Journal: Frontiers in Cell and Developmental Biology

Article Title: Single-cell transcriptome dynamics of the autotaxin-lysophosphatidic acid axis during muscle regeneration reveal proliferative effects in mesenchymal fibro-adipogenic progenitors

doi: 10.3389/fcell.2023.1017660

Figure Lengend Snippet: PF-8380 pharmacological inhibition of ENPP2 impairs satellite cell proliferation and myotube differentiation. (A) Brightfield images of cultured muscle stem cells (i.e., satellite cells). (B) Outline of EdU assay in muscle satellite cells. (C) Flow cytometry detection of EdU labelled cells in combination with DNA fluorescence at 24 h of treatments. (D) Flow cytometry detection of mitotic (phospho-H3 S10 ) labelled cells in combination with DNA fluorescence at 24 h of treatments. (E) Outline of satellite cell differentiation protocol. GM, growth media; DM, differentiation media. (F) Representative laser confocal images of day 3 myotubes after the different treatments. α-Actinin staining is shown in hot yellow, nuclear staining in hot blue, and MF20 in magenta. Scale bar: 100 μm.

Article Snippet: MuSCs were platted at 75,000 cells per cm 2 when EdU (at 10 µM final concentration) or pH3 S10 labelling (Alexa Fluor ® 594 anti-Histone H3 Phospho (Ser10) Antibody, 1:250 dilution, clone 11D8, Cat. No. 650810, Biolegend) was performed as indicated in .

Techniques: Inhibition, Cell Culture, EdU Assay, Flow Cytometry, Fluorescence, Cell Differentiation, Staining

Journal: bioRxiv

Article Title: A novel zebrafish intestinal tumor model reveals a role for cyp7a1 -dependent tumor-liver crosstalk in tumor's adverse effects on host

doi: 10.1101/199349

Figure Lengend Snippet: (A)-(B) Representative images of tumor-bearing fish ( Tg(pInt-Gal4) +/Tg ; Tg(5×UAS:EGFP-P2A-kras G12D ) +/Tg ) and the sibling control ( Tg(pInt-Gal4) +/Tg ; Tg(UAS:EGFP) +/Tg ) at 5 dpf. Bright field (A) and EGFP (B) images are shown. Scale bar indicates 500 μm. (C) qPCR for EGFP-P2A-kras G12D expression in the sibling controls and tumor-bearing fish.The scores are normalized to expression of rpl13a . The data harbors three biological replicates. Error bars represent ± s.e.m. (D)-(I) Representative images of DAPI staining in intestine sections of tumor-bearing fish and the sibling controls at 5 dpf. DAPI (D, E) and EGFP (F, G) images are shown. In the merged images (H, I), DAPI and EGFP signals are shown in blue and green, respectively. Scale bar indicates 100 μm. (J) The number of EGFP and DAPI positive intestinal cells. The number of nuclei was manually counted from single section per individual fish. The data harbors 7 and 11 biological replicates from tumor-bearing fish and the sibling controls, respectively. Error bars represent ± s.e.m. Statistical significance was tested using student’s t -test (unpaired, one-tailed).(K)-(R) Representative images of fluorescent immunohistochemistry for phosphorylated histone H3 (pH3) in intestine sections of tumor-bearing fish and the sibling controls at 5 dpf. pH3 (K, L), DAPI (M, N) and EGFP (O, P) images are shown. White arrows indicate intestinal cells positive for pH3, EGFP, and DAPI. In the merged images (Q, R), pH3, DAPI and EGFP signals are shown in red, blue and green, respectively. Scale bar indicates 100 μm. (S) The number of intestinal cells positive for pH3, EGFP, and DAPI. The number of pH3, EGFP, and DAPI positive cells was counted from single section per individual fish. The data harbors 8 and 6 biological replicates from tumor-bearing fish and the sibling controls, respectively. Error bars represent ± s.e.m.

Article Snippet: Sections were rehydrated by PBS at room temperature for 30 min, and then permeabilized and blocked with 5% normal goat serum in PBS supplemented with 0.5% TritonX-100 (0.5% PBT) for 1 h. Sections were then incubated with the following primary antibodies diluted in 5% normal goat serum in 0.5% PBT at 4°C for overnight: rabbit anti-phosphorylated-Histone H3 (Ser10) (pH3) (EMD Millipore, 06-570; 1:100 dilution) and rabbit anti-E-cadherin (cdh1) (Gene Tex, GTX125890; 1:100 dilution).

Techniques: Control, Expressing, Staining, One-tailed Test, Immunohistochemistry