Journal: Frontiers in Cardiovascular Medicine

Article Title: Experimental renal transplantation in rats improves cardiac dysfunction caused by chronic kidney disease while LVH persists

doi: 10.3389/fcvm.2023.1200323

Figure Lengend Snippet: Changes in markers of cardiac hypertrophy, fibrosis and inflammation in CKD and after renal function restoration by transplantation. mRNA expression of BNP ( A ), FGF23 ( B ), FGFR4 ( C ), fibronectin ( D ), CTGF ( K ) and IL-6 ( L ) was analyzed in hearts from sham, SNx and SNx rat with renal transplant using Real-time PCR. Cardiac fibronectin ( E ) was analyzed by immunohistochemistry using a semi-quantitative score and examples of cardiac fibronectin staining in controls and SNx rats are shown ( F , brown staining). The myofibroblast marker collagen 1 was analyzed semi-quantitatively in heart sections ( G ). Representative pictures of collagen staining in hearts of sham control, SNx and RTx rats are shown ( H , brown staining). Perivascular microscarring was analyzed using a semi-quantitative score ( I ) and collagen 1 stained sections ( J ). Cardiac macrophages were analyzed by counting of ED1-positive cells detected using immunohistochemistry ( M ). * p < 0.05 vs. control, ** p < 0.01 vs. control, *** p < 0.001 vs. control, **** p < 0.0001, # p < 0.05 vs. SNx, ## p < 0.01 vs. SNx, ### p < 0.001 vs. SNx.

Article Snippet: The following primary antibodies were used: a polyclonal rabbit anti-fibronectin antibody (ab2413, Abcam, Cambridge, GB); a polyclonal rabbit anti-collagen 1 antibody (Novus Biologicals, LLC, Centennial, CO, USA) and a monoclonal mouse anti-ED1 antibody (Bio-Rad Laboratories GmbH, Feldkirchen, Germany).

Techniques: Transplantation Assay, Expressing, Real-time Polymerase Chain Reaction, Immunohistochemistry, Staining, Marker, Control

Journal: International Journal of Molecular Sciences

Article Title: Seizures in PPT1 Knock-In Mice Are Associated with Inflammatory Activation of Microglia

doi: 10.3390/ijms23105586

Figure Lengend Snippet: Western blot analysis of glial cell markers and synaptic receptors in hippocampus of PPT1 KI mice and WT mice. ( A ) Representative Western blots ( top ) and the bar graph ( bottom ) show the expression level of Iba-1 in hippocampus of 7-month-old PPT1 KI mice with seizures and age-matched WT mice ( p = 0.04, Permutation t test, n = 6). ( B , C ) Representative Western blots ( top ) and the bar graph ( bottom ) show that expressions of Iba-1 in the hippocampus of PPT1 KI mice and age-matched WT mice from age 1 to 7 months ( B , Iba-1 in PPT1 KI mice, 6 vs. 7 months old, p < 0.001, n = 35; C , Iba-1 in WT mice, n = 35); ( D , E ) Representative Western blots ( top ) and the bar graph ( bottom ) show expression of CD68 in the hippocampus of PPT1 KI mice and age-matched WT mice from age 1 to 7 months ( D , CD68 in PPT1 KI mice, 1 vs. 4 months old, p = 0.049, 6 vs. 7 months old, p = 0.03, n = 21; E , CD68 in WT mice, n = 35); ( F , G ) Representative Western blots ( top ) and the bar graph ( bottom ) show expression of GABA A Rα1 in the hippocampus of PPT1 KI mice and age-matched WT mice from age 1 to 7 months ( F , GABA A Rα1 in PPT1 KI mice, 1 vs. 7 months old, p < 0.01, n = 35; G, GABA A Rα1 in WT mice, n = 35). * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: The primary antibodies used for the immunoblots were as follows: Iba-1 (A19776, 1:1000, ABclonal), CD68 (E-AB-40066,1:1000, Elabscience), glial fibrillary acidic protein (GFAP; 60190-1-Ig, 1:5000, Proteintech); GABA A Rα1 (ab252430, 1:1000, Abcam); GluN2B (21920-1-AP, 1:1000, Proteintech), and β-actin (ab8226, 1:500, Abcam) as a loading control.

Techniques: Western Blot, Expressing

Journal: Journal of experimental & clinical cancer research : CR

Article Title: Gastric cancer-derived exosomal let-7 g-5p mediated by SERPINE1 promotes macrophage M2 polarization and gastric cancer progression.

doi: 10.1186/s13046-024-03269-4

Figure Lengend Snippet: Fig. 2 High SERPINE1 expression in GC cells promotes macrophage M2 polarization. tSNE visualization of nine single-cell clusters partitioned by unsu pervised cluster analysis, SERPINE1 expression of each single-cell, and SERPINE1 expression abundance of different single-cell clusters in the GSE134520 (A–C) and GSE167297 (D–F) datasets. (G) Flow cytometry analysis of the proportion of CD68+CD206+ macrophages in a Transwell co-culture system, with MKN45 and AGS cells overexpressing (oe_SERPINE1) or silencing SERPINE1 (shRNA#3 or sh_SERPINE1#3) in the upper chamber, and THP1 cells treated with PMA in the lower chamber. (H) Immunofluorescence staining of xenograft tumor tissues. Comparison of the proportion of M1 or M2 macrophage infiltra tion. Green indicates F4/80. Red indicates iNOS or Arg1 expression

Article Snippet: THP-1 cells were differentiated into macrophages using 150 ng/mL phorbol 12-myristate 13-acetate (PMA, Sigma) for 24 h and subsequently co-cultured with cancer-derived exosomes or GC cells in 6-well plates with 0.4-μm membranes for 72 h. Harvested macrophages were converted into single-cell suspensions, stained with Elab Fluor 488 anti-human CD68 (Mouse, 1:20, ElabScience) and APC anti-human CD206 (Mouse, 1:20, ElabScience) antibodies, and analyzed for CD68+CD206+ populations by flow cytometry (Accuri C6, BD).

Techniques: Expressing, Flow Cytometry, Co-Culture Assay, shRNA, Immunofluorescence, Staining, Comparison

Journal: Journal of experimental & clinical cancer research : CR

Article Title: Gastric cancer-derived exosomal let-7 g-5p mediated by SERPINE1 promotes macrophage M2 polarization and gastric cancer progression.

doi: 10.1186/s13046-024-03269-4

Figure Lengend Snippet: Fig. 5 SERPINE1-mediated gastric cancer-derived exosomes facilitate the polarization of THP1 cells into M2 macrophages. (A) Schematic representation of the extraction and identification of exosomes and the induction of macrophage polarization. Transmission electron microscopy (B), nanoparticle tracking analysis (C), and western blotting (D) were used to identify the morphology, particle size, and markers of exosomes. (E) Confocal laser scanning microscopy detected Dil-labeled exosomes (red) internalized by DAPI-labeled macrophages (blue). (F–G) Immunofluorescence analysis of the proportion of CD206+ cells in THP1 cells treated with exosomes. (H–I) Flow cytometry analysis of the proportion of CD68+CD206+ cells in THP1 cells treated with exosomes. (J–K) qRT-PCR analysis of M1 markers (iNOS and TNF-α) and M2 markers (TGF-β, IL-10, and Arg-1) in THP1 cells treated with exosomes. (L–N) Transwell migration and invasion assays of GC cells (upper chamber) co-cultured with macrophages (lower chamber) ingesting exosomes

Article Snippet: THP-1 cells were differentiated into macrophages using 150 ng/mL phorbol 12-myristate 13-acetate (PMA, Sigma) for 24 h and subsequently co-cultured with cancer-derived exosomes or GC cells in 6-well plates with 0.4-μm membranes for 72 h. Harvested macrophages were converted into single-cell suspensions, stained with Elab Fluor 488 anti-human CD68 (Mouse, 1:20, ElabScience) and APC anti-human CD206 (Mouse, 1:20, ElabScience) antibodies, and analyzed for CD68+CD206+ populations by flow cytometry (Accuri C6, BD).

Techniques: Derivative Assay, Extraction, Transmission Assay, Electron Microscopy, Western Blot, Confocal Laser Scanning Microscopy, Labeling, Immunofluorescence, Flow Cytometry, Quantitative RT-PCR, Migration, Cell Culture