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Dynamic culture of a thermosensitive collagen hydrogel as an extracellular matrix improves the construction of tissue-engineered peripheral nerve

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Neural Regeneration Research 2014年第14期9卷 1371-1378页

作者： Lanfeng Huang Rui Li Wanguo Liu Jin Dai Zhenwu Du Xiaonan Wang Jianchao Ma Jinsong Zhao Department of Joint Surgery Orthopedics Hospital of the Second Hospital of Jilin University Centre of Hand & Foot Surgery and Reparative & Reconstructive Surgery Orthopedics Hospital of the Second Hospital of Jilin University Department of Orthopedics Surgery the Third Hospital of Jilin University Institute of Orthopedics the Second Hospital of Jilin University Department of Ophthalmology the Second Hospital of Jilin University

Tissue engineering technologies offer new treatment strategies for the repair of peripheral nerve injury, hut cell loss between seeding and adhesion to the scaffold remains inevitable. A thermosensitive collagen hydrogel was used as an extracellular matrix in this study and combined with bone marrow mesenchymal stem cells to construct tissue-engineered peripheral nerve composites in vitro. Dynamic culture was performed at an oscillating frequency of 0.5 Hz and 35° swing angle above and below the horizontal plane. The results demonstrated that bone marrow mesenchymal stem cells formed membrane-like structures around the poly-L-lactic acid scaffolds and exhibited regular alignment on the composite surface. Collagen was used to fill in the pores, and seeded cells adhered onto the poly-L-lactic acid fibers. The DNA content of the bone marrow mesenchymal stem cells was higher in the composites constructed with a thermosensitive collagen hydrogel compared with that in collagen I scaffold controls. The cellular DNA content was also higher in the thermosensitive collagen hydrogel composites constructed with the thermosensitive collagen hydrogel in dynamic culture than that in static culture. These results indicate that tissue-engineered composites formed with thermosensitive collagen hydrogel in dynamic culture can maintain larger numbers of seeded cells by avoiding cell loss during the initial adhe-sion stage. Moreover, seeded cells were distributed throughout the material.

关键词： nerve regeneration peripheral nerve biomaterials extracellular matrix tissueengineering nerve scaffold bone marrow mesenchymal stem cells thermosensitive collagen hydrogel poly-L-lactic acid dynamic culture NSFC grant neural regeneration

Transplantation of Nogo-66 receptor gene-silenced cells in a poly(D,L-lactic-co-glycolic acid) scaffold for the treatment of spinal cord injury

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Neural Regeneration Research 2013年第8期8卷 677-685页

作者： Dong Wang Yuhong Fan Jianjun Zhang Department of Neurosurgery Tianjin Fourth Central Hospital Department of Stomatology First Clinical CollegeHebei North University

Inhibition of neurite growth, which is in large part mediated by the Nogo-66 receptor, affects neural regeneration following bone marrow mesenchymal stem cell transplantation. The tissue engineering scaffold poly（D,L-lactide-co-glycolic acid） has good histocompatibility and can promote the growth of regenerating nerve fibers. The present study used small interfering RNA to silence Nogo-66 receptor gene expression in bone marrow mesenchymal stem cells and Schwann cells, which were subsequently transplanted with poly（D,L-lactide-co-glycolic acid） into the spinal cord lesion regions in rats. Simultaneously, rats treated with scaffold only were taken as the control group. Hematoxylin-eosin staining and immunohistochemistry revealed that at 4 weeks after transplantation, rats had good motor function of the hind limb after treatment with Nogo-66 receptor gene-silenced ceils prus the poly（O,L-lactide-co-glycolic acid） scaffold compared with rats treated with scaffold only, and the number of bone marrow mesenchymal stem cells and neuron-like cells was also increased. At 8 weeks after transplantation, horseradish peroxidase tracing and transmission electron microscopy showed a large number of unmyelinated and myelinated nerve fibers, as well as intact regenerating axonal myelin sheath following spinal cord hemisection injury. These experimental findings indicate that transplantation of Nogo-66 receptor gene-silenced bone marrow mesenchymal stem cells and Schwann cells plus a poly（D,L-lactide-co-glycolic acid） scaffold can significantly enhance axonal regeneration of spinal cord neurons and improve motor function of the extremities in rats following spinal cord injury.

关键词： neural regeneration spinal cord injury bone marrow mesenchymal stem cells Schwann cells poly（D,L-lactide-co-glycolic acid） Nogo-66 receptor gene rats gene silencing grants-supportedpaper photographs-containing paper neuroregeneration

In vivo tracking of neuronal-like cells by magnetic resonance in rabbit models of spinal cord injury

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Neural Regeneration Research 2013年第36期8卷 3373-3381页

作者： Ruiping Zhang Kun Zhang Jianding Li Qiang Liu Jun Xie Department of Radiology First Hospital Shanxi Medical University Department of Radiology Medical Imaging Centre Navy General Hospital Department of Orthopedics Shanxi Dayi Hospital Department of Molecular Biology Shanxi Medical University

In vitro experiments have demonstrated that neuronal-like cells derived from bone marrow mesen- chymal stem cells can survive, migrate, integrate and help to restore the function and behaviors of spinal cord injury models, and that they may serve as a suitable approach to treating spinal cord injury. However, it is very difficult to track transplanted cells in vivo. In this study, we injected su- perparamagnetic iron oxide-labeled neuronal-like cells into the subarachnoid space in a rabbit model of spinal cord injury. At 7 days after cell transplantation, a small number of dot-shaped low signal intensity shadows were observed in the spinal cord injury region, and at 14 days, the number of these shadows increased on T2-weighted imaging. Perl＇s Prussian blue staining detected dot-shaped low signal intensity shadows in the spinal cord injury region, indicative of superpara- magnetic iron oxide nanoparticle-labeled cells. These findings suggest that transplanted neu- ronal-like cells derived from bone marrow mesenchymal stem cells can migrate to the spinal cord injury region and can be tracked by magnetic resonance in vivo. Magnetic resonance imaging represents an efficient noninvasive technique for visually tracking transplanted cells in vivo.

关键词： neural regeneration neuronal-like cells bone marrow mesenchymal stem cells stern cells bone marrow in vivo tracking magnetic resonance transplantation grants-supported paper neuroregeneration

Human adipose-derived mesenchymal stem cells： a better cell source for nervous system regeneration

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Chinese Medical Journal 2014年第2期127卷 329-337页

作者： Han Chao Zhang Liang Song Lin Liu Yang Zou Wei Piao Hua Liu Jing Regenerative Medicine Center the First Affiliated Hospital of Dalian Medical University Dalian Liaoning 116011 China Department of Orthopedics Beijing Friendship Hospital Capital Medical University Beijing 100050 China School of Life Science and Biotechnology Dalian University of Technology Dalian Liaoning 116024 China College of Life Science Liaoning Normal University Dalian Liaoning 116011 China College of Basic Medical Science Dalian Medical University Dalian Liaoning 116027 China Institute of Integrative Medicine Dalian Medical University Dalian Liaoning 116027 China

Background In order to suggest an ideal source of adult stem cells for the treatment of nervous system diseases,MSCs from human adipose tissue and bone marrow were isolated and studied to explore the differences with regard to cell morphology,surface markers,neuronal differentiation capacity,especially the synapse structure formation and the secretion of neurotrophic factors.Methods The neuronal differentiation capacity of human mesenchymal stem cells from adipose tissue （hADSCs） and bone marrow （hBMSCs） was determined based on nissl body and synapse structure formation,and neural factor secretion function.hADSCs and hBMSCs were isolated and differentiated into neuron-like cells with rat brain-conditioned medium,a potentially rich source of neuronal differentiation promoting signals.Specific neuronal proteins and neural factors were detected by immunohistochemistry and enzyme-linked immunosorbent assay analysis,respectively.Results Flow cytometric analysis showed that both cell types had similar phenotypes.Cell growth curves showed that hADSCs proliferated more quickly than hBMSCs.Both kinds of cells were capable of osteogenic and adipogenic differentiation.The morphology of hADSCs and hBMSCs changed during neuronal differentiation and displayed neuronlike cell appearance after 14 days＇ differentiation.Both hADSCs and hBMSCs were able to differentiate into neuron-like cells based on their production of neuron specific proteins including β-tubulin-Ⅲ,neuron-specific enolase （NSE）,nissl bodies,and their ability to secrete brain derived neurotrophic factor （BDNF） and nerve growth factor （NGF）.Assessment of synaptop hysin and growth-associated protein-43 （GAP-43） suggested synapse structure formation in differentiated hADSCs and hBMSCs.Conclusions Our results demonstrate that hADSCs have neuronal differentiation potential similar to hBMSC,but with a higher proliferation capacity than hBMSC.Adipose tissue is abundant,easily available and would be a potential ideal source of adult stem cells for neural-related clinical

关键词： adipose derived stem cells bone marrow mesenchymal stem cells neuronal differentiation rat brain-conditioned medium

Transfection of the glial cell line-derived neurotrophic factor gene promotes neuronal differentiation

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Neural Regeneration Research 2014年第1期9卷 33-40页

作者： Jie Du Xiaoqing Gao Li Deng Nengbin Chang Huailin Xiong Yu Zheng Department of Physiology West China School of Preclinical and Forensic MedicineSichuan University Department of Anatomy Luzhou Medical College Research Center for Preclinical Medicine Luzhou Medical CollegeLuzhou Department of Anatomy Luzhou Medical CollegeLuzhou

Glial cell line-derived neurotrophic factor recombinant adenovirus vector-transfected bone marrow mesenchymal stem cells were induced to differentiate into neuron-like cells using inductive medium containing retinoic acid and epidermal growth factor. Cell viability, micro- tubule-associated protein 2-positive cell ratio, and the expression levels of glial cell line-derived neurotrophic factor, nerve growth factor and growth-associated protein-43 protein in the su- pernatant were significantly higher in glial cell line-derived neurotrophic factor/bone marrow mesenchymal stem cells compared with empty virus plasmid-transfected bone marrow mes- enchymal stem cells. Furthermore, microtubule-associated protein 2, glial cell line-derived neurotrophic factor, nerve growth factor and growth-associated protein743 mRNA levels in cell pellets were statistically higher in glial cell line-derived neurotrophic factor/bone marrow mesen- chymal stem cells compared with empty virus plasmid-transfected bone marrow mesenchymal stem cells. These results suggest that glial cell line-derived neurotrophic factor/bone marrow mesenchymal stem cells have a higher rate of induction into neuron-like cells, and this enhanced differentiation into neuron-like cells may be associated with up-regulated expression of glial cell line-derived neurotrophic factor, nerve growth factor and growth-associated protein-43.

关键词： nerve regeneration bone marrow mesenchymal stem cells cell differentiation neu-ron-like cells glial cell line-derived neurotrophic factor recombinant adenovirus vector transfection retinoic acid epidermal growth factor nerve growth factor growth-associated protein-43 neuralregeneration

Mechanisms of action of neuropeptide Y on stem cells and its potential applications in orthopaedic disorders

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World Journal of stem cells 2020年第9期12卷 986-1000页

作者： Jian-Qun Wu Nan Jiang Bin Yu Department of Orthopedics and Traumatology Huadu District People’s HospitalGuangzhou 510800Guangdong ProvinceChina Division of Orthopaedics and Traumatology Department of OrthopedicsNanfang HospitalSouthern Medical UniversityGuangzhou 510515Guangdong ProvinceChina Guangdong Provincial Key Laboratory of bone and Cartilage Regenerative Medicine Nanfang HospitalSouthern Medical UniversityGuangzhou 510515Guangdong ProvinceChina

Musculoskeletal disorders are the leading causes of disability and result in reduced quality of life.The neuro-osteogenic network is one of the most promising fields in orthopaedic research.Neuropeptide Y(NPY)system has been reported to be involved in the regulations of bone metabolism and homeostasis,which also provide feedback to the central NPY system via NPY receptors.Currently,potential roles of peripheral NPY in bone metabolism remain unclear.Growing evidence suggests that NPY can regulate biological actions of bone marrow mesenchymal stem cells,hematopoietic stem cells,endothelial cells,and chondrocytes via a local autocrine or paracrine manner by different NPY receptors.The regulative activities of NPY may be achieved through the plasticity of NPY receptors,and interactions among the targeted cells as well.In general,NPY can influence proliferation,apoptosis,differentiation,migration,mobilization,and cytokine secretion of different types of cells,and play crucial roles in the development of bone delayed/non-union,osteoporosis,and osteoarthritis.Further basic research should clarify detailed mechanisms of action of NPY on stem cells,and clinical investigations are also necessary to comprehensively evaluate potential applications of NPY and its receptor-targeted drugs in management of musculoskeletal disorders.

关键词： Neuropeptide Y bone marrow mesenchymal stem cells Hematopoietic stem cells Fracture Osteoporosis Osteoarthritis

Differentiation of marrow-derived islet-like cells and their effects on diabetic rats

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Chinese Medical Journal 2010年第22期123卷 3347-3350页

作者： LIU Ge-ling LU Yi-fang LI Wei-juan XIAO Hong-zhen SUN Guo-gui YU Fang XIANG Xiu-xiu ZHANG Hui-qin LIU Xiu-ling SHI Yan-ping LI Sha Department of Endocrinology Tangshan Worker's Hospital Tangshan Hebei 063000 China Department of Radiation Oncology Fourth Hospital of Hebei Medical University Shijiazhuang Hebei 050017 China

In recent years, islet transplantation for diabetes has shown signs of the treatment efficacy, but its application is limited due to lack of donor organizations, sources and immune rejection. bone marrow mesenchymal stem cells （BMSCs） have become a new resource of islet cell substitutes. One focus of the current research is the application of a specific inducing agent or a culture system to get directed differentiation of BMSCs, which may have part characteristics of islet cells and then be used in autologous transplantation for the treatment of diabetes.

关键词： rats bone marrow mesenchymal stem cells induction in vitro

Dynamic mechanical loading facilitated chondrogenic differentiation of rabbit BMSCs in collagen scaffolds

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Regenerative Biomaterials 2019年第2期6卷 99-106页

作者： Wanxu Cao Weimin Lin Hanxu Cai Yafang Chen Yi Man Jie Liang Qiguang Wang Yong Sun Yujiang Fan Xingdong Zhang National Engineering Research Center for Biomaterials Sichuan University29 Wangjiang RoadChengdu 610064China State Key Laboratory of Oral Diseases Department of Oral ImplantologyWest China Hospital of StomatologySichuan UniversityChengdu 610041China Sichuan Testing Center for Biomaterials and Medical Devices Sichuan University29 Wangjiang RoadChengdu 610064China

Mechanical signals have been played close attention to regulate chondrogenic differentiation of bone marrow mesenchymal stem cells(BMSCs).In this study,dynamic mechanical loading simulation with natural frequencies and intensities were applied to the 3D cultured BMSCs–collagen scaffold constructs.We investigated the effects of dynamic mechanical loading on cell adhesion,uniform distribution,proliferation,secretion of extracellular matrix(ECM)and chondrogenic differentiation of BMSCs–collagen scaffold constructs.The results indicated that dynamic mechanical loading facilitated the BMSCs adhesion,uniform distribution,proliferation and secretion of ECM with a slight contraction,which significantly improved the mechanical strength of the BMSCs–collagen scaffold constructs for better mimicking the structure and function of a native cartilage.Gene expression results indicated that dynamic mechanical loading contributed to the chondrogenic differentiation of BMSCs with higher levels of AGG,COL2A1 and SOX9 genes,and prevented of hypertrophic process with lower levels of COL10A1,and reduced the possibility of fibrocartilage formation due to down-regulated COL1A2.In conclusion,this study emphasized the important role of dynamic mechanical loading on promoting BMSCs chondrogenic differentiation and maintaining the cartilage phenotype for in vitro reconstruction of tissue-engineered cartilage,which provided an attractive prospect and a feasibility strategy for cartilage repair.

关键词： bone marrow mesenchymal stem cells collagen I dynamic mechanical loading cartilage tissue engineering

High-precision,gelatin-based,hybrid,bilayer scaffolds using melt electro-writing to repair cartilage injury

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Bioactive Materials 2021年第7期6卷 2173-2186页

作者： Yu Han Bo Jia Meifei Lian Binbin Sun Qiang Wu Benlin Sun Zhiguang Qiao Kerong Dai Department of Orthopedic Surgery Shanghai Key Laboratory of Orthopedic ImplantsShanghai Ninth People’s HospitalShanghai Jiao Tong University School of MedicineShanghai200011China Clinical and Translational Research Center for 3D Printing Technology Shanghai Ninth People’s HospitalShanghai Jiao Tong University School of MedicineShanghai200011China Department of Orthopedics Shanghai General HospitalShanghai Jiao Tong University School of MedicineShanghai200080China Department of Prosthodontics Shanghai Ninth People’s HospitalCollege of StomatologyShanghai Jiao Tong University School of MedicineNational Clinical Research Center for Oral DiseasesShanghai Key Laboratory of Stomatology&Shanghai Research Institute of StomatologyShanghai200011China Department of Orthopedic Surgery Renji HospitalSouth CampusShanghai Jiao Tong University School of MedicineShanghai201112China

Articular cartilage injury is a common disease in the field of orthopedics.Because cartilage has poor self-repairing ability,medical intervention is needed.Using melt electro-writing(MEW)technology,tissue engineering scaffolds with high porosity and high precision can be prepared.However,ordinary materials,especially natural polymer materials,are difficult to print.In this study,gelatin was mixed with poly(lactic-co-glycolic acid)to prepare high-concentration and high-viscosity printer ink,which had good printability and formability.A composite scaffold with full-layer TGF-β1 loading mixed with hydroxyapatite was prepared,and the scaffold was implanted at the cartilage injury site;microfracture surgery was conducted to induce the mesenchyme in the bone marrow.Quality stem cells thereby promoted the repair of damaged cartilage.In summary,this study developed a novel printing method,explored the molding conditions based on MEW printing ink,and constructed a bioactive cartilage repair scaffold.The scaffold can use autologous bone marrow mesenchymal stem cells and induce their differentiation to promote cartilage repair.

关键词： Gelatin-based hybrid bilayer scaffolds Melt electro-writing Cartilage injury bone marrow mesenchymal stem cells