Suppression of ischemia in arterial occlusive disease by JNK-promoted native collateral artery development
- University of Massachusetts Medical School, United States
- Howard Hughes Medical Institute, United States
Figures
Figure 1 with 8 supplements
Enhanced blood perfusion blockade and severe ischemic injury in endothelial JNK-deficient mice upon arterial occlusion.
(A) Control and JNK-deficient primary endothelial cells were examined by immunoblot analysis by probing with antibodies to JNK and GAPDH. (B) Simplified diagram of the medial aspect of the mouse …
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Figure 1—source data 1
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- https://doi.org/10.7554/eLife.18414.004
Figure 1—figure supplement 1
Characterization of endothelial JNK-deficient mice and lung endothelial cells.
(A) The body mass of endothelial JNK-deficient mice and control mice was examined at P0 and P6 (mean ± SEM; n = 12~23). No statistically significant differences in body mass between groups were …
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Figure 1—figure supplement 1—source data 1
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- https://doi.org/10.7554/eLife.18414.006
Figure 1—figure supplement 2
Endothelial JNK-deficient mice have no major perturbations in the hematopoietic system.
(A) Bone marrow, splenocytes, and blood cells isolated from endothelial JNK-deficient mice and control mice were examined by immunoblot analysis by probing with antibodies to JNK and GAPDH. The data …
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Figure 1—figure supplement 2—source data 1
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- https://doi.org/10.7554/eLife.18414.008
Figure 1—figure supplement 3
Normal hypoxia responses and VEGF signaling in JNK-deficient endothelial cells.
(A) Primary MLEC incubated overnight in media with 1% FBS were placed under hypoxia (1% O2). The cells were examined by immunoblot analysis with antibodies to pJNK, JNK, pSer63-cJun, and αTubulin. …
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Figure 1—figure supplement 3—source data 1
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- https://doi.org/10.7554/eLife.18414.010
Figure 1—figure supplement 4
Endothelial JNK is not required for proliferation, migration, and angiogenic responses in vitro.
(A) JNK-deficient and control primary MLEC form similar tubular networks in matrigel. Images are representative of two experiments performed in triplicate with independent primary MLEC preparations. …
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Figure 1—figure supplement 4—source data 1
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- https://doi.org/10.7554/eLife.18414.012
Figure 1—figure supplement 5
Endothelial JNK is not required for in vivo pathologic angiogenesis.
(A,B) Representative confocal images of laser-induced choroidal neovascular (CNV) tufts in control and JNK-deficient mice at day 7 post-treatment stained with iB4, Phalloidin, and DAPI (A). …
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Figure 1—figure supplement 5—source data 1
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- https://doi.org/10.7554/eLife.18414.014
Figure 1—figure supplement 6
Compound JNK-deficiency in endothelial cells causes defects in the response to arterial occlusion.
(A,B) Simplified diagram of the medial aspect of the mouse hindlimb skeletal muscle vasculature indicating the location of the femoral artery ligation site (proximal to the PCFA) (A). Quantitation …
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Figure 1—figure supplement 6—source data 1
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- https://doi.org/10.7554/eLife.18414.016
Figure 1—figure supplement 7
JNK deficient mice show no perturbations in overall cardiovascular function.
(A,B) Analysis of blood pressure and heart rate in WT mice, Mapk8-/- mice, and Mapk9-/- mice (A) and endothelial JNK-deficient mice and control mice (B) demonstrates no statistically significant …
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Figure 1—figure supplement 7—source data 1
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- https://doi.org/10.7554/eLife.18414.018
Figure 1—figure supplement 8
JNK-deficiency causes defects in artery size and connectivity, but not the hypoxia response after femoral artery ligation.
(A) Contrast (Bismuth/gelatin) perfused hindlimb vasculature was examined by µCT analysis at day 4 post-FAL. The images are representative of 7~8 mice analyzed per group. (B–E) Taqman gene …
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Figure 1—figure supplement 8—source data 1
Source data for Figure 1—figure supplement 8.
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- https://doi.org/10.7554/eLife.18414.020
Figure 2
Severe ischemic injury in MLK2/3-deficient mice after femoral artery ligation.
(A) The expression of members of the MLK protein kinase family (MLK1, MLK2, MLK3 & MLK4) in primary MLEC cultures was examined by measurement of Map3k9, Map3k10, Map3k11, and BC021891 mRNA by …
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Figure 2—source data 1
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- https://doi.org/10.7554/eLife.18414.022
Figure 3
MLK2/3-deficient mice exhibit defects in native collateral artery formation.
(A) Representative confocal images (n = 5 mice) of control and MLK2/3-deficient whole mount P6 adductor muscle vasculature stained with antibodies to endomucin (capillary and venous vasculature, …
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Figure 3—source data 1
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- https://doi.org/10.7554/eLife.18414.024
Figure 4
Endothelial JNK is not required for the arteriogenic response of gracilis collaterals in adult mice.
(A) Timeline of tamoxifen administration to induce disruption of Mapk8LoxP and Mapk9LoxP alleles in the vascular endothelium of adult mice prior to FAL and analysis of blood flow by laser doppler …
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Figure 4—source data 1
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- https://doi.org/10.7554/eLife.18414.026
Figure 5 with 1 supplement
Endothelial JNK-deficient mice display abnormal native collateral arteries.
(A,B) Representative confocal images (n = 7 mice) of whole mount adductor muscle vasculature reveals SMA-covered gracilis collateral arteries in P6 control mice, but not JNK-deficient mice (A). …
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Figure 5—source data 1
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- https://doi.org/10.7554/eLife.18414.028
Figure 5—figure supplement 1
Intimate association of gracilis collaterals and peripheral nerves in adductor muscles.
Confocal microscopy of a whole mount adductor muscle stained with antibodies to SMA (green) and Neurofilament-M (red) illustrates the close association of gracilis collateral arteries with …
Figure 6 with 3 supplements
Abnormal retinal vascular development associated with excessive sprouting of tip cells in endothelial JNK-deficient mice.
(A–C) Collages of confocal images of P6 whole mount retinas stained with isolectinB4 (iB4) show reduced vascular extension in JNK-deficient retinas (B,C) compared with littermate control retinas (A).…
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Figure 6—source data 1
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- https://doi.org/10.7554/eLife.18414.031
Figure 6—figure supplement 1
NG2+ pericyte coverage of the P6 retinal vasculature.
Confocal microscopy of whole mount retinal vasculature stained with an antibody to the pericyte marker NG2 (red) and isolectinB4 (green) demonstrated no obvious differences in vessel pericyte …
Figure 6—figure supplement 2
Abnormal retinal vascular development associated with excessive sprouting in endothelial JNK1/2-deficient mice.
(A–C) Collages of confocal images of P6 whole mount retinas stained with iB4 show reduced vascular extension in JNK1/2-deficient retinas (B,C) compared with littermate control retinas (A). The …
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Figure 6—figure supplement 2—source data 1
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- https://doi.org/10.7554/eLife.18414.034
Figure 6—figure supplement 3
Abnormal retinal vascular development associated with excessive sprouting in Map3k10-/- Map3k11-/- mice.
(A–C) Collages of confocal images of P6 whole mount retinas stained with isolectinB4 show reduced vascular extension in MLK2/3-deficient retinas (B,C) compared with littermate control retinas (A). …
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Figure 6—figure supplement 3—source data 1
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- https://doi.org/10.7554/eLife.18414.036
Figure 7 with 1 supplement
Reduced Dll4 / Notch signaling in the JNK-deficient vascular endothelium.
(A) Quantitative RT-PCR analysis of Notch pathway genes revealing reduced expression in JNK-deficient primary endothelial cells compared with control cells (mean ± SEM; n = 4). The data shown are …
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Figure 7—source data 1
Source data for Figure 7.
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- https://doi.org/10.7554/eLife.18414.038
Figure 7—figure supplement 1
RNA-Seq analysis of differentially expressed genes between control and JNK-deficient endothelial cells.
(A) Heatmap of the 781 differentially expressed genes (FPKM > 2; absolute log2 fold change > 0.5; n = 3; q < 0.05) between MLEC cultures isolated from endothelial JNK-deficient mice and control MLEC …
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Figure 7—figure supplement 1—source data 1
Source data for Figure 7—figure supplement 1.
This file contains raw source data used to make the graphs presented in Figure 7—figure supplement 1.
- https://doi.org/10.7554/eLife.18414.040
Figure 8
