Lentivirus-mediated short hairpin RNA interference targeting TNF-alpha in macrophages inhibits particle-induced inflammation and osteolysis in vitro and in vivo.
Lentivirus-mediated short hairpin RNA interference targeting TNF-alpha in macrophages inhibits particle-induced inflammation and osteolysis in vitro and in vivo.
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慢病毒介导的短发夹 RNA 干扰靶向巨噬细胞中的 TNF-α，可在体外和体内抑制颗粒诱导的炎症和骨溶解。
DOI：
10.1186/s12891-016-1290-6
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发表时间：
2016-10-18
影响因子：
2.3
通讯作者：
Ding Y
中科院分区：
文献类型：
--
作者：
Qin CQ;Huang DS;Zhang C;Song B;Huang JB;Ding Y
无菌性松动是关节植入物寿命的重要障碍。假体磨损颗粒被认为在假体周围骨溶解的发生和发展中起核心作用，导致假体无菌性松动。我们研究了慢病毒介导的针对肿瘤坏死因子-α基因的短发夹状RNA在体外和体内对通过巨噬细胞的颗粒诱导的炎症和溶骨改变的抑制作用。筛选和鉴定了一个针对四个候选小鼠肿瘤坏死因子-α基因的siRNA序列，并在体外转录。为了促进肿瘤坏死因子-α-siRNA的高效表达，构建了表达短发夹状RNA(ShRNA)的慢病毒载体。将慢病毒介导的短发夹状RNA导入小鼠巨噬细胞系RAW 264.7。慢病毒转导后24小时加入陶瓷和钛颗粒刺激细胞。用实时荧光定量聚合酶链式反应和酶联免疫吸附试验检测肿瘤坏死因子-α的表达，以mRNA和蛋白水平表示。将慢病毒介导的shRNA混悬液局部注射到小鼠颅骨模型中，然后局部注射颗粒。采用多层螺旋CT扫描，通过测量颅缝宽度评价颅骨溶解情况。巨噬细胞与颗粒共培养时出现伪足。慢病毒介导的shRNA在RAW 264.7中有效地抑制了肿瘤坏死因子-α在基因和蛋白水平的表达。多层螺旋CT扫描显示，慢病毒介导的shRNA明显抑制了小鼠颅骨的骨溶解。我们的研究结果强调，慢病毒介导的针对肿瘤坏死因子-α基因的shRNA在体外和体内都成功地抑制了颗粒诱导的炎症和溶骨性改变。因此，慢病毒介导的基因治疗可能为无菌性关节松动提供一种新的治疗途径。
Aseptic loosening is a significant impediment to joint implant longevity. Prosthetic wear particles are postulated to play a central role in the onset and progression of periprosthetic osteolysis, leading to aseptic loosening of the prosthesis. We investigated the inhibitory effects of a lentivirus-mediated short hairpin RNA that targets the TNF-alpha gene on the particle-induced inflammatory and osteolytic changes via macrophages both in vitro and in vivo. An siRNA sequence targeting the mouse TNF-alpha gene from four candidates, transcribed in vitro, was screened and identified. A lentivirus vector expressing short hairpin RNA (shRNA) was then constructed in order to facilitate efficient expression of TNF-alpha-siRNA. Lentivirus-mediated shRNA was transduced into cells of the mouse macrophage line RAW 264.7. Ceramic and titanium particles were introduced 24 h after lentivirus transduction to stimulate cells. TNF-alpha expression, represented by both mRNA and protein levels, was quantified with real-time PCR and ELISA at all time intervals. Lentivirus-mediated shRNA suspension was locally administered into the murine calvarial model, followed by local injection of particles. A multi-slice spiral CT scan was used to evaluate the osteolysis of the calvaria by detecting the width of the cranial sutures. Macrophages developed pseudopods when co-cultured with particles. Lentivirus-mediated shRNA was shown to effectively inhibit the expression of TNF-alpha at both the mRNA and protein levels in RAW 264.7. The multi-slice spiral CT scan showed that the lentivirus-mediated shRNA significantly suppressed osteolysis of mouse calvaria. Our investigation highlighted the results that lentivirus-mediated shRNA targeting the TNF-alpha gene successfully inhibited particle-induced inflammatory and osteolytic changes both in vitro and in vivo. Therefore, lentivirus-mediated gene therapy may provide a novel therapeutic approach to aseptic joint loosening.
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影响因子：
3.2
作者：
Escors, David;Breckpot, Karine
通讯作者：
Breckpot, Karine
影响因子：
1.1
作者：
Goodman, Stuart B
通讯作者：
Goodman, Stuart B
影响因子：
3.3
作者：
Froelich S;Tai A;Wang P
通讯作者：
Wang P
影响因子：
3.7
作者：
Vicentini FT;Borgheti-Cardoso LN;Depieri LV;de Macedo Mano D;Abelha TF;Petrilli R;Bentley MV
通讯作者：
Bentley MV
影响因子：
6
作者：
Merkel, KD;Erdmann, JM;Teitelbaum, SL
通讯作者：
Teitelbaum, SL