Biomolecular gradients in cell culture systems

Biomolecule gradients have been shown to play roles in a wide range of biological processes including development, inflammation, wound healing, and cancer metastasis. Elucidation of these phenomena requires the ability to expose cells to biomolecule gradients that are quantifiable, controllable, and mimic those that are present in vivo. Here we review the major biological phenomena in which biomolecule gradients are employed, traditional in vitro gradient-generating methods developed over the past 50 years, and new microfluidic devices for generating gradients. Microfluidic gradient generators offer greater levels of precision, quantitation, and spatiotemporal gradient control than traditional methods, and may greatly enhance our understanding of many biological phenomena. For each method, we outline the salient features, capabilities, and applications.     

Resorbable polymer electrospun nanofibers: History,shapes and application for tissue engineering

Resorbable polymer electrospun nanofiber-based materials/devices have high surface-to-volume ratio and often have a porous structure with excellent pore interconnectivity,which are suitable for growth and development of different types of cells.Due to the huge advantages of both resorbable polymers and electrospun nano fibers,re sorbable polymer electrospun nanofibers(RPENs)have been widely applied in the field of tissue engineering.In this paper,we will mainly introduce RPENs for tissue engineering.Firstly,the electrospinning technique and electrospun nanofiber architectures are briefly introduced.Secondly,the application of RPENs in the field of tissue engineering is mainly reviewed.Finally,the advantages and disadvantages of RPENs for tissue engineering are discussed.This review will provide a comprehensive guide to apply resorbable polymer electrospun nanofibers for tissue engineering.     

A review on electrospun nanofibers for multiple biomedical applications

Electrospinning is a well-known technique since 1544 to fabricate nanofibers using different materials like polymers, metals oxides, proteins, and many more. In recent years, electrospinning has become the most popular technique for manufacturing nanofibers due to its ease of use and economic viability. Nanofibers have remarkable properties like high surface-to-volume ratio, variable pore size distribution (10–100 nm), high porosity, low density, and are suitable for surface functionalization. Therefore, electrospun nanofibers have been utilized for numerous applications in the pharmaceutical and biomedical field like tissue engineering, scaffolds, grafts, drug delivery, and so on. In this review article, we will be focusing on the versatility, current scenario, and future endeavors of electrospun nanofibers for various biomedical applications. This review discusses the properties of nanofibers, the background of the electrospinning technique, and its emergence in chronological order. It also covers the various types of electrospinning methods and their mechanism, further elaborating the factors affecting the properties of nanofibers, and applications in tissue engineering, drug delivery, nanofibers as biosensor, skin cancer treatment, and magnetic nanofibers.     

生物高分子静电纺丝研究进展

静电纺丝(电纺)技术是一种制备直径为数十纳米到数微米的纳米纤维的有效方法。由于生物高分子具有良好的生物相容性,近年来国内外对生物高分子的电纺制备进行了大量研究。这种生物高分子纳米纤维在组织工程支架、组织修复等方面有独特的优势。本文对生物高分子——多糖、蛋白质、核酸(DNA)的静电纺丝研究进行了总结。     

Generation of dynamic temporal and spatial concentration gradients using microfluidic devices

This paper describes a microfluidic approach to generate dynamic temporal and spatial concentration gradients using a single microfluidic device. Compared to a previously described method that produced a single fixed gradient shape for each device, this approach combines a simple "mixer module" with gradient generating network to control and manipulate a number of different gradient shapes. The gradient profile is determined by the configuration of fluidic inputs as well as the design of microchannel network. By controlling the relative flow rates of the fluidic inputs using separate syringe pumps, the resulting composition of the inlets that feed the gradient generator can be dynamically controlled to generate temporal and spatial gradients. To demonstrate the concept and illustrate this approach, examples of devices that generate (1) temporal gradients of homogeneous concentrations, (2) linear gradients with dynamically controlled slope, baseline, and direction, and (3) nonlinear gradients with controlled nonlinearity are shown and their limitations are described.     

Electrospinning nanofibers of microbial polyhydroxyalkanoates for applications in medical tissue engineering

Differently to most chemically synthesized medical materials, polyhydroxyalkanoates (PHAs) are intracellular carbon and energy storage granules, which is a family of natural bio-polymers synthesized by microorganism's materials. Due to excellent biocompatibility reasonable biodegradability and versatile material difference, PHAs are well medical biomaterials candidates for applications in tissue engineering and drug delivery, including commercial PHB, PHBV, PHBHHx, PHBVHHx, P34HB and few uncommercial PHAs. Electrospinning nanofibers with the size of 10–10,000 nm can improve the mechanical properties and decrease the crystallinity of PHA, meanwhile simulate the structure and function of native extracellular matrix of cells. Hence, PHAs electrospinning nanofibers as engineered scaffolds have been widely used for tissue engineering scaffolds in cardiovascular, vascular, nerve, bone, cartilage and skin; also, as carriers for application in drug delivery system. In this review, we highlight the extraction and properties of medical PHAs from natural or engineered microorganism, and microstructure, current manufacturing techniques and medical applications of electrospinning nanofibers of PHAs. Moreover, the current challenges and prospects of PHAs electrospinning nanofibers are discussed rationally, providing an insight into developing vibrant fields of PHAs electrospinning nanofibers-based biomedicine.     

Mammalian cell viability in electrospun composite nanofiber structures

Incorporation of mammalian cells into nanofibers (cell electrospinning) and multilayered cell-nanofiber structures (cell layering) via electrospinning are promising techniques for tissue engineering applications. We investigate the viability of 3T3-L1 mouse fibroblasts after incorporation into poly(vinyl alcohol) nanofibers and multilayering with poly(caprolactone) nanofibers and analyze the possible factors that affect cell viability. We observe that cells do not survive cell electrospinning but survive cell layering. Assessing the factors involved in cell electrospinning, we find that dehydration and fiber stretching are the main causes of cell death. In cell layering, the choice of solvent is critical, as residual solvent in the electrospun fibers could be detrimental to the cells.     

Controlled release of dual drugs from emulsion electrospun nanofibrous mats

The purpose of this work is to develop a novel type of tissue engineering scaffold or drugs delivery carrier with the capability of encapsulation and controlled release drugs. In this study, Rhodamine B and Bovine Serum Albumin (BSA) were successfully incorporated into nanofibers by means of emulsion electrospinning. The morphology of composite nanofibers was studied by Scanning Electron Microscopy (SEM). The composite nanofibrous mats made from emulsion electrospinning were characterized by water contact angle measurement and X-ray diffraction. In vitro dual drugs release behaviors from composite nanofibrous mats were investigated. The results indicated that the incorporated drug and/or proteins in composite fibrous mats made from electrospinning could be control released by adjusting the processes of emulsions preparation.     

Silk Fibroin Combined with Electrospinning as a Promising Strategy for Tissue Regeneration

The development of tissue engineering scaffolds is of great significance for the repair and regeneration of damaged tissues and organs. Silk fibroin (SF) is a natural protein polymer with good biocompatibility, biodegradability, excellent physical and mechanical properties and processability, making it an ideal universal tissue engineering scaffold material. Nanofibers prepared by electrospinning have attracted extensive attention in the field of tissue engineering due to their excellent mechanical properties, high specific surface area, and similar morphology as to extracellular matrix (ECM). The combination of silk fibroin and electrospinning is a promising strategy for the preparation of tissue engineering scaffolds. In this review, the research progress of electrospun silk fibroin nanofibers in the regeneration of skin, vascular, bone, neural, tendons, cardiac, periodontal, ocular and other tissues is discussed in detail.     

Alkali‐Mediated Miscibility of Gelatin/Polycaprolactone for Electrospinning Homogeneous Composite Nanofibers for Tissue Scaffolding

Electrospun natural‐synthetic composite nanofibers, which possess favorable biological and mechanical properties, have gained widespread attention in tissue engineering. However, the development of biomimetic nanofibers of hybrids remains a huge challenge due to phase separation of the polymer blends. Here, aqueous sodium hydroxide (NaOH) solution is proposed to modulate the miscibility of a representative natural‐synthetic hybrid of gelatin (GT) and polycaprolactone (PCL) for electrospinning homogeneous composite nanofibers. Alkali‐doped GT/PCL solutions and nanofibers examined at macroscopic, microscopic, and internal molecular levels demonstrate appropriate miscibility of GT and PCL after introducing the alkali dopant. Particularly, homogeneous GT/PCL nanofibers with smooth surface and uniform diameter are obtained when aqueous NaOH solution with a concentration of 10 m is used. The fibers become more hydrophilic and possess improved mechanical properties both in dry and wet conditions. Moreover, biocompatibility experiments show that stem cells adhere to and proliferate better on the alkali‐modified nanofibers than the untreated one. This study provides a facile and effective approach to solve the phase separation issue of the synthetic‐natural hybrid GT/PCL and establishes a correlation of compositionally and morphologically homogeneous composite nanofibers with respect to cell responses.     

聚合物的静电纺丝

静电纺丝法是聚合物溶液或熔体在静电作用下进行喷射拉伸而获得纳米级纤维的纺丝方法.由纳米纤维制得的无纺布,具有孔隙率高、比表面积大、纤维精细程度与均一性高、长径比大等优点,从而赋予了静电纺丝纤维广泛的应用前景,它已在国内外引起了广泛的关注.本文介绍了静电纺丝的装置、基本原理及静电纺丝制备纳米纤维的研究进展,同时也叙述了其在各个领域的应用,最后展望了静电纺丝制备纳米纤维的发展方向及前景.     

In-Vitro Cytotoxicity Study: Cell Viability and Cell Morphology of Carbon Nanofibrous Scaffold/Hydroxyapatite Nanocomposites

Electrospun carbon nanofibers (CNFs), which were modified with hydroxyapatite, were fabricated to be used as a substrate for bone cell proliferation. The CNFs were derived from electrospun polyacrylonitrile (PAN) nanofibers after two steps of heat treatment: stabilization and carbonization. Carbon nanofibrous (CNF)/hydroxyapatite (HA) nanocomposites were prepared by two different methods; one of them being modification during electrospinning (CNF-8HA) and the second method being hydrothermal modification after carbonization (CNF-8HA; hydrothermally) to be used as a platform for bone tissue engineering. The biological investigations were performed using in-vitro cell counting, WST cell viability and cell morphology after three and seven days. L929 mouse fibroblasts were found to be more viable on the hydrothermally-modified CNF scaffolds than on the unmodified CNF scaffolds. The biological characterizations of the synthesized CNF/HA nanofibrous composites indicated higher capability of bone regeneration.     

Generation of linear and non-linear concentration gradients along microfluidic channel by microtunnel controlled stepwise addition of sample solution

The ability to generate stable chemical gradients in microfluidics has important applications, since such gradients are useful in both chemical and biological studies. Growing evidence reveals that many cellular responses are specific to non-linear spatial gradients, hence a need to control complex concentration gradient profiles with and within microfluidics. In this paper, we present a structure-based approach to generate linear and non-linear chemical gradients, with profiles controlled by microtunnels fabricated alongside two main channels. Using single-step photolithography, microtunnels and main channels were fabricated at different heights thus having different fluidic resistance. Through these microtunnels, sample solutions were stepwise dispensed into the buffer stream to generate a chemical gradient profile. By varying the lengths of microtunnels that dictated the volume of sample solutions being dispensed, complex gradient profiles were generated. We have successfully demonstrated the formation of linear, convex and concave gradient profiles and a simple mathematical expression was established to approximate the profiles produced in our microfluidic gradient-generators.     

Surface‐treated and multilayered poly(ε‐caprolactone) nanofiber webs exhibiting enhanced hydrophilicity

To improve the hydrophilic properties of poly(ε‐caprolactone) (PCL) nano/microfiber webs for tissue engineering scaffolds, PCL webs of various structures were fabricated by electrospinning with single or double nozzles connected to an auxiliary electrode. Surface‐modified and layered PCL fiber webs were made by including water‐soluble poly(ethylene oxide) (PEO) in the PCL electrospinning solution (single‐nozzle method) or by electrospinning of alternating PCL and PEO solutions using two nozzles (double‐nozzle method), respectively. When the PEO component within the resulting webs was removed by dissolution with distilled water, the remnant PCL webs exhibited two distinct structures. Those made by the single‐nozzle method consisted of nanofibers with high surface roughness, whereas those made by the double‐nozzle method consisted of stacked layers of PCL nanofibers. Both types of structured PCL web showed improved hydrophilicity characteristics compared with those of nanofiber webs generated from a pure PCL solution using a typical electrospinning process. Cell culturing and scanning electron microscopy showed that the interactions between human dermal fibroblasts and the structured PCL scaffolds were very favorable. © 2007 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 45: 2038–2045, 2007     

The role of rheology of polymer solutions in predicting nanofiber formation by electrospinning

Electrospun polymer nanofibers are gaining increasing importance in tissue engineering, wound dressing and drug delivery. Here, we present a thorough rheological study of polymer solutions in the bulk and at the interface to find correlations between those properties and the electrospinnability of the solutions and the morphology of the resultant nanofibers. Our results indicate that blended solutions of chitosan or alginate with poly(ethylene oxide) (PEO) are appropriate for electrospinning when they form conductive, unstructured fluids displaying plasticity, rather than elasticity, in the bulk and at the interface. The interfacial rheological parameters are three orders of magnitude lower than those in the bulk. We demonstrate for the first time that interfacial, rather than bulk, rheological parameters show improved correlation and can be used to predict the success of the electrospinning process. Using the interfacial parameters of samples with homologous compositions, different groups of solutions can be identified that form smooth nanofibers. However, rheological parameters of the bulk and at the interface provide complimentary information. The bulk parameters are determined by polymer concentration and directly affect jet initiation, while the interfacial behaviour determines the continuation of the jet and fibre formation. We propose that interfacial parameters are indispensible tools for the design of electrospinning experiments.     

Generation of stable complex gradients across two-dimensional surfaces and three-dimensional gels

Many chemical and biological processes are dependent on molecular gradients. We describe a new microfluidic approach that can be used to produce spatiotemporal gradients across two-dimensional surfaces and three-dimensional gels under flow-free conditions. Free diffusion between dynamically replenished flow channels acting as a sink and source is utilized to give rise to stable steady-state gradient profiles. The gradient profile is dictated by the engineered design of the device's gradient-generating region. Different designs can yield both linear and non-linear gradients of varying profiles. More complex gradients can be made by juxtaposing different designs within a single gradient-generating region. By fabricating an array of designs along the gradient-generating region, different gradient profiles can be generated simultaneously, allowing for parallel analysis. Additionally, simple methods of localizing gels into microdevices are demonstrated. The device was characterized by experimentally obtained gradient profiles of fluorescent molecules that corroborated closely with a simulated finite element model.     

Poly(glycerol sebacate) nanofiber scaffolds by core/shell electrospinning

The novel biomaterial poly(glycerol sebacate) (PGS) holds great promise for tissue engineering and regenerative medicine. PGS is a rubbery, degradable polymer much like elastin; however, it has been limited to cast structures. This work reports on the formation of PGS nanofibers in random non-woven mats for use as tissue engineering scaffolds by coaxial core/shell electrospinning. PGS nanofibers are an inexpensive and synthetic material that mimics the chemical and mechanical environment provided by elastin fibers. Poly(lactide) was used as the shell material to constrain the PGS during the curing process and was removed before cell seeding. Human microvascular endothelial cells from skin (HDMEC) were used to evaluate the in-vitro cellular compatibility of the PGS nanofiber scaffolds. [Figure: see text].     

Highly porous core-shell polymeric fiber network

Core-shell nanofibers are of great interest in the field of tissue engineering and cell biology. We fabricated porous core-shell fiber networks using an electrospinning system with a water-immersed collector. We hypothesized that the phase separation and solvent evaporation process would enable the control of the pore formation on the core-shell fiber networks. To synthesize porous core-shell fiber networks, we used polycaprolactone (PCL) and gelatin. Quantitative analysis showed that the sizes of gelatin-PCL core-shell nanofibers increased with PCL concentrations. We also observed that the shapes of the pores created on the PCL fiber networks were elongated, whereas the gelatin-PCL core-shell fiber networks had circular pores. The surface areas of porous nanofibers were larger than those of the nonporous nanofibers due to the highly volatile solvent and phase separation process. The porous core-shell fiber network was also used as a matrix to culture various cell types, such as embryonic stem cells, breast cancer cells, and fibroblast cells. Therefore, this porous core-shell polymeric fiber network could be a potentially powerful tool for tissue engineering and biological applications.     

电纺丝法制备聚乳酸/多壁碳纳米管/羟基磷灰石杂化纳米纤维的研究

电纺丝是一种利用聚合物溶液或熔体在强电场中进行喷射纺丝的加工技术,所制得的纤维、直径一般在数十纳米至几微米之间,比传统方法制得的纤维直径小几个数量级,是获得纳米尺寸长纤维的有效方法之一．     

静电纺丝——从无规纳米纤维膜到取向连续长纱

静电纺丝是通过对聚合物溶液或熔体施加外电场制造纳米纤维的有效方法.电纺过程中,在静电力作用下聚合物射流快速鞭动,形成的纳米纤维无规堆砌,得到无纺布状的无规纳米纤维膜.这种纳米纤维膜具有极大的比表面积,已用于超高效过滤,在刨伤修复、组织工程、水处理等领域有广泛的应用前景.为了进一步拓展纳米纤维在纤维工业、纺织品、微制造等领域的应用,电纺纳米纤维的取向和连续长纱的制备研究受到科学家的重视,文献报道了多种纳米纤维取向方法.本文分析了纳米纤维膜无规堆砌结构的形成机理,总结了纳米纤维取向研究和连续长纱制备研究进展,特别介绍了基于静电作用分析提出的共轭电纺方法,讨论了取向纳米纤维的应用以及纳米纤维未来的研究方向.