Collagen-chitosan scaffold impregnated with bone marrow mesenchymal stem cells for treatment of traumatic brain injury

Combinations of biomaterials and cells can effectively target delivery of cells or other therapeutic factors to the brain to rebuild damaged nerve pathways after brain injury.Porous collagen-chitosan scaffolds were prepared by a freeze-drying method based on brain tissue engineering.The scaffolds were impregnated with rat bone marrow mesenchymal stem cells.A traumatic brain injury rat model was established using the 300 g weight free fall impact method.Bone marrow mesenchymal stem cells/collagen-chitosan scaffolds were implanted into the injured brain.Modified neurological severity scores were used to assess the recovery of neurological function.The Morris water maze was employed to determine spatial learning and memory abilities.Hematoxylin-eosin staining was performed to measure pathological changes in brain tissue.Immunohistochemistry was performed for vascular endothelial growth factor and for 5-bromo-2-deoxyuridine(BrdU)/neuron specific enolase and BrdU/glial fibrillary acidic protein.Our results demonstrated that the transplantation of bone marrow mesenchymal stem cells and collagen-chitosan scaffolds to traumatic brain injury rats remarkably reduced modified neurological severity scores,shortened the average latency of the Morris water maze,increased the number of platform crossings,diminished the degeneration of damaged brain tissue,and increased the positive reaction of vascular endothelial growth factor in the transplantation and surrounding areas.At 14 days after transplantation,increased BrdU/glial fibrillary acidic protein expression and decreased BrdU/neuron specific enolase expression were observed in bone marrow mesenchymal stem cells in the injured area.The therapeutic effect of bone marrow mesenchymal stem cells and collagen-chitosan scaffolds was superior to stereotactic injection of bone marrow mesenchymal stem cells alone.To test the biocompatibility and immunogenicity of bone marrow mesenchymal stem cells and collagen-chitosan scaffolds,immunosuppressive cyclosporine was intravenously injected 12 hours before transplantation and 1-5 days after transplantation.The above indicators were similar to those of rats treated with bone marrow mesenchymal stem cells and collagen-chitosan scaffolds only.These findings indicate that transplantation of bone marrow mesenchymal stem cells in a collagen-chitosan scaffold can promote the recovery of neuropathological injury in rats with traumatic brain injury.This approach has the potential to be developed as a treatment for traumatic brain injury in humans.All experimental procedures were approved by the Institutional Animal Investigation Committee of Capital Medical University,China(approval No.AEEI-2015-035)in December 2015.

Combining acellular nerve allografts with brainderived neurotrophic factor transfected bone marrow mesenchymal stem cells restores sciatic nerve injury better than either intervention alone

In this study, we chemically extracted acellular nerve allografts from bilateral sciatic nerves, and repaired 10-mm sciatic nerve defects in rats using these grafts and brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells. Experiments were performed in three groups： the acellular nerve allograft bridging group, acellular nerve allograft ＋ bone marrow mesenchymal stem cells group, and the acellular nerve allograft ＋ brain-derived neurotrophic factor transfected bone marrow mesenchyrnal stem cells group. Results showed that at 8 weeks after bridging, sciatic functional index, triceps wet weight recovery rate, myelin thickness, and number of myelinated nerve fibers were significantly changed in the three groups. Variations were the largest in the acellular nerve allograft ＋ brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells group compared with the other two groups. Experimental findings suggest that chemically extracted acellular nerve allograft combined nerve factor and mesenchymal stem cells can promote the restoration of sciatic nerve defects. The repair effect seen is better than the single application of acellular nerve allograft or acellular nerve allograft combined mesenchymal stem cell transplantation.

Platelet-rich plasma-induced bone marrow mesenchymal stem cells versus autologous nerve grafting for sciatic nerve repair

Autologous nerve grafting is the gold standard of peripheral nerve repair.We previously showed that autologous platelet-rich plasma（PRP）contains high concentrations of growth factors and can induce in vitro cultured bone marrow mesenchymal stem cells（BMSCs）to differentiate into Schwann cells.Here we used PRP-induced BMSCs combined with chemically extracted acellular nerves to repair sciatic nerve defects and compared the effect with autologous nerve grafting.The BMSCs and chemically extracted acellular nerve promoted target muscle wet weight restoration,motor nerve conduction velocity,and axonal and myelin sheath regeneration,with similar effectiveness to autologous nerve grafting.This finding suggests that PRP induced BMSCs can be used to repair peripheral nerve defects.

Nanofibrous Scaffold Containing Osteoblast-Derived Extracellular Matrix for the Proliferation of Bone Marrow Mesenchymal Stem Cells

Extracellular matrix( ECM) plays a prominent role in establishing and maintaining an appropriate microenvironment for tissue regeneration. The aims of this study were to construct a tissue engineered scaffold by reconstituting osteoblast cell-derived ECM( O-ECM) on the electrospun nanofibrous scaffold,and further to evaluate its subsequent application for promoting the proliferation of bone marrow mesenchymal stem cells( BMSCs). To engineer a biomimetic scaffold, calvarial osteoblasts and electrospun poly-llactic acid( PLLA) nanofibers were prepared and subjected to decellularize for O-ECM deposition. To evaluate and characterize the O-ECM/PLLA scaffold, the morphology was examined and several specific mark proteins of osteoblasts matrix were evaluated.Furthermore,the cell counting kit-8( CCK-8) assay was used to detect the proliferation of the BMSCs cultivated on the O-ECM/PLLA scaffold. The results indicated O-ECM/PLLA scaffold was loaded with Collagen I, Fibronectin, and Laminin, as the composition of the marrow ECM. After decellularization,O-ECM deposition was observed in O-ECM/PLLA scaffold. Moreover,the O-ECM/PLLA scaffold could significantly enhance the proliferation of BMSCs,suggesting better cytocompatibility compared to the other groups tested. Taken together,a biomimetic scaffold based on the joint use of O-ECM and PLLA biomaterials,which represents a promising approach to bone tissue engineering, facilitates the expansion of BMSCs in vitro.

Differentiation of mesenchymal stem cells into neuronal cells on fetal bovine acellular dermal matrix as a tissue engineered nerve scaffold

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.

Full-thickness tissue engineered skin constructed with autogenic bone marrow mesenchymal stem cells

To explore the feasibility of repairing clinical cutaneous deficiency, autogenic bone marrow mesen-chymal stem cells (BMSCs) were isolated and differentiated into epidermal cells and fibroblasts in vitro supplemented with different inducing factors and biomaterials to construct functional tissue- engineered skin. The results showed that after 72 h induction, BMSCs displayed morphologic changes such as typical epidermal cell arrangement, from spindle shape to round or oval; tonofibrils, melano-somes and keratohyaline granules were observed under a transmission electronic microscope. The differentiated cells expressed epidermal stem cell surface marker CK19 (59.66% ± 4.2%) and epidermal cells differentiation marker CK10. In addition, the induced epidermal cells acquired the anti-radiation capacity featured by lowered apoptosis following exposure to UVB. On the other hand, the collagen microfibrils deposition was noticed under a transmission electronic microscope after differentiating into dermis fibroblasts; RT-PCR identified collagen type I mRNA expression in differentiated cells; radioimmunoassay detected the secretion of interleukin-6 (IL-6) and interleukin-8 (IL-8) (up to 115.06 pg/mL and 0.84 ng/mL, respectively). Further in vivo implanting BMSCs with scaffold material short-ened skin wound repair significantly. In one word, autogenic BMSCs have the potential to differentiate into epidermal cells and fibroblasts in vitro, and show clinical feasibility acting as epidermis-like and dermis-like seed cells in skin engineering.

Liver regeneration using decellularized splenic scaffold: a novel approach in tissue engineering

BACKGROUND： The potential application of decellularized liver scaffold for liver regeneration is limited by severe shortage of donor organs. Attempt of using heterograft scaffold is accompanied with high risks of zoonosis and immunological rejection. We proposed that the spleen, which procured more extensively than the liver, could be an ideal source of decellularized scaffold for liver regeneration. METHODS： After harvested from donor rat, the spleen was processed by 12-hour freezing/thawing ×2 cycles, then circulation perfusion of 0.02% trypsin and 3% Triton X-100 sequentially through the splenic artery for 32 hours in total to prepare decellularized scaffold. The structure and component characteristics of the scaffold were determined by hematoxylin and eosin and immumohistochemical staining, scanning electron microscope, DNA detection, porosity measurement, biocompatibility and cytocompatibility test. Recellularization of scaffold by 5×106 bone marrow mesenchymal stem cells（BMSCs） was carried out to preliminarily evaluate the feasibility of liver regeneration by BMSCs reseeding and differentiation in decellularized splenic scaffold.RESULTS： After decellularization, a translucent scaffold, which retained the gross shape of the spleen, was generated. Histological evaluation and residual DNA quantitation revealed the remaining of extracellular matrix without nucleus and cytoplasm residue. Immunohistochemical study proved the existence of collagens I, IV, fibronectin, laminin and elastin in decellularized splenic scaffold, which showed a similarity with decellularized liver. A scanning electron microscope presented the remaining three-dimensional porous structure of extracellular matrix and small blood vessels. The poros-ity of scaffold, aperture of 45.36±4.87 μm and pore rate of 80.14%±2.99% was suitable for cell engraftment. Subcutaneous implantation of decellularized scaffold presented good histocompatibility, and recellularization of the splenic scaffold demonstrated that BMSCs could locate and survive in the decellularized matrix. CONCLUSION： Considering the more extensive organ source and satisfying biocompatibility, the present study indicated that the three-dimensional decellularized splenic scaffold might have considerable potential for liver regeneration when combined with BMSCs reseeding and differentiation.

Tissue engineering using mesenchymal stem cells with periosteal wrap for bone defect repair in rabbits

Aim:Mesenchymal stem cells(MSCs)are an excellent potential source of cells for bone tissue engineering due to their excellent renewal ability and osteogenic differentiation capabilities.This study was designed to evaluate the bone formation properties of a demineralized cancellous bone scaffold seeded with MSCs,with or without periosteum,in a critical size bone defect model in rabbits.Methods:Rabbit culture-expanded bone marrow(BM)-MSCs were seeded onto a human demineralized cancellous bone(HDCB)scaffold.Bone defects measuring 15 mm in length were created in each radius.A total of 56 bone defects in 28 rabbits were randomly assigned to one of the 4 groups for scaffold implantation:Group 1:HDCB graft only;Group 2:periosteum-wrapped HDCB graft;Group 3:HDCB graft seeded with BM-MSCs and Group 4:periosteum-wrapped HDCB graft seeded with BM-MSCs.All rabbits were sacrificed 12 weeks after surgery for gross observation,radiological assessment,histological analyses and biomechanical measurements.Results:New bone(NB)formation and bone healing were successfully achieved,both radiologically and histologically,on demineralized cancellous bone graft seeded with BM-MSCs.Results were improved when BM-MSCs were associated with periosteum.Conclusion:This study demonstrates that repair of bone defects in a rabbit model can be achieved through bone grafting using BM-MSCs,implanted on a demineralized cancellous bone scaffold.The formation of NB was optimized when combined with the preservation of periosteum at the site of injury.

颌骨来源骨髓间充质干细胞成骨分化的特点、优势与应用

背景:颌面部骨组织缺损修复是目前研究的热点与难点,而种子细胞的选择是关键。颌骨来源骨髓间充质干细胞是存在于颌骨内的成体间充质干细胞,在颌面部组织再生方面的应用更具优势。目的:综述颌骨来源骨髓间充质干细胞的生物学特性和成骨分化优势以及药物、体内环境、微小RNA对其成骨分化影响的相关研究进展。方法:利用计算机在PubMed和中国知网进行文献检索。中文检索词为“口腔,骨组织工程,干细胞”,英文检索词为“oral,bone tissue engineering,stem cells”。共检索下载文章405篇,根据纳入与排除标准对文章进行筛选,最终纳入70篇文献进行综述。结果与结论:颌骨来源骨髓间充质干细胞是口腔骨组织工程的优良种子细胞,与长骨骨髓间充质干细胞相比,颌骨来源骨髓间充质干细胞具有更强的增殖能力和成骨分化能力。药物、体内环境、微小RNA均可以调控颌骨来源骨髓间充质干细胞的成骨分化,但目前对颌骨来源骨髓间充质干细胞的研究尚处于初始阶段,因此需要更多论证力较强的研究来证实其在颌面部骨组织再生领域的应用更具优势。

In vitro cartilage production using an extracellular matrix-derived scaffold and bone marrow-derived mesenchymal stem cells

Background Cartilage repair is a challenging research area because of the limited healing capacity of adult articular cartilage.We had previously developed a natural,human cartilage extracellular matrix （ECM）-derived scaffold for in vivo cartilage tissue engineering in nude mice.However,before these scaffolds can be used in clinical applications in vivo,the in vitro effects should be further explored.Methods We produced cartilage in vitro using a natural cartilage ECM-derived scaffold.The scaffolds were fabricated by combining a decellularization procedure with a freeze-drying technique and were characterized by scanning electron microscopy （SEM）,micro-computed tomography （micro-CT）,histological staining,cytotoxicity assay,biochemical and biomechanical analysis.After being chondrogenically induced,the induction results of BMSCs were analyzed by histology and Immunohisto-chemistry.The attachment and viability assessment of the cells on scaffolds were analyzed using SEM and LIVE/DEAD staining.Cell-scaffold constructs cultured in vitro for 1 week and 3 weeks were analyzed using histological and immunohistochemical methods.Results SEM and micro-CT revealed a 3-D interconnected porous structure.The majority of the cartilage ECM was found in the scaffold following the removal of cellular debris,and stained positive for safranin O and collagen Ⅱ.Viability staining indicated no cytotoxic effects of the scaffold.Biochemical analysis showed that collagen content was （708.2±44.7）μg/mg,with GAG （254.7±25.9） μg/mg.Mechanical testing showed the compression moduli （E） were （1.226±0.288） and （0.052±0.007） MPa in dry and wet conditions,respectively.Isolated canine bone marrow-derived stem cells （BMSCs） were induced down a chondrogenic pathway,labeled with PKH26,and seeded onto the scaffold.Immunofluorescent staining of the cell-scaffold constructs indicated that chondrocyte-like cells were derived from seeded BMSCs and excreted ECM.The cell-scaffold constructs contained pink,smooth and translucent cartilage-like tissue after 3 weeks of culture.We observed evenly distributed cartilage ECM proteoglycans and collagen type Ⅱ around seeded BMSCs on the surface and inside the pores throughout the scaffold.Conclusion This study stuggests that a cartilage ECM scaffold holds much promise for in vitro cartilage tissue engineering.

中医药联合医用水凝胶治疗疾病的策略及意义

背景:医用水凝胶是具有三维结构网络的新型功能高分子材料,具有出色的生物相容性,目前已在组织工程领域、药物载体领域有广泛研究,但基于组织工程探究医用水凝胶与中医药结合治疗疾病的研究还处于初期探索阶段。因此,通过对医用水凝胶机制作用的剖析,整合医用水凝胶与中医药在研究中联合应用的文章,进而更好地为科研工作者提供思路,对中医药与医用水凝胶联合应用具有重要意义。目的:基于组织工程研究探讨中医药联合医用水凝胶治疗疾病的策略及意义。方法:利用PubMed和中国知网数据库,检索有关中医药联合医用水凝胶在组织工程中应用的文献,检索时间为2010年1月至2022年11月,英文检索词为“hydrogel,traditional Chinese medicine,drug carrier,tissue engineering”,中文检索词为“医用水凝胶、中医药、药物载体、组织工程”。根据纳入与排除标准对所有文章进行初筛后,最终纳入61篇文章进行综述。结果与结论:①中医药联合医用水凝胶的应用虽然在关节内、组织器官内、软组织伤口和组织工程等方面有所涉及,但除了中医药结合水凝胶敷料在临床应用治疗软组织损伤外,其他方面尚处于基础实验阶段。②中医药联合医用水凝胶的发展有着巨大潜力和发展前景,但对于性能要求较高的凝胶在制造方面存在一定难度,理化性质精确掌握难度较大。③目前综合来看可注射水凝胶凭借着简便易用的特点,其在与中医药联合使用可延伸范围较广,可用于关节、器官和组织工程相关疾病的治疗;智能水凝胶有较高的灵敏度和可逆转化性也可满足特殊环境下的使用;将两者结合的中医药使用过程中还需要明确中药成分的作用机制。④中医药联合医用水凝胶治疗疾病的策略应着手于中医药对器官、组织、细胞的治疗作用联合适当种类的医用水凝胶进行匹配,可弥补传统中医给药方式和频繁给药的不足,在组织工程方面可以用水凝胶负载中药干预后的干细胞,或者同时负载中药和干细胞用于相关疾病的治疗。⑤在中医药联合医用水凝胶应用的未来研究中,还需要考虑:应当确保医用水凝胶生物性能可以量化,以不同材料不同制造工艺把握水凝胶特性,制造出所需要的符合应用条件的医用水凝胶;在中医药方面需要对已知中药单体、中药复方提取物的治疗效果和应用机制全面了解剖析,在更明了的机制下实现中医药与医用水凝胶更多更完美的结合;借助医学科技创新能力的不断提高,医用水凝胶可以创新性地结合中医药其他传统治疗方法比如针灸、推拿和拔罐等方式进行多角度运用。

Mesenchymal stem cells loaded on 3D-printed gradient poly(e-caprolactone)/methacrylated alginate composite scaffolds for cartilage tissue engineering

Cartilage has limited self-repair ability due to its avascular,alymphatic and aneural features.The combination of three-dimensional(3D)printing and tissue engineering provides an up-and-coming approach to address this issue.Here,we designed and fabricated a tri-layered(superficial layer(SL),middle layer(ML)and deep layer(DL))stratified scaffold,inspired by the architecture of collagen fibers in native cartilage tissue.The scaffold was composed of 3D printed depth-dependent gradient poly(e-caprolactone)(PCL)impregnated with methacrylated alginate(ALMA),and its morphological analysis and mechanical properties were tested.To prove the feasibility of the composite scaffolds for cartilage regeneration,the viability,proliferation,collagen deposition and chondrogenic differentiation of embedded rat bone marrow mesenchymal stem cells(BMSCs)in the scaffolds were assessed by Live/dead assay,CCK-8,DNA content,cell morphology,immunofluorescence and real-time reverse transcription polymerase chain reaction.BMSCs-loaded gradient PCL/ALMA scaffolds showed excellent cell survival,cell proliferation,cell morphology,collagen II deposition and hopeful chondrogenic differentiation compared with three individual-layer scaffolds.Hence,our study demonstrates the potential use of the gradient PCL/ALMA construct for enhanced cartilage tissue engineering.

不同静电纺丝膜上骨髓间充质干细胞的黏附、增殖与成血管平滑肌分化

背景:临床上迫切需要小口径人工血管来治疗冠状动脉和外周动脉疾病,目前,血管组织工程已成为制备小口径人工血管的主要方法,选择合适的生物材料和细胞来源是小口径组织工程血管构建成功的关键因素。目的:观察4种静电纺丝膜材料对骨髓间充质干细胞增殖、黏附及分化为血管平滑肌细胞的影响。方法:分离提取SD大鼠骨髓间充质干细胞。将骨髓间充质干细胞分别接种于聚己内酯(PCL)、聚己内酯-透明质酸(PCL-HA)、聚己内酯-丝素蛋白(PCL-SF)、聚己内酯-明胶(PCL-GEL)静电纺丝膜材料上,培养1,3,7 d后,扫描电镜下观察材料上的细胞排布,鬼笔环肽染色观察材料上的细胞增殖与黏附,qRT-PCR检测材料上细胞分泌的CD90、Meflin、转化生长因子βmRNA表达;向血管平滑肌细胞诱导分化7 d后,qRT-PCR检测材料上细胞ɑ-平滑肌肌动蛋白mRNA表达。结果与结论:①扫描电镜下可见骨髓间充质干细胞在4种静电纺丝膜上均沿着静电纺丝膜的纤维走向排列;②鬼笔环肽染色显示,骨髓间充质干细胞在4种静电纺丝膜上分布规律,均沿着纤维走向呈现平行分布,并且PCL-HA、PCL-SF、PCL-GEL静电纺丝膜较PCL静电纺丝膜更有利于骨髓间充质干细胞的增殖、黏附,PCL-SF静电纺丝膜相较于PCL-HA、PCL-GEL静电纺丝膜更有利于骨髓间充质干细胞的增殖、黏附;③qRT-PCR检测显示,4种静电纺丝膜材料均可维持骨髓间充质干细胞CD90和Meflin的mRNA表达,组间比较差异无显著性意义(P>0.05);PCL-HA、PCL-SF、PCL-GEL组培养1,7 d的转化生长因子βmRNA表达高于PCL组(P<0.05),PCL-SF组培养3,7 d的转化生长因子βmRNA表达高于其他3组(P<0.05),PCL-HA组培养7 d的转化生长因子βmRNA表达高于PCL-GEL组(P<0.05);④qRT-PCR检测显示,PCL-SF组ɑ-平滑肌肌动蛋白mRNA表达高于其他3组(P<0.05),PCL-HA组ɑ-平滑肌肌动蛋白mRNA表达高于PCL组(P<0.05);⑤结果表明:相较于PCL、PCL-HA、PCL-GEL静电纺丝膜,PCL-SF静电纺丝膜与骨髓间充质干细胞结合更适合制备小口径组织工程血管。

Repair of sciatic nerve defects using tissue engineered nerves

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.

Comparison of Osteogenesis Between Two Kinds of Stem Cells from Goat Combined Calcium Phosphate Cement in Tissue Engineering

To explore the possible mechanism of osteogenesis for deciduous teeth stem cells (DTSCs) in vivo/ vitro, stem cells from goat deciduous teeth (SGDs) were firstly isolated, induced and transplanted into immunocompromised mice. The SGDs's mineralization pattern and osteogenesis were compared with bone marrow messenchymal stem cells (BMMSCs) from goats. SGDs have similar osteogenic differentiation pattern in vitro and bone-like tissue formation mechanism in vivo to BMMSCs; moreover SGDs have stronger alkaline phosphatase (ALP) gene expression and osteopontin (OPN) gene expression levels than BMMSCs; also SGDs can form more bone-like tissues than BMMSCs when cell-scaffold compounds are transplanted into immunocompromised mice. This pre-clinical study in a large-animal model confirms that DTSCs may be an appropriate source of stem cells in repairing bone defects with tissue engineering.

骨髓间充质干细胞在口腔医学中的应用

在颌面部,由肿瘤、创伤、先天或后天性的急性引起的组织缺损使患者的容貌、正常生理功能及心理健康受到严重伤害,使患者的整体生活质量下降。寻找适当的治疗方法来恢复这些缺损的组织是至关重要的。近年来,随着组织工程的飞速发展,人们对于口腔组织再生技术十分重视。骨髓来源的间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)由于其具有向其他细胞分化的潜力,最常被应用于再生医学。近年来,研究人员开发了许多基于BMSCs的生物支架材料,利用其卓越的性能以满足口腔颌面组织工程的需求。综述BMSCs在口腔医学的研究进展及应用,探讨其在口腔医学中的应用前景。

慢病毒下调酪蛋白激酶2相互作用蛋白1表达的骨髓间充质干细胞修复骨质疏松大鼠牙槽骨缺损

背景:在口腔牙槽骨修复治疗过程中,患者的全身状况尤其是骨质疏松症等系统性疾病往往会影响治疗效果,近来发现酪蛋白激酶2相互作用蛋白1(casein kinase 2-interacting protein-1,CKIP-1)具有负调控成骨的作用,因此有希望作为新的靶点用于治疗骨质疏松。目的:探究CKIP-1表达下调的骨髓间充质干细胞对骨质疏松大鼠牙槽骨缺损的修复作用。方法:(1)从1周龄SD乳鼠股骨及胫骨骨髓中分离培养出骨髓间充质干细胞,慢病毒转染降低骨髓间充质干细胞中的CKIP-1水平;(2)通过维甲酸[70 mg/(kg·d)]灌胃21 d的方式建立骨质疏松SD大鼠模型,将CKIP-1表达下调的骨髓间充质干细胞与明胶海绵复合体、空载体转染的骨髓间充质干细胞与明胶海绵复合体、明胶海绵分别植入骨质疏松大鼠模型牙槽骨缺损内,正常组将明胶海绵植入正常大鼠牙槽骨缺损内。术后正常喂养30 d后,取上颌牙槽骨进行苏木精-伊红染色并通过Lane-Sandhu组织学评分和新生骨百分比评估骨缺损愈合情况,通过q RTPCR技术和免疫组化染色检测各组术区牙槽骨中CKIP-1、碱性磷酸酶、骨钙素的m RNA相对表达量及阳性表达情况。结果与结论:(1)与空载体转染相比,CKIP-1下调后的骨髓间充质干细胞中CKIP-1的m RNA相对表达量减少(P <0.05);(2)与植入空载体转染的骨髓间充质干细胞与明胶海绵复合体组、明胶海绵组相比,植入CKIP-1表达下调的骨髓间充质干细胞与明胶海绵复合体组Lane-Sandhu组织学评分和新生骨百分比显著升高(P <0.05),与正常组相近;碱性磷酸酶、骨钙素表达显著升高(P <0.05);(3)结果说明,局部移植CKIP-1下调的骨髓间充质干细胞对骨质疏松大鼠牙槽骨缺损具有明显的修复作用。

转化生长因子β诱导骨髓间充质干细胞分化为半月板纤维软骨细胞

背景:骨髓间充质干细胞可被诱导分化为纤维软骨细胞作为组织工程中的种子细胞,而细胞因子转化生长因子β(transforming growth factor beta,TGF-β1)和TGF-β3在诱导分化中的作用,目前仍是研究热点。目的:综述TGF-β1和TGF-β3诱导骨髓间充质干细胞分化为软骨细胞的相关文献,为进一步指导TGF-β诱导骨髓间充质干细胞分化为纤维软骨细胞提供理论依据,同时为今后的临床上半月板损伤和退化的治疗提供新策略。方法:计算机检索PubMed、中国知网和万方医学网数据库1990年1月至2022年3月收录的文献。英文数据库检索词:“Transforming Growth Factor beta 1,Transforming Growth Factor beta 3,Bone Marrow Mesenchymal Stem Cells,meniscus,fibrochondrocyte”,中文数据库检索词:“TGF-β1,TGF-β3、骨髓间充质干细胞、半月板、纤维软骨细胞”,根据筛选标准,最终纳入78篇文献进行综述。结果与结论:①TGF-β主要分为TGF-β1、TGF-β2和TGF-β3共3种亚型,其中研究较多的细胞因子是TGF-β1和TGF-β3。②TGF-β主要通过TGF-β/Smad信号通路激活细胞核内的SOX9转录因子进而诱导骨髓间充质干细胞分化为软骨细胞。③TGF-β1可诱导骨髓间充质干细胞向透明软骨细胞分化,伴随培养时间的延长,透明软骨细胞可发生去分化变成纤维软骨细胞;④TGF-β3主要诱导骨髓间充质干细胞向纤维软骨细胞分化,同时促进Ⅰ型胶原和蛋白聚糖合成分泌。⑤目前,关于半月板损伤和退化的治疗,临床上采用的治疗方案均不理想,组织工程的出现为其治疗提供了新的策略,具体方法是将诱导因子和骨髓间充质干细胞植入到半月板损伤部位,可以起到恢复半月板功能及再生纤维软骨组织的作用,但是目前实验尚处于体外和动物模型阶段,其在临床上用于半月板受损的治疗和可用性仍需要后续大量的临床研究去研究探索。

镁基合金支架负载兔源性骨髓间充质干细胞构建组织工程化骨复合支架

背景:随着组织工程学的深入研究与快速发展,组织工程支架为骨组织修复提供了新的契机。目的:探讨骨髓间充质干细胞与镁基合金支架共培养后的细胞增殖情况,为组织工程化骨支架构建提供新思路,进而为组织工程支架临床应用提供依据。方法:无菌条件下抽取兔骨髓液,使用全骨髓贴壁法分离培养骨髓间充质干细胞,进行流式细胞学鉴定和成骨分化能力检测。将第3代骨髓间充质干细胞分别接种于镁基合金支架和壳聚糖-聚己内酯-磷酸三钙支架,第3,10天进行激光共聚焦扫描观察细胞生长情况,第1,4,7,10,14天行MTT检测细胞增殖情况。结果与结论:(1)分离培养的细胞高表达CD44和CD90,几乎不表达CD34;(2)经成骨诱导后,细胞形态逐渐变为多角形,Von Kossa染色可见细胞之间钙结节染成黑色,碱性磷酸酶活性较未诱导对照组升高(P<0.05);(3)镁基合金与骨髓间充质干细胞共培养后,共聚焦显微镜扫描显示细胞可在支架上黏附并呈团块状生长,随着时间延长,镁基合金支架较壳聚糖-聚己内酯-磷酸三钙支架上负载的细胞增殖活跃,细胞数量明显增多,MTT检测显示两种材料与骨髓间充质干细胞共培养至10 d时,细胞生长达峰值;(4)结果表明,兔源性骨髓间充质干细胞具有良好的成骨潜能,与镁基合金支架共培养后,细胞在支架表面生长良好,成功构建了组织工程化镁基种植体。

碳点基材料在骨组织工程中的应用

背景:碳点在改善聚合物和生物陶瓷等材料性能以及促进成骨分化能力方面显示出巨大潜力,因此碳点基支架材料有望成为一种新型的骨修复候选材料。目的:介绍碳点基材料在骨组织工程中的应用进展,旨在为碳点基支架材料的合理设计提供一些研究思路上的参考。方法:由第一作者以“碳点、骨组织工程、骨支架材料”为中文检索词,以“Carbon dots,carbon-based nanomaterials,bone tissue engineering,bone scaffold materials”为英文检索词,检索万方和Web of Science数据库中2014-2022年发表的相关文献,最后对75篇文献进行分析总结。结果与结论:①原始碳点基材料和碳点基复合材料具有合成方法多样、操作简单、材料来源广和绿色环保等优势,然而其在体内的修复骨缺损能力不足以及生物安全性、生物降解以及临床适应性等都需要进一步评估。②碳点基纤维材料具有骨支架所需优异的机械性能、热稳定性以及生物降解性,是一类具有较大优越性及应用潜力的骨修复新材料;目前应用于体内的骨修复能力有限,因此需要和骨形态发生蛋白、生长因子以及血管化因子复合以期进一步改善这一类材料的骨修复能力。③碳点基水凝胶支架材料在修复骨缺损方面展现出了巨大的潜力,是目前报道的修复骨缺损效果最好的一种碳点基支架材料,然而该材料制备工艺复杂且周期长,因而将其推广到临床骨修复应用上还需要进一步探索合成简单易造作且周期短的替代方法。④原始碳点基材料、碳点基复合材料、碳点基纤维材料和碳点基水凝胶材料各有优缺点,目前来看碳点基水凝胶修复骨缺损效果最好。⑤碳点基材料应用于骨组织工程研究仍然处于初期的探索阶段,体内的溶血性、生物降解、代谢过程及预后评估等需要深入研究,同时其机械性能、热稳定性以及促成骨能力均有待进一步提高和改善。⑥未来研究需结合多种测试及表征手段深入分析碳点基材料的促成骨机制,这是设计应用于临床碳点基骨支架材料的前提。