Effects of extracellular matrix analogues on primary human fibroblast behavior

In vitro cell culture is a vital research tool for cell biology, pharmacology, toxicology, protein production, systems biology and drug discovery. Traditional culturing methods on plastic surfaces do not accurately represent the in vivo environment, and a paradigm shift from two-dimensional to three-dimensional (3-D) experimental techniques is underway. To enable this change, a variety of natural, synthetic and semi-synthetic extracellular matrix (ECM) equivalents have been developed to provide an appropriate cellular microenvironment. We describe herein an investigation of the properties of four commercially available ECM equivalents on the growth and proliferation of primary human tracheal scar fibroblast behavior, both in 3-D and pseudo-3-D conditions. We also compare subcutaneous tissue growth of 3-D encapsulated fibroblasts in vivo in two of these materials, Matrigel and Extracel. The latter shows increased cell proliferation and remodeling of the ECM equivalent. The results provide researchers with a rational basis for selection of a given ECM equivalent based on its biological performance in vitro and in vivo, as well as the practicality of the experimental protocols. Biomaterials that use a customizable glycosaminoglycan-based hydrogel appear to offer the most convenient and flexible system for conducting in vitro research that accurately translates to in vivo physiology needed for tissue engineering.     

Cell-based resorption assays for bone graft substitutes

The clinical utilization of resorbable bone substitutes has been growing rapidly during the last decade, creating a rising demand for new resorbable biomaterials. An ideal resorbable bone substitute should not only function as a load-bearing material but also integrate into the local bone remodeling process. This means that these bone substitutes need to undergo controlled resorption and then be replaced by newly formed bone structures. Thus the assessment of resorbability is an important first step in predicting the in vivo clinical function of bone substitute biomaterials. Compared with in vivo assays, cell-based assays are relatively easy, reproducible, inexpensive and do not involve the suffering of animals. Moreover, the discovery of RANKL and M-CSF for osteoclastic differentiation has made the differentiation and cultivation of human osteoclasts possible and, as a result, human cell-based bone substitute resorption assays have been developed. In addition, the evolution of microscopy technology allows advanced analyses of the resorption pits on biomaterials. The aim of the current review is to give a concise update on in vitro cell-based resorption assays for analyzing bone substitute resorption. For this purpose models using different cells from different species are compared. Several popular two-dimensional and three-dimensional optical methods used for resorption assays are described. The limitations and advantages of the current ISO degradation assay in comparison with cell-based assays are discussed.     

Nitric oxide producing coating mimicking endothelium function for multifunctional vascular stents

The continuous release of nitric oxide (NO) by the native endothelium of blood vessels plays a substantial role in the cardiovascular physiology, as it influences important pathways of cardiovascular homeostasis, inhibits vascular smooth muscle cell (VSMC) proliferation, inhibits platelet activation and aggregation, and prevents atherosclerosis. In this study, a NO-catalytic bioactive coating that mimics this endothelium functionality was presented as a hemocompatible coating with potential to improve the biocompatibility of vascular stents. The NO-catalytic bioactive coating was obtained by covalent conjugation of 3,3-diselenodipropionic acid (SeDPA) with glutathione peroxidase (GPx)-like catalytic activity to generate NO from S-nitrosothiols (RSNOs) via specific catalytic reaction. The SeDPA was immobilized to an amine bearing plasma polymerized allylamine (PPAam) surface (SeDPA-PPAam). It showed long-term and continuous ability to catalytically decompose endogenous RSNO and generate NO. The generated NO remarkably increased the cGMP synthesis both in platelets and human umbilical artery smooth muscle cells (HUASMCs). The surface exhibited a remarkable suppression of collagen-induced platelet activation and aggregation. It suppressed the adhesion, proliferation and migration of HUASMCs. Additionally, it was found that the NO catalytic surface significantly enhanced human umbilical vein endothelial cell (HUVEC) adhesion, proliferation and migration. The in vivo results indicated that the NO catalytic surface created a favorable microenvironment of competitive growth of HUVECs over HUASMCs for promoting re-endothelialization and reducing restenosis of stents in vivo.     

A concise total synthesis of secoisolariciresinol

A concise total synthesis of (R,R)-secoisolariciresinol was achieved in four steps, featuring a biomimetic β-β’ radical coupling of two dihydroferulic acid derivatives. The conversion of secoisolariciresinol to other related lignans was ongoing.     

Generation of nanovesicles with sliced cellular membrane fragments for exogenous material delivery

We propose a microfluidic system that generates nanovesicles (NVs) by slicing living cell membrane with microfabricated 500 nm-thick silicon nitride (Si_xN_y) blades. Living cells were sliced by the blades while flowing through microchannels lined with the blades. Plasma membrane fragments sliced from the cells self-assembled into spherical NVs of ∼100–300 nm in diameter. During self-assembly, the plasma membrane fragments enveloped exogenous materials (here, polystyrene latex beads) from the buffer solution. About 30% of beads were encapsulated in NVs, and the generated NVs delivered the encapsulated beads across the plasma membrane of recipient cells, but bare beads could not penetrate the plasma membrane of recipient cells. This result implicates that the NVs generated using the method in this study can encapsulate and deliver exogenous materials to recipient cells, whereas exosomes secreted by cells can deliver only endogenous cellular materials.     

In vitro myogenesis induced by human recombinant elastin-like proteins

Mammalian adult skeletal muscle has a limited ability to regenerate after injury, usage or trauma. A promising strategy for successful regenerative technology is the engineering of bio interfaces that mimic the characteristics of the extracellular matrix. Human elastin-like polypeptides (HELPs) have been synthesized as biomimetic materials that maintain some peculiar properties of the native protein. We developed a novel Human Elastin Like Polypeptide obtained by fusing the elastin-like backbone to a domain present in the α2 chain of type IV collagen, containing two RGD motives. We employed this peptide as adhesion substrate for C2C12 myoblasts and compared its effects to those induced by two other polypeptides of the HELP series. Myoblast adhered to all HELPs coatings, where they assumed morphology and cytoarchitecture that depended on the polypeptide structure. Adhesion to HELPs stimulated at a different extent cell proliferation and differentiation, the expression of Myosin Heavy Chain and the fusion of aligned fibers into multinucleated myotubes. Adhesion substrates significantly altered myotubes stiffness, measured by Atomic Force Microscopy, and differently affected the cells Ca²⁺ handling capacity and the maturation of excitation-contraction coupling machinery, evaluated by Ca²⁺ imaging. Overall, our findings indicate that the properties of HELP biopolymers can be exploited for dissecting the molecular connections underlying myogenic differentiation and for designing novel substrates for skeletal muscle regeneration.     

Facile synthesis of PEI-based crystalline templated mesoporous silica with molecular chirality for improved oral delivery of the poorly water-soluble drug

The aim of this study was to build up a novel chiral mesoporous silica called PEIs@TA-CMS through a facile biomimetic strategy and to explore its potential to serve as a drug carrier for improving the delivery efficiency of poorly water-soluble drug. PEIs@TA-CMS was synthesized by using a chiral crystalline complex associated of tartaric acid and polyethyleneimine (PEIs) as templates, scaffolds and catalysts. The structural features including morphology, size, pore structure and texture properties were systematacially studied. The results showed that PEIs@TA-CMS was monodispersed spherical nanoparticles in a uniformed diameter of 120–130 nm with well-developed pore structure (S_BET: 1009.94 m²/g, pore size <2.21 nm). Then PEIs@TA-CMS was employed as nimodipine (NMP) carrier and compared with the drug carry ability of MCM41. After drug loading, NMP was effectively transformed from the crystalline state to an amorphous state due to the space confinement in mesopores. As expected, PEIs@TA-CMS had superiority in both drug loading and drug release compared to MCM41. It could incorporate NMP with high efficiency, and the dissolution-promoting effect of PEIs@TA-CMS was more obvious because of the unique interconnected curved pore channels. Meanwhile, PEIs@TA-CMS could significantly improve the oral adsorption of NMP to a satisfactory level, which showed approximately 3.26-fold higher in bioavailability, and could effectively prolong the survival time of mice on cerebral anoxia from 10.98 to 17.33 min.     

矿化蚕丝基骨材料的生物相容性及体内降解的观察

目的探讨仿生制备的矿化蚕丝基骨材料在大鼠体内的生物相容性及体内降解性。方法成年雄性SD大鼠16只,体重230-300克。在背部肌肉内和腹腔内植入矿化蚕丝基骨材料,观察时间阶段为2周,4周,6周,8周。结果分别在术后2周,4周,6周,8周,行大体观察,细胞毒性研究,组织学切片观察材料的情况,植入2周后出现炎症反应明显,材料框架基本存在,术后8周大鼠体内植入的矿化蚕丝基骨材料没有明显的排异反应,而且大部分被吸收。结论矿化蚕丝基骨材料具有良好的生物相容性和适宜的生物降解性能,是比较理想的骨缺损的替代材料。     

The critical barrier to progress in dentine bonding with the etch-and-rinse technique

Objectives

The lack of durability in resin-dentine bonds led to the use of chlorhexidine as MMP-inhibitor to prevent the degradation of hybrid layers. Biomimetic remineralisation is a concept-proven approach in preventing the degradation of resin-dentine bonds. The purpose of this study is to examine the integrity of aged resin-dentine interfaces created with a nanofiller-containing etch-and-rinse adhesive after the application of these two approaches.

Methods

The more established MMP-inhibition approach was examined using a parallel in vivo and in vitro ageing design to facilitate comparison with the biomimetic remineralisation approach using an in vitro ageing design. Specimens bonded without chlorhexidine exhibited extensive degradation of the hybrid layer after 12 months of in vivo ageing.

Results

Dissolution of nanofillers could be seen within a water-rich zone within the adhesive layer. Although specimens bonded with chlorhexidine exhibited intact hybrid layers, water-rich regions remained in those hybrid layers and degradation of nanofillers occurred within the adhesive layer. Specimens subjected to in vitro biomimetic remineralisation followed by in vitro ageing demonstrated intrafibrillar collagen remineralisation within hybrid layers and deposition of mineral nanocrystals in nanovoids within the adhesive.

Conclusions

The impact was realized by understanding the lack of an inherent mechanism to remove water from resin-dentine interfaces as the critical barrier to progress in bonding with the etch-and-rinse technique. The experimental biomimetic remineralisation strategy offers a creative solution for incorporating a progressive hydration mechanism to achieve this goal, which warrants its translation into a clinically applicable technique.