本研究的目的是评估槲皮素对牙本质侵蚀和磨损的原位/体内作用。
将人牙本质块(2×2×2mm)嵌入并分配给6组:75μg/mL,150μg/mL和300μg/mL槲皮素(Q75,Q150,Q300);120μg/mL氯己定(CHX,阳性对照);和去离子水和乙醇(阴性对照)。用相应的溶液处理样品2分钟,然后如下进行原位/体内侵蚀/研磨挑战7天:每天4次体内侵蚀,然后在每天的第一次和最后一次侵蚀挑战后进行体内牙刷磨损。牙本质损失通过轮廓术评估。另一个牙本质样本用于通过跟踪其特征拉曼峰的空间分布来评估槲皮素进入牙本质的穿透深度。此外,使用牙本质块(7×1.7×0.7mm)通过原位酶谱检测槲皮素对牙本质衍生的基质金属蛋白酶(MMP)抑制的影响,并计算抑制百分比(%)。此外,潜在的胶原交联相互作用与槲皮素通过拉曼光谱检测,交联度是用九三酮测定法测定的。通过极限微拉伸强度测试(μUTS),使用完全脱矿质的牙本质束(0.5×0.5×10mm)来评估槲皮素对牙本质胶原纤维机械性能的影响。数据采用单因素方差分析和Tukey检验(α=0.05)。
与阴性对照相比,所有治疗方案均显著减少牙本质损失.Q150和Q300的牙本质损失明显少于CHX(均P<0.05)。槲皮素的量随着牙本质深度的增加而减少,最大穿透深度约为25-30µm。原位酶谱显示槲皮素显著抑制牙本质来源的MMPs的活性。Q75和Q150的抑制率明显低于CHX(均P<0.05),但Q300和CHX之间没有发现显着差异(P=0.58)。胶原蛋白与槲皮素的交联相互作用主要涉及氢键,交联度以浓度依赖性方式增加。与对照处理相比,槲皮素处理后的脱矿质牙本质束的μUTS值在统计学上显著增加(所有P<0.05)。
这项研究提供了第一个直接证据,表明槲皮素可以渗透约25-30μm进入牙本质,并通过抑制牙本质衍生的MMP活性以及去矿质有机基质的交联胶原蛋白来进一步防止牙本质侵蚀和磨损。
The aim of the present
study was to evaluate the in situ/in vivo effect of quercetin on dentin erosion and abrasion.
Human dentin blocks (2 × 2 × 2 mm) were embedded and assigned to 6 groups: 75 μg/mL, 150 μg/mL and 300 μg/mL quercetin (Q75, Q150, Q300); 120 μg/mL chlorhexidine (CHX, positive control); and deionized water and ethanol (the negative controls). The specimens were treated with the respective solutions for 2 min and then subjected to in situ/in vivo erosive/abrasive challenge for 7 d as follows: in vivo erosion 4 times a day and then in vivo toothbrush abrasion after the first and last erosive challenges of each day. Dentin loss was assessed by profilometry. An additional dentin specimen was used to evaluate the penetration depth of quercetin into dentin by tracking the spatial distribution of its characteristic Raman peak. Moreover, dentin blocks (7 × 1.7 × 0.7 mm) were used to detect the impact of quercetin on dentin-derived matrix metalloproteinase (MMP) inhibition by in situ
zymography, and the inhibition percentage (%) was calculated. Additionally, the potential collagen crosslinking interactions with quercetin were detected by Raman spectroscopy, and the crosslinking degree was determined with a ninhydrin assay. Fully demineralized dentin beams (0.5 × 0.5 × 10 mm) were used to evaluate the impact of quercetin on the mechanical properties of dentin collagen fibre by the ultimate micro-tensile strength test (μUTS). The data were analysed by one-way analysis of variance and Tukey\'s test (α = 0.05).
Compared to the negative controls, all treatment solutions significantly reduced dentin loss. The dentin loss of Q150 and Q300 was significantly less than that of CHX (all P < 0.05). The amount of quercetin decreased with increasing dentin depth, and the maximum penetration depth was approximately 25-30 µm. In situ
zymography showed that quercetin significantly inhibited the activities of dentin-derived MMPs. The inhibitory percentages of Q75 and Q150 were significantly lower than that of CHX (all P < 0.05), but no significant difference was found between Q300 and CHX (P = 0.58). The collagen crosslinking interactions with quercetin primarily involved hydrogen bonding and the degree of crosslinking increased in a concentration-dependent manner. Statistically significant increases in μUTS values were observed for demineralized dentin beams after quercetin treatment compared with those of the control treatments (all P < 0.05).
This
study provides the first direct evidence that quercetin could penetrate approximately 25-30 µm into dentin and further prevent dentin erosion and abrasion by inhibiting dentin-derived MMP activity as well as crosslinking collagen of the demineralized organic matrix.