This is the first study on the surface modification of a hemocyanin from marine snail Rapana thomasiana (RtH) with series of imidazolium-based amino acid ionic liquids emimAA. We monitored the ...induced by emimAA conformational changes in RtH molecule and evaluated the effect of these ionic liquids (ILs) on the protein thermal stability. The cytotoxicity of all obtained RtH-emimAA complexes was assessed toward breast cancer cells (MCF-7) and murine fibroblasts (3T3).
As a whole, even small amounts of the tested ILs altered the secondary structure of RtH. The thermal denaturation of RtH in presence of emimAA displayed multi-component transitions, which were shifted toward lower temperatures in comparison to those estimated for the native RtH. The profiles of the RtH-IL calorimetric curves show a clear dependence on the structure of the added salts. In addition, all RtH-emimAA complexes exhibited an enhanced antiprofilerative activity of toward MCF-7 cells in comparison to that of the native RtH. The best results are observed for RtH-emimLeu, RtH-emimTrp or RtH-emimIle, which applied in concentration of 700μg/mL inhibited the MCF-7 cell viability (for 24h) by 66, 63 and 53%, respectively. In addition, these IL-RtH complexes were less cytotoxic to 3T3 cells, i.e. they exhibited some cell specificity.
This is the first study on the interactions of ionic liquids with large metalloproteins, in particular hemocyanins (Hcs). At first, complexes of a Hc from Rapana thomasiana (RtH) with a series of ...biocompatible choline amino acid salts CholAA were obtained. Applying UV-vis spectroscopy, Fourier-transformed infrared spectroscopy and differential scanning calorimetry the effect of these organic salts on the structure and thermal stability of RtH was assessed. Then, the cytotoxic effect of RtH–CholAA on breast cancer cells (MCF-7) and 3T3 fibroblast cells (non cancerous) was evaluated. We found that all CholAA induced clear time- and concentration-dependent alterations in the RtH conformation. The conformation and the thermal stability of IL-modified RtH depend strongly on the type of the anion of the tested compounds. All CholAA-modified RtHs exhibited lower thermal stability than the native RtH. At the same time, we established a good correlation between the structure of RtH and its antitumor activity. Namely, RtH–CholAA complexes exhibited enhanced antiproliferative activity toward the MCF-7 cell line. The observed antiproliferative effect was cell specific and the compounds have no effect or in some cases have stimulatory effect on fibroblasts.
Angiogenesis is one of the key processes during development, wound healing and tumor formation. Prerequisite for its existence is the presence of endogenous electrical fields (EFs) generated by ...active ion transport across polarized epithelia and endothelia, and appearance of the transcellular potentials. During angiogenesis cellular factor as endothelial growth factor (VEGF), synthesis of adhesive proteins and membrane metalloproteinases (MMPs) govern the angiogenic response to different external stimuli as biomaterials interactions and/or exogenous EF. Gelatin-based hydrogels with elasticities comparable to human tissues have shown to influence cell behavior as well as cell attachment, protein synthesis, VEGF and MMP's production after the application of EF. Gelatin-based matrices with 3 (G10_LNCO3), 5 (G10_LNCO5), and 8 (G10_LNCO8) fold excess of isocyanate groups per mol of amine groups present in gelatin were used. Human umbilical endothelial cells (HUVEC) (Lonza Basel, Switzerland) and highly invasive breast cancer MDA-MB-231 cells (ATCC®HTB-26TM) were used. For an estimation of the amount of VEGF released from cells a commercially available VEGF ELISA (Thermo Fisher Scientific, Germany) kit was used. Fibronectin (FN) enzyme immunoassay (EIA) was used to analyze the secreted amount of FN by cells seeded on the materials. Secreted MMPs were analyzed by zymography. Gelatin-based hydrogels attracted HUVEC adhesion and diminished the adhesion of MDA-MB-231 cells. The applied direct current (DC) EF induced an almost 5-fold increase in VEGF production by HUVEC seeded on gelatin-based hydrogels, while in contrast, the applied EF decreased the production of VEGF by cancer cells. FN synthesis was elevated in HUVEC cells seeded on gelatin-based materials in comparison to FN synthesis by cancer cells. HUVEC seeded on gelatin hydrogels showed an expression mainly of MMP-2. The application of EF increased the production of MMP-2 in HUVEC seeded on gelatin materials. In contrast, for MDA-MB-231 the production of MMPs on gelatin materials was lower compared to control materials. With the application of EF the levels of MMP-9 decreased but MMP-2 expression raised significantly for gelatin materials. Overall, the results showed that studied gelatin materials suppressed attachment of cancerous cells, as well as suppressed their angiogenic potential revealed by decreased VEGF and MMP production. Thus, this study approved gelatin-based hydrogels with proper elasticity characteristics and different degradation behavior as useful matrices for use in vascular tissue regeneration or in restriction of tumor growth after tumor resection.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
The biocompatibility of hybrid hydroxypropyl cellulose (HPC) hydrogels with different content of embedded silver nanoparticles (AgNPs) is revealed by testing the cytotoxicity and induction of cell ...death. The effect of cytotoxicity and cell death was studied on L929 fibroblasts by using MTT cytotoxicity assay, cell morphological observations and acridine orange/ethidium bromide (AO/EtBr) live cell staining. The results showed good biocompatibility of hydrogels containing AgNPs in the range 0.5-1.5 wt % Ag. Cells incubated with HPC hydrogels with the higher amount of AgNPs (2-2.5 wt % Ag) presented morphological changes corresponding to cell death and increased grade of cytotoxicity revealed by MTT assay. All these data suggest that AgNPs content in HPC materials exibits dose-dependent threshold over which the biocompatibility of the hydrogels is disturbed. Hybrid materials with low silver content - 0.5 wt % to 1.5 wt % Ag proved their biocompatibility and will be suitable candidates for biomedical applications.Key words: antibacterial materials, silver nanoparticles, biocompatibility, cytotoxicity, cell death
In the present study we aimed to evaluate the role of cytotoxicity of erufosine for eliciting changes in cytoskeleton organization and induction of apoptosis in Graffi myeloid tumour cells. The ...cytotoxicity of erufosine was revealed by MTT assay. The effect of erufosine on cytoskeleton and cell nuclei was evaluated by immunostaining for alpha-tubulin and F-actin, as well as by DAPI staining. We show that IC.sub.50 dose for EPC.sub.3 treatment of Graffi tumour cells was obtained at 20 microM. Fluorescent images showed existence of apoptosis at the same EPC.sub.3 concentration. The induction of apoptosis by EPC.sub.3 was accompanied by actin and tubulin reorganization. The obtained results revealed reorganization of actin cytoskeleton and induction of adhesive cell phenotype by erufosine treatment.Key words: erufosine, reorganization of cytoskeleton, apoptosis, Graffi tumour cells
This is the first study on the interactions of ionic liquids with large metalloproteins, in particular hemocyanins (Hcs). At first, complexes of a Hc from
Rapana thomasiana
(RtH) with a series of ...biocompatible choline amino acid salts CholAA were obtained. Applying UV-vis spectroscopy, Fourier-transformed infrared spectroscopy and differential scanning calorimetry the effect of these organic salts on the structure and thermal stability of RtH was assessed. Then, the cytotoxic effect of RtH-CholAA on breast cancer cells (MCF-7) and 3T3 fibroblast cells (non cancerous) was evaluated. We found that all CholAA induced clear time- and concentration-dependent alterations in the RtH conformation. The conformation and the thermal stability of IL-modified RtH depend strongly on the type of the anion of the tested compounds. All CholAA-modified RtHs exhibited lower thermal stability than the native RtH. At the same time, we established a good correlation between the structure of RtH and its antitumor activity. Namely, RtH-CholAA complexes exhibited enhanced antiproliferative activity toward the MCF-7 cell line. The observed antiproliferative effect was cell specific and the compounds have no effect or in some cases have stimulatory effect on fibroblasts.
This is the first study on the interactions of ionic liquids with large metalloproteins, in particular hemocyanins (Hcs).
This is the first study on the interactions of ionic liquids with large metalloproteins, in particular hemocyanins (Hcs). At first, complexes of a Hc from Rapana thomasiana(RtH) with a series of ...biocompatible choline amino acid salts CholAA were obtained. Applying UV-vis spectroscopy, Fourier-transformed infrared spectroscopy and differential scanning calorimetry the effect of these organic salts on the structure and thermal stability of RtH was assessed. Then, the cytotoxic effect of RtH-CholAA on breast cancer cells (MCF-7) and 3T3 fibroblast cells (non cancerous) was evaluated. We found that all CholAA induced clear time- and concentration-dependent alterations in the RtH conformation. The conformation and the thermal stability of IL-modified RtH depend strongly on the type of the anion of the tested compounds. All CholAA-modified RtHs exhibited lower thermal stability than the native RtH. At the same time, we established a good correlation between the structure of RtH and its antitumor activity. Namely, RtH-CholAA complexes exhibited enhanced antiproliferative activity toward the MCF-7 cell line. The observed antiproliferative effect was cell specific and the compounds have no effect or in some cases have stimulatory effect on fibroblasts.
Microbial invasion into the intestinal mucosa after allogeneic hematopoietic cell transplantation (allo-HCT) triggers neutrophil activation and requires antibiotic interventions to prevent sepsis. ...However, antibiotics lead to a loss of microbiota diversity, which is connected to a higher incidence of acute graft-versus-host disease (aGVHD). Antimicrobial therapies that eliminate invading bacteria and reduce neutrophil-mediated damage without reducing the diversity of the microbiota are therefore highly desirable. A potential solution would be the use of antimicrobial antibodies that target invading pathogens, ultimately leading to their elimination by innate immune cells. In a mouse model of aGVHD, we investigated the potency of active and passive immunization against the conserved microbial surface polysaccharide poly-N-acetylglucosamine (PNAG) that is expressed on numerous pathogens. Treatment with monoclonal or polyclonal antibodies to PNAG (anti-PNAG) or vaccination against PNAG reduced aGVHD-related mortality. Anti-PNAG treatment did not change the intestinal microbial diversity as determined by 16S ribosomal DNA sequencing. Anti-PNAG treatment reduced myeloperoxidase activation and proliferation of neutrophil granulocytes (neutrophils) in the ileum of mice developing GVHD. In vitro, anti-PNAG treatment showed high antimicrobial activity. The functional role of neutrophils was confirmed by using neutrophil-deficient LysMcre Mcl1fl/fl
mice that had no survival advantage under anti-PNAG treatment. In summary, the control of invading bacteria by anti-PNAG treatment could be a novel approach to reduce the uncontrolled neutrophil activation that promotes early GVHD and opens a new avenue to interfere with aGVHD without affecting commensal intestinal microbial diversity.
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