益气养阴通络方联合羟氯喹改善干燥综合征模型小鼠颌下腺分泌的实验研究

Experimental study of Yiqi Yangyin Tongluo decoction combined with hydroxychloroquine on improving submandibular gland secretion of Sjogren’s syndrome model mice

目的探讨益气养阴通络方联合羟氯喹对干燥综合征(SS)模型小鼠自发性颌下腺炎改善作用及其可能的机制。方法将40只雌性SS模型非肥胖糖尿病(NOD)小鼠随机分为模型组、羟氯喹组、中药组和联合用药组,各10只,另选10支雌性正常ICR小鼠,作为空白对照组。羟氯喹组给予硫酸羟氯喹灌胃,中药组给予益气养阴通络方颗粒灌胃,联合用药组给予益气养阴通络方颗粒和硫酸羟氯喹灌胃;模型组和空白对照组分别给予0.9%氯化钠注射液灌胃。分别于实验4、8、12周,采用湿重法测量小鼠唾液分泌量;实验12周时采用实时荧光定量聚合酶链反应测定各组小鼠外周血Toll样受体9(TLR-9)mRNA、B淋巴细胞刺激因子(BAFF)mRNA表达差异,比较各组颌下腺质量与指标,并观察各组小鼠颌下腺病理组织学改变。结果实验4、8、12周模型组唾液分泌量均低于空白对照组(均P<0.05)。实验12周,羟氯喹组、中药组、联合用药组唾液分泌量、颌下腺质量及颌下腺指数均高于模型组,且联合用药组均高于羟氯喹组[(26.4±3.8)g比(18.4±4.2)g、(88±11)mg比(80±10)mg、(3.4±0.7)比(2.9±0.8)](均P<0.05)。模型组TLR-9 mRNA、BAFF mRNA表达量均高于空白对照组,但羟氯喹组、中药组及联合用药组均低于模型组,且联合用药组均低于羟氯喹组[(3.4±0.2)比(4.4±0.2)、(3.2±0.3)比(3.7±0.4)](均P<0.05)。联合用药组颌下腺腺泡边界明显、无萎缩、无淋巴细胞浸润,与空白对照组相近。结论益气养阴通络方联合羟氯喹能够增加SS模型NOD小鼠唾液分泌量,可能通过影响TLR-9/干扰素/BAFF信号通路,下调BAFF的阳性表达,减轻颌下腺炎性浸润,延缓颌下腺萎缩。

Objective To investigate the effect of Yiqi Yangyin Tongluo decoction on submandibular gland secretion in Sjgren’s syndrome(SS)model mice and its possible mechanism.Methods Forty SS model non-obese diabetic(NOD)female mice were randomly divided into model group,hydroxychloroquine group,traditional Chinese medicine(TCM)group and combination group,with 10 mice in each group,and 10 female specific pathogen free ICR mice were selected as the blank control group.Hydroxychloroquine group was given sulfate hydroxychloroquine gavage,TCM group was given Yiqi Yangyin Tongluo decoction granules gavage,and combination group was given Yiqi Yangyin Tongluo decoction granules and hydroxychloroquine gavage.The model group and the blank control group were given 0.9%sodium chloride gavage.Wet weight method was used to measure the saliva flow of mice at 4 th,8 th and 12 th weeks respectively.The expression differences of Toll like receptor-9(TLR-9)mRNA and B lymphocyte stimulating factor(BAFF)mRNA in peripheral blood of each group were determined by real-time fluorescence quantitative polymerase chain reaction at 12 th week,and quality and index of submandibular gland were compared and the histopathological changes of submandibular gland were observed in each group.Results The salivary secretion of model group were lower than those of blank control group at 4 th,8 th and 12 th weeks(all P<0.05).At 12 th week,salivary secretion,submandibular gland quality and submandibular gland index in hydroxychloroquine group,TCM group and combination group were higher than those in model group,and those in combination group were higher than those in hydroxychloroquine group[(26.4±3.8)g vs(18.4±4.2)g,(88±11)mg vs(80±10)mg,(3.4±0.7)vs(2.9±0.8)](all P<0.05).The expressions of TLR-9 mRNA and BAFF mRNA in model group were higher than those in blank control group,and those in hydroxychloroquine group,TCM group and combination group were lower than those in model group,and those in combination group were lower than those in hydroxychloroquine group[(3.4±0.2)vs(4.4±0.2),(3.2±0.3)vs(3.7±0.4)](all P<0.05).In combination group,the boundary of submandibular gland acini was obvious,without atrophy and lymphocyte infiltration,which was similar to that of blank control group.Conclusions Combination of Yiqi Yangyin Tongluo decoction and hydroxychloroquine can increase the salivary secretion of SS model NOD mic.It may affect the TLR-9/Interferon/BAFF signaling pathway,down-regulate the positive expression of BAFF,reduce the inflammatory infiltration of the submandibular gland,and delay submandibular gland atrophy.