Adhesion Of Cells Research Articles

Differential Effect of Novel Plant Cystatins on the Adhesive Behaviour of Normal and Cancer Breast Cells

In the present work, we have investigated a novel recombinant cystatin dgECP1 and its mutant form, dgECP1m1, focused on their impact on the adhesive behaviour of two breast cell lines: the cancerous, MDA-MB-231, and the normal, MCF-10A. DgECP1 cystatin is intriguing with its RGD motif, responsible for cell adhesion and typical for mammalian extracellular matrix proteins but uncommon for plant cystatins. The presence of the RGD sequence suggests the potential of the dgECP1 to influence the adhesion of cancer cells and, respectively, cancer metastasis. A mutant form of the dgECP1cystatin, dgECP1m1, where RGD is replaced with HGD tripeptide, was also investigated. We found that both phytocystatins exerted differential effects on the adhesion behaviour of normal and cancer cells. In the case of dgECP1 cystatins, the effect on cancer cell adhesion also depends on the mode of administration of the cystatin to cells. When dgECP1 is pre-adsorbed on a substrate, it improves the attachment of breast cancer cells and induces cell aggregation, which is more typical for normal breast cells, and oppositely suppressed adhesion of cancer cells when added to the medium. The mutant form, dgECP1m1, inhibited cancer cell adhesion independently on the way of administration. On the other hand, both plant cystatins only slightly reduced the adhesion of normal mammary cells pointing to the higher sensitivity of cancer cells to both cystatins. These preliminary results open the possibility of considering the plant cystatin dgECP1 for anti-cancer strategies.

Highly Ordered Nanotube-Like Microstructure on Titanium Dental Implant Surface Fabricated via Anodization Enhanced Cell Adhesion and Migration of Human Gingival Fibroblasts.

Titanium (Ti) surface with nanotubes array via anodization has been used in dental implants to enhance bone regeneration but little research was carried out to evaluate whether the presence of highly ordered or disorderly distributed nanotubes array on titanium surface would have an effect on cell behaviors of gingival fibroblasts. The present study fabricated nanotubes arrays with varied topography under different constant voltage of electrochemical anodization in fluorine-containing electrolyte. Human gingival fibroblasts (HGFs) from extracted third molar were harvested and co-cultured with titanium disks with different nanotubes topography. Then cell behaviors of gingival fibroblasts including cell proliferation, adhesive morphology and cell migration were estimated to investigate the influence of titanium nanotubes on cell biology. Besides, gene and protein expression of adhesion molecule (integrin β1/β4/α6, fibronectin, intracellular adhesion molecule-1 and collagen type I) were detected to evaluate the influence of different surfaces on cell adhesion. Highly ordered arrays of nanotubes with pore diameter of 60 nm and 100 nm were fabricated under 30 and 40 V of anodization (TNT-30 and TNT-40) while disorderedly distributed nanotube arrays formed on the titanium surface under 50 V of anodization (TNT-50). Our results demonstrated that compared with raw titanium surface and disorderly nanotubes, surface with orderly nanotubes array increased cell area and aspect ratio, as well as cell migration ability in the early phase of cell adhesion (p<0.05). Besides, compared with raw titanium surface, gene and protein expression of adhesion molecules were upregulated in nanotubes groups to different extents, no matter whether in an orderly or disorderly array. Within the limitations of our study, we conclude that compared with raw titanium surface, the presence of nanotubes array on titanium surface could enhance cells adhesion and cell migration in the early phase. And compared with disorderly distributed nanotubes, highly ordered nanotubes array might provide a much more favorable surface for gingival fibroblasts to achieve a tight adhesion on the materials.

Hydrophilic/Hydrophobic Janus Nanofibers Containing Compound K for Cartilage Regeneration.

Cartilage regeneration is a challenging issue due to poor regenerative properties of tissues. Electrospun nanofibers hold enormous potentials for treatments of cartilage defects. However, nanofibrous materials used for the treatment of cartilage defects often require physical and/or chemical modifications to promote the adhesion, proliferation, and differentiation of cells. Thus, it is highly desirable to improve their surface properties with functionality. We aim to design hydrophilic, adhesive, and compound K-loaded nanofibers for treatments of cartilage defects. Hydrophilic and adhesive compound K-containing polycaprolactone nanofibers (CK/PCL NFs) were prepared by coatings of gallic acid-conjugated chitosan (CHI-GA). Therapeutic effects of CHI-GA/CK/PCL NFs were assessed by the expression level of genes involved in the cartilage matrix degradation, inflammatory response, and lipid accumulations in the chondrocytes. In addition, Cartilage damage was evaluated by safranin O staining and immunohistochemistry of interleukin-1β (IL-1β) using OA animal models. To explore the pathway associated with therapeutic effects of CHI-GA/CK/PCL NFs, cell adhesion, phalloidin staining, and the expression level of integrins and peroxisome proliferator-activated receptor (PPARs) were evaluated. CHI-GA-coated side of the PCL NFs showed hydrophilic and adhesive properties, whereas the unmodified opposite side remained hydrophobic. The expression levels of genes involved in the degradation of the cartilage matrix, inflammation, and lipogenesis were decreased in CHI-GA/CK/PCL NFs owing to the release of CK. In vivo implantation of CHI-GA/CK/PCL NFs into the cartilage reduced cartilage degradation induced by destabilization of the medial meniscus (DMM) surgery. Furthermore, the accumulation of lipid deposition and expression levels of IL-1β was reduced through the upregulation of PPAR. CHI-GA/CK/PCL NFs were effective in the treatments of cartilage defects by inhibiting the expression levels of genes involved in cartilage degradation, inflammation, and lipogenesis as well as reducing lipid accumulation and the expression level of IL-1β via increasing PPAR.

Enhancing the angiogenesis and osteogenesis of low elastic modulus Ti-Nb-Zr-Sn alloy surface through grafting growth factors-containing gelatin nanoparticles

The study aimed to apply a multiple surface modification for low elastic modulus Ti-24 % Nb-4 % Zr-8 % Sn (Ti2448) alloy, to enhance its biological properties for bone implant applications. We employed a combined sandblasting/acid etching/alkaline etching process to modify the Ti2448 alloy surfaces with mixed nano/submicron/micron-scaled pits and super wettability. We prepared gelatin nanoparticles (GNPs) loaded with growth factors: vascular endothelial growth factor/platelet-derived growth factor (VEGF/PDGF) and/or bone morphogenetic protein-2 (BMP-2). The growth factors-containing GNPs were then sequentially grafted onto the surface-modified Ti2448 alloy in a layered (single layer: BMP-2 or VEGF/PDGF; double layer: inner BMP-2 and outer VEGF/PDGF) or mixed BMP-2/VEGF/PDGF manner. Surface characterizations, including morphology, roughness, surface energy, and cytotoxicity, of Ti2448 alloy were analyzed. For biological response, we investigated the tube formation ability of human umbilical vein endothelial cells (HUVECs) and the cell responses (adhesion, proliferation, and mineralization) of human bone marrow mesenchymal stem cells (hMSCs). The results demonstrated the successful preparation of GNPs with 200∼300 nm particle size and their successful loading with the growth factors using natural crosslinker genipin. The growth factor-containing GNPs were effectively grafted onto the surface-modified hydrophilic Ti2448 alloy without significantly reducing its wettability (water contact angle remained below 40°). The results of the study showed that the GNPs-grafted Ti2448 alloy surfaces, with the GNPs loaded with VEGF/PDGF and/or BMP-2, exhibited enhanced biological properties. The GNPs-grafted surfaces were found to be non-toxic and capable of promoting tube formation of HUVECs, particularly when single VEGF/PDGF layer or double (inner BMP-2/ outer VEGF/PDGF) layer was loaded on GNPs (enhancing the cumulative tube length approximately 1.4 times that of no GNPs-grafted surfaces). Moreover, the GNPs-grafted surfaces enhanced the adhesion-related proteins and extracellular matrix mineralization of hMSCs, particularly with the loading of single VEGF/PDGF or mixed VEGF/PDGF/BMP-2 on GNPs (enhancing αvβ3 integrin patch number > 3.5 times, vinculin patch number > 2 times, and mineralization approximately 1.4 times). Based on these findings, we concluded that the proposed Ti2448 alloy surfaces, when grafted with VEGF/PDGF- and/or BMP-2-containing GNPs, have the potential for bone implant applications. The surface modifications on the Ti2448 alloy may promote angiogenesis in the early stage of bone repair (through VEGF/PDGF) and osteogenesis in the late stage (through BMP-2), thereby accelerating the overall bone healing process.

Anatomical, pathological, and histological features of experimental respiratory infection of birds by biofilm-forming bacteria Staphylococcus aureus.

The pathogenesis of staphylococcal infections is mediated by virulence factors, such as enzymes, toxins, and biofilms, which increase the resistance of microorganisms to host immune system evasion. Testing and searching for standardized multi-level algorithms for the indication and differentiation of biofilms at the early stages of diagnosis will contribute to the development of preventive measures to control the critical points of technology and manage dangerous risk factors for the spread of infectious diseases. This research aimed to study the main stages of Staphylococcus aureus biofilm formation in in vitro experiments and to analyze the dynamics of respiratory syndrome development in chickens infected with these bacteria. Experimental reproduction of the infectious process was performed using laboratory models: 10-day-old White Leghorn chickens (n = 20). Before the experiments, the birds were divided into two groups according to the principle of analogs: Group I (control, n = 10): the birds were intranasally inoculated with 0.5 cm3 of 0.9% NaCl solution; Group II (experiment, n = 10): the birds were intranasally inoculated with a suspension of S. aureus bacteria, 0.5 cm3, concentration 1 billion/cm3. Colonization of individual areas of the substrate under study in vitro occurred gradually from the sedimentation and adhesion of single motile planktonic cells to the attachment stage of microcolony development. Staining preparations with gentian violet due to the "metachromosia" property of this dye are a quick and fairly simple way to differentiate cells and the intercellular matrix of biofilms. Fixation with vapors of glutaraldehyde and osmium tetroxide preserves the natural architecture of biofilms under optical and scanning electron microscopy. Pure cultures of S. aureus microorganisms were isolated from the blood, lungs, small intestine, liver, kidneys, and spleen after 5-10 days during experimental infection of chickens. Clinical signs of respiratory syndrome developed within 5-6 days after infection. Acute and subacute serous-fibrinous airsacculitis, characterized by edema and thickening of the membranes of the air sacs and the presence of turbid, watery, foamy contents in the cavity, was the most characteristic pathomorphological sign. The signs of acute congestive hyperemia and one-sided serous-fibrinous pneumonia developed with significant thickening of fibrinous deposits. In Garder's gland, there was an increase in the number of secretory sections, indicating hypersecretion of the glands. In the lymphoid follicles of Meckel's diverticulum, leukocytes, usually lymphocytes, and pseudoeosinophils were detected. Hydration and heteromorphism of the internal environment of biofilms determine the localization of differentiated cells in a three-dimensional matrix for protection against adverse factors. The most characteristic pathomorphological sign was the development of acute and subacute serous-fibrinous airsacculitis when reproducing the infectious process in susceptible models. There was a significant thickening of fibrinous deposits and signs of acute congestive hyperemia and one or two serous-fibrinous pneumonia developed.

Lentiviral expression of wild-type LAMA3A restores cell adhesion in airway basal cells from children with epidermolysis bullosa

The hallmark of epidermolysis bullosa (EB) is fragile attachment of epithelia due to genetic variants in cell adhesion genes. We describe 16 EB patients treated in the Ear, Nose and Throat department of a tertiary pediatric hospital linked to the United Kingdom’s National EB unit between 1992 and 2023. Patients suffered a high degree of morbidity and mortality from laryngotracheal stenosis. Variants in laminin subunit alpha-3 (LAMA3) were found in 10/15 patients where genotype was available. LAMA3 encodes a subunit of the laminin-332 heterotrimeric extracellular matrix protein complex and is expressed by airway epithelial basal stem cells. We investigated the benefit of restoring wildtype LAMA3 expression in primary EB patient-derived basal cell cultures. EB basal cells demonstrated weak adhesion to cell culture substrates, but could otherwise be expanded similarly to non-EB basal cells. In vitro lentiviral overexpression of LAMA3A in EB basal cells enabled them to differentiate in air-liquid interface cultures, producing cilia with normal ciliary beat frequency. Moreover, transduction restored cell adhesion to levels comparable to a non-EB donor culture. These data provide proof-of-concept for a combined cell and gene therapy approach to treat airway disease in LAMA3-affected EB.

Immobilization of hydroxyapatite on polyetherketoneketone surfaces for improved cell adhesion

A novel approach for the immobilization of hydroxyapatite (HAp) on polyetherketoneketone (PEKK) surface was suggested. HAp immobilization was performed in two steps. On the first step, PEKK surface was swelled in 1,1,1,3,3,3-hexafluoropropan-2-ol (HFIP) vapor. Then, the treated sample was placed in HAp water suspension. Up to 29 ± 3 % of the PEKK surface was coated with HAp. The maximum calcium and phosphorus content on the surface of the modified samples was found at 5.3 ± 0.1 and 0.5 ± 0.1 at. %, respectively. Despite the confirmed HAp immobilization, the hydrophobic character of the PEKK surfaces was preserved. HAp immobilization resulted in the enhanced adhesion of multipotent stem cells to the PEKK surfaces: the number of the adhered cells and cell-covered sample area increased nearly 10-fold and 25-fold, respectively. Thus, the suggested approach of HAp immobilization allows improving cell adhesion to PEKK-based surfaces.

Influence of Zika virus on the cytotoxicity, cell adhesion, apoptosis and inflammatory markers of glioblastoma cells.

Glioblastoma (GBM) is one of the most common types of brain tumor in adults. Despite the availability of treatments for this disease, GBM remains one of the most lethal and difficult types of tumors to treat, and thus, a majority of patients die within 2 years of diagnosis. Infection with Zika virus (ZIKV) inhibits cell proliferation and induces apoptosis, particularly in developing neuronal cells, and thus could potentially be considered an alternative for GBM treatment. In the present study, two GBM cell lines (U-138 and U-251) were infected with ZIKV at different multiplicities of infection (0.1, 0.01 and 0.001), and cell viability, migration, adhesion, induction of apoptosis, interleukin levels and CD14/CD73 cell surface marker expression were analyzed. The present study demonstrated that ZIKV infection promoted loss of cell viability and increased apoptosis in U-138 cells, as measured by MTT and triplex assay, respectively. Changes in cell migration, as determined by wound healing assay, were not observed; however, the GBM cell lines exhibited an increase in cell adhesion when compared with non-tumoral cells (Vero). The Luminex immunoassay showed a significant increase in the expression levels of IL-4 specifically in U-251 cells (MOI 0.001) following exposure to ZIKV. There was no significant change in the expression levels of IFN-γ upon ZIKV infection in the cell lines tested. Furthermore, a marked increase in the percentage of cells expressing the CD14 surface marker was observed in both GBM cell lines compared with in Vero cells; and significantly increased CD73 expression was observed particularly in U-251 cells, when compared with uninfected cells. These findings indicate that ZIKV infection could lead to reduced cell viability, elevated CD73 expression, improved cellular adherence, and higher rates of apoptosis in glioblastoma cells. Further studies are required to explore the potential use of ZIKV in the treatment of GBM.

Investigation of Physicochemical Characteristics of Aspergillus niger Biomass and Examination of Its Ability to Separate Butyl Acetate Isomers.

Aspergillus niger is a species of fungus that is widely found in natural ecosystems and has an important role in various industrial fields and is readily available. To study the adhesion of microbial cells to solid substrates and to improve their properties, physicochemical characterization of microorganisms is extremely important. For this purpose, in this study, the surface properties of A. niger biomass were determined at low cost and with high accuracy by inverse gas chromatography (IGC), a physicochemical characterization technique. IGC experiments were conducted between 303.2 and 328.2K at infinite dilution. Among these temperatures, various organic solvent vapors were passed over the A. niger biomass considered as stationary phase and their retention behavior was studied. Using the raw data, net retention volumes were calculated and retention diagrams were drawn. From the linear retention diagrams, the dispersive surface energy was calculated according to Dorris-Gray (48.73-46.09 mJ/m2), Donnet-Park (47.12-44.50 mJ/m2), Schultz (46.88-42.45 mJ/m2), and Hamieh (76.42-64.06 mJ/m2) methods. With the IGC method, the acidity-basicity parameters of A. niger biomass were determined and it was found that the surface was basic ( ). In the second part of this study, the butyl acetate isomer series, which are difficult to be separated by conventional methods, were effectively separated by the IGC method using A. niger stationary phase.

Instant mucus dressing of PEO reinforced by chitosan nanofiber scaffold for open wound healing

Dressings seamlessly attached to the open wound bed are necessary for fully unleashing the dressing healing ability, as leaving the voids beneath the dressing poses infection hazards. The present study prepared an instant mucus dressing (IMD) of polyethylene oxide (PEO) reinforced by chitosan (CS) nanofiber scaffold, which formed by immersing PEO/CS nanofiber mat in water. The PEO/CS nanofiber mat were fabricated by the solution blow spinning (SBS) method using PEO and CS mixed solutions. Attenuated total reflection Fourier transform infrared spectroscopy (FTIR-ATR), transmission electron microscope (TEM), X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD) and differential scan calorimetry (DSC) analyses indicate that PEO macromolecules formed the most of nanofiber shell due to their lower surface tension while CS macromolecules stayed mainly inside the fiber as the core. When such a PEO/CS nanofiber mat was immersed in water, PEO swelled to form mucus dressing reinforced by CS inside the nanofiber, which was fluidic and able to fully fill the voids on the wound. In vivo rat experiment verified that the dressing significantly accelerated the open wound healing through seamlessly attaching of mucus to the open wound and providing moist environment. The dressings exhibit good platelets and whole blood cells adhesion properties, excellent hemostasis function and no cytotoxicity. This instant mucus dressing provided a new perspective for manufacturing high performance open wound dressings.

Artificial Trachea from Microtissue Engineering and Three-Dimensional Printing for Tracheal Personalized Repair.

Millions of people suffer from tracheal defect worldwide each year, while autograft and allograft cannot meet existing treatment needs. Tissue-engineered trachea substitutes represent a promising treatment for tracheal defect, while lack of precisely personalized treatment abilities. Therefore, development of an artificial trachea that can be used for personalized transplantation is highly desired. In this study, we report the design and fabrication of an artificial trachea based on sericin microsphere (SM) by microtissue engineering technology and three-dimensional (3D) printing for personalized repair of tracheal defect. The SM possessed natural cell adhesion and promoting cell proliferation ability. Then, the microtissue was fabricated by coincubation of SM with chondrocytes and tracheal epithelial cells. This microtissue displayed good cytocompatibility and could support seed cell adhesion and proliferation. After that, this microtissue was individually assembled to form an artificial trachea by 3D printing. Notably, artificial trachea had an encouraging complete cartilaginous and epithelial structure after transplantation. Furthermore, the artificial trachea molecularly resembled native trachea as evidenced by similar expression of trachea-critical genes. Altogether, the work demonstrates the effectiveness of microtissue engineering and 3D printing for individual construction of artificial trachea, providing a promising approach for personalized treatment of tracheal defect.

Ionic Liquid Interface as a Cell Scaffold.

In sharp contrast to conventional solid/hydrogel platforms, water-immiscible liquids, such as perfluorocarbons and silicones, allow the adhesion of mammalian cells via protein nanolayers (PNLs) formed at the interface. However, fluorocarbons and silicones, which are typically used for liquid cell culture, possess only narrow ranges of physicochemical parameters and have not allowed for a wide variety of cell culturing environments. In this paper, it is proposed that water-immiscible ionic liquids (ILs) are a new family of liquid substrates with tunable physicochemical properties and high solvation capabilities. Tetraalkylphosphonium-based ILs are identified as non-cytotoxic ILs, whereon human mesenchymal stem cells are successfully cultured. By reducing the cation charge distribution, or ionicity, via alkyl chain elongation, the interface allows cell spreading with matured focal contacts. High-speed atomic force microscopy observations of the PNL formation process suggest that the cation charge distribution significantly altered the protein adsorption dynamics, which are associated with the degree of protein denaturation and the PNL mechanics. Moreover, by exploiting dissolution capability of ILs, an ion-gel cell scaffold is fabricated. This enables to further identify the significant contribution of bulk subphase mechanics to cellular mechanosensing in liquid-based culture scaffolds.

Characterization and In Vitro Evaluation of Porous Polymer-Blended Scaffolds Functionalized with Tricalcium Phosphate.

Bone tissue is one of the most transplanted tissues. The ageing population and bone diseases are the main causes of the growing need for novel treatments offered by bone tissue engineering. Three-dimensional (3D) scaffolds, as artificial structures that fulfil certain characteristics, can be used as a temporary matrix for bone regeneration. In this study, we aimed to fabricate 3D porous polymer scaffolds functionalized with tricalcium phosphate (TCP) particles for applications in bone tissue regeneration. Different combinations of poly(lactic acid) (PLA), poly(ethylene glycol) (PEG with molecular weight of 600 or 2000 Da) and poly(ε-caprolactone) (PCL) with TCP were blended by a gel-casting method combined with rapid heating. Porous composite scaffolds with pore sizes from 100 to 1500 µm were obtained. ATR-FTIR, DSC, and wettability tests were performed to study scaffold composition, thermal properties, and hydrophilicity, respectively. The samples were observed with the use of optical and scanning electron microscopes. The addition of PCL to PLA increased the hydrophobicity of the composite scaffolds and reduced their susceptibility to degradation, whereas the addition of PEG increased the hydrophilicity and degradation rates but concomitantly resulted in enhanced creation of rounded mineral deposits. The scaffolds were not cytotoxic according to an indirect test in L929 fibroblasts, and they supported adhesion and growth of MG-63 cells when cultured in direct contact.

Multifunctional polyvinyl alcohol/gallic acid functionalized chitosan hydrogels prepared by freeze-thaw method for potential application as wound dressings

The design of multifunctional hydrogel dressings to effectively promote wound healing is highly desirable in clinical trauma and surgery. Inspired by excellent mechanical properties of polyvinyl alcohol (PVA) and strong adhesion of polyphenols, gallic acid was grafted onto chitosan molecular chain (CS-GA) by free radical polymerization in this study, and PVA/CS-GA hydrogels were successfully prepared by repeated freeze-thaw method, without introducing toxic crosslinking agents. The prepared PVA/CS-GA hydrogels could adhere to various substrates, which confirmed excellent universal adhesion properties. The red blood cells adhesion ratio was 67.1%, indicating its potential for promoting wound coagulation. Meanwhile, the mechanical strength of PVA/CS-GA hydrogel was significantly enhanced, exhibiting approximately twice tensile strength and failure stretch compared with PVA hydrogel. Furthermore, attributing to GA molecule, PVA/CS-GA hydrogels not only showed favorable antibacterial activity against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus), but also could scavenge free radicals, showing effective antioxidant activity. The hydrogels also had good biocompatibility, met the basic requirements of biomedical materials and may promote wound repair. It should be noted that the content of GA had significant impact on cytotoxicity. Therefore, multifunctional PVA/CS-GA hydrogels with excellent mechanical, adhesion, antioxidant, antibacterial and biocompatible properties show promising potential applications in the biomedicine field, especially as functional wound dressings.

Preparation and characterization of carboxymethyl cellulose/polyethylene glycol films containing bromelain/curcumin: In vitro evaluation of wound healing activity

AbstractIn this study, a wound‐healing membrane was fabricated based on carboxymethyl cellulose (CMC) and polyethylene glycol (PEG) containing curcumin (Cur) and bromelain (Br). Citric acid (CA) was used as a cross‐linking agent. This membrane showed an ideal degree of swelling, which was significantly dependent on the concentration and duration of cross‐linked CA. The chemical characterization showed the CA cross‐linker mechanism was more associated with chemical reactions with CMC carboxyl groups, and PEG hydroxyl groups played an important role in forming a hybrid polymer network. It greatly enhanced mechanical and adhesive properties, so the stress strength was improved from 29.4 to 38.52 Mpa. The hydrophilicity of the membrane surface according to the water contact angle assay showed the membrane surface is suitable for adhesion, growth, migration, and proliferation of skin cells. The drug delivery assay demonstrated that the Br and Cur were released during 48 h, but the Br followed the burst release in comparison with Cur. Antibacterial properties showed that CMC/PEG‐Cur/Br have ideal antibacterial properties for preventing the growth of bacteria. In summary, the engineered CMC/PEG containing Cur/Br films with desired cell viability properties and antibacterial activity can potentially improve and accelerate skin regeneration for chronic wound healing.Highlights Hydrogel film was prepared based on carboxymethyl cellulose (CMC), polyethylene glycol (PEG), and citric acid as cross‐linking agent. CMC/PEG film were induced controlling curcumin (Cur) and bromelain (Br) release for wound healing process. CMC/PEG film revealed a wide range of physiochemical, mechanical, adhesion, and biological properties behavior. Incorporation of Cur and Br promoted in vitro biocompatibility, L‐929 cells attachments, and cell migration.

The influence of Ras-associated binding protein 23 knockdown on the migration and invasion of esophageal squamous cell carcinoma cells and its mechanism

Objective: To investigate the role and the mechanism of Ras-associated binding protein23 (RAB23) in the migration and invasion of esophageal squamous cell carcinoma (ESCC) cells. Methods: RAB23 mRNA levels were measured in 16 pairs of ESCC and adjacent normal tissues via real-time polymerase chain reactions. RAB23 mRNA levels in the ESCC and adjacent normal tissues of dataset GSE20347 deposited in the Gene Expression Omnibus (GEO) database were also analyzed. Immunohistochemistry (IHC) was used to detect the RAB23 protein expressions in 106 pairs of ESCC and adjacent normal tissues, as well as in the lymph glands and primary tumor tissues of 33 patients with positive lymph nodes and 10 patients with negative lymph nodes. Endogenous RAB23 expression was transiently depleted using siRNAs (si-NC, si-RAB23-1, and si-RAB23-9) or stably reduced using shRNAs (sh-NC and sh-RAB23) in ESCC KYSE30 and KYSE150 cells, and the knockdown efficiency was tested using Western blot assays. Cell counting kit-8 assays and mouse xenograft models were used to test the proliferation of ESCC cells. Transwell assays and tail vein-pulmonary metastasis models in immunocompromised mice were used to examine the migration and invasion of ESCC cells. Cell adhesion assays were used to test the adhesion of ESCC cells. RNA-seq assays were used to analyze how RAB23 knockdown influenced the expression profile of ESCC cells and the implicated signal pathways were confirmed using Western blot assays. Results: The RAB23 mRNA expression in 16 cases of ESCC tissues was 0.009 7±0.008 9, which was markedly higher than that in adjacent normal tissues (0.003 2±0.003 7, P=0.006). GEO analysis on RAB23 expressions in ESCC and adjacent normal tissues showed that the RAB23 mRNA level in ESCC tissues (4.30±0.25) was remarkably increased compared with their normal counterparts (4.10±0.17, P=0.037). Among the 106 pairs of ESCC and tumor-adjacent normal tissues, 51 cases exhibited low expression of RAB23 and 55 cases showed high expression of RAB23, whereas in the paired tumor-adjacent normal tissues 82 cases were stained weakly and 24 strongly for RAB23 protein. These results indicated that RAB23 expression was markedly increased in ESCC tissues (P<0.001). Additionally, only 1 out of 33 primary ESCC tissues with positive lymph nodes showed low RAB23 protein expression. On the other hand, 7 samples of primary ESCC tissues with negative lymph nodes were stained strongly for RAB23 while its level in the other 3 samples was weak. These results showed that RAB23 expression was remarkably increased in primary ESCC tissues with positive lymph nodes compared with those with negative lymph nodes (P=0.024). Further tests showed that 32 out of 33 positive lymph nodes were stained strongly for RAB23, whereas no negative lymph nodes (n=10) exhibited high expression of RAB23 (P<0.001). Both transient and stable knockdown of endogenous RAB23 expression failed to cause detectable changes in the proliferation of KYSE30 cells in vitro and in vivo, but attenuated the migration and invasion of KYSE30 cells as well as the invasion of KYSE150 cells. RAB23 knockdown was found to significantly decrease the number of adhesive KYSE30 cells in the sh-RAB23 group (313.75±89.34) compared with control cells in the sh-NC group (1 030.75±134.29, P<0.001). RAB23 knockdown was also found to significantly decrease the number of adhesive KYSE150 cells in the sh-RAB23 group (710.5±31.74) compared with the number of control cells in the sh-NC group (1 005.75±61.09, P<0.001). RNA-seq assays demonstrated that RAB23 knockdown using two siRNAs targeting RAB23 mRNA markedly impaired focal adhesion-related signal pathways, and decreased the levels of phosphorylated FAK (p-FAK) and phosphorylated paxillin (p-paxillin) in KYSE30 and KYSE150 cells. Conclusions: Significantly increased RAB23 in ESCC tissues positively correlates with lymph node metastasis. Depleted RAB23 expression attenuates focal adhesion-related signal pathways, thus impairing the invasion, metastasis, and adhesion of ESCC cells.

Sulforaphane Inhibits Adhesion and Migration of Cisplatin- and Gemcitabine-Resistant Bladder Cancer Cells In Vitro.

Only 20% of patients with muscle-invasive bladder carcinoma respond to cisplatin-based chemotherapy. Since the natural phytochemical sulforaphane (SFN) exhibits antitumor properties, its influence on the adhesive and migratory properties of cisplatin- and gemcitabine-sensitive and cisplatin- and gemcitabine-resistant RT4, RT112, T24, and TCCSUP bladder cancer cells was evaluated. Mechanisms behind the SFN influence were explored by assessing levels of the integrin adhesion receptors β1 (total and activated) and β4 and their functional relevance. To evaluate cell differentiation processes, E- and N-cadherin, vimentin and cytokeratin (CK) 8/18 expression were examined. SFN down-regulated bladder cancer cell adhesion with cell line and resistance-specific differences. Different responses to SFN were reflected in integrin expression that depended on the cell line and presence of resistance. Chemotactic movement of RT112, T24, and TCCSUP (RT4 did not migrate) was markedly blocked by SFN in both chemo-sensitive and chemo-resistant cells. Integrin-blocking studies indicated β1 and β4 as chemotaxis regulators. N-cadherin was diminished by SFN, particularly in sensitive and resistant T24 and RT112 cells, whereas E-cadherin was increased in RT112 cells (not detectable in RT4 and TCCSup cells). Alterations in vimentin and CK8/18 were also apparent, though not the same in all cell lines. SFN exposure resulted in translocation of E-cadherin (RT112), N-cadherin (RT112, T24), and vimentin (T24). SFN down-regulated adhesion and migration in chemo-sensitive and chemo-resistant bladder cancer cells by acting on integrin β1 and β4 expression and inducing the mesenchymal-epithelial translocation of cadherins and vimentin. SFN does, therefore, possess potential to improve bladder cancer therapy.

An imprint-based approach to replicate nano- to microscale roughness on gelatin hydrogel scaffolds: surface characterization and effect on endothelialization

Biologization of biomaterials with endothelial cells (ECs) is an important step in vascular tissue engineering, aiming at improving hemocompatibility and diminishing the thrombo-inflammatory response of implants. Since subcellular topography in the scale of nano to micrometers can influence cellular adhesion, proliferation, and differentiation, we here investigate the effect of surface roughness on the endothelialization of gelatin hydrogel scaffolds. Considering the micron and sub-micron features of the different native tissues underlying the endothelium in the body, we carried out a biomimetic approach to replicate the surface roughness of tissues and analyzed how this impacted the adhesion and proliferation of human umbilical endothelial cells (HUVECs). Using an imprinting technique, nano and micro-roughness ranging from Sa= 402 nm to Sa= 8 μm were replicated on the surface of gelatin hydrogels. Fluorescent imaging of HUVECs on consecutive days after seeding revealed that microscale topographies negatively affect cell spreading and proliferation. By contrast, nanoscale roughnesses of Sa= 402 and Sa= 538 nm promoted endothelialization as evidenced by the formation of confluent cell monolayers with prominent VE-cadherin surface expression. Collectively, we present an affordable and flexible imprinting method to replicate surface characteristics of tissues on hydrogels and demonstrate how nanoscale roughness positively supports their endothelialization.

Preparation of bilayer tissue-engineered polyurethane/poly-L-lactic acid nerve conduits and their in vitro characterization for use in peripheral nerve regeneration

BackgroundDue to loss of peripheral nerve structure and/or function resulting from trauma, accidents, and other causes, peripheral nerve injuries continue to be a major clinical problem. These injuries can cause partial or total loss of sensory, motor, and autonomic capabilities as well as neuropathic pain. PNI affects between 13 and 23 out of every 100,000 people annually in developed countries. Regeneration of damaged nerves and restoration of function after peripheral nerve injury remain significant therapeutic challenges. Although autologous nerve graft transplantation is a viable therapy option in several clinical conditions, donor site morbidity and a lack of donor tissue often hinder full functional recovery. Biomimetic conduits used in tissue engineering to encourage and direct peripheral nerve regeneration by providing a suitable microenvironment for nerve ingrowth are only one example of the cutting-edge methods made possible by this field. Many innate extracellular matrix (ECM) structures of different tissues can be successfully mimicked by nanofibrous scaffolds. Nanofibrous scaffolds can closely mimic the surface structure and morphology of native ECMs of many tissues.MethodsIn this study, we have produced bilayer nanofibrous nerve conduit based on poly-lactic acid/polyurethane/multiwall carbon nanotube (PLA/PU/MWCNT), for application as composite scaffolds for static nerve tissue engineering. The contact angle was indicated to show the hydrophilicity properties of electrospun nanofibers. The SEM images were analyzed to determine the fiber’s diameters, scaffold morphology, and endometrial stem cell adhesion. Moreover, MTT assay and DAPI staining were used to show the viability and proliferation of endometrial stem cells.ResultsThe constructed bilayer PLA/PU/MWCNT scaffolds demonstrated the capacity to support cell attachment, and the vitality of samples was assessed using SEM, MTT assay, and DAPI staining technique.ConclusionsAccording to an in vitro study, electrospun bilayer PLA/PU/MWCNT scaffolds can encourage the adhesion and proliferation of human endometrial stem cells (hEnSCs) and create the ideal environment for increasing cell survival.

A Cu(Ⅱ)-eluting coating through silk fibroin film on ZE21B alloy designed for in situ endotheliazation biofunction

In the cardiovascular field, coating containing copper used to catalyze NO (nitric oxide) production on non-degradable metal surfaces have shown unparalleled expected performance, but there are few studies on biodegradable metal surfaces. Magnesium-based biodegradable metals have been applied in cardiovascular field in large-scale because of their excellent properties. In this study, the coating of copper loaded in silk fibroin is fabricated on biodegradable ZE21B alloy. Importantly, the different content of copper is set to investigate the effects of on the degradation performance and cell behavior of magnesium alloy. Through electrochemical and immersion experiments, it is found that high content of copper will accelerate the corrosion of magnesium alloy. The reason is the spontaneous micro-batteries between copper and magnesium with the different standard electrode potentials, that is, the galvanic corrosion accelerates the corrosion of magnesium alloy. Moreover, the coating formed through silk fibroin by the right amount copper not only have a protective effect on the ZE21B alloy substrate, but also promotes the adhesion and proliferation of endothelial cells in blood vessel micro-environment. The production of NO catalyzed by copper ions makes this trend more significant, and inhibits the excessive proliferation of smooth muscle cells. These findings can provide guidance for the amount of copper in the coating on the surface of biodegradable magnesium alloy used for cardiovascular stent purpose.