title: The role of miR-135-modified adipose-derived mesenchymal stem cells in bone regeneration.
abstract: Tissue-engineering technology employing genetically-modified mesenchymal stem cells combined with proper scaffolds represents a promising strategy for bone regeneration. Elucidating the underlying mechanisms that govern the osteogenesis of mesenchymal stem cells will give deeper insights into the regulatory patterns, as well as provide more effective methods to enhance bone regeneration. In this study, miR-135 was identified as an osteogenesis-related microRNA that was up-regulated during the osteogenesis of rat adipose-derived stem cells (ADSCs). Gain- and loss-of-function experiments using a lentiviral expression system showed that Homeobox A2 (Hoxa2) was negatively regulated by miR-135, and luciferase reporter assay further indicated that miR-135 repressed Hoxa2 expression through binding to the 3'-untranslated region (3'-UTR) of the Hoxa2 mRNA. In vitro analyses showed that the overexpression of miR-135 significantly enhanced the expression of bone markers and extracellular matrix calcium deposition, whereas the knockdown of miR-135 suppressed these processes. Transduced ADSCs were then combined with poly(sebacoyl diglyceride) (PSeD) scaffold to repair a critical-sized calvarial defects in rats. The results showed that the overexpression of miR-135 significantly promoted new bone formation with higher bone mineral density (BMD) and number of trabeculae (Tb.N), as well as larger areas of newly formed bone and mineralization labeled by tetracycline, calcein and alizarin red. In contrast, the knockdown of miR-135 attenuated these processes. Additionally, immunohistochemical analyses showed that transduced ADSCs participated in new bone formation and a miR-135/Hoxa2/Runx2 pathway might contribute to the regulation of ADSC osteogenesis and bone regeneration. Taken together, our data suggested that miR-135 positively regulated the osteogenesis and bone regeneration of ADSCs both in vitro and in vivo. Thus, the combination of miR-135-modified ADSCs and the PSeD scaffold may serve as a promising and effective method to repair critical-sized bone defects.
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China.
authors.forename: Qing
authors.initials: Q
authors.lastname: Xie
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China.
authors.forename: Zi
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China.
authors.forename: Huifang
authors.initials: H
authors.lastname: Zhou
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China.
authors.forename: Zhang
authors.initials: Z
authors.lastname: Yu
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China.
authors.forename: Yazhuo
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China.
authors.forename: Hao
authors.initials: H
authors.lastname: Sun
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China.
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Bi
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China.
authors.forename: Yefei
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China.
authors.forename: Wodong
authors.initials: W
authors.lastname: Shi
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China. Electronic address: guping2009@hotmail.com.
authors.forename: Ping
authors.initials: P
authors.lastname: Gu
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, PR China. Electronic address: fanxq@sh163.net.
authors.forename: Xianqun
authors.initials: X
authors.lastname: Fan
mesh_terms: D000273:Adipose Tissue / Q000166:cytology*; D000818:Animals; D001483:Base Sequence; D015415:Biomarkers / Q000378:metabolism; D001861:Bone Regeneration; D002113:Calcification, Physiologic; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes / Q000378:metabolism; D005786:Gene Expression Regulation; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000648:ultrastructure; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010012:Osteogenesis / Q000235:genetics; D017207:Rats, Sprague-Dawley; D012886:Skull / Q000000981:diagnostic imaging / Q000473:pathology; D013194:Staining and Labeling; D054457:Tissue Scaffolds / Q000737:chemistry; D055114:X-Ray Microtomography
chemical_list: D015415:Biomarkers; D005456:Fluorescent Dyes; D018398:Homeodomain Proteins; C000600409:MIRN135 microRNA, rat; D035683:MicroRNAs
keywords: ADSCs; Bone regeneration; Osteogenesis; PSeD; miR-135