Growth and decline of the COVID-19 epidemic wave in Italy from March to June 2020.

March 21, 2020 was the ridgeline between the growth of the coronavirus disease 2019 (COVID-19) epidemic wave in Italy and the start of its decline. We analyzed the epidemic patterns from March 1 to June 30. There was a progressive drop of cases from March (104,710) to April (94,888), May (25,705) and June (8110). Likewise, after a slight increase of deaths in April (14,804) compared to March (12,396), a considerable decline occurred in May (5170) and June (1464). Doubling times of cumulative cases grew from 2 to 6 days until March 20 to 2 weeks up to April 5, and thereafter no further doubling occurred until June 30. There was a striking North-South gradient of both cases and deaths. At the end of June, the nine Northern Italian regions or provinces, five central regions, and seven southern regions had contributed to the 81.1%, 12.4%, and 6.5% of the 240,578 national cases, respectively. Lombardy, the most populous region, was by far the most heavily affected one, accounting for the 39.0% of the national cases occurring over the analyzed 4-month period. However, in relative terms, it was preceded by Aosta Valley, the least populous region, less than 1% of the population of both regions having been affected by cases of COVID-19. The curves showing the ratio of daily cumulative cases and deaths to those of the previous day tended to flatten with time by approaching the zero growth but without reaching it, which documents a persisting circulation of severe acute respiratory syndrome coronavirus 2 in the Italian territory.

Rationale for the use of N-acetylcysteine in both prevention and adjuvant therapy of COVID-19.

COVID-19 may cause pneumonia, acute respiratory distress syndrome, cardiovascular alterations, and multiple organ failure, which have been ascribed to a cytokine storm, a systemic inflammatory response, and an attack by the immune system. Moreover, an oxidative stress imbalance has been demonstrated to occur in COVID-19 patients. N- Acetyl-L-cysteine (NAC) is a precursor of reduced glutathione (GSH). Due to its tolerability, this pleiotropic drug has been proposed not only as a mucolytic agent, but also as a preventive/therapeutic agent in a variety of disorders involving GSH depletion and oxidative stress. At very high doses, NAC is also used as an antidote against paracetamol intoxication. Thiols block the angiotensin-converting enzyme 2 thereby hampering penetration of SARS-CoV-2 into cells. Based on a broad range of antioxidant and anti-inflammatory mechanisms, which are herein reviewed, the oral administration of NAC is likely to attenuate the risk of developing COVID-19, as it was previously demonstrated for influenza and influenza-like illnesses. Moreover, high-dose intravenous NAC may be expected to play an adjuvant role in the treatment of severe COVID-19 cases and in the control of its lethal complications, also including pulmonary and cardiovascular adverse events.

Modulation of smoke-induced DNA and microRNA alterations in mouse lung by licofelone, a triple COX-1, COX-2 and 5-LOX inhibitor.

Chronic inflammation plays a crucial role in the carcinogenesis process and, in particular, in smoking-related carcinogenesis. Therefore, anti-inflammatory agents provide an interesting perspective in the prevention of smoking-associated cancers. Among nonsteroidal anti-inflammatory drugs (NSAIDs), licofelone is a triple inhibitor of both cyclooxygenases (COX-1 and COX-2) and of 5-lipooxygenase (5-LOX) that has shown some encouraging results in cancer prevention models. We previously showed that the dietary administration of licofelone, starting after weanling, to Swiss H mice exposed for 4 months to mainstream cigarette smoke since birth attenuated preneoplastic lesions of inflammatory nature in both lung and urinary tract, and had some effects on the yield of lung tumors at 7.5 months of age. The present study aimed at evaluating the early modulation by licofelone of pulmonary DNA and RNA alterations either in smoke-free or smoke-exposed H mice after 10 weeks of exposure. Licofelone protected the mice from the smoke-induced loss of body weight and significantly attenuated smoke-induced nucleotide alterations by decreasing the levels of bulky DNA adducts and 8-hydroxy-2'-deoxyguanosine in mouse lung. Moreover, the drug counteracted dysregulation by smoke of several pulmonary microRNAs involved in stress response, inflammation, apoptosis, and oncogene suppression. However, even in smoke-free mice administration of the drug had significant effects on a broad panel of microRNAs and, as assessed in a subset of mice used in a parallel cancer chemoprevention study, licofelone even enhanced the smoke-induced systemic genotoxic damage after 4 months of exposure. Therefore, caution should be paid when administering licofelone to smokers for long periods.

Epidemiological trends of COVID-19 epidemic in Italy over March 2020: From 1000 to 100 000 cases.

The coronavirus disease 2019 epidemic started in Italy by the end of January 2020 and, after 1 month, it affected 1049 persons. Based on the Italian Ministry of Health data, we reconstructed the daily course of virus-positive cases and deaths over March 2020 for the whole of Italy, 19 regions and 2 provinces. From 29 February to 31 March, there was a 100.9-fold increase in the cumulative number of cases and a 428.6-fold increase in the number of deaths in Italy. When plotted on a semilogarithmic scale, the curves tended to diverge from linearity with 23%, 16%, and 7% average daily increases during the three decades of March. Similarly, the number of deaths decreased from an average daily growth of 19% over the second decade to 10% over the third decade. The correlation coefficients relating the days to cases or deaths over each one of the three decades approached unity. As inferred from the equations of the regression lines relative to the three decades, the doubling times of cases were 3.4, 5.1, and 9.6 days, respectively. The doubling times of deaths over the second and third decades were 4.9 and 7.0 days, respectively. There was a broad geographic variability, with a striking gradient from the North, where 40.8% of cases and 57.9% of deaths occurred in Lombardy, to the South. On the whole, over March there was a trend to epidemic growth decline but the time for the end of the epidemic will depend on a variety of factors and, at present, it is unpredictable.

Carcinogenic response and other histopathological alterations in mice exposed to cigarette smoke for varying time periods after birth.

In spite of the outstanding role of tobacco smoking in human carcinogenesis, it is difficult to reproduce its effects in experimental animals. Based on the knowledge that a variety of mechanisms account for a higher susceptibility to carcinogens early in life, we have developed a murine model in which mainstream cigarette smoke becomes convincingly carcinogenic. The standard model involves exposure to smoke for 4 months, starting after birth, followed by an additional 3-4 months in filtered air. We evaluated herein the time- and dose-dependent response, at 7.5 months of life, of Swiss H mice that had been exposed to smoke for either 1, 2 or 4 months after birth. A one-month exposure, corresponding to a period of intense alveolarization, was sufficient to induce most inflammatory, degenerative and preneoplastic pulmonary lesions, including emphysema and alveolar epithelial hyperplasia, blood vessel proliferation and hemangiomas, reflecting an early proangiogenic role of smoking, and microadenomas bearing ki-67-positive proliferating cells as well as urinary bladder epithelial hyperplasia. Two months of exposure were needed to induce pulmonary adenomas and urinary bladder papillomas in males only, which highlights a protective role of estrogens in urinary bladder carcinogenesis. Four months, which in humans would correspond to the postnatal period, puberty, adolescence and early adulthood, were needed to induce other lesions, including tubular epithelial hyperplasia of kidney, bronchial epithelial hyperplasia and especially pulmonary malignant tumors. These findings highlight the concept that preneoplastic and neoplastic lesions occurring in adulthood can be induced by exposure to smoke early in life.

Aspirin abrogates impairment of mammary gland differentiation induced by early in life second-hand smoke in mice.

Epidemiological studies show that there is limited evidence that tobacco smoking causes breast cancer in humans. In rodents, many tobacco smoke chemicals cause mammary gland tumors. This study evaluated the mammary gland differentiation in mice exposed to environmental cigarette smoke (ECS), using 3R4F Kentucky reference cigarettes, starting after birth and continuing daily for 10 weeks (total particulate exposure 95 mg/m3; CO 610 ppm). We also analyzed the effects of oral administration of non-steroidal anti-inflammatory drugs (NSAIDs), aspirin (1600 mg/kg) or naproxen (320 mg/kg), on mammary gland differentiation, either in unexposed or ECS-exposed mice. The ECS exposure caused delay of mammary glands development. We speculate that this delay may result from aryl hydrocarbon receptor (AHR) signaling activation, which has an antiestrogenic effect and crosstalk to the estrogen metabolism pathway. Similarly, naproxen impaired gland differentiation in unexposed and ECS-exposed mice, while aspirin hindered its development only in unexposed mice. The lack of differentiation induced by the NSAIDs could be explained by their antiestrogenic effect through inhibition of aldo-keto reductases. In ECS-exposed animals, aspirin induced intense lobular formation, which could indicate that aspirin is counteracting the AHR signaling induced by ECS. Based on the differentiation induced by aspirin in ECS-exposed animals, we postulate that these mice would be less susceptible to mammary carcinogenesis. Our results suggest that exposure to smoke at an early age impairs the development of the mammary gland, thus resulting in a longer period of susceptibility and increased risk of breast cancer. However, addition of aspirin can abrogate this effect.

Reduction of hexavalent chromium by fasted and fed human gastric fluid. I. Chemical reduction and mitigation of mutagenicity.

Evaluation of the reducing capacity of human gastric fluid from healthy individuals, under fasted and fed conditions, is critical for assessing the cancer hazard posed by ingested hexavalent chromium [Cr(VI)] and for developing quantitative physiologically-based pharmacokinetic models used in risk assessment. In the present study, the patterns of Cr(VI) reduction were evaluated in 16 paired pre- and post-meal gastric fluid samples collected from 8 healthy volunteers. Human gastric fluid was effective both in reducing Cr(VI), as measured by using the s-diphenylcarbazide colorimetric method, and in attenuating mutagenicity in the Ames test. The mean (±SE) Cr(VI)-reducing ability of post-meal samples (20.4±2.6µgCr(VI)/mL gastric fluid) was significantly higher than that of pre-meal samples (10.2±2.3µgCr(VI)/mL gastric fluid). When using the mutagenicity assay, the decrease of mutagenicity produced by pre-meal and post-meal samples corresponded to reduction of 13.3±1.9 and 25.6±2.8µgCr(VI)/mL gastric fluid, respectively. These data are comparable to parallel results conducted by using speciated isotope dilution mass spectrometry. Cr(VI) reduction was rapid, with >70% of total reduction occurring within 1min and 98% of reduction is achieved within 30min with post-meal gastric fluid at pH2.0. pH dependence was observed with decreasing Cr(VI) reducing capacity at higher pH. Attenuation of the mutagenic response is consistent with the lack of DNA damage observed in the gastrointestinal tract of rodents following administration of ≤180ppm Cr(VI) for up to 90days in drinking water. Quantifying Cr(VI) reduction kinetics in the human gastrointestinal tract is necessary for assessing the potential hazards posed by Cr(VI) in drinking water.

Reduction of hexavalent chromium by fasted and fed human gastric fluid. II. Ex vivo gastric reduction modeling.

To extend previous models of hexavalent chromium [Cr(VI)] reduction by gastric fluid (GF), ex vivo experiments were conducted to address data gaps and limitations identified with respect to (1) GF dilution in the model; (2) reduction of Cr(VI) in fed human GF samples; (3) the number of Cr(VI) reduction pools present in human GF under fed, fasted, and proton pump inhibitor (PPI)-use conditions; and (4) an appropriate form for the pH-dependence of Cr(VI) reduction rate constants. Rates and capacities of Cr(VI) reduction were characterized in gastric contents from fed and fasted volunteers, and from fasted pre-operative patients treated with PPIs. Reduction capacities were first estimated over a 4-h reduction period. Once reduction capacity was established, a dual-spike approach was used in speciated isotope dilution mass spectrometry analyses to characterize the concentration-dependence of the 2nd order reduction rate constants. These data, when combined with previously collected data, were well described by a three-pool model (pool 1 = fast reaction with low capacity; pool 2 = slow reaction with higher capacity; pool 3 = very slow reaction with higher capacity) using pH-dependent rate constants characterized by a piecewise, log-linear relationship. These data indicate that human gastric samples, like those collected from rats and mice, contain multiple pools of reducing agents, and low concentrations of Cr(VI) (<0.7 mg/L) are reduced more rapidly than high concentrations. The data and revised modeling results herein provide improved characterization of Cr(VI) gastric reduction kinetics, critical for Cr(VI) pharmacokinetic modeling and human health risk assessment.

Selective inhibition by aspirin and naproxen of mainstream cigarette smoke-induced genotoxicity and lung tumors in female mice.

The role of nonsteroidal anti-inflammatory drugs (NSAIDs) in smoke-related lung carcinogenesis is still controversial. We have developed and validated a murine model for evaluating the tumorigenicity of mainstream cigarette smoke (MCS) and its modulation by chemopreventive agents. In the present study, the protective effects of the nonselective cyclooxygenase inhibitors aspirin and naproxen were investigated by using a total of 277 Swiss H neonatal mice of both genders. Groups of mice were exposed whole-body to MCS during the first 4 months of life, followed by an additional 3.5 months in filtered air in order to allow a better growth of tumors. Aspirin (1600 mg/kg diet) and naproxen (320 mg/kg diet) were given after weanling until the end of the experiment. After 4 months of exposure, MCS significantly enhanced the frequency of micronucleated normochromatic erythrocytes in the peripheral blood of mice, and naproxen prevented such systemic genotoxic damage in female mice. After 7.5 months, exposure of mice to MCS resulted in the formation of lung tumors, both benign and malignant, and in several other histopathological lesions detectable both in the respiratory tract and in the urinary tract. Aspirin and, even more sharply, naproxen significantly inhibited the formation of lung tumors in MCS-exposed mice, but this protective effect selectively occurred in female mice only. These results lend support to the views that estrogens are involved in smoke-related pulmonary carcinogenesis and that NSAIDs have antiestrogenic properties. The two NSAIDs proved to be safe and efficacious in the experimental model used.

Modulation by aspirin and naproxen of nucleotide alterations and tumors in the lung of mice exposed to environmental cigarette smoke since birth.

Chemoprevention provides an important strategy for cancer control in passive smokers. Due to the crucial role played by smoke-related chronic inflammation in lung carcinogenesis, of special interest are extensively used pharmacological agents, such as nonsteroidal anti-inflammatory drugs (NSAIDs). We evaluated the ability of aspirin and naproxen, inhibitors of both cyclooxygenase-1 and cyclooxygenase -2, to modulate environmental cigarette smoke (ECS)-induced lung carcinogenesis in A/J mice of both genders. Based on a subchronic toxicity study in 180 postweaning mice, we used 1600 mg/kg diet aspirin and 320 mg/kg diet naproxen. In the tumor chemoprevention study, using 320 mice, exposure to ECS started soon after birth and administration of NSAIDs started after weaning. At 10 weeks of life, the NSAIDs did not affect the presence of occult blood in feces. As assessed in a subset of 40 mice, bulky DNA adducts and 8-hydroxy-2'-deoxyguanosine levels were considerably increased in ECS-exposed mice and, irrespective of gender, both NSAIDs remarkably inhibited these nucleotide alterations. After exposure for 4 months followed by 5 months in filtered air, ECS induced a significant increase in the yield of surface lung tumors, the 43.7% of which were adenomas and the 56.3% were adenocarcinomas. Oct-4 (octamer-binding transcription factor 4), a marker of cell stemness, was detected in some adenocarcinoma cells. The NAIDs attenuated the yield of lung tumors, but prevention of ECS-induced lung adenomas was statistically significant only in female mice treated with aspirin, which supports a role for estrogens in ECS-related lung carcinogenesis and highlights the antiestrogenic properties of NSAIDs.

Incidence of infection-associated cancers in Italy and prevention strategies.

Chronic infections and infestations represent major causes of cancer. Overall, Helicobacter pylori, HPV, HBV, and HCV are estimated to account for 15% of all human cancers. We have estimated that cancers associated with 6 pathogens in Italy account for 31,000 yearly cases, 42.0%of which is attributable to H. pylori, 34.7%to HBV and HCV, 19.8%to HPV, 2.9%to KSHV, and 0.2% to EBV. These figures represent 8.5% of all incident cases of cancer in Italy. The implementation of anti-HBV vaccination programs in countries with high endemicity resulted in a significant impact on the incidence of hepatocellular carcinoma, and the availability of antiviral drugs is a real opportunity to drastically reduce the cases attributable to HCV. Primary prevention of cervical cancer mainly involves HPV vaccination; two vaccines (bivalent and quadrivalent) are available and a new vaccine (9-valent) has recently been approved by the FDA. Secondary prevention is based on screening programs that include Pap smear cytology and/or HPV test. To reduce the burden of HIV-associated cancers, prevention programs include primary prevention of HIV infection, early diagnosis and treatment, restoration of immune function, reduction in the prevalence of associated infections and risk factors, and secondary prevention. To date, anti-HBV and anti-HPV vaccinations, eradication of H. pylori infection, treatment of HCV and HIV carriers with antivirals, and HPV-related cancer screening prove to be the most effective strategies for the prevention of infection-associated cancers.

Assay of lapatinib in murine models of cigarette smoke carcinogenesis.

Lapatinib, a dual tyrosine kinase inhibitor targeting the epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER-2), is prescribed for the treatment of patients with metastatic breast cancer overexpressing HER-2. Involvement of this drug in pulmonary carcinogenesis has been poorly investigated. We used murine models suitable to evaluate cigarette smoke-related molecular and histopathological alterations. A total of 481 Swiss H mice were used. The mice were exposed to mainstream cigarette smoke (MCS) during the first four months of life. After 10 weeks, MCS caused an elevation of bulky DNA adducts, oxidative DNA damage and an extensive downregulation of microRNAs in lung. After four months, an increase in micronucleus frequency was observed in peripheral blood erythrocytes. After 7.5 months, histopathological alterations were detected in the lung, also including benign tumors and malignant tumors, and in the urinary tract. A subchronic toxicity study assessed the non-toxic doses of lapatinib, administered daily with the diet after weaning. After 10 weeks, lapatinib significantly attenuated the MCS-related nucleotide changes and upregulated several low-intensity microRNAs in lung. The drug poorly affected the MCS systemic genotoxicity and had modest protective effects on MCS-induced preneoplastic lesions in lung and kidney, when administered under conditions that temporarily mimicked interventions either in current smokers or ex-smokers. On the other hand, it caused some toxicity to the liver. Thus, on the whole, lapatinib appears to have a low impact in the smoke-related lung carcinogenesis models used, especially in terms of tumorigenic response.

Rationale and approaches to the prevention of smoking-related diseases: overview of recent studies on chemoprevention of smoking-induced tumors in rodent models.

Tobacco smoke plays a dominant role in the epidemiology of lung cancer, cancer at other sites, and a variety of other chronic diseases. It is the leading cause of death in developed countries, and the global burden of cancer is escalating in less developed regions. For a rational implementation of strategies exploitable for the prevention smoking-related diseases, it is crucial to elucidate both the mechanisms of action of cigarette smoke and the protective mechanisms of the host organism. The imperative primary prevention goal is to avoid any type of exposure to smoke. Epidemiological studies have shown that a decrease in the consumption of cigarettes can be successful in attenuating the epidemic of lung cancer in several countries. Chemoprevention by means of dietary and/or pharmacological agents provides a complementary strategy aimed at decreasing the risk of developing smoking-associated diseases in addicted current smokers, who are unable to quit smoking, and especially in involuntary smokers and ex-smokers. The availability of new animal models that are suitable to detect the carcinogenicity of cigarette smoke and to assess the underlying molecular mechanisms provides new tools for evaluating both safety and efficacy of putative chemopreventive agents.

DNA damage in exfoliated cells and histopathological alterations in the urinary tract of mice exposed to cigarette smoke and treated with chemopreventive agents.

Cigarette smoke (CS) is convincingly carcinogenic in mice when exposure starts at birth. We investigated the induction and modulation of alterations in the kidney and urinary bladder of CS-exposed mice. A total of 484 strain H Swiss mice were either sham-exposed or exposed since birth to mainstream CS (MCS) for 4 months. Dietary agents, including myo-inositol, suberoylanilide hydroxamic acid, bexarotene, pioglitazone and a combination of bexarotene and pioglitazone, were administered after weaning. Comet analyses showed that, after 2 and 4 months, MCS causes DNA damage in exfoliated urothelial cells, which can be prevented by myo-inositol and the peroxisome proliferator-activated receptor-γ ligand pioglitazone. After 7 months, the 17.6% of MCS-exposed male mice exhibited lesions of the urinary tract versus the 6.1% of sham-exposed mice, which emphasizes the role of sex hormones in urinary tract carcinogenesis. Myo-inositol and the RXR-specific retinoid bexarotene did not affect these alterations. The histone deacetylase inhibitor suberoylanilide hydroxamic acid (Vorinostat) increased the incidence of kidney epithelium hyperplasia. Pioglitazone significantly enhanced the incidence of kidney lesions as compared with mice exposed to MCS only, indicating possible adverse effects of this antidiabetic drug, which were lost upon combination with bexarotene according to a combined chemoprevention strategy. RXR is a heterodymeric partner for peroxisome proliferator-activated receptor-γ, thereby modulating the expression of multiple target genes. In conclusion, there is contrast between the ability of pioglitazone to inhibit DNA damage in exfoliated cells and the alterations induced in the urinary tract of MCS-exposed mice, suggesting the occurrence of non-genotoxic mechanisms for this drug.

Relationships between pulmonary micro-RNA and proteome profiles, systemic cytogenetic damage and lung tumors in cigarette smoke-exposed mice treated with chemopreventive agents.

Assessing the correlation between molecular endpoints and cancer induction or prevention aims at validating the use of intermediate biomarkers. We previously developed murine models that are suitable to detect both the carcinogenicity of mainstream cigarette smoke (MCS) and the induction of molecular alterations. In this study, we used 931 Swiss mice in two parallel experiments and in a preliminary toxicity study. The chemopreventive agents included vorinostat, myo-inositol, bexarotene, pioglitazone and a combination of bexarotene and pioglitazone. Pulmonary micro-RNAs and proteins were evaluated by microarray analyses at 10 weeks of age in male and female mice, either unexposed or exposed to MCS since birth, and either untreated or receiving each one of the five chemopreventive regimens with the diet after weaning. At 4 months of age, the frequency of micronucleated normochromatic erythrocytes was evaluated. At 7 months, the lungs were subjected to standard histopathological analysis. The results showed that exposure to MCS significantly downregulated the expression of 79 of 694 lung micro-RNAs (11.4%) and upregulated 66 of 1164 proteins (5.7%). Administration of chemopreventive agents modulated the baseline micro-RNA and proteome profiles and reversed several MCS-induced alterations, with some intergender differences. The stronger protective effects were produced by the combination of bexarotene and pioglitazone, which also inhibited the MCS-induced clastogenic damage and the yield of malignant tumors. Pioglitazone alone increased the yield of lung adenomas. Thus, micro-RNAs, proteins, cytogenetic damage and lung tumors were closely related. The molecular biomarkers contributed to evaluate both protective and adverse effects of chemopreventive agents and highlighted the mechanisms involved.

Transplacental clastogenic and epigenetic effects of gold nanoparticles in mice.

The broad application of nanotechnology in medicine, biology, and pharmacology is leading to a dramatic increase of the risk of direct contact of nanoproducts, among which gold nanoparticles (AuNP), with the human organism. The present study aimed at evaluating in vivo the genotoxicity of AuNPs with average size of 40 nm and 100 nm. A single intraperitoneal treatment of adult male and female Swiss mice (strain H) with AuNPs, at a dose of 3.3 mg/kg body weight, had no effect on the frequency of micronucleated polychromatic erythrocytes (MN PCEs) in bone marrow. Conversely, the transplacental treatment with AuNP-100 nm, but not with AuNP-40 nm, applied intraperitoneally at a dose of 3.3 mg/kg to pregnant mice on days 10, 12, 14, and 17 of gestation, resulted in a significant increase in the frequency of MN PCEs in both liver and peripheral blood of mouse fetuses. In parallel, the same treatment with AuNP-100 nm, but not with AuNP-40 nm, produced significant changes in microRNA expression. In particular, out of 1281 mouse microRNAs analyzed, 28 were dys-regulated more than two-fold and to a statistically significant extent in fetus lung, and 5 were up-regulated in fetal liver. Let-7a and miR-183 were significantly up-regulated in both organs. The data presented herein demonstrate for the first time the transplacental size-dependent clastogenic and epigenetic effects of AuNPs in mouse fetus, thus highlighting new aspects concerning the putative genotoxicity of AuNPs during a vulnerable period of life.

Genotoxicity and carcinogenicity of the light emitted by artificial illumination systems.

The light delivered by artificial illumination systems, and in particular by halogen quartz bulbs, contains UVA, UVB, and UVC radiation, is genotoxic to both bacterial and human cells and is potently carcinogenic to hairless mice. Since IARC has classified UV radiation in Group 1, any source of UV light poses a carcinogenic hazard to humans. Suitable regulations would be needed in order to control the safety of the light emitted by artificial light sources.

Genotoxic damage in the oral mucosa cells of subjects carrying restorative dental fillings.

A large proportion of the population carries restorative dental fillings containing either classic Hg-based amalgams and/or the more frequently used methacrylates. Both Hg- and resin-based materials have been shown to be released into the buccal cavity and to be spread systemically. In addition, they induce toxic and genotoxic alterations in experimental test systems. Using the comet assay, we previously demonstrated that circulating lymphocytes of subjects with dental fillings have an increased DNA damage. Here, we analyzed the oral mucosa cells of 63 young subjects of both genders, by using both the comet assay and the micronucleus (MN) test and by monitoring cell death markers. The results obtained show that both amalgams and resin-based composite fillings can induce genotoxic damage in human oral mucosa cells, as convincingly and dose-dependently inferred from the results of the MN test and, more marginally, from comet assay data. Lifestyle variables, also including alcohol intake and smoking habits, did not affect the genotoxic response and did not act as confounding factors. Thus, we provide unequivocal evidence for the genotoxicity of both amalgams and resin-based dental fillings in humans not only by testing circulating lymphocytes but also by analyzing oral mucosa cells. These findings are of particular relevance due to the circumstance that subjects with restorative materials are exposed continuously and for long periods of time.

Oxidative stress in the lung of mice exposed to cigarette smoke either early in life or in adulthood.

Birth and early life stages are critical periods characterized by severe alterations of the redox balance and by "physiological" genomic changes in lung cells, which may be responsible for cancer and other diseases in adulthood. Oxidative stress is a major mechanism accounting for the carcinogenicity of cigarette smoke (CS), which becomes more potently carcinogenic in mice when exposure starts at birth and continues early in life. We compared herewith a variety of end-points related to oxidative stress, mitochondrial alterations, and cell turnover in the lung of Swiss H mice, either sham-exposed or CS-exposed for 4 weeks, starting either at birth or at 4 months of age. The results showed that the physiological levels of certain end-points are affected by age. In fact, the baseline proportion of hypodiploid cells and the mitochondrial potential and mass were higher in adults, whereas 8-hydroxy-2'-deoxyguanosine (8-oxo-dGuo) levels, the proportion of necrotic cells, and the extent of autophagy were higher early in life. Adult mice were more responsive to CS by increasing the proportion of necrotic cells and of cells in S/G2 phase, whereas young mice maintained a high extent of autophagy, exhibited a greater increase of lipid peroxidation products and 8-oxo-dGuo levels, and had a higher frequency of micronucleated cells. In addition, exposure to CS affected the mitochondrial potential/mass, especially in young mice. In conclusion, these data provide evidence that oxidative stress and the resulting DNA damage provide a major contribution to the high susceptibility of mice to CS early in life.

Differential carcinogenicity of cigarette smoke in mice exposed either transplacentally, early in life or in adulthood.

Cigarette smoke (CS) plays a dominant role in the epidemiology of human cancer. However, it is difficult to reproduce its carcinogenicity in laboratory animals. Recently, we showed that CS becomes a potent carcinogen in mice when exposure starts soon after birth. In our study, we comparatively evaluated the carcinogenic response to mainstream CS in mice at different ages. Neonatal mice were exposed daily for 4 months to CS, starting within 12 hr after birth, and sacrificed at 8 months. Adult mice were exposed for the same time period (3-7 months) and sacrificed at 11 months. Other mice were exposed transplacentally or both transplacentally and early in life. A total of 351 neonatal mice and 80 adult Swiss H mice were used. With varying intensity depending on age, CS induced pulmonary emphysema, bronchial and alveolar epithelial hyperplasia, blood vessel proliferation and hemangiomas and microadenomas in lung as well as parenchymal degeneration of liver. Histopathological alterations of kidney were only observed in mice exposed to CS early in life. Lung adenomas and malignant tumors of various histopathological nature were detected in neonatally exposed mice but not in adults. Transplacental CS induced the formation of lung adenomas in the offspring 8 months after birth. Previous exposure during pregnancy attenuated CS-related alveolar epithelial hyperplasia induced after birth. In conclusion, the carcinogenic response to CS varies depending on the developmental stage. The early postnatal life and the prenatal life are particularly at risk for the later development of CS-related tumors.