线粒体解偶联蛋白2在大鼠实验性牙周炎相关肾损伤中的作用研究

doi:10.7518/hxkq.2024.2023378

华西口腔医学杂志 ›› 2024, Vol. 42 ›› Issue (4): 502-511.doi: 10.7518/hxkq.2024.2023378

线粒体解偶联蛋白2在大鼠实验性牙周炎相关肾损伤中的作用研究

李琼¹(), 马浩楠¹, 商雅琦¹, 辛禧瑞¹, 刘歆婵², 武洲³, 于维先⁴^,⁵()

^1.吉林大学口腔医院牙周病科，长春 130021
^2.吉林大学口腔医院种植科，长春 130021
^3.日本九州大学齿学研究院，日本福冈 812-8582
^4.吉林大学口腔医院老年口腔科，长春 130021
^5.吉林省牙发育及颌骨重塑与再生重点实验室，长春 130021

收稿日期:2023-11-10 修回日期:2024-02-17 出版日期:2024-08-01 发布日期:2024-07-17
通讯作者: 于维先 E-mail:liqiong21@mails.jlu.edu.cn;ywx@jlu.edu.cn
作者简介:李琼，住院医师，硕士，E-mail：liqiong21@mails.jlu.edu.cn
基金资助:
吉林省科技厅国际合作项目(20220402069GH)

The role of uncoupling protein 2 in experimental periodontitis-associated renal injury in rats

Li Qiong¹(), Ma Haonan¹, Shang Yaqi¹, Xin Xirui¹, Liu Xinchan², Wu Zhou³, Yu Weixian⁴^,⁵()

^1.Dept. of Periodontology, Hospital of Stomatology, Jilin University, Changchun 130021, China
^2.Dept. of Implantology, Hospital of Stomatology, Jilin University, Changchun 130021, China
^3.Institute of Dentistry, Kyushu University, Fukuoka 812-8582, Japan
^4.Dept. of Geriatrics, Hospital of Stomatology, Jilin University, Changchun 130021, China
^5.Jilin Provincial Key Laboratory of Tooth Development and Bone Remodeling, Changchun 130021, China

Received:2023-11-10 Revised:2024-02-17 Online:2024-08-01 Published:2024-07-17
Contact: Yu Weixian E-mail:liqiong21@mails.jlu.edu.cn;ywx@jlu.edu.cn
Supported by:
International Cooperation Project of Science and Technology Department of Jilin Province(20220402069GH)

摘要/Abstract

摘要：

目的探索线粒体解偶联蛋白2（UCP2）在结扎诱导的实验性牙周炎相关肾损伤中的变化，研究UCP2对牙周炎诱导肾损伤的影响。方法选取12只Wistar雄性大鼠随机分为2组：对照组和牙周炎组。使用0.2 mm正畸结扎丝结扎在大鼠上颌第一磨牙的牙颈部，建立牙周炎模型。8周后观察2组大鼠口内情况，检测牙周临床指标牙龈出血指数（BI）、牙周探诊深度（PD）、牙齿松动度（TM）。收集大鼠上颌骨并扫描Micro CT，观察牙槽骨吸收情况，记录组织矿物质密度（TMD）、骨矿物质密度（BMD）、骨体积分数（BV/TV）、骨小梁厚度（Tb.Th）、骨小梁骨分离度（Tb.Sp）等参数，测量上颌第一磨牙釉牙骨质界到牙槽嵴顶的距离（CEJ-ABC）。冻存大鼠肾组织以检测氧化应激指标丙二醛（MDA）、谷胱甘肽（GSH）、超氧化物歧化酶（SOD），并进行实时定量聚合酶链反应（qRT-PCR）观察UCP2、核因子红系2相关因子2（Nrf2）抗体、过氧化物酶体增殖物激活受体γ辅激活因子-1α（PGC-1α）基因表达情况。使用大鼠牙龈组织进行免疫组织化学染色，观察牙龈组织UCP2表达情况。使用大鼠固定的肾组织进行苏木精-伊红（HE）、过碘酸雪夫（PAS）、MitoSOX Red、JC-1、免疫组织化学染色，观察大鼠肾组织病理学表现、活性氧（ROS）水平、线粒体膜电位水平及UCP2、Nrf2、PGC-1α蛋白表达情况。收集大鼠血清检测肾脏功能指标血尿素氮（BUN）、肌酐（Cre）、白蛋白（Alb）的表达。结果相较于对照组，牙周炎组大鼠第一磨牙周围的牙龈组织发红肿胀，质地松软，临床检查探诊出血且PD增加，牙齿松动，牙槽骨吸收，TMD、BMD、BV/TV、Tb.Th指数降低，Tb.Sp指数及CEJ-ABC增加，牙龈UCP2蛋白表达升高。相较于对照组，牙周炎组大鼠肾组织内MDA、ROS水平上升，MMP、GSH和SOD水平下降，UCP2基因及蛋白表达升高，Nrf2、PGC-1α基因及蛋白表达下降。肾脏功能指标BUN、Cre、Alb两组差异无统计学意义。结论 UCP2可通过氧化应激在牙周炎诱导的肾损伤中发挥作用。

关键词: 牙周炎, 肾损伤, 线粒体解偶联蛋白2

Abstract:

Objective This study aims to explore changes in uncoupling protein 2 (UCP2) in experimental periodontitis-associated renal injury induced by ligation and investigate the effect of UCP2 on renal injury induced by periodontitis. Methods Twelve Wistar male rats were randomly divided into two groups: control and periodontitis groups. A periodontal model was built by ligating the maxillary first molars area with 0.2 mm orthodontic ligature wire. After 8 weeks, the intraoral condition of the rats was observed and periodontal clinical indices such as gingival bleeding index (BI), periodontal probing depth (PD), and tooth mobility (TM) were detected. The maxillary bone was scanned by Micro CT to observe the alveolar bone resorption. The tissue mineral density (TMD), bone mineral density (BMD), bone volume fraction (BV/TV), trabecular thickness (Tb.Th), trabecular bone separation (Tb.Sp) were recorded, and the distance from the enamel bone boundary to the alveolar crest (CEJ-ABC) of the maxillary first molar was measured. The oxidative stress indexes such as malondialdehyde, glutathione (GSH), and superoxide dismutase (SOD) were detected using frozen rat kidney tissue. The gene expression of UCP2, nuclear factor erythroid 2-related factor 2 (Nrf2), and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) was observed by quantitative real-time polymerase chain reaction (qRT-PCR) test. The gingival tissue of the rats was used for immunohistochemical staining to observe the expression of the UCP2 protein. The fixed rat kidney tissue was used for hematoxylin-eosin (HE), periodic acid-schiff (PAS), MitoSOX Red, JC-1, and immunohistochemical staining to observe the renal histopathology, the level of reactive oxygen species (ROS), the level of mitochondrial membrane potential, and the expression of UCP2, Nrf2, and PGC-1α protein. Rat serum was collected to detect renal function indices, namely, blood urea nitrogen (BUN), creatinine (Cre), and albumin (Alb). Results Compared with the control group, the periodontitis group showed red, swollen, and soft gingival tissue, with gingival probing bleeding, periodontal PD increased, tooth loosening, alveolar bone resorption, decreased TMD, BMD, BV/TV, and Tb.Th indices, and increased Tb.Sp index, CEJ-ABC, and gingival UCP2 protein expression. Compared with the control group, the levels of MDA and ROS in the kidney tissue of periodontitis rats and the gene and protein expression of UCP2 increased, and the levels of MMP, GSH, and SOD and the gene and protein expression of Nrf2 and PGC-1α decreased. Renal functional indices, namely, BUN, Cre, and Alb, were not significantly different between the two groups. Conclusion UCP2 may play a role in renal injury induced by periodontitis through oxidative stress.

Key words: periodontitis, renal injury, uncoupling protein 2

中图分类号:

R781.4

李琼, 马浩楠, 商雅琦, 辛禧瑞, 刘歆婵, 武洲, 于维先. 线粒体解偶联蛋白2在大鼠实验性牙周炎相关肾损伤中的作用研究[J]. 华西口腔医学杂志, 2024, 42(4): 502-511.

Li Qiong, Ma Haonan, Shang Yaqi, Xin Xirui, Liu Xinchan, Wu Zhou, Yu Weixian. The role of uncoupling protein 2 in experimental periodontitis-associated renal injury in rats[J]. West China Journal of Stomatology, 2024, 42(4): 502-511.

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参考文献 26

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基因	引物序列（5’-3’）	碱基数/bp	基因号
UCP2	F：CAAGACCATTGCACGAGAGGA R：GAGGTTGGCTTTCAGGAGAGTAT	139	NM_019354.3
Nrf2	F：TTGGCAGAGACATTCCCATTTGTA R：GAGCTATCGAGTGACTGAGCCTG	116	NM_031789.3
PGC-1ɑ	F：GCACTGACAGATGGAGACGTGA R：TCATTGTAGCTGAGCTGAGTGTTGG	149	NM_031347.1
β-actin	F：GGAGATTACTGCCCTGGCTCCTA R：GACTCATCGTACTCCTGCTTGCTG	150	NM_031144.3

牙周临床指标	对照组	牙周炎组
PD/mm	0.21±0.06	2.43±0.29***
BI	0.00±0.00	2.33±0.76**
TM	0.00±0.00	1.17±0.29**

肾脏功能指标	对照组	牙周炎组
BUN/（mmol/L）	6.04±1.86	6.07±0.48
Cre/（μmol/L）	25.55±7.94	26.43±3.28
Alb/（g/L）	19.90±1.70	20.89±2.63

功能指标	对照组	牙周炎组
MDA/（nmol/mgprot）	3.17±0.23	4.62±0.12***
GSH/（μmol/gprot）	33.97±1.58	24.19±3.47*
SOD/（U/mgprot）	74.00±4.79	59.84±7.03*

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线粒体解偶联蛋白2在大鼠实验性牙周炎相关肾损伤中的作用研究

The role of uncoupling protein 2 in experimental periodontitis-associated renal injury in rats

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