侧壁开窗法上颌窦底提升术植入Bio-Oss后骨生成的mRNA表达谱

摘要

目的:探讨侧壁开窗法上颌窦底提升术这种特殊的引导性骨再生(guided bone regeneration,GBR)过程的转录组特点.方法:3只Beagle犬,随机选择一侧经侧壁开窗行上颌窦底提升术,同时植入Bio-Oss骨粉,术后2周,对另一侧用相同的方法行上颌窦底提升术.第2次术后2周,收集植入的Bio-Oss骨粉组织块,提取总RNA,全基因组表达谱芯片检测分析上颌窦底提升后2周和4周时的差异表达基因.利用基因本体论(gene ontolo-gY,GO)对差异表达的基因进行功能分类,以及京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Ge-nomes,KEGG)进行信号通路分析.结果:在2周时表达升高的基因与刺激反应、免疫-炎症反应和细胞增殖有关,同时,与神经元分化、破骨细胞分化和早期血管化过程相关的基因也高表达.在4周时,与细胞代谢和发育相关的基因表达升高,重要的是,与血管生成和骨骼发育相关的基因都高表达,与骨骼发育相关的基因包括骨生成相关的生长和分化因子[转化生成因子β1(transforming growth factor beta 1,TGFB1)和骨形态发生蛋白4(bone morphogenetic protein 4,BMP4)]、软骨和骨胞外基质[软骨黏附素(chondroadherin,CHAD)、Ⅰ型胶原α1链(collagentypeⅠ,alpha 1 chain,COL1A1)、Ⅰ型胶原α2链(collagen type Ⅰ,alpha 2 chain,COL1A2)、V型胶原α2链(collagen type Ⅴ,alpha2 chain,COL5A2)、核心蛋白聚糖(decorin,DCN)、骨连接蛋白(osteonecfin,SPARC)和骨钙素(osteocalcin,BGLAP)]等相关的基因.信号通路分析显示4周时表达升高的基因与转化生长因子β/骨形态发生蛋白(transforming growth factor beta/bone morphogenetic protein,TGF-β/BMP)信号通路有关.无翅型MMTV整合位点家族(wingless-type MMTV integration site family,WNT)信号通路中的关键基因WNT5A和卷曲蛋白受体9(frizzled receptor 9,FZD9)在2周时表达升高,而连环蛋白β1(catenin beta 1,CTNNB1)在4周时表达升高.结论:侧壁开窗法上颌窦底提升术2周后转录表达谱变化以免疫-炎症反应和破骨反应下调、骨生成相关基因表达升高为特点,骨再生的过程还与神经生成和血管生成过程相关.在骨再生的过程中,TGF-β/BMP和WNT信号通路起重要作用,WNT信号通路的作用复杂.%AIM:To explore the characteristics of transcriptome associated with the unique guided bone regeneration (GBR) of maxillary sinus floor elevation by the lateral window approach.METHODS:One side of the upper jaw was randomly selected for maxillary sinus floor elevation with Bio-Oss by the lateral window approach in 3 Beagle dogs.The other side of maxillary sinus was elevated by the same approach 2 weeks later.Both Bio-Oss particles and ingrown tissueswere harvested 2 weeks after the second operation.Total RNA was extracted and microarray analysis was carried out to identify the differences of the transcriptome at the 2nd and 4th weeks.Functional classification by gene ontology (GO) and signaling pathway analysis by Kyoto Encyclopedia of Genes and Genomes (KEGG) were applied to determine the differentially expressed genes.RESULTS:The genes up-regulated at the 2nd week were associated with stimulus response,immuno-inflammatory response and cell proliferation.Moreover,the genes related to neuron differentiation,osteoclast differentiation and early vascularization process were also up-regulated at this time point.At the 4th week,up-regulated genes were associated with cellular metabolism and development,importantly including angiogenesis and skeletal development.The genes associated with skeletal development were growth and differentiation factors [transforming growth factor beta 1 (TGFB1) and bone morphogenetic protein 4 (BMP4)],extracellular matrix proteins [chondroadherin (CHAD),collagen type Ⅰ,alpha 1 chain (COL1A1),collagen type Ⅰ,alpha 2 chain (COL1A2),collagen type Ⅴ,alpha 2 chain (COL5A2),decorin (DCN),osteonectin (SPARC) and osteocalcin (BGLAP)] and so on.Furthermore,it was found that some of the up-regulated genes at the 4th week were associated with the signaling of transforming growth factor beta/bone morphogenetic protein (TGF-β/BMP) shown by KEGG analysis.The upstream genes of wingless-type MMTV integration site family (WNT) signaling,WNTSA and frizzled receptor 9 (FZD9),were up-regulated at the 2nd week,while the important downstream gene catenin beta 1 (CTNNB1) was up-regulated at the 4th week.CONCLUSION:After 2 weeks of maxillary sinus floor elevation by the lateral window approach,the transcriptome is characterized by down-regulation of the genes associated with immuno-inflammatory response and bone resorption,and up-regulation of the genes associated with osteogenesis.Neurogenesis and angiogenesis are closely related to bone regeneration.The TGF-β/BMP and WNT signaling pathways play an important role in the bone regeneration process.The effect of WNT signaling is complicated.