| Objective (1) To construct a plastic engineered bone in vitro. (2) Toobserve bone generation in vivo of the plastic engineered bone andeffectiveness of the plastic engineered bone to repair bone defect. (3) Toevaluate the bone generation ability of serial plastic engineered bonesconstructed by different methods. Methods ?After obtaining osteoblastsfrom infant rabbit skulls and preparing granular bio-derived bones,construct plastic engineered bone with osteoblasts,alginate gel and granularbio-derived bones. (2) 44 Japanese white rabbits were randomly divided intogroup A(n=28), group B(n=12) and group C(n=4).Bilateral cranialperiosteum and full skull defects 10 mm in diameter were created and the leftdefects of group A( group A1) were filled with the plastic engineeredbone.The right defects of group A (group A2)were filled with the mixture ofalginate gel and granular bio-derived bones.The defects of group B wereuntreated as control groups and group C as normal controls.Boneregeneration in vivo ,defect repairing and vascularization of plasticengineered bone were studied by means of gross observation, X ray,electronic scan, histology, immunohistochemistry, biomechanics, measuringthe amount of Ca and P in repairing tissue and perfusion of vessel. (3)88Japanese white rabbits were randomly divided into group D(n=20), groupE(n=20), group F(n=20), group G(n=20) and group H(n=8).Bilateral cranialperiosteum and full skull defects 10 mm in diameter were created and the leftdefects of group D were filled with osteoblasts-alginate gel (group Dl) with right defects filled by alginate gel as control(group D2).The left defects of group E were filled with the compound constructed by alginate gel-osteoblasts-granular bio-derived bones(group El) and the right defects were filled with the compound constructed by alginate gel-granular bio-derived bones as control(group E2). The left defects of group F were filled with the compound composed of alginate gel-osteoblasts-SIS powers (group Fl) and the right defects were filled with the compound composed of alginate gel- SIS powers as control(group F2). The left defects of group G were filled with the compound composed of alginate gel-osteoblasts- granular bio-derived bones -SIS powers (group Gl) and the right defects were filled with the compound composed of alginate gel- granular bio-derived bones-SIS powers as control(group G2).Bilateral defects of group H were untreated as blank control.The bone generation and defect repairing were observed by means of gross observation ,X way, histology, biomechanics and measuring Ca and P amount in defect-repairing tissue.Results ? The plastic engineered bone constructed by osteoblast-alginate gel-granular bio-derived bone was paste-like material which may fill bone defect with any shape. (2) After repairing defects with the plastic engineered bone,at 2 week time point alginate gel was absorbed partially and was replaced by fibrous tissue with osteoid tissue blocks in it.There were new bone formation around the granular bio-derived bone and near the cranial defect edge. Great many BrdU positive cells in and around the newly formed bones.At 4 week time point,new bone formed in fibrous tissue and around the granular bio-derived bones increased and expressed I type collagen. At 6 week time point, thegranular bio-derived bones were absorbed partially and the new bonesformed in fibrous tissue ,around the granular bio-derived bones and nearcranial defect edge were connected mutually to form bone structure lookinglike weave bone with chaos array. At 12 week time point,the cranial defectswere repaired nearly completely by bone tissue .Radiogram showed that highdensity bone callus crowded the whole defects and histology demonstratedthat the repairing tissue was composed of bone trabeculae with marrow cavityformation .There were little amount of residual alginate gel scattering in therepairing tissue and the granular bio-derived bones had already beenabsorbed completely. The bone area percentage was 40.92 +19.36%.The Caand P amounts were respectively 7.30±0.93mg and 3.11 ±0.42mg.Maximalcompression load beared by repairing tissue was 37.33 + 2.95 N. Vesselperfusion showed that the whole area of defect was vascularized completely.Repairing tissues of group A2 were composed of bone tissue in marginal areaand soft fibrous tissue in central part with partial bone heal ratio of 4/8. Thebone formation area percentage was 18.51 ±6.01%,statistically lower thangroup Al(p<0.05).Radiogram showed that high density bone callusdistributed in the marginal area in the majority. The Ca and P amounts wererespectively 4.89 + 0.27mg and 2.05 + 0.25mg,statistically lower than groupAl(p<0.05).Maximal compression load beared by repairing tissue was 30.97+ 0.27 N, statistically lower than group Al(p<0.05).The defects of group Bwere mostly repaired by membranous fibrous tissue with little new bone inthe marginal area. The bone area percentage was 12.72 ± 9.46%,statisticallylower than group Al(p<0.05). Radiogram showed that high density bonecallus mostly distributed in the marginal area near the defect edge. The Caand P amounts were respectively 3.54 ± 0.45mg and 1.78 ± 0.53mg,statistically lower than group Al(p<0.05).Maximal compression load beared by repairing tissue was 29.5 + 2.05 N, statistically lower than group Al(p<0.05). (3) At 12 week time point,the most repairing type of defects of group Dl was incomplete bone healing ,with histological bone area percentage of 23.56 + 8.22%. The Ca and P amounts were respectively 5.29 + 0.17mg and 2.34±0.35mg. Maximal compression load beared by repairing tissue was 31.08 + 3.65N. The most repairing type of defects of group D2 was no bone healing ,with histological bone area percentage of 12.23 + 8.63%. The most repairing type of defects of group El was complete bone healing ,with histological bone area percentage of 40.92 + 19.26% statistically more than group Dl(p<0.05). The Ca and P amounts were respectively 7.30 + 0.93 mg and 3.11+0.42 mg. Maximal compression load beared by the repairing tissue was 37.33 +2.95N statistically more than group Dl(p<0.05). The most repairing type of defects of group E2 was partial bone healing ,with histological bone area percentage of 18.51+6.01%. The most repairing type of defects of group Fl was incomplete bone healing ,with histological bone area percentage of 25.42 + 11.41%. Maximal compression load beared by the repairing tissue was 32.35 + 1.98N. The most repairing type of defects of group F2 was no bone healing ,with histological bone area percentage of 15.54 + 7.97%. The most repairing type of defects of group Gl was complete bone healing ,with histological bone area percentage of 42.62 + 17.66% statistically more than group Dl(p<0.05). The Ca and P amounts were respectively 8.15 + 0.43 mg and 3.83 + 0.37 mg. Maximal compression load beared by the repairing tissue was 38.37±2.15N statistically more thangroup Dl(p<0.05). The most repairing types of defects of group G2 were incomplete bone healing and partial bone healing,with histological bone area percentage of 19.61 + 7.21%. The most repairing type of defects of group H was no bone healing ,with histological bone area percentage of 12.72 + 9.46% statistically lower than group Dl,El,Fl,Gl(p<0.05). The Ca and P amounts were respectively 3.54 + 0.45 mg and 1.78 + 0.53 mg. Maximal compression load beared by the repairing tissue was 29.5 + 2.05 N statistically more than group El and Gl(p<0.05).Conclusions ? It is easy to fill small bone defects such as bone gap, round defect with the plastic engineered bone constructed by osteoblaste-alginate gel-granular bio-derived bone. ?The plastic engineered bone showed good ability to form bone tissue in vivo and could repair bone defect nearly completely with new bone mostly composed of bone trabeculae.The way of forming bone is endochondral ossification. ?Implanted osteoblastes are major functional cells. ? The granular bio-derived bone has osteoconductive and osteoinductive properties and may be absorbed completely in 12 weeks. ? The alginate gel could not be absorbed completely in 12 weeks. ? Depending on the quality and quantity of new bone formed in vivo,4 different plastic engineered bones which were respectively constructed by osteoblaste-alginate gel-SIS powder-granular bio-derived bones, osteoblaste-alginate gel-granular bio-derived bones, osteoblaste-alginate gel-SIS powder and osteoblaste-alginate gel could be ranked as following: the osteoblaste-alginate gel-SIS powder-granular bio-derived bone group be superior to the osteoblaste-alginate gel- granular bio-derived bone group, the osteoblaste-alginate gel- granular bio-derived bone group be superior to theosteoblaste-alginate gel-SIS powder group,and the osteoblaste-alginate gel-SIS powder group be superior to osteoblaste-alginate gel group.? SIS powder way augment bone formation of the plastic engineered bone by accelerating vessel regeneration.
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