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背景:牙周组织的修复再生取决于牙周膜细胞的数量和增殖分化能力。牙周膜细胞具有多向的分化潜能,可分化成成牙骨质细胞、成骨细胞和成纤维细胞,形成牙骨质、牙槽骨和牙周膜,实现牙周组织再生。目的:观察体外培养人牙周膜细胞增殖和分化过程中传统中药双黄补水提液对其功能的影响。设计:观察性实验。单位:无锡市第三人民医院中心实验室。材料:牙周膜组织(由健康青少年患者因矫正畸形门诊就诊时自愿提供);黄连、黄岑、骨碎补(中国药品检定所提供)。方法:实验于2003-07/10在无锡市第三人民医院中心实验室完成。分别将粉碎的黄连、黄岑、骨碎补和蒸馏水1∶10(m∶V)混合,沸水回流5h。收集提取液,粗滤后残渣再次沸水回流3h。合并2次提取液,旋转蒸发浓缩,所得中药水提液浓度为3kg/L。分黄连组、黄芩组、骨碎补组、黄连加黄岑组、黄连加骨碎补组、黄岑加骨碎补组、双黄补组和对照组8组进行实验。通过体外培养人牙周膜细胞,应用双黄补提取液作为辅助因子,采用MTT比色法测定细胞增殖情况,羟脯氨酸法测定胶原蛋白占总蛋白比值。主要观察指标:牙周膜细胞增殖的吸光度值及胶原蛋白占总蛋白的比值。结果:①牙周膜细胞增殖的测定结果:除黄连组外其他各组均明显促进人牙周膜细胞增殖,与对照组比较,差异明显(P<0.05)。双黄补组细胞的吸光度值增大最为明显。随着作用时间的延长,细胞的吸光度值逐渐增大,在第5天达到最大。不同时间各组间比较,差异有显著性意义(P<0.05)。②各组牙周膜细胞胶原蛋白占总蛋白的比值:除黄连组外其他各组均明显促进比值增大,与对照组比较,差异明显(P<0.05)。双黄补组比值增大最为明显。随着作用时间的延长,比值逐渐增大,在第5天达到最大。不同时间各组间比较,差异有显著性意义(P<0.05)。结论:双黄补水提液可明显促进人牙周膜细胞增殖,明显提高胶原蛋白在总蛋白中的比值,可作为人牙周膜细胞增殖的理想的中药辅助因子。
BACKGROUND: The regeneration of periodontal tissue depends on the number of periodontal ligament cells and their ability to proliferate and differentiate. Periodontal ligament cells have multi-directional differentiation potential and can differentiate into cementoblasts, osteoblasts, and fibroblasts, forming cementum, alveolar bone, and periodontal ligament to achieve periodontal tissue regeneration. OBJECTIVE: To observe the effect of Shuanghuang Replenishing Water Extract on the function of cultured human periodontal ligament cells in vitro. Design: Observational experiment. Unit: Central Laboratory of the Third People’s Hospital of Wuxi City. MATERIALS: Periodontal ligament tissue (provided voluntarily by healthy adolescents during a correctional outpatient visit); Rhizoma Coptidis, Astragalus membranaceus, Rhizoma Drynaria (recruited by the China National Drug Authentication Institute). METHODS: The experiment was performed at the Central Laboratory of the Third People’s Hospital of Wuxi City from July to October 2003. The crushed Rhizoma Coptidis, Astragalus membranaceus, Rhizoma Drynariae, and distilled water were mixed 1:10 (m:V) and refluxed in boiling water for 5 h. The extract was collected and the crude residue was boiled again under reflux for 3 h. The extracts were combined twice and concentrated by rotary evaporation. The concentration of the aqueous extract of the Chinese medicine was 3 kg/L. The experimental group was divided into 8 groups: Huanglian group, Huangqi group, Guliao group, Huanglian plus Huangqi group, Huanglian plus Guliao group, Huangqi plus Gugebu group, Shuanghuangbu group and control group. By in vitro culture of human periodontal ligament cells, Shuanghuangbu extract was used as a cofactor, cell proliferation was measured by MTT colorimetry, and the ratio of collagen to total protein was determined by hydroxyproline method. MAIN OUTCOME MEASURES: Absorbance values of periodontal ligament cell proliferation and the ratio of collagen to total protein. RESULTS: 1 The results of the periodontal ligament cell proliferation assay showed that all groups except the Huanglian group significantly promoted the proliferation of human periodontal ligament cells, which was significantly different from that of the control group (P<0.05). The absorbance of cells in Shuanghuangbu group increased most obviously. With the prolongation of the action time, the absorbance of the cells gradually increased and reached the maximum on the fifth day. There was a significant difference between the groups at different times (P<0.05). 2 The ratio of collagen in total periodontal ligament cells in all groups: In addition to the Huanglian group, the proportions of all promoted obviously increased, compared with the control group, the difference was significant (P<0.05). The increase of the ratio of Shuanghuangbu group is most obvious. With the extension of the action time, the ratio gradually increased and reached the maximum on the fifth day. There was a significant difference between the groups at different times (P<0.05). Conclusion: Shuanghuangshui water extract can significantly promote the proliferation of human periodontal ligament cells and significantly increase the ratio of collagen protein in total protein. It can be used as an ideal auxiliary factor for human periodontal ligament cell proliferation.