Na+/K+-ATPase α1 Identified as an Abundant Protein in the Blood-Labyrinth Barrier That Plays an Essential Role in the Barrier Integrity

Abstract

BackgroundThe endothelial-blood/tissue barrier is critical for maintaining tissue homeostasis. The ear harbors a unique endothelial-blood/tissue barrier which we term “blood-labyrinth-barrier”. This barrier is critical for maintaining inner ear homeostasis. Disruption of the blood-labyrinth-barrier is closely associated with a number of hearing disorders. Many proteins of the blood-brain-barrier and blood-retinal-barrier have been identified, leading to significant advances in understanding their tissue specific functions. In contrast, capillaries in the ear are small in volume and anatomically complex. This presents a challenge for protein analysis studies, which has resulted in limited knowledge of the molecular and functional components of the blood-labyrinth-barrier. In this study, we developed a novel method for isolation of the stria vascularis capillary from CBA/CaJ mouse cochlea and provided the first database of protein components in the blood-labyrinth barrier as well as evidence that the interaction of Na+/K+-ATPase α1 (ATP1A1) with protein kinase C eta (PKCη) and occludin is one of the mechanisms of loud sound-induced vascular permeability increase.Methodology/Principal FindingsUsing a mass-spectrometry, shotgun-proteomics approach combined with a novel “sandwich-dissociation” method, more than 600 proteins from isolated stria vascularis capillaries were identified from adult CBA/CaJ mouse cochlea. The ion transporter ATP1A1 was the most abundant protein in the blood-labyrinth barrier. Pharmacological inhibition of ATP1A1 activity resulted in hyperphosphorylation of tight junction proteins such as occludin which increased the blood-labyrinth-barrier permeability. PKCη directly interacted with ATP1A1 and was an essential mediator of ATP1A1-initiated occludin phosphorylation. Moreover, this identified signaling pathway was involved in the breakdown of the blood-labyrinth-barrier resulting from loud sound trauma.Conclusions/SignificanceThe results presented here provide a novel method for capillary isolation from the inner ear and the first database on protein components in the blood-labyrinth-barrier. Additionally, we found that ATP1A1 interaction with PKCη and occludin was involved in the integrity of the blood-labyrinth-barrier.

Perivascular-resident macrophage-like melanocytes in the inner ear are essential for the integrity of the intrastrial fluid–blood barrier

“…The major tight junction-associated proteins in the strial capillaries are occludins, claudins, ZOs, and adherens-junction proteins (23,24). The present study found that tight-junction and adhesion proteins were down-regulated substantially in the absence of PVM/Ms.…”

“…In an earlier mass spectrometry study, strial capillaries were found rich in tight-junction and cell-adhesion proteins (24). The major tight junction-associated proteins in the strial capillaries are occludins, claudins, ZOs, and adherens-junction proteins (23,24).…”

“…The major tight junctionassociated proteins in the intrastrial fluid-blood barrier are occludin, claudins, zonula occludens, and adherens-junction proteins (22). Several tight-and adherens-junction proteins, including ZO-1, occludin, and vascular endothelial cadherin (vecadherin), have been found in the intrastrial fluid-blood barrier (23,24). In the in vitro models, mRNA levels for zo-1, occludin, and ve-cadherin, assessed with quantitative RT-PCR (qRT-PCR), were decreased dramatically in the absence of PVM/Ms (n = 3; P < 0.05) (Fig.…”

Evaluation of Sustained-Release Steroid Hydrogels in a Guinea Pig Model for Noise-Induced Hearing Loss

“…The systemic application of glucocorticoids causes side effects such as hyperglycemia and osteoporosis [Cope and Bova, 2008;Trune and Canlon, 2012]. Additionally, the efficacy of systemic glucocorticoid treatment of conditions affecting the inner ear is limited by the low blood flow to the cochlea and the blood-perilymph barrier, which result in low perilymph concentrations of the active compound [Nakashima et al, 2003;Yang et al, 2011]. To minimize side effects and achieve higher perilymph drug concentrations, administration of glucocorticoids directly to the round window membrane via intratympanic injection has been preclinically evaluated and translated into clinical practice [Bird et al, 2011;Chou et DOI: 10.1159/000487662 al., 2013; Han et al, 2015;Shea et al, 2012].…”

Recent Research Trend for Ménièreʼs Disease

“…The BLB within the perilymph-filled areas that are part of the blood-perilymph barrier include the cochlear spiral ligament, spiral limbus, modiolus, osseous spiral lamina, vestibular stromal vasculature and the sub-epithelial dark cell area 13 . The vestibular BLB unit in the rodent is similar to the rodent strial BLB unit at the light microscopic level: composed of vascular endothelial cells, pericytes and perivascular-resident macrophage-like melanocytes Recent studies have implicated loss of the integrity of the BLB in several inner ear pathologies including acoustic trauma, autoimmune inner ear disease, and presbycusis 3,7,8,[14][15][16] . Meniere's disease is a disabling syndrome of fluctuating hearing loss, episodic vertigo, and ear fullness, the etiology and pathophysiology of which remain poorly understood.…”