| Objective: To study bone induction activity of P17derived from Bone morphogeneticprotein2.Methods: P17was made through FMOC/tBu solid-phase synthesis method. Take asmall amount of P17and analyze it by spectrometry and chromatographic method todetermine P17molecular weight and purity. Cultivate SD rat mesenchymal cells invitro and prepare two kinds of P17saline solution,10μg/ml and30μg/ml. These twokinds of solution were added to the mesenchymal stem cell complete medium. After2weeks and3weeks, observe the cell growth condition under a microscope and detectthe expression of alkaline phosphatase.Results: The average molecular weight of P17was2023.67, consistent with thetheoretical design of peptide molecular weight; High Pressure Liquid Chromatograph(HPLC) showed the activity of synthetic peptide purity was96%, meeting therequirement of experiment. With the extension of cultivation time, the ALP expressionof the two groups continued to elevate. After2and3weeks, the ALP content of30μg/ml group were higher than that of10μg/ml group.Conclusion:BMP-2related peptide P17prepared by solid-phase peptide synthesismethod has induction activity in vitro. The induction activity of30μg/ml group isbetter than10μg/ml group. Objective: Prepare an injectable artificial bone repair material, α-calcium sulphatehemihydrate (α-CSH) and mineralized collagen (nHAC). Test the physical andchemical properties of the different components; Add BMP-2related peptide intoCSH/nHAC, test its release properties in vitro.Methods:(1). α-CSH was mixed with mineralized collagen with different content.2gsamples were mixed with deionized water according to different liquid/solid ratio,moved to a5ml syringe. Record the liquid-solid ratio under which the material cankeep good injectable propertities.(2). Study the curing properties of composites withdifferent liquid/solid ratio, mineralized collagen content and different content of CSD.(3). Test the compression properties of the materials in a universal material testingmachine.(4). Put the material in the simulated body fluid after solidification and testits degradation rate.(5). Prepare cylindrical material loaded with P17, put it intosimulated body fluid and test the release properties in vitro.Results:(1) As the content of mineralized collagen increases, the liquid/solid ratio alsoneed to increase in order to keep good injection properties.(2) With the mineralizedcollagen content and the liquid/solid ratio increase, the solidification time increases.The addition of CSD can significantly shorten the solidification time. Thesolidification time can be adjusted between4.3±0.6min to169±22.3min.(3) As theincrease of liquid/solid ratio and the content of mineralized collagen, the compressionproperties of this composites decreased from18.3±2.3Mpa to1.5±0.5Mpa.(4) Withthe increase of solid/liquid ratio, the material degradation rate increased; the moremineralized collagen in the composite, the slower its degradation rate was; Addingcoagulant and P17has no significant impact on the degradation rate.(5) This materialcan be used as a carrier of P17. P17was released in a sustained way in vitro and it cancontinue to release for more than21days. Conclusion: Calcium sulfate/mineralized collagen is a kind of self-curing and in situinjectable bone repair material and it can be used as the carrier of BMP-2relatedpeptide. Objective: To study the ability of BMP-2related peptide/mineralized collagen/calciumsulfate to repair bone defect.Methods:45New Zealand white rabbit were randomly divided into three groups, theblank group (group A), calcium sulfate/mineralized collagen group (group B),P17/calcium sulfate/mineralized collagen group (group C). Femoral condyle bonedefects(depth of10mm, diameter of7mm) were made in all rabbits. The materialswere implanted into the bone defect respectively.4,8,12weeks after surgery, X-ray,histological observation and Micro CT were taken to compare the ability of boneregeneration between three groups.Results: The gross observation: all animals woke up1hour after operation, couldstand in the first day. After surgery for2days, the rabbits could walk freely. Knee jointmoved well. X-ray: Group A:12weeks, bone defect was obvious, the edge of bonedefect edge was blur and a small amount of new bone formed. Group B:4weeks aftersurgery, some material degraded, there was still some material existing, bone defectedge were blur, a small amount of new bone formed;8weeks after operation, thematerials almost degraded completely, and the defect area was still visible, but repaireffect was superior to group A;12weeks after surgery, most of the bone defect wasrepaired, no obvious material was observed. Group C: The repair effect was better thanthe other two groups at each time points. At12weeks bone defect was completelyrepaired. Micro CT: The new bone formed from the edges of the defect. The boneregeneration of group C was obviously better than the other two groups,12weeks after surgery, bone defect in group A was still obvious. Histological observation: In HEstaining, from4weeks to12weeks, new bone formation area percentage of group Aincreased from6.20±7.6%to15.40±7.30%; for group B it increased from32.20±6.26%to41.60±6.26%, for group C38.00±8.97%to67.20±6.89%. The newbone formation of group A and group B were significantly less than that of group Cand the difference was statistically significant (p<0.05). In Sirius red staining, type Icollagen fiber area of group A and group B was obviously less than group C and thedifference was statistically significant (p<0.05).Conclusion: The repair effect of P17/CSH/nHAC is better than CSH/nHAC.P17/CSH/nHAC material is a kind of ideal bone tissue engineering material and it caneffectively promote the bone defect repair. |
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