水通道蛋白5对干燥综合征大鼠TLR4/MyD88/NF-κB信号的影响

doi:10.19723/j.issn.1671-167X.2025.05.011

摘要/Abstract

摘要： 目的: 探讨水通道蛋白5(aquaporin 5, AQP5)对干燥综合征(Sjögren syndrome, SS)大鼠Toll样受体4(Toll-like receptor 4, TLR4)/髓样分化因子88(myeloid differentiation factor 88, MyD88)/核因子κB(nuclear factor κB, NF-κB)信号的影响。方法: 从基因表达综合数据库(Gene Expression Omnibus, GEO)中提取SS基因表达数据集GSE406611和GSE84844, 使用R软件分析AQP5的mRNA表达。构建大鼠SS模型, 将造模成功的大鼠随机分为SS组、SS+NC组和SS+pc组, 每组各10只; 取10只正常大鼠作为Normal组。SS+NC组大鼠于颌下腺位置每日一次皮下注射10 μg的rno-pcDNA3.1-AQP5-NC, SS+pc组大鼠于颌下腺位置每日一次皮下注射10 μg的rno-pcDNA3.1-AQP5, 连续注射28 d。采用酶联免疫吸附试验(enzyme-linked immunosorbent assay, ELISA)试剂盒检测大鼠血清中肿瘤坏死因子-α(tumor necrosis factor-α, TNF-α)和白细胞介素-1β(interleukin-1β, IL-1β)的含量, 采用高通量测序确定靶点基因, 采用实时荧光定量PCR(quantitative real-time PCR, qPCR)和Western blot检测大鼠颌下腺组织中AQP5、TLR4、MyD88、NF-κB的mRNA水平和蛋白表达情况。结果: 在SS数据集GSE406611和GSE84844中, AQP5的mRNA表达明显降低。与Normal组相比, SS组大鼠血清中TNF-α和IL-1β的含量明显升高, TLR4、MyD88、NF-κB的mRNA和蛋白表达明显升高, AQP5的mRNA和蛋白表达明显降低; 过表达AQP5后, SS+pc组大鼠血清中TNF-α和IL-1β的含量明显降低, TLR4、MyD88、NF-κB的mRNA和蛋白表达明显降低, AQP5的mRNA和蛋白表达明显升高, 差异均有统计学意义(均P < 0.05)。结论: AQP5参与SS的进展, AQP5的表达升高能明显抑制炎症性应激, 减轻颌下腺组织病理损伤, 这可能与抑制TLR4/MyD88/NF-κB信号的传导有关。

关键词: 水通道蛋白质5, 干燥综合征, Toll样受体4, 髓样分化因子88, NF-κB, 信号通路

Abstract: Objective: To investigate the effect of aquaporin 5 (AQP5) on Toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88)/nuclear factor κB (NF-κB) signaling pathway in Sjögren syndrome (SS) rats. Methods: The SS gene expression data sets GSE406611 and GSE84844 were extracted from the Gene Expression Omnibus (GEO), and the AQP5 mRNA expression was analyzed by R software. The rat SS model was constructed. The successfully modeled rats were divided into SS group, SS+NC group, and SS+pc group, 10 rats in each group; and 10 rats were set as Normal group. The rats in the SS+NC group were injected with 10 μg of rno-pcDNA3.1-AQP5-NC at the submandibular gland, subcutaneously every day for 28 days. The rats in the SS+pc group were injected with 10 μg of rno-pcDNA3.1-AQP5 at the submandibular gland, subcutaneously every day for 28 days. The enzyme-linked immunosorbent assay (ELISA) kit was used to detect the content of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in the serum. High-throughput sequencing was used to identify the target genes. Quantitative real-time PCR (qPCR) and Western blot were used to detect the mRNA and protein expressions of AQP5, TLR4, MyD88, and NF-κB in the rat submandibular gland tissue. Results: In the SS dataset GSE406611 and GSE84844, the mRNA expression of AQP5 in SS was significantly reduced. Compared with the Normal group, the content of TNF-α and IL-1β in the serum, the mRNA and protein expressions of TLR4, MyD88, and NF-κB in the SS group were significantly increased, the mRNA and protein expressions of AQP5 were significantly decreased. After overexpression of AQP5, the content of TNF-α and IL-1β in the serum, the mRNA and protein expressions of TLR4, MyD88, and NF-κB in the SS+pc group were significantly decreased, the mRNA and protein expressions of AQP5 were significantly increased. The differences were statistically significant (all P < 0.05). Conclusion: The expression of AQP5 is involved in the progression of SS. Increasing the expression of AQP5 can significantly inhibit inflammatory stress and reduce the pathological damage of submandibular gland tissue. This may be related to the inhibition of TLR4/MyD88/NF-κB conduction.

Key words: Aquaporin 5, Sjögren syndrome, Toll-like receptor 4, Myeloid differentiation factor 88, NF-kappa B, Signaling pathway

中图分类号:

R593.2

朱丽秀, 陈仁利, 周素娟, 林烨, 汤一榕, 叶桢. 水通道蛋白5对干燥综合征大鼠TLR4/MyD88/NF-κB信号的影响[J]. 北京大学学报（医学版）, 2025, 57(5): 875-883.

Lixiu ZHU, Renli CHEN, Sujuan ZHOU, Ye LIN, Yirong TANG, Zhen YE. Effect of aquaporin 5 on TLR4/MyD88/NF-κB signaling pathway in Sjögren syndrome rats[J]. Journal of Peking University (Health Sciences), 2025, 57(5): 875-883.

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Gene		Sequence
rno-AQP5	Forward	GGACCTGACCTGCCGTCTAG
rno-AQP5	Reverse	TAGCCCAGGATGCCCTTGAG
rno-TLR4	Forward	CCATGAGGCACATTGTTACG
rno-TLR4	Reverse	AAGTGCTTCACCACCTGCTT
rno-MyD88	Forward	CTAGCGACAAGCCATACACG
rno-MyD88	Reverse	GTAGCCGAATCGTAGCCAGA
rno-NF-κB	Forward	GTGGGTTCAGATGAGGAGGA
rno-NF-κB	Reverse	TCTGGTCCAAATAGGCTTGG
rno-GAPDH	Forward	TGTAATAATTGTAGCCAAGTAAATCTCC
rno-GAPDH	Reverse	AAGTAACCATTTTTCAAAACATTCAAG

Normal vs. SS				SS+NC vs. SS+pc
Up-regulated	Score	Down-regulated	Score	Up-regulated	Score	Down-regulated	Score
rno-TLR4	0.993 4	rno-AQP5	0.998 2	rno-AQP5	0.999 3	rno-TLR4	0.998 7
rno-MyD88	0.990 3	rno-ALOX15B	0.976 1	rno-AGTR1	0.976 4	rno-MyD88	0.973 4
rno-NF-κB	0.985 1	rno-ADCY7	0.962 5	rno-AGTR2	0.951 7	rno-NF-κB	0.950 5
rno-ATIC	0.963 7	rno-CACNA1S	0.948 9	rno-CALCA	0.934 8	rno-SPP1	0.893 9
rno-BDKRB1	0.952 9	rno-CAMK2B	0.930 3	rno-CAT	0.930 5	rno-SIRT1	0.891 6
rno-CCL2	0.941 8	rno-NCAM1	0.922 9	rno-NOS1	0.915 3	rno-TLR2	0.887 2
rno-CCR5	0.930 7	rno-NADK	0.917 7	rno-NOS2	0.909 1	rno-HTRIA	0.885 7
rno-IRF4	0.922 4	rno-MED1	0.901 4	rno-NR3C1	0.893 3	rno-GRM5	0.876 1
rno-JUN	0.913 8	rno-MC4R	0.899 0	rno-MTOR	0.877 2	rno-GNAT3	0.863 9
rno-LCK	0.892 9	rno-NCOA1	0.887 3	rno-KIT	0.861 7	rno-ESR1	0.852 8