cell markers (SFTPC or ABCA3) in COPD and never-smoker lung sections. B . Violin plots for COL4A1, COL4A2, and COL4A3 in AT1 clusters. C. Violin plots for FKBP5 in selected cell types. a) present dataset; b) combined publicly-available datasets.

Article Snippet: Antibodies and reagents were as follows: anti-PD-L1 (rabbit, 1:1000, #ab205921, Abcam, Cambridge, UK); anti-TNC (rabbit, 1:100, #HPA004823, Sigma-Aldrich); anti-CCL2 (rabbit, 1:100, #HPA019163, Sigma-Aldrich); anti-RAGE (rabbit, 1:1000, #ab216329, Abcam); anti-SFTPC (rabbit, 1:1000, #HPA010928, Sigma-Aldrich); anti-ABCA3 (mouse, 1:1000, #WMAB-ABCA3-17, Seven Hills Bioreagents, OH, USA); anti- macrophage inflammatory protein 3 alpha (rabbit, 1:1000, #ab224188, Abcam); anti- CXCL1 (mouse, 1:100, #MAB275, R&D Systems, MN, USA); anti-CXCL8 (mouse, 1:100, #MAB208, R&D Systems); and Hoechst 33242 (1:1000, #H342, Sigma-Aldrich).

Techniques:

Journal: medRxiv

Article Title: Anomalous epithelial variations and ectopic inflammatory response in chronic obstructive pulmonary disease

doi: 10.1101/2020.12.03.20242412

Figure Lengend Snippet: A. 100% stacked bar charts of the percentage of cell populations for AT1 subtype clusters, AT2 subtype clusters, and basal lineage clusters. B. Bar charts displaying the percentages of subpopulations such as AT1-B, AT2-A, AT2-B, club, goblet, and basal clusters across patient states. C. a UMAP plot of epithelial cells focusing on the AT2-C cluster (left). The cell population of the AT2-C cluster (right). The y-axis represents the ratio of cells in the AT2-C cluster across patient states. Brackets () represent the percentage of cells of the AT2-C cluster based on epithelial cells in each of the patient states. D. Violin plots of representative inflammatory-related genes displaying the AT2-C (iAT2) cluster. E. Violin plots for CXCL1 and CXCL8 in AT2 cells in combined publicly-available datasets. F. Representative images immunostained with CD274 (PD-L1), CXCL1 and CXCL20 with AT2 cell markers (SFTPC or ABCA3) in COPD and never-smoker lung sections. Scale bars, 25 μm (left), 12.5 μm (right).

Article Snippet: Antibodies and reagents were as follows: anti-PD-L1 (rabbit, 1:1000, #ab205921, Abcam, Cambridge, UK); anti-TNC (rabbit, 1:100, #HPA004823, Sigma-Aldrich); anti-CCL2 (rabbit, 1:100, #HPA019163, Sigma-Aldrich); anti-RAGE (rabbit, 1:1000, #ab216329, Abcam); anti-SFTPC (rabbit, 1:1000, #HPA010928, Sigma-Aldrich); anti-ABCA3 (mouse, 1:1000, #WMAB-ABCA3-17, Seven Hills Bioreagents, OH, USA); anti- macrophage inflammatory protein 3 alpha (rabbit, 1:1000, #ab224188, Abcam); anti- CXCL1 (mouse, 1:100, #MAB275, R&D Systems, MN, USA); anti-CXCL8 (mouse, 1:100, #MAB208, R&D Systems); and Hoechst 33242 (1:1000, #H342, Sigma-Aldrich).

Techniques:

Journal: Frontiers in Microbiology

Article Title: Probing Clostridium difficile Infection in Complex Human Gut Cellular Models

doi: 10.3389/fmicb.2019.00879

Figure Lengend Snippet: Production of C. difficile spores, toxins, and host responses to infection. (A) Colony counts of spores recovered after heat treatment, and total cells in the cell-associated C. difficile fraction (infected cell lysates) in the 2D epithelial model. Data shown are the mean of three independent experiments and error bars indicate SD, * p < 0.05 as determined by two-way ANOVA. (B) ELISA for C. difficile toxins A and B shows increased toxin production after extended infection. Toxins were measured from medium obtained from the apical compartment containing uninfected cell layers incubated for 24 or 48 h (Control) or cells infected with C. difficile for 3, 6, 24, or 48 h. Data shown are the mean of three independent experiments and error bars indicate SD, * p < 0.05 as determined by two-way ANOVA. Gray line represents the sensitivity of the test at 0.5 ng/ml. (C) ELISA for human IL-8 indicates increased IL-8 production at 24 and 48 h p.i. IL-8 was measured in medium obtained from the basolateral compartment containing uninfected cell layers incubated for 3, 6, 24, or 48 h (Control) or cells infected with C. difficile for 3–48 h. Gray line represents the limit of detection at 32 pg/ml. Data shown are the mean of three independent experiments and error bars indicate SD, * p < 0.05, ** p < 0.01, and *** p < 0.001, **** p < 0.0001 as determined by one-way ANOVA with Tukey’s test for multiple comparison.

Article Snippet: IL-8 production was also determined by analysis of basolateral supernatants from the VDC using a human IL-8 ELISA kit (R&D systems, Minneapolis, MN, United States) following the manufacturer’s instruction.

Techniques: Infection, Enzyme-linked Immunosorbent Assay, Incubation, Control, Comparison

Journal: Frontiers in Microbiology

Article Title: Probing Clostridium difficile Infection in Complex Human Gut Cellular Models

doi: 10.3389/fmicb.2019.00879

Figure Lengend Snippet: C. difficile spores and toxin production, and host response to infection in the 3D-VDC model. (A) Colony counts of spores and total cells in the host cell-associated C. difficile fraction (infected cell lysates). Data shown are the mean of three independent experiments and error bars indicate SD, ns, not significant as determined by two-way ANOVA. (B) Toxin A and B levels from apical compartment supernatants as determined by ELISA in the 3D model. Data shown are the mean of three independent experiments and error bars indicate SD, ns, not significant as determined by two-way ANOVA. Gray line represents the sensitivity of the test at 0.5 ng/ml. (C) Human IL-8 levels in supernatants from the basolateral compartments with uninfected cells incubated for 24 h or with cells infected with C. difficile for 3 and 24 h, as determined by ELISA. Gray line represents the limit of detection at 32 pg/ml. Data shown are the mean of three independent experiments and error bars indicate SD, ** p < 0.01, as determined by the one-way ANOVA with Tukey’s test for multiple comparison.

Article Snippet: IL-8 production was also determined by analysis of basolateral supernatants from the VDC using a human IL-8 ELISA kit (R&D systems, Minneapolis, MN, United States) following the manufacturer’s instruction.

Techniques: Infection, Enzyme-linked Immunosorbent Assay, Incubation, Comparison