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Flow-induced HDAC1 phosphorylation and nuclear export in angiogenic sprouting

View Article: PubMed Central - PubMed

ABSTRACT

Angiogenesis requires the coordinated growth and migration of endothelial cells (ECs), with each EC residing in the vessel wall integrating local signals to determine whether to remain quiescent or undergo morphogenesis. These signals include vascular endothelial growth factor (VEGF) and flow-induced mechanical stimuli such as interstitial flow, which are both elevated in the tumor microenvironment. However, it is not clear how VEGF signaling and mechanobiological activation due to interstitial flow cooperate during angiogenesis. Here, we show that endothelial morphogenesis is histone deacetylase-1- (HDAC1) dependent and that interstitial flow increases the phosphorylation of HDAC1, its activity, and its export from the nucleus. Furthermore, we show that HDAC1 inhibition decreases endothelial morphogenesis and matrix metalloproteinase-14 (MMP14) expression. Our results suggest that HDAC1 modulates angiogenesis in response to flow, providing a new target for modulating vascularization in the clinic.

No MeSH data available.


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Inhibition of p-HDAC1 nuclear export inhibits angiogenic invasion.(a) Angiogenic invasion into the 3-D collagen matrix under static conditions (no VEGF gradient) (top row) is not inhibited by a 24 h LMB (20 nM) treatment. Under flow and a VEGF gradient (bottom row), LMB treatment results in reduced sprouting and invasion. Dashed arrow indicates the direction of flow. Control (DMSO) devices (both static and flow) proceeded with extensive sprouting. (Scale bar, 100 μm). (b) Quantification of the % of invasion area under static and flow in response to 24 h of continuous leptomycin B (LBM) treatment. Data represent mean ± SEM; *p < 0.05.
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f4: Inhibition of p-HDAC1 nuclear export inhibits angiogenic invasion.(a) Angiogenic invasion into the 3-D collagen matrix under static conditions (no VEGF gradient) (top row) is not inhibited by a 24 h LMB (20 nM) treatment. Under flow and a VEGF gradient (bottom row), LMB treatment results in reduced sprouting and invasion. Dashed arrow indicates the direction of flow. Control (DMSO) devices (both static and flow) proceeded with extensive sprouting. (Scale bar, 100 μm). (b) Quantification of the % of invasion area under static and flow in response to 24 h of continuous leptomycin B (LBM) treatment. Data represent mean ± SEM; *p < 0.05.

Mentions: Interestingly, our results showed that interstitial flow resulted in increased localization of t- and p-HDAC1 to the cytoplasm (Fig. 3e), as quantified by the area fraction of cytoplasmic t- and p-HDAC1 (Fig. 3f). Inhibition of CRM1 (exportin 1)-dependent transport with leptomycin B (LMB) (20 nM) prevented the increase in cytoplasmic localization (Fig. 3e,f), suggesting that nuclear export of HDAC1 plays a role in the cytoplasmic p-HDAC1 localization. To examine whether nuclear export of HDAC1 is required for angiogenic sprouting, ECs in the microfluidic device were continuously treated with LMB (20 nM) for 24 h in the presence of interstitial flow or under static conditions. Exogenous VEGF was also included in the media. Under flow conditions, inhibition of protein nuclear export significantly reduced invasion into the collagen matrix (Fig. 4a,b), while under static conditions it had no significant effect (Fig. 4a,b). Together these results suggest that CRM1-mediated nuclear export of HDAC1 is part of the mechanism that modulates interstitial flow-induced angiogenic sprouting.


Flow-induced HDAC1 phosphorylation and nuclear export in angiogenic sprouting
Inhibition of p-HDAC1 nuclear export inhibits angiogenic invasion.(a) Angiogenic invasion into the 3-D collagen matrix under static conditions (no VEGF gradient) (top row) is not inhibited by a 24 h LMB (20 nM) treatment. Under flow and a VEGF gradient (bottom row), LMB treatment results in reduced sprouting and invasion. Dashed arrow indicates the direction of flow. Control (DMSO) devices (both static and flow) proceeded with extensive sprouting. (Scale bar, 100 μm). (b) Quantification of the % of invasion area under static and flow in response to 24 h of continuous leptomycin B (LBM) treatment. Data represent mean ± SEM; *p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5037418&req=5

f4: Inhibition of p-HDAC1 nuclear export inhibits angiogenic invasion.(a) Angiogenic invasion into the 3-D collagen matrix under static conditions (no VEGF gradient) (top row) is not inhibited by a 24 h LMB (20 nM) treatment. Under flow and a VEGF gradient (bottom row), LMB treatment results in reduced sprouting and invasion. Dashed arrow indicates the direction of flow. Control (DMSO) devices (both static and flow) proceeded with extensive sprouting. (Scale bar, 100 μm). (b) Quantification of the % of invasion area under static and flow in response to 24 h of continuous leptomycin B (LBM) treatment. Data represent mean ± SEM; *p < 0.05.
Mentions: Interestingly, our results showed that interstitial flow resulted in increased localization of t- and p-HDAC1 to the cytoplasm (Fig. 3e), as quantified by the area fraction of cytoplasmic t- and p-HDAC1 (Fig. 3f). Inhibition of CRM1 (exportin 1)-dependent transport with leptomycin B (LMB) (20 nM) prevented the increase in cytoplasmic localization (Fig. 3e,f), suggesting that nuclear export of HDAC1 plays a role in the cytoplasmic p-HDAC1 localization. To examine whether nuclear export of HDAC1 is required for angiogenic sprouting, ECs in the microfluidic device were continuously treated with LMB (20 nM) for 24 h in the presence of interstitial flow or under static conditions. Exogenous VEGF was also included in the media. Under flow conditions, inhibition of protein nuclear export significantly reduced invasion into the collagen matrix (Fig. 4a,b), while under static conditions it had no significant effect (Fig. 4a,b). Together these results suggest that CRM1-mediated nuclear export of HDAC1 is part of the mechanism that modulates interstitial flow-induced angiogenic sprouting.

View Article: PubMed Central - PubMed

ABSTRACT

Angiogenesis requires the coordinated growth and migration of endothelial cells (ECs), with each EC residing in the vessel wall integrating local signals to determine whether to remain quiescent or undergo morphogenesis. These signals include vascular endothelial growth factor (VEGF) and flow-induced mechanical stimuli such as interstitial flow, which are both elevated in the tumor microenvironment. However, it is not clear how VEGF signaling and mechanobiological activation due to interstitial flow cooperate during angiogenesis. Here, we show that endothelial morphogenesis is histone deacetylase-1- (HDAC1) dependent and that interstitial flow increases the phosphorylation of HDAC1, its activity, and its export from the nucleus. Furthermore, we show that HDAC1 inhibition decreases endothelial morphogenesis and matrix metalloproteinase-14 (MMP14) expression. Our results suggest that HDAC1 modulates angiogenesis in response to flow, providing a new target for modulating vascularization in the clinic.

No MeSH data available.


Related in: MedlinePlus