Electrospinning is a versatile technique for the construction of microfibrous and nanofibrous structures with considerable potential in applications ranging from textile manufacturing to tissue engineering scaffolds. In the simplest form, electrospinning uses a high voltage of tens of thousands volts to draw out ultrafine polymer fibers over a large distance. However, the high voltage limits the flexible combination of material selection, deposition substrate, and control of patterns. Prior studies show that by performing electrospinning with a well-defined "near-field" condition, the operation voltage can be decreased to the kilovolt range, and further enable more precise patterning of fibril structures on a planar surface. In this work, by using solution dependent "initiators", we demonstrate a further lowering of voltage with an ultralow voltage continuous electrospinning patterning (LEP) technique, which reduces the applied voltage threshold to as low as 50 V, simultaneously permitting direct fiber patterning. The versatility of LEP is shown using a wide range of combination of polymer and solvent systems for thermoplastics and biopolymers. Novel functionalities are also incorporated when a low voltage mode is used in place of a high voltage mode, such as direct printing of living bacteria; the construction of suspended single fibers and membrane networks. The LEP technique reported here should open up new avenues in the patterning of bioelements and free-form nano- to microscale fibrous structures.

Recreating tissue-specific microenvironments of the extracellular matrix (ECM) is of broad interest for the fields of tissue engineering and organ-on-a-chip. Both the biochemical and biophysical signatures of the engineered ECM interplay to affect cell response. Currently, there are limited biomaterials processing methods which allow to design ECM membrane properties flexibly and rapidly. Solvents and additives used in many existing processes also induced unwanted ECM protein degradation and toxic residues. This paper presents a solution fibre spinning technique, where careful selection of the solution combination led to well-preserved ECM proteins with tuneable composition. This technique also provides a highly versatile approach to fabricate ECM fibres and membranes, leading to designable fibre Young's modulus for over two orders of magnitude.

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Cellular behavior is strongly influenced by the architecture and pattern of its interfacing extracellular matrix (ECM). For an artificial culture system which could eventually benefit the translation of scientific findings into therapeutic development, the system should capture the key characteristics of a physiological microenvironment. At the same time, it should also enable standardized, high throughput data acquisition. Since an ECM is composed of different fibrous proteins, studying cellular interaction with individual fibrils will be of physiological relevance. In this study, we employ near-field electrospinning to create ordered patterns of collagenous fibrils of gelatin, based on an acetic acid and ethyl acetate aqueous co-solvent system. Tunable conformations of micro-fibrils were directly deposited onto soft polymeric substrates in a single step. We observe that global topographical features of straight lines, beads-on-strings, and curls are dictated by solution conductivity; whereas the finer details such as the fiber cross-sectional profile are tuned by solution viscosity. Using these fibril constructs as cellular assays, we study EA.hy926 endothelial cells' response to ROCK inhibition, because of ROCK's key role in the regulation of cell shape. The fibril array was shown to modulate the cellular morphology towards a pre-capillary cord-like phenotype, which was otherwise not observed on a flat 2-D substrate. Further facilitated by quantitative analysis of morphological parameters, the fibril platform also provides better dissection in the cells' response to a H1152 ROCK inhibitor. In conclusion, the near-field electrospun fibril constructs provide a more physiologically-relevant platform compared to a featureless 2-D surface, and simultaneously permit statistical single-cell image cytometry using conventional microscopy systems. The patterning approach described here is also expected to form the basics for depositing other protein fibrils, seen among potential applications as culture platforms for drug screening.

We used samples from rheumatoid arthritis (RA) patients to examine whether Anti-citrullinated protein antibodies (ACPAs) alter macrophage subset distribution and promote RA development. Macrophage subset distributions and interferon regulatory factor 4 (IRF4) and IRF5 expressions were analyzed. ACPAs were purified by affinity column. After RA and osteoarthritis (OA) patients' macrophages were cocultured with ACPAs, macrophage subsets and IRF4 and IRF5 expressions were measured. Small interfering RNAs (siRNAs) were transfected into ACPA-activated cells to suppress IRF4 or IRF5. Fluorescence-activated cell sorting (FACS), Western blot, and immunohistochemistry were performed. Macrophage subset disequilibrium occurred in RA patient synovial fluids. IRF4 and IRF5 were all expressed in the synovial fluid and synovium. ACPAs (40 IU/ml) could induce macrophages to polarize to M1 subsets, and the percentage of increased M1/M2 ratio of RA patients was higher than that of the OA patients. ACPAs also induce IRF4 and IRF5 protein expressions. IRF5 siRNA transfection impaired ACPA activity significantly. We demonstrated that macrophage subset disequilibrium occurred in RA patients. ACPAs induced IRF5 activity and led to M1 macrophage polarization.

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