Pioglitazone as a modulator of the chemoresistance of renal cell adenocarcinoma to methotrexate.

Kidney cancer is one of the most lethal urological malignancies associated with a high risk of mortality. Recent studies have shown that several antidiabetic drugs may limit the risk of the growth of different types of cancer. Pioglitazone (PIO) belongs to a novel class of antidiabetic drugs called thiazolidinediones (TZDs), which are commonly used in the treatment of type 2 diabetes. This drug has been demonstrated to exert an inhibitory effect on cell growth in colon, prostatic, breast and pancreatic cancer lines. The aim of the present study was to assess the inhibitory effect of PIO on the proliferation of the renal adenocarcinoma cell line 769­P. In addition, the proapoptotic potential of combined treatment with PIO and methotrexate (MTX) was evaluated, as well as the impact of the above drugs on the cell cycle of the 769­P cells. The present study showed that PIO efficaciously inhibited the proliferation and viability of renal cancer cells, and it induced sub­G1 cell cycle arrest and a decrease in the number of cells in the G2 phase, which indicated cytotoxic activity. PIO also exhibited proapoptotic properties at the lowest dose applied (10 µM). Furthermore, combined therapy with PIO and MTX increased the sensitivity of tumor cells to MTX while at the same time this combined therapy did not exhibit a cytotoxic effect to normal kidney cells. In renal adenocarcinoma cells, the combination of the above cytostatic agent at the lowest dose administered (MTX, 5 µM) with the peroxisome proliferator­activated receptor Î³ agonist PIO exhibited better efficacy in triggering the process of apoptosis than that displayed by MTX alone.

Cytotoxic activity of methanolic fractions of different Marrubium spp. against melanoma cells is independent of antioxidant activity and total phenolic content.

The Marrubium genus (horehound) has proved to be an abundant source of biologically active compounds, but there is little knowledge about its potential anticancer activity. Moreover, some Marrubium species have not been the subject of study in this regard. In this study, we performed comparative analysis of phenolic acid (PhA) content and total phenolic content in fractions obtained from methanolic extracts of Marrubium vulgare L. (common horehound), Marrubium cylleneum Boiss. & Heldr. and Marrubium friwaldskyanum Boiss herbs. We examined the cytotoxicity of these fractions against a human melanoma cancer cell line (A375) and normal human skin fibroblasts (BJ) using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide test, cell cycle analysis and real-time monitoring of cell viability. We detected caffeic, p-coumaric, ferulic and gentisic acids among the PhAs. Although the extracts obtained demonstrated low total phenolic content and did not show significant antioxidative properties, the nonhydrolyzed PhA fraction exhibited cytotoxic activity against a human melanoma cancer cell line, without affecting normal fibroblasts. Both acidic and alkaline hydrolysis abolished this activity, indicating that the esterified forms of phenolic compounds caused the observed cytotoxic effects. Further investigation of these compounds may facilitate the development of novel drugs for cancer treatment.

Ethanol extracts of Allium sp. regulate cyclooxygenase-2 and E-cadherin expression in gastric cancer MKN74 cell line and enhance doxorubicin toxicity.

BACKGROUND: Gastric cancer (GC) remains one of the leading causes of cancer-related death. Its aetiology is multifactorial, but the major risk factor is a high in salt diet. During gastric carcinogenesis, cadherin-1 (CDH1) down-expression and cyclooxygenase 2 (COX2) overexpression may be observed. The intensity of these alterations contributes to the GC invasion, its metastases and poor prognosis. As the diet plays a significant role in the aetiology of GC, it is reasonable to include the nutritional chemoprevention agents. One of the plant genus demonstrating chemoprotective properties is Allium genus, which includes garlic. The relationship between CDH1 and COX2 in GC cells treated with Allium species extract has never been evaluated. METHODS: In this study, the MKN28 and MKN74 GC cell lines were treated with ethanol extracts of Allium angulosum L., Allium lusitanicum Lam., Allium sativum L. (from Malaysia and Poland), Allium tibeticum Rendle and Allium ursinum L. The cytotoxicity of the extracts and their influence on COX2 and CDH1 mRNA and protein expression were evaluated as well as their influence on doxorubicin's (DOX) efficacy - a drug that has been used in GC treatment. RESULTS: Among the tested species, ethanol extracts of A. sativum L. (Poland and Malaysia), A. tibeticum Rendle and A. ursinum L. influenced the levels of CDH1 and COX2, but only in the MKN74 cell line. Thus, it is possible that tumours with increased COX2 expression will be more susceptible to garlic treatment. Observed phenomenon was independent of Allium extract's toxicity. In comparison to DOX, tested extracts were more toxic. Moreover, A. sativum revealed synergistic effect with the drug. CONCLUSION: In conclusion, the results indicate the potential application of Allium genus to GC chemoprevention and treatment support through CDH restoration and COX2 downregulation. This issue needs further investigations as it might be used in clinics.

Edaravone presents antidepressant-like activity in corticosterone model of depression in mice with possible role of Fkbp5, Comt, Adora1 and Slc6a15 genes.

Chronic exposure to environmental-like stress leads to dysregulation of hypothalamic-pituitary-adrenal (HPA) axis and to appearance of oxidative stress, which is implicated in the development of depression-like behaviour. Edaravone (3-methyl-1-phenyl-2-pyrazoline-5-one) exhibits a neuroprotective effect attributed to the potent free radical scavenging. This study was designed to assess antidepressant-like activity of edaravone based on behavioural tests in the animal model of depression. Furthermore, to elucidate its mechanisms, the expression of Fkbp5, Comt, Adora and Slc6a15 genes involved in turnover of neurotransmitters was analysed. In order to evaluate the antioxidant features of edaravone, DNA's oxidative damage was determined. The mice were injected subcutaneously (sc) with 40 mg/kg corticosterone, chronically for 21 days. Paroxetine (10 mg/kg) (a selective serotonin reuptake inhibitor) and edaravone (10 mg/kg) were administered separately (ip) 30 min prior to the corticosterone injection. After 21-days of treatment with respective drugs, the mice were decapitated and the prefrontal cortex was rapidly dissected and used for determination of DNA's oxidative damage and the real-time PCR analysis. Edaravone ameliorated behavioural impairments in sucrose preference test (SPT) and forced swim test (FST). A possible role in Fkbp5, Comt, Adora1 and Slc6a15 genes' expression in mediating this effect is postulated. Both edaravone and paroxetine have no effect on corticosterone-induced DNA's oxidative damage.

Apigenin and hesperidin augment the toxic effect of doxorubicin against HepG2 cells.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common malignancies, with an increasing incidence. Despite the fact that systematic chemotherapy with a doxorubicin provides only marginal improvements in survival of the HCC patients, the doxorubicin is being used in transarterial therapies or combined with the target drug - sorafenib. The aim of the study was to evaluate the effect of natural flavonoids on the cytotoxicity of the doxorubicin against human hepatocellular carcinoma cell line HepG2. METHODS: The effect of apigenin and its glycosides - cosmosiin, rhoifolin; baicalein and its glycosides - baicalin as well as hesperetin and its glycosides - hesperidin on glycolytic genes expression of HepG2 cell line, morphology and cells' viability at the presence of doxorubicin have been tested. In an attempt to elucidate the mechanism of observed results, the fluorogenic probe for reactive oxygen species (ROS), the DNA oxidative damage, the lipid peroxidation and the double strand breaks were evaluated. To assess impact on the glycolysis pathway, the mRNA expression for a hexokinase 2 (HK2) and a lactate dehydrogenase A (LDHA) enzymes were measured. The results were analysed statistically with the one-way analysis of variance (ANOVA) and post hoc multiple comparisons. RESULTS: The apigenin and the hesperidin revealed the strongest effect on the toxicity of doxorubicin. Both flavonoids simultaneously changed the expression of the glycolytic pathway genes - HK2 and LDHA, which play a key role in the Warburg effect. Although separate treatment with doxorubicin, apigenin and hesperidin led to a significant oxidative DNA damage and double strand breaks, simultaneous administration of doxorubicin and apigenin or hesperidin abolished these damage with the simultaneous increase in the doxorubicin toxicity. CONCLUSION: The obtained results indicate the existence of a very effective cytotoxic mechanism in the HepG2 cells of the combined effect of doxorubicin and apigenin (or hesperidin), not related to the oxidative stress. To explain this synergy mechanism, further research is needed, The observed intensification of the cytotoxic effect of doxorubicin by this flavonoids may be a promising direction of the research on the therapy of hepatocellular carcinoma, especially in a chemoembolization.

Inhibition of glycolysis disrupts cellular antioxidant defense and sensitizes HepG2 cells to doxorubicin treatment.

Increased glucose consumption is a known hallmark of cancer cells. Increased glycolysis provides ATP, reducing agents and substrates for macromolecular synthesis in intensely dividing cells. Therefore, inhibition of glycolysis is one strategy in anticancer therapy as well as in improved efficacy of conventional anticancer chemotherapeutic agents. One such agent is doxorubicin (DOX), but the mechanism of sensitization of tumor cells to DOX by inhibition of glycolysis has not been fully elucidated. As oxidative stress is an important phenomenon accompanying DOX action and antioxidant defense is closely related to energy metabolism, the aim of the study was the evaluation of oxidative stress markers and antioxidant abilities of cancer cells treated with DOX while glycolysis is inhibited. HepG2 cells were treated with DOX and one of three glycolysis inhibitors: 2-deoxyglucose, dichloroacetate or 3-promopyruvate. To evaluate the possible interaction mechanisms, we assessed mRNA expression of selected genes related to energy metabolism and antioxidant defense; oxidative stress markers; and reduced glutathione (GSH) and NADPH levels. Additionally, glutamine consumption was measured. It was demonstrated that the chemotherapeutic agent and glycolysis inhibitors induced oxidative stress and associated damage in HepG2 cells. However, simultaneous treatment with both agents resulted in even greater lipid peroxidation and a significant reduction in GSH and NADPH levels. Moreover, in the presence of the drug and an inhibitor, HepG2 cells had a reduced ability to take up glutamine. These results indicated that cells treated with DOX while glycolysis was inhibited had significantly reduced ability to produce NADPH and antioxidant defenses.

Altered expression of genes involved in brain energy metabolism as adaptive responses in rats exposed to chronic variable stress; changes in cortical level of glucogenic and neuroactive amino acids.

Brain metabolism is closely associated with neuronal activity and enables the accurate synthesis and function of neurotransmitters. Although previous studies have demonstrated that chronic stress is associated with the overproduction of reactive oxygen species (ROS), which leads to oxidative stress and the disruption of glucose metabolism, the molecular mechanisms and cerebral gluconeogenesis in depression have not yet been completely elucidated. In order to examine this subject, the present study evaluated changes in the expression of selected genes involved in the glycolytic pathway and the levels of glucogenic and neuroactive amino acids in the brain of rats exposed to chronic variable stress. Male Wistar rats (50­55 days old, weighing 200­250 g) were divided into two groups: control and stressed, and the rats in the stressed group were exposed to stress conditions for 40 days. Depressive­like states were observed and recorded by measuring the body weight and forced swim test (FST). The mRNA levels of Slc2a3 (coding GLUT3) and Tfam (activator of mitochondrial transcription and a participant in mitochondrial genome replication) were markedly increased, while a decrease in the expression of Ldhb and GAPDH was also observed. These modifications were associated with the redirection of glucose metabolism to appropriate defensive pathways under chronic stress conditions, and an increased ability to maintain mitochondrial function as potential adaptive responses. A marked reduction of glucogenic and neuroactive amino acids levels indicate the support of energy metabolism by stimulation of the gluconeogenesis pathway. The findings of the present study provide a novel insight into the molecular and biochemical events that impact the development of depression under chronic stress conditions, and they may identify novel targets for therapeutic intervention.

Novel 2,3­disubstituted 1,3­thiazolidin­4­one derivatives as potential antitumor agents in renal cell adenocarcinoma.

Cancer represents one of the main causes of mortality in developed countries. In particular, the overall survival of patients with renal cell carcinoma (RCC) remains poor and the available cytostatic agents are insufficient. Therefore, there is an urgent requirement to identify more effective and safer anticancer drugs. Recently, the evaluation of antitumor activity appeared to be promising for thiazolidinone derivatives. The present study presents the synthesis and the cytotoxicity assays of 1,3­thiazolidin­4­ones. The newly synthesized substances were screened in vitro against selected cancer human renal cell adenocarcinoma cells (769­P), human hepatoblastoma­derived cells (HepG2) and normal green monkey kidney cells (GMK) as a reference cell line. N­[2­(4­methylphenyl)­4­oxo­1,3­thiazolidin­3­yl]acetamide and N­[2­(4­methylphenyl)­4­oxo­1,3­thiazolidin­3­yl]benzamide displayed significant antiproliferative activity towards 769­P. To elucidate the mechanisms of the cytotoxic actions, additional studies on the cell cycle and apoptosis were performed. The aforementioned compounds were responsible for G1 cell cycle arrest and the decrease in cell distribution in the G2 phase in a dose­dependent manner, which prevents mitotic divisions of the 769­P cells. In addition, these novel 2,3­disubstituted 1,3­thiazolidin­4­ones slightly induced apoptosis in 769­P in a dose­dependent manner. It was hypothesized that the 4­methylphenyl group at position 2 of the thiazolidin­4­one scaffold may be regarded as a promising moiety for further development of this group of compounds. Therefore, benzamide moiety appeared to be crucial for triggering cells to apoptotic cell death.

Effect of Shot Peening on the Mechanical Properties and Cytotoxicity Behaviour of Titanium Implants Produced by 3D Printing Technology.

Structural discontinuities characterize the implants produced directly from metal powders in 3D printing technology. Mainly, the surface defects should be subjected to procedures associated with surface layer modification (likewise shot peening) resulting in the increase of the implant service life maintaining optimal biocompatibility. Therefore, the purpose of the present study was to investigate the effect of type of shot used for the peening process on the Ti-6Al-4V implants functional properties as well as the biological properties. The components were produced by DMLS (direct metal laser sintering) additive technology. The surfaces of titanium specimens have been subjected to the shot peening process by means of three different shots, i.e., CrNi steel shot, crushed nut shells, and ceramic balls shot. Then, the specimens have been subjected to profilometric analysis, microhardness tests, and static strength testing as well as to the assessment of biocompatibility in respect of cytotoxicity using human BJ fibroblasts. The shot peening process causes the strengthening of surface layer and the increase of strength parameters. Furthermore, the test results indicate good biocompatibility of surfaces being tested, and the effect of shot peening process on the titanium alloy cytotoxicity is acceptable. At the same time, most favourable behaviour in respect of cytotoxicity has been found in the case of surfaces modified by means of ceramic balls > nut shells > CrNi steel shot correspondingly.

Resveratrol Limits Lipogenesis and Enhance Mitochondrial Activity in HepG2 Cells.

PURPOSE: The aim of this study was to evaluate the effect of resveratrol on de novo lipogenesis in HepG2 cells caused by high glucose concentrations. Increased lipogenesis in the liver is the main reason for the development of nonalcoholic fatty liver disease (NAFLD) - currently one of the most common chronic liver diseases. In developed countries, this disease is mostly associated with nutritional disorders, resulting from the increasing consumption of monosaccharides. Resveratrol is a natural polyphenol with a promising potential for NAFLD treatment. METHODS: The steatosis of HepG2 cells was visualized using the intracellular lipid staining by Nile Red dye with a fluorescence microscope. This study also evaluated the effect of resveratrol on the mitochondrial activity (MitoTracker Green staining), dsDNA (Hoechst 33342 staining) and the viability of HepG2 cells treated with high glucose concentrations (25 and 33 mM). RESULTS: Current study showed that high glucose concentrations induced fat-overloading in HepG2 cells (microvacuolar steatosis occurred in most of the cells). Resveratrol (20 µM) limits the steatosis induction in HepG2 cells by glucose and increased the mitochondrial activity of cells. Resveratrol did not affect the viability of HepG2 cells. CONCLUSION: This beneficial effect could be helpful in the treatment of NAFLD.

Withdrawal of caffeine after its chronic administration modifies the antidepressant-like activity of atypical antidepressants in mice. Changes in cortical expression of Comt, Slc6a15 and Adora1 genes.

RATIONALE: Depressed patients often present increased consumption of caffeine. OBJECTIVES: We aimed to investigate the effects of chronic treatment with caffeine (5 mg/kg, twice daily for 14 days) on the activity of single, ineffective doses of agomelatine (20 mg/kg) or mianserin (10 mg/kg) given on day 15 alone or simultaneously with caffeine. METHODS: We used the forced swim test (FST), tail suspension test (TST), and locomotor activity test in mice and quantitative real-time PCR analysis of the selected genes in the cerebral cortex (Cx). RESULTS: There were no changes in the immobility time between mice that received saline and caffeine for 14 days. Administration of agomelatine or mianserin on day 15 did not produce an antidepressant-like effect, but such effect was observed after administration of agomelatine or mianserin simultaneously with caffeine on day 15, in both mice that received saline and caffeine for 14 days. In mice treated with caffeine for 14 days, joint administration of agomelatine or mianserin and caffeine on day 15 decreased solute carrier family 6, member 15 (Slc6a15), messenger RNA (mRNA) level in the Cx, compared to the group which received only the respective antidepressant on this day. Moreover, in mice treated with caffeine for 14 days, joint administration of mianserin and caffeine on day 15 decreased adenosine A1 receptor (Adora1) and catechol-O-methyltransferase (Comt) mRNA level in the Cx, compared to the group which received mianserin without caffeine on this day. CONCLUSIONS: Withdrawal of caffeine after its chronic intake can modify the activity of antidepressants. Adora1, Slc6a15, and Comt may be involved in the antidepressant-like effect observed after joint administration of caffeine and mianserin or agomelatine, following chronic treatment with caffeine.

Chronic Variable Stress Is Responsible for Lipid and DNA Oxidative Disorders and Activation of Oxidative Stress Response Genes in the Brain of Rats.

Chronic environmental stress is associated with reactive oxygen species (ROS) overproduction and the pathogenesis of depression. The purpose of this study was to evaluate biochemical and molecular changes associated with ROS generation in the brains of rats submitted to chronic variable stress. Male Wistar rats (50-55 days old, weighing 200-250 g) were divided in two groups (n = 10): control and stressed. Rats in the stressed group were exposed to stress conditions for 40 days. The animals were decapitated and the brain samples were collected. In prefrontal cortex, we measured the following biochemical parameters: lipid peroxidation and concentration of glutathione-GSH, GSSG, GSH/GSSG ratio, glutathione peroxidase, and glutathione reductase activities. In the hippocampus marker of DNA, oxidative damage and expression of DNA-repairing genes (Ogg1, MsrA) and gene-encoding antioxidative transcriptional factor (Nrf2) were determined. The results demonstrate indirect evidence of ROS overproduction and presence of oxidative stress. They also reveal disruption of oxidative defense systems (decreased GR activity, diminished GSH/GSSG ratio, and decreased Nrf2 expression) and activation of the oxidative DNA repair system (increased Ogg1 and MsrA expression). Together, the presented data suggest that independent activation of oxidative stress response genes occurs in chronic variable stress conditions.

Chronic treatment with caffeine and its withdrawal modify the antidepressant-like activity of selective serotonin reuptake inhibitors in the forced swim and tail suspension tests in mice. Effects on Comt, Slc6a15 and Adora1 gene expression.

Recent preclinical and clinical data suggest that low dose of caffeine enhances the effects of common antidepressants. Here we investigated the effects of chronic administration of caffeine (5mg/kg, twice daily for 14days) and its withdrawal on day 15th on the activity of per se ineffective doses of fluoxetine (5mg/kg) and escitalopram (2mg/kg) given on day 15th. We found decreased immobility time in the forced swim and tail suspension tests in mice in which caffeine was administered simultaneously with antidepressants on day 15th following a 14-day caffeine treatment and no alterations in the spontaneous locomotor activity. A decrease in the level of escitalopram and an increase in the level of caffeine in serum were observed after concomitant administration of these compounds, while the joint administration of caffeine and fluoxetine was not associated with changes in their levels in serum or brain. Caffeine withdrawal caused a decrease in Adora1 mRNA level in the cerebral cortex (Cx). Administration of escitalopram or fluoxetine followed by caffeine withdrawal caused an increase in this gene expression, whereas administration of escitalopram, but not fluoxetine, on day 15th together with caffeine caused a decrease in Adora1 mRNA level in the Cx. Furthermore, antidepressant-like activity observed after joint administration of the tested drugs with caffeine was associated with decreased Slc6a15 mRNA level in the Cx. The results show that withdrawal of caffeine after its chronic intake may change activity of antidepressants with concomitant alterations within monoamine, adenosine and glutamate systems.

The beneficial effects of resveratrol on steatosis and mitochondrial oxidative stress in HepG2 cells.

Nonalcoholic fatty liver disease (NAFLD) is currently one of the most common chronic liver diseases, especially in developed countries. One group of substances with a potential use in the treatment of NAFLD are plant polyphenols, represented by resveratrol. The aim of this study was to evaluate the effect of resveratrol on steatosis and oxidative stress in HepG2 cells. The steatosis of cells was carried out using free fatty acids: oleic or palmitic acid and their mixtures. Steatosis was visualized using the intracellular lipid staining by Nile Red dye with a fluorescence microscope. This study also determined the viability of cells and mitochondrial membrane potential. The current study showed that fatty acids and their mixtures induced fat overloading in HepG2 cells. In the group of cells incubated with oleic acid (OA), observed changes were moderate with prevailing micro-vesicular steatosis. In case of cells incubated with palmitic acid (PA) and the mixtures of fatty acids, micro- and macro-vacuolar steatosis occurred in most of the cells. Resveratrol decreased steatosis in HepG2 cells induced by OA, PA, as well as their mixtures, and in most of experimental groups did not reduce cells viability. Resveratrol reduced the oxidative stress in HepG2 cells treated with fatty acids mixtures.

Evaluation of the phytochemical composition and protective activities of methanolic extracts of Centaurea borysthenica and Centaurea daghestanica (Lipsky) Wagenitz on cardiomyocytes treated with doxorubicin.

Centaurea L. is a genus of the family Asteraceae that comprises over 600 taxa. Representatives of the Centaurea genus were used as natural medications for many diseases. Methanolic-aqueous extracts from aerial parts of two Centaurea species: C. borysthenica Gruner and C. daghestanica (Lipsky) Wagenitz were studied for their polyphenolic composition and potential protective effect on cardiomyocytes treated with doxorubicin. Effectiveness of doxorubicin in cancer therapy is limited by a dose-dependent cardiotoxicity. Oxidative stress is a widely recognized mechanism of this phenomenon. One of the most important strategies has been an application of drug together with antioxidant agents. A cardioprotective effect of selected extracts of Centaurea species was suspected in this study. Cell viability, oxidative stress, and mitochondrial membrane potential analyses showed protective activity of the methanolic extract of C. borysthenica and C. daghestanica on rat cardiomyocytes treated with doxorubicin. Although C. borysthenica is more effective as a cardiomyocyte protective agent, in higher concentrations it weakened the drug activity. C. daghestanica extract did not change the doxorubicin efficacy in the evaluated experiment. Interestingly, both tested extracts were cytotoxic for myeloma cells. The detected antioxidant activity of the studied extracts can be used in the prevention of doxorubicin-induced cardiotoxicity.

The Positive Synergism of CPT and MK-801 in Behavioral Tests and in Reduction of Environmental Stress and Redox Signaling Changes in Mice Cerebral Cortex.

BACKGROUND: Depressive disorders are associated with oxidative stress. Therefore, it is interesting if antidepressants can affect redox equilibrium and signaling. The first step of our study was to determine the influence of the adenosine system on the antidepressant-like activity of noncompetitive antagonist of the NMDA (N-methyl-d-aspartate) receptor complex - dizocilpine (MK- 801). To this aim, two behavioral tests commonly used to assess the antidepressant capability of drugs - the forced swim test (FST) and tail suspension test (TST), were performed. Locomotor activity was estimated to verify and exclude false positive/negative results in the FST and TST. To examine whether antidepressants affect redox equilibrium, we have investigated lipid peroxidation products (LPO), GSH (glutathione), GSSG (glutathione disulfide), NADP+ (nicotinamide adenine dinucleotide phosphate) and NADPH (reduced nicotinamide adenine dinucleotide phosphate) in the cerebral cortex of mice following administration of CPT (8-cyclopentyl-1,3-dimethylxanthine) and MK-801 (dizocilpine) under environmental stress conditions. METHOD: The experiments were carried out using male Albino Swiss mice (25-30 g). The drugs were administered ip., alone and simultaneously, 60 min before tests. RESULTS: The behavioural tests results showed that CPT (3 mg/kg) potentiated the antidepressant-like activity of MK-801 (0.05 mg/kg) and the observed effects were not due to the increase in mice locomotor activity. Positive synergism of CPT and MK-801 in reduction of environmental stress conditions was revealed. In this group an increase in GSH and GSSG without changes in GSH/GSSG ratio and reduction of LPO was found. The level of lipid peroxidation products was also decreased in group receiving CPT and MK-801 separately. CONCLUSION: Examined antidepressant agents may increase antioxidant defences however further studies are needed with different range of time.

An outline of main factors of drug resistance influencing cancer therapy.

Drug resistance in cancer therapy is a multifactorial phenomenon that determines remission or progression. It is known that resistance to used anticancer drugs may be the consequence of drug transport to the cell or intracellular distribution. It may also be the result of its molecular target structural change, apoptosis inhibition or increase in some enzymes activity, e.g. pentose phosphate pathway enzymes. Intrinsic (pre-existed) drug resistance is related to the phenotype of cancer as well as normal cells. Acquired, after partial administration of chemotherapy, type of drug resistance in addition to the starting phenotype is closely linked to the development of new more aggressive clones and adaptive processes. In both, the intrinsic and acquired resistance, role play also mutations. These may be partially spontaneous, but in terms of acquired resistance, they are mostly induced by the exposure to the drugs. The article mentions some traditional mechanisms related to the acquisition of resistance by cancer cells during therapy, through the protein transporters, apoptosis deregulation, angiogenesis and the impact of the tumour microenvironment. We focused however on some more alternative ways of therapy resistance, such as, hypoxia and tumour acidification, cancer stem cells (CSCs), exosomes and radiotherapy resistance. A concise summary of the drug resistance presented in the paper may be an important aspect in studies to increase the effectiveness of cancer therapies.

Tirapazamine has no Effect on Hepatotoxicity of Cisplatin and 5-fluorouracil but Interacts with Doxorubicin Leading to Side Changes in Redox Equilibrium.

Tirapazamine is a hypoxia-activated prodrug which was shown to exhibit up to 300 times greater cytotoxicity under anoxic in comparison with aerobic conditions. Thus, the combined anticancer therapy of tirapazamine with a routinely used anticancer drug seems to be a promising solution. Because tirapazamine undergoes redox cycle transformation in this study, the effect of tirapazamine on redox hepatic equilibrium, lipid status and liver morphology was evaluated in rats exposed to cisplatin, doxorubicin and 5-fluorouracil. Rats were intraperitoneally injected with tirapazamine and a particular cytostatic. The animals were killed, and blood and liver were collected. Hepatic glucose, total cholesterol, triglycerides, NADH, NADPH glutathione and the activity of glucose-6-phosphate dehydrogenase were determined. Liver morphology and the immune expression of HMG-CoA-reductase were also assessed. Glucose, total cholesterol, triglycerides, bilirubin concentrations and the activity of aspartate and alanine aminotransferases were determined in the plasma. Tirapazamine displayed insignificant interactions with cisplatin and 5-fluorouracil referring to hepatic morphology and biochemical parameters. However, tirapazamine interacts with doxorubicin, thus leading to side changes in redox equilibrium and lipid peroxidation, but those effects are not severe enough to exclude that drug combination from further studies. Thus, tirapazamine seems to be a promising agent in successive studies on anticancer activity in similar schedules.

The effect of a number of H9C2 rat cardiomyocytes passage on repeatability of cytotoxicity study results.

The embryonic cardiomyocyte cell line H9C2 is commonly used in numerous in vitro studies, including cardiotoxicity analyses of new drugs. So far no results were published for studies on cell parameters variability during the cell line ageing process. For this reason the aim of the study was to evaluate the effect of a number of H9C2 rat embryonic cardiomyocytes passages on repeatability of study results for selected cytotoxicity parameters, with doxorubicin as a model toxic agent. The cultures were passaged twenty-five times. Cells from passage 1, 5, 10, 15, 20 and 25 were treated with doxorubicin for 24 h. Then drug cytotoxicity was evaluated with the MTT test and additionally the nuclear factor erythroid 2-related factor (Nrf2) gene expression was examined. The analysis of oxidative stress intensity and cell morphology was also assessed. The microscopic appearance of cells indicates that untreated cardiomyocytes morphology changes as well as sensitivity to toxic effects increases with the number of passages. Also an increase in oxidative stress in cells occurs with further passaging of cardiomyocytes. Statistical significance of differences in conducted tests results depended on doxorubicin concentration but in many cases the H9C2 line was found to be a reliable in vitro model only for the first five passages. For this reason it is important to take into consideration that further culturing of cardiomyocytes may not ensure repeatability of study results due to the culture ageing.

The differential effects of green tea on dose-dependent doxorubicin toxicity.

BACKGROUND: Doxorubicin (DOX) is an anticancer drug displaying cardiac and hepatic adverse effects mostly dependent on oxidative stress. Green tea (GT) has been reported to play a protective role in diseases resulting from oxidative stress. OBJECTIVE: The objective of this study was to evaluate if GT protects against DOX-induced oxidative stress, heart and liver morphological changes, and metabolic disorders. METHODS: Male Wistar rats received intraperitoneal injection of DOX (1.0 or 2.0 mg/kg b.w.) for 7 weeks or concomitantly GT extract soluble in drinking water. RESULTS: There were multidirectional effects of GT on blood metabolic parameters changed by DOX. Among all tested biochemical parameters, statistically significant protection of GT against DOX-induced changes was revealed in case of blood fatty acid-binding protein, brain natriuretic peptide, and superoxide dismutase. CONCLUSION: DOX caused oxidative stress in both organs. It was inhibited by GT in the heart but remained unchanged in the liver. DOX-induced general toxicity and histopathological changes in the heart and in the liver were mitigated by GT at a higher dose of DOX and augmented in rats treated with a lower dose of the drug.