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Recently, green microalgae have gained importance due to their nutritional and bioactive compounds, which makes them some of the most promising and innovative functional foods. The aim of this study was to evaluate the chemical profile and the in vitro antioxidant, antimicrobial and antimutagenic activity of an aqueous extract of the green microalga Ettlia pseudoalveolaris, obtained from the freshwater lakes of the Ecuadorian Highlands. Human microvascular endothelial cells (HMEC-1) were used to determine the ability of the microalga to reduce the endothelial damage caused by hydrogen peroxide-induced oxidative stress. Furthermore, the eukaryotic system Saccharomyces cerevisiae was used to evaluate the possible cytotoxic, mutagenic and antimutagenic effect of E. pseudoalveolaris. The extract showed a notable antioxidant capacity and a moderate antibacterial activity mostly due to the high content in polyphenolic compounds. It is likely that the antioxidant compounds present in the extract were also responsible for the observed reduction in endothelial damage of HMEC-1 cells. An antimutagenic effect through a direct antioxidant mechanism was also found. Based on the results of in vitro assays, E. pseudoalveolaris proved to be a good source of bioactive compounds and antioxidant, antibacterial and antimutagenic capacities making it a potential functional food.

Final results regarding brain and heart tumors in Sprague-Dawley rats exposed from prenatal life until natural death to mobile phone radiofrequency field representative of a 1.8 GHz GSM base station environmental emission The Ramazzini Institute (RI) performed a life-span carcinogenic study on Sprague-Dawley rats to evaluate the carcinogenic effects of RFR in the situation of far field, reproducing the environmental exposure to RFR generated by 1.8 GHz GSM antenna of the radio base stations of mobile phone. Male and female Sprague-Dawley rats were exposed from prenatal life until natural death to a 1.8 GHz GSM far field of 0, 5, 25, 50 V/m with a whole-body exposure for 19 h/day. In 2018, we reported the final results regarding brain and heart tumors. A statistically significant increase in the incidence of heart Schwannomas was observed in treated male rats at the highest dose (50 V/m). Furthermore, an increase in the incidence of malignant glial tumors was observed in treated female rats at the highest dose (50 V/m), although not statistically significant. The RI findings are consistent with the NTP results on RFR exposure, as both reported an increase in the incidence of brain and heart tumors of the same histotype of those observed in some epidemiological studies on cell phone users. These studies provide sufficient evidence to call for the reevaluation of IARC conclusions regarding the carcinogenic potential of RFR in humans.

BACKGROUND AND AIM: Glyphosate (G) is set for re-approval in the European Union in late 2022 and the full risk posed by glyphosate-based herbicides (GBHs) is still unclear.The Ramazzini Institute (...

Herein we characterized the bioactive metabolites of the aqueous extract of Kavolì®, a commercial product composed of a mixture of Brassica oleracea leaves, and assessed its potential ameliorating effects in a rat model of non-alcoholic fatty liver disease (NAFLD). Kavolì® extract showed high levels of bioactive compounds and strong in vitro antioxidant activities. Chlorogenic and neochlorogenic acids were identified as the most representative polyphenols. The administration of brassica extract to steatotic rats significantly ameliorated the levels of blood lipids and transaminases, and lipid content and inflammatory markers in liver. Oxidative stress parameters were significantly improved in both liver and brain of steatotic rats. Moreover, plasma and feces levels of short chain fatty acids (SCFAs) were bring back close to control values by Kavolì® treatment, in spite of high fat diet/streptozotocin (HFD/STZ)-induced alterations. The efficacy of Kavolì® in treating hypercholesterolemia, reducing the level of inflammation and cardiovascular disease biomarkers, steatosis and oxidative stress parameters, as well as the ability in modulating SCFAs levels is probably related to the bioactive compounds of the water extract administered to the rat model of NAFLD. In particular, the ameliorating effects are largely attributable to the high content in polyphenols observed in our study.

Numerosi studi sperimentali suggeriscono un ruolo chemiopreventivo della vitamina E (VE) nei confronti di numerose patologie tumorali incluso il cancro alla prostata (CA). Tuttavia, le evidenze epidemiologiche più recenti non confermano tale ipotesi, e lo studio clinico SELECT ha addirittura evidenziato un aumento dell’incidenza di CA alla prostata in pazienti sani al momento dell’arruolamento, ai quali era stata somministrata VE giornalmente alla dose di 400 UI/die. Essendo noto in letteratura come alcune isoforme del citocromo P450 (CYP) siano sovra espresse in lesioni prostatiche maligne, e che la VE, agisca come induttore del CYP in alcuni tessuti, abbiamo ipotizzato che l’aumento dell’incidenza di CA possa coinvolgere un meccanismo co-cancerogenico legato all’induzione CYP. L’esposizione a VE produce una marcata induzione del CYP in cellule epiteliali di prostata RWPE-1, incluse le isoforme responsabili dell’attivazione degli Idrocarburi Policiclici Aromatici (IPA), accompagnata da un significativo aumento delle specie reattive dell’ossigeno (ROS). Inoltre, il pathway pro-infiammatorio COX-2-dipendente risulta incrementato, caratteristica che si osserva in diverse forme tumorali incluso quelle che interessano la prostata. Il fenomeno è stato confermato nel modello animale; è stato osservato un aumento delle monoossigenasi e dell’espressione genica di alcune isoforme CYP a livello prostatico unito a una maggiore concentrazione di ROS, in linea con marcatori di danno ossidativo a livello proteico e lipidico, nella prostata di ratto. L’esposizione di cellule IMR90 a VE ha evidenziato un effetto genotossico riportando un’aumentata frequenza dei micronuclei (MN). Il potenziale co-cancerogeno di VE è stato poi confermato dallo studio della trasformazione cellulare mediate l’uso di cellule BALB/c 3T3. Cellule pre-incubate con VE e successivamente esposte a benzo[a]pirene mostrano un significativo incremento della frequenza di trasformazione rispetto a quello senza della vitamina. Il presente studio dimostra come la VE possa, in determinate condizioni, agire come co-cancerogeno tramite un meccanismo CYP-dipendente, causando danno al DNA e promovendo la trasformazione cellulare.

A large number of basic researches and observational studies suggested the cancer preventive activity of vitamin E, but large-scale human intervention trials have yielded disappointing results and actually showed a higher incidence of prostate cancer although the mechanisms underlying the increased risk remain largely unknown. Here we show through in vitro and in vivo studies that vitamin E produces a marked inductive effect on carcinogen-bioactivating enzymes and a pro-oxidant status promoting both DNA damage and cell transformation frequency. First, we found that vitamin E in the human prostate epithelial RWPE-1 cell line has the remarkable ability to upregulate the expression of various phase-I activating cytochrome P450 (CYP) enzymes, including activators of polycyclic aromatic hydrocarbons (PAHs), giving rise to supraphysiological levels of reactive oxygen species. Furthermore, our rat model confirmed that vitamin E in the prostate has a powerful booster effect on CYP enzymes associated with the generation of oxidative stress, thereby favoring lipid-derived electrophile spread that covalently modifies proteins. We show that vitamin E not only causes DNA damage but also promotes cell transformation frequency induced by the PAH-prototype benzo[a]pyrene. Our findings might explain why dietary supplementation with vitamin E increases the prostate cancer risk among healthy men.

Introduction The dietary supplements use has increased over the past years in the global landscape and vitamins are by far the products which record the greater appeal. Following the well-rooted idea that the vitamin E promotes health status, many of people assume it not to treat diagnosed deficiencies, but to prevent new occurrences of several diseases, including cancer. However, recent trials and meta-analysis showed no significant effects against cancer aetiology, whereas data from SELECT study reported an increased risk of prostate cancer among healthy men. To date, the mechanism underlying these observations is largely unknown. In this work, we show in in vitro and in vivo models, how vitamin E through co-mutagenic and pro-oxidant mechanisms promotes both DNA damage and cell transformation frequency. Materials and Methods In vitro models Human prostate cells RWPE-1 were exposed to α-tocopherol (100 μM) for 48 h. The gene expression of COX2, CYPA1, CYPA4, CYPB6, CYPC9, and CYP4F2 gene was studied. PGE2 were determinate by ELISA test. The measure of reactive oxygen species (ROS) was carried out by the use of the Electron Paramagnetic Resonance (EPR) radical-probe technique. Human fibroblasts IMR90 were exposed to α-tocopherol (100 μM) for 48 h then cells were fixed, stained with DAPI. Micronuclei were scored. IMR90 cultured as previously described were employed for the Immunofluorescence determination of 53BP1 and γH2AX foci. BALB/c 3T3 cells were treated with α-tocopherol (100 μM), after 48 h exposition the medium was removed and cells were incubated with benzo[a]pyrene with a final concentrations ranged from 1 µg/ml to 0.01 µg/ml. Cells were maintained in culture for 4 weeks with twice weekly medium changes, then fixed and stained. Positive transformation foci were counted. In vivo model Vitamin E was administered to Sprague-Dawley rats at doses of 100 or 200 mg/kg b.w. dose daily for 7 or 14 consecutive days. Controls received vehicle (corn oil). Prostate was resected and the CYP1A1 CYP1A2, CYP1B1 gene expression was studied along with the catalytic activity through specific probes. ROS yields was measured by EPR spectroscopy. Finally, the malonyldialdehyde (MDA) development, as a marker of lipid peroxidation as well as protein carbonylation were assessed. Results Here we show that in RWPE-1 cell line vitamin E produces an overexpression of various phase-I bioactivating cytochrome P450 (CYP) isoforms, including activators of polycyclic aromatic hydrocarbons (PAHs), such as CYPA1, CYPA4, CYPB6, CYPC9, and CYP4F2 coupled with a significant yield of radical oxygen species (ROS). These observations have been supported by an in vivo model; in vitamin E supplemented rats we reported a significant booster effect on CYP superfamily along with oxidative damage to lipids and proteins. Furthermore, we show that vitamin E causes DNA damage as exemplified through micronuclei and H2AX - 53BP1 foci based-assays. Finally, we find that the pre-exposure of BALB/c 3T3 cells with vitamin E significantly increase the transformation frequency of the PAH-prototype benzo[a]pyrene. Discussion and Conclusions Taken together, these results can contribute to explain the increased cancer risk found in the SELECT trial. Our data support the hypothesis that vitamin E could act as a co-mutagenic/co-carcinogenic agent increasing the bioactivation of chemicals joint with its pro-oxidant activity resulting in higher risk of cancer through DNA insults and epigenetic alterations. Our study point out one more time the potentially harmful effects of recommending supplementations with micronutrient as a chemopreventive strategy on a large scale.

Lymphomas and leukaemias involving the lung have in some cases been hard to distinguish from respiratory tract infection in Sprague-Dawley (SD) rats from long-term bioassays. In order to differentiate between tumours and immune cell infiltrates, updated pathological criteria and nomenclature were used and immunohistochemistry (IHC) was applied to haematopoietic and lymphoid tissue tumours (HLTs) in the original prenatal long-term Aspartame (APM) study performed by the Ramazzini Institute (RI). All 78 cases of HLTs from treated and control groups were re-examined based on light microscopic morphological characteristics and subjected to a panel of IHC markers including Ki67, CD3, PAX5, CD20, CD68, TdT, CD45, CD14 and CD33. The analysis confirmed the diagnoses of HLTs in 72 cases, identified 3 cases of preneoplastic lesions (lymphoid hyperplasia), and categorized 3 cases as inflammatory lesions. A statistically significant increase in total HLTs (p = 0.006), total lymphomas (p = 0.032) and total leukaemias (p = 0.031) in treated female rats was confirmed (high dose vs control), and a statistically significant linear trend for each HLT type was also observed. After the HLT cases re-evaluation, the results obtained are consistent with those reported in the previous RI publication and reinforce the hypothesis that APM has a leukaemogenic and lymphomatogenic effect.

Background In 2011, IARC classified radiofrequency radiation (RFR) as possible human carcinogen (Group 2B). According to IARC, animals studies, as well as epidemiological ones, showed limited evidence of carcinogenicity. In 2016, the NTP published the first results of its long-term bioassays on near field RFR, reporting increased incidence of malignant glial tumors of the brain and heart Schwannoma in rats exposed to GSM – and CDMA – modulated cell phone RFR. The tumors observed in the NTP study are of the type similar to the ones observed in some epidemiological studies of cell phone users. Objectives The Ramazzini Institute (RI) performed a life-span carcinogenic study on Sprague-Dawley rats to evaluate the carcinogenic effects of RFR in the situation of far field, reproducing the environmental exposure to RFR generated by 1.8 GHz GSM antenna of the radio base stations of mobile phone. This is the largest long-term study ever performed in rats on the health effects of RFR, including 2448 animals. In this article, we reported the final results regarding brain and heart tumors. Methods Male and female Sprague-Dawley rats were exposed from prenatal life until natural death to a 1.8 GHz GSM far field of 0, 5, 25, 50 V/m with a whole-body exposure for 19 h/day. Results A statistically significant increase in the incidence of heart Schwannomas was observed in treated male rats at the highest dose (50 V/m). Furthermore, an increase in the incidence of heart Schwann cells hyperplasia was observed in treated male and female rats at the highest dose (50 V/m), although this was not statistically significant. An increase in the incidence of malignant glial tumors was observed in treated female rats at the highest dose (50 V/m), although not statistically significant. Conclusions The RI findings on far field exposure to RFR are consistent with and reinforce the results of the NTP study on near field exposure, as both reported an increase in the incidence of tumors of the brain and heart in RFR-exposed Sprague-Dawley rats. These tumors are of the same histotype of those observed in some epidemiological studies on cell phone users. These experimental studies provide sufficient evidence to call for the re-evaluation of IARC conclusions regarding the carcinogenic potential of RFR in humans.

Polyphenol-rich foods could have a pivotal function in the prevention of oxidative stress-based pathologies and antibacterial action. The purpose of this study was to investigate the in vitro antimicrobial activity, as well as the in vitro and In Vivo antioxidant capacities of wild Prunus spinosa L. fruit (PSF) from the southeast regions of Italy. The total phenolic content (TPC) was quantified, and the single polyphenols were analyzed by HPLC-DAD, showing high rutin and 4-hydroxybenzoic acid levels, followed by gallic and trans-sinapic acids. PSF extract demonstrated antimicrobial activity against some potentially pathogenic Gram-negative and Gram-positive bacteria. Besides, we investigated the cellular antioxidant activity (CAA) and the hemolysis inhibition of PSF extract on human erythrocytes, evidencing both a good antioxidant power and a marked hemolysis inhibition. Furthermore, an In Vivo experiment with oxidative stress-induced rats treated with a high-fat diet (HFD) and a low dose of streptozotocin (STZ) demonstrated that PSF has a dose-dependent antioxidant capacity both in liver and in brain. In conclusion, the wild Italian Prunus spinosa L. fruit could be considered a potentially useful material for both nutraceutical and food industries because of its antioxidant and antimicrobial effects.

The proliferation of cellular antennas and other radiofrequency radiation (RFR) generating devices of the last decades has led to more and more concerns about the potential health effects from RFR exposure. Since the 2011 classification as a possible carcinogen by the International Agency for Research on Cancer (IARC), more experimental studies have been published that support a causal association between RFR exposure and health hazards. As regard cancer risk, two long-term experimental studies have been recently published by the US National Toxicology Program (NTP) and the Italian Ramazzini Institute (RI). Despite important experimental differences, both studies found statistically significant increases in the development of the same type of very rare glial malignant tumors. In addition to carcinogenicity, reproductive organs might be particularly exposed, as well as sensitive to RFR. In this work, we reviewed the currently available evidence from in vivo studies on carcinogenicity and reproductive toxicity studies in order to summarize the contribution of experimental research to the prevention of the adverse effects of RFR on human health.

The cancer hazard associated with exposure to cell phone radiofrequency radiation (RFR) was examined using lifetime exposure in Sprague-Dawley (SD) rats at the Ramazzini Institute (RI), Italy. There were increased incidences of gliomas in the brain and schwannomas in the heart. In order to understand the translational relevance of these rat tumors for human disease, we examined the top 23 orthologous cancer genes mutated in human gliomas using a custom built a next-generation sequencing gene panel for rats based on Illumina’s TruSeq Custom Amplicon Technology. SD rat tissues (gliomas =15; cardiac schwannomas=9, interim (1 year) sacrificed non-tumor brain tissues from RFR exposed rats =30, control brain and heart control tissues from lifetime exposure=10 each, and age-matched control brain tissues from interim sacrificed rats=10) from the RI-RFR cancer bioassay were examined in this study. The deepSNV R-package with various filtering criteria and the read depth of >1000x were used to identify single-nucleotide variants (SNVs) in the rat gliomas and schwannomas. At 5% allelic frequency, there were an average of 43 SNVs per rat glioma and point mutations were detected in 9 genes (Arid1a, Cic, Tert promoter, Tp53, Atrx, Nf1, Pdgfra, Pi3kr1, and Setd2) in at least 3 or more rat gliomas based on population frequency. Five genes (Nf1, Tert promoter, Setd2, Arid1a, and Pdgfra) harbored SNVs in interim sacrificed brain tissues that were also present in RFR-exposed gliomas from lifetime exposure. Interestingly, no mutations were detected in hotspot regions of Idh1, Idh2, Egfr or Braf. In contrast to most human gliomas which harbor mutations in IDH1 and IDH2 genes, the rat gliomas seem to be Idh1 wild type with mutations in the other glioma-related genes. Primary cardiac tumors are extremely rare in humans. With this targeted NGS panel (at allelic frequency of 2.5%), there were an average of 146 SNVs per rat cardiac schwannoma and unique point mutations were detected in Cic, Egfr, Arid1a, Nf1, Setd2, Cdkn2a, Erbb2, Atrx, Pdgfra, and Notch1 in 3 or more cardiac schwannomas. A subset of the SNVs (Arid1a, Tp53, and Nf1) in rat gliomas was confirmed in human gliomas in the COSMIC database supporting the translational relevance of the rat gliomas for human disease. In addition, several SNVs from rat gliomas and cardiac schwannomas were found in various human cancers including carcinomas, hematopoietic neoplasms, melanomas and neuroendocrine tumors. In conclusion, the rat gliomas appear to share genetic alterations with a subtype of IDH1 wildtype human gliomas and rat primary cardiac schwannomas also harbor mutations in some of the queried cancer genes. Citation Format: Ramesh C. Kovi, Andrea Vornoli, Ashley Brooks, Thai Vu T. Ton, Miaofei Xu, Eva Tibaldi, Federica Gnudi, Jian-Liang Li, Robert C. Sills, John R. Bucher, Fiorella Belpoggi, Arun R. Pandiri. Genetic profiling of rat gliomas and cardiac schwannomas from cell phone radiofrequency radiation exposure using a targeted next-generation sequencing gene panel [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PO-084.

The spice turmeric (Curcuma longa L.) has a long history of use as an anti-inflammatory agent. The active component curcumin induces a variety of diverse biological effects and forms a series of degradation and metabolic products in vivo. Our hypothesis is that the field of toxicogenomics provides tools that can be used to characterize the mode of action and toxicity of turmeric components and to predict turmeric-drug interactions. Male Sprague-Dawley rats were treated for 4 days with turmeric root containing about 3% curcumin (comparable to what people consume in the fresh or dried root) or a fraction of turmeric enriched for curcumin (∼74%) and liver tissue collected for gene expression analysis. Two doses of each agent were added to the diet, corresponding to 540 and 2700 mg/kg body weight/day of turmeric. The transcriptomic effects of turmeric on rat liver tissue were examined using 3 programs, ToxFx Analysis Suite, in the context of a large drug database, Ingenuity Pathway and NextBio analyses. ToxFx analysis indicates that turmeric containing about 3% or 74% curcumin represses the expression of cholesterol biosynthetic genes. The dose of 400 mg/kg b.w./day curcumin induced the Drug Signature associated with hepatic inflammatory infiltrate. Ingenuity analysis confirmed that all 4 turmeric treatments had a significant effect on cholesterol biosynthesis, specifically the Cholesterol biosynthesis superpathway and Cholesterol biosynthesis 1 and 2. Among the top 10 up or downregulated genes, all 4 treatments downregulated PDK4; while 3 treatments downregulated ANGPTL4 or FASN. These findings suggest curcumin may enhance the anticancer effects of certain classes of statins, which we confirmed with biological assays. Given this enhancement, lower levels of statins may be required, and even be desirable. Our findings also warn of possible safety issues, such as potential inflammatory liver effects, for patients who ingest a combination of certain classes of statins and curcumin. Transcriptomic analysis suggests that turmeric is worthwhile to study to prevent and treat cancer and lipid disorders. Our approach lays new groundwork for studies of the mode of action and safety of herbal medicines and can also be used to develop a methodology to standardize herbal medicines.

Electronic cigarettes (e-cigs) are devices designed to deliver nicotine in a vaping solution rather than smoke and without tobacco combustion. Perceived as a safer alternative to conventional cigarettes, e-cigs are aggressively marketed as lifestyle-choice consumables, thanks to few restrictions and a lack of regulatory guidelines. E-cigs have also gained popularity among never-smokers and teenagers, becoming an emergent public health issue. Despite the burgeoning worldwide consumption of e-cigs, their safety remains largely unproven and it is unknown whether these devices cause in vivo toxicological effects that could contribute to cancer. Here we demonstrate the co-mutagenic and cancer-initiating effects of e-cig vapour in a rat lung model. We found that e-cigs have a powerful booster effect on phase-I carcinogen-bioactivating enzymes, including activators of polycyclic aromatic hydrocarbons (PAHs), and increase oxygen free radical production and DNA oxidation to 8-hydroxy-2′-deoxyguanosine. Furthermore, we found that e-cigs damage DNA not only at chromosomal level in peripheral blood, such as strand breaks in leucocytes and micronuclei formation in reticulocytes, but also at gene level such as point mutations in urine. Our results demonstrate that exposure to e-cigs could endanger human health, particularly among younger more vulnerable consumers.

Polyphenol-rich foods could have a pivotal function in the prevention of oxidative stress-based pathologies and antibacterial action. The purpose of this study was to investigate the in vitro antimicrobial activity, as well as the in vitro and In Vivo antioxidant capacities of wild Prunus spinosa L. fruit (PSF) from the southeast regions of Italy. The total phenolic content (TPC) was quantified, and the single polyphenols were analyzed by HPLC-DAD, showing high rutin and 4-hydroxybenzoic acid levels, followed by gallic and trans-sinapic acids. PSF extract demonstrated antimicrobial activity against some potentially pathogenic Gram-negative and Gram-positive bacteria. Besides, we investigated the cellular antioxidant activity (CAA) and the hemolysis inhibition of PSF extract on human erythrocytes, evidencing both a good antioxidant power and a marked hemolysis inhibition. Furthermore, an In Vivo experiment with oxidative stress-induced rats treated with a high-fat diet (HFD) and a low dose of streptozotocin (STZ) demonstrated that PSF has a dose-dependent antioxidant capacity both in liver and in brain. In conclusion, the wild Italian Prunus spinosa L. fruit could be considered a potentially useful material for both nutraceutical and food industries because of its antioxidant and antimicrobial effects.