Journal: Communications Biology

Article Title: Fortilin deficiency induces anti-atherosclerotic phenotypes in macrophages and protects hypercholesterolemic mice against atherosclerosis

doi: 10.1038/s42003-025-08425-w

Figure Lengend Snippet: A.U. arbitrary unit, RT-qPCR reverse transcription quantitative polymerase chain reaction, WT THP1 WT-fortilin cells, KO THP1 KO-fortilin cells, oxLDL oxidized low density lipoprotein, αSMA alpha smooth muscle cell actin, CD68 Cluster of Differentiation 68, TCE, 2,2,2-trichloroethanol total protein staining, GAPDH glyceraldehyde-3-phosphate dehydrogenase, phMΦ primary human macrophages, CTL control, phMΦ transduced by control short hairpin RNA (shRNA control ) lentiviral particles, KD knockdown, phMΦ transduced by shRNA fortilin lentiviral particles. a Expression levels of fortilin in WT and KO cells with and without oxLDL stimulation. b – f Status of expression of VSMC markers (αSMA, SM22α, CNN1) ( b – d ) and MФ markers (Mac2, CD68) ( e , f ) in WT and KO cells as analyzed by RT-qPCR (n = 3, 3). g Western blot analysis of VSMC and MΦ markers in WT and KO cells at their baselines and upon oxLDL stimulation. h Successful KD of fortilin using shRNA fortilin lentivirus particles in phMΦ. Status of expression of VSMC markers ( i – k ) and MΦ markers ( l , m ) in CTL and KD phMΦ cells as analyzed by RT-qPCR (n = 3, 3). Data are expressed as mean ± s.d., P values determined by one-way ANOVA with Tukey multiple comparisons ( a – f , h – m ) are shown. The findings in ( g ) were confirmed in three independent experiments.

Article Snippet: The following primary antibodies were used: Rabbit anti-fortilin (MBL International, Woburn, MA, USA; Catalog #: PM017; 1:1,000 dilution; used for the experiments depicted in Fig. ); Rabbit anti-fortilin (Abcam, Waltham, MA, USA; Clone EPR5540, ab133568; 1:2,000 dilution; Figs. , ); Mouse anti-GAPDH (Fitzgerald, Acton, MA, USA; Clone: 10R-G109a; 1:10,000 dilution; Fig. ); mouse anti-GAPDH (Santa Cruz Biotechnology, Inc. Dallas, TX, USA; Clone: 6C5; Catalog #: sc-32233; 1:1,000 dilution; Figs. , ); Rabbit anti-human calponin-1 (CNN1) mAb (Cell Signaling Technology (CST), Danvers, MA, USA; Clone D8L2T, Catalog #: 17819; 1:1,000 dilution; Fig. ); Recombinant rabbit anti-human α-smooth muscle actin (SMA) antibody (Abcam, clone EPR5368, Catalog #: ab124964; 1:1,000 dilution; Fig. ); Rabbit anti-human SM22α (Transgelin/TAGLN) polyclonal Ab (CST, Catalog #: 40471; 1:1,000 dilution; Fig. ); Rabbit anti-human TGF-β1 polyclonal Ab (Abcam, Catalog #: ab92486; 1:1,000 dilution; Figs. , ); Rabbit anti-Mac2 (Galectin-3/LGALS3) mAb (CST, Clone: D4I2R, Catalog #: 8785; 1:1,000 dilution; Fig. ); Rabbit anti-human CD68 mAb MultiMabTM(CST, multiple clones, Catalog #: 86985; 1:1,000 dilution; Fig. ).

Techniques: Quantitative RT-PCR, Reverse Transcription, Real-time Polymerase Chain Reaction, Staining, Control, shRNA, Knockdown, Expressing, Western Blot

Journal: Communications Biology

Article Title: Fortilin deficiency induces anti-atherosclerotic phenotypes in macrophages and protects hypercholesterolemic mice against atherosclerosis

doi: 10.1038/s42003-025-08425-w

Figure Lengend Snippet: A.U. arbitrary unit, RT-qPCR reverse transcription quantitative polymerase chain reaction, WT THP1 WT-fortilin cells, KO THP1 KO-fortilin cells, oxLDL oxidized low density lipoprotein, αSMA alpha smooth muscle cell actin, CD68 Cluster of Differentiation 68, TCE, 2,2,2-trichloroethanol total protein staining, GAPDH glyceraldehyde-3-phosphate dehydrogenase, phMΦ primary human macrophages, CTL control, phMΦ transduced by control short hairpin RNA (shRNA control ) lentiviral particles, KD knockdown, phMΦ transduced by shRNA fortilin lentiviral particles. a Expression levels of fortilin in WT and KO cells with and without oxLDL stimulation. b – f Status of expression of VSMC markers (αSMA, SM22α, CNN1) ( b – d ) and MФ markers (Mac2, CD68) ( e , f ) in WT and KO cells as analyzed by RT-qPCR (n = 3, 3). g Western blot analysis of VSMC and MΦ markers in WT and KO cells at their baselines and upon oxLDL stimulation. h Successful KD of fortilin using shRNA fortilin lentivirus particles in phMΦ. Status of expression of VSMC markers ( i – k ) and MΦ markers ( l , m ) in CTL and KD phMΦ cells as analyzed by RT-qPCR (n = 3, 3). Data are expressed as mean ± s.d., P values determined by one-way ANOVA with Tukey multiple comparisons ( a – f , h – m ) are shown. The findings in ( g ) were confirmed in three independent experiments.

Article Snippet: The following primary antibodies were used: Rabbit anti-fortilin (MBL International, Woburn, MA, USA; Catalog #: PM017; 1:1,000 dilution; used for the experiments depicted in Fig. ); Rabbit anti-fortilin (Abcam, Waltham, MA, USA; Clone EPR5540, ab133568; 1:2,000 dilution; Figs. , ); Mouse anti-GAPDH (Fitzgerald, Acton, MA, USA; Clone: 10R-G109a; 1:10,000 dilution; Fig. ); mouse anti-GAPDH (Santa Cruz Biotechnology, Inc. Dallas, TX, USA; Clone: 6C5; Catalog #: sc-32233; 1:1,000 dilution; Figs. , ); Rabbit anti-human calponin-1 (CNN1) mAb (Cell Signaling Technology (CST), Danvers, MA, USA; Clone D8L2T, Catalog #: 17819; 1:1,000 dilution; Fig. ); Recombinant rabbit anti-human α-smooth muscle actin (SMA) antibody (Abcam, clone EPR5368, Catalog #: ab124964; 1:1,000 dilution; Fig. ); Rabbit anti-human SM22α (Transgelin/TAGLN) polyclonal Ab (CST, Catalog #: 40471; 1:1,000 dilution; Fig. ); Rabbit anti-human TGF-β1 polyclonal Ab (Abcam, Catalog #: ab92486; 1:1,000 dilution; Figs. , ); Rabbit anti-Mac2 (Galectin-3/LGALS3) mAb (CST, Clone: D4I2R, Catalog #: 8785; 1:1,000 dilution; Fig. ); Rabbit anti-human CD68 mAb MultiMabTM(CST, multiple clones, Catalog #: 86985; 1:1,000 dilution; Fig. ).

Techniques: Quantitative RT-PCR, Reverse Transcription, Real-time Polymerase Chain Reaction, Staining, Control, shRNA, Knockdown, Expressing, Western Blot

Journal: Communications Biology

Article Title: Fortilin deficiency induces anti-atherosclerotic phenotypes in macrophages and protects hypercholesterolemic mice against atherosclerosis

doi: 10.1038/s42003-025-08425-w

Figure Lengend Snippet: CM conditioned media, WT THP1 WT-fortilin cells, KO THP1 KO-fortilin cells, oxLDL oxidized low density lipoprotein, UCDMSCs human umbilical cord derived mesenchymal stem cells, M media, F.C. fold change, α-TGF-β1-MB anti-TGF-β1-antibody-coated magnetic beads, ID-CM CM immunodepleted of TGF-β1, TGF-β1 ID media with TGF-β1 immunodepleted of TGF-β1. Time-course of RT-qPCR of RNA from UCDMSCs treated by either M only, M + TGF-β1, CM of THP1 KO-fortilin , or CM of THP1 WT-fortilin ( a ) for VSMC markers (αSMA ( b ); SM22α ( c ); and CNN1 ( d )), a differentiation marker (CD45; e ); and a marker of undifferentiated cells (CD105, f ) (n = 3 for each group). g ELISA of CM from THP1 WT-fortilin cells and THP1 KO-fortilin cells with and without oxLDL stimulation (n = 4 for each group). RT-qPCR of RNA from UCDMSCs treated by either M only with and without TGF-β1 ID, M + TGF-β1 with and without TGF-β1 ID, CM of THP1 KO-fortilin with and without TGF-β1 ID, or CM of THP1 WT-fortilin with and without TGF-β1 ID ( h ) for VSMC markers αSMA ( i ), SM22α ( j ), and CNN1 ( k ); a differentiation marker (CD45, l ); and a marker of undifferentiated cells (CD105, m ) (n = 3 for each group). Data are expressed as mean ± s.d. P values determined by one way ANOVA ( b – g , i – m ) are shown.

Article Snippet: The following primary antibodies were used: Rabbit anti-fortilin (MBL International, Woburn, MA, USA; Catalog #: PM017; 1:1,000 dilution; used for the experiments depicted in Fig. ); Rabbit anti-fortilin (Abcam, Waltham, MA, USA; Clone EPR5540, ab133568; 1:2,000 dilution; Figs. , ); Mouse anti-GAPDH (Fitzgerald, Acton, MA, USA; Clone: 10R-G109a; 1:10,000 dilution; Fig. ); mouse anti-GAPDH (Santa Cruz Biotechnology, Inc. Dallas, TX, USA; Clone: 6C5; Catalog #: sc-32233; 1:1,000 dilution; Figs. , ); Rabbit anti-human calponin-1 (CNN1) mAb (Cell Signaling Technology (CST), Danvers, MA, USA; Clone D8L2T, Catalog #: 17819; 1:1,000 dilution; Fig. ); Recombinant rabbit anti-human α-smooth muscle actin (SMA) antibody (Abcam, clone EPR5368, Catalog #: ab124964; 1:1,000 dilution; Fig. ); Rabbit anti-human SM22α (Transgelin/TAGLN) polyclonal Ab (CST, Catalog #: 40471; 1:1,000 dilution; Fig. ); Rabbit anti-human TGF-β1 polyclonal Ab (Abcam, Catalog #: ab92486; 1:1,000 dilution; Figs. , ); Rabbit anti-Mac2 (Galectin-3/LGALS3) mAb (CST, Clone: D4I2R, Catalog #: 8785; 1:1,000 dilution; Fig. ); Rabbit anti-human CD68 mAb MultiMabTM(CST, multiple clones, Catalog #: 86985; 1:1,000 dilution; Fig. ).

Techniques: Derivative Assay, Magnetic Beads, Quantitative RT-PCR, Marker, Enzyme-linked Immunosorbent Assay