The Influence And Mechanism Of Actived δ Opium Receptor To Apoptosis Of The Rat Osteoblast Which Has Been Induced By Starvative Blood Serum

ObjectiveThrough the rat osteoblast has been cultured in vitro to study the influenceand mechanism of actived δ opium receptor to apoptosis of the rat osteoblastwhich has been induced by starvative blood serum.MethodsIn the condition of asepsis,take out of the skull of24h aged rat,culturingthe primary generation osteoblast out of body.Using microscope to observe thecell modality and multiplication.After AKP coloration,using Gomori amendedBJP method to appraise osteoblast. through replacing to the culture mediumwithout calf serum,come into being the serum starvative condition,to inducethe osteoblast apoptosis model;The whole experiment divided into5groups,including control group: cultured osteoblast out of body;Model group:the osteoblast apoptosis model; DADLE group:giving1μmol·L^-1DADLE tocultivating48h;DADLE+NAL group:in the starvative condition,giving1μmol·L^-1DADLE and10μmol·L^-1Naltrindole to culture48h;DADLE+GFX group:in thestarvative condition,giving1μmol·L^-1DADLE and10μmol·L^-1GF109203X toculture48h.the survival rate of the cell has been determined by the MTTMethod. The cell cycle has been analyzed by flow cytometry（FCM）. The rateof cellular apoptosis has been determined by the paired labeled technique of Annexin V-FITC/PI. The expression of PKC and Caspase-3have beendetermined by the Western blot.ResultsThe generation of osteoblasts was separated to Cranial cover bone ofNewborn rat, morphology is Small round sphere, visible bright Cell membrane,black blob-shaped is the cell nuclei inside that. The osteoblast has beencultured3-4days, cell fusion a single. After AKP coloration, using Gomoriamended BJP method to appraise osteoblast, Around the cell membrane isvisible dark brown and subtle grain. Culturing the primary generationosteoblast48h, showed significant apoptosis phenomenon with cell activitydecreasing and the apoptosis rat of the cell reducing,percentage of S+G2+Mdecreasing. Meanwhile,The expression of Caspase-3increases and Theexpression of PKC reduces. Compared with control group,it has a significantmean（P﹤0.01）.Apoptosis of osteoblast which has been induced by starvativeblood serum has been restrain significantly. So the survival rate of the cellincreases and the apoptosis rat of the cell reduces. So the survival rate of thecell increases and the apoptosis rat of the cell reduces. The percentage of S+G2+M increases. The expression of Caspase-3reduces and The expression ofPKC increaces. Compared with control group,it has a significant mean（P﹤0.01）,but this phenomenon can be restrained by Naltrindole and GF109203X.DADLE+NAL group and DADLE+GFX group showed that the survival rate ofthe cell reduces and the apoptosis rat of the cell increases,the percentage ofS+G2+M reduces,The expression of Caspase-3reduces,which compared withDADLE group has significant mean（P﹤0.01）,compared with Model group hasno significant mean.ConclusionsThe apoptosis of osteoblast which has been induced by starvative bloodserum can be protected by δ opium receptor which has been activated byDADLE, and this protection has been achieved by the PKC way.