牙周炎对肥胖小鼠肾脏病变的影响及其机制探讨

发布时间：2018-06-08 15:19

本文选题：肥胖 + 牙周炎　；参考：《南方医科大学》2017年博士论文

【摘要】：许多研究都已分别证实肥胖与慢性肾病具有相关性、牙周炎与慢性肾病具有相关关系。但尚缺乏在肥胖这种代谢异常状态下牙周病对肾脏影响的直接证据。本研究使用牙周炎复合肥胖小鼠模型,旨在探讨在肥胖状态下,牙周炎是否对肾脏病变的产生存在影响及其可能的机制。为牙周炎是否影响肾脏疾病的发生寻找直接的证据。为临床多学科联合诊断,治疗和预防提供依据。本文内容共分为三部分:一牙周炎对肥胖小鼠肾脏损害的影响目的:建立饮食诱导型肥胖伴实验性牙周炎的小鼠模型,研究牙周炎对肥胖小鼠肾脏功能及肾脏病损是否存在影响。方法:275只小鼠随机分为高脂饲料组(HF组)和低脂饲料组(LF组)两组。饲料诱导时间分为8周、16周、30周三个时间点。每个饲料诱导时间结束时,再将每个饲料组分为牙周炎组(P组)和非牙周炎组(对照组,C组),P组采用丝线结扎法诱导实验性牙周炎,C组做假性结扎对照处理。处死小鼠取材。取30周小鼠,检测血清肾功能指标血清总蛋白、白蛋白、血尿素氮、肌酐,并对组织切片行HE染色及三种特殊染色(MASSON染色、PAS染色、改良六胺银染色)。结果:HF组比LF组血清总蛋白、白蛋白水平均有显著下降。而HFP和HFC两组间无明显差异,同样,LF组有无牙周炎干预(LFP和LFC),二者之间也没有明显差异。HFP组的血清肌酐明显高于LFP和LFC,而HFC、LFP和LFC三组之间无明显差异,HFP和HFC之间也无明显差异。HF组的血清尿素氮数值偏高,但四组之间没有统计学差异。肾组织病理改变可见LFP组织学特征与LFC接近,表现为正常肾脏的组织学特点。HFP和HFC组小鼠可见肾小球局灶性毛细血管袢消失,系膜细胞增生,肾小球基底膜及球周纤维增生。肾小管上皮细胞玻璃样或空泡变性,管间质纤维增生。以上病理特征在HFP组与HFC组小鼠情况近似,但HFP组的肾小管间质纤维增生更为严重。结论:本研究成功建立了不同程度的饮食诱导型肥胖复合牙周炎的小鼠模型。正常小鼠患单纯牙周病并不会导致肾脏损害,肥胖才是肾脏损害的主要因素。在DIO30周这种严重肥胖的情况下,牙周炎可能会加剧肾小管间质纤维化的病变。二不同肥胖/牙周炎症状态下对肾脏损害的差异目的:利用DI08周、16周、30周及牙周感染5d、10d小鼠,探索在不同肥胖状态下及不同牙周炎症状态下,牙周感染对肥胖小鼠肾脏损害的影响。方法:对所有分组小鼠组织切片行HE染色及三种特殊染色(MASSON染色、PAS染色、改良六胺银染色)。结果:从DI08周至16周至30周,肥胖小鼠肾小球均有系膜细胞增生和毛细血管基底膜增厚,严重程度近似,但受累肾小球数目及范围增加,肾小管上皮细胞均有空泡变性,范围逐渐扩大。肾小球周纤维增生逐渐加重,肾小管间质纤维化程度加重、范围加大。DI016周小鼠牙周感染10d与5d肾脏病变无明显区别,30周小鼠牙周感染10d比5d肾小管间质纤维化程度更加严重。结论:随着肥胖状态增加,肾脏损害加重。可能在DIO30周这种重度肥胖状态下,小鼠牙周感染时间越长,对肾小管间质纤维化的影响越大。三牙周炎影响肾脏损害的可能机制目的:检测各组小鼠肾脏组织中TGF-β1,MMP-2,TIMP-1蛋白和基因转录水平的表达差异,进而探讨肥胖状态下时慢性牙周炎影响肾脏损害可能的病理机制。方法:采用免疫组化检测小鼠肾脏组织中TGF-β1,MMP-2,TIMP-1蛋白的表达,进行半定量分析,另采用实时定量PCR技术进行TGF-β1,MMP-2,TIMP-1 mRNA基因表达差异分析。结果:免疫组化显示,HFP、HFC、LFP、LFC四组肾脏组织中TGF-β1、TIMP1蛋白有统计学差异,每两组均差异明显,HFP组表达量最高。MMP2表达与之相反,LF两组表达较高。实时定量PCR结果显示,肥胖是主要影响因素,HFP、HFC、LFP、LFC 四组之间 TGF-β1、MMP2、TIMP1 mRNA 表达有统计学差异。结论:在肥胖状态下,牙周干预促进了肾脏组织TGF-β1、TIMP-1这两个促进纤维化抑制降解蛋白的表达,同时抑制了降解EDM的主要水解酶MMP-2的表达,这可能是牙周炎影响肥胖小鼠肾脏病变的机制之一。
[Abstract]:Many studies have confirmed the correlation between obesity and chronic kidney disease. Periodontitis is associated with chronic kidney disease. But there is no direct evidence of the impact of periodontitis on the metabolic disorder of obesity. This study uses periodontitis compound obesity mice model to explore whether periodontitis is the kidney in the condition of obesity. The existence of the influence and possible mechanisms of the formation of visceral lesions. It provides a direct evidence for the effect of periodontitis on the occurrence of renal diseases. It provides a basis for clinical multidisciplinary combined diagnosis, treatment and prevention. The content of this article is divided into three parts: the effect of periodontitis on kidney damage in obese mice: the establishment of diet induced obesity with real disease The effect of periodontitis on renal function and renal impairment in obese mice was studied. Methods: 275 mice were randomly divided into high fat diet group (HF group) and low fat diet group (group LF). The feed induction time was divided into 8 weeks, 16 weeks, 30 Wednesday time points. Each feed induction time was over, and each feed was then fed. The material group was divided into periodontitis group (group P) and non periodontitis group (control group, group C). Group P was used to induce experimental periodontitis by wire ligation, and group C was treated with pseudoligature. The mice were sacrificed for 30 weeks. The serum total protein, albumin, blood urea nitrogen and creatinine were detected in the serum kidney function index, and the tissue sections were stained with HE and three special types. Staining (MASSON staining, PAS staining, improved six amine silver staining). Results: the level of serum total protein and albumin decreased significantly in group HF than in group LF, but there was no significant difference between groups of HFP and HFC. Similarly, there was no periodontitis intervention (LFP and LFC) in the LF group (LFP and LFC), and the serum creatinine in the two group was significantly higher than that of LFP and LFC. There was no significant difference between the three groups. There was no significant difference in the serum urea nitrogen between the HFP and the HFC groups, but there was no statistical difference between the four groups. The histopathological changes of the renal tissue showed that the histological features of the LFP were close to LFC, and the histological features of the normal kidneys were found to disappear in the glomerular focal capillary loops in the group.HFP and the HFC group. Mesangial cell proliferation, glomerular basement membrane and peripheral fibrous hyperplasia. Renal tubular epithelial cells were glassy or vacuolar degeneration, tubulointerstitial fibrous hyperplasia. The above pathological features were similar in group HFP to HFC mice, but in group HFP, tubulointerstitial fiber hyperplasia was more serious. Conclusion: This study successfully established a different degree of dietary induced fertilizer. A mouse model of fat complex periodontitis. Normal periodontitis does not cause renal damage in normal mice. Obesity is the main factor in renal damage. Periodontitis may exacerbate renal tubulointerstitial fibrosis in DIO30 weeks. Two different obesity / periodontitis differences in renal damage Using DI08 weeks, 16 weeks, 30 weeks and periodontal infection 5D, 10d mice, to explore the effect of periodontal infection on kidney damage in obese mice under different obesity and periodontitis. Methods: HE staining and three special staining (MASSON staining, PAS staining, improved six amine silver staining) were carried out in all the tissue sections of the group mice. Results: from DI08 From 16 weeks to 30 weeks, the glomeruli of the obese mice had mesangial cell proliferation and capillary basement membrane thickening. The severity of the glomeruli was similar, but the number and scope of the glomeruli were increased. The renal tubular epithelial cells were vacuolated and expanded gradually. The proliferation of peripheral fibrous fibrosis was gradually aggravated and the degree of renal tubulointerstitial fibrosis was aggravated. There was no obvious difference between 10d and 5D in the periodontal infection of mice in the large.DI016 week. The periodontal infection of the mice in 30 weeks was more severe than that of the 5D renal tubule. Conclusion: with the increase of obesity, the renal damage is aggravated. The longer the periodontal infection time is, the longer the periodontal infection of the mice is, the fibrosis of the renal tubules. The possible mechanism of the effect of three periodontitis on renal damage: the difference in the expression of TGF- beta 1, MMP-2, TIMP-1 protein and gene transcription level in kidney tissues of mice was detected, and the possible pathological mechanism of chronic periodontitis in the condition of obesity was explored. Methods: immunohistochemistry was used to detect T in the kidney tissues of mice. GF- beta 1, MMP-2, TIMP-1 protein expression, semi quantitative analysis, and real-time quantitative PCR technology for TGF- beta 1, MMP-2, TIMP-1 mRNA gene expression difference analysis. Results: immunohistochemistry showed that HFP, HFC, LFP, LFC four groups of renal tissue beta 1, every two groups are different, the highest expression of the two groups On the contrary, the expression of LF two groups was higher. Real-time quantitative PCR results showed that obesity was the main influencing factor. The expression of TGF- beta 1, MMP2, TIMP1 mRNA was statistically different between groups of HFP, HFC, LFP and LFC. Conclusion: in the obese state, periodontal intervention promoted the TGF- beta 1 of the kidney tissue, TIMP-1 these two promote the expression of fibrotic inhibition degrading protein. It inhibited the expression of MMP-2, a major hydrolytic enzyme that degrades EDM, which may be one of the mechanisms of periodontitis affecting the kidney of obese mice.
【学位授予单位】：南方医科大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R781.4;R692

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