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TRPV4 Activation Contributes Functional Recovery from Ischemic Stroke via Angiogenesis and Neurogenesis

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Abstract

The endothelial transient receptor potential cation channel subfamily V member 4 (TRPV4) plays a crucial role in vascular remodeling; however, TRPV4-mediated angiogenesis after ischemic neuronal death as a neurorestorative strategy has not yet been thoroughly examined. In this study, we first tested whether TRPV4 activation can improve functional recovery in rats subjected to transient brain ischemia. The possible mechanisms for TRPV4 activation-promoted functional recovery were explored. A TRPV4 agonist, 4α-phorbol 12,13-didecanoate (4α-PDD), was intravenously injected via the tail vein at 6 h and 1, 2, 3, 4 days after ischemic stroke. The treatment reduced infarct volume by almost 50% (14.7 ± 3.7 vs. 29.2 ± 6.2%; p < 0.0001) and improved functional outcomes (p = 0.03) on day 5. To explore the therapeutic mechanism, we measured endothelial nitric oxide synthase (eNOS) expression and phosphorylation, vascular endothelial growth factor A (VEGFA) signaling, and neural stem/progenitor cells (NPCs). TRPV4 activation significantly increased eNOS expression and phosphorylation (serine 1177) by more than 2-fold in the ischemic region. The expressions of VEGFA and VEGF receptor-2 were significantly higher in the treated animals, especially an increase of the proangiogenic VEGFA164a isoform while a decrease of the antiangiogenic VEGFA165b isoform. We evaluated angiogenesis by detecting microvessel density in ischemic region. Using the immunohistochemistry staining, we found that 4α-PDD treatment caused a 3.4-fold increase of microvessel density (p < 0.0001). In addition, NPC proliferation and migration in the ischemic hemisphere were increased by 3-fold and 5-fold, respectively. In conclusion, our data suggest that TRPV4 activation by 4α-PDD may improve poststroke functional improvement through angiogenesis and neurogenesis.

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Abbreviations

4α-PDD:

4α-Phorbol 12,13-didecanoate

BBB:

Blood–brain barrier

EPCs:

Endothelial progenitor cells

ECs:

Endothelial cells

FSS:

Fluid shear stress

GS:

Garcia score

eNOS:

Endothelial nitric oxide synthase

NPCs:

Neural stem/progenitor cells

NO:

Nitric oxide

SVZ:

Subventricular zone

tMCAO:

Transient middle carotid artery occlusion

TRPV4:

Transient receptor potential vanilloid 4

VEGFA:

Vascular endothelial growth factor A

VEGFR2:

VEGF receptor-2

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Acknowledgements

We thank JY. Wang (Division of Colorectal Surgery, Department of Surgery, Kaohsiung Medical University) for technical support. This work was supported by the following funding: Ministry of Science and Technology (Taiwan, R.O.C.; MOST 103-2314-B-075-076-MY3, MOST 104-2745-B-037-001, MOST 105-2314-B-039-050, MOST 103-2314-B-037-027-MY2) and National Health Research Institutes (Taiwan, R.O.C.; NHRI-EX106-10605PI).

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Authors and Affiliations

  1. Department of Physical Medicine and Rehabilitation, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan

    Chun-Kai Chen

  2. Graduate Institute of Medicine, Collage of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan

    Chun-Kai Chen

  3. Department of Medical Research, China Medical University Hospital, Taichung, Taiwan

    Po-Yuan Hsu, Tzu-Ming Wang & Suh-Hang H. Juo

  4. Division of Colorectal Surgery, Department of Surgery, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan

    Zhi-Feng Miao

  5. Department of Neurology, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan

    Ruey-Tay Lin

  6. Graduate Institute of Biomedical Sciences, China Medical University, Taichung, Taiwan

    Suh-Hang H. Juo

  7. Institute of New Drug Development, China Medical University, Taichung, Taiwan

    Suh-Hang H. Juo

  8. Brain Disease Research Center, China Medical University, Taichung, Taiwan

    Suh-Hang H. Juo

  9. Center for Myopia and Eye Disease, China Medical University, Taichung, Taiwan

    Suh-Hang H. Juo

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  1. Chun-Kai Chen
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  2. Po-Yuan Hsu
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Corresponding author

Correspondence to Suh-Hang H. Juo.

Ethics declarations

All experimental procedures were approved by the Institutional Animal Ethical Committee Kaohsiung Medical University and were conducted according to the Guide for the Care and Use of Laboratory Animal of the National Institute of Health.

Conflict of Interest

The authors declare that they have no conflict of interest.

Electronic Supplementary Material

Supplementary Fig. 1

Schematic illustration of the experimental design. Transient middle cerebral artery occlusion (tMCAO) was inducted by 2-h occlusion in right internal carotid artery. 4a-phorbol-12,13-didecanoate (4α-PDD) was used as an agonist for endothelial transient receptor potential vanilloid 4.* represents the test points for neurological deficits score (NDS). ** represents the test points for infarct volume. # represents the test points for Garcia score (GS). (GIF 15 kb)

High resolution image (TIFF 76 kb)

Supplementary Fig. 2

Delayed deaths are not observed with 4α-PDD treatment. Mortality was assessed during 5 d after tMCAO in 4α-PDD group (n = 23), and control group (treated with DMSO, n = 26). Mortality at day 5 was not significantly different. (GIF 34 kb)

High resolution image (TIFF 206 kb)

Supplementary Table S1

(DOCX 13 kb)

Supplementary Table S2

(DOCX 14 kb)

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Chen, CK., Hsu, PY., Wang, TM. et al. TRPV4 Activation Contributes Functional Recovery from Ischemic Stroke via Angiogenesis and Neurogenesis. Mol Neurobiol 55, 4127–4135 (2018). https://doi.org/10.1007/s12035-017-0625-0

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  • DOI: https://doi.org/10.1007/s12035-017-0625-0

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