Department of Otorhinolaryngology-Head and Neck Surgery, School of Medicine, University of Occupational and Environmental Health, Kitakyushu, Japan.
Faculty of Public Health, Vinh Medical University, Vinh, Vietnam.
Int Arch Allergy Immunol. 2022;183(6):579-590. doi: 10.1159/000521765. Epub 2022 Jan 31.
The mucociliary transport function of the airway epithelium is largely dependent on ciliary beating. The control signal of ciliary beating is thought to be intracellular Ca2+. We herein investigated the expression of T-type voltage-gated calcium channel (VGCC), a generator of intracellular Ca2+ oscillation, in the human nasal mucosa.
The inferior turbinate was collected from patients with chronic hypertrophic rhinitis. The expression of T-type VGCC α1 subunits was examined by immunohistochemistry, transmission immunoelectron microscopy, Western blot, and real-time reverse transcription-polymerase chain reaction (RT-PCR). Participation of T-type VGCC in the ciliary beat regulation was examined by pharmacological inhibition tests using specific blockers of T-type VGCC in ex vivo measurements of the ciliary beat frequency (CBF) and ATP release and in intracellular Ca2+ imaging of isolated ciliated cells.
Immunohistochemical staining showed the expressions of T-type VGCC α1 subunits, Cav3.1 and Cav3.3, on the surface of the epithelial cells. At the ultrastructural level, immunoreactivity for Cav3.1 was localized on the surface of the cilia, and that for Cav3.3 was localized in the cilia and at the base of the cilia. The existence of Cav3.1 and Cav3.3 was confirmed at the protein level by Western blot and at the transcriptional level by real-time RT-PCR. Specific blockers of T-type VGCC, mibefradil and NNC 55-0396, significantly inhibited CBF. These blockers also inhibited a CBF increase induced by 8-bromo-cAMP/8-bromo-cGMP and significantly lowered the intracellular Ca2+ level of isolated ciliated cells in a time-dependent manner. On the other hand, the ATP release from the nasal mucosa was not changed by mibefradil or NNC 55-0396.
These results indicate that T-type VGCC α1 subunits, Cav3.1 and Cav3.3, exist at the cilia of the nasal epithelial cells and participate in the regulation of ciliary beating and that these channels act downstream of cAMP/cGMP.
气道上皮的黏液纤毛转运功能在很大程度上依赖于纤毛的摆动。纤毛摆动的控制信号被认为是细胞内 Ca2+。本文研究了 T 型电压门控钙通道（VGCC），即细胞内 Ca2+ 震荡的发生器，在人鼻黏膜中的表达。
从慢性肥厚性鼻炎患者的下鼻甲中采集标本。通过免疫组织化学、透射电镜免疫电镜、Western blot 和实时逆转录聚合酶链反应（RT-PCR）检测 T 型 VGCC α1 亚单位的表达。通过使用 T 型 VGCC 特异性阻断剂在体外测量纤毛摆动频率（CBF）和 ATP 释放以及分离的纤毛细胞内 Ca2+ 成像来检测 T 型 VGCC 在纤毛摆动调节中的作用。
免疫组织化学染色显示 T 型 VGCC α1 亚单位 Cav3.1 和 Cav3.3 在上皮细胞表面表达。在超微结构水平上，Cav3.1 的免疫反应性定位于纤毛表面，而 Cav3.3 的免疫反应性定位于纤毛和纤毛基部。Western blot 和实时 RT-PCR 证实了 Cav3.1 和 Cav3.3 的存在。T 型 VGCC 的特异性阻断剂 mibefradil 和 NNC 55-0396 显著抑制 CBF。这些阻断剂还抑制了 8-溴-cAMP/8-溴-cGMP 诱导的 CBF 增加，并以时间依赖的方式显著降低分离的纤毛细胞内的 Ca2+水平。另一方面，mibefradil 或 NNC 55-0396 并未改变鼻黏膜中的 ATP 释放。
这些结果表明，T 型 VGCC α1 亚单位 Cav3.1 和 Cav3.3 存在于鼻上皮细胞的纤毛中，并参与纤毛摆动的调节，并且这些通道作用于 cAMP/cGMP 的下游。
