Optimizing Attachment of Human Mesenchymal Stem Cells on Poly(ε-caprolactone) Electrospun Yarns

Research into biomaterials and tissue engineering often includes cell-based in vitro investigations, which require initial knowledge of the starting cell number. While researchers commonly reference their seeding density this does not necessarily indicate the actual number of cells that have adhered to the material in question. This is particularly the case for materials, or scaffolds, that do not cover the base of standard cell culture well plates. This study investigates the initial attachment of human mesenchymal stem cells seeded at a known number onto electrospun poly(ε-caprolactone) yarn after 4 hr in culture. Electrospun yarns were held within several different set-ups, including bioreactor vessels rotating at 9 rpm, cell culture inserts positioned in low binding well plates and polytetrafluoroethylene (PTFE) troughs placed within petri dishes. The latter two were subjected to either static conditions or positioned on a shaker plate (30 rpm). After 4 hr incubation at 37 ^oC, 5% CO₂, the location of seeded cells was determined by cell DNA assay. Scaffolds were removed from their containers and placed in lysis buffer. The media fraction was similarly removed and centrifuged – the supernatant discarded and pellet broken up with lysis buffer. Lysis buffer was added to each receptacle, or well, and scraped to free any cells that may be present. The cell DNA assay determined the percentage of cells present within the scaffold, media and well fractions. Cell attachment was low for all experimental set-ups, with greatest attachment (30%) for yarns held within cell culture inserts and subjected to shaking motion. This study raises awareness to the actual number of cells attaching to scaffolds irrespective of the stated cell seeding density.     

Smart scaffolds in bone tissue engineering: A systematic review of literature

AIM: To improve osteogenic differentiation and attachment of cells.METHODS: An electronic search was conducted inPub Med from January 2004 to December 2013. Studies which performed smart modifications on conventional bone scaffold materials were included. Scaffolds with controlled release or encapsulation of bioactive molecules were not included. Experiments which did not investigate response of cells toward the scaffold(cell attachment, proliferation or osteoblastic differentiation) were excluded. RESULTS: Among 1458 studies, 38 met the inclusion and exclusion criteria. The main scaffold varied extensively among the included studies. Smart modifications included addition of growth factors(group Ⅰ-11 studies), extracellular matrix-like molecules(group Ⅱ-13 studies) and nanoparticles(nano-HA)(group Ⅲ-17 studies). In all groups, surface coating was the most commonly applied approach for smart modification of scaffolds. In group I, bone morphogenetic proteins were mainly used as growth factor stabilized on polycaprolactone(PCL). In group Ⅱ, collagen 1 in combination with PCL, hydroxyapatite(HA) and tricalcium phosphate were the most frequent scaffolds used. In the third group, nano-HA with PCL and chitosan were used the most. As variable methods were used, a thorough and comprehensible compare between the results and approaches was unattainable.CONCLUSION: Regarding the variability in methodology of these in vitro studies it was demonstrated that smart modification of scaffolds can improve tissue properties.     

Mechanisms of iron–sulfur cluster assembly: the SUF machinery

Biosynthesis of iron-sulfur clusters is a cellular process which depends on complex protein machineries. Escherichia coli contains two such biosynthetic systems, ISC and SUF. In this review article we specifically make a presentation of the various Suf proteins and discuss the molecular mechanisms by which these proteins work together to assemble Fe and S atoms within a scaffold and to transfer the resulting cluster to target proteins. An erratum to this article can be found at     

干细胞3D支架的研究进展

干细胞是一类具有无限增殖潜能,可自我更新的细胞,不同的培养或诱导条件下能够分化成为不同的细胞类型。目前,随着医学治疗手段及干细胞研究的不断发展,发现干细胞可应用于很多疾病的治疗,干细胞临床应用日益广泛,逐渐成为科研工作者研究的热点。然而干细胞移植进入生物体后繁殖效率很低,无法达到需求的量,因此如何对干细胞在生物体外进行大量培养扩增、在生物体内如何更稳定地增殖分化成为迫切需要解决的难题。当前干细胞最主要的培养方法仍是2D培养,2D培养无法模拟体内的3D微环境,繁殖效率较低,正是由于2D培养局限性太多,促使国内外学者对3D培养技术和3D支架材料进行深入探索,并取得了大量成果。对3D培养技术在干细胞中的发展与应用进行概述和展望。     

A Mechanobiology-based Algorithm to Optimize the Microstructure Geometry of Bone Tissue Scaffolds

Complexity of scaffold geometries and biological mechanisms involved in the bone generation process make the design of scaffolds a quite challenging task. The most common approaches utilized in bone tissue engineering require costly protocols and time-consuming experiments. In this study we present an algorithm that, combining parametric finite element models of scaffolds with numerical optimization methods and a computational mechano-regulation model, is able to predict the optimal scaffold microstructure. The scaffold geometrical parameters are perturbed until the best geometry that allows the largest amounts of bone to be generated, is reached. We study the effects of the following factors: (1) the shape of the pores; (2) their spatial distribution; (3) the number of pores per unit area. The optimal dimensions of the pores have been determined for different values of scaffold Young''s modulus and compression loading acting on the scaffold upper surface.Pores with rectangular section were predicted to lead to the formation of larger amounts of bone compared to square section pores; similarly, elliptic pores were predicted to allow the generation of greater amounts of bone compared to circular pores. The number of pores per unit area appears to have rather negligible effects on the bone regeneration process. Finally, the algorithm predicts that for increasing loads, increasing values of the scaffold Young''s modulus are preferable.The results shown in the article represent a proof-of-principle demonstration of the possibility to optimize the scaffold microstructure geometry based on mechanobiological criteria.     

Discovery of a series of benzopyrimidodiazepinone TNK2 inhibitors via scaffold morphing

The protein kinase TNK2 (ACK1) is an emerging drug target for a variety of indications, in particular for cancer where it plays a key role transmitting cell survival, growth and proliferative signals via modification of multiple downstream effectors by unique tyrosine phosphorylation events. Scaffold morphing based on our previous TNK2 inhibitor XMD8-87 identified urea 17 from which we developed the potent and selective compound 32. A co-crystal structure was obtained showing 32 interacting primarily with the main chain atoms of an alanine residue of the hinge region. Additional H-bonds exist between the urea NHs and the Thr205 and Asp270 residues.     

Minced Tissue in Compressed Collagen: A Cell-containing Biotransplant for Single-staged Reconstructive Repair

Conventional techniques for cell expansion and transplantation of autologous cells for tissue engineering purposes can take place in specially equipped human cell culture facilities. These methods include isolation of cells in single cell suspension and several laborious and time-consuming events before transplantation back to the patient. Previous studies suggest that the body itself could be used as a bioreactor for cell expansion and regeneration of tissue in order to minimize ex vivo manipulations of tissues and cells before transplanting to the patient. The aim of this study was to demonstrate a method for tissue harvesting, isolation of continuous epithelium, mincing of the epithelium into small pieces and incorporating them into a three-layered biomaterial. The three-layered biomaterial then served as a delivery vehicle, to allow surgical handling, exchange of nutrition across the transplant, and a controlled degradation. The biomaterial consisted of two outer layers of collagen and a core of a mechanically stable and slowly degradable polymer. The minced epithelium was incorporated into one of the collagen layers before transplantation. By mincing the epithelial tissue into small pieces, the pieces could be spread and thereby the propagation of cells was stimulated. After the initial take of the transplants, cell expansion and reorganization would take place and extracellular matrix mature to allow ingrowth of capillaries and nerves and further maturation of the extracellular matrix. The technique minimizes ex vivo manipulations and allow cell harvesting, preparation of autograft, and transplantation to the patient as a simple one-stage intervention. In the future, tissue expansion could be initiated around a 3D mold inside the body itself, according to the specific needs of the patient. Additionally, the technique could be performed in an ordinary surgical setting without the need for sophisticated cell culturing facilities.