电离辐射对大鼠腮腺旁细胞分泌途径损伤及紧密连接蛋白claudin-4的影响

doi:10.19439/j.sjos.2022.04.005

上海口腔医学 ›› 2022, Vol. 31 ›› Issue (4): 359-366.doi: 10.19439/j.sjos.2022.04.005

电离辐射对大鼠腮腺旁细胞分泌途径损伤及紧密连接蛋白claudin-4的影响

姚清婷^1,2, 吴言辉^1,2, 刘少华^3,4, 许辉¹, 李军¹, 石亮^3,4

1.新疆维吾尔自治区人民医院口腔颌面外科,新疆乌鲁木齐 830001;
2.新疆医科大学,新疆乌鲁木齐 830054;
3.山东大学齐鲁医院口腔颌面外科,山东济南 250012;
4.山东大学口腔医学研究所,山东济南 250012

收稿日期:2021-01-28 修回日期:2021-04-17 出版日期:2022-08-25 发布日期:2022-08-30
通讯作者: 石亮,E-mail：shiliang1976@126.com
作者简介:姚清婷 (1993-),女,硕士研究生,E-mail：yaoqingting@126.com
基金资助:
山东省重点研发计划项目（GG201709220011）; 山东省自然科学基金（ZR2020MH187）

Damage on paracellular pathway and effect on tight junction protein claudin-4 in rat parotid glands caused by ionizing radiation

YAO Qing-ting^1,2, WU Yan-hui^1,2, LIU Shao-hua^3,4, XU Hui¹, LI Jun¹, SHI Liang^3,4

1. Department of Oral and Maxillofacial Surgery, People's Hospital of Xinjiang Uygur Autonomous Region. Urumqi 830001, Xinjiang Uygur Autonomous Region;
2. Xinjiang Medical University. Urumqi 830054, Xinjiang Uygur Autonomous Region;
3. Department of Oral and Maxillofacial Surgery, Qilu Hospital of Shandong University. Jinan 250012, Shandong Province;
4. Institute of Stomatology, Shandong University. Jinan 250012, Shandong Province, China

Received:2021-01-28 Revised:2021-04-17 Online:2022-08-25 Published:2022-08-30

摘要/Abstract

摘要： 目的: 探讨电离辐射对大鼠腮腺旁细胞分泌功能损伤及紧密连接 (tight junction,TJ) 蛋白claudin-4的影响与潜在作用机制。方法: 24只8周龄雄性Wistar大鼠,随机分为对照组（6只）和照射组（照射后1周组、4周组、12周组,每组6只）。照射组一次性20 Gy射线局部照射实验侧腮腺区。采用Schirmer实验,检测各组腺体唾液静息分泌量;H-E染色,光镜下观察腺体组织病理变化;透射电镜观察TJ超微结构改变;免疫荧光染色和蛋白质印迹检测毒蕈碱型乙酰胆碱受体（M受体）亚型M3、水通道蛋白5（AQP5）及claudin-4的蛋白表达。采用SPSS 23.0软件包对数据进行统计学分析。结果: 照射后1、4、12周,腺体静息分泌量较对照组显著减少（P<0.05）;12周较1周、4周时减少更显著（P<0.05）。组织学观察可见,照射后腺体早期间质血管扩张、充血,后期腺泡细胞计数显著减少（P<0.05）;TJ结构模糊、电子密度降低、宽度减少（P<0.05）。免疫荧光染色和蛋白质印迹检测结果显示,M3受体及AQP5蛋白在照射后1、4、12周表达显著下调,而claudin-4蛋白表达显著升高。结论: 电离辐射后腮腺旁细胞分泌功能降低,TJ结构改变,claudin-4表达上调,可能参与腺体分泌功能损伤。

关键词: 电离辐射, 紧密连接, Claudin-4, 旁细胞途径, 腮腺

Abstract: PURPOSE: To investigate the effect and the potential mechanism of ionizing radiation on secretory function and tight junction (TJ) protein claudin-4 in paracellular pathway of rat parotid glands. METHODS: Twenty four 8-week-old male Wistar rats were randomly divided into one control group(n=6) and three irradiation groups (i.e., 1-week group post-irradiation, 4-week group post-irradiation, and 12-week group post-irradiation, 6 rats in each group). The experimental glands of irradiation groups were exposed to X-radiation in one-time single doses of 20 Gy. The residual salivary secretion of parotid glands was measured by Schirmer's test. The pathological changes of gland tissues were observed under light microscope after hematoxylin-eosin(H-E) staining. The changes of TJs ultrastructure were observed under transmission electron microscopy (TEM). Immunofluorescence staining and Western blot were used to detect the expression levels of muscarinic acetylcholine M3 subtype receptor, aquaporin 5 (AQP5), and claudin-4 proteins. The experimental results were analyzed with SPSS 23.0 software package for one-way analysis of variance. RESULTS: The residual salivary secretion of irradiation group glands at 1, 4, and 12 weeks after irradiation was reduced compared with that of the control group(P<0.05), and the residual salivary secretion of irradiation group at 12 weeks was significantly lower than that at 4 weeks(P<0.05). Histologically, the dilation and congestion of interstitial vessels were observed at early stage after irradiation, and significant reduced number of acinar cells was found at late stage(P<0.05). In the irradiation groups, the ultrastructures of TJ were fuzzy, the electron density was decreased, and the TJ width at 1, 4, and 12 weeks was reduced compared with that in the control group. Immunofluorescence staining and Western blot indicated that the protein expression levels of M3 and AQP5 were down-regulated; however, the protein expression levels of claudin-4 were significantly increased at 1, 4, and 12 weeks after irradiation. CONCLUSIONS: After ionizing radiation, decreased secretory function of paracellular pathway, alterations in TJ structures, and up-regulation of claudin-4 expression may be involved in the mechanism of hyposecretion in rat parotid glands after irradiation.

Key words: Ionizing radiation, Tight junction, Claudin-4, Paracellular pathway, Parotid gland

中图分类号:

R739.8

姚清婷, 吴言辉, 刘少华, 许辉, 李军, 石亮. 电离辐射对大鼠腮腺旁细胞分泌途径损伤及紧密连接蛋白claudin-4的影响[J]. 上海口腔医学, 2022, 31(4): 359-366.

YAO Qing-ting, WU Yan-hui, LIU Shao-hua, XU Hui, LI Jun, SHI Liang. Damage on paracellular pathway and effect on tight junction protein claudin-4 in rat parotid glands caused by ionizing radiation[J]. Shanghai Journal of Stomatology, 2022, 31(4): 359-366.

参考文献

[1] Jensen SB, Vissink A, Limesand KH, et al.Salivary gland hypofunction and xerostomia in head and neck radiation patients[J]. J Natl Cancer Inst Monogr, 2019, 2019(53): 95-106.
[2] Vissink A, 's-Gravenmade EJ, Ligeon EE, et al. A functional and chemical study of radiation effects on rat parotid and submandibular/sublingual glands[J]. Radiat Res, 1990, 124(3): 259-265.
[3] Zhang T, Liu C, Ma S, et al.Protective effect and mechanism of action of rosmarinic acid on radiation-induced parotid gland injury in rats[J]. Dose Response, 2020, 18(1): 1559325820907782.
[4] Zhang XM, Huang Y, Cong X, et al.Parasympathectomy increases resting secretion of the submandibular gland in minipigs in the long term[J]. Cell Physiol, 2019, 234(6): 9515-9524.
[5] Cong X, Zhang Y, Li J, et al.Claudin-4 is required for modulation of paracellular permeability by muscarinic acetylcholine receptor in epithelial cells[J]. J Cell Sci, 2015, 128(12): 2271-2286.
[6] Krane CM, Melvin JE, Nguyen HV, et al.Salivary acinar cells from aquaporin 5-deficient mice have decreased membrane water permeability and altered cell volume regulation[J]. J Biol Chem, 2001, 276(26): 23413-23420.
[7] Ding C, Li L, Su YC, et al.Adiponectin increases secretion of rat submandibular gland via adiponectin receptors-mediated AMPK signaling[J]. PLoS One, 2013, 8(5): e63878.
[8] Yokoyama M, Narita T, Sakurai H, et al.Maintenance of claudin-3 expression and the barrier functions of intercellular junctions in parotid acinar cells via the inhibition of Src signaling[J]. Arch Oral Biol, 2017, 81: 141-150.
[9] Ulmansky M, Sela J, Dishon T, et al.A technique for the intubation of the parotid duct in rats[J]. Arch Oral Biol, 1972, 17(3): 609-612.
[10] López-Jornet P, Camacho-Alonso F, Bermejo-Fenoll A.A simple test for salivary gland hypofunction using oral Schirmer's test[J]. J Oral Pathol Med, 2006, 35(4): 244-248.
[11] Qi W, Cong X, Zhang XM, et al.Parasympathectomy increases resting salivary secretion in normal and irradiated submandibular glands of rats[J]. Eur J Oral Sci, 2017, 125(2): 110-118.
[12] Zeilstra LJ, Vissink A, Konings AW, et al.Radiation induced cell loss in rat submandibular gland and its relation to gland function[J]. Int J Radiat Biol, 2000, 76(3): 419-429.
[13] Li Z, Zhao D, Gong B, et al.Decreased saliva secretion and down-regulation of AQP5 in submandibular gland in irradiated rats[J]. Radiat Res, 2006, 165(6): 678-687.
[14] Segawa A.Tight junctional permeability in living cells: dynamic changes directly visualized by confocal laser microscopy[J]. J Electron Microsc (Tokyo), 1994, 43(5): 290-298.
[15] Hashimoto S, Murakami M.Morphological evidence of paracellular transport in perfused rat submandibular glands[J]. J Med Invest, 2009, 56(Suppl): 395-397.
[16] Coppes RP, Zeilstra LJ, Kampinga HH, et al.Early to late sparing of radiation damage to the parotid gland by adrenergic and muscarinic receptor agonists[J]. Br J Cancer, 2001, 85(7): 1055-1063.
[17] Avila JL, Grundmann O, Burd R, et al.Radiation-induced salivary gland dysfunction results from p53-dependent apoptosis[J]. Int J Radiat Oncol Biol Phys, 2009, 73(2): 523-529.
[18] Konings AW, Coppes RP, Vissink A.On the mechanism of salivary gland radiosensitivity[J]. Int J Radiat Oncol Biol Phys, 2005, 62(4): 1187-1194.
[19] Ishikawa Y, Yuan Z, Inoue N, et al.Identification of AQP5 in lipid rafts and its translocation to apical membranes by activation of M3 mAChRs in interlobular ducts of rat parotid gland[J]. Am J Physiol Cell Physiol, 2005, 289(5): C1303-C1311.
[20] Takakura K, Takaki S, Takeda I, et al.Effect of cevimeline on radiation-induced salivary gland dysfunction and AQP5 in submandibular gland in mice[J]. Bull Tokyo Dent Coll, 2007, 48(2): 47-56.
[21] Baker OJ.Current trends in salivary gland tight junctions[J]. Tissue Barriers, 2016, 4(3): e1162348.
[22] Cong X, Zhang XM, Zhang Y, et al.Disruption of endothelial barrier function is linked with hyposecretion and lymphocytic infiltration in salivary glands of Sjögren's syndrome[J]. Biochim Biophys Acta Mol Basis Dis, 2018, 1864(10): 3154-3163.
[23] Zhang LW, Cong X, Zhang Y, et al.Interleukin-17 impairs salivary tight junction integrity in Sjögren's syndrome[J]. J Dent Res, 2016, 95(7): 784-792.
[24] Min SN, Wu LL, Zhang YY, et al.Disruption of tight junction structure contributes to secretory dysfunction in IgG4-related sialadenitis[J]. J Mol Histol, 2020, 51(1): 33-46.
[25] Huang Y, Mao QY, Shi XJ, et al.Disruption of tight junctions contributes to hyposalivation of salivary glands in a mouse model of type 2 diabetes[J]. J Anat, 2020, 237(3): 556-567.
[26] Nighot PK, Blikslager AT.ClC-2 regulates mucosal barrier function associated with structural changes to the villus and epithelial tight junction[J]. Am J Physiol Gastrointest Liver Physiol, 2010, 299(2): G449-G456.
[27] Peppi M, Ghabriel MN.Tissue-specific expression of the tight junction proteins claudins and occludin in the rat salivary glands[J]. J Anat, 2004, 205(4): 257-66.
[28] Zhang XM, Huang Y, Zhang K, et al.Expression patterns of tight junction proteins in porcine major salivary glands: a comparison study with human and murine glands[J]. J Anat, 2018, 233(2): 167-176.
[29] Cook DI, Dinudom A, Komwatana P, et al.Control of Na+ transport in salivary duct epithelial cells by cytosolic Cl- and Na+[J]. Eur J Morphol, 1998, 36(Suppl): 67-73.
[30] Qu W, Zhang Lj, Ao Jf.Radiotherapy induces intestinal barrier dysfunction by inhibiting autophagy[J]. ACS Omega, 2020, 5(22): 12955-12963.
[31] Shi L, Xiao M, Dai ML, et al.Ischemia preconditioning protects rat submandibular glands from ischemia/reperfusion injuries[J].Eur J Oral Sci, 2014, 122(5): 324-331.
[32] Narita T, Qi B, Murakami M, et al.Pilocarpine induces the residual secretion of salivary fluid in perfused submandibular glands of rats[J]. PLoS One, 2019, 14(8): e0221832.

电离辐射对大鼠腮腺旁细胞分泌途径损伤及紧密连接蛋白claudin-4的影响

Damage on paracellular pathway and effect on tight junction protein claudin-4 in rat parotid glands caused by ionizing radiation

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价

[1]	马学花, 徐江, 夏飞飞, 方桃莉, 孙志鹏. 腮腺深叶良性肿瘤：与定位和手术入路相关的分类[J]. 上海口腔医学, 2024, 33(4): 387-392.
[2]	陈倩倩, 余强, 孙晶晶, 熊屏. 12例Warthin瘤样黏液表皮样癌影像学分析[J]. 上海口腔医学, 2024, 33(2): 219-224.
[3]	杨成龙, 杨蒙, 王宇峰, 宋晨成, 杜观环, 唐国瑶. 具核梭杆菌来源的外膜囊泡对口腔上皮细胞闭合蛋白Claudin-4表达的影响[J]. 上海口腔医学, 2023, 32(2): 126-131.
[4]	肖华, 陶晓峰, 朱文静, 陈倩倩. 19例腮腺腺泡细胞癌的MRI影像表现分析[J]. 上海口腔医学, 2022, 31(3): 322-326.
[5]	徐万林, 朱云, 卢浩, 丁继平, 姚原, 刘胜文, 张陈平, 杨雯君. 腮腺区广泛型复发性多形性腺瘤的面神经处理及术后放疗经验：附10例报告[J]. 上海口腔医学, 2021, 30(6): 667-672.
[6]	齐鑫, 谢敬生, 王梓欣. 腮腺保留式调强放疗对唾液组成、流量及口干的影响[J]. 上海口腔医学, 2020, 29(4): 396-399.
[7]	常攀辉, 艾克白尔·吐日洪, 李崴嵬. mTOR及Tfr1 mRNA在腮腺黏液表皮样癌组织中的表达及与预后的关系[J]. 上海口腔医学, 2019, 28(5): 499-503.
[8]	郑重阳, 张新宇, 曹锐, 刘浏, 胡永杰. 超声刀辅助下腮腺良性肿瘤功能性包膜外切除术的临床效果评价[J]. 上海口腔医学, 2019, 28(4): 402-407.
[9]	胡赟, 张艳胜. CD44v6与COX-2在腮腺多形性腺瘤及癌在多形性腺瘤中的表达及临床意义[J]. 上海口腔医学, 2019, 28(4): 412-416.
[10]	高明华, 尹雪莱, 胡永杰, 盛美春, 王忠华, 郑重阳, 张新宇, 吴海威. 52例腮腺区神经鞘瘤临床治疗及预后分析[J]. 上海口腔医学, 2018, 27(4): 411-414.
[11]	张蔚蒨, 龚霞, 熊屏. 超声弹性成像在干燥综合征唾液腺病变中的应用价值[J]. 上海口腔医学, 2018, 27(1): 73-78.
[12]	赵志国, 高丹, 王瑾, 张力平. 896例腮腺肿瘤临床回顾分析[J]. 上海口腔医学, 2017, 26(6): 605-609.
[13]	高振然, 马松波, 张国琴. C形小切口治疗腮腺后下极良性肿瘤48例临床分析[J]. 上海口腔医学, 2017, 26(6): 666-668.
[14]	刘华, 周驰, 纪均. 不同剂量右美托咪定对腮腺手术患者全麻苏醒质量的影响[J]. 上海口腔医学, 2016, 25(3): 368-372.
[15]	王智明，叶明，杨东升，马颖，苏欣，张维庆，潘遥，关乔丹. 45例腮腺囊肿临床病理分析[J]. 上海口腔医学, 2016, 25(2): 199-203.