Mammalian Ste20-like kinase 1 inhibition as a cellular mediator of anoikis in mouse bone marrow mesenchymal stem cells

Figure 1 Mouse bone marrow mesenchymal stem cells exhibit susceptibility to anoikis under detachment conditions. A: Terminal-deoxynucleoitidyl Transferase Mediated Nick End Labeling (TUNEL)-positive cells (apoptotic cells) of mouse bone marrow mesenchymal stem cells (mBMSCs) cultured in precoated Poly-2-hydroxyethyl methacrylate conditions for 0 h, 12 h, 24 h, 36 h and 48 h using flow cytometry; B: Fluorescence staining images of dead cells; C: Reactive oxygen species (ROS) levels were measured using the ROS probe 2,7-Dichlorodihydrofluorescein diacetate by flow cytometry at 0 h, 12 h, 24 h, 36 h and 48 h; D: Quantitative analysis of the rate of TUNEL-positive cells; E: Quantification of live cells using a thiazolyl blue tetrazolium bromide assay; F: Quantitative analysis of the intracellular ROS level; G: Cell adhesion was expressed as fold changes between 0 h groups; H and I: Mammalian Ste20-like kinase 1 (Mst1), microtubules associated protein 1 light chain 3 II/I, p62, Beclin1, phospho-focal adhesion kinase, and activated caspase 3 expression levels at 0 h, 12 h, 24 h, 36 h and 48 h were evaluated by Western blot analysis; The expression of protein was expressed as the fold change relative to glyceraldehyde-3-phosphate dehydrogenase. Values are the mean ± SD of three independent experiments in each case. TUNEL: Terminal-deoxynucleoitidyl Transferase Mediated Nick End Labeling; ROS: Reactive oxygen species; Mst1: Mammalian Ste20-like kinase 1; LC3-II/I: Microtubules associated protein 1 light chain 3 II/I; p-FAK: Phospho-focal adhesion kinase; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase. ^cP < 0.01; ^bP < 0.05; ^aP > 0.05.