Mini-encyclopedia of mitochondria-relevant nutraceuticals protecting health in primary and secondary care-clinically relevant 3PM innovation.

Despite their subordination in humans, to a great extent, mitochondria maintain their independent status but tightly cooperate with the "host" on protecting the joint life quality and minimizing health risks. Under oxidative stress conditions, healthy mitochondria promptly increase mitophagy level to remove damaged "fellows" rejuvenating the mitochondrial population and sending fragments of mtDNA as SOS signals to all systems in the human body. As long as metabolic pathways are under systemic control and well-concerted together, adaptive mechanisms become triggered increasing systemic protection, activating antioxidant defense and repair machinery. Contextually, all attributes of mitochondrial patho-/physiology are instrumental for predictive medical approach and cost-effective treatments tailored to individualized patient profiles in primary (to protect vulnerable individuals again the health-to-disease transition) and secondary (to protect affected individuals again disease progression) care. Nutraceuticals are naturally occurring bioactive compounds demonstrating health-promoting, illness-preventing, and other health-related benefits. Keeping in mind health-promoting properties of nutraceuticals along with their great therapeutic potential and safety profile, there is a permanently growing demand on the application of mitochondria-relevant nutraceuticals. Application of nutraceuticals is beneficial only if meeting needs at individual level. Therefore, health risk assessment and creation of individualized patient profiles are of pivotal importance followed by adapted nutraceutical sets meeting individual needs. Based on the scientific evidence available for mitochondria-relevant nutraceuticals, this article presents examples of frequent medical conditions, which require protective measures targeted on mitochondria as a holistic approach following advanced concepts of predictive, preventive, and personalized medicine (PPPM/3PM) in primary and secondary care.

Biochemical Properties of Atranorin-Induced Behavioral and Systematic Changes of Laboratory Rats.

Atranorin (ATR) is a secondary metabolite of lichens. While previous studies investigated the effects of this substance predominantly in an in vitro environment, in our study we investigated the basic physicochemical properties, the binding affinity to human serum albumin (HSA), basic pharmacokinetics, and, mainly, on the systematic effects of ATR in vivo. Sporadic studies describe its effects during, predominantly, cancer. This project is original in terms of testing the efficacy of ATR on a healthy organism, where we can possibly attribute negative effects directly to ATR and not to the disease. For the experiment, 24 Sprague Dawley rats (Velaz, Únetice, Czech Republic) were used. The animals were divided into four groups. The first group (n = 6) included healthy males as control intact rats (♂INT) and the second group (n = 6) included healthy females as control intact rats (♀INT). Groups three and four (♂ATR/n = 6 and ♀ATR/n = 6) consisted of animals with daily administered ATR (10mg/kg body weight) in an ethanol-water solution per os for a one-month period. Our results demonstrate that ATR binds to HSA near the binding site TRP214 and acts on a systemic level. ATR caused mild anemia during the treatment. However, based on the levels of hepatic enzymes in the blood (ALT, ALP, or bilirubin levels), thiobarbituric acid reactive substances (TBARS), or liver histology, no impact on liver was recorded. Significantly increased creatinine and lactate dehydrogenase levels together with increased defecation activity during behavioral testing may indicate the anabolic effect of ATR in skeletal muscles. Interestingly, ATR changed some forms of behavior. ATR at a dose of 10 mg/kg body weight is non-toxic and, therefore, could be used in further research.

5-Fluorouracil Treatment of CT26 Colon Cancer Is Compromised by Combined Therapy with IMMODIN.

Due to the physiological complexity of the tumour, a single drug therapeutic strategy may not be sufficient for effective treatment. Emerging evidence suggests that combination strategies may be important to achieve more efficient tumour responses. Different immunomodulators are frequently tested to reverse the situation for the purpose of improving immune response and minimizing chemotherapy side effects. Immodin (IM) represents an attractive alternative to complement chemotherapy, which can be used to enhance the immune system after disturbances resulting from the side effects of chemotherapy. In the presented study, a model of CT26 tumor-bearing mice was used to investigate the effect of single IM or its combination with 5-fluorouracil (5-FU) on colon cancer cells. Our results highlight that the beneficial role of IM claimed in previous studies cannot be generalised to all chemotherapeutic drugs, as 5-FU toxicity was not increased. On the contrary, the chemotherapeutic anti-cancer efficacy of 5-FU was greatly compromised when combined with IM. Indeed, the combined treatment was significantly less effective regarding the tumour growth and animal survival, most probably due to the increased number of tumour-associated macrophages, and increased 5-FU cytotoxic effect related to kidneys and the liver.

Addition of palm olein to lard-supplemented diet indicates myocardial dysfunction and augments oxidative stress by authophagy-lysosome pathway in rats.

This study evaluated a prolonged effect of palm oil addition to lard-supplemented diet (PLD) on the oxidative status, lysosomal enzyme activities, markers of hepatotoxicity and basic lipid profile in female rats. Female Sprague-Dawley rats received PLD (10% of total fat: 7.5% from palm oil and 2.5% from lard), and the control group received lard-supplemented diet (2.5% fat) from 28 days of age for 14 weeks. Histopathological evaluation of the liver from animals fed the PLD showed slight steatosis and signs of mild chronic inflammation. Reduction of extramedullary hematopoiesis and an increased ratio of red/white pulp were observed in the spleen. PLD induced oxidative stress (evaluated in the liver, heart, spleen, muscle and kidney) evidenced by an increase in conjugated dienes and malondialdehyde in all tissues except the muscle; protein carbonyl derivatives were increased as well. The changes in the antioxidant enzyme activities in the evaluated tissues were ambiguous except for the prominent increase in the heart. Lysosomal enzyme activities showed a tendency to increase in the heart and kidney and to decrease in the muscle and spleen. The De Ritis ratio, which is a biomarker of hepatotoxicity, was higher in the heart from animals fed the PLD. The palm oil addition to the lard-supplemented diet-induced prominent oxidative stress, particularly in myocardial tissue with involvement of the authophagy-lysosome pathway.

Dietary Fat and Cancer-Which Is Good, Which Is Bad, and the Body of Evidence.

A high-fat diet (HFD) induces changes in gut microbiota leading to activation of pro-inflammatory pathways, and obesity, as a consequence of overnutrition, exacerbates inflammation, a known risk factor not only for cancer. However, experimental data showed that the composition of dietary fat has a greater impact on the pathogenesis of cancer than the total fat content in isocaloric diets. Similarly, human studies did not prove that a decrease in total fat intake is an effective strategy to combat cancer. Saturated fat has long been considered as harmful, but the current consensus is that moderate intake of saturated fatty acids (SFAs), including palmitic acid (PA), does not pose a health risk within a balanced diet. In regard to monounsaturated fat, plant sources are recommended. The consumption of plant monounsaturated fatty acids (MUFAs), particularly from olive oil, has been associated with lower cancer risk. Similarly, the replacement of animal MUFAs with plant MUFAs decreased cancer mortality. The impact of polyunsaturated fatty acids (PUFAs) on cancer risk depends on the ratio between ω-6 and ω-3 PUFAs. In vivo data showed stimulatory effects of ω-6 PUFAs on tumour growth while ω-3 PUFAs were protective, but the results of human studies were not as promising as indicated in preclinical reports. As for trans FAs (TFAs), experimental data mostly showed opposite effects of industrially produced and natural TFAs, with the latter being protective against cancer progression, but human data are mixed, and no clear conclusion can be made. Further studies are warranted to establish the role of FAs in the control of cell growth in order to find an effective strategy for cancer prevention/treatment.

Fluctuations of Histone Chemical Modifications in Breast, Prostate, and Colorectal Cancer: An Implication of Phytochemicals as Defenders of Chromatin Equilibrium.

Natural substances of plant origin exert health beneficiary efficacy due to the content of various phytochemicals. Significant anticancer abilities of natural compounds are mediated via various processes such as regulation of a cell's epigenome. The potential antineoplastic activity of plant natural substances mediated by their action on posttranslational histone modifications (PHMs) is currently a highly evaluated area of cancer research. PHMs play an important role in maintaining chromatin structure and regulating gene expression. Aberrations in PHMs are directly linked to the process of carcinogenesis in cancer such as breast (BC), prostate (PC), and colorectal (CRC) cancer, common malignant diseases in terms of incidence and mortality among both men and women. This review summarizes the effects of plant phytochemicals (isolated or mixtures) on cancer-associated PHMs (mainly modulation of acetylation and methylation) resulting in alterations of chromatin structure that are related to the regulation of transcription activity of specific oncogenes, which are crucial in the development of BC, PC, and CRC. Significant effectiveness of natural compounds in the modulation of aberrant PHMs were confirmed by a number of in vitro or in vivo studies in preclinical cancer research. However, evidence concerning PHMs-modulating abilities of plant-based natural substances in clinical trials is insufficient.

Metformin and melatonin improve histopathological outcome of NMU-induced mammary tumors in rats.

OBJECTIVE: Numerous reports showed inhibition of carcinogenesis after metformin (MF) and melatonin (MEL) administration. However, most in vivo studies used standard diet type, with relatively low fat content. As increase in fat intake may have a considerable impact on malignant transformation, we evaluated the effects of these two substances in a model of mammary carcinogenesis in rats fed a high-fat diet (10%). METHODS: Mammary tumors were induced by N-methyl-N-nitrosourea (NMU) in female rats of sensitive Sprague-Dawley strain. MF was administered in a diet (0.2%), MEL was administered in drinking water (20 mg/L). The chemoprevention was initiated 12 days prior to tumor initiation, both substances were administered through the termination of the experiment on 16th week after carcinogen application. Analysis of basic parameters of tumor growth, histopathological profile, and serum IGF-1 level were performed together with immunohistochemical detection of Ki67 (proliferation marker) and caspase-3 and BCL-2 (apoptosis markers) in mammary cancer cells. RESULTS: Although neither tumor incidence nor frequency were changed after MF and/or MEL administration, MF and MEL decreased high-grade/low-grade (HG/LG) tumor ratio. MEL decreased proliferation in mammary cancer cells; positive correlations between histological grade and Ki67 expressions were found after single administration of both MF and MEL. Serum IGF-1 levels were reduced to the level of intact rats in all groups receiving chemoprevention. CONCLUSIONS: MF and MEL administration did not inhibit growth of NMU-induced mammary tumors in rats in a significant manner but both substances ameliorated tumor histopathological profile. Surprisingly, combined treatment had no such effect.

Melatonin May Increase Anticancer Potential of Pleiotropic Drugs.

Melatonin (N-acetyl-5-methoxytryptamine) is not only a pineal hormone, but also an ubiquitary molecule present in plants and part of our diet. Numerous preclinical and some clinical reports pointed to its multiple beneficial effects including oncostatic properties, and as such, it has become one of the most aspiring goals in cancer prevention/therapy. A link between cancer and inflammation and/or metabolic disorders has been well established and the therapy of these conditions with so-called pleiotropic drugs, which include non-steroidal anti-inflammatory drugs, statins and peroral antidiabetics, modulates a cancer risk too. Adjuvant therapy with melatonin may improve the oncostatic potential of these drugs. Results from preclinical studies are limited though support this hypothesis, which, however, remains to be verified by further research.

Liver antioxidant and aerobic status improves after metformin and melatonin administration in a rat model of high-fat diet and mammary carcinogenesis.

Oxidative stress is involved in the development of various cancers. In the present study, the effect of long-term administration of peroral antidiabetic metformin and pineal hormone melatonin on liver antioxidant and aerobic status in female Sprague-Dawley rats carrying mammary tumors induced by N-methyl-N-nitrosourea was evaluated. Both substances were administered in a preventive and curative manner (12 days before and 16 weeks after the carcinogen application). Carcinogen administration induced oxidative stress: the level of thiobarbituric acid reactive substances (TBARS) considered as a marker of reactive oxygen species (ROS) generation in liver increased as well as the level of oxidatively modified protein content (OMP; aldehyde and ketone derivates). Metformin administration restored succinate dehydrogenase and lactate dehydrogenase activity and associated ROS production and OMP content to the level of intact rats, with predominant activation of superoxide dismutase (SOD) and glutathione reductase (GR). Melatonin alone and in combination with metformin also decreased TBARS content. OMP content decreased in all groups receiving chemoprevention. The rise in total antioxidant capacity after melatonin and particularly metformin and melatonin combination might result from the initiation of anaerobic metabolism and increasing SOD, GR, and glutathione peroxidase activity. Long-term administration of metformin and melatonin exerts antioxidant properties in liver, especially in combination.

Melatonin and Metformin Diminish Oxidative Stress in Heart Tissue in a Rat Model of High Fat Diet and Mammary Carcinogenesis.

The aim of this study was to determine the effects of long-term administration of the oral antidiabetic metformin or the pineal hormone melatonin, and a combination thereof, in preventing oxidative stress in the heart tissue of female Sprague-Dawley rats with mammary tumors induced by N-methyl-N-nitrosourea (NMU) (50 mg/kg) given on the 42nd postnatal day. Metformin and melatonin were administered 12 days before and 16 weeks after the carcinogen. During the experiment, all animals were fed a high fat diet (10% total fat, 2.5% from lard, and 7.5% from palm oil). The findings are that mammary carcinogenesis generated oxidative stress. Reactive oxygen species (ROS) content, estimated from thiobarbituric acid reactive substances (TBARS), oxidatively modified protein content (aldehyde and ketone derivatives), and the activity of the antioxidant enzymes superoxide dismutase, glutathione reductase, and glutathione peroxidase were all augmented. Metformin caused a decrease in oxidative stress in the heart, accompanied by a decrease in diene conjugates, the elimination of ROS (stable total antioxidant status), and the activation of catalase and glutathione reductase. Melatonin caused an increase in total antioxidant status and a substantial reduction in ROS as estimated from aldehyde and ketone derivatives, lipid peroxidation at the initial (diene conjugates) and terminal stages (TBARS), and increased catalase and glutathione peroxidase activities. Metformin and melatonin combined reversed the effects of NMU on oxidative stress. In conclusion, melatonin reduces the level of oxidative stress in the heart tissue, caused by NMU carcinogenesis and a high fat diet, significantly stronger than metformin.

Metformin and melatonin inhibit DMBA-induced mammary tumorigenesis in rats fed a high-fat diet.

The data from in-vitro and in-vivo studies show that both peroral antidiabetic metformin (MF) and pineal hormone melatonin (MT) inhibit the growth of many cancers, including breast cancer. However, most in-vivo studies used standard-type diet with low fat content. Therefore, in this study, we evaluated the chemopreventive effect of MF and MT in an in-vivo model of breast cancer in rats on a high-fat diet (10% of total fat). Mammary carcinogenesis was induced by 7,12-dimethylbenz[a]anthracene (DMBA) in female Sprague-Dawley rats. Chemoprevention with MF (administered in a diet, 0.2%) and MT (administered in tap water, 20 mg/l) was induced 20 days before the carcinogen administration through the termination of the experiment (14 weeks after carcinogen administration). Tumor growth parameters were analyzed together with histopathological examination and immunohistochemical detection of KI67 (proliferation marker), caspase-3, BAX, BCL-2 (apoptosis markers), and CD24 and CD44 (cancer stem cell markers) in mammary tumor samples. The combination of chemopreventive agents decreased tumor incidence by 29%. Cumulative tumor volume was lower in all groups treated with chemoprevention. Histopathology did not show significant changes in high-grade/low-grade tumor ratio. Immunohistochemistry showed increased expression of BAX in the combination group, and caspase-3 expression increased in both MT and combination groups. MT, and particularly the MF and MT combination, inhibited DMBA-induced mammary tumor growth in rats by apoptosis stimulation in cancer cells. Our results indicate that MT supplements in patients treated with MF may have a considerable effect on the incidence of breast cancer.

Immodin and its immune system supportive role in paclitaxel therapy of 4T1 mouse breast cancer.

It is evident that standard chemotherapy agents may have an impact on both tumor and host immune system. Paclitaxel (PTX), a very potent anticancer drug from a taxane family, has achieved prominence in clinical oncology for its efficacy against a wide range of tumors including breast cancer. However, significant toxicity, such as myelosuppression, limit the effectiveness of Paclitaxel-based treatment regimens. Immodin (IM) is low molecular dialysate fraction of homogenate made from human leukocytes. It contains a mixture of substances from which so far have been described e.g. Imreg 1 and Imreg 2 formed by the dipeptide tyrosine-glycine and the tripeptide tyrosine-glycine-glycine, respectively. The aim of this study was to explore immunopharmacological activities of IM, using the strongly immunogenic 4T1 mouse breast cancer model, and evaluate its effect on the reactivity and the efficiency of PTX cancer therapy. The results highlight a potentially beneficial role for IM in alleviating PTX-induced toxicity, especially on the nonspecific immunity, during breast cancer therapy. Co-treatment exhibited an antitumor effect including reduced tumor growth, prolonged survival of tumor bearing mice, increased number of monocytes and lymphocytes in peripheral blood. In spleens, IM+PTX therapy elevated proportion of whole lymphocytes in the account of myelo-monocytic cells characteristic with low expression of CD11c+ and bearing Fc receptor (CD16/32) as well as T-lymphocytes, NK cells and dendritic cells. Accumulation of tumor-associated granulocytes in stroma of PTX-treated group and intensive 4T1-necrosis/apoptosis in tumors after co-treatment were also recorded. These findings suggest the possibility of using IM alongside PTX treatment for maintaining the immune system functions and increasing patient survival.

Antitumor effect of the combination of manumycin A and Immodin is associated with antiplatelet activity and increased granulocyte tumor infiltration in a 4T1 breast tumor model.

Manumycin A is a natural antibiotic isolated from Streptomyces parvulus with broad range of biological activities including antineoplastic activity in several in vitro and in vivo cancer models. Immodin [dialyzable leukocyte extract (DLE)] is a dialysate released from disintegrated blood leukocytes of healthy donors which exerts immunonormalizing effects on cell-mediated immune responses. The aim of the present study was to explore the antitumor potential of the combination of manumycin A and Immodin in an experimental breast cancer model. Experiments were carried using a 4T1 tumor-bearing BALB/c mouse model. Survival analysis, tumor growth, hematological and biochemical profiles, leukocyte differential, phagocytic activity of leukocytes and histology of the primary tumor were examined. The combination treatment suppressed the tumor growth and prolonged the survival of tumor-bearing mice, decreased the number of monocytes, plateletes and plateletcrit in peripheral blood of the tumor-bearing mice and increased the infiltration of neutrophils and eosinophils in the primary tumor. Moreover, individual therapies enhanced the phagocytic activity of monocytes and neutrophils. These findings demonstrate the antitumor effect of the combination of manumycin A and Immodin in 4T1 tumor-bearing mice associated with strong antiplatelet activity and innate immunity activation.

Role of high-fat diet on the effect of pioglitazone and melatonin in a rat model of breast cancer.

The risk of cancer may be modulated by drugs with pleiotropic effects and diet has been implicated in the efficacy of treatment. The oncopreventive effects of the antidiabetic drug pioglitazone (PIO) and the anti-insomnia drug melatonin (MT), in vivo, have been proven before, but using a standard-type diet. This study evaluated the impact of a high-fat diet on their efficacy in chemically induced mammary carcinogenesis in Sprague-Dawley rats. Mammary tumours were induced by N-methyl-N-nitrosourea (50 mg/kg, intraperitoneal, on the 41st postnatal day). PIO and MT administration was initiated 11 days before the carcinogen application and lasted until the termination of the experiment at 16 weeks. PIO was administered in a diet (10% fat) at a concentration of 100 ppm and MT was administered in tap water (20 mg/l). PIO, MT and the combination did not significantly alter the basic tumour growth parameters. However, histopathology showed a decrease in the high-grade/low-grade tumour ratio, particularly in animals that received combined treatment (P<0.01). Semiquantitative immunohistochemistry indicated the proapoptotic effect of chemoprevention, particularly in the drug combination group (P<0.01), but no changes in tumour cell proliferation and angiogenesis were recorded. Results were evaluated by one-way analysis of variance or the Mann-Whitney U-test, respectively. PIO and MT, alone or in combination, administered to rats fed a high-fat diet reduced the proportion of high-grade tumours and promoted apoptosis in an in-vivo breast cancer model, although it did not suppress tumour growth. The impact of high dietary fat content on the chemopreventive efficacy of these and other substances should be considered in human studies.

Fruit peel polyphenols demonstrate substantial anti-tumour effects in the model of breast cancer.

PURPOSE: Fruit and vegetable intake is inversely correlated with cancer; thus, it is proposed that an extract of phytochemicals as present in whole fruits, vegetables, or grains may have anti-carcinogenic properties. Thus, the anti-tumour effects of fruit peel polyphenols (Flavin7) in the chemoprevention of N-methyl-N-nitrosourea-induced mammary carcinogenesis in female rats were evaluated. METHODS: Lyophilized substance of Flavin7 (F7) was administered at two concentrations of 0.3 and 3 % through diet. The experiment was terminated 14 weeks after carcinogen administration, and mammary tumours were removed and prepared for histopathological and immunohistochemical analysis. In addition, using an in vitro cytotoxicity assay, apoptosis and proliferation after F7 treatment in human breast adenocarcinoma (MCF-7) cells were performed. RESULTS: High-dose F7 suppressed tumour frequency by 58 % (P < 0.001), tumour incidence by 24 % (P < 0.05), and lengthened latency by 8 days (P > 0.05) in comparison with the control rats, whereas lower dose of F7 was less effective. Histopathological analysis of tumours showed significant decrease in the ratio of high-/low-grade carcinomas after high-dose F7 treatment. Immunohistochemical analysis of rat carcinoma cells in vivo found a significant increase in caspase-3 expression and significant decrease in Bcl-2, Ki67, and VEGFR-2 expression in the high-dose group. Both doses demonstrated significant positive effects on plasma lipid metabolism in rats. F7 significantly decreased survival of MCF-7 cells in vitro in MTT assay by dose- and time-dependent manner compared to control. F7 prevented cell cycle progression by significant enrichment in G1 cell populations. Incubation with F7 showed significant increase in the percentage of annexin V-/PI-positive MCF-7 cells and DNA fragmentation. CONCLUSIONS: Our results reveal a substantial tumour-suppressive effect of F7 in the breast cancer model. We propose that the effects of phytochemicals present in this fruit extract are responsible for observed potent anti-cancer activities.

Antineoplastic effects of Chlorella pyrenoidosa in the breast cancer model.

OBJECTIVES: There has been considerable interest in both clinical and preclinical research about the role of phytochemicals in the reduction of risk for cancer in humans. The aim of this study was to determine the antineoplastic effects of Chlorella pyrenoidosa in experimental breast cancer in vivo and in vitro. METHODS: In this experiment, the antineoplastic effects of C. pyrenoidosa in the chemoprevention of N-methyl-N-nitrosourea-induced mammary carcinogenesis in female rats were evaluated. Chlorella powder was administered through diet at concentrations of 0.3% and 3%. The experiment was terminated 14 wk after carcinogen administration. At autopsy, mammary tumors were removed and prepared for histopathological and immunohistochemical analysis. In vitro cytotoxicity assay, parameters of apoptosis, and proliferation after chlorella treatment in human breast adenocarcinoma (MCF-7) cells were carried out. RESULTS: Basic parameters of experimental carcinogenesis, mechanism of action (biomarkers of apoptosis, proliferation, and angiogenesis), chosen metabolic variables, and side effects after long-term chlorella treatment in animals were assessed. Chlorella at higher concentration suppressed tumor frequency by 61% (P < 0.02) and lengthened tumor latency by 12.5 d (P < 0.02) in comparison with the controls. Immunohistochemical analysis of rat tumor cells showed caspase-7 expression increase by 73.5% (P < 0.001) and vascular endothelial growth factor receptor-2 expression decrease by 19% (P = 0.07) after chlorella treatment. In a parallel in vitro study, chlorella significantly decreased survival of MCF-7 cells in a dose-dependent manner. In chlorella-treated MCF-7 cells, a significant increase in cells having sub-G0/G1 DNA content and significant increase of early apoptotic and late apoptotic/necrotic cells after annexin V/PI staining assay were found. Decreases in mitochondrial membrane potential and increasing reactive oxygen species generation were observed in the chlorella-treated MCF-7 cells. CONCLUSIONS: This study is the first report on the antineoplastic effects of C. pyrenoidosa in experimental breast cancer in vivo and in vitro.

Combination of Pitavastatin and melatonin shows partial antineoplastic effects in a rat breast carcinoma model.

Our previous results indicated significant tumor-suppressive effects of different statins in rat mammary carcinogenesis. The purpose of this experiment was to examine the chemopreventive effects of Pitavastatin alone and in combination with the pineal hormone melatonin in the model of N-methyl-N-nitrosourea-induced mammary carcinogenesis in female Sprague-Dawley rats. Pitavastatin was administered dietary (10mg/kg) and melatonin in an aqueous solution (20µg/ml). Chemoprevention began 7 days prior to carcinogen administration and subsequently continued for 15 weeks until autopsy. At autopsy, mammary tumors were removed and prepared for histopathological and immunohistochemical analysis. Compared to controls, Pitavastatin alone reduced average tumor volume by 58% and lengthened latency by 8 days; on the other hand, the drug increased tumor frequency by 23%. Combined administration of Pitavastatin with melatonin decreased tumor frequency by 23%, tumor volume by 44% and lengthened tumor latency by 5.5 days compared to control animals. The analysis of carcinoma cells showed significant increase in caspase-3 expression in both treated groups and a tendency of increased caspase-7 expression after Pitavastatin treatment alone. Significant expression decrease of Ki67 was found in carcinoma cells from both treated groups. Compared to control carcinoma cells, Pitavastatin alone increased VEGF expression by 41%, however melatonin totally reversed its undesirable effect. Pitavastatin combined with melatonin significantly increased femur compact bone thickness in animals. Pitavastatin alone decreased plasma triglycerides and total cholesterol levels, however it significantly increased levels of glucose. In summary, our results show a partial antineoplastic effect of Pitavastatin combined with melatonin in the rat mammary gland carcinoma model.

Melatonin potentiates the anti-tumour effect of pravastatin in rat mammary gland carcinoma model.

Previous studies in the field of cancer research have suggested a possible role for statins in the reduction of risk in certain malignancies. The purpose of these studies was to examine the chemopreventive effects of pravastatin alone and in combination with pineal hormone melatonin in the N-methyl-N-nitrosourea-induced mammary carcinogenesis model. Pravastatin was given orally (1 00 mg/kg) and melatonin was added to the water (20 µg/ml). Chemoprevention began seven days prior to carcinogen administration and subsequently continued for 15 weeks until autopsy. At autopsy, mammary tumours were removed and prepared for histopathological and immunohistochemical analysis. Parameters of experimental carcinogenesis, mechanism of action (biomarkers of apoptosis, angiogenesis and proliferation) and side effects after long-term treatment in animals were assessed. Pravastatin alone suppressed tumour frequency by 20.5% and average tumour volume by 15% compared with controls. Combined administration of the drugs decreased tumour frequency by 69% and lengthened tumour latency by nine days compared with control animals. The ration between high and low grade carcinomas was apparently reduced in both treated groups. The analysis of carcinoma cells showed significant expression increase in caspase-3 and caspase-7 after pravastatin treatment; however, combined treatment even more pronounced increase in the expression of both caspases. Regarding VEGFR-2 expression, a small effect in carcinomas of both treated groups was found. In plasma metabolism evaluation, pravastatin alone significantly decreased levels of glucose and triacylglycerols. Our results suggest a mild anti-neoplastic effect of pravastatin in this rat mammary gland carcinoma model. Statins co-administered with other suitable drug (e.g. melatonin) should be further evaluated for tumour-preventive properties.

Resveratrol enhances the chemopreventive effect of celecoxib in chemically induced breast cancer in rats.

Resveratrol and celecoxib were used as chemopreventive agents in animal models of carcinogenesis, and exert antiproliferative and proapoptotic effects on cancer cells. Therefore, the aim of this study was to evaluate whether combining resveratrol with celecoxib may exert more potent anticarcinogenic effects than the single agents. Mammary carcinogenesis was initiated in 70 female Sprague-Dawley rats with N-methyl-N-nitrosourea (NMU). The chemoprevention with resveratrol, celecoxib, and their combination started 2 weeks before the first carcinogen dose and lasted until the end of the experiment. Tumor incidence and frequency, latency period, tumor volume, the expression of cyclooxygenase 2 (COX2) and growth differentiation factor 15 (GDF15), and also the formation of reactive oxygen species were analyzed using different methods. In addition, the levels of resveratrol and its metabolites in blood and selected tumor tissues were determined by high-performance liquid chromatography. Finally, the anticancer effects of the reagents were studied in the human breast cancer cell line MCF-7. Celecoxib as a single agent significantly decreased tumor frequency, prolonged tumor latency, and decreased the total number of malignant tumors compared with the NMU conditions. Tumor volume was nonsignificantly reduced (0.68±0.25 vs. 0.93±0.28 cm3). Importantly, the addition of resveratrol to celecoxib reduced tumor volume by 60% compared with celecoxib alone (from 0.68±0.25 to 0.27±0.07 cm3, P<0.05). Furthermore, the combination of resveratrol and celecoxib reduced tumor frequency by 29% compared with celecoxib alone (P=0.53). Tumor latency was not influenced by this combination compared with celecoxib alone (126.56±3.45 vs. 120.71±4.08 days). In addition, COX2 mRNA and immunoreactive protein stained on tumor sections were reduced and GDF15 protein increased significantly by the combination studied compared with the NMU conditions. In agreement with these data, a significant reduction in reactive oxygen species in blood lymphocytes of the combination was detected, which may have contributed toward the cancer-preventive effects of this application. This study showed that in NMU-induced mammary cancer in rats, the combination of resveratrol and celecoxib led to a significant reduction in all tumor parameters. In addition, in terms of tumor volume, the combination was more efficient than celecoxib as a single agent.

Preventive effects of probiotic bacteria Lactobacillus plantarum and dietary fiber in chemically-induced mammary carcinogenesis.

AIM: The purpose of the present study was to evaluate the chemopreventive efficacy of a new probiotic bacterial strain, Lactobacillus plantarum LS/07 (PRO), prebiotic oligofructose-enriched inulin (PRE) and PRO-PRE combination in a rat model of breast cancer. MATERIALS AND METHODS: Mammary carcinogenesis was induced by 7,12-dimethylbenz[a]anthracene (DMBA). Daily oral administration of PRO (at a dose of 8.4×10(8) c.f.u./rat) and PRE (in the diet, 20 g/kg) started two weeks before the first DMBA dose and lasted until the end of the experiment (16 weeks). RESULTS: Administration of PRO, PRE and PRO-PRE combination significantly suppressed the tumor frequency, increased Cd4(+) T-cells in tumor tissue and reduced serum tumor necrosis factor-α concentration. In PRO and PRO-PRE groups, the decline of Cd8(+) T-cells in blood and their increase in tumor tissue was observed. CONCLUSION: Long-term administration of Lactobacillus plantarum LS/07 with and without inulin is effective against breast cancer, at least partially, through immunomodulatory mechanisms.