The First Clinical Medical College, Nanjing University of Chinese Medicine, Nanjing 210023, China.
Ophthalmology Department, the Affiliated Hospital of Gansu University of Chinese Medicine, Lanzhou 730000.
Zhen Ci Yan Jiu. 2023 Oct 25;48(10):1025-1032. doi: 10.13702/j.1000-0607.20230061.
To observe the effect of acupuncture on the ocular surface symptoms and the protein expression of vasoactive intestinal peptide (VIP) / cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) / aquaporin 5(AQP5) signaling pathway in lacrimal gland tissue of aqueous tear deficiency (ATD) type dry eye model, so as to investigate its mechanism underlying improvement of ATD.
British shorthair guinea pigs were randomly divided into blank control, model, acupuncture, sham-acupuncture and medication group, with 8 guinea pigs in each group. The ATD model was established by subcutaneous injection of scopolamine hydrobromide (0.6 mg/dose, 4 times/d for 10 days). For guinea pigs of the acupuncture group, filiform needles were inserted into bilateral "Jingming"(BL1), "Cuanzhu"(BL2), "Sizhukong"(TE23), "Taiyang"(EX-HN5), and "Tongziliao"(GB1) for 15 min. For guinea pigs of the sham-acupuncture group, a blunt filiform needle was used to repeatedly prick (not pierce) the skin of the same acupoints mentioned above. The treatment in the above two groups was conducted once daily for 14 days. The guinea pigs in the medication group received administration of sodium hyaluronate eye drops in both eyes, three times a day for 14 days. The objective tests of tear film break-up time (BUT), corneal fluorescein staining score (FLS) and phenol red thread (PRT) test were conducted before and after modeling and after the intervention. After the intervention, the lacrimal index (weight of lacrimal gland/body weight) was calculated. Histopathological changes of the lacrimal gland were observed after H.E. staining. The expression of AQP5 in the lacrimal gland were detected by immunofluorescence, and the contents of VIP and AQP5 in the lacrimal gland were measured by ELISA, the protein expression levels of VIP, cAMP, PKA, p-PKA and AQP5 in the lacrimal gland were detected by Western blot.
In comparison with the blank control group, the PRT, BUT, lacrimal index, AQP5 immunoactivity, contents of VIP and AQP5, and protein expression levels of VIP, cAMP, PKA, p-PKA and AQP5 were significantly decreased(<0.01, <0.05), and FLS was obviously increased (<0.01) in the model group . Compared to the model group, the PRT, BUT, lacrimal index, AQP5 immunoactivity, contents of VIP and AQP5, and expression levels of VIP and AQP5 in both acupuncture and medication groups, and the expression levels of cAMP, PKA, p-PKA in the acupuncture group were considerably increased (<0.01, <0.05), while the FLS was markedly decreased in both acupuncture and medication groups (<0.01, <0.05). Compared with the medication group, the acupuncture group had increased PRT (<0.05).
Acupuncture intervention is effective in reducing ocular surface damage and promoting tear secretion in guinea pigs with ATD, which may be related to its function in activating VIP/cAMP/PKA signaling, and promoting the expression of AQP5 in the lacrimal gland.
观察针刺对干眼症模型中泪液缺乏型（ATD）眼表症状和血管活性肠肽（VIP）/环磷酸腺苷（cAMP）/蛋白激酶 A（PKA）/水通道蛋白 5（AQP5）信号通路中蛋白表达的影响，探讨针刺改善 ATD 的作用机制。
将英国短毛豚鼠随机分为空白对照组、模型组、针刺组、假针刺组和药物组，每组 8 只。采用皮下注射氢溴酸东莨菪碱（0.6mg/剂量，每天 4 次，连续 10 天）建立 ATD 模型。针刺组豚鼠双侧“睛明”（BL1）、“攒竹”（BL2）、“四白”（TE23）、“太阳”（EX-HN5）和“瞳子髎”（GB1）进针 15 分钟。假针刺组豚鼠使用钝针反复针刺（不刺穿）上述相同穴位的皮肤。两组治疗均每日 1 次，共 14 天。药物组豚鼠双眼滴用玻璃酸钠滴眼液，每天 3 次，共 14 天。造模前后及干预后进行泪膜破裂时间（BUT）、角膜荧光素染色评分（FLS）和酚红棉线试验（PRT）的客观检查。干预后计算泪液指数（泪腺重量/体重）。HE 染色观察泪腺组织病理学变化。免疫荧光法检测泪腺中 AQP5 的表达，ELISA 法检测泪腺中 VIP 和 AQP5 的含量，Western blot 法检测泪腺中 VIP、cAMP、PKA、p-PKA 和 AQP5 的蛋白表达水平。
与空白对照组相比，模型组 PRT、BUT、泪液指数、AQP5 免疫活性、VIP 和 AQP5 含量以及 VIP、cAMP、PKA、p-PKA 和 AQP5 蛋白表达水平均显著降低（<0.01，<0.05），FLS 明显升高（<0.01）。与模型组相比，针刺组和药物组的 PRT、BUT、泪液指数、AQP5 免疫活性、VIP 和 AQP5 含量以及 VIP、AQP5 蛋白表达水平和针刺组 cAMP、PKA、p-PKA 蛋白表达水平均显著升高（<0.01，<0.05），而针刺组和药物组的 FLS 均显著降低（<0.01，<0.05）。与药物组相比，针刺组的 PRT 升高（<0.05）。
针刺干预可减轻 ATD 豚鼠眼表损伤，促进泪液分泌，其作用机制可能与激活 VIP/cAMP/PKA 信号通路，促进泪腺 AQP5 表达有关。
