Jiacai He

Blood vessel formation in the tissue-engineered bone with the constitutively active form of HIF-1α mediated BMSCs

The successful clinical outcome of the implanted tissue-engineered bone is dependent on the establishment of a functional vascular network. A gene-enhanced tissue engineering represents a promising approach for vascularization. Our previous study indicated that hypoxia-inducible factor-1α (HIF-1α) can up-regulate the expression of vascular endothelial growth factor (VEGF) and stromal-derived factor 1 (SDF-1) in bone mesenchymal stem cells (BMSCs). The angiogenesis is a co-ordinated process that requires the participation of multiple angiogenic factors. To further explore the angiogenic effect of HIF-1α mediated stem cells, in this study, we systematically evaluated the function of HIF-1α in enhancing BMSCs angiogenesis in vitro and in vivo. A constitutively active form of HIF-1α (CA5) was inserted into a lentivirus vector and transduced into BMSCs, and its effect on vascularization and vascular remodeling was further evaluated in a rat critical-sized calvarial defects model with a gelatin sponge (GS) scaffold. The expression of the key angiogenic factors including VEGF, SDF-1, basic fibroblast growth factor (bFGF), placental growth factor (PLGF), angiopoietin 1 (ANGPT1), and stem cell factor (SCF) at both mRNAs and proteins levels in BMSCs were significantly enhanced by HIF-1α overexpression compared to the in vitro control group. In addition, HIF-1α-over expressing BMSCs showed dramatically improved blood vessel formation in the tissue-engineered bone as analyzed by photography of specimen, micro-CT, and histology. These data confirm the important role of HIF-1α in angiogenesis in tissue-engineered bone. Improved understanding of the mechanisms of angiogenesis may offer exciting therapeutic opportunities for vascularization, vascular remodeling, and bone defect repair using tissue engineering strategies in the future.

Repair of Critical-Sized Rat Calvarial Defects Using Genetically Engineered Bone Marrow-Derived Mesenchymal Stem Cells Overexpressing Hypoxia-Inducible Factor-1ᆇ§

The processes of angiogenesis and bone formation are coupled both temporally and spatially during bone repair. Bone marrow‐derived mesenchymal stem cells (BMSCs) have been effectively used to heal critical‐size bone defects. Enhancing their ability to undergo angiogenic and osteogenic differentiation will enhance their potential use in bone regeneration. Hypoxia‐inducible factor‐1α (HIF‐1α) has recently been identified as a major regulator of angiogenic‐osteogenic coupling. In this study, we tested the hypothesis that HIF‐1α gene therapy could be used to promote the repair of critical‐sized bone defects. Using lentivirus‐mediated delivery of wild‐type (HIF) or constitutively active HIF‐1α (cHIF), we found that in cultured BMSCs in vitro, HIF and cHIF significantly enhanced osteogenic and angiogenic mRNA and protein expression when compared with the LacZ group. We found that HIF‐1α‐overexpressing BMSCs dramatically improved the repair of critical‐sized calvarial defects, including increased bone volume, bone mineral density, blood vessel number, and blood vessel area in vivo. These data confirm the essential role of HIF‐1α modified BMSCs in angiogenesis and osteogenesis in vitro and in vivo. STEM CELLS 2011;29:1380–1390

The Bone-Forming Effects of HIF-1α-Transduced BMSCs Promote Osseointegration with Dental Implant in Canine Mandible

The presence of insufficient bone volume remains a major clinical problem for dental implant placement to restore the oral function. Gene-transduced stem cells provide a promising approach for inducing bone regeneration and enhancing osseointegration in dental implants with tissue engineering technology. Our previous studies have demonstrated that the hypoxia-inducible factor-1α (HIF-1α) promotes osteogenesis in rat bone mesenchymal stem cells (BMSCs). In this study, the function of HIF-1α was validated for the first time in a preclinical large animal canine model in term of its ability to promote new bone formation in defects around implants as well as the osseointegration between tissue-engineered bone and dental implants. A lentiviral vector was constructed with the constitutively active form of HIF-1α (cHIF). The ectopic bone formation was evaluated in nude mice. The therapeutic potential of HIF-1α-overexpressing canine BMSCs in bone repair was evaluated in mesi-implant defects of immediate post-extraction implants in the canine mandible. HIF-1α mediated canine BMSCs significantly promoted new bone formation both subcutaneously and in mesi-implant defects, including increased bone volume, bone mineral density, trabecular thickness, and trabecular bone volume fraction. Furthermore, osseointegration was significantly enhanced by HIF-1α-overexpressing canine BMSCs. This study provides an important experimental evidence in a preclinical large animal model concerning to the potential applications of HIF-1α in promoting new bone formation as well as the osseointegration of immediate implantation for oral function restoration.

Iron oxide nanoclusters for T 1 magnetic resonance imaging of non-human primates

Iron-oxide-based contrast agents for magnetic resonance imaging (MRI) had been clinically approved in the United States and Europe, yet most of these nanoparticle products were discontinued owing to failures to meet rigorous clinical requirements. Significant advances have been made in the synthesis of magnetic nanoparticles and their biomedical applications, but several major challenges remain for their clinical translation, in particular large-scale and reproducible synthesis, systematic toxicity assessment, and their preclinical evaluation in MRI of large animals. Here, we report the results of a toxicity study of iron oxide nanoclusters of uniform size in large animal models, including beagle dogs and the more clinically relevant macaques. We also show that iron oxide nanoclusters can be used as T1 MRI contrast agents for high-resolution magnetic resonance angiography in beagle dogs and macaques, and that dynamic MRI enables the detection of cerebral ischaemia in these large animals. Iron oxide nanoclusters show clinical potential as next-generation MRI contrast agents.Uniform iron oxide nanoparticle clusters are highly biocompatible and can be used as contrast agents for high-resolution magnetic resonance angiography of large animals.

A pH-Responsive Yolk-Like Nanoplatform for Tumor Targeted Dual-Mode Magnetic Resonance Imaging and Chemotherapy

Incorporation of T1 and T2 contrast material in one nanosystem performing their respective MR contrast role and simultaneously serving as an efficient drug delivery system (DDS) has a significant potential application for clinical diagnosis and chemotherapy of cancer. However, inappropriate incorporation always encountered many issues, such as low contact area of T1 contrast material with water-proton, inappropriate distance between T2 contrast material and water molecule, and undesirable disturbance of T2 contrast material for T1 imaging. Those issues seriously limited the T1 or T2 contrast effect. In this work, we developed a yolk-like Fe3O4@Gd2O3 nanoplatform functionalized by polyethylene glycol and folic acid (FA), which could efficiently exert their tumor targeted T1-T2 dual-mode MR imaging and drug delivery role. First, this nanoplatform possessed a high longitudinal relaxation rate (r1) (7.91 mM-1 s-1) and a stronger transverse relaxation rate (r2) (386.5 mM-1 s-1) than that of original Fe3O4 (268.1 mM-1 s-1). Second, cisplatin could be efficiently loaded into this nanoplatform (112 mg/g) and showed pH-responsive release behavior. Third, this nanoplatform could be effectively internalized by HeLa cells with time and dosage dependence. Fourth, the FA receptor-mediated nanoplatform displayed excellent T1-T2 dual mode MR contrast enhancement and anticancer activity both in vitro and in vivo. Fifth, no apparent toxicity for vital organs was observed with systemic delivery of the nanoplatform in vivo. Thus, this nanoplatform could be a potential nanotheranostic for tumor targeted T1-T2 dual-mode MR imaging and chemotherapy.

Human Stem Cells Overexpressing miR-21 Promote Angiogenesis in Critical Limb Ischemia by Targeting CHIP to Enhance HIF-1α Activity

Critical limb ischemia (CLI) is a severe blockage in the arteries of the lower extremities. However, the effective and optimal treatment for CLI remains to be elucidated. Previous therapeutic research is mainly focused on proangiogenic growth factors administrations. Recently, miR‐21 has been revealed to play a crucial role in angiogenesis. Thus, we hypothesize that miR‐21 over‐expression in human umbilical cord blood‐derived mesenchymal stem cells (UCBMSCs) can effectively treat CLI. Herein, UCBMSCs were transduced with lentivirus‐miR‐21‐Luciferase (Lenti‐miR‐21) or lentivirus‐ LacZ‐Luciferase (Lenti‐LacZ). The results indicated that miR‐21 induced UCBMSCs proliferation, migration, and angiogenesis in vitro. Subsequently, general observation and laser Doppler perfusion imaging were introduced to detect perfusion in muscles of CLI‐nude mice on 1, 4, 7, 14, and 28 day postoperation. There was a significant improvement in blood vessels of the ischemic limb in Lenti‐miR‐21 group at 7 day compared with the saline or Lenti‐LacZ groups. At 28 day, histological analysis confirmed that UCBMSCs over‐expressing miR‐21 increased neovascularization in CLI. Furthermore, carboxyl terminus of Hsc70‐interacting protein (CHIP) was found to be the target gene for miR‐21‐mediated activation of hypoxia‐inducible factor 1α (HIF‐1α) in UCBMSCs. In summary, our study demonstrated that over‐expressing miR‐21 in UCBMSCs could improve neovascularization in CLI through enhancing HIF‐1α activity by targeting CHIP, which may hold great therapeutic promise in treating CLI. Stem Cells 2016;34:924–934

Superhero Rictor promotes cellular differentiation of mouse embryonic stem cells

Embryonic stem cells (ESCs) hold great promise for regenerative medicine. To harness the full therapeutic potential of ESCs, better understanding of the molecular mechanisms underlying the maintenance and differentiation of ESCs is required. Mammalian target of rapamycin (mTOR) is a serine/threonine protein kinase that integrates growth factor receptor signaling with cellular growth and proliferation. Dysregulation of mTOR signaling has been linked to various human diseases including cancer and metabolic syndromes. However, little is known regarding the function of mTOR signaling in the regulation of ES cell differentiation. Here we report that Rictor, a key component of mTORC2, functions as a novel ES cell differentiation promoting factor. Mechanistically, Rictor is able to interact with Prkch and facilitate Prkch phosphorylation at Ser-642. Upon phosphorylation, Prkch promotes Klf4 phosphorylation and inhibits Klf4-dependent E-cadherin expression, thereafter leading to the ES cell differentiation. These findings reveal a novel Rictor–Prkch–Klf4 pathway that plays an important role in the regulation of ES cell differentiation.

Functional reconstruction of critical-sized load-bearing bone defects using a Sclerostin-targeting miR-210-3p-based construct to enhance osteogenic activity

A considerable amount of research has focused on improving regenerative therapy strategies for repairing defects in load-bearing bones. The enhancement of tissue regeneration with microRNAs (miRNAs) is being developed because miRNAs can simultaneously regulate multiple signaling pathways in an endogenous manner. In this study, we developed a miR-210-based bone repair strategy. We identified a miRNA (miR-210-3p) that can simultaneously up-regulate the expression of multiple key osteogenic genes in vitro. This process resulted in enhanced bone formation in a subcutaneous mouse model with a miR-210-3p/poly-l-lactic acid (PLLA)/bone marrow-derived stem cell (BMSC) construct. Furthermore, we constructed a model of critical-sized load-bearing bone defects and implanted a miR-210-3p/β-tricalcium phosphate (β-TCP)/bone mesenchymal stem cell (BMSC) construct into the defect. We found that the load-bearing defect was almost fully repaired using the miR-210-3p construct. We also identified a new mechanism by which miR-210-3p regulates Sclerostin protein levels. This miRNA-based strategy may yield novel therapeutic methods for the treatment of regenerative defects in vital load-bearing bones by utilizing miRNA therapy for tissue engineering. STATEMENT OF SIGNIFICANCE The destroyed maxillofacial bone reconstruction is still a real challenge for maxillofacial surgeon, due to that functional bone reconstruction involved load-bearing. Base on the above problem, this paper developed a novel miR-210-3p/β-tricalcium phosphate (TCP)/bone marrow-derived stem cell (BMSC) construct (miR-210-3p/β-TCP/BMSCs), which lead to functional reconstruction of critical-size mandible bone defect. We found that the load-bearing defect was almost fully repaired using the miR-210-3p construct. In addition, we also found the mechanism of how the delivered microRNA activated the signaling pathways of endogenous stem cells, leading to the defect regeneration. This miRNA-based strategy can be used to regenerate defects in vital load-bearing bones, thus addressing a critical challenge in regenerative medicine by utilizing miRNA therapy for tissue engineering.