Abstract:

Objective: The objective of this paper was to establish a mouse model with chronic periodontitis, to investigate the therapeutic effect of Puerarin on chronic periodontitis mice and its effect on periodontal tissue growth. Methods: The experimental mice were randomly divided into five groups: control group, model group, low-dose Puerarin group, medium- dose Puerarin group and high-dose Puerarin group respectively. Except for the control group, in the other groups, the orthodontic wire ligation method was used to establish chronic periodontitis models and 2 μL Porphyromonas gingivalis solution and 15 μL 1 mg/mL LPS normal saline were evenly injected into all periodontal tissues on the 3rd, 5th, 7th and 9th days after modeling. Four weeks after modeling, mice in low-, medium-, and high-dose Puerarin groups were given 200, 400 and 800 mg/kg by gavage. The control group and model group were given the same amount of CMC-Na solution. The administration solution was prepared once a day for 2 weeks. The activity status and performance of the experimental mice were observed. The concentrations of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β) and interleukin-6(IL-6) in peripheral serum were detected by enzyme-linked immunosorbent assay(ELISA). The expression levels of p38 mitogen activated protein kinase (p38MAPK) and phosphorylated p38 mitogen activated protein kinase (P-p38MAPK) in periodontal tissues of mice were detected by Western Blot test. Results: Four weeks after the establishment of chronic periodontitis model in mice, the appetite of the experimental mice decreased and the activity state was low. The symptoms of chronic periodontitis were manifested: gingiva was obviously swollen, the gingival tissue was soft, some of them ulcerated, gingival bleeding was obvious and spontaneous bleeding tendency was found. The results of enzyme-linked immunosorbent assay showed: versus the control group, the levels of TNF-α, IL-1β and IL-6 in the model group were significantly increased(P<0.01). Conversely, versus the model group, the levels of TNF-α, IL-1β and IL-6 in the middle and high dose Puerarin groups were significantly decreased(P<0.05). The results of Western Blot showed: compared with the control group, the protein expression of p38MAPK and P-p38MAPK in the gingival tissue of the model group was significantly increased; compared with the model group, the expression levels of p38MAPK and P-p38MAPK in the gingival tissue of the middle and high dose Puerarin groups were significantly decreased(P<0.01). Conclusion: The Puerarin can alleviate the symptoms of chronic periodontitis in mice, reduce the periodontal inflammatory reaction, and has a significant improvement and treatment effect on chronic periodontitis in mice. It may be concluded via p38MAPK signaling pathway, the Puerarins play such a therapeutic role.

Key words: chronic periodontitis, Puerarin, p38 mitogen activated protein kinase, mice

摘要：

目的： 建立慢性牙周炎小鼠模型，探讨葛根素对慢性牙周炎小鼠的治疗作用及其对牙周组织生长情况的影响，并探讨其作用机制。方法： 将实验小鼠随机分为对照组、模型组、葛根素低剂量组、葛根素中剂量组和葛根素高剂量组。除对照组外，其余各组均采用正畸钢丝结扎法建立慢性牙周炎模型，并分别在建模后的第3、5、7 和 9 天将2 μL 牙龈卟啉菌液和15 μL 1 mg/mL的脂多糖（lipopolysaccharide，LPS）0.9%氯化钠溶液均匀注射到全部牙周组织。建模4周后，对葛根素低、中、高剂量组小鼠进行灌胃给药，剂量分别为200、400、800 mg/kg，对照组和模型组小鼠给予等量的羧甲基纤维素钠（sodium carboxyl methyl cellulose，CMC-Na）溶液，给药溶液现用现配，每天1次，共治疗2周。观察各组实验小鼠的活动状态及表现，通过酶联免疫吸附试验检测各组小鼠外周血清中肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）、白细胞介素-1β（interleukin-1β，IL-1β）和白细胞介素-6（interleukin-6，IL-6）的浓度，通过Western Blot（蛋白质印迹法）实验检测小鼠牙周组织中p38丝裂原活化蛋白激酶（p38MAPK）和磷酸化的p38丝裂原活化蛋白激酶（P-p38MAPK）的表达水平。结果： 建立小鼠慢性牙周炎模型4周后，实验小鼠食欲减退，活动状态低迷，表现出慢性牙周炎症状：牙龈明显红肿、肥大，牙龈组织松软，部分出现溃疡现象，探诊出血明显，并有自发出血倾向。酶联免疫吸附试验检测结果显示：与对照组比较，模型组小鼠外周血清中的TNF-α、IL-1β和IL-6的水平显著升高（P<0.01）；与模型组比较，葛根素低、中、高剂量组小鼠外周血清中TNF-α、IL-1β和IL-6的水平降低（P<0.05）。Western Blot 检测结果显示：与对照组比较，模型组小鼠牙龈组织中p38MAPK和P-p38MAPK的蛋白表达水平显著升高；与模型组比较，葛根素低、中、高剂量组小鼠牙龈组织中p38MAPK和P-p38MAPK的蛋白表达水平显著降低（P<0.01）。结论： 葛根素能够缓解小鼠慢性牙周炎的症状，减轻牙周炎症反应，对于小鼠慢性牙周炎有显著的改善和治疗作用，且可能是通过p38MAPK信号通路发挥治疗作用。

关键词: 慢性牙周炎, 葛根素, p38丝裂原活化蛋白激酶, 小鼠