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1.
Int J Biol Macromol ; 79: 459-68, 2015 Aug.
Article in English | MEDLINE | ID: mdl-26003302

ABSTRACT

In the report, three bioactive fractions from Cerrena unicolor: laccase (LAC), endopolysaccharides (c-EPL), and low molecular weight (ex-LMS) were tested for the first time towards their antiviral, immunostimulatory, cytotoxic and antiproliferative effect. The immunomodulatory activity was studied by means of THP-1-derived macrophages able to synthesize and secrete IL-6 and TNF-α. We used cervical carcinoma cell lines SiHa (ATCC, HTB-35) and CaSki (ATCC, CRL 1550) to determine antitumor activity and human skin fibroblasts (HSF) as a control. SiHa and L929 cell lines were used in the antiviral activity assay to propagate HHV-1 and EMCV, respectively. LAC was the most active against HSV at an early stage of viral replication, whereas the activity of laccase against EMCV was evident after incubation of the virus with LAC before and after the adsorption step. Moreover, the investigations showed that the fungal c-EPL fraction stimulated the production and secretion of TNF-α and IL-6 by THP-1-derived macrophages up to a level of 2000 pg/ml and 400 pg/ml, respectively. It was indicated for the first time that the LAC and ex-LMS fractions exhibited anticancer activity. This resulted from their cytotoxic or antiproliferative action against the investigated tumor cells at concentrations above 250 µg/ml and 10 µg/ml, respectively.


Subject(s)
Antineoplastic Agents/isolation & purification , Antiviral Agents/isolation & purification , Fungal Polysaccharides/isolation & purification , Fungal Proteins/isolation & purification , Immunologic Factors/isolation & purification , Laccase/isolation & purification , Polyporaceae/chemistry , Antineoplastic Agents/pharmacology , Antiviral Agents/pharmacology , Cell Line , Cell Line, Tumor , Cell Survival/drug effects , Encephalomyocarditis virus/drug effects , Encephalomyocarditis virus/physiology , Epithelial Cells/drug effects , Epithelial Cells/pathology , Fibroblasts/drug effects , Fibroblasts/pathology , Fibroblasts/virology , Fungal Polysaccharides/pharmacology , Fungal Proteins/pharmacology , Herpesvirus 1, Human/drug effects , Herpesvirus 1, Human/physiology , Humans , Immunologic Factors/pharmacology , Interleukin-6/biosynthesis , Interleukin-6/metabolism , Laccase/pharmacology , Macrophages/drug effects , Macrophages/pathology , Macrophages/virology , Polyporaceae/metabolism , Tumor Necrosis Factor-alpha/biosynthesis , Tumor Necrosis Factor-alpha/metabolism , Virus Replication/drug effects
2.
Arch Immunol Ther Exp (Warsz) ; 63(3): 197-205, 2015 Jun.
Article in English | MEDLINE | ID: mdl-25193979

ABSTRACT

The role of endogenous animal opioids in the biology of cancer is widely recognized but poorly understood. This is, among others, because of the short half-life of these peptides, which are quickly inactivated by endopeptidases, e.g., neutral endopeptidase (NEP, CD10). It has been established that NEP is engaged in the modulation of the tumor microenvironment, among others that of colon cancer, by exerting influence on cell growth factors, the extracellular matrix and other biologically active substances. Although there are some discrepancies among the findings on the role of both opioids and NEP in cancer development, authors agree that their role seems to depend on the origin, stage and grade of tumor, and even on the method of examination. Moreover, recently, natural inhibitors of NEP, such as sialorphin, opiorphin and spinorphin have been detected. Their analgesic activity has been established. It is interesting to ask whether there is a relationship among opioid peptides, tumor-associated NEP and its inhibitors.


Subject(s)
Analgesics, Opioid/pharmacology , Analgesics/pharmacology , Colonic Neoplasms/metabolism , Neprilysin/metabolism , Animals , Carcinogenesis , Humans , Oligopeptides/pharmacology , Protein Precursors/pharmacology , Salivary Proteins and Peptides/pharmacology , Tumor Microenvironment
3.
Biomed Res Int ; 2014: 743812, 2014.
Article in English | MEDLINE | ID: mdl-25114920

ABSTRACT

A new exopolysaccharide preparation isolated from stationary cultures of the white rot fungus Ganoderma applanatum (GpEPS) was tested in terms of its bioactive properties including its cytotoxic and immunostimulatory effect. The results indicate that the tested GpEPS (at concentrations above 22.85 µg/mL and 228.5 µg/mL) may exhibit selective activity against tumor cells (cell lines SiHa) and stimulate production of TNF-α THP-1-derived macrophages at the level of 752.17 pg/mL. The GpEPS showed antibacterial properties against Staphyloccoccus aureus and a toxic effect against Vibrio fischeri cells (82.8% cell damage). High cholesterol-binding capacity and triglycerides-binding capacity (57.9% and 41.6% after 24 h of incubation with the tested substances, resp.) were also detected for the investigated samples of GpEPS.


Subject(s)
Anti-Bacterial Agents/pharmacology , Antineoplastic Agents/pharmacology , Cytostatic Agents/pharmacology , Fungal Polysaccharides/pharmacology , Ganoderma/chemistry , Aliivibrio fischeri/drug effects , Anti-Bacterial Agents/chemistry , Antineoplastic Agents/chemistry , Cell Line, Tumor , Cell Proliferation/drug effects , Cytostatic Agents/chemistry , Fungal Polysaccharides/chemistry , Humans , Immunologic Factors , Microbial Viability/drug effects
4.
J Basic Microbiol ; 54(3): 232-46, 2014 Mar.
Article in English | MEDLINE | ID: mdl-23456635

ABSTRACT

The influence of infection with an entomopathogenic strain of Pseudomonas aeruginosa on Galleria mellonella hemocytes was investigated. Extensive bacteriaemia developed 18 h after infection. This was correlated with significant changes in morphology, viability and the spreading ability of immunocompetent hemocytes, namely granulocytes and plasmatocytes. Since bacteriaemia developed, membrane blebbing, cytoplasm vacuolization, cell and organelle swelling, and chromatin condensation were observed among others. These features are typical for apoptotic and autophagal cell death. A gradually increasing level of procaspase and its activation as well as lack of DNA degradation were also detected. Propidium iodide and acridine orange staining indicated that hemocytes become dead ultimately. Infection of G. mellonella larvae with P. aeruginosa also caused significant changes in the arrangement of the actin cytoskeleton in the hemocytes, which might be correlated with their restricted spreading ability.


Subject(s)
Hemocytes/cytology , Moths/microbiology , Pseudomonas aeruginosa/physiology , Animals , Apoptosis , Cytoskeleton/pathology , Hemocytes/ultrastructure , Larva/cytology , Larva/microbiology , Moths/cytology
5.
Leuk Res ; 37(5): 586-94, 2013 May.
Article in English | MEDLINE | ID: mdl-23434428

ABSTRACT

The mechanisms involved in anti-myeloma activity of statins combined with thalidomide were studied in multiple myeloma (MM) cells. In addition, the effect of p38 MAPK inhibition on the induction of apoptosis in MM cells by the combination of thalidomide and simvastatin was investigated. Thalidomide was observed to significantly potentiate the antiproliferative activity of statins and enhance the proapoptotic effect of simvastatin and lovastatin. What is more, the combination of thalidomide with statins inhibited cell migration and decreased the production of VEGF and MMP-9 in MM cells more effectively than each of these drugs used separately. The combination of simvastatin and thalidomide augmented caspase 8 and 3 activation, and the additional application of p38 MAPK inhibitor resulted in enhanced apoptosis of MM cells concomitant with increased caspase 9 and 3 activation, as well as JNK phosphorylation. The results suggest that p38 inhibitors together with the combination of simvastatin and thalidomide have the potential to be used in MM treatment.


Subject(s)
Apoptosis/drug effects , Hydroxymethylglutaryl-CoA Reductase Inhibitors/pharmacology , Immunosuppressive Agents/pharmacology , Multiple Myeloma/drug therapy , Protein Kinase Inhibitors/pharmacology , Simvastatin/pharmacology , Thalidomide/pharmacology , p38 Mitogen-Activated Protein Kinases/antagonists & inhibitors , Caspases/biosynthesis , Cell Line, Tumor , Enzyme Activation/drug effects , Gene Expression Regulation, Leukemic/drug effects , Humans , MAP Kinase Kinase 4/metabolism , Multiple Myeloma/enzymology , Multiple Myeloma/pathology , Phosphorylation/drug effects , Vascular Endothelial Growth Factor A/biosynthesis , p38 Mitogen-Activated Protein Kinases/metabolism
6.
J Gastroenterol ; 48(2): 222-37, 2013 Feb.
Article in English | MEDLINE | ID: mdl-22722906

ABSTRACT

BACKGROUND: Butein has been reported to prevent and partly reverse liver fibrosis in vivo; however, the mechanisms of its action are poorly understood. We, therefore, aimed to determine the antifibrotic potential of butein. METHODS: We assessed the influence of the incubation of hepatic stellate cells (HSCs) and hepatoma cells (HepG2) with butein on sensitivity to ethanol- or acetaldehyde-induced toxicity; the production of reactive oxygen species (ROS); the expression of markers of HSC activation, including smooth muscle α-actin (α-SMA) and procollagen I; and the production of transforming growth factor-ß1 (TGF-ß1), metalloproteinases-2 and -13 (MMP-2and MMP-13), and tissue inhibitors of metalloproteinases (TIMPs). The influence of butein on intracellular signals in HSCs; i.e., nuclear factor-κB (NFκB), c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (p38 MAPK) induced by ethanol was estimated. RESULTS: Butein protected HSCs and HepG2 cells against ethanol toxicity by the inhibition of ethanol- or acetaldehyde-induced production of ROS when cells were incubated separately or in co-cultures; butein also inhibited HSC activation measured as the production of α-SMA and procollagen I. As well, butein downregulated ethanol- or acetaldehyde-induced HSC migration and the production of TGF-ß, TIMP-1, and TIMP-2; decreased the activity of MMP-2; and increased the activity of MMP-13. In ethanol-induced HSCs, butein inhibited the activation of the p38 MAPK and JNK transduction pathways as well as significantly inhibiting the phosphorylation of NF κB inhibitor (IκB) and Smad3. CONCLUSIONS: The results indicated that butein inhibited ethanol- and acetaldehyde-induced activation of HSCs at different levels, acting as an antioxidant and inhibitor of ethanol-induced MAPK, TGF-ß, and NFκB/IκB transduction signaling; this result makes butein a promising agent for antifibrotic therapies.


Subject(s)
Antioxidants/pharmacology , Chalcones/pharmacology , Ethanol/antagonists & inhibitors , Hepatic Stellate Cells/drug effects , Oxidative Stress/drug effects , Acetaldehyde/antagonists & inhibitors , Acetaldehyde/pharmacology , Actins/biosynthesis , Animals , Cell Death/drug effects , Cells, Cultured , Coculture Techniques , Collagen Type I/biosynthesis , Drug Evaluation, Preclinical/methods , Ethanol/pharmacology , Hep G2 Cells , Hepatic Stellate Cells/metabolism , Humans , MAP Kinase Kinase 4/physiology , NF-kappa B/physiology , Oxidative Stress/physiology , Rats , Reactive Oxygen Species/metabolism , Signal Transduction/drug effects , Signal Transduction/physiology , Transforming Growth Factor beta/physiology , p38 Mitogen-Activated Protein Kinases/physiology
7.
J Insect Sci ; 12: 88, 2012.
Article in English | MEDLINE | ID: mdl-23421724

ABSTRACT

Abstract Susceptibility of proteins and peptides present in immune hemolymph of Galleria mellonella Fabricius (Lepidoptera: Pyralidae) larvae to proteolytic degradation by purified elastase B of Pseudomonas aeruginosa was studied. Results showed that apoLp-III protein was gradually digested by elastase B in vitro. Additionally, polipeptides with molecular mass 6.5 and 4 kDa were degraded after treatment with the studied enzyme. The lack of these peptides and the decrease in anti-Escherichia coli activity could indicate that inducible antimicrobial peptides were digested by elastase B. On the contrary, no change in the lysosome activity level was observed in immune hemolymph incubated with elastase B. Thus, elastase B might contribute to the pathogenesis of P. aeruginosa.


Subject(s)
Apolipoproteins/metabolism , Bacterial Proteins/toxicity , Insect Proteins/metabolism , Metalloendopeptidases/toxicity , Moths/immunology , Pseudomonas aeruginosa/enzymology , Animals , Escherichia coli/immunology , Hemolymph/metabolism , Immunity, Humoral , Larva/growth & development , Larva/immunology , Moths/growth & development , Proteolysis
8.
J Invertebr Pathol ; 107(1): 16-26, 2011 May.
Article in English | MEDLINE | ID: mdl-21236262

ABSTRACT

The role of Pseudomonas aeruginosa elastase B in activation of the humoral immune response in Galleria mellonella larvae was investigated. The results of our study showed that elastase B injected at a sublethal concentration was responsible for eliciting the humoral immune response in G. mellonella larvae. The insects exhibited increased antibacterial activity, namely, we observed appearance of antimicrobial peptides and a higher level of lysozyme in cell-free hemolymph. Elastase B seems to be a more potent elicitor than thermolysin because similar maximal antibacterial activity levels were observed at a 5-fold lower concentration. We also demonstrated that there were differences in the kinetics of induction of antimicrobial activity between thermolysin and elastase B. The maximum level was observed 18h post challenge of thermolysin and 38h after injection of elastase B. It was also shown that, 24h after elastase injection, the relative levels of apoLp-III in the hemolymph significantly increased in comparison with control G. mellonella larvae. The activation of immune responses in metalloproteinase-challenged larvae involved synthesis of metalloproteinase inhibitors which increased the survival rates of insects both against the lethal dose of thermolysin as well as against viable pathogenic bacterial cells of P. aeruginosa.


Subject(s)
Bacterial Proteins/immunology , Immunity, Humoral/immunology , Moths/immunology , Moths/microbiology , Pancreatic Elastase/immunology , Animals , Electrophoresis, Polyacrylamide Gel , Hemolymph/immunology , Immunoblotting , Pseudomonas Infections/immunology , Pseudomonas aeruginosa/immunology , Pseudomonas aeruginosa/metabolism , Thermolysin/immunology
9.
Comp Biochem Physiol B Biochem Mol Biol ; 152(2): 118-23, 2009 Feb.
Article in English | MEDLINE | ID: mdl-18996217

ABSTRACT

The antibacterial activity of hemolymph from Galleria mellonella infected with entomopathogenic strain of Pseudomonas aeruginosa and non-pathogenic bacterium Escherichia coli was studied. In vivo, the antimicrobial activity appeared shortly after P. aeruginosa infection, reached the maximum level 18 h postinjection, while 30 h later only trace activity was noted. The activity induced by E. coli sustained on the high level until 48 h after infection. We also noted that the antimicrobial activity level induced by the non-pathogenic bacterium was higher in comparison to that measured in insects infected with the pathogenic strain of P. aeruginosa. The results of our in vitro studies indicated that inducible antimicrobial peptides of G. mellonella larvae were digested by P. aeruginosa elastase B. After 1 h incubation of cell-free hemolymph of immune-challenged larvae with elastase B, no antibacterial activity was observed. It was also shown that elastase B degraded synthetic cecropin B while in the presence of 6 mM EDTA antibacterial activity of cell-free hemolymph as well as cecropin B, was not changed which confirmed that the activity was abolished by the metalloprotease.


Subject(s)
Anti-Bacterial Agents/metabolism , Hemolymph/microbiology , Moths/microbiology , Pseudomonas Infections/microbiology , Pseudomonas aeruginosa/physiology , Animals , Antimicrobial Cationic Peptides/metabolism , Escherichia coli/drug effects , Hemolymph/drug effects , Larva/drug effects , Larva/microbiology , Pancreatic Elastase/pharmacology , Pseudomonas aeruginosa/drug effects , Pseudomonas aeruginosa/enzymology
10.
J Invertebr Pathol ; 97(1): 14-9, 2008 Jan.
Article in English | MEDLINE | ID: mdl-17681528

ABSTRACT

The level of apoLp-III in fat body, hemocytes and plasma from Galleria mellonella larvae infected with Pseudomonas aeruginosa was studied. It was found that the amount of 18kDa protein present in fat body and hemocytes decreased progressively with time after infection. In the case of plasma, an increase in apoLp-III content was observed during the first 19h after infection and then decreased significantly after prolonged infection time. The decreased level of apoLp-III in plasma 24h after infection was accompanied by the appearance of smaller than 18kDa immunoreactive polypeptides. Four intermediate forms with molecular mass of, respectively, 15, 13.3, 12 and 9.5kDa were detectable. The size of polypeptides detected in experiments performed in vivo is comparable with the degradation products of apoLp-III produced by serine protease IV in vitro. In addition, the total proteolytic activity of plasma increased progressively during infection time. The results of our studies suggest that a significant part of total proteolytical activity in the plasma of infected G. mellonella larvae can be attributed to proteases produced by P. aeruginosa during pathogenesis. We discuss the possibility that protease IV of P. aeruginosa is responsible for apoLp-III degradation in vivo.


Subject(s)
Apolipoproteins/metabolism , Moths/metabolism , Moths/microbiology , Pseudomonas Infections/metabolism , Animals , Electrophoresis, Polyacrylamide Gel , Immunoblotting , Peptide Hydrolases/metabolism , Pseudomonas aeruginosa/pathogenicity
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