应用原子力显微镜研究淫羊霍苷诱导人脐带间充质干细胞成骨分化

利用碱性磷酸酶(ALP)染色和钙结节(Vonkossa)染色的方法对诱导21 d的淫羊霍苷诱导人脐带间充质干细胞进行鉴定;应用原子力显微镜(AFM)观察淫羊霍苷的形貌和人脐带间充质干细胞诱导0、5、10、15、21 d后的细胞形貌。结果表明,经成骨诱导分化21 d后,ALP染色呈强阳性,Vonkossa染色可见明显钙结节。AFM分析表明,淫羊霍苷在盖玻片上呈分散状分布,在细胞表面上聚集并呈微米域分布。实验发现,由于吸附在细胞表面时,被细胞膜分子包裹,更有利于在细胞表面的吸附,进入细胞内部,细胞表面的淫羊霍苷颗粒较在盖玻片上时增大,由淫羊霍苷颗粒进入细胞后在细胞表面留下一些小孔,可知其通过进入细胞内部诱导成骨分化。分化后,细胞表面有小突触,是由成骨分化后细胞内形成钙结节造成。     

胎儿脐带血铅水平与钙、锌、铁的关系研究

目的研究胎儿脐带血中的铅与钙、锌、铁的含量以及两两之间的相关性。方法采用原子吸收光谱法对398例脐带血铅和钙、锌、铁的含量进行测定,并对它们进行相关性分析。结果 398例脐带血铅和钙、锌、铁的质量浓度分别为铅(37.96±17.78)μg/L、钙(1.51±0.29)mmol/L、锌(30.01±11.14)μmol/L、铁(8.63±1.39)mmol/L。线性回归分析显示:铅与钙(r=-0.563,P0.01)、铅与锌(r=-0.424,P0.01)有相关性,铅与铁无显著相关(r=0.018,P0.05)。结论随着胎儿体内血铅水平的上升,二价元素钙、锌含量有下降趋势,即中毒元素铅会干扰胎儿体内的必需二价元素的代谢。     

Investigation of the Effective Action Distance Between Hematopoietic Stem/Progenitor Cells and Human Adipose-Derived Stem Cells During Their In Vitro Co-culture

The in vitro suitable action distance between umbilical cord blood-derived hematopoietic stem/progenitor cells and its feeder cell, human adipose-derived stem cells, during their co-culture, was investigated through a novel transwell co-culture protocol, in which the distance between the two culture chambers where each cell type is growing can be adjusted from 10 to 450 μm. The total cell number was determined with a hemacytometer, and the cell morphology was observed under an inverted microscope each day. After 7 days of co-culture, the fold-expansion, surface antigen expression of CD34(+) and CFU-GM assay of the hematopoietic mononuclear cells (MNCs) were analyzed. The results showed that there was an optimal communication distance at around 350 μm between both types of stem cells during their in vitro co-culture. By using this distance, the UCB-MNCs and CD34(+) cells were expanded by 15.1?±?0.2 and 5.0?±?0.1-fold, respectively. It can therefore be concluded that the optimal action distance between stem cells and their supportive cells, when cultured together for 7 days, is of around 350 μm.     

用于人类脂肪干细胞的纯化和体外长期培养的聚合物涂层

采用紫外固化法制备了基于丙烯酸酯类水凝胶的聚合物涂层(PC),并用X射线光电子能谱(XPS)、水接触角(WCA)和原子力显微镜(AFM)分别对PC进行了化学组成和表面性能的表征.在PC表面进行了人类脂肪干细胞(h ASC)的体外长期培养扩增,得到的第3代细胞的生物学表征结果表明,干细胞在PC表面能正常黏附生长,流式细胞仪检测发现干细胞对特征标记物CD49d,CD73,CD105的阳性显性比例较高,对HLA-DR和CD31几乎不显性,说明扩增的干细胞具有h ASC特征.对PC上扩增的干细胞进行诱导分化,并用油红O、茜素红和阿利新蓝分别进行染色分析,结果表明,该干细胞保留了h ASC的多能特性:能分化为成脂、成骨和成软骨细胞.含有单体甲基丙烯酰氧乙基三甲基氯化铵(DMC)、甲基丙烯酸环己酯(CHMA)和甲基丙烯酸-2-(二乙氨基)乙酯(DEAEMA)的PC2(质量比为3∶1∶2)在用于h ASC体外长期培养时,比其它PC和TCP更有利于细胞的黏附和增殖,纯化细胞,保持其多能性.实时荧光定量PCR(RT-q PCR)的分析表明PC2上得到的细胞更容易向成骨和成软骨细胞分化.     

Anchoring Supramolecular Polymers to Human Red Blood Cells by Combining Dynamic Covalent and Non-Covalent Chemistries

Understanding cell/material interactions is essential to design functional cell-responsive materials. While the scientific literature abounds with formulations of biomimetic materials, only a fraction of them focused on mechanisms of the molecular interactions between cells and material. To provide new knowledge on the strategies for materials/cell recognition and binding, supramolecular benzene-1,3,5-tricarboxamide copolymers bearing benzoxaborole moieties are anchored on the surface of human erythrocytes via benzoxaborole/sialic-acid binding. This interaction based on both dynamic covalent and non-covalent chemistries is visualized in real time by means of total internal reflection fluorescence microscopy. Exploiting this imaging method, we observe that the functional copolymers specifically interact with the cell surface. An optimal fiber affinity towards the cells as a function of benzoxaborole concentration demonstrates the crucial role of multivalency in these cell/material interactions.     

Aligned Poly(ε‐caprolactone) Nanofibers Guide the Orientation and Migration of Human Pluripotent Stem Cell‐Derived Neurons,Astrocytes, and Oligodendrocyte Precursor Cells In Vitro

Stem cell transplantations for spinal cord injury (SCI) have been studied extensively for the past decade in order to replace the damaged tissue with human pluripotent stem cell (hPSC)‐derived neural cells. Transplanted cells may, however, benefit from supporting and guiding structures or scaffolds in order to remain viable and integrate into the host tissue. Biomaterials can be used as supporting scaffolds, as they mimic the characteristics of the natural cellular environment. In this study, hPSC‐derived neurons, astrocytes, and oligodendrocyte precursor cells (OPCs) are cultured on aligned poly(ε‐caprolactone) nanofiber platforms, which guide cell orientation to resemble that of spinal cord in vivo. All cell types are shown to efficiently spread over the nanofiber platform and orient according to the fiber alignment. Human neurons and astrocytes require extracellular matrix molecule coating for the nanofibers, but OPCs grow on nanofibers without additional treatment. Furthermore, the nanofiber platform is combined with a 3D hydrogel scaffold with controlled thickness, and nanofiber‐mediated orientation of hPSC‐derived neurons is also demonstrated in a 3D environment. In this work, clinically relevant materials and substrates for nanofibers, fiber coatings, and hydrogel scaffolds are used and combined with cells suitable for developing functional cell grafts for SCI repair.

     

Anchoring Supramolecular Polymers to Human Red Blood Cells by Combining Dynamic Covalent and Non‐Covalent Chemistries

Understanding cell/material interactions is essential to design functional cell‐responsive materials. While the scientific literature abounds with formulations of biomimetic materials, only a fraction of them focused on mechanisms of the molecular interactions between cells and material. To provide new knowledge on the strategies for materials/cell recognition and binding, supramolecular benzene‐1,3,5‐tricarboxamide copolymers bearing benzoxaborole moieties are anchored on the surface of human erythrocytes via benzoxaborole/sialic‐acid binding. This interaction based on both dynamic covalent and non‐covalent chemistries is visualized in real time by means of total internal reflection fluorescence microscopy. Exploiting this imaging method, we observe that the functional copolymers specifically interact with the cell surface. An optimal fiber affinity towards the cells as a function of benzoxaborole concentration demonstrates the crucial role of multivalency in these cell/material interactions.