活性维生素D3联合力学拉伸对成骨细胞MC3T3-E1增殖、分化及OPG和RANKL表达的影响

目的体外模拟成骨细胞在体内的生存环境,考察活性维生素D3(VD3)、力学拉伸以及两者联合对成骨细胞MC3T3-E1增殖、分化及破骨细胞抑制因子(OPG)和破骨细胞分化因子(RANKL)表达的影响。方法将10 nmol/L VD3、间断性力学拉伸以及两者联合作用于成骨细胞。流式细胞术检测细胞增殖。荧光探针试剂盒检测碱性磷酸酶(ALP)活性。实时定量PCR检测核心转录因子Runx2、OPG、RANKL mRNA水平,Western blotting检测其蛋白表达。结果 VD3抑制成骨细胞增殖,力学拉伸不能改变这种抑制效应。力学拉伸和VD3单独作用成骨细胞均能增加ALP活性及提高ALP、Runx2 mRNA水平,但当联合刺激后这些指标均降低,且成强度依赖性。力学拉伸增加OPG/RANKL比值,增强成骨作用,联合VD3后,OPG/RANKL比值下降。结论力学拉伸能有效诱导成骨分化,增加骨形成。VD3与力学拉伸联合抑制成骨细胞增殖和分化,并通过增加RANKL表达而影响骨重建。研究结果为骨质疏松及相关骨疾病的理论和治疗提供有意义的探索。     

活性维生素D3联合力学拉伸对成骨细胞MC3T3-E1增殖、分化及OPG和RANKL表达的影响（英文）

目的体外模拟成骨细胞在体内的生存环境,考察活性维生素D3(VD3)、力学拉伸以及两者联合对成骨细胞MC3T3-E1增殖、分化及破骨细胞抑制因子(OPG)和破骨细胞分化因子(RANKL)表达的影响。方法将10 nmol/L VD3、间断性力学拉伸以及两者联合作用于成骨细胞。流式细胞术检测细胞增殖。荧光探针试剂盒检测碱性磷酸酶(ALP)活性。实时定量PCR检测核心转录因子Runx2、OPG、RANKL mRNA水平,Western blotting检测其蛋白表达。结果 VD3抑制成骨细胞增殖,力学拉伸不能改变这种抑制效应。力学拉伸和VD3单独作用成骨细胞均能增加ALP活性及提高ALP、Runx2 mRNA水平,但当联合刺激后这些指标均降低,且成强度依赖性。力学拉伸增加OPG/RANKL比值,增强成骨作用,联合VD3后,OPG/RANKL比值下降。结论力学拉伸能有效诱导成骨分化,增加骨形成。VD3与力学拉伸联合抑制成骨细胞增殖和分化,并通过增加RANKL表达而影响骨重建。研究结果为骨质疏松及相关骨疾病的理论和治疗提供有意义的探索。     

Apelin对小鼠MC3T3-E1成骨样细胞RANKL/OPG表达的影响

目的探讨脂肪因子Apelin对小鼠MC3T3-E1成骨样细胞RANKL/OPG表达的影响,为Apelin参与调节骨代谢提供理论依据。方法用real time PCR检测RANKL/OPG m RNA的表达,用Western blot方法检测RANKL/OPG蛋白的表达。结果不同浓度Apelin(0.4n M,1n M,10n M)干预MC3T3-E1细胞48h后,与对照组相比可明显促进OPG m RNA及蛋白的表达(P0.01),同时可明显抑制RANKL m RNA及蛋白的表达(P0.01),其中当实验组Apelin浓度为1n M时,效果最明显。结论 Apelin可作用于MC3T3-E1成骨样细胞,从而影响RANKL/OPG的表达,为骨质疏松的防治提供新靶点。     

人多发性骨髓瘤RPMI 8226细胞条件培养液诱导破骨前体细胞的分化成熟

目的: 探讨人多发性骨髓瘤RPMI 8226细胞条件培养液对破骨前体细胞RAW264.7的影响。方法: Western blotting检测可溶性核因子κB 受体激活剂配体(soluble receptor activator of NF-κB ligand, sRANKL)的蛋白表达。抗酒石酸酸性磷酸酶(tartrate－resistant acid phosphatase, TRAP)染色观察RAW264.7细胞的分化成熟情况。RT-PCR法检测RAW264.7细胞TRAP和cathepsin K mRNA的表达。结果: Western blotting法证实RPMI 8226细胞条件培养液中含有sRANKL。TRAP染色发现RPMI 8226细胞条件培养液能诱导RAW264.7细胞分化为TRAP阳性的多核成熟破骨细胞。人抑制性RANKL单克隆抗体拮抗30%RPMI 8226细胞条件培养液中sRANKL的作用,且具有剂量依赖性。30% RPMI 8226细胞条件培养液能刺激RAW264.7细胞上调TRAP和cathepsin K mRNA表达。结论: 人多发性骨髓瘤细胞RPMI 8226条件培养液中的sRANKL具有促RAW264.7破骨前体细胞分化成TRAP阳性的多核成熟破骨细胞的生物活性。人抑制性RANKL单克隆抗体阻断30%RPMI 8226细胞条件培养液中sRANKL的诱导分化作用,且具有浓度依赖性。     

双氢青蒿素抑制核因子κB受体激动剂配体诱导RAW264.7细胞分化破骨细胞的研究

目的 探讨双氢青蒿素(DA)对核因子κB受体激动剂配体(RANKL)诱导RAW264.7细胞形成破骨细胞的影响。方法 通过细胞计数试剂盒(CCK-8)确定不同浓度(1、5、10、50、100、200 μmol/L)DA对RAW264.7细胞的生存影响。分别用50 ng/mL RANKL及50 ng/mL RANKL+5、10、50、100 μmol/L DA诱导RAW264.7细胞形成破骨细胞,共3 d。对形成的破骨细胞用抗酒石酸酸性磷酸酶(TRAP)染色并计数,TRAP染色阳性且细胞核数目＞3个认为是成熟的破骨细胞。50 ng/mL RANKL及50 ng/mL RANKL+10、100 μmol/L DA培养RAW264.7细胞24 h,用Trizol试剂提取总RNA,并使用荧光实时定量PCR检测破骨细胞分化相关基因NFATc1、c-fos及Cathepsin K的表达。结果 1、5、10、50、100 μmol/L DA对RAW264.7细胞毒性较小,细胞存活率均＞90%。TRAP染色显示,5、10、50、100 μmol/L DA可以减少RANKL诱导成熟破骨细胞的数目,并呈剂量依赖关系(F=139.156, P＜0.01)。实时荧光定量PCR结果显示,10、100 μmol/L DA具有下调破骨细胞分化关键基因NFATc1和c-fos表达的作用,且100 μmol/L抑制作用比10 μmol/L明显,但两种浓度DA对Cathepsin K的表达无明显影响。结论 DA对RAW 264.7细胞毒性较低,通过下调RAW 264.7细胞NFATc1和c-fos基因表达抑制RANKL诱导破骨细胞形成。DA可以作为治疗骨质破坏性疾病的潜在药物。     

瘦素通过抑制PPARγ的表达抑制RAW264.7巨噬细胞向破骨细胞分化

目的探讨瘦素对可溶性核因子κB配体的受体活化因子(sRANKL)诱导RAW264.7巨噬细胞向破骨细胞分化的影响及其机制。方法抗酒石酸酸性磷酸酶(TRAP)染色检测瘦素对sRANKL诱导RAW264.7向破骨细胞分化的形态学影响;实时荧光定量PCR检测TRAP及过氧化物酶体增殖物激活受体γ(PPARγ)的mRNA表达;Western blot法检测TRAP及PPARγ的蛋白表达。结果瘦素和sRANKL联合处理组与单独sRANKL组相比,TRAP染色破骨细胞数目明显减少;TRAP的mRNA和蛋白表达量明显降低;PPARγ的mRNA和蛋白表达量明显降低,且与瘦素浓度呈梯度相关。结论瘦素可能通过抑制PPARγ的表达来抑制RAW264.7细胞向破骨细胞分化。     

CCR1参与肺癌分泌因子诱导破骨细胞生成

目的建立肺癌细胞与单核/巨噬细胞间接共培养模型,观察在肺癌细胞条件培养基作用下抑制前体破骨细胞(RAW264.7)的CCR1对破骨细胞生成的影响。方法收集肺癌细胞A549的条件培养基,将条件培养基以不同浓度加入RAW264.7培养基中共培养,在共培养体系中加或不加CCR1特异性拮抗剂BX471。细胞培养至第5天进行TRAP染色,提取不同条件作用后RAW264.7细胞的总RNA,采用Real-time PCR检测CCR1及破骨细胞标志基因cathepsin K、TRAP mRNA的表达,使用细胞免疫荧光和Western blot检测不同时期RAW264.7细胞中CCR1及Cathepsin K蛋白的表达量。结果加入A549细胞条件培养基的RAW264.7细胞TRAP阳性细胞显著增多,条件培养基明显上调RAW264.7的CCR1、Cathepsin K、TRAP的表达且呈浓度依赖,CCR1特异性拮抗剂BX471可抑制肺癌分泌因子对破骨细胞的活化。结论肺癌细胞分泌因子可促进破骨前体细胞向破骨细胞分化,CCR1参与了肺癌细胞分泌因子对破骨细胞的活化过程。     

乌司他丁对骨水泥颗粒诱导MC3T3-E1鼠前成骨细胞凋亡的干预作用

背景:前期研究发现,乌司他丁对RANKL诱导RAW264.7细胞活化为成熟破骨细胞具有一定的抑制作用,并可降低基质金属蛋白酶9的表达,但其对聚甲基丙烯酸甲酯骨水泥促成骨细胞凋亡效应是否有干预作用尚不清楚。目的:探讨乌司他丁对骨水泥颗粒诱导MC3T3-E1鼠前成骨细胞凋亡及Ⅰ型胶原、骨钙素、基质金属蛋白酶2 mRNA表达的影响。方法:取第六七代生长状态良好的MC3T3-E1鼠前成骨细胞,分4组培养,骨水泥组加入1 g/L的聚甲基丙烯酸甲酯骨水泥悬液;高、低剂量乌司他丁组在加入1 g/L的聚甲基丙烯酸甲酯骨水泥悬液后,再分别加入5 000,500 U/mL的乌司他丁;设置单独培养的细胞为空白对照。MTT法检测细胞增殖活性,茜素红染色检测细胞矿化程度,流式细胞仪检测细胞凋亡率,半定量RT-PCR检测细胞中Ⅰ型胶原、骨钙素、基质金属蛋白酶2 mRNA的表达。结果与结论:与空白对照组比较,骨水泥抑制了MC3T3-E1细胞增殖活性(P0.05),促进了细胞凋亡(P0.05),在加入不同浓度乌司他丁后,细胞增殖活性逐渐增强(P0.05),凋亡率降低(P0.05),且呈剂量时间依赖关系;骨水泥降低了细胞中Ⅰ型胶原、骨钙素表达(P0.05),升高了基质金属蛋白酶2表达(P0.05),在加入5 000 U/mL乌司他丁后,细胞基质金属蛋白酶2表达降低,Ⅰ型胶原、骨钙素表达升高(P0.05)。骨水泥组无矿化结节,其他组均有矿化结节形成。结果表明乌司他丁对骨水泥诱导MC3T3-E1鼠前成骨细胞凋亡具有一定的抑制作用,可促进成骨细胞中Ⅰ型胶原、骨钙素的生成,并降低基质金属蛋白酶2的表达水平。     

建立小鼠成骨与破骨细胞相互作用的共育新体系

背景：众所周知,骨重建是骨组织中重要的生物学反应过程,其中成骨细胞与破骨细胞发挥了关键作用。但目前,关于骨重建中成骨与破骨细胞间信号传递的深层机制还不清楚。 目的：利用transwell技术,在体外建立一种成骨与破骨细胞的新型共育体系,为深入研究骨重建中成骨与破骨细胞的相互作用提供成熟的实验模型。 方法：采用MC3T3-E1成骨样细胞株与RAW264.7破骨前体细胞株,进行体外成骨与破骨细胞的诱导分化,并利用Transwell共培养板(0.4 µm聚酯膜)建立成骨与破骨细胞的共育体系。共培养6 d后,通过测定细胞活性和碱性磷酸酶(ALP)活力分析成骨细胞的增殖和分化活性,利用抗酒石酸酸性磷酸酶(TRAP)染色、甲苯胺蓝(TB)染色、TRAP活性测定及扫描电镜技术观察破骨细胞的分化及骨吸收功能。 结果与结论：共培养体系中成骨样细胞的无限增殖能力减弱,而分化活性明显增强,同时破骨前体细胞被诱导分化为成熟的破骨细胞,并具有一定的骨吸收功能。因此,该共培养体系可用于骨重建中成骨与破骨细胞间信号通路的深层研究。     

Asxl1在炎性微环境中对成骨细胞增殖分化的作用

背景:研究表明Asxl1的缺失可导致骨质发育不全、骨质缺损类疾病的发生,但目前在根尖周炎环境下该因子与骨破坏之间的关系暂无相关报道。目的:探讨炎性微环境下Asxl1对成骨细胞增殖分化的影响。方法:实验选用脂多糖刺激MC3T3-E1细胞建立体外炎性微环境,通过CCK-8实验筛取脂多糖最佳质量浓度和最佳作用时间,然后用20 mg/L脂多糖刺激MC3T3-E1细胞24 h,免疫荧光检测Asxl1的蛋白表达水平,Real Time-PCR检测Asxl1 mRNA的表达水平。为进一步验证Asxl1基因在炎性微环境中影响成骨细胞的增殖与分化,脂多糖刺激形成炎性微环境后转染Asxl1-SiRNA 24 h,采用CCK-8检测细胞增殖活性,RealTime-PCR检测Asxl1及成骨相关基因ALP和RUNX2 mRNA的表达水平。结果与结论:①脂多糖刺激MC3T3-E1细胞后,Asxl1蛋白和mRNA表达水平呈降低趋势;②脂多糖刺激MC3T3-E1细胞后,转染Asxl1-SiRNA 24 h,细胞增殖活性下降趋势明显,Asxl1基因及成骨相关基因ALP和RUNX2 mRNA的表达水平明显降低;③结果提示,Asxl1可能通过参与炎性反应过程,影响成骨细胞的增殖与分化,进而参与骨破坏进程。     

Different effects of carbon ion and γ-irradiation on expression of receptor activator of NF-kB ligand in MC3T3-E1 osteoblast cells

We investigated the effects of carbon ion and γ-irradiation on osteoblastic MC3T3-E1 cells by comparing mRNA expression levels for RANKL and osteoprotegerin by RT-PCR. MC3T3-E1 cells were irradiated with 2, 4, or 6 Gy of carbon ions or γ-rays, and total RNA was harvested 1, 2, 3, 5, or 7 days after irradiation. The RANKL mRNA/OPG mRNA ratio in carbon ion-irradiated MC3T3-E1 cells was lower, while in γ-irradiated MC3T3-E1 cells this ratio was higher than in non-irradiated cells. To evaluate osteoclastogenesis of MC3T3-E1 cells, carbon ion-or γ-irradiated cells were co-cultured with non-irradiated cells from murine bone marrow. Staining for tartrate-resistant acid phosphatase (TRAP) in co-cultures showed that carbon ion irradiation suppressed osteoclastogenesis. This result is consistent with the lower RANKL/OPG mRNA ratio for carbon ion-irradiated cells. These results suggest that carbon ion irradiation acts primarily on osteoblastic cells, leading to a decrease in the RANKL/OPG mRNA ratio. This effect, in turn, leads to a decrease in osteoclastogenesis and osteoclast activity, which results in an increase in bone volume. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 142, No. 11, pp. 566–572, November, 2006     

Compressive Force Induces Osteoclast Differentiation via Prostaglandin E2 Production in MC3T3-E1 Cells

In orthodontic tooth movement, prostaglandin E₂ (PGE₂) released from osteoblasts can alter the normal process of bone remodeling. We previously showed that compressive force (CF) controls bone formation by stimulating the production of PGE₂ and Ep2 and/or Ep4 receptors in osteoblasts. The present study was undertaken to examine the effect of CF on the production of PGE₂, cyclooxygenase-2 (COX-2), macrophage colony-stimulating factor (M-CSF), receptor activator of NF-κB ligand (RANKL), and osteoprotegerin (OPG) using osteoblastic MC3T3-E1 cells and to examine the indirect effect of CF on osteoclast differentiation using RAW264.7 cells as osteoclast precursors. MC3T3-E1 cells were cultured with or without continuous CF (1.0 or 3.0 g/cm²) for 24 hr, and PGE₂ production was determined using ELISA. The expression of COX-2, M-CSF, RANKL, and OPG genes and proteins was determined using real-time PCR and ELISA, respectively. Osteoclast differentiation was estimated using tartrate-resistant acid phosphatase (TRAP) staining of RAW 264.7 cells cultured for 10 days with conditioned medium from CF-treated MC3T3-E1 cells and soluble RANKL. As CF increased, PGE₂ production and the expression of COX-2, M-CSF, and RANKL increased, whereas OPG expression decreased. The number of TRAP-positive cells increased as CF increased. Celecoxib, a specific inhibitor of COX-2, blocked the stimulatory effect of CF on TRAP staining and the production of PGE₂, M-CSF, RANKL, and OPG. These results suggest that CF induces osteoclast differentiation by increasing M-CSF production and decreasing OPG production via PGE₂ in osteoblasts.     

Expression of osteoprotegerin and RANK ligand in breast cancer bone metastasis

Bone destruction is primarily mediated by osteoclastic bone resorption, and cancer cells stimulate the formation and activation of osteoclasts next to metastatic foci. Accumulating evidences indicate that receptor activator of NF-kB ligand (RANKL) is the ultimate extracellular mediator that stimulates osteoclast differentiation into mature osteoclasts. In contrast, osteoprotegerin (OPG) inhibits osteoclast development. In order to elucidate a mechanism for cancer-induced osteoclastogenesis, cells from a human breast cancer line, MDA-MB-231, were directly co-cultured with ST2, MC3T3-E1, or with primary mouse calvarial cells. Osteoclast-like cells and tartarate resistant acid phosphatase (TRAP) activities were then quantitated. We examined these cell lines and samples from breast cancer by RT-PCR for the expressions of OPG and RANKL mRNA. Compared to controls, co-culture of MDA-MB-231 cells with stromal or osteoblastic cells induced an increase in number of osteoclasts and TRAP activities. MDA-MB-231 cells alone or breast cancer samples did not express RANKL mRNA. However, co-culture of these cancer cells with stromal or osteoblastic cells induced RANKL mRNA expression and decreased OPG mRNA expression. These experiments demonstrate that direct interactions between breast cancer and stromal or osteoblastic cells induce osteoclastogenesis in vitro through modulating RANKL expression.     

小分子肽对成骨前体细胞MC3T3-E1骨保护素和核转录因子κB受体活化因子配体表达的影响

背景：体内实验显示,小分子肽能明显增加去卵巢大鼠的骨钙含量,使其骨密度增加,能很好地预防骨质疏松。同时体外实验显示,小分子肽能促进小鼠成骨细胞和成骨前体细胞MC3T3-E1增殖、分化、矿化,并且可能是通过抑制核转录因子 p50和p65的表达来起作用。而小分子肽对骨保护素/核转录因子κB受体活化因子配体的影响尚不明确。 目的：观察小分子肽对MC3T3-E1在增殖、分化、矿化过程中骨保护素和RANKL表达的影响。 方法：以体积分数10%胎牛血清的DMEM培养液为空白对照组,50,100 mg/L质量浓度小分子肽作用小鼠成骨前体细胞MC3T3-E1,分别于作用3,6,12,18,24,30 d后,收集细胞提取蛋白,Western Blot检测骨保护素和核转录因子κB受体活化因子配体蛋白的表达。 结果与结论：50,100 mg/L小分子肽作用MC3T3-E1后能明显促进作用骨保护素的表达(P < 0.01),而对核转录因子κB受体活化因子配体无明显影响。小分子肽作用后MC3T3-E1中骨保护素/核转录因子κB受体活化因子配体的比值要明显高于空白对照组(P < 0.01)。因此,认为小分子肽可以通过增加骨保护素的表达来影响骨保护素/核转录因子κB受体活化因子配体系统,间接地抑制破骨细胞的数量和功能。     

Co-culture with periodontal ligament stem cells enhanced osteoblastic differentiation of MC3T3-E1 cells and osteoclastic differentiation of RAW264.7 cells

Objectives: Periodontal ligament stem cells (PDLSCs) are characterized by having multipotential differentiation and immunoregulatory properties, which are the main mechanisms of PDLSCs-mediated periodontal regeneration. Periodontal or bone regeneration requires coordination of osteoblast and osteoclast, however, very little is known about the interactions between PDLSCs and osteoblast-like cells or osteoclast precursors. In this study, the indirect co-culture approach was introduced to preliminarily elucidate the effects of PDLSCs on differentiation of osteoblast-like cells and osteoclast precursors in vitro. Materials and methods: Human PDLSCs were obtained from premolars extracted and their stemness was identified in terms of their colony-forming ability, proliferative capacity, cell surface epitopes and multi-lineage differentiation potentials. A noncontact co-culture system of PDLSCs and preosteoblastic cell line MC3T3-E1 or osteoclast precursor cell line RAW264.7 was established, and osteoblastic differentiation of MC3T3-E1 and osteoclastic differentiation of RAW264.7 were evaluated. Results: PDLSCs exhibited features of mesenchymal stem cells. Further investigation through indirect co-culture system showed that PDLSCs enhanced ALP activity, expressions of ALP, Runx2, BSP, OPN mRNA and BSP, OPN proteins and mineralization matrix deposition in MC3T3-E1. Meanwhile, they improved maturation of osteoclasts and expressions of TRAP, CSTK, TRAF6 mRNA and TRAP, TRAF6 proteins in RAW264.7. Conclusions: PDLSCs stimulates osteoblastic differentiation of osteoblast precursors and osteoclastic differentiation of osteoclast precursors, at least partially, in a paracrine fasion.     

盐酸小檗碱通过PI3K/Akt信号通路促进小鼠前成骨细胞MC3T3-E1的分化

目的:研究盐酸小檗碱对小鼠前成骨细胞系MC3T3-E1分化与矿化的调控作用及其机制。方法:MC3T3-E1细胞给予不同浓度(0、1、5、10和20 mg/L)的盐酸小檗碱刺激3 d,CCK-8法检测细胞活性。不同浓度的盐酸小檗碱分别干预3 d和7 d,检测细胞碱性磷酸酶(ALP)活性。进一步将实验随机分为4组:对照组、盐酸小檗碱组、盐酸小檗碱+LY249002(PI3K/Akt通路抑制剂)组及LY249002组。干预2 d后,采用real-time PCR检测成骨细胞分化相关因子ALP、骨钙素(OCN)、骨桥蛋白(OPN)及Runt相关转录因子2(Runx2)的mRNA表达情况,采用Western blot检测PI3K/Akt信号通路相关蛋白p-Akt的表达水平。将MC3TC-E1细胞用矿化培养基诱导21 d,茜素红染色检测其矿化情况。结果:与对照组相比,不同浓度的盐酸小檗碱对细胞活性的影响没有明显差异;不同浓度的盐酸小檗碱处理MC3T3-E1细胞后ALP活性有不同程度升高。Real-time PCR结果表明,盐酸小檗碱(5 mg/L)促进ALP、OCN、OPN及Runx2的mRNA表达(P 0. 01),而LY294002能抑制这些分化相关因子的表达。Western blot检测结果表明,盐酸小檗碱(5 mg/L)促进p-Akt蛋白的表达(P 0. 01),其作用被LY249002抑制。茜素红染色发现盐酸小檗碱组矿化明显,但LY294002能抑制盐酸小檗碱的促进作用。结论:盐酸小檗碱可以促进小鼠前成骨细胞的分化与矿化,其机制可能与其激活PI3K/Akt信号通路有关。     

周期性牵张力介导BMP9通过PI3K/AKT信号通路调控人牙周膜成纤维细胞成骨分化

目的　研究在周期性牵张力介导下，骨形态发生蛋白9（bone morphogenetic proteins 9，BMP9）能否激活PI3K/AKT信号通路调控人牙周膜成纤维细胞（human periodontal ligament fibroblasts，hPDLFs）成骨分化。 方法　利用FlexCell系统对体外培养的hPDLFs加载正弦波、形变率10%、频率0.5 Hz的周期性牵张力，免疫荧光法检测细胞骨架蛋白的表达及分布，qPCR检测RUNX2、OCN、OPN、OSX mRNA的表达情况，免疫印迹法检测成骨标志蛋白OCN、OPN的表达，加入PI3K信号抑制剂LY294002后AKT、P-AKT以及OCN、OPN的变化。 结果　与对照组比较，6 h、12 h组的细胞骨架蛋白表达增多且呈受力方向分布，OCN、OPN表达上调，差异有统计学意义（P<0.05），qPCR检测mRNA的表达趋势与蛋白检测结果一致。 结论　周期性牵张力介导BMP9可以通过PI3K/AKT信号通路调控hPDLFs成骨分化。