Fibroblast Cell Line L929 Research Articles

In vitro cellular viability studies on a concentrated surfactant-based wound dressing.

In this study, three cellular cytotoxic assays (direct contact assay, extraction assay, and cell insert assay) were applied to evaluate the effects of a concentrated surfactant gel preserved with antimicrobials and a concentrated surfactant gel with 1% silver sulfadiazine on both the mouse fibroblast cell line L929 and human dermal fibroblasts (HDFa). Also, the in vitro wound model was wounded by a 100 μL pipette tip and used to assess cell migration and wound closure after treatment with both gels. A needle-scratched membrane disruption model was used to preliminarily evaluate membrane stabilisation and the membrane-resealing effects of concentrated surfactant gels. It was demonstrated that the concentrated surfactant gel preserved with antimicrobials was not toxic to both L929 and HDFa. However, the concentrated surfactant gel with 1% silver sulfadiazine demonstrated a degree of cytotoxicity to both cell types. After treatment with a concentrated surfactant gel preserved with antimicrobials, cell movement to close the scratch gap was enhanced at 24 and 48 hours. The results also showed that cells treated with the concentrated surfactant gel preserved with antimicrobials decreased cell necrosis and improved cell resistance of the f-actin rearrangement after a needle scratch. The results demonstrated that a concentrated surfactant gel preserved with antimicrobials is non-cytotoxic and has ability to accelerate wound closure by enhancing cell mobility. Furthermore, the concentrated surfactant gel appeared to stabilise the plasma membrane and demonstrated a resealing ability and helped to retain the plasma membrane integrity and enhanced wound healing.

Bioactive agar-based functional composite film incorporated with copper sulfide nanoparticles

Copper sulfide nanoparticles (CuS NP) were prepared using copper acetate and thiourea as a source of Cu2+ and S2−, respectively, cornstarch as a stabilizing agent and ammonia as a hydrolyzing agent under a mild condition of sulfidation (70 °C for 1 h). The present work describes the synthesis and preparation of bioactive agar/CuS NP nanocomposite film. CuS NP was characterized using UV-vis spectroscopy, X-ray diffraction pattern, scanning electron microscopy (SEM) and transmission electron microscopy (TEM). CuS NP was roughly spherical and irregular with a size in the range of 2–10 nm with an average of 4.6 ± 1.3 nm. Agar-based films with different concentrations (0.25, 0.50, 1.0 and 2.0 wt%) of CuS NP were prepared and characterized using SEM and Fourier transforms infrared spectroscopy (FTIR), and the film properties such as UV-barrier, mechanical, water vapor barrier, swelling ratio, water solubility, hydrophobicity, thermal stability, antibacterial properties and cytotoxicity were investigated. The CuS NP was well dispersed in the polymer matrix to form compatible nanocomposite films. The swelling ratio and moisture content of the composite films decreased while the water solubility increased slightly after the addition of CuS NP. The composite films showed significantly increased UV-barrier without much sacrifice of transparency, and they also showed increased mechanical strength and water vapor barrier properties. Also, in vitro analysis showed excellent biocompatibility of CuS NP and nanocomposite films on skin fibroblast L929 cell lines with cell viability above 90%. Also, they exhibited distinctive antibacterial activity against food-borne pathogenic bacteria, E. coli and some activity against L. monocytogenes.

Biological evaluation of bioavailable amphiphilic polymeric conjugate based-on natural products: diosgenin and curcumin

The aim of this study was to design and synthesize amphiphilic supramolecular structures consisting of curcumin and diosgenin-terminated poly (ethylene glycol). Diosgenin, a natural plant extract, has proven to possess significant biological activities such as anti-tumor, anti-diabetes, anti-inflammatory, anti-cancer, and effect on improving memory in Alzheimer’s disease. While curcumin, the main compound of the spice turmeric, is one of the natural products that exhibits a wide range of pharmacological activities. But poor water solubility and low bioavailability of these materials have limited their clinical applications. Herein, novel amphiphilic conjugate poly (ethylene glycol) diosgenin-curcumin with improved solubility and bioavailability were developed. Cytotoxicity/anticancer effects on human fibroblast cells and human breast cancer cells are investigated with prospective potential applications in cancer therapeutics. The synthesized polymeric conjugate demonstrated non-toxic to L929 (fibroblast) cell lines but highly toxic to MCF-7 breast cancer cell lines.

Ciprofloxacin‐Loaded Ceramic/Polymer Composite Coatings on Ti with Improved Antibacterial and Corrosion Resistance Properties for Orthopedic Applications

AbstractPast few decades witnessed the upcoming of osteomyelitis which is a challenging infectious bone disease. Complications related to this infection is encountered by local drug delivery via orthopedic surgery. In this work, TiO2‐SiO2 based bioceramics coatings were fabricated on Ti alloy by anodization method using sulphuric acid and Sodium silicofluoride (SSF) as an electrolyte. Plasma polymerized ethylenediamine (PPEDA) thin film deposited on composites by inductively coupled plasma chemical vapor deposition and followed by ciprofloxacin were used as a model drug by electrodeposition method. The resultant coatings were characterized by FTIR, XRD, FESEM, EDAX and AFM techniques. The mechanical behavior of the composite coatings was evaluated using thickness and hardness tester and electrochemical properties were studied in Ringer's solution. Similarly, CIP@PPEDA/TiO2‐SiO2 composite coating on Ti alloy shows good corrosion resistance and better antibacterial efficacy as well as L929 fibroblast cell line, which exhibited a high cell adhesion and high cell viability compared than TiO2‐SiO2 and PPEDA/TiO2‐SiO2 coatings. All these results suggest that, this newly developed composite coating exhibits superior bioactivity and improved anticorrosion performance on Ti specimen and can be a potential candidate for orthopedic applications.

Wound-healing potential of methanolic extract of Rhaphidophora korthalsii leaves possibly mediated by collagen and fibronectin expression in L929 cell line

Background: Rhaphidophora korthalsii is a climber shrub well known for its therapeutic properties in traditional medicine. The immunomodulatory properties of the plant extract have been investigated on mice splenocytes, human peripheral blood, and human peripheral blood mononuclear cell natural killer cells. The plant extract has also shown antioxidant effect and elevated macrophage production and has also been known to have anticancer properties. Yet, its wound-healing ability is not explored. Aims and Objectives: The objective of this research study is to evaluate the wound-healing potential of the plant extract on mouse fibroblast cell line L929 through scratch assay and the potential effect of the plant extract on the expression of two key wound-healing factors, fibronectin and collagen 1. Materials and Methods: L929 was exposed to various concentrations of R. korthalsii methanol extract to check the cytotoxic effect of the extract. In vitro scratch assay was performed on the treated cells to assess the wound-healing ability of the extract. Further, the effect of the extract on the expression of wound-healing factors, fibronectin and collagen 1, was studied through a flow cytometric analysis. Results: R. korthalsii leaf extract showed no toxicity on the L929 fibroblast cells. The treated cells showed ~93% wound closure after 48 h of incubation. The treatment of cells with the leaf extract, also, upregulated the expression of the wound-healing factors, fibronectin and collagen 1, i.e., 76.30% and 96.09% of the cells expressed them, respectively. Conclusion: R. korthalsii is a non-toxic plant with wound-healing properties. The phytochemicals in this plant may be responsible for its abilities. Thus, it can serve as an alternative therapy against synthetic drugs for wound healing.

Facile synthetic Photoluminescent Graphene Quantum dots encapsulated β-cyclodextrin drug carrier system for the management of macular degeneration: Detailed analytical and biological investigations

Drug administration by effective nano-carriers is an emerging and growing technology in the field of bio-medicine and particularly Age –related macular degeneration (AMD). This developed nanomaterials based methods with drug administration maximizes the biocompatibility and systemically increases drug delivery profile for the drugs. Herein, we described the effective drug molecules delivery profiles by the hydrothermally synthesized graphene quantum dots (GQDs) encapsulated with supramolecular β-cyclodextrin (β-CD) as a drug delivery system for AMD. The drug release profiles were analysed and plotted by two different types of drugs ((Bevacizumab (Bev) and Ranibizumab (Ran))) and compounds displayed an initial burst delivery percentage of 55.7 ± 1.6% and 52.2 ± 2.6, respectively, within 15 min. After 1 h, 94.2% (Ran) and 93.1% (Bev) of loaded drug molecules were released from the β-CD encapsulated GQDs in sustained manner. The biocompatibility of the synthesized carriers was investigated quantitatively and qualitatively with the mouse Fibroblast L929 cell line. The biological cell analysis observed by calculated cell count and green fluorescence visualization has been clearly confirmed the samples are non-toxic and highly compatible to the cells with more than 90% cell viability after 5 days cell culture. The observed material properties and biological results demonstrated that the suitability of the developed nano-carriers for the drug delivery system in the AMD.

Antitumor effect of chondroitin AC lyase (PsPL8A) from Pedobacter saltans on melanoma and fibrosarcoma cell lines by in vitro analysis.

PGs are involved in cellular communication and cancer biology. The role of CS in melanoma and fibrosarcoma cell lines was explored by using chondroitin AC lyase (PsPL8A). The proliferation of mouse fibroblast L929, human melanoma (SK-Mel 28) and fibrosarcoma (HT-1080) cell lines after treatment with chondroitin AC lyase (PsPL8A) was studied by MTT assay. The mode of cell death was studied by Annexin-V FITC using flow cytometry and fluorescence microscopy. The alteration in mitochondrial cell potential was studied by JC-1 dye using fluorescence microscopy and flow cytometry. Treatment of L929 cells with PsPL8A imparts no cytotoxicity and showed no alteration in proliferation with nearly 95-98% cell viability. An overall 58% and 59% inhibition of SK-Mel 28 and HT-1080 cell proliferation was observed with 1.3 μM of PsPL8A after 24 h of incubation. The PsPL8A (1.3 μM) treated SK-Mel 28 and HT-1080 cells showed significant green fluorescence with annexin-V FITC under fluorescence microscopy and 56.6% and 35.5% apoptosis, respectively by flow cytometry analysis. The results of fluorescence microscopy and flow cytometry of SK-Mel 28 and HT-1080 upon treatment with PsPL8A (1.3 μM) for 24 h, gave green fluorescence due to dissipation of mitochondrial potential with JC-1 dye. Chondroitin AC lyase (PsPL8A) displayed anti-tumor potential against human melanoma SK-Mel 28 and fibrosarcoma HT-1080 cell lines, while the mouse fibroblast L929 cells were unaffected.

Cefuroxime conjugated chitosan hydrogel for treatment of wound infections.

Hydrogels are excellent drug delivery systems for the treatment of chronic wound infections. However, the problem of high burst release still remains a challenge that needs to be tackled. In terms of antibiotic release from the hydrogels, as the drug payload depletes it could act as a substrate for bacterial seeding which can create a life-threatening condition. Therefore, to provide the sustained effect of an antibiotic at the localized site via hydrogel matrix, we prepared a chitosan (CS) hydrogel system in which cefuroxime (CEF) is covalently conjugated with chitosan polymer via an ester linkage. To prepare the cefuroxime conjugated chitosan hydrogel, the formulations were optimized using different concentrations of cefuroxime, 0% (CS/CEF_0), 5% (CS/CEF_5), 10% (CS/CEF_10) and 20% (CS/CEF_20) w/w of chitosan. Fourier Transform Infra-red Spectroscopy (FTIR) confirmed the conjugation of cefuroxime and chitosan. The drug release studies showed that the release of cefuroxime was higher in the phosphate buffer (pH 7.4) with esterase enzyme and alkaline medium (pH 10) compared to phosphate buffer (pH 7.4) alone. Hemolysis assay was performed to demonstrate the hemocompatibility of the prepared hydrogel samples. The cell viability study using the L929 fibroblast and MG63 osteosarcoma cell lines revealed that synthesized hydrogel is biocompatible. Furthermore, a potent antibacterial activity for the extended time period proved the biological efficacy of a hydrolyzed cefuroxime. Thus, CS/CEF_5, CS/CEF_10, and CS/CEF_20 hydrogels have a promising future in the treatment of chronic wound infections.

Chitosan/poly(ethylene glycol)/hyaluronic acid biocompatible patches obtained by electrospraying

Electrospray is a promising technique to scale-up production of microparticles and nanoparticles. In this study, electrospraying was used in order to produce candidate biopatches (CPH) by using chitosan, poly(ethylene glycol) (PEG) and hyaluronic acid (HA). Four different ratios of polymer blend compositions (CPH1, CPH2, CPH3 and CPH4) were tested by dissolving in 2% acetic acid solution (Ac.A.). The HA amount in each blend was kept the same to designate the optimum surface with different chitosan/PEG ratios for electrospray process. Fourier-transform infrared (FTIR) microscopy, scanning electron microscopy (SEM), and atomic force microscopy (AFM) studies showed that obtained patches had highly adhesive surfaces with the aid of heterogeneously distributed micro- and nano-particles. Additionally, video images of FTIR microscopy and AFM images proved that all surfaces have similar heterogeneity except CPH2. The most homogenous surface was obtained by CPH3. Patches were directly subjected to antibacterial tests against ten different types of gram positive and gram negative bacteria using disc diffusion assay (Kirby–Bauer method). Extraordinarily there was no antibacterial property of patches coated with microparticles. Finally, biocompatibility studies were performed by using mouse fibroblast L929 cell lines (ATTC number CCL-1) to test cell adhesion and proliferation properties of the patches. Results of 72 h viability tests proved the electrospray of ternary blends had displayed good biocompatibility; in particular, CPH3 had the highest cell viability.

Potential bactericidal activity of S. nux-vomica–ZnO nanocomposite against multidrug-resistant bacterial pathogens and wound-healing properties

Multidrug resistance in bacterial strains has become the greatest challenge for healthcare professionals for treating non-healing ulcers such as diabetic foot infections (DFI). Plant-mediated synthesis of S. nux-vomica–ZnO nanocomposite appears as a potential new alternative therapeutic agent that might be capable of tackling antibiotic-resistant bacterial pathogens and for treating a non-healing ulcer. The aim of the study was to investigate the antibacterial potential of S. nux-vomica–ZnO nanocomposite biosynthesised from Strychnos nux-vomica against multidrug-resistant organisms (MDROs) from DFU, wound-healing properties, and cytotoxic effects. The antibacterial potential was assessed by minimum inhibitory concentration (MIC)/ minimum bactericidal concentration (MBC) assays, time-kill kinetics, protein-leakage, and flow cytometric analysis. The wound-healing properties were assessed by scratch assay on mouse L929 fibroblastic cell line to quantify cell migration towards the injured area. Cytotoxicity was assessed using 3-[4,5-dimethyl-2-thiazol-yl]-2,5-diphenyl- 2H-tetrazolium bromide (MTT) cellular viability assay on the L929 cell line and human embryonic kidney epithelial (HEK-293) cell line. Strychnos nux-vomica–ZnO nanocomposite at a size range of 10–12 nm exhibited significant bactericidal potency at a concentration of 100–200 μg/ml against MDR–Methicillin-resistant Staphylococcus aureus, MDR–Escherichia coli, MDR–Pseudomonas aeruginosa, MDR–Acinetobacter baumannii, and also against standard bacterial strains S. aureus ATCC 29213, E. coli ATCC 25922, P. aeruginosa ATCC 27853, E. faecalis ATCC 29212. S. nux-vomica–ZnO nanocomposite also exhibited wound-healing and reduced cytotoxic properties at the antimicrobially active concentrations. Our findings thus suggested remarkable bactericidal properties of S. nux-vomica–ZnO nanocomposite and can be further exploited towards for the development of an antibacterial agent against the threatening superbugs

Lanyamycin, a macrolide antibiotic from Sorangium cellulosum, strain Soce 481 (Myxobacteria).

Lanyamycin (1/2), a secondary metabolite occurring as two epimers, was isolated from the myxobacterium Sorangium cellulosum, strain Soce 481. The structures of both epimers were elucidated from HRESIMS and 1D and 2D NMR data and the relative configuration of their macrolactone ring was assigned based on NOE and vicinal 1H NMR coupling constants and by calculation of a 3D model. Lanyamycin inhibited HCV infection into mammalian liver cells with an IC50 value of 11.8 µM, and exhibited a moderate cytotoxic activity against the mouse fibroblast cell line L929 and the human nasopharyngeal cell line KB3 with IC50 values of 3.1 and 1.5 μM, respectively, and also suppressed the growth of the Gram-positive bacterium Micrococcus luteus.

Development of Amoxicillin-Loaded Electrospun Polyurethane/Chitosan/ $\beta$ -Tricalcium Phosphate Scaffold for Bone Tissue Regeneration.

Biocompatible nanocomposite electrospun fibers containing Polyurethane/Chitosan/ $\beta $ -Tri calcium phosphate with diverse concentrations were designed and produced through the electrospinning process for bone tissue engineering applications. After the production process, density measurement, viscosity, electrical conductivity, and tensile strength measurement tests were carried out as physical analyses of blended solutions. The chemical structural characterization was scrutinized using Fourier transform infrared spectrometer (FTIR), and scanning electron microscopy (SEM) was used to observe the morphological details of developed electrospun scaffolds. Cell viability, attachment, and proliferation were performed using a L929 fibroblast cell line. Based on the physical, SEM, FTIR analysis, and cell culture studies, preferable nanofiber composition was selected for further studies. Amoxicillin (AMX) was loaded to that selected nanofiber composition for examination of the drug release. In comparison with other studies on similar AMX controlled products, higher drug loading and encapsulation efficiencies were obtained. It has been clearly found that the developed nanofiber composites have potential for bone tissue engineering applications.

Chitosan/polyethylene glycol fumarate blend films for wound dressing application: in vitro biocompatibility and biodegradability assays

Blending is one of the effective approaches in preparing tailored materials with a wide range of properties. Thus, chitosan-based polymers have been fabricated and used as wound dressings since they possess better properties than those of the constituent materials. The objective of this work was to evaluate the biocompatibility and biodegradability of biodegradable blend films based on polyethylene glycol-co-fumarate (PEGF) and chitosan (Ch). The blend films of Ch/PEGF were prepared by solution casting/solvent evaporation method. Degradation behavior of these blend films was evaluated in a simulated fluid at physiological pH supplemented with lysozyme at a concentration similar to that in human serum by weight loss of the films and changes in the pH of media. When the pH of incubation media was analyzed, with an increase of PEGF content in the blend films, the degradation rate increased accordingly. The pH of the media of samples was not significantly changed at any measured time point and all films kept their integrities during 28 days. The biocompatibility of the films and cell behavior on the surface of these films were investigated by in vitro tests. Biological assessment using mouse fibroblast cell line L929 on the blend films of Ch/PEGF indicated that films supported the attachment, spreading and proliferation of cells. Since the Ch/PEGF films are biocompatible with the tailored biodegradation rate, they might have a great prospective position in the application of wound dressings.

Preparation and characterization of magnetic core–shell iron oxide@glycyrrhizic acid nanoparticles in ethanol–water mixed solvent

In this research, we used the co-precipitation method to synthesize glycyrrhizic acid coated iron oxide magnetic nanoparticles. The aim of this study is to investigate how different amounts of glycyrrhizic acid affect the structural, biological, magnetic, and hyperthermic characteristics of the synthesized magnetic nanoparticles. The synthesis was conducted under different glycyrrhizic acid concentrations in water with the presence of ethanol to generate coated nanoparticles with different amounts of coating agent. The characteristics of the synthesized nanoparticles were examined by several devices including X-ray diffractometer, transmission electron microscope, field-emission scanning electron microscope, vibrating sample magnetometer, Fourier transform infrared spectra, and thermal gravimetric analyzer. The cytotoxicity of synthesized nanoparticles was examined by MTT assay using L929 fibroblast cell line. The results indicated the enhanced biocompatibility of the coated iron oxide nanoparticles due to the presence of glycyrrhizic acid. The comparison of the coated samples shows that the samples with higher amounts of coating agent were more biocompatible. The possibility of using the synthesized magnetic nanoparticles for medical hyperthermia was examined by performing hyperthermia process on a nanofluid made up of the nanoparticles dispersed in water using a high-frequency alternating magnetic field generator and the results confirm the effectiveness of the synthesized nanoparticles in the elevation of the solutions temperature.

Identification and preliminary structure-activity relationship studies of novel pyridyl sulfonamides as potential Chagas disease therapeutic agents

Chagas disease is a neglected pathology responsible for about 12,000 deaths every year across Latin America. Although six million people are infected by the Trypanosoma cruzi, current therapeutic options are limited, highlighting the need for new drugs. Here we report the preliminary structure activity relationships of a small library of 17 novel pyridyl sulfonamide derivatives. Analogues 4 and 15 displayed significant potency against intracellular amastigotes with EC50 of 5.4 µM and 8.6 µM. In cytotoxicity assays using mice fibroblast L929 cell lines, both compounds indicated low toxicity with decent selectivity indices (SI) >36 and >23 respectively. Hence these compounds represent good starting points for further lead optimization.

One step preparation of polymeric maltitol particles, from a sugar molecule, maltitol for biomedical applications

Here, a facile method for the preparation of polymeric particles from a sugar molecule, maltitol (ML) in single step was reported via microemulsion polymerization/crosslinking technique with a high yield, 89 ± 6%. Furthermore, poly(Maltitol) (p(ML)) particles were chemically modified to induce different physicochemical characteristic using different anionic and cationic modifying agents such as taurine (TA) and diethylenetriamine (DETA) to prepare m-p(ML)/TA and m-p(ML)/DETA. The blood compatibility of the p(ML) particles and their modified forms were tested with fresh blood, and found that p(ML) and m-(ML)/DETA particles are blood compatible with about 5% hemolysis, and above 80% blood clotting indices. Moreover, the cytotoxicity results against L929 fibroblast cell line revealed that p(ML) based particle are biocompatible up to 100 μg/mL with 85% cell viability regardless of their nature, and at 200 μg/mL dosage of p(ML), m-p(ML)/TA and m-p(ML)/DETA particles, the cell viabilities were determined as 83.33, 64.03 and 73.89% on for L929 fibroblast cells implying the slightly less biocompatibility nature of m-p(ML)/TA in accord with the blood compatibility results. Additionally, the apoptotic and necrotic indices were determination for p(ML) based particles on L929 fibroblast cells, and the results revealed that these particles do not induce any and stress on the cells. Also, it does not have genotoxic effect as determined against CHO cells.

Hyfraxins A and B, cytotoxic ergostane-type steroid and lanostane triterpenoid glycosides from the invasive ash dieback ascomycete Hymenoscyphus fraxineus

A virulent culture of Hymenoscyphus fraxineus, the causal agent of ash dieback, was investigated for its production of secondary metabolites in a 70 L batch fermentation. Chemical analysis of the mycelial extract by means of flash chromatography and preparative HPLC led to the isolation of a new ergostane-type steroid (1) and a new related lanostane triterpenoid (2), both revealing the same glycosylation pattern. While their planar structures were elucidated by HR-ESIMS and NMR data, relative stereochemistry was assigned by ROESY correlations in conjunction with H,H and C,H coupling constants. Absolute configuration was determined based on ROESY correlations between the aglycons and the sugar moieties, which were identified in both cases as d-mannose by GC/MS analysis of the trimethylsilylated derivatives. The isolated compounds, for which we propose the trivial names hyfraxins A (1) and B (2), were found to be cytotoxic against the mouse fibroblast cell line L929 and exhibited moderate to weak activity against Gram-positive bacteria.

Aptamer-functionalized magnetic graphene oxide nanocarrier for targeted drug delivery of paclitaxel

Targeted drug delivery has come out as a golden system owing to its capability to reduce side effects and increase efficacy. The application of graphene oxide (GO) based nanomaterials for targeted delivery has recently attracted several researchers because of their advantageous properties. Herein, we have attempted to prepare aptamer-conjugated magnetic graphene oxide (MGO) nanocarrier, which can specifically target tumor cells. MGO nanocarriers were prepared by attaching Fe3O4 on the layer of GO and then, aptamer (APT) was linked as a targeting moiety. Paclitaxel (PAC), an anti-cancer drug, was also loaded on the nanocarrier. PAC loading and in vitro release results revealed a very good loading performance with entrapment efficiency 95.75% and high pH-responsive release. Cellular toxicity assay showed MGO nanocarriers are biocompatible having cell viability greater than 80% for L-929 fibroblast cell line. Besides, high cytotoxic effect was observed for PAC and PAC loaded MGO (MGO@PAC) on MCF-7 cancer cells, at different drug doses. Furthermore, flow cytometry investigation reveals that the obtained nanocarrier can specifically bind to MCF-7 cancer cells. Therefore, based on the results the prepared superparamagnetic nanocarrier could be considered as a promising agent for cancer drug delivery systems.

Novel hemostatic biomolecules based on elastin-like polypeptides and the self-assembling peptide RADA-16

BackgroundSafe and effective hemostatic materials are important for reducing mortality resulting from excessive hemorrhage. In this work, new biomaterials with hemostatic effects were created by fusing the gene coding for RADA-16, a self-assembling peptide with the sequence RADARADARADARADA, to the 3′-end of the open reading frame (ORF) encoding elastin-like polypeptides through gene recombination.ResultsThe fusion proteins, termed 36R, 60R and 96R, were solubly over-expressed in Escherichia coli BL21 (DE3) based on genetic manipulation of the high-efficiency prokaryotic expression vector pET28a (+) and bacterial transformation. Western Blot analysis showed that the over-expressed proteins were the target fusion proteins. The target proteins 36R with 94.72% purity, 60R with 96.91% purity and 96R with 96.37% purity were prepared using an inverse phase transition cycle at 65 °C followed by His-tag affinity chromatography. The proliferation results of the mouse fibroblast cell line L929 and hippocampus neuron cell line HT22 indicated that the fusion proteins did not cause obvious cell toxicity. The lyophilized spongy film of the purified 36R, 60R and 96R could stop the hemorrhage of a 2 × 2 mm bleeding wound in the mouse liver after 27.21 ± 1.92 s, 18.65 ± 1.97 s and 15.85 ± 1.21 s, respectively. The hemostasis time was 21.23 ± 1.84 s for rat-tail collagen and 14.44 ± 1.33 s for RADA-16 lyophilized on gauze. The hemostatic time of three treated groups were all significantly superior to that of the negative control without any hemostasis treatment, which spontaneously stopped bleeding after 37.64 ± 1.34 s. Statistical analysis showed that the spongy film with purified 96R exhibited an exciting hemostatic effect that was superior to rat-tail collagen and close to that of RADA-16 lyophilized on gauze.ConclusionsThese results revealed that the fusion proteins achieved by gene recombination technology could serve as a promising hemostatic material.

Controlled release of mitomycin C from PHEMAH–Cu(II) cryogel membranes

Molecular imprinting technique was used for the preparation of antibiotic and anti-neoplastic chemotherapy drug (mitomycin C) imprinted cryogel membranes (MMC-ICM). The membranes were synthezied by using metal ion coordination interactions with N-methacryloyl-(l)-histidine methyl ester (MAH) functional monomer and template molecules (i.e. MMC). The 2-hydroxyethyl methacrylate (HEMA) monomer and methylene bisacrylamide (MBAAm) crosslinker were used for the preparation of mitomycin C imprinted cryogel membranes by radical suspension polymerization technique. The imprinted cryogel membranes were characterized by scanning electron microscopy (SEM), Brunauer–Emmett–Teller (BET), Fourier transform infrared spectroscopy-attenuated total reflectance (FTIR-ATR) and swelling degree measurements. Cytotoxicity of MMC-ICMs was investigated using mouse fibroblast cell line L929. Time-dependent release of MMC was demonstrated within 150 h from cryogel membranes. Cryogels demonstrated very high MMC loading efficiency (70–80%) and sustained MMC release over hours.