Anti f4 80 70076

Palmitate (PA) (P0500) was obtained from Sigma-Aldrich (St. Louis, MO). LPC (Sigma, St. Louis, MO) was dissolved as previously described (8 (link)). Primary antisera employed for the studies include anti-alpha smooth muscle actin (αSMA) (ab124964) and anti-fibronectin (ab2413) antibodies from Abcam (Cambridge, MA), anti-GAPDH (MAB374) from Millipore Sigma, anti-β-actin (sc-47778) from Santa Cruz Biotechnologies (Santa Cruz, CA) and anti-Lyve1 (AF2125) from R&D systems (Minneapolis, MN), anti-F4/80 (70076) and anti-phospho Yap (Ser127) (4911) from Cell Signaling Technology (Danvers, MA), anti-Yap1 (sc-101199) from Santa Cruz Biotechnologies (Santa Cruz, CA), anti-type I collagen antibody (1310-01) from Southern Biotech. Recombinant human VCAM1 (rhVCAM1) (862-VC) was obtained from R&D systems.

Immunohistochemistry was performed on mouse testis tissue sections using standard protocols: after being dewaxed and rehydrated, the paraffin sections were repaired with sodium citrate repair solution (P0083; Beyotime, China). Next, sections were incubated with anti-LC3B (18725-1-AP, Proteintech, USA; 1:200) and anti-F4/80 (70076, Cell Signaling Technology; 1:200) primary antibodies overnight at 4°C. The slices were then treated with biotinylated secondary antibodies and incubated with peroxidase-linked streptavidin (Nakasugi Golden Bridge, China). Hematoxylin was used as a counterstaining agent; adding 3,3'-diaminobenzidine (DAB) allowed the staining to be visualized with a microscope (Nikon, Japan).

Dulbecco’s modified Eagle’s medium (DMEM) and fetal bovine serum (FBS) were obtained from Gibco (Grand Island, NY, USA). Matrigel was purchased from Corning, Inc. (Corning, NY, USA). Anti-mouse PD-1 antibody (clone RMP1-14) and isotype control IgG2a antibody were obtained from BioXCell (West Lebanon, NH, USA). Anti-CD3 (#ab16669) and anti-TGF-β1 (#ab215715) antibodies were purchased from Abcam (Cambridge, MA, USA). Anti-CD8 (#98941) and anti-F4/80 (#70076) antibodies were purchased from Cell Signaling (Beverly, MA, USA). Anti-granzyme B antibody (#14-8822-82) was purchased from Invitrogen (Carlsbad, CA, USA). PE/Cy7 anti-CD3 (#552774), BV605 Anti-CD8 (#563152), BV711 anti-CD4 (#563050), APC anti-CD335 (NKp46) (#137608), APC/Cy7 anti-CD45 (#103116), BV421 anti-CD11b (#562605), APC anti-GR-1 (#108412), PE/Cy7 anti-F4/80 (#123114), Alexa488 anti-MHC Class II (#107616), PE anti-CD206 (#141706), and TruStain FcX™ PLUS (#156604) antibodies were purchased from Biolegend (San Diego, CA, USA).