脂联素干预糖尿病牙周炎模型小鼠牙周组织及血浆中的脂联素水平
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摘要
背景:低水平表达的脂联素在牙周炎和糖尿病的发生与发展进程之中均发挥着明显的促进作用。目的:探讨脂联素注射对糖尿病并发牙周炎病理模型小鼠的治疗作用及其机制。方法:60只8周龄的昆明系小鼠,由北京维通利华动物实验技术有限公司提供。小鼠被平均随机分为3组,其中模型组和脂联素组中所有小鼠均接受一次性的链脲佐菌素腹腔注射从而建立小鼠糖尿病模型,于牙周结扎部位涂抹联合菌液以建立小鼠牙周炎模型。正常对照组和糖尿病牙周炎模型组小鼠每日1次接受1 mL生理盐水腹腔注射,脂联素组小鼠则每日接受1次15μg/kg的脂联素腹腔注射。治疗持续6周,酶联免疫吸附法(ELISA)检测小鼠血清及牙周组织中脂联素的水平、血糖及血浆胰岛素的水平;观察小鼠的牙周探诊深度以及牙槽骨垂直吸收高度;苏木精-伊红染色检测牙周组织病理变化;TUNEL法检测牙龈上皮细胞凋亡水平变化,RT-PCR和Western blot检测RAGE、活性氧基因和蛋白表达。结果与结论:①脂联素组小鼠的血清和牙周组织中的脂联素水平较正常对照组和模型组明显升高(P<0.05);②脂联素组较模型组小鼠血糖水平明显降低(P <0.05),血清胰岛素水平明显升高(P <0.05);③脂联素组小鼠的牙周探诊深度和牙槽骨垂直吸收高度较模型组明显减轻,牙龈上皮细胞凋亡率也明显降低;④RAGE、活性氧基因和蛋白表达水平脂联素组较模型组明显减轻(P <0.05);⑤结果说明,脂联素干预可以提高糖尿病牙周炎病理模型小鼠牙周组织及血浆的脂联素水平,并可以明显改善小鼠的血糖状态;减轻小鼠牙周组织的病理变化及牙周细胞的凋亡,此系通过调节RAGE-ROS信号通路发挥以上调节作用。
BACKGROUND: Low level of adiponectin plays a significant role in the occurrence and development of periodontitis and diabetes mellitus.OBJECTIVE: To investigate the therapeutic effect of adiponectin injection on diabetic periodontitis model mice and its mechanism.METHODS: Sixty 8-week old Kunming mice(provided by Beijing Weitong Lihua Animal Experimental Technology Co., Ltd.) were randomly divided into three groups: control group, model group and adiponectin group. The mice in the model and adiponectin groups received intraperitoneal injection of streptozotin to establish a mouse model of diabetes mellitus, and then the periodontitis model was established by applying the combined bacterial fluid to the ligation site. The mice in the control and model groups received intraperitoneal injection of 1 mL normal saline once daily, while adiponectin group received intraperitoneal injection of 15 μg/kg adiponectin once daily, for 6 consecutive weeks. The three experimental groups of mice were treated for 6 weeks, after which physiological and biochemical indicators were tested.The levels of adiponectin in serum and periodontal tissue, the blood sugar and plasma insulin levels were then detected by ELISA. The depth of periodontal examination and the vertical absorption of alveolar bone were observed. The pathological changes of periodontal tissue were detected by hematoxylin-eosin staining. The changes of apoptosis level of gingival epithelial cells were detected by TUNEL method. The expression levels of RAGE and reactive oxygen species mRNA and protein were detected by RT-PCR and western blot assay.RESULTS AND CONCLUSION:(1) The levels of adiponectin in serum and periodontal tissue in the adiponectin group were significantly higher than those in the control and model groups(P < 0.05).(2) The blood sugar level in the adiponectin group was significantly lower than that in the model group(P < 0.05) and the serum insulin level was significantly increased(P < 0.05).(3) The depth of periodontal examination and vertical absorption of alveolar bone were decreased significantly in the adiponectin group compared with the model group, and the apoptotic rate of gingival epithelial cells was decreased significantly.(4) The expression levels of RAGE and reactive oxygen species mRNA and protein in the adiponectin group were significantly lower than those in the model group(P < 0.05).(5) These results suggest that adiponectin can improve the adiponectin level in periodontal tissue and plasma in the diabetic periodontitis model mice and can significantly improve the blood sugar status, and reduce the pathological changes of periodontal tissue and the apoptosis of periodontal cells possibly via regulating RAGE-ROS signaling pathway.
BACKGROUND: Low level of adiponectin plays a significant role in the occurrence and development of periodontitis and diabetes mellitus.OBJECTIVE: To investigate the therapeutic effect of adiponectin injection on diabetic periodontitis model mice and its mechanism.METHODS: Sixty 8-week old Kunming mice(provided by Beijing Weitong Lihua Animal Experimental Technology Co., Ltd.) were randomly divided into three groups: control group, model group and adiponectin group. The mice in the model and adiponectin groups received intraperitoneal injection of streptozotin to establish a mouse model of diabetes mellitus, and then the periodontitis model was established by applying the combined bacterial fluid to the ligation site. The mice in the control and model groups received intraperitoneal injection of 1 mL normal saline once daily, while adiponectin group received intraperitoneal injection of 15 μg/kg adiponectin once daily, for 6 consecutive weeks. The three experimental groups of mice were treated for 6 weeks, after which physiological and biochemical indicators were tested.The levels of adiponectin in serum and periodontal tissue, the blood sugar and plasma insulin levels were then detected by ELISA. The depth of periodontal examination and the vertical absorption of alveolar bone were observed. The pathological changes of periodontal tissue were detected by hematoxylin-eosin staining. The changes of apoptosis level of gingival epithelial cells were detected by TUNEL method. The expression levels of RAGE and reactive oxygen species mRNA and protein were detected by RT-PCR and western blot assay.RESULTS AND CONCLUSION:(1) The levels of adiponectin in serum and periodontal tissue in the adiponectin group were significantly higher than those in the control and model groups(P < 0.05).(2) The blood sugar level in the adiponectin group was significantly lower than that in the model group(P < 0.05) and the serum insulin level was significantly increased(P < 0.05).(3) The depth of periodontal examination and vertical absorption of alveolar bone were decreased significantly in the adiponectin group compared with the model group, and the apoptotic rate of gingival epithelial cells was decreased significantly.(4) The expression levels of RAGE and reactive oxygen species mRNA and protein in the adiponectin group were significantly lower than those in the model group(P < 0.05).(5) These results suggest that adiponectin can improve the adiponectin level in periodontal tissue and plasma in the diabetic periodontitis model mice and can significantly improve the blood sugar status, and reduce the pathological changes of periodontal tissue and the apoptosis of periodontal cells possibly via regulating RAGE-ROS signaling pathway.
引文
[1]Oliver RC,Tervonen T.Diabetes--a risk factor for periodontitis in adults?. J Periodontol. 1994;65(5 Suppl):530-538.
[2]Daudt LD,Musskopf ML,Mendez M,et al.Association between metabolic syndrome and periodontitis:a systematic review and meta-analysis. Braz Oral Res. 2018;32:e35.
[3]康健,李新月.牙周炎与糖尿病关系的研究进展[J].医学综述,2017,23(18):3670-3674.
[4]赵博,王永兰.糖尿病牙周炎昆明系模型小鼠牙周组织中脂联素的表达[J].中国组织工程研究,2015,19(5):721-726.
[5]Kishida K,Funahashi T,Shimomura I.Adiponectin as a routine clinical biomarker. Best Pract Res Clin Endocrinol Metab.2014;28(1):119-130.
[6]Lim S,Quon MJ,Koh KK.Modulation of adiponectin as a potential therapeutic strategy. Atherosclerosis. 2014;233(2):721-728.
[7]权善花.2型糖尿病血清脂联素与骨密度变化及其相关因素的研究[D].延边:延边大学,2014.
[8]陈亮,魏民,刘玉杰.脂联素对骨性关节炎影响的研究进展[J].中华老年多器官疾病杂志,2017,16(8):620-623.
[9]刘努,贺涛,张馨艺,等.牙周基础治疗对2型糖尿病伴牙周炎患者疗效的研究进展[J].口腔疾病防治,2014,22(5):278-280.
[10]Taylor JJ,Preshaw PM,Lalla E.A review of the evidence for pathogenic mechanisms that may link periodontitis and diabetes. J Clin Periodontol. 2013;40 Suppl 14:S113-134.
[11]Antonoglou G,Knuuttila M,Nieminen P,et al.Serum osteoprotegerin and periodontal destruction in subjects with type 1 diabetes mellitus. J Clin Periodontol. 2013;40(8):765-770.
[12]Wang X,Yu S,Hu J P,et al.Streptozotocin-induced diabetes increases amyloid plaque deposition in AD transgenic mice through modulating AGEs/RAGE/NF-IoB pathway. Int J Neurosci 2014;124(8):601-608.
[13]Kay AM, Simpson CL, Stewart JA Jr.The Role of AGE/RAGE Signaling in Diabetes-Mediated Vascular Calcification. J Diabetes Res. 2016;2016:6809703
[14]Fisman EZ, Tenenbaum A. Adiponectin:a manifold therapeutic target for metabolic syndrome, diabetes, and coronary disease?. Cardiovasc Diabetol. 2014;13:103.
[15]Ghoshal K, Bhattacharyya M. Adiponectin:Probe of the molecular paradigm associating diabetes and obesity. World J Diabetes. 2015;6(1):151-166.
[16]李渊,房伟,王辉,等.脂联素在炎症疾病中的作用[J].中国医药指南,2016,14(22):32-33.
[17]李志高.牙周干预和老年2型糖尿病牙周炎患者脂联素以及炎症因子之间的相互作用机制的研究[J].中国医药科学, 2017,7(8):234-237.
[18]张静隆,朱迪,王贺林,等.脂联素抑制炎症小体NLRP3表达减轻高糖高脂诱导内皮细胞损伤[J].心脏杂志,2016,26(6):634-637.
[19]黄焱,郑卫东,修惠平,等.吡哆胺对糖尿病小鼠视皮质AGE-RAGE系统的影响[J].中国组织化学与细胞化学杂志, 2014,23(1):55-59.
[20]杨圣菊,孟国梁,顾黎雄,等.糖尿病小鼠皮肤组织中晚期糖基化终末产物的表达及其对胶原纤维的影响[J].中华皮肤科杂志, 2014,47(11):785-789.
[21]Fukami K,Yamagishi S,Okuda S.Role of AGEs-RAGE system in cardiovascular disease. Curr Pharm Des. 2014;20(14):2395-2402.
[22]Tesch G,Sourris K C,Summers S A,et al.Deletion of bone-marrow-derived receptor for AGEs(RAGE)improves renal function in an experimental mouse model of diabetes.Diabetologia.2014;57(9):1977-1985.
[23]Chen J,Jing J,Yu S,et al.Advanced glycation endproducts induce apoptosis of endothelial progenitor cells by activating receptor RAGE and NADPH oxidase/JNK signaling axis. Am J Transl Res. 2016;8(5):2169-2178.
[24]皇甫长梅.晚期糖基化终产物通过RAGE/ROS通路影响肾小管上皮细胞自噬-溶酶体系统[D].广州:暨南大学,2015.
[25]Ohtsu A,Shibutani Y,Seno K,et al.Advanced glycation end products and lipopolysaccharides stimulate interleukin-6secretion via the RAGE/TLR4-NF-κB-ROS pathways and resveratrol attenuates these inflammatory responses in mouse macrophages. Exp Ther Med. 2017;14(5):4363-4370.
[26]王敏华,邓淑丽,刘云西,等.二甲双胍对牙周致病菌内毒素所致饮食诱导肥胖小鼠高血糖的改善作用[J].中国临床药理学杂志,2018,34(8):945-948.
[27]刘海涛.慢性牙周炎患者基础治疗前后血清及龈沟液脂联素检测及临床意义研究[J].贵州医药,2018,42(2):232-233.
[28]颜明,牙周基础治疗对伴2型糖尿病的中、重度牙周炎患者脂联素和代谢水平的影响[J].中国医学创新,2018,15(10):99-102.
[29]张杰,梁伯贵,王柏灿,等.牙周治疗对2型糖尿病伴牙周炎患者炎症因子、脂联素及糖化血红蛋白的影响[J].现代实用医学, 2017,29(4):527-528.
[30]刘洋,王国芳,岳二丽,等.罗格列酮对牙周炎大鼠牙龈脂联素受体表达的影响[J].实用口腔医学杂志,2017,33(1):19-22.
[2]Daudt LD,Musskopf ML,Mendez M,et al.Association between metabolic syndrome and periodontitis:a systematic review and meta-analysis. Braz Oral Res. 2018;32:e35.
[3]康健,李新月.牙周炎与糖尿病关系的研究进展[J].医学综述,2017,23(18):3670-3674.
[4]赵博,王永兰.糖尿病牙周炎昆明系模型小鼠牙周组织中脂联素的表达[J].中国组织工程研究,2015,19(5):721-726.
[5]Kishida K,Funahashi T,Shimomura I.Adiponectin as a routine clinical biomarker. Best Pract Res Clin Endocrinol Metab.2014;28(1):119-130.
[6]Lim S,Quon MJ,Koh KK.Modulation of adiponectin as a potential therapeutic strategy. Atherosclerosis. 2014;233(2):721-728.
[7]权善花.2型糖尿病血清脂联素与骨密度变化及其相关因素的研究[D].延边:延边大学,2014.
[8]陈亮,魏民,刘玉杰.脂联素对骨性关节炎影响的研究进展[J].中华老年多器官疾病杂志,2017,16(8):620-623.
[9]刘努,贺涛,张馨艺,等.牙周基础治疗对2型糖尿病伴牙周炎患者疗效的研究进展[J].口腔疾病防治,2014,22(5):278-280.
[10]Taylor JJ,Preshaw PM,Lalla E.A review of the evidence for pathogenic mechanisms that may link periodontitis and diabetes. J Clin Periodontol. 2013;40 Suppl 14:S113-134.
[11]Antonoglou G,Knuuttila M,Nieminen P,et al.Serum osteoprotegerin and periodontal destruction in subjects with type 1 diabetes mellitus. J Clin Periodontol. 2013;40(8):765-770.
[12]Wang X,Yu S,Hu J P,et al.Streptozotocin-induced diabetes increases amyloid plaque deposition in AD transgenic mice through modulating AGEs/RAGE/NF-IoB pathway. Int J Neurosci 2014;124(8):601-608.
[13]Kay AM, Simpson CL, Stewart JA Jr.The Role of AGE/RAGE Signaling in Diabetes-Mediated Vascular Calcification. J Diabetes Res. 2016;2016:6809703
[14]Fisman EZ, Tenenbaum A. Adiponectin:a manifold therapeutic target for metabolic syndrome, diabetes, and coronary disease?. Cardiovasc Diabetol. 2014;13:103.
[15]Ghoshal K, Bhattacharyya M. Adiponectin:Probe of the molecular paradigm associating diabetes and obesity. World J Diabetes. 2015;6(1):151-166.
[16]李渊,房伟,王辉,等.脂联素在炎症疾病中的作用[J].中国医药指南,2016,14(22):32-33.
[17]李志高.牙周干预和老年2型糖尿病牙周炎患者脂联素以及炎症因子之间的相互作用机制的研究[J].中国医药科学, 2017,7(8):234-237.
[18]张静隆,朱迪,王贺林,等.脂联素抑制炎症小体NLRP3表达减轻高糖高脂诱导内皮细胞损伤[J].心脏杂志,2016,26(6):634-637.
[19]黄焱,郑卫东,修惠平,等.吡哆胺对糖尿病小鼠视皮质AGE-RAGE系统的影响[J].中国组织化学与细胞化学杂志, 2014,23(1):55-59.
[20]杨圣菊,孟国梁,顾黎雄,等.糖尿病小鼠皮肤组织中晚期糖基化终末产物的表达及其对胶原纤维的影响[J].中华皮肤科杂志, 2014,47(11):785-789.
[21]Fukami K,Yamagishi S,Okuda S.Role of AGEs-RAGE system in cardiovascular disease. Curr Pharm Des. 2014;20(14):2395-2402.
[22]Tesch G,Sourris K C,Summers S A,et al.Deletion of bone-marrow-derived receptor for AGEs(RAGE)improves renal function in an experimental mouse model of diabetes.Diabetologia.2014;57(9):1977-1985.
[23]Chen J,Jing J,Yu S,et al.Advanced glycation endproducts induce apoptosis of endothelial progenitor cells by activating receptor RAGE and NADPH oxidase/JNK signaling axis. Am J Transl Res. 2016;8(5):2169-2178.
[24]皇甫长梅.晚期糖基化终产物通过RAGE/ROS通路影响肾小管上皮细胞自噬-溶酶体系统[D].广州:暨南大学,2015.
[25]Ohtsu A,Shibutani Y,Seno K,et al.Advanced glycation end products and lipopolysaccharides stimulate interleukin-6secretion via the RAGE/TLR4-NF-κB-ROS pathways and resveratrol attenuates these inflammatory responses in mouse macrophages. Exp Ther Med. 2017;14(5):4363-4370.
[26]王敏华,邓淑丽,刘云西,等.二甲双胍对牙周致病菌内毒素所致饮食诱导肥胖小鼠高血糖的改善作用[J].中国临床药理学杂志,2018,34(8):945-948.
[27]刘海涛.慢性牙周炎患者基础治疗前后血清及龈沟液脂联素检测及临床意义研究[J].贵州医药,2018,42(2):232-233.
[28]颜明,牙周基础治疗对伴2型糖尿病的中、重度牙周炎患者脂联素和代谢水平的影响[J].中国医学创新,2018,15(10):99-102.
[29]张杰,梁伯贵,王柏灿,等.牙周治疗对2型糖尿病伴牙周炎患者炎症因子、脂联素及糖化血红蛋白的影响[J].现代实用医学, 2017,29(4):527-528.
[30]刘洋,王国芳,岳二丽,等.罗格列酮对牙周炎大鼠牙龈脂联素受体表达的影响[J].实用口腔医学杂志,2017,33(1):19-22.