Department of Comprehensive Care, Tufts University School of Dental Medicine, Boston, Massachusetts, United States.
Schepens Eye Research Institute/Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, United States.
Invest Ophthalmol Vis Sci. 2021 Nov 1;62(14):25. doi: 10.1167/iovs.62.14.25.
We reported that oxytocin (OXT), added to freshly prepared lacrimal gland lobules, induced myoepithelial cell (MEC) contraction. In other systems, OXT activates phospholipase C (PLC) generating Inositol 1,4,5-trisphosphate (IP3) which increases intracellular calcium concentration ([Ca2+]i) causing contraction. The aim of the current study was to investigate the role of this pathway in OXT-induced contraction of MEC.
Tear volume was measured using the cotton thread method. Lacrimal gland MEC were isolated and propagated from α-smooth muscle actin (SMA)-green fluorescent protein (GFP) mice, in which MEC express GFP making them easily identifiable. RNA and protein samples were prepared for RT-PCR and Western blotting for G protein expression. Changes in [Ca2+]i were measured in Fura-2 loaded MEC using a ratio imaging system. MEC contraction was monitored in real time and changes in cell size were quantified using ImageJ software.
OXT applied either topically to surgically exposed lacrimal glands or delivered subcutaneously resulted in increased tear volume. OXT stimulated lacrimal gland MEC contraction in a dose-dependent manner, with a maximum response at 10-7 M. MEC express the PLC coupling G proteins, Gαq and Gα11, and their activation by OXT resulted in a concentration-dependent increase in [Ca2+]i with a maximum response at 10-6 M. Furthermore, the activation of the IP3 receptor to increase [Ca2+]i is crucial for OXT-induced MEC contraction since blocking the IP3 receptor with 2-APB completely abrogated this response.
We conclude that OXT uses the PLC/Ca2+ pathway to stimulate MEC contraction and increase lacrimal gland secretion.
我们曾报道过，在新制备的泪腺小叶中添加催产素（OXT）可诱导肌上皮细胞（MEC）收缩。在其他系统中，OXT 通过激活磷脂酶 C（PLC）产生三磷酸肌醇（IP3），从而增加细胞内钙离子浓度（[Ca2+]i），导致收缩。本研究旨在探讨该途径在 OXT 诱导的 MEC 收缩中的作用。
采用棉线法测量泪液量。从α-平滑肌肌动蛋白（SMA）-绿色荧光蛋白（GFP）小鼠中分离和培养泪腺 MEC，其中 MEC 表达 GFP，使其易于识别。通过 RT-PCR 和 Western 印迹法制备 RNA 和蛋白质样品，以检测 G 蛋白表达。使用比率成像系统测量 Fura-2 负载的 MEC 中的 [Ca2+]i 变化。实时监测 MEC 收缩，并使用 ImageJ 软件定量细胞大小变化。
OXT 局部应用于手术暴露的泪腺或皮下给药均可增加泪液量。OXT 以剂量依赖性方式刺激泪腺 MEC 收缩，最大反应在 10-7 M。MEC 表达 PLC 偶联 G 蛋白，Gαq 和 Gα11，OXT 激活它们可导致 [Ca2+]i 浓度依赖性增加，最大反应在 10-6 M。此外，激活 IP3 受体以增加 [Ca2+]i 对于 OXT 诱导的 MEC 收缩至关重要，因为用 2-APB 阻断 IP3 受体可完全消除这种反应。
我们得出结论，OXT 使用 PLC/Ca2+途径刺激 MEC 收缩并增加泪腺分泌。
