Experiment Study On Effects Of Strorage Time In Vitro To The Healing Of NVBG With Implants

Mandible segmental defects,caused from tumors,traumas or inflammation,affect patients' life quality seriously.Transplantation of Nonvascularized bone graft with immediate implants was applied to rehabilitate mandible successfully, the patients' life quality was improved obviously.But there were still arguments about the success of this operation style,some scholars opposed this style.They still insisted that nonvascularised bone graft in vivo undergo long creeping substitution and healing process was too slow accompanying with critical bone absorption.So the success of the operation style lack of theorical inevitability and regulation.This article aimed to study the effects of storage time in vitro on the healing of nonvascularized bone and have a primary probe of its inevitability and regulation on the basis of experiments.1 Effects of different storage time on nonvascularized bone graft's vitality in vitroObjective: To study bone cell vitality and proliferation ability of nonvasucularized bone graft under different storage time in vitro stored in different preserving medium.Methods: Block bone grafts were acquired from both sides of canine ilia and a dental implant was implanted in the experimental iliac graft.Grafts were stored separately in the air ,natural salt solution and DMEM contained 10% FBS.Small pieces of bone about 3mm×3mm size were cut off respectively from the rim and central(or around the implant) of the block graft at the 20 minute,40th minute,60 minute, 80 minute, 100 minute, 120 minute, 150 minute after they were cutted.Then single cell solution was prepared for cell vitality test and cell adhensive rate by typran rejection experiment and MTT colorimetry.Compare cell vitality,cell adhensive rate and cell proliferation ability of NVBG stored in different preserving medium.Results: Cell vitality,cell adhensive rate and cell growth rate all dropped when the NVBG storage time was prolonged in every storage medium. Cell vitality,cell adhensive rate and cell growth rate of NVBG stored in the air was remarkably lower than them stored in N.S and DMEM. But there was no remarkable differentia between NVBG stored in N.S and in DMEM. At 20min and 40min cell vitality,cell adhensive rate and cell growth rate of NVBG stored in N.S and DMEM in vitro were remarkably higher than at 80min and upwards.The parameters of NVBG brim were remarkably higher than them of NVBG center.The The parameters of the center of NVBG without implants were especially low compared with the center of the NVBG with implants.Conclusions: N.S and DMEM was better storage medium than air.The NVBG stored in N.S or DMEM could maintain good cell vitality,cell adhensive rate andproliferation power.Especially when the storage time in vitro was no more than 40 minutes.When the NVBG was stored for more than 80 minutes in N.S or DMEM or it was stored in the air the parameters would descend remarkably. 2 Experimental observation of nonvascularized bonegraft trans-planted in muscle sackObjective: Observe the effects of storage time in vitro to revascularization and osteogensis of nonvasularised bone graft transplanted in muscle.Methods: By aseptic manipulation both i lia were cut off and stored in N.S under normal atmospheric t-emperature.The NVBGs were separated to 1.5cm X 1.2cm size pi eces and one implant was implanted to each piece after extern al periosteum and soft tissue were removed.Muscle sack was p repared on the right of back.When the storage time was less t han 40 minutes and over 80 minutes the NVBG piece was res pectively placed in the prepared muscle sack.At the 3rd,7th,14th, 28th,56th day after operation dogs were sacrificed and the NVB Gs were acquired.To make routine tissue slice and HE,PAS andALP pigmentatioin were processed.The slices were observed a nd the imaging was analysed by M550 imagiometry.Parameters were analysed with SPSS vll.O and t test was carried out bet ween the parameters.Results: Angiogenesis and osteogenesis we re found in every group.At the first week positive reaction of PAS and ALP were found in ST40 group,while until the secon d week positive reaction could be found in ST80 group.At thesixth week positive reaction became weak and at the 8th weekthe reaction became negative.Imaging analysis indicated reacti on of ALP and PAS were remarkably stronger in ST40 group than in ST80 group in the lst,2ndweek(p<0.01).At the 6th week r eaction of ALP and PAS were weaker in ST40 group than in ST80 group (p>0.05).Expression of VIII factor was remarkably st ronger in ST40 group than in ST80 group at the 2nd and 4th w eek(p<0.01).Conclusions: NVBG had some ability of osteogene sis in non-bony reception region.Osteogenesis and angiogenesisof ST40 group were better than them of ST80 group. 3 The effects of rh-bFGF on biological behavior to nonvascularized bone graftObjective: To observe the effects of rh-bFGF to the proliferation in vitro and osteogenesis in vivo of NVBG under different storage time.To confirm different biological function of rh-bFGF to NVBG of different storage time in vitro.Methods: 1. By aseptic manipulation both ilia were cut off and stored in N.S under normal atmospheric temperature and one implant was implanted one side ilium.At the 20th,40th,60th,80th minute 3mm X 3mm size bone was separately cutted off from the brim of NVBG. Cut the bone to pieces and digest it to get cell solution.Calculate cells and adjust the desity to lX104/ml by DMEM.Then inoculate the cell solution to 96-hole utensil, 15 holes of experimental group and 15 holes of control group for every time point. lOug/L rh-bFGF was used to cultivate the cells of experimental group.At the 2nd,4th,6th,8th and 10th day three holes of every group of one time point were tested OD and relative growth rate was calculated.lOug/L rh-bFGF was also used to the hatch of NVBG and cell adhensive rate was calculated.All the parameters of two groups were compared with / test.2. By aseptic manipulation both ilia were cut off and stored in N.S under normal atmospheric temperature.The NVBGs wereseparated to 1.5cm X 1.2cm size pieces and one implant was implanted to each piece after external periosteum and soft tissue were removed.Muscle sack was prepared on the right of back.The rh-bFGF was used on the surface of two pieces by 100AU/cm2 as experimental group.When the storage time was less than 40 minutes and over 80 minutes the NVBG piece was respectively placed in the prepared muscle sack,the left for experimental group and the right for the controlled group.At the 3rd,7th,14th,28th,56th day after operation dogs were sacrificed and the NVBGs were acquired.To make routine tissue slice and HE,PAS and ALP pigmentatioin were processed.The slices were observed and the imaging was analysed by M550 imagiometry.Parameters were analysed with SPSS vll.O and t test was carried out between the parameters.Results: The rh-bFGF could remarkably accelerate the adhensive rate and the proliferation of NVBG of every group.The ST40 group were promoted remarkably better than the ST80 group (p<0.05) .The rh-bFGF could remarkably promote NVBG angiogenesis and osteogenesis.At 4th and 8th the expression of VI-factor of NVBG applied with rh-bFGF was remarkably stronger than it of NVBG not applied with rh-bFGF in ST40 group (p<0.01) .While in ST80 group the differentia was not remarkable(p>0.01).Conclusion: The rh-bFGF could promote the proliferation and adhensive rate in vitro.The function of rh-bFGF to 20min and 40min group were more remarkable than to 80min group.The rh-bFGF could promote the angiogenesis and osteogenesis of NVBG when transplanted in non bony region.The rh-bFGF promote the angiogenesis and osteogenesis of NVBG of ST40 group remarkably excelled ST80 group.