Journal: Scientific Reports

Article Title: The role of anthrax toxin protein receptor 1 as a new mechanosensor molecule and its mechanotransduction in BMSCs under hydrostatic pressure

doi: 10.1038/s41598-019-49100-5

Figure Lengend Snippet: Effects of different pressure stimulating conditions on the expression of ANTXR1 and the chondrogenic potential of BMSC sheets. ( a ) RT-PCR was performed to determine the mRNA expression levels of ANTXR1, integrinβ1, and 3 chondrogenic markers in BMSC sheets treated with 8 different hydrostatic pressure conditions for 1 h. ( b ) BMSC sheets were stimulated with 8 different hydrostatic pressure conditions for 1 h. Extracts from the cytoplasmic fractions were analyzed by western blotting analysis to determine the protein expression levels of ANTXR1, integrinβ1, and 3 chondrogenic markers, Sox-9, aggrecan, and Col-II. Expression levels relative to that for GAPDH were derived using Quantity One density analysis. BMSCs treated with no pressure was used as a control. ( c ) Quantitative analysis of the western blotting bands by ImageJ software. Four independent assays were performed for each group. Data presented as the mean ± SD. # P < 0.05 represents a significant increase compared with the control group. * P < 0.05 represents a significant difference compared with the indicated groups. △ P < 0.05 represents a significant decrease compared with the control group.

Article Snippet: The primer sequences for ANTXR1 and integrinβ1 (Sango Biotech, Shanghai, China) are listed in the Supplementary Materials, Table .

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot, Derivative Assay, Control, Software

Journal: Scientific Reports

Article Title: The role of anthrax toxin protein receptor 1 as a new mechanosensor molecule and its mechanotransduction in BMSCs under hydrostatic pressure

doi: 10.1038/s41598-019-49100-5

Figure Lengend Snippet: Establishment of ANTXR1-downregulated or integrinβ1-downregulated BMSC sheets. ( a ) Fluorescent microscopy images of BMSC sheets transfected with ANTXR1-shRNA at MOI = 40. Scale bar = 500 µm. ( b ) RT-PCR was performed to determine the mRNA expression levels of ANTXR1. ( c , d ) Western blotting was performed to determine the protein expression levels of ANTXR1, followed by the quantitative analysis of the western blotting bands by ImageJ software. ( e ) Fluorescent microscopy images of BMSC sheets transfected with integrinβ1-shRNA at MOI = 40. Scale bar = 500 µm. ( f ) RT-PCR was performed to determine the mRNA expression levels of integrinβ1. ( g – h ) Western blotting was performed to determine protein expression levels of integrinβ1, followed by the quantitative analysis of the western blotting bands by ImageJ software.

Article Snippet: The primer sequences for ANTXR1 and integrinβ1 (Sango Biotech, Shanghai, China) are listed in the Supplementary Materials, Table .

Techniques: Microscopy, Transfection, shRNA, Reverse Transcription Polymerase Chain Reaction, Expressing, Western Blot, Software

Journal: Scientific Reports

Article Title: The role of anthrax toxin protein receptor 1 as a new mechanosensor molecule and its mechanotransduction in BMSCs under hydrostatic pressure

doi: 10.1038/s41598-019-49100-5

Figure Lengend Snippet: Knockdown of ANTXR1 or integrinβ1 partly inhibits mechiancally promoted chondrogenic differentiation of BMSs. ( a ) RT-PCR was performed to determine the mRNA expression levels of Sox-9, aggrecan and Col-II. ( b ) Western blotting was performed to determine the protein expression levels of Sox-9, aggrecan and Col-II. ( c ) Quantitative analysis of the western blotting bands by ImageJ software. Four independent assays were performed for each group. Data presented as the mean ± SD. # P < 0.05 represents a significant increase compared with the control group. * P < 0.05 represents a significant difference compared with the indicated groups. △ P < 0.05 represents a significant decrease compared with the control group.

Article Snippet: The primer sequences for ANTXR1 and integrinβ1 (Sango Biotech, Shanghai, China) are listed in the Supplementary Materials, Table .

Techniques: Knockdown, Reverse Transcription Polymerase Chain Reaction, Expressing, Western Blot, Software, Control

Journal: Scientific Reports

Article Title: The role of anthrax toxin protein receptor 1 as a new mechanosensor molecule and its mechanotransduction in BMSCs under hydrostatic pressure

doi: 10.1038/s41598-019-49100-5

Figure Lengend Snippet: Association between ANTXR1 and integrinβ1. ( a ) RT-PCR was performed to determine the mRNA expression levels of ANTXR1 and integrinβ1. ( b ) Western blotting was performed to determine the protein expression levels of ANTXR1 and integrinβ1. ( c ) Quantitative analysis of the western blotting bands by ImageJ software. ( d ) Interactions between ANTXR1 and integrinβ1 were demonstrated by Co-IP assays. Western blots of the inputs and immunoprecipitates were analyzed using the indicated antibodies. Four independent assays were performed for each group. Data presented as the mean ± SD. # P < 0.05 represents a significant increase compared with the control group. * P < 0.05 represents a significant difference compared with the indicated groups. △ P < 0.05 represents a significant decrease compared with the control group.

Article Snippet: The primer sequences for ANTXR1 and integrinβ1 (Sango Biotech, Shanghai, China) are listed in the Supplementary Materials, Table .

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Western Blot, Software, Co-Immunoprecipitation Assay, Control

Journal: Scientific Reports

Article Title: The role of anthrax toxin protein receptor 1 as a new mechanosensor molecule and its mechanotransduction in BMSCs under hydrostatic pressure

doi: 10.1038/s41598-019-49100-5

Figure Lengend Snippet: ANTXR1 interacted with integrinβ1 and the cytoskeleton. ( a ) Co-localization between ANTXR1 and integrinβ1 was detected by immunofluorescence in BMSCs from the 0 kPa control group and from the group subjected to 120 kPa of static pressure for 1 h. ANTXR1 was stained red, integrinβ1 was stained green, and nuclei were stained blue. Scale bar = 100 µm. ( b ) Co-localization between ANTXR1 and the F-actin was detected by immunofluorescence in BMSCs from the 0 kPa control group and from the group subjected to 120 kPa of static pressure for 1 h. ANTXR1 was stained red, F-actin was stained green by phalloidin and nuclei were stained blue. Scale bar = 100 µm. All experiments were repeated at least three times.

Article Snippet: The primer sequences for ANTXR1 and integrinβ1 (Sango Biotech, Shanghai, China) are listed in the Supplementary Materials, Table .

Techniques: Immunofluorescence, Control, Staining

Journal: Nature Communications

Article Title: Angiopoietin-2–integrin α5β1 signaling enhances vascular fatty acid transport and prevents ectopic lipid-induced insulin resistance

doi: 10.1038/s41467-020-16795-4

Figure Lengend Snippet: a Diagram depicting EC-specific RiboTag transcriptome analysis. Inducible recombination of the RiboTag allele led to expression of hemaglutanin (HA)-tagged ribosomal protein (reddish orange) specifically in ECs. Ribosome-bound transcripts were immunoprecipitated (IP) from homogenized whole tissues with anti-HA antibody-coupled magnetic beads. Extracted mRNAs were analyzed by RNA-Seq analysis. b Generation of RiboTag ∆EC mouse, inducible and specific tagging of ribosomes in ECs in 6-week-old mice, and their analyses 2 weeks later. Red arrowheads indicate daily injections of tamoxifen. c Representative images of HA-tagged ribosomal protein in VE-Cadherin expressing ECs in adipose tissues of RiboTag ∆EC mouse. Scale bars, 50 μm. d Comparisons of perilipin and pecam expression in isolated mRNA of ECs from different organs of RiboTag ∆EC mouse. e RNA-seq expression heatmap of ITGα5, ITGβ1, and Tie2 in isolated ECs from different organs using RiboTag ∆EC mouse. n = 5 for each group. f Representative images of ITGα5β1 expression in various adipose tissues of WT mice. Scale bars, 50 μm. g Representative images of Tie2 expression in SAT and BAT of Tie2-GFP mice. Magnified views of dotted-line box are shown in lower panels. Scale bars, 50 μm.

Article Snippet: Specific pathogen-free C57BL/6J, Adiponectin- Cre , Integrinβ1 flox/flox , Tie2 -GFP, and RiboTag mice were purchased from Jackson Laboratory (Jackson Labs, Bar Harbor, ME).

Techniques: Expressing, Immunoprecipitation, Magnetic Beads, RNA Sequencing, Isolation