Peripheral nerve injuries（PNI） are caused by a range of etiologies and result in a broad spectrum of disability. While nerve autografts are the current gold standard for the reconstruction of extensive nerve damage,...Peripheral nerve injuries（PNI） are caused by a range of etiologies and result in a broad spectrum of disability. While nerve autografts are the current gold standard for the reconstruction of extensive nerve damage, the limited supply of autologous nerve and complications associated with harvesting nerve from a second surgical site has driven groups from multiple disciplines, including biomedical engineering, neurosurgery, plastic surgery, and orthopedic surgery, to develop a suitable or superior alternative to autografting. Over the last couple of decades, various types of scaffolds, such as acellular nerve grafts（ANGs）, nerve guidance conduits, and non-nervous tissues, have been filled with Schwann cells, stem cells, and/or neurotrophic factors to develop tissue engineered nerve grafts（TENGs）. Although these have shown promising effects on peripheral nerve regeneration in experimental models, the autograft has remained the gold standard for large nerve gaps. This review provides a discussion of recent advances in the development of TENGs and their efficacy in experimental models. Specifically, TENGs have been enhanced via incorporation of genetically engineered cells, methods to improve stem cell survival and differentiation, optimized delivery of neurotrophic factors via drug delivery systems（DDS）, co-administration of platelet-rich plasma（PRP）, and pretreatment with chondroitinase ABC（Ch-ABC）. Other notable advancements include conduits that have been bioengineered to mimic native nerve structure via cell-derived extracellular matrix（ECM） deposition, and the development of transplantable living nervous tissue constructs from rat and human dorsal root ganglia（DRG） neurons. Grafts composed of non-nervous tissues, such as vein, artery, and muscle, will be briefly discussed.展开更多
The purpose of this study was to assess fetal bovine acellular dermal matrix as a scaffold for supporting the differentiation of bone marrow mesenchymal stem cells into neural cells fol-lowing induction with neural di...The purpose of this study was to assess fetal bovine acellular dermal matrix as a scaffold for supporting the differentiation of bone marrow mesenchymal stem cells into neural cells fol-lowing induction with neural differentiation medium. We performed long-term, continuous observation of cell morphology, growth, differentiation, and neuronal development using several microscopy techniques in conjunction with immunohistochemistry. We examined speciifc neu-ronal proteins and Nissl bodies involved in the differentiation process in order to determine the neuronal differentiation of bone marrow mesenchymal stem cells. The results show that bone marrow mesenchymal stem cells that differentiate on fetal bovine acellular dermal matrix display neuronal morphology with unipolar and bi/multipolar neurite elongations that express neuro-nal-speciifc proteins, includingβIII tubulin. The bone marrow mesenchymal stem cells grown on fetal bovine acellular dermal matrix and induced for long periods of time with neural differen-tiation medium differentiated into a multilayered neural network-like structure with long nerve ifbers that was composed of several parallel microifbers and neuronal cells, forming a complete neural circuit with dendrite-dendrite to axon-dendrite to dendrite-axon synapses. In addition, growth cones with filopodia were observed using scanning electron microscopy. Paraffin sec-tioning showed differentiated bone marrow mesenchymal stem cells with the typical features of neuronal phenotype, such as a large, round nucleus and a cytoplasm full of Nissl bodies. The data suggest that the biological scaffold fetal bovine acellular dermal matrix is capable of supporting human bone marrow mesenchymal stem cell differentiation into functional neurons and the subsequent formation of tissue engineered nerve.展开更多
The aim of this study was to fabricate biomatrix/polymer hybrid scaffolds using an electrospinning technique. Then tissue engineered heart valves were engineered by seeding mesenchymal stromal cells （MSCs） onto the ...The aim of this study was to fabricate biomatrix/polymer hybrid scaffolds using an electrospinning technique. Then tissue engineered heart valves were engineered by seeding mesenchymal stromal cells （MSCs） onto the scaffolds. The effects of the hybrid scaffolds on the proliferation of seed cells, formation of extracellular matrix and mechanical properties of tissue engineered heart valves were investigated. MSCs were obtained from rats. Porcine aortic heart valves were decellularized, coated with poly（3-hydroxybutyrate-co-4-hydroxybutyrate） using an electrospinning technique, and reseeded and cultured over a time period of 14 days. In control group, the decellularized valve scaffolds were reseeded and cultured over an equivalent time period. Specimens of each group were examined histologically （hematoxylin-eosin [HE] staining, immunohistostaining, and scanning electron microscopy）, biochemically （DNA and 4-hydroxyproline） and mechanically. The results showed that recellularization was comparable to the specimens of hybrid scaffolds and controls. The specimens of hybrid scaffolds and controls revealed comparable amounts of cell mass and 4-hydroxyproline （P〉0.05）. However, the specimens of hybrid scaffolds showed a significant increase in mechanical strength, compared to the controls （P〈0.05）. This study demonstrated the superiority of the hybrid scaffolds to increase the mechanical strength of tissue engineered heart valves. And compared to the decellularized valve scaffolds, the hybrid scaffolds showed similar effects on the proliferation of MSCs and formation of extracellular matrix. It was believed that the hybrid scaffolds could be used for the construction of tissue engineered heart valves.展开更多
Objectives To investigate the effects of epoxy chloropropan on the expression of matrix metalloproteinases-9 （MMP-9）in creating tissue engineered heart valves（TEHV）,on the tissue structures of TEHV,and to study th...Objectives To investigate the effects of epoxy chloropropan on the expression of matrix metalloproteinases-9 （MMP-9）in creating tissue engineered heart valves（TEHV）,on the tissue structures of TEHV,and to study the effects of epoxy chloropropan on the calcification of TEHV.Methods The porcine aortic valve leaflets were digested and decellularized by using detergent and trypsin.Those treated with 0.3% glutaraldehyde for 48 hours were the control group;those treated with 3% epoxy choloropropan for 24 hours were the experimental group.The cultured human bone marrow mesenchymal stem cells（hBMSCs）were seeded onto the decellularized scaffolds of TEHV.The histological studies were done with pathological sections and scanning electron microscopy and reverse transcriptase-polymerase chain reaction（RT-PCR）were used to detect the expression of MMP-9.Results In the experimental group.the histology showed that the BMSCs grew well into the pores and formed a confluent layer in decellularized scaffolds;RT-PCR indicated significantly attenuated expressions of MMP-9,compared with the control（P〈0.05）.Conclusion The decellularized porcine aortic valves treated with 3% epoxy chloropropan may inhibit the expression of MMP-9;therefore epoxy chloropropan may prevent the calcification of tissue engineered heart valves.展开更多
In this study, we constructed tissue-engineered nerves with acellular nerve allografts in Sprague-Dawley rats, which were prepared using chemical detergents-enzymatic digestion and mechanical methods, in combination w...In this study, we constructed tissue-engineered nerves with acellular nerve allografts in Sprague-Dawley rats, which were prepared using chemical detergents-enzymatic digestion and mechanical methods, in combination with bone marrow mesenchymal stem cells of Wistar rats cultured in vitro, to repair 15 mm sciatic bone defects in Wistar rats. At postoperative 12 weeks, electrophysiological detection results showed that the conduction velocity of regenerated nerve after repair with tissue-engineered nerves was similar to that after autologous nerve grafting, and was higher than that after repair with acellular nerve allografts. Immunohistochemical staining revealed that motor endplates with acetylcholinesterase-positive nerve fibers were orderly arranged in the middle and superior parts of the gastrocnemius muscle; regenerated nerve tracts and sprouted branches were connected with motor endplates, as shown by acetylcholinesterase histochemistry combined with silver staining. The wet weight ratio of the tibialis anterior muscle at the affected contralateral hind limb was similar to the sciatic nerve after repair with autologous nerve grafts, and higher than that after repair with acellular nerve allografts. The hind limb motor function at the affected side was significantly improved, indicating that acellular nerve allografts combined with bone marrow mesenchymal stem cell bridging could promote functional recovery of rats with sciatic nerve defects.展开更多
Neural tissue engineering is premised on the integration of engineered living tissue with the host nervous system to directly restore lost function or to augment regenerative capacity following ner- vous system injury...Neural tissue engineering is premised on the integration of engineered living tissue with the host nervous system to directly restore lost function or to augment regenerative capacity following ner- vous system injury or neurodegenerative disease. Disconnection of axon pathways - the long-distance fibers connecting specialized regions of the central nervous system or relaying peripheral signals - is a common feature of many neurological disorders and injury. However, functional axonal regenera- tion rarely occurs due to extreme distances to targets, absence of directed guidance, and the presence of inhibitory factors in the central nervous system, resulting in devastating effects on cognitive and sensorimotor function. To address this need, we are pursuing multiple strategies using tissue engi- neered ＂living scaffolds＂, which are preformed three-dimensional constructs consisting of living neural cells in a defined, often anisotropic architecture. Living scaffolds are designed to restore function by serving as a living labeled pathway for targeted axonal regeneration - mimicking key developmental mechanisms- or by restoring lost neural circuitry via direct replacement of neurons and axonal tracts. We are currently utilizing preformed living scaffolds consisting of neuronal dusters spanned by long axonal tracts as regenerative bridges to facilitate long-distance axonal regeneration and for targeted neurosurgical reconstruction of local circuits in the brain. Although there are formidable challenges in predinical and clinical advancement, these living tissue engineered constructs represent a promising strategy to facilitate nervous system repair and functional recovery.展开更多
Objective:Urethral stricture is a highly prevalent disease and has a continued ris-ing incidence.The global burden of disease keeps rising as there are significant rates of recur-rence with the existing management opt...Objective:Urethral stricture is a highly prevalent disease and has a continued ris-ing incidence.The global burden of disease keeps rising as there are significant rates of recur-rence with the existing management options with the need for additional repeat procedures.Moreover,the existing treatment options are associated with significant morbidity in the pa-tient.Long segment urethral strictures are most commonly managed by augmentation urethro-plasty.We explored the potential for the application of an acellular tissue engineered bovine pericardial patch in augmentation urethroplasty in a series of our patients suffering from ure-thral stricture disease.The decreased morbidity due to the avoidance of harvest of buccal mu-cosa,decreased operative time and satisfactory postoperative results make it a promising option for augmentation urethroplasty.Methods:Nine patients with long segment anterior urethral strictures(involving penile and/or bulbar urethra and stricture length>4 cm)were included in the study after proper informed consent was obtained.Acellular tissue engineered indigenous bovine pericardial patch was used for urethroplasty using dorsal onlay technique.Results:A total of nine patients underwent tissue engineered indigenous pericardial patch ur-ethroplasty for long segment urethral strictures,mostly catheter injury induced or associated with balanitis xerotica obliterans.Median follow-up was 8 months(range:2-12 months).Out of nine patients,eight(88.9%)were classifed as success and one(11.1%)was classified as fail-ure.Conclusion:Our study brings a product of tissue engineering,already being used in the cardio-vascular surgery domain,into the urological surgery operating room with satisfactory results achieved using standard operating techniques of one stage urethroplasty.展开更多
To study the osteogenic ability of tissue-engineered bone constructed by compounding zinc-sintered bovine cancellous bone with rabbit marrow stromal cells (MSCs) in vivo,the zinc-sintered bovine cancellous bone of bet...To study the osteogenic ability of tissue-engineered bone constructed by compounding zinc-sintered bovine cancellous bone with rabbit marrow stromal cells (MSCs) in vivo,the zinc-sintered bovine cancellous bone of beta-tricalcium phosphate (TCP) type was prepared by sintering the fresh calf cancellous bone twice and then loading it with zinc-ion.The rabbit MSCs were cultured,induced and seeded onto the zinc-sintered bovine cancellous bones.The tissue-engineered bones were then implanted into the rabbits' back muscles.The newly formed bone tissues were observed by histological methods and the areas of new osseous tissues were measured at the end of the 4th and 8th week.The zinc-sintered bovine cancellous bones alone were implanted on the other side as control.The osteogenic activity of MSCs was identified by alkaline phosphatase (ALP) staining and calcification nod chinalizarin staining.At the end of 4th week,a small amount of new bone tissues was observed.At the end of 8th week,there were many newly formed bone mature tissues.Moreover,the area of the latter was significantly larger than that of the former(P<0.01),while in the control group there was no new bone formation.The tissue-engineered bone,which was constructed by combining zinc-sintered bovine cancellous bone with MSCs,has satisfactory osteogenic capabilities in vivo.展开更多
To enhance the adhesion of seeding-cells to the biomaterial scaffolds, the PEG-hydrogels were modified. Porcine aortic valves were decellularized with Triton X-100 and trypsin. The cells were encapsulated into the PEG...To enhance the adhesion of seeding-cells to the biomaterial scaffolds, the PEG-hydrogels were modified. Porcine aortic valves were decellularized with Triton X-100 and trypsin. The cells were encapsulated into the PEG-hydrogels to complete the process of the cells attaching to the acellular porcine aortic valves. Herein, the autologous mesenchymal stem cells （MSCs） of goats were selected as the seeding-cells and the tendency of MSCs toward differentiation was observed when the single semilunar TEHV had been implanted into their abdominal aortas. Furthermore, VEGF, TGF-β1, and the cell adhesive peptide motif RGD were incorporated. Light and electron microscopy observations were performed. Analysis of modified PEG-hydrogels TEHV＇s （PEG-TEHV） tensile strength, and the ratio of reendothelial and mural thrombosis revealed much better improvement than the naked acellular porcine aortic valve （NAPAV）. The data illustrated the critical importance of MSC differentiation into endothelial and myofibroblast for remodeling into native tissue. Our results indicate that it is feasible to reconstruct TEHV efficiently by combining modified PEG-hydrogels with acellular biomaterial scaffold andautologous MSCs cells.展开更多
Objective:Unlike other tissues,myocardium has not substitute whick can be used to repair damaged cardiac tissue.This paper proposes engineering 3-D myocardium-like tissue constructs in vitro with bone mesenchymal stem...Objective:Unlike other tissues,myocardium has not substitute whick can be used to repair damaged cardiac tissue.This paper proposes engineering 3-D myocardium-like tissue constructs in vitro with bone mesenchymal stem cells(BMSCs) of infant and poly-lactic-co-glycolic acid(PLGA)in vitro.Methods:Bone marrow was obtained from the sternal marrow cavum outflow of infant with congenital heart disease (CHD)undergoing cardiac operation.BMSCs were obtained by density gradient centrifugation.The cells in passages two were induced in DMED with 10 umol/L 5- Azacytidine(5-Aza)for 24 h.When the induced BMSCS were cultured nearly into filled,the cells were planted in the scaffold of PLGA in 5.5×106 cells/cm2.The cell- scaffold complex has been cultured in the shake cultivation for 1 week,then the complex has been planted in the dorse of the nude mouse.When the experiment had been finished,the histology,immunology,real time PCR and so on were done.Results: The BMSCs of infant with congenital heart disease have the properties of the stable growth and the rapid proliferation.The immunohistochemistry showed that tissue engineered myocardium constructed in vitro expressed some cardiac related proteins such asα-actin,Cx-43,Desmine,cTNI and so on.The transparent myofilaments,gap junctions and intercalated disk-like structure formation could be observed in the 3D tissue-like constructs by transmission electron microscope(TEM).The engineered myocardium-like tissue had the auto-myocardial property as assessed by real time- PCR and so on.Conclusion:The engineered myocardial tissue-like constructs could be built with infant BMSCs and PLGA in vitro.Our results may provide the first step on the long road toward engineering myocardial material for repairing the defect or augmenting the tract in CHD,such as ventricular septal defect,tetralogy of Fallot and so on.展开更多
The inherent complexities of excitable cardiac,nervous,and skeletal muscle tissues pose great challenges in constructing artificial counterparts that closely resemble their natural bioelectrical,structural,and mechani...The inherent complexities of excitable cardiac,nervous,and skeletal muscle tissues pose great challenges in constructing artificial counterparts that closely resemble their natural bioelectrical,structural,and mechanical properties.Recent advances have increasingly revealed the beneficial impact of bioelectrical microenvironments on cellular behaviors,tissue regeneration,and therapeutic efficacy for excitable tissues.This review aims to unveil the mechanisms by which electrical microenvironments enhance the regeneration and functionality of excitable cells and tissues,considering both endogenous electrical cues from electroactive biomaterials and exogenous electrical stimuli from external electronic systems.We explore the synergistic effects of these electrical microenvironments,combined with structural and mechanical guidance,on the regeneration of excitable tissues using tissue engineering scaffolds.Additionally,the emergence of micro/nanoscale bioelectronics has significantly broadened this field,facilitating intimate interactions between implantable bioelectronics and excitable tissues across cellular,tissue,and organ levels.These interactions enable precise data acquisition and localized modulation of cell and tissue functionalities through intricately designed electronic components according to physiological needs.The integration of tissue engineering and bioelectronics promises optimal outcomes,highlighting a growing trend in developing living tissue construct-bioelectronic hybrids for restoring and monitoring damaged excitable tissues.Furthermore,we envision critical challenges in engineering the next-generation hybrids,focusing on integrated fabrication strategies,the development of ionic conductive biomaterials,and their convergence with biosensors.展开更多
Craniofacial muscles are essential components of the skeletal muscular system that contribute to important physiological processes.Severe trauma can induce craniofacial volumetric muscle loss(VML),which impairs muscle...Craniofacial muscles are essential components of the skeletal muscular system that contribute to important physiological processes.Severe trauma can induce craniofacial volumetric muscle loss(VML),which impairs muscle regeneration,causes facial muscular deformities and functional disability,and leads to psychosocial consequences such as isolation and depression.Conventional therapies involving muscle flap transposition or autologous tissue grafting achieve morphological repair but are ineffective in restoring muscle function,resulting in donor site injury and sensory deficit.In this study,we successfully constructed a biomimetically engineered muscle tissue that integrates myofiber alignment,effective innervation,and blood perfusion to promote multi-tissue regeneration in the masseter area in vivo,enabling functional regeneration.Using light-controlled micropatterning technology,we constructed mature muscle fibers with oriented alignment and established a neuromuscular co-culture system for in vitro neuromuscular junction reconstruction.Furthermore,we designed and fabricated a vascular network structure to promote tissue vascularization using hydrogel as the vehicle for assembling the composite engineered tissue.Using this technology,the shape and dimension of the constructed entity can be customized to address various muscle defects,enabling individualized repair.This study offers a promising novel strategy for tissue regeneration that breaks through the current challenges in the treatment of craniofacial VML.展开更多
Objective： To observe the effect of tissue engineered nerves in repairing peripheral nerve defects （ about 1. 5 cm in length） in rats to provide data for clinical application. Methods： Glycerinated sciatic nerves...Objective： To observe the effect of tissue engineered nerves in repairing peripheral nerve defects （ about 1. 5 cm in length） in rats to provide data for clinical application. Methods： Glycerinated sciatic nerves （2 cm in length） from 10 Sprague Dawiey （SD） rats （aged 4 months） were used to prepare homologous dermal acellular matrix. Other 10 neonate SD rats （aged 5-7 days） were killed by neck dislocation. After removing the epineurium, the separated sciatic nerve tracts were cut into small pieces, then digested by 2.5 g/L trypsin and 625 U/nd collagenase and cultured in Dulbecco＇ s modified Eagle＇ s medium （DMEM） for 3 weeks. After proliferation, the Schwann cells （SCs） were identified and prepared for use. And other 40 female adult SD rats （ weighing 200 g and aged 3 months） with sciatic nerve defects of 1.5 cm in length were randomly divided into four groups： the defects of 10 rats bridged with proliferated SCs and homologous dermal acellular matrix （the tissue engineered nerve group, Group A）, 10 rats with no SCs but homologous dermal acellular matrix with internal scaffolds （Group B ）, 10 with autologous nerves （Group C ）, and the other 10 with nothing （the blank control group, Group D）. The general status of the rats was observed, the wet weight of triceps muscle of calf was monitored, and the histological observation of the regenerated nerves were made at 12 weeks after operation. Results： The wounds of all 40 rats healed after operation and no death was found. No foot ulceration was found in Groups A, B and C, but 7 rats suffered from foot ulceration in Group D. The triceps muscles of calf were depauperated in the experimental sides in all the groups compared with the uninjured sides, which was much more obvious in Group D. The wet weight of triceps muscle of calf and nerve electrophysiologic monitoring showed no statistical difference between Group A and Group C, but statistical difference was found between Groups A and B and Groups B and D. And significant statistical difference was found between Group B and Group D. Obvious compound muscle （ or motor） action potential （CMAP） could be evoked in Group A and Group C, but the evoked amplitude was very low in Group B and Group D. The axons of regenerated nerves penetrated through the whole graft in Group A and Group C, and partly penetrated through the graft in Group B, but did not penetrate in Group D. The two tips of the separated sciatic nerves of Groups A , B , and C were connected together, without formation of neuroma. But those of Group D were not connected together and neuroma formed in 6 rats. Conclusions： Tissue engineered nerves can be used for repairing long defects of the peripheral nerves of rats and ideal repairing effects can be obtained.展开更多
Extracellular vesicles from skin-derived precursor Schwann cells(SKP-SC-EVs)promote neurite outgrowth in culture and enhance peripheral nerve regeneration in rats.This study aimed at expanding the application of SKPSC...Extracellular vesicles from skin-derived precursor Schwann cells(SKP-SC-EVs)promote neurite outgrowth in culture and enhance peripheral nerve regeneration in rats.This study aimed at expanding the application of SKPSC-EVs in nerve grafting by creating a chitosan/PLGA-based,SKP-SC-EVs-containing tissue engineered nerve graft(TENG)to bridge a 40-mm long sciatic nerve defect in dogs.SKP-SC-EVs contained in TENGs significantly accelerated the recovery of hind limb motor and electrophysiological functions,supported the outgrowth and myelination of regenerated axons,and alleviated the denervation-induced atrophy of target muscles in dogs.To clarify the underlying molecular mechanism,we observed that SKP-SC-EVs were rich in a variety of miRNAs linked to the axon growth of neurons,and miR-30b-5p was the most important among others.We further noted that miR-30b-5p contained within SKP-SC-EVs exerted nerve regeneration-promoting effects by targeting the Sin3a/HDAC complex and activating the phosphorylation of ERK,STAT3 or CREB.Our findings suggested that SKP-SC-EVs-incorporating TENGs represent a novel type of bioactive material with potential application for peripheral nerve repair in the clinic.展开更多
Background We previously showed that nano-hydroxyapatite/carboxymethyl chitosan （n-Ha/CMCS） displayed excellent mechanical properties, good degradation rates and exceptional biocompatibility, with negligible toxicit...Background We previously showed that nano-hydroxyapatite/carboxymethyl chitosan （n-Ha/CMCS） displayed excellent mechanical properties, good degradation rates and exceptional biocompatibility, with negligible toxicity. The aim of this study was to determine the effect of the same composite with vascular endothelial growth factor （VEGF）transfected bone marrow stromal cells （BMSCs） in a rabbit radial defect model.展开更多
基金supported,in part,by a research grant from Baylor Scott&White Health Central Texas Foundation and NIH grant R01-NS067435(JHH)
文摘Peripheral nerve injuries（PNI） are caused by a range of etiologies and result in a broad spectrum of disability. While nerve autografts are the current gold standard for the reconstruction of extensive nerve damage, the limited supply of autologous nerve and complications associated with harvesting nerve from a second surgical site has driven groups from multiple disciplines, including biomedical engineering, neurosurgery, plastic surgery, and orthopedic surgery, to develop a suitable or superior alternative to autografting. Over the last couple of decades, various types of scaffolds, such as acellular nerve grafts（ANGs）, nerve guidance conduits, and non-nervous tissues, have been filled with Schwann cells, stem cells, and/or neurotrophic factors to develop tissue engineered nerve grafts（TENGs）. Although these have shown promising effects on peripheral nerve regeneration in experimental models, the autograft has remained the gold standard for large nerve gaps. This review provides a discussion of recent advances in the development of TENGs and their efficacy in experimental models. Specifically, TENGs have been enhanced via incorporation of genetically engineered cells, methods to improve stem cell survival and differentiation, optimized delivery of neurotrophic factors via drug delivery systems（DDS）, co-administration of platelet-rich plasma（PRP）, and pretreatment with chondroitinase ABC（Ch-ABC）. Other notable advancements include conduits that have been bioengineered to mimic native nerve structure via cell-derived extracellular matrix（ECM） deposition, and the development of transplantable living nervous tissue constructs from rat and human dorsal root ganglia（DRG） neurons. Grafts composed of non-nervous tissues, such as vein, artery, and muscle, will be briefly discussed.
基金supported by a grant from Construction Project of Gansu Provincial Animal Cell Engineering Center,No.0808NTGA013Program for Innovative Research Team in University of Ministry of Education of China,No.IRT13091
文摘The purpose of this study was to assess fetal bovine acellular dermal matrix as a scaffold for supporting the differentiation of bone marrow mesenchymal stem cells into neural cells fol-lowing induction with neural differentiation medium. We performed long-term, continuous observation of cell morphology, growth, differentiation, and neuronal development using several microscopy techniques in conjunction with immunohistochemistry. We examined speciifc neu-ronal proteins and Nissl bodies involved in the differentiation process in order to determine the neuronal differentiation of bone marrow mesenchymal stem cells. The results show that bone marrow mesenchymal stem cells that differentiate on fetal bovine acellular dermal matrix display neuronal morphology with unipolar and bi/multipolar neurite elongations that express neuro-nal-speciifc proteins, includingβIII tubulin. The bone marrow mesenchymal stem cells grown on fetal bovine acellular dermal matrix and induced for long periods of time with neural differen-tiation medium differentiated into a multilayered neural network-like structure with long nerve ifbers that was composed of several parallel microifbers and neuronal cells, forming a complete neural circuit with dendrite-dendrite to axon-dendrite to dendrite-axon synapses. In addition, growth cones with filopodia were observed using scanning electron microscopy. Paraffin sec-tioning showed differentiated bone marrow mesenchymal stem cells with the typical features of neuronal phenotype, such as a large, round nucleus and a cytoplasm full of Nissl bodies. The data suggest that the biological scaffold fetal bovine acellular dermal matrix is capable of supporting human bone marrow mesenchymal stem cell differentiation into functional neurons and the subsequent formation of tissue engineered nerve.
基金supported by grants from National Natural Sciences Foundation of China (No.30571839,30600608 and 30872540)National High Technology Research and Development Program ("863" Program) of China (No.2009AA-03Z420)
文摘The aim of this study was to fabricate biomatrix/polymer hybrid scaffolds using an electrospinning technique. Then tissue engineered heart valves were engineered by seeding mesenchymal stromal cells （MSCs） onto the scaffolds. The effects of the hybrid scaffolds on the proliferation of seed cells, formation of extracellular matrix and mechanical properties of tissue engineered heart valves were investigated. MSCs were obtained from rats. Porcine aortic heart valves were decellularized, coated with poly（3-hydroxybutyrate-co-4-hydroxybutyrate） using an electrospinning technique, and reseeded and cultured over a time period of 14 days. In control group, the decellularized valve scaffolds were reseeded and cultured over an equivalent time period. Specimens of each group were examined histologically （hematoxylin-eosin [HE] staining, immunohistostaining, and scanning electron microscopy）, biochemically （DNA and 4-hydroxyproline） and mechanically. The results showed that recellularization was comparable to the specimens of hybrid scaffolds and controls. The specimens of hybrid scaffolds and controls revealed comparable amounts of cell mass and 4-hydroxyproline （P〉0.05）. However, the specimens of hybrid scaffolds showed a significant increase in mechanical strength, compared to the controls （P〈0.05）. This study demonstrated the superiority of the hybrid scaffolds to increase the mechanical strength of tissue engineered heart valves. And compared to the decellularized valve scaffolds, the hybrid scaffolds showed similar effects on the proliferation of MSCs and formation of extracellular matrix. It was believed that the hybrid scaffolds could be used for the construction of tissue engineered heart valves.
文摘Objectives To investigate the effects of epoxy chloropropan on the expression of matrix metalloproteinases-9 （MMP-9）in creating tissue engineered heart valves（TEHV）,on the tissue structures of TEHV,and to study the effects of epoxy chloropropan on the calcification of TEHV.Methods The porcine aortic valve leaflets were digested and decellularized by using detergent and trypsin.Those treated with 0.3% glutaraldehyde for 48 hours were the control group;those treated with 3% epoxy choloropropan for 24 hours were the experimental group.The cultured human bone marrow mesenchymal stem cells（hBMSCs）were seeded onto the decellularized scaffolds of TEHV.The histological studies were done with pathological sections and scanning electron microscopy and reverse transcriptase-polymerase chain reaction（RT-PCR）were used to detect the expression of MMP-9.Results In the experimental group.the histology showed that the BMSCs grew well into the pores and formed a confluent layer in decellularized scaffolds;RT-PCR indicated significantly attenuated expressions of MMP-9,compared with the control（P〈0.05）.Conclusion The decellularized porcine aortic valves treated with 3% epoxy chloropropan may inhibit the expression of MMP-9;therefore epoxy chloropropan may prevent the calcification of tissue engineered heart valves.
基金financially sponsored by the Natural Science Foundation of Liaoning Province,No.201102135
文摘In this study, we constructed tissue-engineered nerves with acellular nerve allografts in Sprague-Dawley rats, which were prepared using chemical detergents-enzymatic digestion and mechanical methods, in combination with bone marrow mesenchymal stem cells of Wistar rats cultured in vitro, to repair 15 mm sciatic bone defects in Wistar rats. At postoperative 12 weeks, electrophysiological detection results showed that the conduction velocity of regenerated nerve after repair with tissue-engineered nerves was similar to that after autologous nerve grafting, and was higher than that after repair with acellular nerve allografts. Immunohistochemical staining revealed that motor endplates with acetylcholinesterase-positive nerve fibers were orderly arranged in the middle and superior parts of the gastrocnemius muscle; regenerated nerve tracts and sprouted branches were connected with motor endplates, as shown by acetylcholinesterase histochemistry combined with silver staining. The wet weight ratio of the tibialis anterior muscle at the affected contralateral hind limb was similar to the sciatic nerve after repair with autologous nerve grafts, and higher than that after repair with acellular nerve allografts. The hind limb motor function at the affected side was significantly improved, indicating that acellular nerve allografts combined with bone marrow mesenchymal stem cell bridging could promote functional recovery of rats with sciatic nerve defects.
基金support provided by the U.S.Army Medical Research and Materiel Command through the Joint Warfighter Medical Research Program(#W81XWH-13-13207004)Axonia Medical,Inc.+3 种基金Department of Veterans Affairs(RR&D Merit Review#B1097-I)National Institutes of Health(NINDS T32-NS043126)Penn Medicine Neuroscience Centerthe National Science Foundation(Graduate Research Fellowship DGE-1321851)
文摘Neural tissue engineering is premised on the integration of engineered living tissue with the host nervous system to directly restore lost function or to augment regenerative capacity following ner- vous system injury or neurodegenerative disease. Disconnection of axon pathways - the long-distance fibers connecting specialized regions of the central nervous system or relaying peripheral signals - is a common feature of many neurological disorders and injury. However, functional axonal regenera- tion rarely occurs due to extreme distances to targets, absence of directed guidance, and the presence of inhibitory factors in the central nervous system, resulting in devastating effects on cognitive and sensorimotor function. To address this need, we are pursuing multiple strategies using tissue engi- neered ＂living scaffolds＂, which are preformed three-dimensional constructs consisting of living neural cells in a defined, often anisotropic architecture. Living scaffolds are designed to restore function by serving as a living labeled pathway for targeted axonal regeneration - mimicking key developmental mechanisms- or by restoring lost neural circuitry via direct replacement of neurons and axonal tracts. We are currently utilizing preformed living scaffolds consisting of neuronal dusters spanned by long axonal tracts as regenerative bridges to facilitate long-distance axonal regeneration and for targeted neurosurgical reconstruction of local circuits in the brain. Although there are formidable challenges in predinical and clinical advancement, these living tissue engineered constructs represent a promising strategy to facilitate nervous system repair and functional recovery.
文摘Objective:Urethral stricture is a highly prevalent disease and has a continued ris-ing incidence.The global burden of disease keeps rising as there are significant rates of recur-rence with the existing management options with the need for additional repeat procedures.Moreover,the existing treatment options are associated with significant morbidity in the pa-tient.Long segment urethral strictures are most commonly managed by augmentation urethro-plasty.We explored the potential for the application of an acellular tissue engineered bovine pericardial patch in augmentation urethroplasty in a series of our patients suffering from ure-thral stricture disease.The decreased morbidity due to the avoidance of harvest of buccal mu-cosa,decreased operative time and satisfactory postoperative results make it a promising option for augmentation urethroplasty.Methods:Nine patients with long segment anterior urethral strictures(involving penile and/or bulbar urethra and stricture length>4 cm)were included in the study after proper informed consent was obtained.Acellular tissue engineered indigenous bovine pericardial patch was used for urethroplasty using dorsal onlay technique.Results:A total of nine patients underwent tissue engineered indigenous pericardial patch ur-ethroplasty for long segment urethral strictures,mostly catheter injury induced or associated with balanitis xerotica obliterans.Median follow-up was 8 months(range:2-12 months).Out of nine patients,eight(88.9%)were classifed as success and one(11.1%)was classified as fail-ure.Conclusion:Our study brings a product of tissue engineering,already being used in the cardio-vascular surgery domain,into the urological surgery operating room with satisfactory results achieved using standard operating techniques of one stage urethroplasty.
文摘To study the osteogenic ability of tissue-engineered bone constructed by compounding zinc-sintered bovine cancellous bone with rabbit marrow stromal cells (MSCs) in vivo,the zinc-sintered bovine cancellous bone of beta-tricalcium phosphate (TCP) type was prepared by sintering the fresh calf cancellous bone twice and then loading it with zinc-ion.The rabbit MSCs were cultured,induced and seeded onto the zinc-sintered bovine cancellous bones.The tissue-engineered bones were then implanted into the rabbits' back muscles.The newly formed bone tissues were observed by histological methods and the areas of new osseous tissues were measured at the end of the 4th and 8th week.The zinc-sintered bovine cancellous bones alone were implanted on the other side as control.The osteogenic activity of MSCs was identified by alkaline phosphatase (ALP) staining and calcification nod chinalizarin staining.At the end of 4th week,a small amount of new bone tissues was observed.At the end of 8th week,there were many newly formed bone mature tissues.Moreover,the area of the latter was significantly larger than that of the former(P<0.01),while in the control group there was no new bone formation.The tissue-engineered bone,which was constructed by combining zinc-sintered bovine cancellous bone with MSCs,has satisfactory osteogenic capabilities in vivo.
文摘To enhance the adhesion of seeding-cells to the biomaterial scaffolds, the PEG-hydrogels were modified. Porcine aortic valves were decellularized with Triton X-100 and trypsin. The cells were encapsulated into the PEG-hydrogels to complete the process of the cells attaching to the acellular porcine aortic valves. Herein, the autologous mesenchymal stem cells （MSCs） of goats were selected as the seeding-cells and the tendency of MSCs toward differentiation was observed when the single semilunar TEHV had been implanted into their abdominal aortas. Furthermore, VEGF, TGF-β1, and the cell adhesive peptide motif RGD were incorporated. Light and electron microscopy observations were performed. Analysis of modified PEG-hydrogels TEHV＇s （PEG-TEHV） tensile strength, and the ratio of reendothelial and mural thrombosis revealed much better improvement than the naked acellular porcine aortic valve （NAPAV）. The data illustrated the critical importance of MSC differentiation into endothelial and myofibroblast for remodeling into native tissue. Our results indicate that it is feasible to reconstruct TEHV efficiently by combining modified PEG-hydrogels with acellular biomaterial scaffold andautologous MSCs cells.
基金The Tackle Key Problems in Science and Technology, Shanxi Province grant number: 20080311061-2
文摘Objective:Unlike other tissues,myocardium has not substitute whick can be used to repair damaged cardiac tissue.This paper proposes engineering 3-D myocardium-like tissue constructs in vitro with bone mesenchymal stem cells(BMSCs) of infant and poly-lactic-co-glycolic acid(PLGA)in vitro.Methods:Bone marrow was obtained from the sternal marrow cavum outflow of infant with congenital heart disease (CHD)undergoing cardiac operation.BMSCs were obtained by density gradient centrifugation.The cells in passages two were induced in DMED with 10 umol/L 5- Azacytidine(5-Aza)for 24 h.When the induced BMSCS were cultured nearly into filled,the cells were planted in the scaffold of PLGA in 5.5×106 cells/cm2.The cell- scaffold complex has been cultured in the shake cultivation for 1 week,then the complex has been planted in the dorse of the nude mouse.When the experiment had been finished,the histology,immunology,real time PCR and so on were done.Results: The BMSCs of infant with congenital heart disease have the properties of the stable growth and the rapid proliferation.The immunohistochemistry showed that tissue engineered myocardium constructed in vitro expressed some cardiac related proteins such asα-actin,Cx-43,Desmine,cTNI and so on.The transparent myofilaments,gap junctions and intercalated disk-like structure formation could be observed in the 3D tissue-like constructs by transmission electron microscope(TEM).The engineered myocardium-like tissue had the auto-myocardial property as assessed by real time- PCR and so on.Conclusion:The engineered myocardial tissue-like constructs could be built with infant BMSCs and PLGA in vitro.Our results may provide the first step on the long road toward engineering myocardial material for repairing the defect or augmenting the tract in CHD,such as ventricular septal defect,tetralogy of Fallot and so on.
基金financially supported by the National Natural Science Foundation of China(Nos.52125501,52405325)the Key Research Project of Shaanxi Province(Nos.2021LLRH-08,2024SF2-GJHX-34)+5 种基金the Program for Innovation Team of Shaanxi Province(No.2023-CX-TD17)the Postdoctoral Fellowship Program of CPSF(No.GZB20230573)the Postdoctoral Project of Shaanxi Province(No.2023BSHYDZZ30)the Basic Research Program of Natural Science in Shaanxi Province(No.2021JQ-906)the China Postdoctoral Science Foundationthe Fundamental Research Funds for the Central Universities。
文摘The inherent complexities of excitable cardiac,nervous,and skeletal muscle tissues pose great challenges in constructing artificial counterparts that closely resemble their natural bioelectrical,structural,and mechanical properties.Recent advances have increasingly revealed the beneficial impact of bioelectrical microenvironments on cellular behaviors,tissue regeneration,and therapeutic efficacy for excitable tissues.This review aims to unveil the mechanisms by which electrical microenvironments enhance the regeneration and functionality of excitable cells and tissues,considering both endogenous electrical cues from electroactive biomaterials and exogenous electrical stimuli from external electronic systems.We explore the synergistic effects of these electrical microenvironments,combined with structural and mechanical guidance,on the regeneration of excitable tissues using tissue engineering scaffolds.Additionally,the emergence of micro/nanoscale bioelectronics has significantly broadened this field,facilitating intimate interactions between implantable bioelectronics and excitable tissues across cellular,tissue,and organ levels.These interactions enable precise data acquisition and localized modulation of cell and tissue functionalities through intricately designed electronic components according to physiological needs.The integration of tissue engineering and bioelectronics promises optimal outcomes,highlighting a growing trend in developing living tissue construct-bioelectronic hybrids for restoring and monitoring damaged excitable tissues.Furthermore,we envision critical challenges in engineering the next-generation hybrids,focusing on integrated fabrication strategies,the development of ionic conductive biomaterials,and their convergence with biosensors.
基金supported by the National Natural Science Foundation of China(Nos.82122014,82071085,82020108011,and 82301031)the Zhejiang Provincial Natural Science Foundation of China(No.LR21H140001)+2 种基金the National Key Research and Development Program of China(No.2018YFA0703000)the Medical Technology and Education of Zhejiang Province of China(No.2018KY501)the Fundamental Research Funds for the Central Universities(No.2022QZJH55).
文摘Craniofacial muscles are essential components of the skeletal muscular system that contribute to important physiological processes.Severe trauma can induce craniofacial volumetric muscle loss(VML),which impairs muscle regeneration,causes facial muscular deformities and functional disability,and leads to psychosocial consequences such as isolation and depression.Conventional therapies involving muscle flap transposition or autologous tissue grafting achieve morphological repair but are ineffective in restoring muscle function,resulting in donor site injury and sensory deficit.In this study,we successfully constructed a biomimetically engineered muscle tissue that integrates myofiber alignment,effective innervation,and blood perfusion to promote multi-tissue regeneration in the masseter area in vivo,enabling functional regeneration.Using light-controlled micropatterning technology,we constructed mature muscle fibers with oriented alignment and established a neuromuscular co-culture system for in vitro neuromuscular junction reconstruction.Furthermore,we designed and fabricated a vascular network structure to promote tissue vascularization using hydrogel as the vehicle for assembling the composite engineered tissue.Using this technology,the shape and dimension of the constructed entity can be customized to address various muscle defects,enabling individualized repair.This study offers a promising novel strategy for tissue regeneration that breaks through the current challenges in the treatment of craniofacial VML.
文摘Objective： To observe the effect of tissue engineered nerves in repairing peripheral nerve defects （ about 1. 5 cm in length） in rats to provide data for clinical application. Methods： Glycerinated sciatic nerves （2 cm in length） from 10 Sprague Dawiey （SD） rats （aged 4 months） were used to prepare homologous dermal acellular matrix. Other 10 neonate SD rats （aged 5-7 days） were killed by neck dislocation. After removing the epineurium, the separated sciatic nerve tracts were cut into small pieces, then digested by 2.5 g/L trypsin and 625 U/nd collagenase and cultured in Dulbecco＇ s modified Eagle＇ s medium （DMEM） for 3 weeks. After proliferation, the Schwann cells （SCs） were identified and prepared for use. And other 40 female adult SD rats （ weighing 200 g and aged 3 months） with sciatic nerve defects of 1.5 cm in length were randomly divided into four groups： the defects of 10 rats bridged with proliferated SCs and homologous dermal acellular matrix （the tissue engineered nerve group, Group A）, 10 rats with no SCs but homologous dermal acellular matrix with internal scaffolds （Group B ）, 10 with autologous nerves （Group C ）, and the other 10 with nothing （the blank control group, Group D）. The general status of the rats was observed, the wet weight of triceps muscle of calf was monitored, and the histological observation of the regenerated nerves were made at 12 weeks after operation. Results： The wounds of all 40 rats healed after operation and no death was found. No foot ulceration was found in Groups A, B and C, but 7 rats suffered from foot ulceration in Group D. The triceps muscles of calf were depauperated in the experimental sides in all the groups compared with the uninjured sides, which was much more obvious in Group D. The wet weight of triceps muscle of calf and nerve electrophysiologic monitoring showed no statistical difference between Group A and Group C, but statistical difference was found between Groups A and B and Groups B and D. And significant statistical difference was found between Group B and Group D. Obvious compound muscle （ or motor） action potential （CMAP） could be evoked in Group A and Group C, but the evoked amplitude was very low in Group B and Group D. The axons of regenerated nerves penetrated through the whole graft in Group A and Group C, and partly penetrated through the graft in Group B, but did not penetrate in Group D. The two tips of the separated sciatic nerves of Groups A , B , and C were connected together, without formation of neuroma. But those of Group D were not connected together and neuroma formed in 6 rats. Conclusions： Tissue engineered nerves can be used for repairing long defects of the peripheral nerves of rats and ideal repairing effects can be obtained.
基金supported by the Major Research Plan of the National Natural Science Foundation of China(92068112)the National Key Research and Development Program of China(2017YFA0104700)+1 种基金the National Natural Science Foundation of China(82201509)the National Major Project of Research and Development(2022YFA1105500).
文摘Extracellular vesicles from skin-derived precursor Schwann cells(SKP-SC-EVs)promote neurite outgrowth in culture and enhance peripheral nerve regeneration in rats.This study aimed at expanding the application of SKPSC-EVs in nerve grafting by creating a chitosan/PLGA-based,SKP-SC-EVs-containing tissue engineered nerve graft(TENG)to bridge a 40-mm long sciatic nerve defect in dogs.SKP-SC-EVs contained in TENGs significantly accelerated the recovery of hind limb motor and electrophysiological functions,supported the outgrowth and myelination of regenerated axons,and alleviated the denervation-induced atrophy of target muscles in dogs.To clarify the underlying molecular mechanism,we observed that SKP-SC-EVs were rich in a variety of miRNAs linked to the axon growth of neurons,and miR-30b-5p was the most important among others.We further noted that miR-30b-5p contained within SKP-SC-EVs exerted nerve regeneration-promoting effects by targeting the Sin3a/HDAC complex and activating the phosphorylation of ERK,STAT3 or CREB.Our findings suggested that SKP-SC-EVs-incorporating TENGs represent a novel type of bioactive material with potential application for peripheral nerve repair in the clinic.
文摘Background We previously showed that nano-hydroxyapatite/carboxymethyl chitosan （n-Ha/CMCS） displayed excellent mechanical properties, good degradation rates and exceptional biocompatibility, with negligible toxicity. The aim of this study was to determine the effect of the same composite with vascular endothelial growth factor （VEGF）transfected bone marrow stromal cells （BMSCs） in a rabbit radial defect model.
