牙周炎伴类风湿性关节炎患者牙龈浆细胞表型及RANKL表达特点分析

doi:10.13481/j.1671-587X.20230326

摘要/Abstract

摘要：

目的分析类风湿性关节炎（RA）对牙周炎患者牙龈浆细胞表型及核因子κB受体活化因子配体（RANKL）表达的影响，阐明其可能的机制。方法选择于本科就诊的60例研究对象，按入组标准分为健康组、牙周炎组和牙周炎+RA组，每组20例。记录各组研究对象牙龈指数（GI）、探诊深度（PD）、探诊出血指数（BOP）和临床附着丧失量（CAL）。采用实时荧光定量PCR（RT-qPCR）法检测各组研究对象牙龈组织中增殖诱导配体（APRIL）和B淋巴细胞刺激因子（BLyS） mRNA表达水平。采用流式细胞术分析各组研究对象牙龈组织中CD38+和CD138+细胞百分率，采用流式细胞术和酶联免疫吸附测定（ELISA）法检测各组研究对象牙龈浆细胞中RANKL+细胞百分率和RANKL水平，抗酒石酸酸性磷酸酶（TRAP）染色观察各组研究对象破骨样细胞生长情况，RT-qPCR法检测各组研究对象牙龈浆细胞中RANKL、核因子κB受体活化因子（RANK）和肿瘤坏死因子受体相关分子6（TRAF6）mRNA表达水平。结果与健康组比较，牙周炎组和牙周炎+RA组患者GI、PD、BOP和CAL均升高（P<0.05）；与牙周炎组比较，牙周炎+RA组患者PD和CAL升高（P<0.05）。与健康组比较，牙周炎组和牙周炎+RA组患者牙龈组织中APRIL和BLyS mRNA表达水平均升高（P<0.05）；与牙周炎组比较，牙周炎+RA组患者APRIL mRNA表达水平升高（P<0.05）。与健康组比较，牙周炎组和牙周炎+RA组患者牙龈组织中CD38+和CD138+细胞百分率升高（P<0.05）；与牙周炎组比较，牙周炎+RA组患者牙龈组织中CD138+细胞百分率升高（P<0.05）。与健康组比较，牙周炎组和牙周炎+RA组患者牙龈浆细胞中RANKL+细胞百分率升高（P<0.05）；与牙周炎组比较，牙周炎+RA组患者牙龈浆细胞中RANKL+细胞百分率升高（P<0.05）。与健康组比较，牙周炎组和牙周炎+RA组患者牙龈浆细胞中RANKL水平升高（P<0.05）。与健康组比较，牙周炎组和牙周炎+RA组正常诱导TRAP+破骨样细胞数增加（P<0.05）；与牙周炎组比较，牙周炎+RA组正常诱导TRAP+破骨样细胞数增加（P<0.05）；与正常诱导破骨样细胞比较，各组anti-RANKL诱导的TRAP+破骨样细胞数均减少（P<0.05）。与健康组比较，牙周炎组和牙周炎+RA组患者牙龈浆细胞中RANKL、RANK和TRAF6 mRNA表达水平均升高（P<0.05）；与牙周炎组比较，牙周炎+RA组患者牙龈浆细胞中RANKL、RANK和TRAF6 mRNA表达水平均升高（P<0.05）。结论牙周炎伴RA患者牙龈浆细胞分化水平及其诱导破骨样细胞分化能力均高于单纯牙周炎患者，可能是RA导致牙周炎患者牙周组织破坏加剧的潜在原因。

关键词: 牙周炎, 类风湿性关节炎, 浆细胞, 核因子κB受体活化因子配体

Abstract:

Objective To analyze the effect of rheumatoid arthritis （RA） on the phenotypes of plasma cells of gingiva and expression of receptor activator of nuclear factor-κB ligand（RANKL） in the patients with periodontitis，and to clarify its possible mechanism. Methods Sixty subjects who came to our hospital were selected and divided into healthy group， periodontitis group and periodontitis+RA group according to inclusion criteria， and there were 20 subjects in each group.The gingival index （GI）， probing depth （PD）， bleeding on probing （BOP） and clinical attachment loss （CAL） of the subjects were recorded. The expression levels of proliferation inducing ligand（APRIL） and B lymphocyte stimulator（BLyS） in gingiva tissue of the subjects in various groups were detected by real-time fluoresence quantatitive PCR（RT-qPCR） method；the percentages of CD38+ cells， CD138+ cells in gingiva tissue of the subjects in various groups were analyzed by flow cytometry；the percentages of RANKL+ cells in the gingival plasma cells and the RANKL levels in plasma cells of gingiva of the subjects in various groups were detected by flow cytometry and enzyme-linked immunosorbent assay（ELISA） methods； the growth of osteoclasts in the subjects in various groups was observed by anti-tartrate acid phosphatase （TRAP） staining； the expression levels of RANKL， receptor activator of nuclear factor-κB（RANK） and tumor necrosis factor associated factor 6（TRAF6） mRNA in plasma cells of gingiva of the subjects in various groups were detected by RT-qPCR method. Results Compared with healthy group， GI， PD， BOP and CAL of the patients in periodontitis group and periodontitis+RA group were significantly increased （P<0.05）； compared with periodontitis group，the PD and CAL of the patients in periodontitis+RA group were increased（P<0.05）.Compared with healthy group， the expression levels of APRIL and BLyS mRNA in gingiva tissue of the patients in periodontitis group and periodontitic+PA group were increased（P<0.05）；compared with periodontitis group，the expression level of APRIL mRNA of the patients in periodontitis+RA group was increased（P<0.05）.Compared with healthy group， the percentages of CD38+ cells and CD138+ cells in gingiva tissue of the patients in periodontitis group and periodontitis+RA group were significantly increased（P<0.05）；compared with periodontitis group， the percentage of CD138+ cells in gingiva tissue of the patients in periodontitis+RA group was increased （P<0.01）.Compared with healthy group， the percentages of RANKL+ cells in plasma cells of gingiva of the patients in periodontitis group and periodontitis+RA group were increased（P<0.05）；compared with periodontitis group， the percentage of RANKL+ cells in the plasma cells of gingiva the patients in periodontitis+RA group was increased（P<0.05）. Compared with healthy group， the percentages of RANKL in plasma cells of gingiva of the patients in periodontitis group and periodontitis+RA group were increased（P<0.05）. Compared with healthy group， the number of normal induced TRAP+osteoblast-like cells of the patients in periodontitis group and periodontitis+RA group were increased（P<0.05）； compared with periodontitis group， the number of normal induced TRAP+ osteoblast-like cells of the patients in periodontitis+RA group was increased （P<0.05）； compared with the normal induced cells， the numbers of anti-RANKL induced TRAP+osteoblast-like cells in various groups were decreased（P<0.05）.Compared with healthy group，the expression levels of RANKL， RANK， and TRAF6 mRNA in the plasma cells of gingiva of the patients in periodontitis group and periodontitis+RA group were significantly increased （P<0.05）；compared with periodontitis group， the expression levels of RANKL， RANK， and TRAF6 mRNA in the plasma cells of gingiva of the patients in periodontitis+RA group were increased （P<0.05）. Conclusion The differentiation level and the ability on promoting osteoclastogenesis of plasma cells of gingival of the patients with periodontitis complicated with RA are higher than those in the patients with single periodontitis， which may be a potential cause for RA aggravating periodontal tissue destruction in the patients with periodontitis.

Key words: Periodontits, Rheumatoid arthritis, Plasma cell, Receptor activator of nuclear factor-κB ligand

中图分类号:

R781.4

于艳,于程程,韩亚琨. 牙周炎伴类风湿性关节炎患者牙龈浆细胞表型及RANKL表达特点分析[J]. 吉林大学学报(医学版), 2023, 49(3): 757-764.

Yan YU,Chengcheng YU,Yakun HAN. Analysis on phenotypes of plasma cells of gingiva and expression characteristics of RANKL in patients with periodontitis complicated with rheumatoid arthritis[J]. Journal of Jilin University(Medicine Edition), 2023, 49(3): 757-764.

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Group	GI	PD(l/mm)	BOP	CAL(l/mm)
Health	0.40±0.55	2.00±1.00	2.00±1.58	1.20±0.44
Periodontitis	2.40±0.89^*	5.40±1.82^*	13.80±3.11^*	4.20±1.48^*
Periodontitis+RA	2.60±0.55^*	6.40±2.41^*△	12.00±3.81^*	4.80±1.64^*△

Group	APRIL mRNA	BLyS mRNA
Health	1.00±0.00	1.00±0.00
Periodontitis	5.94±1.52^*	10.20±1.48^*
Periodontitis+RA	7.95±1.01^*△	10.70±2.09^*

Group	Percentage of CD38+ cells	Percentage of CD138+ cells
Health	21.80±5.32	15.70±6.97
Periodontitis	53.20±10.10^*	33.90±7.51^*
Periodontitis+RA	54.90±7.86^*	38.30±5.35^*△

Group	Number of osteoclasto-like cells
Group	Normal induction	Anti-RANKL induction
Health	1.6±1.1	0.6±0.2^#
Periodontitis	4.8±2.8^*	0.6±0.4^#
Periodontitis+RA	7.6±3.1^*△	0.8±0.5^#

Group	RANKL mRNA	RANK mRNA	TRAF6 mRNA
Health	1.00±1.00	1.00±1.00	1.00±1.00
Periodontitis	2.96±0.96^*	2.74±0.87^*	2.07±0.45^*
Periodontitis+RA	4.41±0.99^*△	4.89±1.88^*△	3.18±1.29^*△