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Immunomodulation is an important phenomenon in the normal mammalian host response toward an injury, and plays a critical role in tissue regeneration and regenerative medicine. Different phenotypes of macrophages show an array of activation states compassing pro-inflammatory to pro-alleviating cells, which are the critical players to modulate immune response and tissue regeneration. In this study, macrophage membranes of different phenotypes (macrophages (M0), classically activated macrophages (M1) and alternatively activated macrophages (M2)) were coated onto poly-ε-caprolactone (PCL) nanofibers to acquire exterior surface proteins and similar functions of the natural membranes. In vitro results unveiled that these nanofibers, especially the M2-PCL nanofibers, can suppress the activities of inflammatory markers such as TNF-α and IL-1β, and stimulate anti-inflammatory markers such as Arg-1, IL-10 and TGF-β. In a C57BL/6 mouse model, the macrophage membrane-coated nanofibers, especially the M2-PCL nanofibers, displayed minimal cellular infiltration and low collagen deposition, increased anti-inflammatory CD206 and decreased inflammatory CD86 levels. The M2-PCL nanofibers most effectively neutralized inflammatory chemokines, regulated the expression of inflammation-associated genes as well as anti-inflammatory genes, and showed strong immunomodulatory effects than the PCL, M0-PCL and M1-PCL nanofibers. STATEMENT OF SIGNIFICANCE: Different types of macrophage membrane-functionalized PCL nanofibers were successfully prepared and well characterized. They inherited the surface proteins imitating the source macrophages, and played an important role in limiting cellular infiltration and collagen deposition. These different macrophages and their membrane-coated nanofibers (M0-PCL, M1-PCL and M2-PCL) behaved like their respective source cells. The M2 mimicking M2-PCL nanofibers effectively polarized macrophages to M2 phenotype and decreased the expression of inflammation-associated chemokines and promoted the anti-inflammation in vitro and in vivo, which is critical for tissue regeneration. The mice implanted with the bio-mimicking M2-PCL nanofibers effectively inhibited toll like receptors signaling induced NF-kB and IRF-5 and their target genes such as Edn-1, IL-6, iNOS, TNF-α, etc. compared to the PCL, and M0-PCL and M1-PCL macrophage membrane-coated nanofibers.

Regeneration and functional recovery of peripheral nerves remain formidable due to the inefficient physical and chemical cues in the available nerve guidance conduits (NGCs). Introducing micropatterns and bioactive substances into the inner wall of NGCs can effectively regulate the behavior of Schwann cells, the elongation of axons, and the phenotype of macrophages, thereby aiding the regeneration of injured nerve. In this study, linear micropatterns with ridges and grooves of 3/3, 5/5, 10/10, and 30/30 μm were created on poly(d,l-lactide-co-caprolactone) (PLCL) films following with surface aminolysis and electrostatic adsorption of graphene oxide (GO) nanosheets. The GO-modified micropatterns could significantly accelerate the collective migration of Schwann cells (SCs) and migration of SCs from their spheroids in vitro. Moreover, the SCs migrated directionally along the stripes with a fastest rate on the 3/3-GO film that had the largest cell adhesion force. The neurites of N2a cells were oriented along the micropatterns, and the macrophages tended to differentiate into the M2 type on the 3/3-GO film judged by the higher expression of Arg 1 and IL-10. The systematic histological and functional assessments of the regenerated nerves at 4 and 8 weeks post-surgery in vivo confirmed that the 3/3-GO NGCs had better performance to promote the nerve regeneration, and the CMAP, NCV, wet weight of gastrocnemius muscle, positive S100β and NF200 area percentages, and average myelinated axon diameter were more close to those of the autograft group at 8 weeks. This type of NGCs thus has a great potential for nerve regeneration.

Cellular metabolism plays a major role in the regulation of inflammation. The inflammatory macrophages undergo a wide-range of metabolic rewriting due to the production of significant amount of itaconate metabolite from cis-aconitate in the tricarboxylic acid cycle. This itaconate molecule has been recently described as a promising immunoregulator. However, its function and mode of action on macrophages and tissue repair and regeneration are yet unclear. Herein, the itaconate-derivative dimethyl itaconate (DMI) suppresses the IL-23/IL-17 inflammatory axis-associated genes and promotes antioxidant nuclear factor erythroid 2-related factor 2 target genes. The poly-e-caprolactone (PCL)/DMI nanofibers implanted in mice initially maintain inflammation by suppressing anti-inflammatory activity and particular inflammation, while at later stage promotes anti-inflammatory activity for an appropriate tissue repair. Furthermore, the PCL/DMI nanofiber patches show an excellent myocardial protection by reducing infarct area and improving ventricular function via time-dependent regulation of myocardium-associated genes. This study unveils potential DMI macrophage modulatory functions in tissue microenvironment and macrophages rewriting for proper tissue repair.

The excessive reactive oxygen species (ROS) and hypoxia deteriorate the inflammation-related diseases such as myocardial infarction (MI), and thereby deter the normal tissue repair and recovery and further lead to severe fibrosis and malfunction of tissues and organs. In particular, the MI has become one of the leading causes of death nowadays. In this study, a novel type of injectable hydrogel with dual functions of ROS scavenging and O

Regeneration and functional recovery of peripheral nerves remain formidable due to the inefficient physical and chemical cues in the available nerve guidance conduits (NGCs). Introducing micropatterns and bioactive substances into the inner wall of NGCs can effectively regulate the behavior of Schwann cells, the elongation of axons, and the phenotype of macrophages, thereby aiding the regeneration of injured nerve. In this study, linear micropatterns with ridges and grooves of 3/3, 5/5, 10/10, and 30/30 μm were created on poly(d,l-lactide

Cell migration plays a pivotal role in many pathological and physiological processes. So far, most of the studies have been focused on 2-dimensional cell adhesion and migration. Herein, the migration behaviors of cell spheroids in 3D hydrogels obtained by polymerization of methacrylated hyaluronic acid (HA-MA) and fibrinogen (Fg) with different ratios were studied. The Fg could be released to the medium gradually along with time prolongation, achieving the dynamic change of hydrogel structures and properties. Three types of cell spheroids, i.e., endothelial cell (EC), smooth muscle cell (SMC), and EC-SMC spheroids, were prepared with 10,000 cells in each, whose diameters were about 343, 108, and 224 μ m, respectively. The composite hydrogels with an intermediate ratio of Fg allowed the fastest 3D migration of cell spheroids. The ECs-SMCs migrated longest up to 3200 μ m at day 14, whereas the SMC spheroids migrated slowest with a distance of only ~400 μ m at the same period of time. The addition of free RGD or anti-CD44 could significantly reduce the migration distance, revealing that the cell-substrate interactions take the major roles and the migration is mesenchymal dependent. Moreover, addition of anti-N-cadherin and MMP inhibitors also slowed down the migration rate, demonstrating that the degradation of hydrogels and cell-cell interactions are also largely involved in the cell migration. RT-PCR measurement showed that expression of genes related to cell adhesion and antiapoptosis, and angiogenesis was all upregulated in the EC-SMC spheroids than single EC or SMC spheroids, suggesting that the use of composite cell spheroids is more promising to promote cell-substrate interactions and maintenance of cell functions.

Cell migration plays an important role in many physiological and biological processes , which is influenced by both physicochemical properties of surrounding matrix and signal gradient generated by neighboring/remote cells. Here we aim to develop a co-culture system of immune cells and smooth muscle cells (SMCs) based on the combination of Transwell and cell-responsive hydrogels. This model can be used to study the cell invasion into hydrogels in dynamic physiological conditions , with better mimicking of the in vivo microenvironment . Methacrylic anhydride-modified hyaluronic acid (MA-HA) macromolecules were crosslinked by matrix metalloproteinases (MMPs) sensitive peptides (MMP SP) to fabricate a cell-degradable hydrogel mimicking dynamic extracellular matrix (ECM). The migration of SMCs into the MMP-sensitive hydrogel was investigated under the existence of U937 cells, a type of macrophage-like cells. The invasion distance of SMCs in the MMP-sensitive hydrogels was much longer than that in the MMP-insensitive ones both in vitro and in vivo. The impact of hydrogel degradability and inductive signal gradient generated by U937 cells on cell invasion was compared, revealing that the degradability plays a major role in regulating cell invasion into the 3D hydrogels. Further mechanism investigation revealed that the expressions of cell migration-related genes and proteins were significantly up-regulated in the MMP-sensitive hydrogels compared to those in the MMP-insensitive hydrogels.

Selective adhesion and migration of smooth muscle cells (SMCs) over fibroblasts (FIBs) is required to prevent adventitia fibrosis in vascular regeneration. In this study, a uniform cell-resisting layer of poly(ethylene glycol) (PEG) with a density gradient of azide groups was generated on a substrate by immobilizing two kinds of PEG molecules in a gradient manner. A density gradient of alkynyl-functionalized Val-Ala-Pro-Gly (VAPG) peptides was then prepared on the PEG layer via click chemistry. The VAPG density gradient was characterized by fluorescence imaging, revealing the gradual enhancement of the fluorescent intensity along the substrate direction. The adhesion and mobility of SMCs were selectively enhanced on the VAPG density gradient, leading to directional migration toward the higher peptide density (up to 84%). In contrast, the adhesion and mobility of FIBs were significantly weakened. The net displacement of SMCs also significantly increased compared with that on tissue culture polystyrene (TCPS) and that of FIBs on the gradient. The mitogen-activated protein kinase (MAPK) signaling pathways related to cell migration were studied, showing higher expressions of functional proteins from SMCs on the VAPG-modified surface in a density-dependent manner. For the first time the selective adhesion and directional migration of SMCs over FIBs was achieved by an elaborative design of a gradient surface, leading to a new insight in design of novel vascular regenerative materials. Statement of Significance Selective cell adhesion and migration guided by regenerative biomaterials are extremely important for the regeneration of targeted tissues, which can avoid the drawbacks of incorrect and uncontrolled responses of tissue cells to implants. For example, selectivity of smooth muscle cells (SMCs) over fibroblasts (FIBs) is required to prevent adventitia fibrosis in vascular regeneration. Herein we prepare a uniform cell-repelling layer, on which SMCs-selective Val-Ala-Pro-Gly (VAPG) peptides are immobilized in a continuous manner. Selective adhesion and enhanced and directional migration of SMCs over FIBs are achieved by the interplay of cell-repelling layer and gradient SMCs-selective VAPG peptides, paving a new way for the design of novel vascular grafts with enhanced biological performance.

The design of hyaluronic acid (HA)-based and stimuli-responsive hydrogels to elicit highly controlled and tunable cell response and behaviors is a major field of interest in tissue engineering and regenerative medicine. The pH-responsive hydrogel can respond to pH variation during wound healing, which may in turn regulate the tissue regeneration process. In this study, a double-network hydrogel cross-linked with vinyl double bonds and Schiff base was prepared, whose properties were further adjusted by incubation in pH 7.4 and pH 5 buffers. The endothelial cells (ECs) migrated much deeper into the softer HA hydrogel pre-treated with pH 5 buffer than the stiffer hydrogel. By contrast, the mesenchymal stem cells (MSCs) migrated easily into the stiffer hydrogel. The ECs highly expressed RhoA and non-muscle myosin (NM) II genes in the softer hydrogel, which may facilitate amoeboid migration. Meanwhile, the MSCs were stiffer than the ECs, and highly expressed Rac1, RhoA, vinculin, NM II, hyaluronidase (HYAL) 2 and CD44 genes in the stiffer hydrogel, which facilitate mesenchymal migration. These results provide important clues for revealing the different migration strategies of the ECs and MSCs in HA hydrogels with different stiffness, and suggest that the mechanical properties and the network structure of hydrogels play an important role in regulating the three-dimensional migration process of these cells.

The recruitment of endogenous mesenchymal stem cells (MSCs), as an alluring approach for in situ tissue regeneration, always accompanies with other types of cells. Therefore, it is of enormous value to bestow a substrate with the property of selective capture to MSCs. However, it was reported that when MSCs are cultured on a substrate with excessive affinity, their stemness diminished. Therefore, constructing a substrate with the balanced ability of selective capture and stemness maintenance becomes a big challenge. In this study, an Aptamer 19S (Apt19S)-modified substrate was fabricated by grafting Apt19S on a PEGylated glass substrate. The X-ray photoelectron spectroscopy results verified that the antifouling poly(ethylene glycol) (PEG) layer was created. Tracking by ellipsometry, the thicknesses of PEG layers were proved to increase with PEG concentration. The results of the quartz crystal microbalance also validated that the Apt19S densities could be modulated by the concentrations of the Apt19S solution. The results of the cell adhesion assay indicated that the modification of Apt19S caused a significant increase in the adhesion ratio and area of rBMSCs. Selective adhesion was confirmed by coculture of rBMSCs with macrophages and NIH3T3 cells, demonstrating that a higher proportion of rBMSCs adhered to the Apt19S-modified substrate. The results of specific capture were further confirmed by a flow model to simulate the body fluid flow. The comprehensive results of reverse transcription polymerase chain reaction, immunofluorescence staining, proliferation capacity, and differentiation assay showed that the stemness of rBMSCs was maintained better on a substrate with the appropriate Apt19S density. All of these results indicated that Apt19S modification is an effective strategy to endow a substrate with the specific capture ability of MSCs, and the balance between selective capture and stemness maintenance can be achieved by the precise regulation of the aptamer density.

Capture of endothelial progenitor cells (EPCs) in situ has been considered as a promising strategy for the rapid endothelialization and long-term patency of artificial blood vessels and implant devices. In this study, a CD133+ EPC capture surface was fabricated by grafting CD133 antibody (a more specific EPC surface marker than CD34) and Arg-Glu-Asp-Val (REDV) peptideon the methacrylate-grafted hyaluronic acid (MA-HA) and heparin-hybridized (MA-HA&Heparin) resisting layer. Vascular endothelial growth factor (VEGF) was further conjugated to the immobilized heparin. This engineered surface showed good hemocompatibility and significantly higher ability of capturing CD133+ EPCs from human peripheral blood mononuclear cells (PBMCs) and obviously upregulated the expression of endothelial cell (EC) marker genes of EPCs such as VEGF receptor 2 (VEGFR2), CD31, VE-cadherin, and von Willebrand factor (vWF), facilitating the differentiation of EPCs into ECs. The dramatically enhanced EPC proliferation on this surface was dependent on the integrin-VEGFR synergistic signaling, as ERK1/2 phosphorylation was only significantly enhanced on the REDV and VEGF co-immobilized surface. This study highlights a new surface coating strategy for blood-contact materials based on the specific EPC capturing and rapid endothelialization. STATEMENT OF SIGNIFICANCE: Capture of endothelial progenitor cells (EPCs) in situ is a promising strategy for the rapid endothelialization and long-term patency of artificial blood vessels and scaffolds. More specific capture of EPCs by targeting CD133 rather than CD34 can better reduce the risk of inflammation and restenosis. On the other hand, an appropriate microenvironment for EPC proliferation is equally important for endothelialization, which is rarely considered by the existing EPC capture strategies. In this study, the capture ratio of EPCs was significantly increased by simultaneously grafting CD133 antibody and VEGF on a MA-HA and heparin-hybridized antifouling layer. Further, proliferation of EPCs after capture was significantly promoted by grafting VEGF and REDV peptide through the integrin-VEGFR synergistic signaling. This study highlights a new strategy for the surface coating of blood-contact materials based on specific EPC capture and rapid endothelialization.

The biodegradable polymer microparticles with different surface morphology and chemical compositions may influence significantly the behaviors of cells, and thereby further the performance of tissue regeneration in vivo. In this study, multi-stage hierarchical textures of poly(D,L-lactic-co-glycolide) (PLGA)/PLGA-b -PEG (poly(ethylene glycol)) microspheres with a diameter as large as 50–100 μm are fabricated based on interfacial instability of an emulsion. The obtained fuzzy structures on the microspheres are sensitive to annealing, which are changed gradually to a smooth one after treatment at 37 °C for 6 d or 80 °C for 1 h. The surface microstructures that are chemically dominated by PEG can be stabilized against annealing by dopamine deposition. By the combination use of annealing and dopamine deposition, a series of microspheres with robust surface topologies are facilely prepared. The fuzzy microstructures and dopamine deposition show a synergetic role to enhance cell-material interaction, leading to a larger number of adherent bone marrow-derived mesenchymal stem cells (BMSCs), A549 and MC 3T3 cells. The fuzzy microspheres with dopamine deposition can significantly promote bone regeneration 12 w post surgery in vivo, as revealed by micro-CT, histological, western blotting and RT-PCR analyses.

The excessive reactive oxygen species (ROS) and hypoxia deteriorate the inflammation-related diseases such as myocardial infarction (MI), and thereby deter the normal tissue repair and recovery and further lead to severe fibrosis and malfunction of tissues and organs. In particular, the MI has become one of the leading causes of death nowadays. In this study, a novel type of injectable hydrogel with dual functions of ROS scavenging and O2 generating is fabricated for MI treatment in vivo. The hydrogel is formed within 3 s from the synthetic ROS-cleavable hyperbranched polymers and methacrylate hyaluronic acid (HA-MA) under UV-irradiation. Addition of biocompatible and applicable catalase in vivo enables the further transition of H2 O2 , a major type of ROS, to O2 and H2 O. Results of rat MI model demonstrate that this hydrogel can significantly remove excessive ROS, inhibit cell apoptosis, increase M2/M1 macrophage ratio, promote angiogenesis, reduce infarcted area, and improve cardiac functions. With the appropriate degradation rate, simple structure and composition without cell seeding, and very excellent MI therapeutic effect, this ROS scavenging and O2 generating hydrogel has a great promise to be applied clinically.

The immune system responds immediately to tissue trauma and to biomaterial implants under the participation of M1/M2 macrophages polarization. The surface properties of biomaterials can significantly influence the tissue repair progress through modulating the macrophage functions. In this study, the surface of poly(propylene fumarate) polyurethane films (PPFU) is grafted with a same density of enantiomeric poly-l-lysine (PPFU-g-PLL) and poly-d-lysine (PPFU-g-PDL), leading to a similar level of enhanced surface wettability for the PPFU-g-PLL and PPFU-g-PDL. The polylysine-grafted PPFU can restrict the M1 polarization, whereas promote M2 polarization of macrophages in vitro, judging from the secretion of cytokines and expression of key M1 and M2 related genes. Comparatively, the PPFU-g-PDL has a stronger effect in inducing M2 polarization in vivo, resulting in a thinner fibrous capsule surrounding the implant biomaterials. The CD44 and integrins of macrophages participate in the polarization process probably by activating focal adhesion kinase (FAK) and Rho-associated protein kinase (ROCK), and downstream PI3K/Akt1/mTOR signal axis to up regulate M2 related gene expression. This study confirms for the first time that polylysine coating is an effective method to regulate the immune response of biomaterials, and the polylysine-modified thermoplastic PPFU with the advantage to promote M2 polarization may be applied widely in regenerative medicine.

External stimuli-responsive biomaterials represent a type of promising candidates for addressing the complexity of biological systems. In this study, a platform based on the combination of temperature-sensitive polymers and a nanotube array was developed for loading sphingosine 1-phosphate (S1P) and regulating the migration of endothelial cells (ECs) at desired conditions. The localized release dosage of effectors could be controlled by the change of environmental temperature. At a culture temperature above the lower critical solution temperature, the polymer "gatekeeper" with a collapsed conformation allowed the release of S1P, which in turn enhanced the migration of ECs. The migration rate of single cells was significantly enhanced up to 58.5%, and the collective migration distance was also promoted to 25.1% at 24 h and 33.2% at 48 h. The cell morphology, focal adhesion, organization of cytoskeleton, and expression of genes and proteins related to migration were studied to unveil the intrinsic mechanisms. The cell mobility was regulated by the released S1P, which would bind with the S1PR1 receptor on the cell membrane and trigger the Rho GTPase pathway.

The gradient localization of biological cues is of paramount importance to guide directional migration of cells. In this study, poly(2-hydroxyethyl methacrylate-co-glycidyl methacrylate)-block- poly(2-hydroxyethyl methacrylate) (P(HEMA-co-GMA)-b-PHEMA) brushes with a uniform underneath P(HEMA-co-GMA) layer and a gradient thickness of PHEMA blocks were prepared by using surface-initiated atom-transfer radical polymerization and a dynamically controlled polymerization process. The polymer chains were subsequently functionalized with the cell-adhesive arginine-glycine-aspartic acid (RGD) peptides by reaction with the glycidyl groups, and their structures and properties were characterized by X-ray photoelectron spectrometry (XPS), quartz crystal microbalance with dissipation (QCM-D) and air contact angle. Adhesion and migration processes of smooth muscle cells (SMCs) were then studied. Compared with those on the sufficiently exposed RGD surface, the cell adhesion and mobility were well maintained when the RGD peptides were localized at 18.9 nm depth, whereas the adhesion, spreading and migration rate of SMCs were significantly impaired when the RGD peptides were localized at a depth of 38.4 nm. On the RGD depth gradient surface, the SMCs exhibited preferential orientation and enhanced directional migration toward the direction of reduced thickness of the second PHEMA brushes. Half of the cells were oriented within ± 30° to the x-axis direction, and 72% of the cells moved directionally at the optimal conditions. Cell adhesion strength, arrangement of cytoskeleton, and gene and protein expression levels of adhesion-related proteins were studied to corroborate the mechanisms, demonstrating that the cell mobility is regulated by the complex and synergetic intracellular signals resulted from the difference in surface properties. Statement of Significance Cell migration is of paramount importance for the processes of tissue repair and regeneration. So far, the gradient localization of biological cues perpendicular to the substrate, which is the usual case for the biological signaling molecules to locate in ECM in vivo, has been scarcely studied, and has not been used to guide the directional migration of cells. In this study, we prepare a depth gradient of RGD peptides along the polymer chains, which is used to guide the directional migration of SMCs after a second hydrophilic bock is prepared in a gradient manner. For the first time the directional migration of SMCs is achieved under the guidance of a depth gradient of RGD ligands. The mechanisms of different cell migration abilities are further discussed based on the results of cell adhesion, cell adhesion force, cytoskeleton alignment and expression of relative proteins and genes. This work paves a new strategy by fabricating a gradient polymer brushes with immobilized bioactive molecules to dominate the directional cell migration, and elucidates the mechanisms underlining the biased migration along RGD depth localization gradients, shedding a light for the design of novel biomaterials to control and guide cell migration and invasion.

The interplay of microstructures and biological cues is critical to regulate the behaviors of Schwann cells (SCs) in terms of cellular spatial arrangement and directional migration as well as neurite orientation for bridging the proximal and distal stumps of the injured peripheral nervous system. In this study, stripe micropatterns having ridges/grooves of width 20/20 and 20/40 μm were fabricated on the surface of maleimide-functionalized biodegradable poly(ester carbonate) (P(LLA-MTMC)) films by the polydimethylsiloxane mold-pressing method, respectively. The laminin-derived CQAASIKVAV peptides end-capped with an SH group were then grafted by the thiol-ene click reaction under mild conditions to obtain micropatterned and peptide-grafted films. SCs cultured on these films, especially on the 20/40-μm film, displayed faster and aligned adhesion as well as a larger number of elongated cells with a higher length-to-width (L/W) ratio along the stripe direction than those on the flat-pep film. The migration rate of SCs was significantly enhanced in parallel to the stripe direction with a large net displacement. The micropatterned and peptide-grafted films, especially the 20/40-μm film, could promote SC proliferation and nerve growth factor (NGF) secretion in a manner similar to that of the peptide-grafted planar film. Moreover, the neurites of rat pheochromocytoma 12 (PC12) cells sprouted along the ridges with a longer average length on the micropatterned and peptide-grafted films. The synergistic effect of physical patterns and biological cues was evaluated by considering the results of cell adhesion force; immunofluorescence staining of vinculin; fluorescence staining of F-actin and the nucleus; as well as gene expression of neural cadherin (NCAD), neurocan (NCAN), and myelin protein zero (P0). Statement of Significance The interplay of microstructures and biological cues is critical to regulate the behaviors of Schwann cells (SCs) and nerve cells, and thereby the regeneration of peripheral nerve system. In this study, the combined micropatterning and CQAASIKVAV grafting endowed the modified P(LLA-MTMC) films with both contact guidance and bioactive chemical cues to enhance cell proliferation, directional alignment and migration, longer net displacement and larger NGF secretion, and stronger neurite outgrowth of SCs and PC12 cells. Hence, the integration of physical micropatterns and bioactive molecules is an effective way to obtain featured biomaterials for the regeneration of nerves and other types of tissues.

A disintegrin and metalloproteinase 33 (ADAM33) has been identified as a susceptibility gene for asthma, but details of the causality are not fully understood. We hypothesize that soluble ADAM33 (sADAM33) overexpression can alter the mechanical behaviors of airway smooth muscle cells (ASMCs) via regulation of the cell's contractile phenotype, and thus contributes to airway hyperresponsiveness (AHR) in asthma. To test this hypothesis, we either overexpressed or knocked down the sADAM33 in rat ASMCs by transfecting the cells with sADAM33 coding sequence or a small interfering RNA (siRNA) that specifically targets the ADAM33 disintegrin domain, and subsequently assessed the cells for stiffness, contractility and traction force, together with the expression level of contractile and proliferative phenotype markers. We also investigated whether these changes were dependent on Rho/ROCK pathway by culturing the ASMCs either in the absence or presence of ROCK inhibitor (H1152). The results showed that the ASMCs with sADAM33 overexpression were stiffer and more contractile, generated greater traction force, exhibited increased expression levels of contractile phenotype markers and markedly enhanced Rho activation. Furthermore these changes were largely attenuated when the cells were cultured in the presence of H-1152. However, the knock-down of ADAM33 seemed insufficient to influence majority of the mechanical behaviors of the ASMCs. Taken together, we demonstrated that sADAM33 overexpression altered the mechanical behaviors of ASMCs in vitro, which was most likely by promoting a hypercontractile phenotype transition of ASMCs through Rho/ROCK pathway. This revelation may establish the previously missing link between ADAM33 expression and AHR, and also provide useful insight for targeting sADAM33 in asthma prevention and therapy.

The review aimed to assess the current research status of polymer brushes and artificial cilia, based on the analysis of polymer brush properties and the structure and function of natural cilia. The application of polymer brushes to develop artificial cilia has attracted increasing research interest, with rheological analysis often employed as an entry point for polymer mechanics. The present review assessed whether combing polymer brushes with artificial cilia, along with appropriate designing of high‑polymer materials and use of suitable equipment, may to the development of a polymer net‑structured material to imitate the morphology and functionality of a natural cilium system.

Scientific reports 6, 30451 (2016). doi:10.1038/srep30451, Published by Macmillan Publishers Limited, part of Springer Nature, [London]

Airway inflammation has been suggested as the pathological basis in asthma pathogenesis. Recruitment of leukocytes from the vasculature into airway sites is essential for induction of airway inflammation, a process thought to be mediated by a disintegrin and metalloprotease 8 (ADAM8). However, there is an apparent controversy about whether ADAM8 helps or hampers transmigration of leukocytes through endothelium in airway inflammation of asthma. This review outlines the current contradictory concepts concerning the role of ADAM8 in airway inflammation, particularly focusing on the recruitment of leukocytes during asthma, and attempts to bridge the existing experimental data on the basis of the functional analysis of different domains of ADAM8 and their endogenous processing in vivo. We suggest a possible hypothesis for the specific mechanism by which ADAM8 regulates the transmigration of leukocytes to explain the disparity existing in current studies, and we also raise some questions that require future investigations.

A disintegrin and metalloproteinase 33 (ADAM33) has been identified as an asthma susceptibility gene; however, the role of ADAM33 in the pathogenesis and progression of asthma remains to be elucidated. As ADAM33 is predominantly expressed in airway smooth muscle cells (ASMCs), it is feasible to investigate whether ADAM33 protein expression is correlated with ASMC mechanics that are ultimately responsible for airway hyperresponsiveness in asthma. To determine this, Sprague Dawley rats were sensitized with ovalbumin (OVA) for up to 12 weeks to simulate asthma symptoms. Subsequently, ASMCs were isolated from the rats and cultured in vitro. The protein expression of ADAM33 and cytoskeletal proteins (including F‑actin and vinculin), cell stiffness and contractility, as well as traction force were measured. The results demonstrated that compared with the non‑sensitized rats, the protein expression of ADAM33 in ASMCs from the OVA‑sensitized rats increased in a time‑dependent manner, reaching a maximum level at 4 weeks of sensitization and gradually subsiding as OVA sensitization continued (P

Chronic inflammation and remodeling of the bron - chial wall are basic hallmarks of asthma. During the process of bronchial wall remodeling, inflammatory factors , such as trans- forming growth factor-β1 (TGF-β1), are known to induce the differentiation of fibroblasts into myofibroblasts, which leads to excessive synthesis and secretion of extracellular matrix (ECM) proteins, thus thickening and stiffening the basement membrane. However, it has not been thoroughly studied whether or not substrate stiffening affects the TGF-β1-induced myofibrobl ast differentiation. In the present study, the influence of substrate stiffness on the process of bronchial fibroblast differentiation into myofibroblasts in the presence of TGF-β1 was investigated. To address this question, we synthesized polydimethylsiloxane (PDMS) substrates with varying degrees of stiffness (Young's modulus of 1, 10 and 50 kPa, respectively). We cultured bronchial fibroblasts on the substrates of varying stiffness in media containing TGF- β1 (10 ng/ml) to stimulate the differentiation of fibroblasts into myofibroblasts. Myofibroblast differentiation was examined using semi-quantitative RT-PCR for the expression of α-smooth muscle actin (α-SMA) mRNA and collagen I mRNA, the enzyme-linked immunosorbent assay method was used to assess the expression of collagen I protein and western blotting to assess the expression of α-SMA protein. The optical magnetic twisting cytometry (OMTC) method was used for the changing of cell mechanical proper- ties. Our findings suggest that when fibroblasts were incubated with TGF-β1 (10 ng/ml) on substrate of varying stiffness, the differentiation of fibroblasts into myofibroblasts was enhanced by increasing substrate stiffness. Compared with those cultured on substrate with Young's modulus of 1 kPa, the mRNA and protein expression of collagen I and α-SMA of fibroblasts cultured on substrates with Young's modulus of 10 and 50 kPa were increased. Furthermore, with the increase of substrate stiffness, the cell stiffness and contractility were also increased, which also indicated further aggravation of asthma. This finding may help better understand the underlying mechanisms of hyperplasia of myofibroblasts in asthma, which has a marked significance in the therapy of asthma.

ADAM (a disintegrin and metalloprotease) genes have been identified in various tissues and species, and recently associated with several important human diseases such as tumor and asthma. Although various biological processes have been known for the ADAM family in different species including fertilization, neurogenesis, infection and inflammation, little is known about its detailed phylogenetic and molecular evolutionary history. In this study, the ADAMs of Xenopus (Silurana) tropicalis, Mus musculus, Rattus norvegicus, and Homo sapiens were collected and analyzed by using the Bayesian analysis and gene synteny analysis to establish a comprehensive phylogenetic relationship and evolutionary drive of this gene family. It was found that there were more ADAMs in the two rodents than in the amphibian, suggesting an expansion of the ADAM gene family during the early evolution of mammals. All ADAMs from this expansion were retained in both the rodents, but other duplication events occurred subsequently in the two rodents, respectively, leading to the classification of rodent ADAMs as classes I, II and III. Moreover, these duplicated ADAM genes in the rodents were found to be driven by positive selection, which might be the major force to retain them in the genome. Importantly, it was also found that orthologs of ADAM3 and 5 have been lost in humans. These results not only provide valuable information of the evolution of ADAM genes, but may also help in understanding the role of ADAM genes in the pathobiology of relevant diseases.