The Effects Of Cementum Preservation On The Differentiation Of Human Periodontal Ligament Cells

Objective:Cementum that coveraged on the superficial of dental root was lamellar mineralization connective tissue.Investigation indicated that cementum basilaris substantia contain cementum attachment protein(CAP) and attachment protein-23 (CP-23).They were known as cementum specificity molecule to possess many biology effectiven.Therefor the effect of cementum promotion periodontal tissue regeneration was more and more to be think highly.In this experiment,we studied the influence of remaining cementum on the accretion and differentiation of periodontal ligament cells and discussed the feasibility of remaining cementum in parodontosis treatment.Materials and methods:1.HPDLCs were isolated from the premolars extracted due to orthodontic therapy and cultured.2.Root slices were made from the periodontal ligament denuded premolars.In group-Cementum(PDLCs+C),the cementum was preserved,while in group-Dentin (PDLCs+D),the cementum was scaled completely from the root surface and the dentin was exposed.3.The second passage HPDLCs were inoculated on the two groups of root slices.After a 7-day co-culture,the cells growing on the root slices were harvested .Total cellular RNA of the two groups was extracted by using Ttrizol reagent respectively.Real-time reverse transcriptase polymerase chain reaction(RT-PCR) was performed for the measurement of mRNA expression of cementum attachment protein (CAP) and cementum protein 23(CP-23).β-action was used as an internal control. The amount of mRNA was calculated for each sample from the standard curve using the instrument software.Results were expressed as the ratio of the amount of tested mRNA divided byβ-action mRNA.4.Then collected the second generation.Cultivated the cells seven days on dentine or cementum slices in vitro respectively,and then putthe cells-slice complexes inside semipermeable membranes.Then the complexes were imbed inthe bodies of athymic mice.Executed athymic mice and extracted the slices after six or eightweeks. Fixed,decalcified the tissue and carried out HE dyeing to observe the disparity of newformative hard tissue in dentine or cementum slices.Transmission electron microscope(TEM) was used to observe ultramicrostructure of organiza.Results:1.For CAP mRNA expression,the ratio is 0.00375 for PDLCs+C and 0.00325 for PDLCs+D.For CP-23 mRNA expression,the ratio is 0.000073 for PDLCs+C and 0.000000662 for PDLCs+D.Statistical analysis indicated that expression difference of PDLCs+C and PDLCs+D had statistical significance.2.In HE dyeing image that sixteen of twenty cementum slices formed cementum basilaris substantia on the surface cementum slices surface,but there was not structure formation on the dentine slices surface,and only there was fibroplasia.3.Transmission electron microscope(TEM) of dentine or cementum slices which were extracted from the bodies of athymic mice,cells were cohered on the surface of new formative hard tissue,entoblast anachromasis,intracytoplasm of cells was bound to rough endoplasmic reticulum.There were evident circumsciption and conjunction close-up between new formative hard tissue and inhere cementum. Conclusions:Our study suggests that the pre-existent cementum maybe promote the HPDLCs to differentiate into cementoblasts.Experiments can suggests that the pre-existent cementum maybe promote the regeneration in periodontal tissue.