Journal: eLife

Article Title: In vivo transcriptomic profiling using cell encapsulation identifies effector pathways of systemic aging

doi: 10.7554/eLife.57393

Figure Lengend Snippet: ( a ) Flow cytometry strategy used to identify live and apoptotic cells. CytoCalcein is sequestered in the cytoplasm of live cells, while apoptosis is measured by labeling of cell surface phosphatidylserine using Apopxin. ( b ) Quantification of live and apoptotic human (hskMPs) or mouse (mskMPs) skeletal muscle progenitors embedded into different biomaterials and maintained in growth media for 4 or 10 days. A = Growth factor reduced Matrigel, B = Hydrogel, C = MaxGel extracellular matrix. Values were obtained from pooled cells of n = 6 replicates. ( c ) Schematic outlining the construction of the adaptor hub used for mounting and loading of the PES hollow fiber capsules. ( d , e ) Schematics and graphical representations of the parts of the 3D printed capsule cutting and sealing platform. The photograph in the upper left of ( f ) shows the assembled platform loaded with a capsule mounted to the adaptor hub. ( g ) Quantification of Pax7 and MyoD positive hskMPs over a time course of 10 days in 2D culture. Graphs represent means ± s.e.m. n = 6 replicates for each time point. ****p < 0.0001, ***p < 0.001 by one-way ANOVA followed by Bonferroni post hoc test.

Article Snippet: For transcriptomic profiling of hskMPs in vitro, the cells were seeded into human fibronectin coated dishes (Corning, 356008) in human skeletal muscle myoblast growth medium (Zenbio, SKM-M).

Techniques: Flow Cytometry, Labeling

Journal: eLife

Article Title: In vivo transcriptomic profiling using cell encapsulation identifies effector pathways of systemic aging

doi: 10.7554/eLife.57393

Figure Lengend Snippet: ( a , b ) Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-based quantification of apoptosis in encapsulated human (hskMPs) or mouse (mskMPs) skeletal muscle progenitors maintained in growth media for 10 days. ( c , d ) DNA staining-based quantification of hskMP or mskMP numbers in cross sections of capsules maintained for 4, 8, or 10 days in growth media. ( e , f ) Representative DNA stainings of cross sections from a capsule containing hskMPs or mskMPs maintained 10 days in growth media. Scale bars: 75 µm. ( g ) Representative Pax7 immunostainings of cross sections from capsules containing hskMPs or mskMPs maintained 10 days in growth media. Scale bars: 150 µm. ( h , i ) Quantification of Pax7 positive cells in capsules containing hskMPs or mskMPs maintained for 4, 8, or 10 days in growth media. ( j ) Representative MyoD immunostainings of cross sections from capsules containing hskMPs or mskMPs maintained 10 days in growth media. Scale bars: 150 µm. ( k , l ) Quantification of MyoD positive cells in capsules containing hskMPs or mskMPs maintained for 4, 8, or 10 days in growth media. ( m–p ) Quantification of Pax7 and MyoD in cross sections from capsules maintained for 4 days under proliferative (Prolif.) conditions in growth media or in differentiation (Diff.) media. ( q ) Representative myosin heavy chain (MHC) immunostainings of cross sections from capsules containing hskMPs or mskMPs maintained for 4 days in differentiation media. Scale bars: 75 µm. ( r , s ) Quantification of MHC positive cells in capsules containing hskMPs or mskMPs maintained for 4 days in growth media compared to differentiation media. All graphs represent means ± s.e.m. n ≥ 3 cross sections from different capsules were quantified for each experiment and time point. ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05. Two-way comparisons were made with a Student’s t-test and multiple comparisons by one-way ANOVA followed by Bonferroni post hoc test.

Article Snippet: For transcriptomic profiling of hskMPs in vitro, the cells were seeded into human fibronectin coated dishes (Corning, 356008) in human skeletal muscle myoblast growth medium (Zenbio, SKM-M).

Techniques: TUNEL Assay, Staining