富血小板血浆凝胶复合材料修复颈椎缺损的实验研究
摘要
目的:探讨富血小板血浆凝胶复合材料经成骨诱导后对比格犬颈椎缺损的修复能力,为组织工程化骨的建立和修复颈椎缺损中的应用提供实验依据。
方法:(1)比格犬骨髓间充质干细胞的分离、纯化、扩增,富血小板血浆及凝胶的制备。(2)将传至第3代的骨髓间充质干细胞分别接种于富血小板血浆凝胶/强化磷酸三钙复合支架材料和单纯强化磷酸三钙支架上并体外成骨诱导培养1周。(3)经前路制作比格犬颈椎缺损模型,并分别将制备好的骨髓间充质干细胞复合单纯强化磷酸三钙支架(Ⅰ组)、犬自体髂骨(Ⅱ组)、骨髓间充质干细胞复合强化磷酸三钙/富血小板血浆凝胶支架材料(Ⅲ组)植入颈椎缺损处。观察植入物周边组织反应、局部缺损修复和骨痂形成情况和表面变化情况。(4)术后4、8、12周行X线片检查,观察X线片变化情况。(5)术后4、8、12周取材行HE染色及骨钙素蛋白免疫组织化学染色观察表达情况。(6)RT-PCR检测体内各组在4、8、12周时OCN、ALP、Col1A1的mRNA的表达情况。
结果:(1)各组材料复合体植入比格犬颈椎缺损后,伤口愈合好,没有全身和局部的炎症和毒性反应。植入材料部位均未见明显炎症及排斥反应。(2)术后4、8、12周摄X线片结果显示Ⅱ组及Ⅲ组12周时基本达到骨质愈合,而Ⅰ组仍未见材料完全吸收、骨性愈合情况,材料-骨质界面清晰,材料周缘出现负影。(3)术后4、8、12周取材行HE染色及骨钙素蛋白免疫组织化学染色可见各组均可观察到新生软骨成骨情况,强化磷酸三钙/富血小板血浆凝胶复合材料组表达较强化磷酸三钙材料组更强。(4)各组在4、8、12周各组均可见OCN、ALP和Col1A1的mRNA表达条带出现。比较4、8、12周各组目的条带和内参灰度值,8、12周时Ⅱ组和Ⅲ组各项表达强于Ⅰ组,Ⅱ组和Ⅲ组之间无明显区别。
结论:富血小板血浆凝胶/强化磷酸三钙复合支架材料在体内显著促进骨髓间充质干细胞成骨效果,并能持续稳定表达成骨特异性表型,是一种较为理想的骨组织工程支架材料,在组织相容性、降解速度及成骨效果等方面均比单纯强化磷酸三钙支架优秀。富血小板血浆凝胶/强化磷酸三钙复合支架材料能够有效的修复犬颈椎缺损,为组织工程骨在颈椎缺损修复中的应用提供了一定的实验基础和论依据。
Objective: To investigate the capacity of repairing cervical bone defect of beagle dogs with platelet-rich plasma gel composite materials after being osteogenic induced , so as to provide experimental basis for establishing tissue-engineered bone and repairing cervical defects.
Method: (1) Separation, purification, amplification of the beagle bone marrow mesenchymal stem cells, and preparation of platelet-rich plasma gel. (2) Choose the 3rd generation bone marrow mesenchymal stem cells respectively combine with platelet-rich plasma gel / beta-tricalcium phosphate composite scaffold material, simply beta-tricalcium phosphate composite scaffold and cultured with osteogenesis induced in vitro for 1 week. (3) Produced Beagle cervical bone defect model by anterior operative approach, and combined the prepared bone marrow mesenchymal stem cells with tricalcium phosphate alone enhanced stent (Ⅰgroup), canine autologous bone (Ⅱgroup), BMSCs compound strengthen the tricalcium phosphate / platelet-rich plasma gel scaffold (Ⅲgroup) implanted cervical defect respectively. Then observe the tissue reaction around the implant, partial defect repair and bone formation and surface changes. (4) X-ray inspection was used at 4,8,12 week. Observe any changes in X-ray film. (5)Obtain the sample at 4,8,12 weeks and use HE staining and immunohistochemical to observe the osteocalcin protein expression. (6) RT-PCR was used to detect the expression of mRNA regarding OCN, ALP, CollA1 in the body by each groups at 4,8,12 week.
Result: (1) There was no systemic and local inflammatory and toxicity response in each group of complex materials which implanted in beagle dogs with cervical bone defects,and the wound healing in each group was well. No significant inflammation and rejection occurred in implant sites. (2) The postoperative X-ray at 4,8,12 weeks displayed that groupⅡand groupⅢreached bone healing in 12 weeks, but the group I did not show complete absorption, bone healing, the materials - bone interface clear, material negative impact peripheral. (3) HE staining and osteocalcin protein immunohistochemical staining showed that endochondral bone emerged in each group, and the expression of enhanced tricalcium phosphate / autologous platelet-rich plasma gel composite material was stronger than the group of tricalcium phosphate group in 4,8,12 weeks. (4) The OCN, ALP and the mRNA expression of CollA1 emerging bands could be seen in 4,8,12 weeks. Compared the purpose band and the reference strip gray value of of each group, the expression of the groupⅡand groupⅢwere stronger than group I in 8,12 weeks .there is no any significant difference between groupsⅡandⅢ.
Conclusion: Platelet-rich plasma gel / beta-tricalcium phosphate composite scaffold is an ideal material for bone tissue engineering scaffold material which has a satisfactory effect especially in histocompatibility, degradation rate and the capacity of osteogenics. Platelet-rich plasma gel / beta-tricalcium phosphate composite scaffold material can effectively promote osteoblastic effect of BMSCs after revulsant in vivo, in which osteoblastic phenotype are stably expressed . Platelet-rich plasma gel / beta-tricalcium phosphate composite scaffold material can effectively repair the bone defect of cervical in dogs which provide a experimental and theoretical basis for tissue engineering to repair bone defects in the cervical spine.
方法:(1)比格犬骨髓间充质干细胞的分离、纯化、扩增,富血小板血浆及凝胶的制备。(2)将传至第3代的骨髓间充质干细胞分别接种于富血小板血浆凝胶/强化磷酸三钙复合支架材料和单纯强化磷酸三钙支架上并体外成骨诱导培养1周。(3)经前路制作比格犬颈椎缺损模型,并分别将制备好的骨髓间充质干细胞复合单纯强化磷酸三钙支架(Ⅰ组)、犬自体髂骨(Ⅱ组)、骨髓间充质干细胞复合强化磷酸三钙/富血小板血浆凝胶支架材料(Ⅲ组)植入颈椎缺损处。观察植入物周边组织反应、局部缺损修复和骨痂形成情况和表面变化情况。(4)术后4、8、12周行X线片检查,观察X线片变化情况。(5)术后4、8、12周取材行HE染色及骨钙素蛋白免疫组织化学染色观察表达情况。(6)RT-PCR检测体内各组在4、8、12周时OCN、ALP、Col1A1的mRNA的表达情况。
结果:(1)各组材料复合体植入比格犬颈椎缺损后,伤口愈合好,没有全身和局部的炎症和毒性反应。植入材料部位均未见明显炎症及排斥反应。(2)术后4、8、12周摄X线片结果显示Ⅱ组及Ⅲ组12周时基本达到骨质愈合,而Ⅰ组仍未见材料完全吸收、骨性愈合情况,材料-骨质界面清晰,材料周缘出现负影。(3)术后4、8、12周取材行HE染色及骨钙素蛋白免疫组织化学染色可见各组均可观察到新生软骨成骨情况,强化磷酸三钙/富血小板血浆凝胶复合材料组表达较强化磷酸三钙材料组更强。(4)各组在4、8、12周各组均可见OCN、ALP和Col1A1的mRNA表达条带出现。比较4、8、12周各组目的条带和内参灰度值,8、12周时Ⅱ组和Ⅲ组各项表达强于Ⅰ组,Ⅱ组和Ⅲ组之间无明显区别。
结论:富血小板血浆凝胶/强化磷酸三钙复合支架材料在体内显著促进骨髓间充质干细胞成骨效果,并能持续稳定表达成骨特异性表型,是一种较为理想的骨组织工程支架材料,在组织相容性、降解速度及成骨效果等方面均比单纯强化磷酸三钙支架优秀。富血小板血浆凝胶/强化磷酸三钙复合支架材料能够有效的修复犬颈椎缺损,为组织工程骨在颈椎缺损修复中的应用提供了一定的实验基础和论依据。
Objective: To investigate the capacity of repairing cervical bone defect of beagle dogs with platelet-rich plasma gel composite materials after being osteogenic induced , so as to provide experimental basis for establishing tissue-engineered bone and repairing cervical defects.
Method: (1) Separation, purification, amplification of the beagle bone marrow mesenchymal stem cells, and preparation of platelet-rich plasma gel. (2) Choose the 3rd generation bone marrow mesenchymal stem cells respectively combine with platelet-rich plasma gel / beta-tricalcium phosphate composite scaffold material, simply beta-tricalcium phosphate composite scaffold and cultured with osteogenesis induced in vitro for 1 week. (3) Produced Beagle cervical bone defect model by anterior operative approach, and combined the prepared bone marrow mesenchymal stem cells with tricalcium phosphate alone enhanced stent (Ⅰgroup), canine autologous bone (Ⅱgroup), BMSCs compound strengthen the tricalcium phosphate / platelet-rich plasma gel scaffold (Ⅲgroup) implanted cervical defect respectively. Then observe the tissue reaction around the implant, partial defect repair and bone formation and surface changes. (4) X-ray inspection was used at 4,8,12 week. Observe any changes in X-ray film. (5)Obtain the sample at 4,8,12 weeks and use HE staining and immunohistochemical to observe the osteocalcin protein expression. (6) RT-PCR was used to detect the expression of mRNA regarding OCN, ALP, CollA1 in the body by each groups at 4,8,12 week.
Result: (1) There was no systemic and local inflammatory and toxicity response in each group of complex materials which implanted in beagle dogs with cervical bone defects,and the wound healing in each group was well. No significant inflammation and rejection occurred in implant sites. (2) The postoperative X-ray at 4,8,12 weeks displayed that groupⅡand groupⅢreached bone healing in 12 weeks, but the group I did not show complete absorption, bone healing, the materials - bone interface clear, material negative impact peripheral. (3) HE staining and osteocalcin protein immunohistochemical staining showed that endochondral bone emerged in each group, and the expression of enhanced tricalcium phosphate / autologous platelet-rich plasma gel composite material was stronger than the group of tricalcium phosphate group in 4,8,12 weeks. (4) The OCN, ALP and the mRNA expression of CollA1 emerging bands could be seen in 4,8,12 weeks. Compared the purpose band and the reference strip gray value of of each group, the expression of the groupⅡand groupⅢwere stronger than group I in 8,12 weeks .there is no any significant difference between groupsⅡandⅢ.
Conclusion: Platelet-rich plasma gel / beta-tricalcium phosphate composite scaffold is an ideal material for bone tissue engineering scaffold material which has a satisfactory effect especially in histocompatibility, degradation rate and the capacity of osteogenics. Platelet-rich plasma gel / beta-tricalcium phosphate composite scaffold material can effectively promote osteoblastic effect of BMSCs after revulsant in vivo, in which osteoblastic phenotype are stably expressed . Platelet-rich plasma gel / beta-tricalcium phosphate composite scaffold material can effectively repair the bone defect of cervical in dogs which provide a experimental and theoretical basis for tissue engineering to repair bone defects in the cervical spine.
引文
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[1]Fennis JP,Stoelinga PJ,Jansen JA.Mandibular reconstruction:a histological and histomorphometric study on the use of autogenous scaffolds,particulate cortico-cancellous bone grafts and platelet rich plasma in goats.Int J Oral Maxillofac Surg,2004,33:48-55.
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[4]Okuda K,Kawase T,Momose M,et al.Platelet-rich plasma contains high levels of platelet-derived growth factor and transforming growth factor-beta and modulates the proliferation of periodontally related cells in vitro.J Periodontol,2003,74:849-57.
[5] Eppley BL, Woodell JE, Higgins J. Platelet quantification and growth factor analysis from platelet-rich plasma: implications for wound healing. Plast Reconstr Surg, 2004,114:1502-8.
[6] Langer R, Vacanti JP. Tissue engineering. Science, 1993,260: 920-926.
[7] Lynch SE, Genco RJ, Marx RE, et al. Tissue engineering: applications in maxillofacial surgery and periodontics [J].Carol stream IL: Quintessence,1999,11-15.
[8] Bielecki, T.M., Gazdzik, T.S., Arendt, J., Szczepanski, T.,Krol, W., and Wielkoszynski, T. Antibacterial effect of autologous platelet gel enriched with growth factors and other active substances: an in-vitro study. [J] Bone Joint Surg Br 2007.89,417-420,
[9] Lindeboom, J.A., K.R. Mathura, I.H. Aartman,F.H. Kroon, D.M. Milstein, C.Ince Influence of the application of platelet-enriched plasma in oral mucosal wound healing.[J]Clin Oral Implants Res 2007,18:133-139
[10] Marx RE, Carlson ER, Eichstaedt RM, et al. Platelet2rich p lasma: growth factor enhancement for bone grafts [J]. Oral Surg Oral Med Pathol Oral Radiol Endod, 1998, 85 (6): 638 - 646.;
[11] Landesberg R, Roy M, Glickman RS. Quantification of growth factor levels using a simplified method of platelet-rich plasma gel preparation. J Oral Maxillofac Surg, 2000,58:297-300; discussion 300-1.
[12] Carlson ER. Bone grafting the jaws in the 21st century: the use of platelet-rich plasma and bone morphogenetic protein. Alpha Omegan, 2000,93:26-30.
[13] Arpornmaeklong P, Kochel M, Depprich R, et al. Influence of platelet-rich plasma (PRP) on osteogenic differentiation of rat bone marrow stromal cellsl An in vitro study. Int J Oral Maxillofac Surg, 2004, 33 (4): 60-70
[14] Bouletreau PJ, Warren SM, Spector JA, et al. Factors in the fracture microenvironment induce primary osteoblast angiogenic cytokine production.Plast Reconstr Surg, 2002,110:139-48.
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[19]张长青,袁霆,曾炳芳,等。富血小板血浆促进骨缺损修复的实验研究.中国修复重建外科杂志,2003,17(5):355-358.
[20]Piche JE,Graves DT.Study of the growth factor requirements of human bone-derived cells:a comparison with human fibroblasts.Bone,1989,10:131-8.
[21]Lai CF,Cheng SL.Signal transductions induced by bone morphogenetic protein-2 and transforming growth factor-beta in normal human osteoblastic cells.J Biol Chem,2002,277:15514-22.
[22]Allan Mishra,.,Padmaja Tummala,.,Aaron King,Byung Lee,,Mark Kraus,Victor Tse,Christopher R.Jacobs.Buffered Platelet-Rich Plasma Enhances MesenchymalStem Cell Proliferation and Chondrogenic Differentiation[J]TISSUE ENGINEERING:Part CVolume 15,11,2009.
[23]Kocaoemer,A.,Kern,S.,Klueter,H.,and Bieback,K.HumanAB-serum as well as thrombin-activated platelet-rich-plasma are suitable alternatives to fetal calf serum for the expansion of mesenchymal stem cells.[J]Stem Cells 2007.25,1270-1278.
[24]Vogel,J.P.,Szalay,K.,Geiger,F.,Kramer,M.,Richter,W.,and Kasten,P.Platelet-rich plasma improves expansion of human mesenchymal stem cells and retains differentiation capacity and in vivo bone formation in calcium phosphate.[J]Platelets2006.17,462-469.
[25]Kim ES,Park EJ,Choung PH1Platelet concentration and its effect on bone formation in calvarial defects:an experimental study in rabbits.J Prothet Dent,2001,86(4):428-433.
[26]Kim SG,Kim WK,Park JC,et al.A comparative study of osseointegration of Avana imp lants in a demineralized freez-dried bone alone orwith platelet-rich plasma.J Oral Maxillofac Surg,2002,60(9):1018-1025.
[27]Fennis JP,Stoelinga PJ,Jansen JA.Mandibular reconstruction:A clinical and radiographic animal study on the use of autogenous scaffolds and platelet-rich plasma[J].Int J Oral Maxillofac Surg,2002,31(3):281-286.
[28]Camargo PM,Lekovic V,WeinlaenderM,et al.Platelet-rich plasma and bovine porous bone mineral combined with guided tissue regeneration in the treatment of intrabony defect in humans[J].J Periodontal Res,2002,37(4):300-306.
[29]Fontana S,Olmedo DG,Linares J,et al.Effect of platelet-rich plasma on the perimplant bone response:an experiment study.Implant Dent,2004,13(1):73-78.
[30]Lucarelli E,Fini M,Beccheroni A,et al.Stromal stem Cells and platelet-rich plasma improve bone allograft integration.Clin Othop Relat Res,2005,435:62-68.
[31]Kovacs K,Velich N,Huszar T,et al.Mistomorphometric and densitometric evaluation of the effects of platelet-rich plasma on the remodeling of beta-tricalcium phosphate in beagle dogs[J].J Craniofac Surg,2005,16(1):150-154.
[32]陶海荣,张长青,曾炳芳,等.磷酸三钙结合富血小板血浆修复股骨头坏死的研究.中国修复重建外科杂志,2005,19(3):170-173.
[33]Fujimori T.The effect of platelet-rich plasma combined with autogenous bone graft for bone regeneration in bone defects[J].Kokubyo Gakkai Zasshi,2005,72(1):90-97.
[34]Tomoyasu A,Higashio K,Kanomata K,et al.Platelet-rich plasma stimulates osteoblastic differentiation in the presence of BMPs.Biochem Biophys Res Commun.2007 Sep 14;361(1):62-7.
[35]Hokugo A,Ozeki M,Kawakami O,et al,Augmented bone regeneration activity of platelet-rich plasma by biodegradable gelatin hydrogel.Tissue Eng.2005 Jul-Aug;11(7-8):1224-1257.
[36] Nikolidakis D, Van den Dolder J, Joop G.C, et al. The Effect of Platelet-Rich Plasma on the Bone Healing around Calcium Phosphate-Coated and Non-coated Oral Implants in Trabecular Bone. Tissue Eng. 2006 Sept;9(12):2555-2563.
[37] Jensen TB, Rahbek 0, Overgaard S, S(?)balle K. No effect of platelet-rich plasma with frozen or processed bone allograft around noncemented implants. Int Orthop. 2005 Apr;29(2):67-72.
[38] Van den Dolder J, Mooren R, Vloon AP, et al.Platelet-rich plasma:quantification of growth factor levels and the effect on growth and differentiation of rat bone marrow cells.Tissue Eng. 2006 Nov;12(11):3067-73.
[39] Kasten P, Vogel J, Luginbuhl R, et al. Influence of platelet-rich plasma on osteogenic differentiation of mesenchymal stem cells and ectopic bone formation in calcium phosphate ceramics. Cells Tissues Organs. 2006;183(2):68-79.
[40] Tajima N, Sotome S, Marukawa E, et al. A three-dimensional cell-loading system using autologous plasma loaded into a porous β-tricalcium-phosphate block promotes bone formation at extraskeletal sites in rats. Materials Science and Engineering C 27 (2007) 625-632.
[41] Oyama T, Nishimoto S, Tsugawa T, et al. Efficacy of platelet-rich plasma in alveolar bone grafting. J Oral Maxillofac Surg, 2004,62:555-8.
[42] RobionyM, Polini F, Costa F, et al. Osteogenesis distraction and platelet-rich plasma for bone restoration of the severely atrophic mandible: preliminary results. J Oral Maxillofac Surg, 2002,60 (6): 630-635.
[43] Oyama T, Nishimoto S, Tsugawa T, et al. Efficacy of platelet-rich plasma in alveolar bone graftingl J Oral Maxillofac Surg, 2004, 62(5): 555-558.
[44] Mazor Z, PelegM, Garg AK, et al.Platelet-rich plasma for bone graft enhancement in sinus floor augmentation with Simultaneous implant placement: patient series study. Implant Dent, 2004,13 (1): 65-72.
[45] Kitoh H, Kitakoji T, Tsuchiya H, et al. Transplantation of marrow-derived mesenchymal stem cells and platelet-rich plasma during distraction osteogen- esis-a preliminary result of three cases.Bone,2004,35:892-8.
[46]Kitoh H,Kitakoji T,Tsuchiya H,et al.Transplantation of culture expanded bone marrow ceils and platelet rich plasma in distraction osteogenesis of the long bones.Bone 40(2007) 522-528.
[47]Michael D.Barnett Jr,MD.Use of Platelet-Rich Plasma and Bone Marrow-Derived Mesenchymal Stem Cells in Foot and Ankle Surgery.Techniques in Foot and Ankle Surgery 6(2):89-94,2007.
[48]Hanna R,Trejo PM,Weltman RL.Treatment of intrabony defects with bovine-derived xenograft alone and in combination with platelet-rich plasma:a randomized clinical trial.J Periodontol,2004,75:1668-77.
[49]Lowery GL,Kulkarni S,Pennisi AE.Use of autologous growth factors in lumbar spinal fusion[J].Bone,1999,25(2 Supp 1):47-50.
[50]Castro FP Jr.Role of activated growth factors in lumbar spinal fusions[J].J Spinal Disord Tech,2004,17(5):380-384.
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[16]王玺,β-磷酸三钙/生物玻璃多孔复合支架的制备与强化研究,中南大学粉末冶金研究院2006年硕士毕业论文。
[17]Okuda K,Kawase T,Momose M,et al.Platelet-rich plasma contains high levels of platelet-derived growth factor and transforming growth factor-beta and modulates the proliferation of periodontally related cells in vitro.J Periodontol,2003,74:849-57.
[18]Eppley BL,Woodell JE,Higgins J.Platelet quantification and growth factor analysis from platelet-rich plasma:implications for wound healing.Plast Reconstr Surg,2004,114:1502-8.
[19]Bielecki,T.M.,Gazdzik,T.S.,Arendt,J.,Szczepanski,T.,Krol,W.,and Wielkoszynski,T.Antibacterial effect of autologous platelet gel enriched with growth factors and other active substances:an in-vitro study.[J]Bone Joint Surg Br 2007.89,417-420,
[20]Lindeboom,J.A.,K.R.Mathura,I.H.Aartman,F.H.Kroon,D.M.Milstein,C.Ince Influence of the application of platelet-enriched plasma in oral mucosal wound healing.[J]Clin Oral Implants Res 2007,18:133-139
[21]Allan Mishra,.,Padmaja Tummala,.,Aaron King,.,Byung Lee,,Mark Kraus,Victor Tse,Christopher R.Jacobs.Buffered Platelet-Rich Plasma Enhances MesenchymalStem Cell Proliferation and Chondrogenic Differentiation[J]TISSUE ENGINEERING:Part C Volume 15,Number 00,2009.
[22]Anja D,Antonia Z,Ewa K S,Klaus M S,Karl-Heinz F.Influence of Platelet-Rich Plasma on Chondrogenic Differentiation and Proliferation of Chondrocytes and Mesenchymal Stem Cells[J]Cells Tissues Organs 2009;189:317-326.
[23]Fennis JP,Stoelinga PJ,Janson JA.Mandibular reconstruction:a histological and histomerphometric study on the use of autogenous scaffolds,particulate cortico-cancellous bone grafts and platelet rich plasma in goats.Int J Oral Maxillofac Surg, 2004,33(1): 48-55.
[24] Marx RE, Carlson ER, Eichstaedt RM, et al. Platelet2rich p lasma: growth factor enhancement for bone grafts [J]. Oral Surg Oral Med Pathol Oral Radiol Endod, 1998, 85 (6): 638 - 646.
[25] Bouletreau PJ, Warren SM, Spector JA, et al. Factors in the fracture microenvironment induce primary osteoblast angiogenic cytokine production. Plast Reconstr Surg, 2002,110:139-48.
[26] Carlson ER. Bone grafting the jaws in the 21 st century: the use of platelet-rich plasma and bone morphogenetic protein. Alpha Omegan, 2000,93:26-30.
[27] Landesberg R, Roy M, Glickman RS. Quantification of growth factor levels using a simplified method of platelet-rich plasma gel preparation. J Oral Maxillofac Surg, 2000,58:297-300;
[28] Arpornmaeklong P, Kochel M, Depprich R, et al. Influence of platelet-rich plasma (PRP) on osteogenic differentiation of rat bone marrow stromal cellsl An in vitro study. Int J Oral Maxillofac Surg, 2004,33 (4): 60-70.
[29] Pieri F, Lucarelli E, Corinaldesi G, Iezzi G, Piattelli A, Giardino R, Bassi M,Donati D, Marchetti C. Mesenchymal stem cells and platelet-rich plasma enhance bone formation in sinus grafting: a histomorphometric study in minipigs.[J] Clin Periodontol 2008; 35: 539-546.
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[33] Nikolidakis D, Van den Dolder J, Joop GC, et al. The Effect of Platelet-Rich Plasma on the Bone Healing around Calcium Phosphate-Coated and Non-coated Oral Implants in Trabecular Bone. Tissue Eng. 2006 Sept; 9(12):2555-2563.
[34] Van den Dolder J, Mooren R, Vloon AP, et al.Platelet-rich plasma:quantification of growth factor levels and the effect on growth and differentiation of rat bone marrow cells.Tissue Eng. 2006 Nov;12(11):3067-73.
[35] Tajima N, Sotome S, Marukawa E, et al. A three-dimensional cell-loading system using autologous plasma loaded into a porous P-tricalcium-phosphate block promotes bone formation at extraskeletal sites in rats. Materials Science and Engineering C 27 (2007)625-632.
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[1]Fennis JP,Stoelinga PJ,Jansen JA.Mandibular reconstruction:a histological and histomorphometric study on the use of autogenous scaffolds,particulate cortico-cancellous bone grafts and platelet rich plasma in goats.Int J Oral Maxillofac Surg,2004,33:48-55.
[2]Kassolis JD,Reynolds MA.Evaluation of the adjunctive benefits of platelet-rich plasma in subantral sinus augmentation.J Craniofac Surg,2005,16:280-7.
[3]Assoian RK,Grotendorst GR,Miller DM,et al,Cellular transformation by coordinated action of three peptide growth factors from human platelet,Nature,1984,309(5971):804-810
[4]Okuda K,Kawase T,Momose M,et al.Platelet-rich plasma contains high levels of platelet-derived growth factor and transforming growth factor-beta and modulates the proliferation of periodontally related cells in vitro.J Periodontol,2003,74:849-57.
[5] Eppley BL, Woodell JE, Higgins J. Platelet quantification and growth factor analysis from platelet-rich plasma: implications for wound healing. Plast Reconstr Surg, 2004,114:1502-8.
[6] Langer R, Vacanti JP. Tissue engineering. Science, 1993,260: 920-926.
[7] Lynch SE, Genco RJ, Marx RE, et al. Tissue engineering: applications in maxillofacial surgery and periodontics [J].Carol stream IL: Quintessence,1999,11-15.
[8] Bielecki, T.M., Gazdzik, T.S., Arendt, J., Szczepanski, T.,Krol, W., and Wielkoszynski, T. Antibacterial effect of autologous platelet gel enriched with growth factors and other active substances: an in-vitro study. [J] Bone Joint Surg Br 2007.89,417-420,
[9] Lindeboom, J.A., K.R. Mathura, I.H. Aartman,F.H. Kroon, D.M. Milstein, C.Ince Influence of the application of platelet-enriched plasma in oral mucosal wound healing.[J]Clin Oral Implants Res 2007,18:133-139
[10] Marx RE, Carlson ER, Eichstaedt RM, et al. Platelet2rich p lasma: growth factor enhancement for bone grafts [J]. Oral Surg Oral Med Pathol Oral Radiol Endod, 1998, 85 (6): 638 - 646.;
[11] Landesberg R, Roy M, Glickman RS. Quantification of growth factor levels using a simplified method of platelet-rich plasma gel preparation. J Oral Maxillofac Surg, 2000,58:297-300; discussion 300-1.
[12] Carlson ER. Bone grafting the jaws in the 21st century: the use of platelet-rich plasma and bone morphogenetic protein. Alpha Omegan, 2000,93:26-30.
[13] Arpornmaeklong P, Kochel M, Depprich R, et al. Influence of platelet-rich plasma (PRP) on osteogenic differentiation of rat bone marrow stromal cellsl An in vitro study. Int J Oral Maxillofac Surg, 2004, 33 (4): 60-70
[14] Bouletreau PJ, Warren SM, Spector JA, et al. Factors in the fracture microenvironment induce primary osteoblast angiogenic cytokine production.Plast Reconstr Surg, 2002,110:139-48.
[15] Horner A, Bord S, Kemp P, et al. Distribution of platelet-derived growth factor (PDGF) A chain mRNA, protein, and PDGF-alpha receptor in rapidly forming human bone. Bone, 1996,19:353-62.
[16]Lind M.Growth factor stimulation of bone healing.Effects on osteoblasts,osteomies,and implants fixation.Acta Orthop Scand Suppl,1998,283:2-37.
[17]Lai CF,Cheng SL.Signal transductions induced by bone morphogenetic protein-2 and transforming growth factor-beta in normal human osteoblastic cells.J Biol Chem,2002,277(18):15514-15522.
[18]Arpornmaeklong P,Kochel M,Depprich R,et al.Influence of platelet-rich plasma(PRP) on osteogenic differentiation of rat bone marrow stromal cells.An in vitro study.Int J Oral Maxillofac Surg,2004,33:60-70.
[19]张长青,袁霆,曾炳芳,等。富血小板血浆促进骨缺损修复的实验研究.中国修复重建外科杂志,2003,17(5):355-358.
[20]Piche JE,Graves DT.Study of the growth factor requirements of human bone-derived cells:a comparison with human fibroblasts.Bone,1989,10:131-8.
[21]Lai CF,Cheng SL.Signal transductions induced by bone morphogenetic protein-2 and transforming growth factor-beta in normal human osteoblastic cells.J Biol Chem,2002,277:15514-22.
[22]Allan Mishra,.,Padmaja Tummala,.,Aaron King,Byung Lee,,Mark Kraus,Victor Tse,Christopher R.Jacobs.Buffered Platelet-Rich Plasma Enhances MesenchymalStem Cell Proliferation and Chondrogenic Differentiation[J]TISSUE ENGINEERING:Part CVolume 15,11,2009.
[23]Kocaoemer,A.,Kern,S.,Klueter,H.,and Bieback,K.HumanAB-serum as well as thrombin-activated platelet-rich-plasma are suitable alternatives to fetal calf serum for the expansion of mesenchymal stem cells.[J]Stem Cells 2007.25,1270-1278.
[24]Vogel,J.P.,Szalay,K.,Geiger,F.,Kramer,M.,Richter,W.,and Kasten,P.Platelet-rich plasma improves expansion of human mesenchymal stem cells and retains differentiation capacity and in vivo bone formation in calcium phosphate.[J]Platelets2006.17,462-469.
[25]Kim ES,Park EJ,Choung PH1Platelet concentration and its effect on bone formation in calvarial defects:an experimental study in rabbits.J Prothet Dent,2001,86(4):428-433.
[26]Kim SG,Kim WK,Park JC,et al.A comparative study of osseointegration of Avana imp lants in a demineralized freez-dried bone alone orwith platelet-rich plasma.J Oral Maxillofac Surg,2002,60(9):1018-1025.
[27]Fennis JP,Stoelinga PJ,Jansen JA.Mandibular reconstruction:A clinical and radiographic animal study on the use of autogenous scaffolds and platelet-rich plasma[J].Int J Oral Maxillofac Surg,2002,31(3):281-286.
[28]Camargo PM,Lekovic V,WeinlaenderM,et al.Platelet-rich plasma and bovine porous bone mineral combined with guided tissue regeneration in the treatment of intrabony defect in humans[J].J Periodontal Res,2002,37(4):300-306.
[29]Fontana S,Olmedo DG,Linares J,et al.Effect of platelet-rich plasma on the perimplant bone response:an experiment study.Implant Dent,2004,13(1):73-78.
[30]Lucarelli E,Fini M,Beccheroni A,et al.Stromal stem Cells and platelet-rich plasma improve bone allograft integration.Clin Othop Relat Res,2005,435:62-68.
[31]Kovacs K,Velich N,Huszar T,et al.Mistomorphometric and densitometric evaluation of the effects of platelet-rich plasma on the remodeling of beta-tricalcium phosphate in beagle dogs[J].J Craniofac Surg,2005,16(1):150-154.
[32]陶海荣,张长青,曾炳芳,等.磷酸三钙结合富血小板血浆修复股骨头坏死的研究.中国修复重建外科杂志,2005,19(3):170-173.
[33]Fujimori T.The effect of platelet-rich plasma combined with autogenous bone graft for bone regeneration in bone defects[J].Kokubyo Gakkai Zasshi,2005,72(1):90-97.
[34]Tomoyasu A,Higashio K,Kanomata K,et al.Platelet-rich plasma stimulates osteoblastic differentiation in the presence of BMPs.Biochem Biophys Res Commun.2007 Sep 14;361(1):62-7.
[35]Hokugo A,Ozeki M,Kawakami O,et al,Augmented bone regeneration activity of platelet-rich plasma by biodegradable gelatin hydrogel.Tissue Eng.2005 Jul-Aug;11(7-8):1224-1257.
[36] Nikolidakis D, Van den Dolder J, Joop G.C, et al. The Effect of Platelet-Rich Plasma on the Bone Healing around Calcium Phosphate-Coated and Non-coated Oral Implants in Trabecular Bone. Tissue Eng. 2006 Sept;9(12):2555-2563.
[37] Jensen TB, Rahbek 0, Overgaard S, S(?)balle K. No effect of platelet-rich plasma with frozen or processed bone allograft around noncemented implants. Int Orthop. 2005 Apr;29(2):67-72.
[38] Van den Dolder J, Mooren R, Vloon AP, et al.Platelet-rich plasma:quantification of growth factor levels and the effect on growth and differentiation of rat bone marrow cells.Tissue Eng. 2006 Nov;12(11):3067-73.
[39] Kasten P, Vogel J, Luginbuhl R, et al. Influence of platelet-rich plasma on osteogenic differentiation of mesenchymal stem cells and ectopic bone formation in calcium phosphate ceramics. Cells Tissues Organs. 2006;183(2):68-79.
[40] Tajima N, Sotome S, Marukawa E, et al. A three-dimensional cell-loading system using autologous plasma loaded into a porous β-tricalcium-phosphate block promotes bone formation at extraskeletal sites in rats. Materials Science and Engineering C 27 (2007) 625-632.
[41] Oyama T, Nishimoto S, Tsugawa T, et al. Efficacy of platelet-rich plasma in alveolar bone grafting. J Oral Maxillofac Surg, 2004,62:555-8.
[42] RobionyM, Polini F, Costa F, et al. Osteogenesis distraction and platelet-rich plasma for bone restoration of the severely atrophic mandible: preliminary results. J Oral Maxillofac Surg, 2002,60 (6): 630-635.
[43] Oyama T, Nishimoto S, Tsugawa T, et al. Efficacy of platelet-rich plasma in alveolar bone graftingl J Oral Maxillofac Surg, 2004, 62(5): 555-558.
[44] Mazor Z, PelegM, Garg AK, et al.Platelet-rich plasma for bone graft enhancement in sinus floor augmentation with Simultaneous implant placement: patient series study. Implant Dent, 2004,13 (1): 65-72.
[45] Kitoh H, Kitakoji T, Tsuchiya H, et al. Transplantation of marrow-derived mesenchymal stem cells and platelet-rich plasma during distraction osteogen- esis-a preliminary result of three cases.Bone,2004,35:892-8.
[46]Kitoh H,Kitakoji T,Tsuchiya H,et al.Transplantation of culture expanded bone marrow ceils and platelet rich plasma in distraction osteogenesis of the long bones.Bone 40(2007) 522-528.
[47]Michael D.Barnett Jr,MD.Use of Platelet-Rich Plasma and Bone Marrow-Derived Mesenchymal Stem Cells in Foot and Ankle Surgery.Techniques in Foot and Ankle Surgery 6(2):89-94,2007.
[48]Hanna R,Trejo PM,Weltman RL.Treatment of intrabony defects with bovine-derived xenograft alone and in combination with platelet-rich plasma:a randomized clinical trial.J Periodontol,2004,75:1668-77.
[49]Lowery GL,Kulkarni S,Pennisi AE.Use of autologous growth factors in lumbar spinal fusion[J].Bone,1999,25(2 Supp 1):47-50.
[50]Castro FP Jr.Role of activated growth factors in lumbar spinal fusions[J].J Spinal Disord Tech,2004,17(5):380-384.