In vivo transcriptomic profiling using cell encapsulation identifies effector pathways of systemic aging
- Nestlé Institute of Health Sciences, Nestlé Research, Switzerland
- School of Life Sciences École Polytechnique Fédérale de Lausanne (EPFL), Switzerland
- Vascular Pathophysiology Area, Centro Nacional de Investigaciones Cardiovasculares, Spain
- Département de Pharmacologie-Physiologie, Institut de Pharmacologie de Sherbrooke, Centre de Recherche du Centre Hospitalier Universitaire de Sherbrooke, Faculté de Médecine et des Sciences de la Santé, Université de Sherbrooke, Canada
- Department of Experimental and Health Sciences, Pompeu Fabra University, CIBERNED and ICREA, Barcelona, Spain, Spain
Figures
Figure 1 with 1 supplement
Polyethersulfone (PES) hollow fiber capsules for transcriptomic profiling of long-range systemic signals.
(a) Scheme outlining interactions in the tissue niche in the context of the systemic circulation. Using cell encapsulation in PES hollow fiber membranes (PES-HFM), direct effects of long-range …
Figure 1—figure supplement 1
Description of the capsule loading and sealing platform and cellular characterization.
(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 …
Figure 2 with 1 supplement
In vitro characterization of encapsulated myogenic progenitors.
(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 …
Figure 2—figure supplement 1
Comparison of encapsulated cells to 2D culture.
(a-d) Representative phalloidin and MitoTracker stainings of hskMPs and mskMPs in cross sections of capsules (3D) and in 2D culture. Scale bars: 50 µm and 25 µm for inserts. (e) Quantification of …
Figure 3 with 2 supplements
In vivo characterization of encapsulated myogenic progenitors.
(a) Schematic of the capsule implantation strategy in mice. In case of hskMPs, an osmotic pump supplying immunosuppressant was implanted at the contralateral flank. (b) Photograph of the capsules in …
Figure 3—figure supplement 1
Enzymatically liberated encapsulated cells and staining for infiltration.
(a) Representative bright field images of hskMPs and mskMPs in 2D culture after enzymatic liberation from capsules. Scale bars: 40 µm. (b) Representative CD31 immunostainings of cross sections of …
Figure 3—figure supplement 2
Inflammatory profiling after capsule implantation.
(a) Multiplexed enzyme-linked immunosorbent assay (ELISA) array signal heatmaps for levels of inflammatory factors in serum of mice that underwent surgical implantation of polyethersulfone (PES) …
Figure 4 with 5 supplements
Transcriptomic profiling of systemic aging.
(a) Gene set enrichment analysis (GSEA) of age-induced genes in encapsulated hskMPs or mskMPs compared to the young control after 10 days in vivo. Shaded bars represent gene sets overlapping between …
Figure 4—figure supplement 1
RNA yield from capsules and validation of Myc and E2F target genes.
(a, b) RNA yield from capsules containing hskMPs or mskMPs after 10 days in vivo in young and aged mice. For each replicate two capsules from one mouse were pooled. Graphs represent means ± s.e.m. …
Figure 4—figure supplement 2
Inflammatory profiling in young and aged mice.
(a) Multiplexed enzyme-linked immunosorbent assay (ELISA) array heatmaps for levels of inflammatory factors in plasma of young and aged mice. n ≥ 5 mice. Asterisks indicate factors changed with a …
Figure 4—figure supplement 3
Proliferation, senescence, and differentiation of encapsulated cells after implantation in young and aged mice.
(a, b) Representative Ki-67 immunostainings and quantification of cross sections of capsules containing hskMPs after 10 days of exposure to the systemic environment in young and aged mice. (c, d) …
Figure 4—figure supplement 4
Transcriptomic profiling of cells exposed to young and aged serum in 2D culture.
(a) Gene set enrichment analysis (GSEA) of genes induced in hskMPs in 2D culture in the presence of aged human serum for 4 or 10 days compared to the young condition. Gene sets that were also …
Figure 4—figure supplement 5
Proliferation, senescence, and differentiation of cells after exposure to young and aged serum in 2D culture.
(a, b) Representative Ki-67 immunostainings and quantification of 2D cultured hskMPs exposed to young and aged human serum for 4 days. (c, d) Representative β-gal immunostainings and quantification …
Additional files
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Supplementary file 1
Tables.
(a) Multiplexed enzyme-linked immunosorbent assay (ELISA) quantification of inflammatory factors in serum of mice that underwent surgical implantation of polyethersulfone (PES) hollow fiber capsules compared to untreated controls 10 days after the procedure. n ≥ 7 mice. Each value represents averages in pg/ml from n = 4 technical replicates for each factor. TNF-Rl and MIP-1g were excluded from the analysis since signals were outside of the detection range. (b) Gene sets increased with age in encapsulated human skeletal muscle progenitors (hskMPs) compared to the young control after 10 days in vivo. Data are derived from capsules of n ≥ 5 mice in each age group. GST p-value (Pval) = Wilcoxon gene set test p-value. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure. GST FDR = Wilcoxon gene set test false discovery rate. (c) Gene sets increased with age in encapsulated mouse skeletal muscle progenitors (mskMPs) compared to the young control after 10 days in vivo. Data are derived from capsules of n ≥ 5 mice in each age group. GST Pval = Wilcoxon gene set test p-value. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure. (d) Significantly enriched Hallmark Myc V1 target genes when comparing encapsulated hskMPs in aged mice to the young control after 10 days in vivo. Data are derived from capsules of n ≥ 5 mice in each age group. Pval < 1%. (e) Significantly enriched Hallmark E2F target genes when comparing encapsulated hskMPs in aged mice to the young control after 10 days in vivo. Data are derived from capsules of n ≥ 5 mice in each age group. Pval < 1%. (f) Multiplexed ELISA array quantification of levels of inflammatory factors in plasma of mice young and aged mice. n ≥ 5 mice. Each value represents averages in pg/ml from n = 4 technical replicates for each factor. TNF-Rl and MIP-1g were excluded from the analysis since signals were outside of the detection range. (g) Gene sets decreased with age in encapsulated hskMPs compared to the young control after 10 days in vivo. Data are derived from capsules of n ≥ 5 mice in each age group. GST Pval = Wilcoxon gene set test p-value. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure. (h) Gene sets decreased with age in encapsulated mskMPs compared to the young control after 10 days in vivo. Data are derived from capsules of n ≥ 5 mice in each age group. GST Pval = Wilcoxon gene set test p-value. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure. (i) Gene sets increased with age in niche-resident freshly isolated mouse muscle stem cells compared to the young control. Data from n = 8 young or aged mice. GST Pval = Wilcoxon gene set test p-value. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure. (j) Gene sets decreased with age in niche-resident freshly isolated mouse muscle stem cells compared to the young control. Data from n = 3 young or aged mice. GST Pval = Wilcoxon gene set test p-value. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure. (k) Gene sets increased after 4 days in 2D hskMP culture exposed to aged human serum compared to the young human serum. GST Pval = Wilcoxon gene set test p-value. Data are derived from hskMPs exposed to human serum from n = 3 different young or aged donors. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure. (l) Gene sets increased after 10 days in 2D hskMP culture exposed to aged human serum compared to the young human serum. Data are derived from hskMPs exposed to human serum from n = 3 different young or aged donors. GST Pval = Wilcoxon gene set test p-value. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure. (m) Gene sets decreased after 4 days in 2D hskMP culture exposed to aged human serum compared to the young human serum. Data are derived from hskMPs exposed to human serum from n = 3 different young or aged donors. GST Pval = Wilcoxon gene set test p-value. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure. (n) Gene sets decreased after 10 days in 2D hskMP culture exposed to aged human serum compared to the young human serum. Data are derived from hskMPs exposed to human serum from n = 3 different young or aged donors. GST Pval = Wilcoxon gene set test p-value. GST FDR = Adjusted p-value using the Benjamini-Hochberg procedure.
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