The combination of induced pluripotent stem cells and bioscaffolds holds promise for spinal cord regeneration

Spinal cord injuries（SCIs） are debilitating conditions for which no effective treatment currently exists. The damage of neural tissue causes disruption of neural tracts and neuron loss in the spinal cord. Stem cell replacement offers a solution for SCI treatment by providing a source of therapeutic cells for neural function restoration. Induced pluripotent stem cells（i PSCs） have been investigated as a potential type of stem cell for such therapies. Transplantation of i PSCs has been shown to be effective in restoring function after SCIs in animal models while they circumvent ethical and immunological concerns produced by other stem cell types. Another approach for the treatment of SCI involves the graft of a bioscaffold at the site of injury to create a microenvironment that enhances cellular viability and guides the growing axons. Studies suggest that a combination of these two treatment methods could have a synergistic effect on functional recovery post-neural injury. While much progress has been made, more research is needed before clinical trials are possible. This review highlights recent advancements using i PSCs and bioscaffolds for treatment of SCI.

The effect of porosity on the mechanical properties of 3D-printed triplyperiodic minimal surface (TPMS) bioscaffold

Prevailing tissue degeneration caused by musculoskeletal maladies poses a great demand on bioscaffolds,which are artificial,biocompatible structures implanted into human bodies with appropriate mechanical properties.Recent advances in additive manufacturing,i.e.,3D printing,facilitated the fabrication of bioscaffolds with unprecedented geometrical complexity and size flexibility and allowed for the fabrication of topologies that would not have been achieved otherwise.In our work,we explored the effect of porosity on themechanical properties of a periodic cellular structure.The structure was derived from the mathematically created triply periodic minimal surface(TPMS),namely the Sheet-Diamond topology.First,we employed a series of software including MathMod,Meshmixer,Netfabb and Cura to design the model.Then,we utilized additive manufacturing technology to fabricate the cellular structures with designated scale.Finally,we performed compressive testing to deduce the mechanical properties of each cellular structure.Results showed that,in comparison with the highporosity group,the yield strength of the low-porosity group was 3 times higher,and the modulus was 2.5 times larger.Our experiments revealed a specific relationship between porosity and Young’s modulus of PLA-made Sheet-Diamond TPMS structure.Moreover,it was observed that the high-and low-porosity structures failed through distinctive mechanisms,with the former breaking down via buckling and the latter via micro-fracturing.

Matrix bound vesicles and miRNA cargoes are bioactive factors within extracellular matrix bioscaffolds

Injury to central nervous system （CNS） tissues in adult mam- mals often leads to neuronal loss, scarring, and permanently lost neurologic functions, and this default healing response is increasingly linked to a pro-inflammatory innate immune response. Extracellular matrix （ECM） technology can reduce inflammation, while increasing functional tissue remodeling in various tissues and organs, including the CNS.

Bioscaffolds embedded with regulatory modules for cell growth and tissue formation:A review

The demand for artificial organs has greatly increased because of various aging-associated diseases and the wide need for organ transplants.A recent trend in tissue engineering is the precise reconstruction of tissues by the growth of cells adhering to bioscaffolds,which are three-dimensional(3D)structures that guide tissue and organ formation.Bioscaffolds used to fabricate bionic tissues should be able to not only guide cell growth but also regulate cell behaviors.Common regulation methods include biophysical and biochemical stimulations.Biophysical stimulation cues include matrix hardness,external stress and strain,surface topology,and electromagnetic field and concentration,whereas biochemical stimulation cues include growth factors,proteins,kinases,and magnetic nanoparticles.This review discusses bioink preparation,3D bioprinting(including extrusion-based,inkjet,and ultraviolet-assisted 3D bioprinting),and regulation of cell behaviors.In particular,it provides an overview of state-of-the-art methods and devices for regulating cell growth and tissue formation and the effects of biophysical and biochemical stimulations on cell behaviors.In addition,the fabrication of bioscaffolds embedded with regulatory modules for biomimetic tissue preparation is explained.Finally,challenges in cell growth regulation and future research directions are presented.

Microfluidic bioscaffolds for regenerative engineering

The construction of biological scaffolds in vitro plays an important role in regenerative engineering,which has always been the focus of research.The application of scaffold materials combined with cells/bioactive factors is considered to have great potential for tissue regeneration.Various strategies have been used for constructing biological scaffolds,including electrospinning/spraying,direct laser writing,solvent casting,and microfluidic technology.Among them,considering the advantages of the safety,low cost,and highly controllable process-ing characteristics,microfluidic technology has become a promising method for the construction of biological scaffolds.This review overviews the recent research progress of microfluidic bioscaffolds for regenerative engi-neering.Firstly,we introduce some typical natural polymers which are widely used to construct bioscaffolds.Then,the bioscaffolds with different structures templated by microfluidic droplets and fibers are described.Fur-therly,we talk about the different applications of microfluidic bioscaffolds in wound healing,drug delivery,bone regeneration,and nerve regeneration.Finally,the challenges and future prospects of the development of microfluidic bioscaffolds for regenerative engineering are discussed.

外泌体复合生物支架在牙髓再生中的应用

现有根管治疗技术缺乏理想的长期效果,世界范围内对牙髓再生的关注日益增长,牙髓再生已成为口腔医学领域的重点研究方向。外泌体是一类来源于核内体的多囊泡结构物,随着细胞生物学和组织工程技术的快速发展,外泌体复合生物支架应用于牙髓再生领域已成为新趋势,其中植物来源外泌体和牙髓干细胞来源外泌体在牙髓再生领域的应用具有广阔前景,对外泌体复合生物支架在牙髓再生中应用的研究可为生物支架搭载外泌体实现牙髓组织的再生和功能重建提供新思路。

组织工程血管脱细胞支架主要组织相容性复合体Ⅰ抗原性的免疫组化观察

目的 :探讨组织工程血管全生物支架的免疫原性的检测方法。 方法 :取猪颈总动脉 ,剥离外膜 ,剪成 2 cm长的血管段 ,随机分为 3组 :对照组、脱细胞组和脱细胞加碱处理组 ,每组 5段。以免疫组化方法检测血管脱细胞处理前后主要组织相容性复合体 (MHC )的变化情况 ,并将检测结果作图像分析。 结果 :对照组血管壁 MHC 高表达 ,脱细胞处理后 MHC 明显减少。图像分析得出阳性颗粒面积与视场面积比 :对照组为 (2 4 .0 9± 6 .0 9) % ,脱细胞组为 (6 .71± 2 .1 8) % ,脱细胞加碱处理组为 (1 .6 2± 0 .76 ) % ,组间比较差别均有显著意义 (P<0 .0 1 )。 结论 :MHC

小鼠胰岛与肝脏脱细胞支架三维共培养的实验研究

目的 :观测原代小鼠胰岛在小鼠肝脏脱细胞支架上的生长及功能,探讨组织工程学治疗糖尿病的新方法。方法 :运用灌注法制备结构完整的全肝脏脱细胞支架并检测,运用胶原酶P灌注消化法获取小鼠胰岛细胞并行结构和功能完整性分析;将分离纯化的原代小鼠胰岛经门静脉灌入全肝脏脱细胞支架,在三维培养体系培养后进行相关检测。结果:肝脏脱细胞支架检测未见细胞结构,胶原结构保存完整。支架内剩余DNA定量检测结果为(38±11)ng/mg ds DNA,生物相容性良好。双硫腙(dithizone,DTZ)染色胰岛呈现特异性猩红色,葡萄糖刺激实验提示在高糖组中培养的原代小鼠胰岛胰岛素分泌功能明显高于低糖组,差异具有统计学意义(P<0.01);将胰岛移植入肝脏脱细胞支架培养5 d后胰岛定植,且胰岛素基因表达显示脱细胞支架三维培养中胰岛素表达水平与平面培养相比,差异具有统计学意义(P<0.01)。结论:肝脏脱细胞支架三维培养体系中的胰岛细胞比传统平面培养具有更佳的细胞活力及更有效的胰岛素分泌功能。

三维打印聚乳酸基石墨烯复合材料生物支架的工艺与性能研究

概述了生物支架的特征,通过试验分析了不同三维打印参数对聚乳酸基石墨烯复合材料生物支架的工艺与性能影响,确认在喷头直径0.2 mm、打印温度230℃、打印速度40 mm/s的条件下,所打印出的生物支架质量较为理想。

生物支架在肩袖损伤修复应用中的研究进展

肩袖撕裂损伤是造成肩关节功能下降、疼痛常见的一种疾病，尽管目前对于肩袖撕裂损伤的研究进展以及外科治疗手段都有了很大的进步，但肩袖修复的失败率依然为20％～90％，因此临床上需要能有效地提高力学强度并能刺激患者内在愈合潜能的修补材料。近年来应用于肩袖修复的生物支架材料已成为研究的热点，文中就目前治疗肩袖损伤的生物支架材料研究进展进行概述。

Extracellular matrices for gastrointestinal surgery:Ex vivo testing and current applications

AIM:To assess the effects of bile and pancreatic juice on structural and mechanical resistance of extracellular matrices(ECMs) in vitro.METHODS:Small-intestinal submucosa(SIS),porcine dermal matrix(PDM),porcine pericardial matrix(PPM) and bovine pericardial matrix(BPM) were incubated in human bile and pancreatic juice in vitro.ECMs were examined by macroscopic observation,scanning electron microscopy(SEM) and testing of mechanical resistance.RESULTS:PDM dissolved within 4 d after exposure to bile or pancreatic juice.SIS,PPM and PDM retained their integrity for > 60 d when incubated in either digestive juice.The effect of bile was found to be far more detrimental to mechanical stability than pancreatic juice in all tested materials.In SIS,the loss of mechanical stability after incubation in either of the digestive secretions was less distinct than in PPM and BPM [mFmax 4.01/14.27 N(SIS) vs 2.08/5.23 N(PPM) vs 1.48/7.89 N(BPM)].In SIS,the extent of structural damage revealed by SEM was more evident in bile than in pancreatic juice.In PPM and BPM,structural damage was comparable in both media.CONCLUSION:PDM is less suitable for support of gastrointestinal healing.Besides SIS,PPM and BPM should also be evaluated experimentally for gastrointestinal indications.

异丙肾上腺素在去细胞化肝脏再细胞化过程中的作用研究

目的通过制备去细胞化肝脏生物支架,研究自主神经递质对再细胞化过程的影响。方法使用全器官灌注法制备C57小鼠去细胞化肝脏生物支架,行HE和Massion染色鉴定并检测DNA残留量;RT-PCR和免疫荧光化学检测再细胞化肝脏中种子细胞(L02细胞株)肾上腺素能受体及亚型表达情况;检测平面以及生物支架内异丙肾上腺素对种子细胞增殖的作用,以及对再细胞化肝脏白蛋白分泌和尿素合成水平的影响。结果成功制备去细胞化肝脏生物支架,L02细胞株表达肾上腺素能受体,并且β2受体亚型表达相对较高;异丙肾上腺素显著促进平面及再细胞化肝脏内L02细胞株增殖,并增强再细胞化肝脏分泌白蛋白和尿素。结论异丙肾上腺素可促进去细胞化肝脏生物支架再细胞化进程。

门静脉输注胰岛至大鼠肝脏去细胞支架的结果考察

目的考察经门静脉移植后的胰岛细胞团在肝脏血管中停留的位置及栓塞部位细胞团的相关情况。方法取成年SD大鼠,从门静脉在体灌注去污剂十二烷基硫酸钠溶液使肝脏去细胞化,再输注经二苯基硫卡巴腙染色后的大鼠胰岛,在体视显微镜和倒置显微镜下观察去细胞后的肝脏内部的血管和胰岛在肝脏内的栓塞情况。结果大鼠肝脏在体去细胞后,可得到完全透明、清晰的肝脏支架,肝内血管分支丰富,经灌注(800±250)IEQ胰岛细胞团后,胰岛主要栓塞在大鼠肝脏边缘血管末梢,有时一个血管末梢栓塞有多个细胞团。结论本方法模拟大鼠门静脉内胰岛移植,呈现了胰岛在肝脏内血管的驻留情况,有利于进一步研究改进胰岛在肝内移植后的存活和功能发挥。

去细胞肌肉生物支架与人脐带间充质干细胞的相容性

背景:去细胞肌肉生物支架联合人脐带间充质干细胞移植将是治疗脊髓损伤的一项重要措施。但两者是否具有良好的相容性,人脐带间充质干细胞能否在去细胞肌肉生物支架中长期存活并均匀分布,尚未得到证实。目的:观察大鼠去细胞肌肉生物支架与人脐带间充质干细胞的相容性。方法:改良化学法制备大鼠去细胞肌肉生物支架,将第3代人脐带间充质干细胞Hoechest33342荧光标记后分为3组进行实验,细胞+支架组、细胞+支架大鼠体内组和单纯细胞组。分别应用苏木精-伊红、Masson染色方法观察去细胞肌肉生物支架的组织形态,以荧光倒置相差显微镜和扫描电镜观察人脐带间充质干细胞的吸附和生长情况。结果与结论:人脐带间充质干细胞与去细胞肌肉生物支架充分附着,生长增殖活跃,细胞在支架内分布均匀。细胞+支架体内组与细胞+支架组相比在移植后1-7d人脐带间充质干细胞数量差异无显著性意义(P>0.05),在移植14d细胞+支架体内组人脐带间充质干细胞数量大于细胞+支架组(P<0.05)。提示去细胞肌肉生物支架与人脐带间充质干细胞有较好的相容性,体内环境更有利于细胞增殖和两者融合。

三种方法构建去细胞肌肉生物支架:组织学和生物力学比较

背景:目前构建去细胞肌肉生物支架的方法有化学方法、物理冻融法和冷冻法。目的:比较化学方法、物理冻融法和改良化学方法构建去细胞肌肉生物支架组织学和生物力学差异。方法:将6只SD大鼠竖脊肌截成36段,随机分3组(n=12),分别采用化学方法、物理冻融方法、改良化学方法构建去细胞肌肉生物支架。将荧光标记的骨髓间充质干细胞接种于3组支架上进行组织学与生物力学检测。结果与结论:苏木精-伊红染色显示3组支架均可见平行排列结构,化学方法组连续性较物理冻融组、改良化学组差;Masson染色显示3组支架结构主要为胶原纤维,物理冻融组残留一些肌纤维,间隙不均。物理冻融组第7天荧光标记种子细胞计数明显少于化学方法组、改良化学组(P<0.01),第14天物理冻融组少于改良化学组(P<0.05);扫描电镜可见细胞贴附各组支架生长、大量存活;物理冻融组最大载荷高于化学方法组(P<0.05),与改良化学组无明显差别,各组弹性模量无差异;物理冻融组孔隙率低于化学方法组、改良化学组(P<0.05)。表明改良化学方法可以兼顾支架的组织学和生物力学特性,彻底清除肌细胞,较多保留更完整的细胞外基质。

A nanostructured look of collagen apatite porosity into human mineralized collagen fibril

Bone tissue is a hierarchical material characterized at nanoscale by the mineralized collagen fibril,a recurring structure mainly composed of apatite minerals,collagen and water.Bone nanostructure has a fundamental role in determining the mechanical behavior of the tissue and its mass transport properties.Diffusion phenomenon allows to maintain an adequate supply of metabolites in the mechanisms of bone remodeling,adaptation and repair.Several analytical and computational models have been developed to analyze and predict bone tissue behavior.However,the fine replication of the natural tissue still represents a challenge.Insights on the structural organization at nanoscale and on the influence of apatite mineral crystals on the diffusion coefficient lead to outline the functional conditions for the development of biomimetic strategies for bone tissue engineering.Thorough understanding of bone nanostructure is essential to improve longevity of bioscaffolds and to decrease the risk of failure by controlling their mechanical and biological performance.

骨髓间充质干细胞-海螵蛸复合支架的细胞毒性评估

背景:作为骨组织工程的修复材料,应具有良好的生物相容性及降解吸收性,许多学者在此方面也做了较深入的研究,但中药复合细胞生物支架的研究较少。目的:参照《医疗器械生物学评价标准》,对兔骨髓间充质干细胞-海螵蛸生物复合支架进行体外细胞毒性检测,评估复合支架的毒性等级,为临床应用生物支架提供实验依据。方法:依照ISO标准按“材料面积∶浸提介质体积=3-6 cm 2∶1 mL”制作骨髓间充质干细胞-海螵蛸生物复合支架材料浸提液。制备L-929细胞悬液,将细胞浓度调整至1×107 L-1进行接种培养,设置阳性对照组(含苯酚的DMEM培养液)、实验组(材料浸提液)、阴性对照组(新鲜DMEM培养液)。培养24,48,72 h采用MTT法检测L-929细胞吸光度值,计算各组细胞的相对增殖率,评估复合支架毒性等级。结果与结论:实验组、阴性对照组、阳性对照组吸光度值在不同时间点不完全相同(P=0.000<0.01),各时间点内比较,实验组与阴性对照组吸光度值均明显高于阳性对照组(P<0.01);兔骨髓间充质干细胞-海螵蛸生物支架细胞毒性为1级。结果说明兔骨髓间充质干细胞-海螵蛸生物支架无明显毒性作用,符合生物材料应用要求。

脐带间充质干细胞在骨组织工程中的应用与进展

背景:由于脐带间充质干细胞作为种子细胞具有来源广泛、取材方便、免疫原性低、成骨分化潜能及较强的增殖和自我更新能力等诸多优点,近年来关于其在骨组织工程中的应用研究越来越多。目的:对脐带间充质干细胞在分离培养、成骨诱导及其支架等方面进行综述。方法:由第一作者检索CNKI数据库和PubMed数据库的相关文章,检索词为“人脐带间充质干细胞,分离,培养,成骨分化,支架,骨组织工程”“human umbilical cord stem cell,culture,isolation,osteogenic differentiation,scaffold,bone tissue engineering”,查阅2004至2020年来有关的文献,最终纳入104篇文献。结果与结论:脐带间充质干细胞的分离、培养方法不一;无血清或动物血清替代物培养体系及细胞共培养技术取得了很大进展;3D培养系统将是未来发展的方向。脐带间充质干细胞成骨诱导分化机制仍不明确,需深入研究阐明。至今,开发取长补短、性能更优的复合支架仍是研究的焦点;生物打印技术突破了微米级精确调控支架内部复杂结构打造个体化支架的难题,为骨组织工程带来巨大希望;设计和制造出具有多方面理想组成(包括生物相容性、微观及宏观高孔隙率、机械性能和相关生物可吸收性等)及减少临床治疗不良反应的支架仍是骨组织工程中的重要挑战之一。

载重组人骨形态发生蛋白2/聚乳酸缓释微球的复合生物支架修复超临界骨缺损

背景:前期研究制备了丝素/壳聚糖/纳米羟基磷灰石复合生物支架与重组人骨形态发生蛋白2/聚乳酸缓释系统。目的:观察载重组人骨形态发生蛋白2/聚乳酸缓释微球的复合生物支架,修复犬桡骨超临界骨缺损的效果。方法:采用真空冷冻干燥及化学交联法制备丝素蛋白/壳聚糖/纳米羟基磷灰石生物支架,采用复乳挥发法制备重组人骨形态发生蛋白2/聚乳酸缓释微球,将二者整合后制备成载缓释微球的三维生物支架。取8只成年犬(遵义医科大学实验动物中心提供),制作桡骨临界骨缺损模型(桡骨中段截骨3 cm),其中4只骨缺损处植入载缓释微球的三维生物支架与自体松质骨(实验组1),另4只仅植入自体松质骨(对照组1);另取8只成年犬,制作桡骨超临界骨缺损模型(桡骨中段截骨5cm),其中4只骨缺损处植入载缓释微球的三维生物支架与自体松质骨(实验组2),另4只仅植入自体松质骨(对照组2)。术后4,8,12周进行X射线检查;术后12周,取出缺损区桡骨,行组织病理学观察。实验已通过遵义医科大学伦理委员会审查,批准号:伦审(2016)2-071号。结果与结论:①X射线:术后12周,实验组1缺损处髓腔基本再通,骨皮质形成,对照组1缺损处基本完成骨连接,但未完成髓腔再通与皮质骨塑形;实验组2髓腔基本再通,骨皮质塑形与骨长入基本完成,对照组2未完成骨连接,无皮质骨塑形;②组织学观察:实验组1骨缺损处已完成骨连接,可观察到骨皮质形成及完全髓腔再通,对照组1可见骨组织形成,其间可见大量纤维组织,无皮质骨,髓腔未完成再通;实验组2骨缺损处已有皮质骨形成及完成髓腔再通,对照组2可见少量骨组织,无皮质骨形成及髓腔再通;并且实验组1骨修复效果优于实验组2;③结果表明载重组人骨形态发生蛋白2缓释微球的丝素蛋白/壳聚糖/纳米羟基磷灰石三维生物支架,能有效修复一定范围内的犬桡骨临界及超临界骨缺损。

去细胞组织水凝胶化的研究进展

通过去细胞组织可制备细胞外基质Extracellular matrix,ECM生物支架,在临床应用中大量应用于构建结构化和功能化的组织模型,ECM材料易溶解,可与水凝胶材料很好的融合,不论是体外作为培养支架材料,还是体内作为注射填充材料,都避免了形状无规则的缺点。然而ECM水凝胶促进细胞分化、增殖的机制尚未完全研究清楚,但是与天然材料保留了许多结构和生物学信号有很大关系。该文献初步阐述了ECM水凝胶的制备、部分物理、化学和生物学特性等研究进展,以及在心、脑两种器官的体内临床应用,初步讨论了ECM水凝胶的某些研究和应用前景。