Effects Of Osteogenic Growth Peptide And Insulin-like Growth Factor-I On Biological Function In Cultured Human Periodontal Ligament Cell

Objective:This experiment is by the means of culturing in vitro human periodontal ligament cells,human PDLCs.Then we will use osteogenic growth peptide,OGP and insulin-like growth factor-I,IGF-1 alone or in pairs on human PDLCs , observe the human PDLCs proliferation ,changes of Alkaline phosphatase activities in the cells and changes of totle protein content to discuss the effect of two kinds of growth factor used alone or in pairs on human PDLCs which was cultured in vitro , and it will provide Theoretical basis in the use of clinical cure of periodontal disease.Methods:1. the culture in vitro and test of human PDLCs:Take the freash dentes premolares between 12 to 18 years which were needed to remove because of orthodontic needs ,and make sure that they must be Periodontal health and without caries.Scrape take one third of Periodontal tissue of fang with blade , foster human PDLCs with Tissue mass method .Take the third generation cells and use sABC method to take the Immunohistochemical stains of Vimentin and Kerati , which will tell us the origin of the cells.2.Use one of the OGP or IGF-1 on human PDLCs at a time ,after three days fostering use MTT to observe the proliferative activity of human PDLCs, chose each of the Biggest powerfully concentration and the minimum powerfully concentration of the effection of OGP and IGF-1 worked on human PDLCs after statistic analysis.3.Let OGP and IGF-1 in the two conditions in 2,work on human PDLCs alone then in pair .Use MTT to test the cell proliferation , assay Absorbency value(A),and take comparison among groups on the first , third , sixth day;Use alkaline phosphatase detection kit to test the activity of alkaline phosphatase ,assay Absorbency value(A) , and take comparison among groups;Use Coomassie brilliant blue colorimetric method to test content of total protein in cells , assay Absorbency value(A), and take comparison among groups.Result:1. When using tissue mass method on the seventh to ninth day, some cells freed themselvs from organization, Immunohistochemical stains tells that the Vimentin are positive and the Kerati are negative.2.On the third day ,Absorbency value ofdifferent concentration experimental group cells of OGP are very different from control group(P<0.05).Biggest powerfully concentration is 10-9mol/L,Minimum powerfully concentration is 10-11mol/L.3.On the third day ,Absorbency value ofdifferent concentration experimental group cells of IGF-1 are very different from control group(P<0.05).Biggest powerfully concentration is 100ng/ml,Minimum powerfully concentration is 1ng/ml.4.When OGP and IGF-1 are worked in pairs on human PDLCs,Promote proliferation role are very different from control group(P<0.05), and also very different from the conditiong when they work aolne(P<0.05), Promote proliferation works from the first day to the sixth day .5.when OGP and IGF-1 are worked in pairs on human PDLCs, in the cell supernatant or in the Cell Lysis Solution ,the Absorbency value(A) of activities of ALP are very different from control group(P<0.05).and also very different from the conditiong when they work aolne(P<0.05)The promote of ALP active strengthening works from the first day to the sixth day.6.When OGP and IGF-1 are worked in pairs on human PDLCs,the Absorbency value(A) of total protein content are very different from control group(P<0.05),and also very different from the conditiong when they work aolne(P<0.05) .The promote of total protein content works from the first day to the sixth day.Conclusion:1.The experimental results proves that when OGP is worked alone on human PDLCs , it brings the proliferation of human PDLCs, strengthen the activities of ALP in cells and increase the Total protein content.2.The experimental results proves that when IGF-1 is worked alone on human PDLCs , it brings the proliferation of human PDLCs, strengthen the activities of ALP in cells and increase the Total protein content. 3.The experimental results proves that when OGP and IGF-1 are combined on human PDLCs , it brings the proliferation of human PDLCs, strengthen the activities of ALP in cells and increase the Total protein content.The two growth factors combined with a synergistic effect.