[Alpha-1 Antitrypsin Affects U0126-Induced Cytotoxicity in Colon Cancer Cell Line (HCT116)].

Alpha-1-antitrypsin (AAT), an acute phase protein, is the principal circulatory anti-protease. This multifunctional protein is encoded by the SERPINA1 gene. Although AAT was recognised as a potential tumour marker, its role in cancer biology remains unknown. Given that it has been demonstrated that AAT has an anti-apoptotic property against non-malignant cells, we aimed to investigate whether AAT affects apoptosis in a colon cancer cell line (HCT116). The presence of AAT in the HCT116 cell culture antagonized cytotoxicity of blockers of MEK1/2, PI3K/Akt pathways as well as NF-κB. The dominantly recovered cell viability was observed in the co-treatment with MEK1/2 inhibitor U0126. In addition, it was revealed that AAT almost completely abolished U0126-induced apoptosis through maintenance of the autophagy process. Our study revealed for the first time that the observed cyto-protection triggered by AAT was accompanied by sustained autophagy which opposed apoptosis. These results may contribute to understanding of the role of AAT in cancer development and evaluation of efficacy of cancer therapy.

Cell-type dependent response of melanoma cells to aloe emodin.

Intrinsic characteristics of melanoma cells such as expression of inducible nitric oxide synthase (iNOS), redox status, and activity of signaling pathways involved in proliferation, differentiation and cell death define the response of the cells to the diverse treatments. In this context we compared the effectiveness of herbal antaquinone aloe emodin (AE) against mouse B16 melanoma and human A375, different in initial activity of ERK1/2, constitutive iNOS expression and basal level of reactive oxygen species (ROS). Both cell lines are sensitive to AE treatment. However, while the agent induces differentiation of B16 cells toward melanocytes, in A375 cells promoted massive apoptosis. Differentiation of B16 cells, characterized by enhanced melanin production and tyrosinase activity, was mediated by H(2)O(2) production synchronized with rapid p53 accumulation and enhanced expression of cyclins D1 and D3. Caspase mediated apoptosis triggered in A375 cells was accompanied with Bcl-2 but not iNOS down-regulation. In addition, opposite regulation of Akt-ERK1/2 axis in AE treated B16 and A375 cells correlated with different outcome of the treatment. However, AE in a dose-dependent manner rescued both B16 and A375 cells from doxorubicin- or paclitaxel-induced killing. These data indicate that caution is warranted when AE is administrated to the patients with conventional chemotherapy.

Anti-tumor effect of Coriolus versicolor methanol extract against mouse B16 melanoma cells: in vitro and in vivo study.

Numerous studies have shown immunostimulatory and anti-tumor effects of water and standardized aqueous ethanol extracts derived from the medicinal mushroom, Coriolus versicolor, but the biological activity of methanol extracts has not been examined so far. In the present study we investigated the anti-tumor effect of C. versicolor methanol extract (which contains terpenoids and polyphenols) on B16 mouse melanoma cells both in vitro and in vivo. In vitro treatment of the cells with the methanol extract (25-1600 microg/ml) reduced melanoma cell viability in a dose-dependent manner. Furthermore, in the presence of the methanol extract (200 microg/ml, concentration IC(50)) the proliferation of B16 cells was arrested in the G(0)/G(1) phase of the cell cycle, followed by both apoptotic and secondary necrotic cell death. In vivo methanol extract treatment (i.p. 50 mg/kg, for 14 days) inhibited tumor growth in C57BL/6 mice inoculated with syngeneic B16 tumor cells. Moreover, peritoneal macrophages collected 21 days after tumor implantation from methanol extract-treated animals exerted stronger tumoristatic activity ex vivo than macrophages from control melanoma-bearing mice. Taken together, our results demonstrate that C. versicolor methanol extract exerts pronounced anti-melanoma activity, both directly through antiproliferative and cytotoxic effects on tumor cells and indirectly through promotion of macrophage anti-tumor activity.

Interleukin-17 stimulates inducible nitric oxide synthase-dependent toxicity in mouse beta cells.

The influence of the proinflammatory cytokine interleukin (IL)-17 on inducible nitric oxide (NO) synthase (iNOS)-mediated NO release was investigated in the mouse insulinoma cell line MIN6 and mouse pancreatic islets. IL-17 markedly augmented iNOS mRNA/protein expression and subsequent NO production induced in MIN6 cells or pancreatic islets by different combinations of interferon-gamma, tumor necrosis factor-alpha, and IL-1beta. The induction of iNOS by IL-17 was preceded by phosphorylation of p38 mitogen-activated protein kinase (MAPK), and inhibition of p38 MAPK activation completely abolished IL-17-stimulated NO release. IL-17 enhanced the NO-dependent toxicity of proinflammatory cytokines toward MIN6 cells, while IL-17-specific neutralizing antibody partially reduced the NO production and rescued insulinoma cells and pancreatic islets from NO-dependent damage induced by activated T cells. Finally, a significant increase in blood IL-17 levels was observed in a multiple low-dose streptozotocin model of diabetes, suggesting that T cell-derived IL-17 might be involved in NO-dependent damage of beta cells in this disease.

Aloe emodin decreases the ERK-dependent anticancer activity of cisplatin.

The present study describes the ability of an anthraquinone derivative aloe emodin (AE) to reduce the cytotoxic activity of the platinum(II)-based anticancer agent cisplatin toward murine L929 fibrosarcoma and C6 glioma cell lines. The protective effect of AE was demonstrated by MTT and crystal violet assays for cell viability, and involved supression of cisplatin-induced apoptosis and necrosis, as assessed by lactate dehydrogenase release and flow cytometric analysis of DNA fragmentation or phosphatidylserine exposure. Cell-based ELISA and Western blot analysis revealed that AE abolished cisplatin-triggered activation of extracellular signal-regulated kinase (ERK) in tumor cells, while activation of c-Jun N-terminal kinase was not significantly altered. A selective blockade of ERK activation with PD98059 mimicked the protective effect of AE treatment in both tumor cell lines. Moreover, AE failed to protect tumor cells against the ERK-independent toxicity of the Pt(IV)-based complex tetrachloro(O,O-dibutyl-ethylenediamine-N,N'-di-3-propanoate)platinum(IV). Taken together, these data indicate that herbal anthraquinone AE can downregulate the anticancer activity of cisplatin by blocking the activation of ERK in tumor cells.

Anti-glioma action of aloe emodin: the role of ERK inhibition.

The effect of aloe emodin (AE), a herbal anthraquinone derivative, on the rat C6 glioma cell line was investigated. In addition to cell cycle block and caspasedependent apoptosis, AE led to the formation of intracytoplasmic acidic vesicles indicative for autophagic cell death. Moreover, differentiation of surviving cells toward the astrocytic lineage was confirmed by typical morphological changes and increased expression of glial fibrillary acidic protein (GFAP). AE did not affect the activation of mitogen-activated protein kinase p38, Jun-N-terminal kinase, or transcription factor NF-kappaB, but markedly inhibited the activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) in C6 cells. A selective inhibitor of ERK activation, PD98059, mimicked the effects of AE on glioma cell morphology and GFAP expression, but failed to induce either apoptosis or autophagy. Taken together, these results indicate that the anti-glioma action of AE involves ERK-independent induction of both apoptosis and autophagy, as well as ERK inhibition-mediated differentiation of glioma cells.

Aloe-emodin prevents cytokine-induced tumor cell death: the inhibition of auto-toxic nitric oxide release as a potential mechanism.

Aloe-emodin (AE) is a plant-derived hydroxyanthraquinone with potential anticancer activity. We investigated the ability of AE to modulate survival of mouse L929 fibrosarcoma and rat C6 astrocytoma cells through interference with the activation of inducible nitric oxide (NO) synthase (NOS) and subsequent production of tumoricidal free radical NO. Somewhat surprisingly, AE in a dose-dependent manner rescued interferon-gamma + interleukin-1-stimulated L929 cells from NO-dependent killing by reducing their autotoxic NO release. The observed protective effect was less pronounced in C6 cells, due to their higher sensitivity to a direct toxic action of the drug. AE-mediated inhibition of tumor cell NO release coincided with a reduction in cytokine-induced accumulation of transcription and translation products of genes encoding inducible NOS and its transcription factor IRF-1, while activation of NF-kappaB remained unaltered. These data indicate that the influence of AE on tumor growth might be more complex that previously recognized, the net effect being determined by the balance between the two opposing actions of the drug: its capacity to directly kill tumor cells, but also to protect them from NO-mediated toxicity.

Down-regulation of multiple low dose streptozotocin-induced diabetes by mycophenolate mofetil.

The new immunosuppressive agent mycophenolate mofetil (MMF) has been shown recently to exert a protective effects in certain animal models of autoimmunity, including diabetes in diabetes-prone bio-breeding (BB) rats. In the present study, the immunomodulatory potential of MMF was investigated in autoimmune diabetes induced by multiple low doses of streptozotocin (MLD-STZ) in genetically susceptible DA rats 20 mg STZ/kg body weight (b.w.) for 5 days] and CBA/H mice (40 mg STZ/kg b.w. for 5 days). In both species, short time treatment of animals with MMF (25 mg/kg) during the early development of the disease, as well as continuous MMF treatment, prevented the appearance of hyperglycaemia and inflammatory infiltrates in the pancreatic tissue. Moreover, clinical manifestations of diabetes were suppressed by application of the drug after the onset of clinical symptoms. Treatment with guanosine (1 mg/kg) in parallel with MMF completely reversed MMF activity in vivo, indicating that inhibition of inosine monophosphate dehydrogenase (IMPDH) was responsible for the observed suppressive effects. MMF-mediated protection from diabetes correlated with reduced ex vivo spontaneous spleen mononuclear cell (MNC) proliferation and defective adhesive cell interactions. MMF-treated animals also had lower local production of IFN-gamma, as well as IL-12 and nitric oxide (NO) production by peripheral tissues (spleen and peritoneal cells), compared to that in control diabetic groups, while IL-10 level was elevated. Together, these data demonstrate that MMF interferes with autoimmune process in streptozotocin-induced diabetes at multiple levels, including lymphocyte proliferation and adhesion, as well as pro/anti-inflammatory cytokine balance.

The content of total fatty acids in the serum of healthy children from the Zagreb area.

A study of the fatty acid content was performed by gas chromatography in the sera of 26 healthy children aged 3-13 years and in the sera of 7 children aged up to three years, both from the Zagreb area. The following parameters were calculated for each subject and each fatty acid individually: mmol/l, mol%, g/l, mass% and mean molecular mass. On the basis of the results, the values of the parameters for the whole group of subjects were calculated. The values of these parameters for some characteristic groups of fatty acids (saturated, unsaturated, monounsaturated and long-chain polyunsaturated fatty acids) were also calculated and their interrelationship is presented. The ratio of fatty acids according to age groups (0-1, 1-3 and 3-13 years) is also shown. Fatty acids C 18:2, C 16:0 and C 18:1 with 32.87 mol%, 22.56 mol% and 20.79 mol%, respectively, were most frequently observed in the children's sera, which correlated well with the generally known data. Essential fatty acids were present in the following amounts: C 20:4, 5.49 mol%; C 22:5, 0.26 mol%; and C 22:6, 0.82 mol%. The sera of the children aged up to one year were found to contain significantly higher amounts of short-chain fatty acids and lower amounts of C 18:2 and C 20:4 than the sera from children older than one year.

[The chemical war in Croatia]. / Kemijski rat protiv Hrvatske.

During the present war against the Republic of Croatia, chemical weapons have been used by the Yugoslav Federal Army (YFA) against both civilians and Croatian Army soldiers. The use of irritants was suspected (Vukovar, Bogdanovci and Vinkovci, October-November, 1991; Solin in the Split area, September, 1991) and proved (Velika Gorica in the Zagreb area, September, 1991; Cakovec, November, 1991) in many cases. The use of psychochemical incapacitating agents (Bilje near Osijek, July, 1991), as well as of psychostimulants in YFA own soldiers (Zadar, August, 1991) has been suspected on clinical findings or laboratory tests. The use of acetylcholnesterase inhibitors was proved in one aggressor's diversion (Zadar, Krusevo, July, 1991). Phosphorus from projectiles and fuming boxes caused poisoning and skin burns due to incineration (Vukovar, November, 1991). YFA used the civilian's fear of chemical and biological weapons, throwing untoxic substances all over the Croatian territory. Great ecocide problems have occurred with massive industry devastation (Sisak, Osijek, October, 1991-January, 1992), with enumerous amounts of toxic substances released into the soil and river aquatoria.