The influence of chronic and subacute exposure to lead on the levels of prolactin, leptin, osteopontin, and follistatin in humans.

This study was designed to determine the levels of prolactin, leptin, osteopontin, and follistatin in workers chronically and subacutely exposed to lead compounds. The examined population consisted of three groups. The first group was composed of 56 male workers who were chronically exposed to lead for 13.38 ± 10.38 years. The second group served as a control group and consisted of 24 male administrative workers, while the third group included 32 male workers exposed to lead for 40 ± 3 days. The levels of leptin, osteopontin, and prolactin were significantly lower in the group of workers chronically exposed to lead than in the control group by 42%, 26%, and 41%, respectively. The levels of follistatin did not differ between those groups. The levels of all measured hormones did not change after a short-term exposure to lead compared to baseline. Chronic lead exposure is associated with significantly decreased level of prolactin, leptin, and osteopontin. Lead-induced changes in the levels of these hormones may disturb many functions of the human body, including the immune response, metabolism, reproduction, and bone turnover.

ALA-induced photodynamic effect on vitality, apoptosis, and secretion of vascular endothelial growth factor (VEGF) by colon cancer cells in normoxic environment in vitro.

BACKGROUND: Cancer therapy is often based on combination of conventional methods of cancer treatment with immunotherapy. Photodynamic therapy (PDT) is one of the immunomodulating methods used in oncology. We examined how PDT influences the secretory activity of colon cancer cells in vitro, especially the secretion of vascular endothelial growth factor (VEGF) in aerobic conditions. METHODS: We used two cancer cell lines with different malignancy potentials: a metastatic SW620 line and a non-metastatic SW480 line. In the first stage of the experiment, we exposed each cell line to three different concentrations of photosensitizer's precursor: 5-aminolevulinic acid (ALA) and varying levels of light radiation, after which we assessed cell viability and apoptosis induction in these lines, using the MTT and LDH assays. Then, we determined the secretion of VEGF by these cells in aerobic conditions and under the ALA-PDT parameters at which cells presented the highest viability. RESULTS: Photodynamic treatment with ALA did not influence on VEGF secretion by the non-metastatic SW480 cells, but caused a decrease in VEGF secretion by the metastatic SW 620 cell line by 29% (p<0.05). SW 620 cell line secreted more actively VEGF than the SW480 cells, both before and after photo dynamic therapy (p<0.05). CONCLUSION: The outcome of this in vitro study presented a beneficial effect of ALA-PDT, resulting in a decrease of VEGF secretion in the more malignant SW620 cell lines. Further studies should be considered to confirm the clinical relevance of this finding.

[Tumour-stroma interactions in urothelial cancer]. / Tumor-Stroma-Interaktionen im Harnblasenkarzinom.

BACKGROUND: The histopathological structure of malignant tumours involves two essential compartments - the tumour parenchyma with the actual transformed cells, and the supportive tumour stroma. The latter consists of specialized mesenchymal cells, such as fibroblasts, macrophages, lymphocytes and vascular cells, as well as of their secreted products, including components of the extracellular matrix, matrix modifying enzymes and numerous regulatory growth factors and cytokines. In consequence, the tumour stroma has the ability to influence virtually all aspects of tumour development and progression, including therapeutic response. AIM: In this article we review the current knowledge of tumor stroma interactions in urothelial carcinoma and present various experimental systems that are currently in use to unravel the biological basis of these heterotypic cell interactions. RESULTS: For urothelial carcinoma, an extensive tumour stroma is quite typical and markers of activated fibroblasts correlate significantly with clinical parameters of advanced disease. Another clinically important variable is provided by the stromal expression of syndecan-1. CONCLUSION: Integration of markers of activated stroma into clinical risk evaluation could aid to better stratification of urothelial bladder carcinoma patients. Elucidation of biological mechanisms underlying tumour-stroma interactions could provide new therapeutical targets.

Administration of low doses of tumor necrosis factor-alpha protects rat liver from ischaemic damage and reperfusion injury.

Liver ischaemia and reperfusion (IR) injury is a significant clinical problem. The aim of our study was to investigate the protective effect of tumor necrosis factor-alpha (TNF-alpha) on rat liver ischaemia-reperfusion injury. A TNF-alpha dose of 3 microg/kg body weight was injected into rats that had undergone partial (70%) ischaemia and reperfusion. The activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), total blood antioxidant level (using the FRAP test), and the concentrations of TNF-alpha, myeloperoxidase (MPO) and malondialdehyde (MDA) in liver homogenates after 1, 6, and 72 hours of reperfusion were measured. It was demonstrated that, rats subjected to IR, the administration of small doses of TNF-alpha significantly reduced ALT and AST activities after 60- minute liver ischaemia and 1 or 6 hour of reperfusion. The strongest reductions in ALT and AST activities were seen after 1 hour of reperfusion (30% and 35%, respectively). Exogenous TNF-alpha reduced the release of this cytokine in all observed periods, with the greatest reduction observed after 1 hour of reperfusion. Decreases in MPO concentration (by 40-45% in all periods of observation), as a marker of hepatic neutrophil infiltration, and in MDA concentration, the end-product of lipid peroxidation (by 55-60% at all time points), accompanied the reduction of TNF-alpha release. The administration of TNF-alpha to the rats after IR did not alter total plasma antioxidant potential, as assayed by the FRAP test, after 1 hour of reperfusion; however, at the later times a marked increase (approximately 40-50%) occurred. We demonstrated that intraperitoneal injections of small doses of TNF-alpha protect rat livers from IR injury. The mechanism of this protection is related to reductions in the release of TNF-alpha during IR after injection of this cytokine, resulting in reductions in oxidative stress and inflammation during the later phase of reperfusion.

Influence of 5-aminoisoquinolin-1-one (5-AIQ) on neutrophil chemiluminescence in rats with transient and prolonged focal cerebral ischemia and after reperfusion.

Stimulation of neutrophils by different factors increases their oxidative activity and the free radicals produced can report on the degree of activation. Poly(adenosine 5'-diphosphate ribose)polymerase-1 (PARP-1), a nuclear enzyme activated by strand breaks in DNA, plays an important role in the tissue injury associated with ischaemia-reperfusion injury and inflammation. 5-aminoisoquinolin-1-one (5-AIQ) is a potent inhibitor of PARP-1 activity in vitro and in vivo in rats. Acute (80 min) and prolonged (24h) focal cerebral ischaemia was induced in rats by obstruction of the median cerebral artery, with or without reperfusion, with or without administration of 5-AIQ. The oxidative activity of neutrophils was measured by chemiluminescence. Administration of 5-AIQ.HCl (3.0 mg kg(-1) b.w. - i.v.) caused a significant decrease in the oxidative activity of neutrophils in the group which had experienced chronic ischaemia for 24h but had no significant effect in the group which had received 80 min ischaemia, when compared to the control group. Increase of the oxidative activity of neutrophils was confirmed in rats with prolonged cerebral ischaemia, followed by reperfusion. 5-AIQ probably may decrease this activity through inhibition of PARP-1 in focus of local ischaemia as well as hence lowering the expression of inflammatory mediators by activated neutrophils.

Effects of ethanol extract of propolis (EEP) and its flavones on inducible gene expression in J774A.1 macrophages.

Propolis, a bee-hive product, has been used in folk medicine for centuries, and recently in modern medicine as an anti-inflammatory and immunomodulatory agent. These activities would be mainly due to phenolic compounds such as flavonoids, especially flavone derivatives. The present study examined the effect of ethanol extract of propolis (EEP) and selected flavone derivatives (chrysin, galangin, kaempferol and quercetin) on interleukin-1beta (IL-1beta) and inducible nitric oxide synthase (iNOS) gene expression in lipopolysaccharide (LPS)-induced J774A.1 macrophages. Treatment of cells with EEP significantly suppressed both IL-1beta mRNA (P<0.02) and iNOS mRNA (P<0.001) expression. The concentrations of cytokine in cell culture supernatants and cell lysates and nitric oxide (NO) generation were reduced in a dose-dependent manner. The tested phenolic compounds significantly decreased the IL-1beta mRNA level and IL-1beta protein concentration (P<0.05) (excluding galangin), iNOS mRNA level and NO production (P<0.001). The most potent inhibitor of the IL-1beta synthesis and NO generation was chrysin. These results indicate that EEP exerts its inhibitory effect on the IL-1beta and iNOS gene expression in J774A.1 macrophages at the transcriptional level. Tested flavone derivatives contribute to the anti-inflammatory activity of propolis.

Effect of flavone derivatives on interleukin-1beta (IL-1beta) mRNA expression and IL-1beta protein synthesis in stimulated RAW 264.7 macrophages.

It is known that the redox status of cells affects gene expression. Flavones, as natural antioxidants, efficiently modulate this status and may play a role in the regulation of inducible gene expression of inflammatory mediators. This study was designed to investigate the effect of five flavone derivatives variously substituted with hydroxyl groups (chrysin, galangin, kaempferol, quercetin and myricetin) on interleukin-1beta (IL-1beta) gene expression in stimulated RAW 264.7 macrophages. The cells were incubated with tested hydroxyflavones and stimulated with lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). Then, the following were estimated: the level of IL-1beta mRNA in these cells and the concentration of IL-1beta protein in cell-culture supernatants and cell lysates. Each of the tested compounds significantly decreased IL-1beta mRNA expression. The most potent inhibitor was chrysin (hydroxyflavone with two hydroxyl groups and a weak antioxidant activity). The effects of galangin and kaempferol were similar. Myricetin (hydroxyflavone with a strong antioxidant activity) significantly decreased the level of IL-1beta mRNA, but it had no effect on the IL-1beta protein synthesis. The results indicated that hydroxyflavones could modulate the IL-1beta gene expression in activated RAW 264.7 macrophages via inhibiting gene transcription. This action seems unlikely to be the result of antioxidant properties of tested compounds.

[Evaluation of selected immunological parameters in alcohol dependence syndrome]. / Ocena wybranych parametrów immunologicznych u pacjentów z zespolem zaleznosci alkoholowej.

In 28 patients with alcohol dependence syndrome (ADS) the study of selected immunological parameters (percentage of peripheral blood lymphocytes CD3+, CD4+, CD8+, CD19+; lymphocyte transformation without and with phytohemagglutinin (PHA), concanavalin A (Con A), pokeweed mitogen (PWM); chemiluminescence of peripheral blood granulocytes stimulated with phorbol myristate acetate (PMA)) and Instytut Mérieux' skin tests (Multitest CMI) were performed. The results of immunological parameters were connected with activity of liver enzymes (aspartate aminotransferase (ASP), alanine aminotransferase (ALT), gamma-glutamyltransferase (GGT). The differences of reactivity of immune system in the tested groups of patients were observed.

Immunological parameters in patients suffering from alcohol-dependence syndrome.

Alcohol abuse is a major cause of abnormal liver development and activity. In addition to enzymatic malfunction, alcohol and its metabolites induce changes in the levels of some liver antigens, resulting in immunological disturbance. The purpose of the present study is to correlate the severity of liver function impairment with the length of alcohol abuse, in order to be able to use such tests as indicative of the severity of Alcohol Dependence Syndrome. Thirty-one alcohol abusers were allocated to three groups on the basis of the levels of their liver enzymes, and were tested for a variety of immunological parameters and skin reactions. The data indicate that even though not all immunological values measured differed significantly from the control values, in those that did (granulocytes, lymphocytes, CD4/CD8 ratio, C3, IgG, IgM and some skin positive reactions), the biggest difference was between the healthy volunteers and the group with the longest abuse period. It is suggested that changes in selected immunological parameters in alcohol abusers may indicate the severity of their liver dysfunction.

The effects of taxol (paclitaxel) on chemiluminescence of neutrophils, macrophages and J.774.2 cell line.

Taxol (paclitaxel) is a chemotherapeutic diterpene with promising anticancer activity that blocks cell division by preventing microtubule depolymerization. Furthermore, recent studies have shown that taxol has other intracellular effects that may contribute to its effect, particularly in macrophages. The signal transduction mechanisms by which taxol stimulates macrophages to anticancer activity are not clear. The purpose of this study was to determine the effect of taxol on chemiluminescence (an indicator of the production of free radicals) of neutrophils, macrophages and murine macrophage J.774.2 cells. The chemiluminescence was measured in the presence of taxol and/or phorbol myristate acetate (PMA) as a stimulant. Taxol stimulated chemiluminescence (without PMA) of neutrophils and macrophages but not of J.774.2 cells, and modulated chemiluminescence of the cells stimulated with PMA.

Inhibition of neutrophils' chemiluminescence by ethanol extract of propolis (EEP) and its phenolic components.

Ethanol extract of propolis (EEP), a natural beehive product, has been known for centuries for a variety of beneficial traditional medical properties, among which an anti-inflammatory effect is a major one. Now that most of its components have been isolated and recently identified, we tested 19 of them (all phenolic compounds) for their degree of anti-inflammatory activity. This was performed by evaluating the luminol-enhanced chemiluminescence, formed after their scavenging free radicals, generated by neutrophils that had been stimulated by phorbol myristate acetate. Caffeic-acid-phenylethyl-ester abolished the chemiluminescence completely at a concentration of 10 microM, while three flavone derivatives and three flavonols (galangin, kaempferol and kaempferid) diminished this chemiluminescence by 73-93% at the same concentration. These results indicate that some of the phenolic components of the ethanol extract of propolis are its active components in exerting its renowned anti-inflammatory activity.

[Evaluation of selected immunological and biochemical parameters of blood in alcohol dependency]. / Ocena wybranych parametrów immunologicznych i biochemicznych krwi w zespole zalenosci alkoholowej.

The aim of this study was evaluation of selected immune humoral indicators in connection with biochemical parameters of blood in 28 alcohol dependent men. Increased activity of liver enzymes: AspAT in 53,6% of patients, A1AT in 46,4%, GGTP in 25% were found. Macrocytosis in 29% of patients was observed. There were also abnormal changes of immune proteins concentration: IgM, IgG, IgA, C3, C4 respectively in 60,7%, 46,4%, 21,4%, 42,9%, 10,7% of patients. In the group of patients with normal values of AspAT, GGTP, MCV; abnormal levels of humoral indicators concentration were observed. The correlation between immune proteins concentration and biochemical parameters was not found. The authors conclude that changes of determined immunological parameters may be used as prognostic indicators in disturbances in the alcoholics' immunity system.

Inhibition of nitric oxide (NO.) production in murine macrophages by flavones.

The effect of flavone (2-phenylbenzopyran-4-one) and three amino-substituted flavones on the production of nitrite by murine activated peritoneal macrophages was studied in vitro. Activated peritoneal macrophages obtained from mice pre-treated with concanavalin A (Con A) (in vivo), after exposure in vitro to lipopolysaccharide (LPS) at a concentration of 100 ng/ml, produced nitrite (20.3 +/- 2.5 nmol/10(6) cells), as measured after 24 hr by the Griess reaction. Stimulation of production of nitrite was inhibited by NG-monomethyl-L-arginine, suggesting that nitrite was formed via nitric oxide (NO.) as a product of metabolism of arginine. Stimulation was inhibited by flavone and the aminoflavones (20-100 microM). 3'-amino-4'-hydroxyflavone was the most potent inhibitor of nitrite production. Genistein (5,7-dihydroxy- 3-(4-hydroxy-phenyl)-4H-1-benzopyran-4-one) also inhibited production of nitrite, by a mechanism that appears not to involve protein tyrosine kinases. These results suggest that the flavones can modulate the immune responses and the inflammatory reactions by controlling production of nitric oxide.

Modulation of luminol-dependent chemiluminescence of murine macrophages by flavone and its synthetic derivatives.

The effect of flavone (CAS 525-82-6, 2-phenylbenzopyran-4-one, 1), flavone-8-acetic acid (CSA 87626-55-9, FAA, 2) and 10 substituted flavones on the luminol-dependent chemiluminescence of murine macrophages was studied in vitro. The synthetic derivatives were variously substituted with halo, nitro, amino, hydroxy and methoxy substituents in the 3' and 4' positions. Chemiluminescence was used in this study as an indicator for the production of reactive oxygen species by macrophages, stimulated in vitro by phorbol myristate acetate (PMA). All flavones except FAA (2) showed more than 20% inhibition at 10 mumol/l or 100 mumol/l. 3'-Amino-4'-hydroxyflavone (8) was the most potent inhibitor. The IC50s for inhibition of chemiluminescence were 4.2 +/- 1.1 mumol/l, 5.0 +/- 1.0 mumol/l and 3.3 +/- 1.4 mumol/l for resident, elicited and LPS-Poly I:C-primed macrophages, respectively. Small but statistically significant enhancements of chemiluminescence were caused by low concentrations of flavone (1), FAA (2) and 4'-methoxyflavone (6). These results suggest that modulation of the chemiluminescent capacity of macrophages depends on the nature of the substituents and the concentration of the flavones.

Structure-activity relationship in the ability of flavonols to inhibit chemiluminescence.

Fourteen flavonoids were evaluated for their ability to inhibit chemiluminescence, either of neutrophils that had been briefly exposed to both luminol and phorbol-myristate acetate or to an enzymatic system with H2O2, luminol and horseradish peroxidase. Using chemiluminescence as the quantitative parameter, it can be concluded that the hydroxyl group in position 3 of the flavonols is vital for their inhibitory effect, and that two hydroxyl groups on the phenyl ring are optimal for such an effect. It was also noted that the C2-C3 double bond is essential for the flavonols' anti-oxidative effect. It is suggested that the ability of flavonols to suppress chemiluminescence is reciprocally correlated with their lipophilicity.

Synergistic effect of ethanolic extract of propolis and antibiotics on the growth of staphylococcus aureus.

Ethanolic extract of propolis (EEP), known to possess marked antibacterial activity, was incubated with 8 different common antibiotics in culture medium containing a fixed amount of a standard strain of Staphylococcus aureus. The antibiotic compounds used were: penicillin G, doxycycline, streptomycin, cloxacillin, chloramphenicol, cefradine, ampicillin and polymyxin B. They were used in varying levels, ranging between 0.000005-125.0 micrograms/ml or units, resp. Firstly, their minimal inhibitory concentrations were established in the absence of EEP, than EEP was added in concentrations up to 600 micrograms/ml. EEP had a marked synergistic effect on the antibacterial activity of streptomycin and cloxacillin, and a moderate synergistic effect on the others, except ampicillin.

Modulating luminol-dependent chemiluminescence of neutrophils by flavones.

The effect of 14 flavones on luminol-dependent chemiluminescence of neutrophils was studied in vitro. Chemiluminescence was used in this study as an indicator for the production of a reactive oxygen species by neutrophils, stimulated by phorbol myristate acetate. While flavone-8-acetic acid, and most of the compounds tested, inhibited chemiluminescence, flavone and its 5-hydroxy-7-methoxy derivatives enhanced it by up to 150%. The most active inhibitors of photon emission were the glycosides. These results indicate that lipophilicity and some structural determinants modulate the chemiluminescent capacity of neutrophils.

Effect of cinnamic and acrylic acids' derivatives on luminol-enhanced chemiluminescence of neutrophils.

Fourteen derivatives of cinnamic and acrylic acids were evaluated for their ability to modulate chemiluminescence, evoked by neutrophils that had been exposed to luminol and phorbol-myristate-acetate. Compounds with one or two hydroxyl groups on the phenyl ring demonstrated significant inhibition of the chemiluminescence, but this inhibition was diminished by methoxylation. Saturation of the double bond in the aliphatic chain of cinnamic acid at C6-enhanced the chemiluminescence to a small degree. All three acrylic acid derivatives demonstrated a marked inhibition of the luminol chemiluminescence, indicating that characteristics of the heterocyclic ring is of utmost importance in this activity.

Anti-oxidant property of ethanolic extract of propolis (EEP) as evaluated by inhibiting the chemiluminescence oxidation of luminol.

Ethanolic extract of propolis (EEP) has remarkable medical properties, including protection of mice against gamma irradiation. Its anti-oxidative effect has been attributed to its radical scavenging ability. This manuscript demonstrates the ability of increasing amounts of EEP to inhibit luminol-H2O2 chemiluminescence in vitro, and suggests that its anti-oxidative capacity is partly due to its high content of flavonoids.

The ability of ethanolic extract of propolis (EEP) to protect mice against gamma irradiation.

Ethanolic extract of propolis (EEP) was tested as a protective agent against gamma irradiation in mice. The mice were exposed to 6 Gy gamma irradiation from a 60Co source, and were treated intraperitoneally with EEP, administered before and after their irradiation. While the non-treated mice expired within 12 weeks, the mice that received a series of EEP treatments survived the irradiation, and their leucocyte count as well as their spleens' plaque-forming activity returned to normal. It is suggested that an antioxidant and a free radical scavenger in the EEP are responsible for the radiation protective effect of the extract of this natural product.