PKD1通过IL-6调控体内引导骨膜成骨过程以修复大段骨缺损及其机制研究

The requirement of large segment bone defects are still the major challenge in medicine. The way of in-vivo bone tissue engineering has been gradually accepted, and bring hope for translational medicine. In previously study, our group have first report and successfully constructed large segment osteoid tissue that has the morphology and function of bone by the technique of “guided periosteum osteogenesis” by pig in vivo (Sci rep, 2016). However, the mechanism of this process is unclear, and it limited of long period of guided bone generation. We have found the high expression PKD1 and IL-6, in the periosteal cells of guided periosteum osteogenesis group with the help of high throughput sequencing technology and proteomics test in pre-experiment. Meanwhile, the p38-MAPK and STAT3 pathways of IL-6 upstream were significantly enriched. As the latest reference report, PKD1 regulated in the process of embryonic bone development just through the pathway of p38-MAPKand STAT3. On this basis, to promote “PKD1 regulates the process of in-vivo guided periosteum osteogenesis through IL-6” as the hypothesis, by means of tissue microarray, lentivirus cell model, and protein chip method, our group will verify the important role of PKD1 in the progress of in-vivo guided periosteum osteogenesis; explore the relevance of the specific transduction signaling pathways of PKD1 and IL-6 to clarify the mechanism. This study will help to deepen the understanding of the process of in-vivo guided “periosteum-bone regeneration chamber” osteogenesis, and provide the specific intervention targets and experimental basis for clinical treatment.

大段骨缺损的修复始终是临床治疗难题。近年来体内组织工程构建方式渐受认可，为临床转化带来希望。前期本课题组率先成功引导猪肋骨骨膜成骨，于体内构建出具有骨组织学形态及功能的大段骨组织（Sci rep,2016）。但此过程机制尚不明确且成骨周期较长。为此，预实验通过高通量测序及差异蛋白组学检测发现，引导骨膜成骨组PKD1、IL-6高表达，而剩余上调基因主要富集于IL-6的上游p38-MAPK和STAT3等通路。最新研究报道PKD1恰可通过p38-MAPK和STAT3途径调控胚胎骨发育过程，因此本项目将以“PKD1通过IL-6调控体内引导骨膜成骨过程”为假说，借助组织芯片、慢病毒干扰及蛋白质液相质谱等技术，多维度验证PKD1在体内引导骨膜成骨过程中的作用；探究PKD1与IL-6在此过程中的特异性关联以明确机制。从而加深对体内引导“骨膜-骨再生室”成骨过程的认识，为临床提供特异性干预靶点及实验依据。

大段骨缺损的修复是临床治疗难题，始终缺乏令人满意的治疗手段。本课题组前期率先在猪体内，成功引导新生了具有骨组织学形态及功能的大段骨组织，并用于缺损修复。.为了进一步明确该过程的发生机制，本研究通过高通量测序及单细胞测序检测发现，引导骨膜成骨过程的早期，体内主要启动补体活化、细胞趋化及骨组织新生等生物学功能通路调控成骨过程，在这期间骨膜干细胞与免疫细胞的细胞通讯发挥了重要作用，例如IL10、IL18等重要免疫因子均承担了一定角色；后期主要以上调骨组织塑形和重建通路为主导，期间成骨细胞与破骨细胞的动态平衡发挥了重要作用。.另外，项目组通过测序数据发现在骨膜引导成骨的早期过程中，SFRP2显著上调，并对此进行了基因以及蛋白水平的验证。通过组织芯片、慢病毒干扰等技术，验证了SFRP2在体内外引导骨膜成骨的过程中，具有影响骨膜细胞、BMSC成骨的作用。以上研究结果明确了引导骨膜成骨过程中，SFRP2起到了调控成骨的关键作用。该研究加深了对体内引导“骨膜-骨再生室”成骨过程的认识，并为临床提供特异性干预靶点及实验依据。