Our long-term objective is to study the vascular biology of the human blood-nerve barrier. Specifically, we are interested in studying the role of GDNF and its signaling pathways in restoring blood-nerve barrier function following injury. This project is designed to address a fundamental question: Is GDNF sufficient and essential to the recovery of the blood-nerve barrier following peripheral nerve injury and degeneration? Based on our exciting new preliminary data, we propose the following hypothesis: GDNF sufficiently restores human endoneurial endothelial cell tight junction function following injury primarily by cytoskeletal reorganization that results in more continuous intercellular contacts and fewer intercellular gaps. This occurs without significantly enhancing adherens and tight junction associated protein expression in vitro. GDNF sufficiently restores human endoneurial endothelial cell tight junction barrier function following injury via RET-Tyrosine kinase/ Ras-MAP kinase signaling pathways in vitro. Using a novel conditional GDNF knockout mouse model, we hypothesize that GDNF is essential for the early and persistent restoration of blood-nerve barrier function following sciatic nerve crush injury. GDNF sufficiently restored human blood-nerve barrier resistance following serum withdrawal in vitro in a dose-dependent manner. Our preliminary data specifically implicates the RET-Tyrosine kinase/ Ras-MAP kinase signaling pathways in vitro. Membrane localization of F-actin microfilaments and more continuous intercellular adherens and tight junctions occurred following GDNF treatment of the serum-deprived blood-nerve barrier. We did not observe significant changes in -catenin, VE cadherin, ZO-1, occludin and claudin-5 expression or alterations in claudin-5 tyrosine phosphorylation compared to the untreated injured blood-nerve barrier in vitro. In order to address these hypotheses, we will confirm the sufficient effect of GDNF in restoring blood-nerve barrier function using well-established endothelial barrier experimental assays. Similarly, we will determine that the RET-Tyrosine kinase/ Ras-MAP kinase signaling pathway is responsible for the GDNF-mediated effect following serum withdrawal in vitro, using specific cell-permeable inhibitors in the above experimental assays. Finally, we will evaluate effect of GDNF in restoring blood-nerve barrier function following peripheral nerve injury in vivo using a novel tamoxifen-inducible conditional GDNF knockout mouse strain. Very little is known about the vascular biology of the human blood-nerve barrier. Phenotypic and functional differences between vascular endothelial cells provide the rationale to specifically study the biology of the human blood-nerve barrier in health and disease. Alternations in blood-nerve barrier function have been implicated in many peripheral neuropathies and neuropathic pain. Restoration of blood-nerve barrier function by GDNF could accelerate recovery from these prevalent, disabling medical problems.

Public Health Relevance

The purpose of our work is to discover how a molecule called glial cell-derived neurotrophic factor (GDNF) may restore blood-nerve barrier function in peripheral nerves following injury. Peripheral neuropathies affect over 20 million people in the United States and millions more worldwide. By studying how GDNF helps restore blood-nerve barrier function; we will obtain important new knowledge that can lead to better treatments for peripheral neuropathies and neuropathic pain. This could significantly improve the lives of patients and their families; and reduce the financial burden on the health care system nationally and internationally.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
7R01NS075212-03
Application #
8811628
Study Section
Brain Injury and Neurovascular Pathologies Study Section (BINP)
Program Officer
Owens, David F
Project Start
2012-07-01
Project End
2017-03-31
Budget Start
2013-09-01
Budget End
2014-03-31
Support Year
Fiscal Year
2013
Total Cost
$660,661
Indirect Cost
$224,830
Name
University of Alabama Birmingham
Department
Neurology
Type
Schools of Medicine
DUNS #
063690705
City
Birmingham
State
AL
Country
United States
Zip Code
35294
Yuan, Furong; Yosef, Nejla; Lakshmana Reddy, Chetan et al. (2014) CCR2 gene deletion and pharmacologic blockade ameliorate a severe murine experimental autoimmune neuritis model of Guillain-Barré syndrome. PLoS One 9:e90463
Yosef, Nejla; Ubogu, Eroboghene E (2013) An immortalized human blood-nerve barrier endothelial cell line for in vitro permeability studies. Cell Mol Neurobiol 33:175-86
Chiang, Sharon; Ubogu, Eroboghene E (2013) The role of chemokines in Guillain-Barre syndrome. Muscle Nerve 48:320-30
Ubogu, Eroboghene E (2013) The molecular and biophysical characterization of the human blood-nerve barrier: current concepts. J Vasc Res 50:289-303
Reddy, Chetan Lakshmana; Yosef, Nejla; Ubogu, Eroboghene E (2013) VEGF-A165 potently induces human blood-nerve barrier endothelial cell proliferation, angiogenesis, and wound healing in vitro. Cell Mol Neurobiol 33:789-801
Ubogu, Eroboghene E (2013) Chemokine-dependent signaling pathways in the peripheral nervous system. Methods Mol Biol 1013:17-30