| Part 1 The effects of extracts of Propolis on the proliferation and differentiation of human periodontal ligament fibroblasts in vitroObjective:This study investigates the effects of propolis on the proliferation and osteogenic differentiation of human periodontal ligament fibroblasts(HPDLFs)in vitro,while to determine the optimal concentration of propolis extracts.Methods:Human periodontal ligament fibroblasts(HPDLFs)were cultured in vitro using the primary tissue explant method.Using the 4～6 passages for the experiment.The cells were divided into ten groups:eight groups were exposed to an extracts of propolis in various concentrations of 150μg/ml,200μg/ml,250μg /ml,300μg/ml,350μg/ml and 400μg/ml,450μg/ml,500μg/ml respectively;one group was exposed to 100ng/ml of basic fibroblast growth factor(bFGF) as positive control;the final group was exposed to DMEM containing 1%fetal bovine serum as negative control.After exposure to the various mediums, proliferation of HPDLFs was measured with the MTT colorimeteric assay,alkaline phosphatase activity was determined with the enzyme kinetic method,osteocalcin was determined with radio-immunological method.Finally using one-way ANOVA to analyze the data.Result:The propolis extracts at concentrations of 250ug/ml,300 ug/ml,350 ug/ml, 400 ug/ml,450 ug/ml and 500 ug/ml can promoted proliferation of HPDLFs,and have the significance(P＜0.05).Also the 450ug/ml concentration is better than the positive control(P＜0.05);Contrast with control group,250μg/ml,300μg/ml,350μg/ml,400μg/ml concentration of propolis extracts can accelerate the expression of ALP(P＜0.05);250μg/ml,300μg/ml,350μg/ml level of propolis extract can also improve the expression of OC(P＜0.05).Conclusion:Under these experimental conditions,high concentration of extracts of propolis were able to significantly increase HPDLFs proliferation and osteogenic differentiation activity.It may be a useful active agent in topical periodontal preparations to promote the reparation and growth of periodontal ligament in the treatment of periodontal disease. Part 2 Experiment of security of the propolis extractsExperiment 1 The oral mucous membrane irritation test of the propolis extractsObjective:The purpose of this study was to estimate the potential oral mucous membrane irritation of the propolis extracts,and to provide the evidence for clinical application.Methods:Using white rabbits with its average weight 2.1kg for the experimental animals.Put the small cotton wool soaked with propolis extracts on the palate mucous on the right side for 15 minutes under anesthesia,the other side of the palate mucous placed fresh plaster as the negative control.The test position were observed after removing the wool for 1h,24h,and the animals were sacrificed and their mucous surgically removed and processed for histological evaluation.Result:The oral mucous membrane contacting the propolis extracts and the negative control materials showed no engorgement,tumidness,erosion,haemorrhage, or ulceration.The histopathological findings were also normal.There was no significant difference between the experimental sides and the negative control sides.Conclusion:There was no oral mucous membrane irritation of the propolis extracts.Experiment 2 The short-term systemic toxicity test(oral route) of the extracts of propolisObjective:The goal of this study was to estimate the potential systemic toxicity of the propolis extracts,and to provide the evidence for clinical application.Methods:Twenty 130～150g Sprague-Dawley rats,half of which were male,were randomly divided into the negative control group and the experimental group,5 of 10 rats in each group were male.The negative control group was taken the normal saline, and the experimental group was taken the extracts of propolis,1000mg/kg per day for 7 days.The rats were observed for 7 days,while their weight and the food consumption were recorded.The food use ratio,the weight relatively increasing ratio were taken to statistics.Results:During the 14 days,none of the rats showed the clinical toxicity symptom. There was no statistically significant difference about the weight,the weight relatively increasing ratio between the experimental group and the negative control group(P＞0.05).The food use ratio of the experimental group was approximately consistent with the control group.Conclusion:There was no short-term systemic toxicity of the propolis extractsExperiment 3 The skin sensitization test of the extracts of propolisObjective:The aim of this study was to estimate the potential sensitization of the propolis extracts,and to provide the evidence for clinical application.Methods:Fifteen 300～500g albino guinea pigs,were randomly divided into the experimental group(10 animals) and the negative control group(5 animals).Using the normal saline for the negative control.Respectively injected normal saline maceration extract of the extract propolis,Freund complete adjuvant(FCA),the emulsion of the extract propolis and FCA.After 3 stages(intradermal induction,local inducement,stimulating contact),the result of stimulation were observed in 24 and 48 hours. Results:24h and 48h after stimulated to contact,7 albino guinea pigs of experimental group showed very slight erythema,and no erythema or oedema for the other 3 albino guinea pigs.The negative control showed no sensitization reaction.Conclusion:There was sensitization of the extraction of the extracts of propolis...
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