Sekulic Marijana, Puche Raoul, Bodmer Daniel, Petkovic Vesna
Department of Biomedicine, University Hospital Basel, University of Basel, Basel, Switzerland.
University Hospital Basel, Clinic for Otorhinolaryngology, Basel, Switzerland.
Front Mol Neurosci. 2023 Sep 21;16:1243370. doi: 10.3389/fnmol.2023.1243370. eCollection 2023.
Hearing loss is one of the 10 leading causes of disability worldwide. No drug therapies are currently available to protect or restore hearing. Inner ear auditory hair cells and the blood-labyrinth barrier (BLB) are critical for normal hearing, and the BLB between the systemic circulation and stria vascularis is crucial for maintaining cochlear and vestibular homeostasis. BLB defects are associated with inner ear diseases that lead to hearing loss, including vascular malformations, inflammation, and Meniere's disease (MD). Antibodies against proteins in the inner ear and cytokines in the cochlea, including IL-1α, TNF-α, and NF-kβ, are detected in the blood of more than half of MD patients. There is also emerging evidence of inner ear inflammation in some diseases, including MD, progressive sensorineural hearing loss, otosclerosis, and sudden deafness. Here, we examined the effects of TNF-α, IL6, and LPS on human stria vascularis-derived primary endothelial cells cultured together with pericytes in a Transwell system. By measuring trans-endothelial electrical resistance, we found that TNF-α causes the most significant disruption of the endothelial barrier. IL6 had a moderate influence on the barrier, whereas LPS had a minimal impact on barrier integrity. The prominent effect of TNF-α on the barrier was confirmed in the expression of the major junctional genes responsible for forming the tight endothelial monolayer, the decreased expression of and . We further tested permeability using 2 μg of daptomycin (1,619 Da), which does not pass the BLB under normal conditions, by measuring its passage through the barrier by HPLC. Treatment with TNF-α resulted in higher permeability in treated samples compared to controls. LPS-treated cells behaved similarly to the untreated cells and did not show differences in permeability compared to control. The endothelial damage caused by TNF-α was confirmed by decreased expression of an essential endothelial proteoglycan, syndecan1. These results allowed us to create an inflammatory environment model that increased BLB permeability in culture and mimicked an inflammatory state within the stria vascularis.
听力损失是全球十大致残原因之一。目前尚无药物疗法可用于保护或恢复听力。内耳听觉毛细胞和血迷路屏障（BLB）对正常听力至关重要，而全身循环与血管纹之间的BLB对于维持耳蜗和前庭内环境稳定至关重要。BLB缺陷与导致听力损失的内耳疾病有关，包括血管畸形、炎症和梅尼埃病（MD）。超过一半的MD患者血液中可检测到针对内耳蛋白质和耳蜗细胞因子（包括IL-1α、TNF-α和NF-kβ）的抗体。在包括MD、进行性感音神经性听力损失、耳硬化症和突发性耳聋在内的一些疾病中，也有越来越多的证据表明存在内耳炎症。在此，我们在Transwell系统中研究了TNF-α、IL6和脂多糖（LPS）对与周细胞共培养的人血管纹来源的原代内皮细胞的影响。通过测量跨内皮电阻，我们发现TNF-α对内皮屏障的破坏最为显著。IL6对屏障有中度影响，而LPS对屏障完整性的影响最小。TNF-α对屏障的显著作用在负责形成紧密内皮单层的主要连接基因的表达中得到证实，即 和 的表达降低。我们进一步使用2μg达托霉素（1619Da）测试通透性，在正常情况下该药物不会通过BLB，通过高效液相色谱法测量其通过屏障的情况。与对照组相比，TNF-α处理导致处理后的样品通透性更高。LPS处理的细胞与未处理的细胞表现相似，与对照组相比通透性没有差异。TNF-α导致的内皮损伤通过必需的内皮蛋白聚糖syndecan1的表达降低得到证实。这些结果使我们能够创建一个炎症环境模型，该模型增加了培养物中BLB的通透性，并模拟了血管纹内的炎症状态。
