Novel Aptamer-Functionalized Nanoparticles Enhances Bone Defect Repair By Improving Stem Cell Recruitment

Abstract

Meng Wang,1 Haibin Wu,2,3 Qiao Li,4 Ying Yang,2 Fengyu Che,2 Guoxia Wang,2 Liyu Zhang2,3 1Department of Orthopaedics, The NO. 946 Hospital of PLA, YiNing, XinJiang 86-835000, People’s Republic of China; 2Shaanxi Institute of Pediatric Diseases, Xi’an Children’s Hospital, Xi’an, Shaanxi 86-710003, People’s Republic of China; 3Key Laboratory of Environment and Genes Related to Diseases, Ministry of Education, Xi’an Jiaotong University Health Science Center, Xi’an, Shaanxi 86-710061, People’s Republic of China; 4Clinical Laboratory, Xi’an Children’s Hospital, Xi’an, Shaanxi 86-710003, People’s Republic of ChinaCorrespondence: Liyu ZhangShaanxi Institute of Pediatric Diseases, Xi’an Children’s Hospital, 69 West JuYuan Alley, LianHu, Xi’an, Shaanxi 86-710003, People’s Republic of ChinaTel [phone]Fax [phone]Email [email protected]: The restoration and repair method in the clinic of delayed fracture healing and non-union after comminuted fractures are urgently needed to improve the prognosis of patients. The recruitment of endogenous stem cells has been considered a promising approach in bone defect repair.Propose: The aim of this study was to generate a de novel MSCs aptamer and developed the first, feasible, economical, bio-compatible, and functional MSCs aptamer-directed nanoparticles without complex manufacture to recruit mesenchymal stem cells (MSCs) for bone defect regeneration.Methods: Whole-cell SELEX was used to generate a de novel MSCs aptamer. Flow cytometry was applied to assess the binding specificities, affinities and sorting abilities of the aptamers. Nano-Aptamer Ball (NAB) was constructed by NHS/EDC reaction. The diameter and zeta of NAB were assessed by dynamic light scattering. CCK8 assay was utilized to evaluate whether NAB could cause non-specific cytotoxicity and induce cell proliferation. To evaluate the bone repair capacity of NAB, histomorphological staining, alizarin red and micro X-ray were used to observe the repair degree of defect in vivo. ELISA was used to detect osteopontin (OPN), osteocalcin (BGP) by, and alkaline phosphatase (ALP) in peripheral blood.Results: MSCs aptamer termed as HM69 could bind with MSCs with high specificity and Kd of 9.67 nM, while has minimal cross-reactivities to other negative cells. HM69 could capture MSCs with a purity of >89%. In vitro, NAB could bind and capture MSCs effectively, whereas did not cause obvious cytotoxicity. In vivo, serum OPN, BGP, and ALP levels in the NAB group of rats were increased at both 2 and 4 weeks, indicating the repair and osteogenesis generation. The healing of bone defects in the NAB group was significantly better than control groups, the defects became blurred, and local trabecular bone growth could be observed in X-ray. The organized hematoma and cell growth in the bone marrow of the NAB group were more vigorous in bone sections staining.Conclusion: These suggested that HM69 and HM69-functionalized nanoparticles NAB exhibited the ability to recruit MSCs both in vitro and in vivo and achieved a better outcome of bone defect repair in a rat model. The findings demonstrate a promising strategy of using aptamer-functionalized bio-nanoparticles for the restoration of bone defects via aptamer-introduced homing of MSCs.Keywords: comminuted fractures, bone defect, aptamer, mesenchymal stem cells, nanoparticles

Keywords

• comminuted fractures
• bone defect
• aptamer
• mesenchymal stem cells
• nanoparticles