Cis-lycopene is more bioavailable than trans-lycopene in vitro and in vivo in lymph-cannulated ferrets.

Lycopene is the predominant carotenoid in tomatoes and tomato-based foods and is also a predominant carotenoid in human serum and tissues. Intake of lycopene-rich foods was recently associated with decreased risk for several chronic diseases. The observation that serum and tissue lycopene is more than 50% cis-lycopene, whereas tomatoes and tomato-based foods contain mainly all-trans-lycopene, has led to the hypothesis that cis-isomers of lycopene are more bioavailable. We tested this hypothesis both in vitro (study 1) and in vivo (study 2). In study 1, bile acid micelles containing crystalline lycopene were prepared. The crystalline lycopene used for these analyses was 54.4% cis-lycopene. The optically clear micelle preparation contained 75.9% cis-lycopene in repeated analyses. In study 2, mesenteric lymph duct cannulated ferrets were used to study the in vivo absorption of lycopene from LycoredTM (an ethyl acetate extract of tomatoes containing 5% lycopene by weight; of which 91% was all-trans lycopene). Before being anesthetized, male ferrets (n = 7) were dosed orally with 40 mg lycopene per kg body weight in soybean oil. Lymph secretions were collected, on ice, for 2 h. The residual stomach and small intestinal contents, mucosa lining, lymph secretion and serum were analyzed by HPLC. Whereas the dose, stomach and intestinal contents contained 6.2-17.5% cis-lycopene, the mesenteric lymph secretions contained significantly more, 77.4%, cis-lycopene (P < 0.01). These studies demonstrate that in ferrets, cis-isomers of lycopene are more bioavailable than trans-lycopene probably because cis-isomers are more soluble in bile acid micelles and may be preferentially incorporated into chylomicrons.     

Nutrikinetic studies of food bioactive compounds: from in vitro to in vivo approaches

ABSTRACT

Poor absorption is an important cause of costly late-stage failures in functional food development, and therefore, it has become widely appreciated that pharmacokinetic parameters should be considered as early as possible in the functional food development process. In many cases, the molecular structure of bioactive ingredients is known, but information is lacking on how they interact with other food components, what their fate is upon consumption, what they do in the body and what their target site is. This information is of major importance, as the biological effects of food bioactive compounds (CBAs) are ultimately determined by their bioavailability and their temporal and spatial distribution in the body. In this chapter, we propose the phases to perform nutrikinetic studies of food CBAs from the simplest in vitro assays, applicable in early stages of the development of a functional food, to human intervention studies, which are required by the European Food Safety Authority and are aimed to establish the dose–exposure relationship (pharmacokinetic studies) and at last the exposure–response relationship (pharmacodynamic studies).     

The most prevalent errors in screening studies

The article presents theoretical foundations of screening research in epidemiology. In planing of screening study several features of the disease should be considered such as severity and prevalence of studied disease as well as availability of treatment for it and also availability of appropriate tests which enable screening program at the affordable cost. Evaluation of screening may be based on the number of early diagnoses or on the difference in survival time between people who are screened and those who are not. Any analysis of the effects of screening program may have pitfalls leading to incorrect assessment. The most prevalent errors such as selection bias and measurement error are discussed.     

Benzotriazole is antiestrogenic in vitro but not in vivo

Benzotriazole (BT) is an anticorrosive agent well known for its use in aircraft deicing and antifreeze fluids but also used in dishwasher detergents. It is highly persistent in the environment; therefore, BT is frequently found in runoff emanating from large airports as well as in the surrounding groundwater. In addition, BT has recently been found to be ubiquitous in Swiss wastewater treatment plant effluents and their receiving waters; however, very little chronic toxicity data is available on which to base a sound ecological risk assessment of this chemical. In vitro assays conducted using a recombinant yeast (anti-) estrogen assay indicated that BT possessed clear antiestrogenic properties. This chemical was approximately 100-fold less potent than Tamoxifen, which was used as a positive control. A subsequent in vivo study, however, involving analysis of vitellogenin induction and somatic indices in adult fathead minnows (Pimephales promelas) exposed to BT at concentrations of 10, 100, and 1,000 mug/L for two weeks showed no evidence of antiestrogenic activity by this compound. The possibility exists that higher concentrations of BT may yet induce the type of activity observed in vitro, although the concentrations used here already far exceed those reported in surface-water samples. Furthermore, adverse effects may be observed in fish or other organisms exposed to BT for a longer period than employed here, although such studies are costly and unlikely to be included in standard risk assessment procedures. A rigorous investigation of the chronic toxicity of BT is imperative.     

Naringenin from cooked tomato paste is bioavailable in men

Naringenin has been shown to exert antiestrogenic, cholesterol-lowering and antioxidant activities, as well as an indirect modulation on the metabolism of many xenobiotics. It is one of the most abundant polyphenols in tomato. Given the widespread consumption of tomato (Lycopersicum esculentum) and tomato-based products, this study was designed to determine whether plasma levels of naringenin were detectable in five men after consumption of a test meal containing 150 mg of cooked tomato paste. Naringenin intake with the test meal was 3.8 mg. Blood was drawn from fasting subjects and 2, 4, 6, 8 and 24 h after the meal. To compare the results with a control, the same meal without tomato paste (control meal) was administered to the same subjects 2 wk later. Analyses were performed using high-performance liquid chromatography coupled with a CoulArray electrochemical detector. The peak plasma concentration was 0.12 +/- 0.03 micro mol/L 2 h after the meal. Unconjugated naringenin was not detected. Naringenin was not detected in plasma at any time after consumption of the control meal. In addition to naringenin, we detected rutin and chlorogenic acid in tomato paste, but these polyphenols and their derivatives (quercetin and caffeic acid) were not detected in plasma at any time. To the best of our knowledge, this is the first study demonstrating naringenin bioavailability in humans after consumption of a meal containing cooked tomato paste.     

Contraceptive efficacy of antimicrobial peptide Nisin: in vitro and in vivo studies

Sexually transmitted infections and unplanned pregnancies present a great risk to the reproductive health of women. Therefore, female-controlled vaginal products directed toward disease prevention and contraception are needed urgently. In the present study, efforts were made to evaluate the contraceptive potential of Nisin. The effect of Nisin on sperm motility was assessed under in vitro and in vivo conditions. The results showed that sperm motility was completely inhibited with Nisin. The minimum effective concentration of Nisin required to immobilize sperm (80-100 x 10(6)) in vitro within 20 s was found to be 50 microg in rat, 200 microg in rabbit and 300-400 microg in monkey and human. The effect on sperm motility was observed to be dose- and time-dependent. Intravaginal administration of Nisin (200 microg) before mating during proestrus-estrous transition phase caused complete arrest of sperm motility and blockage of conception. Subacute toxicity studies in rats indicated that, repetitive intravaginal application of Nisin at the dose of 200 microg for 14 consecutive days induced no abnormalities either in the length of estrous cycle or in the morphology of vaginal epithelial cells. No histopathological abnormalities in vaginal tissue or any change in blood and serum biochemical profiles were observed. Furthermore, no adverse effects were observed on subsequent reproductive performance, neonate survival and development of pups. It is suggested that Nisin, with its antibacterial and spermicidal activities, could be developed as a potent vaginal contraceptive for future use in humans.     

Cytogenetic studies of herbicide interactions in vitro and in vivo using atrazine and linuron

The herbicides atrazine and linuron, found in Wisconsin's groundwater, were tested alone and in combination, both in vivo and in vitro, to determine their individual and combined genotoxic effects. Human lymphocytes exposed in vitro to either 1 g/ml linuron or 0.001 g/ml atrazine showed little chromosome damage, whereas significant chromosome damage was observed in lymphocytes simultaneously exposed to 0.5 g/ml linuron and 0.0005 g/ml atrazine, suggesting at least an additive model. In another experiment, mice were fed 20 g/ml atrazine, 10 g/ml linuron, or a combination of 10 g/ml atrazine and 5 g/ml linuron in their drinking water for 90 days, after which bone marrow cells and cultured splenocytes were examined for chromosomal damage. None of the treatment groups showed chromosome damage in bone marrow, whereas the cultured splenocytes demonstrated damage in all treatment groups. These experiments suggest that, prior to assessing the risk of a herbicide, it may be necessary to test it in combinations which mimic the mixtures which would occur under field conditions, such as in contaminated groundwater.     

Potential bias due to prevalent diseases in prospective studies

In prospective studies, subjects found to have the disease under investigation at the initial screening examination are commonly excluded from analyses. However, the possibility of bias due to prevalent conditions other than the disease of interest is usually not considered. In the present study, an algebraic development enables analysis of the effects of inclusion and exclusion of subjects with certain prevalent conditions upon risk estimates. Hypothetical data are presented for which an association between a risk factor and an incident disease could become null or even reversed after removing subjects with certain prevalent diseases. Bias appears even when the only association present is between risk factor and total disease incidence. Data from the Honolulu Heart Study also have been used to illustrate this finding, examining the association between coronary heart disease (CHD) incidence and smoking. Decisions regarding the inclusion or exclusion of subjects with prevalent diseases requires prior knowledge of alteration of usual risk factors levels by individuals with these diseases. Simply removing all subjects with prevalent diseases might on the contrary create bias. Therefore, people with prevalent diseases should be screened for potential alteration of their risk factor levels as a result of the diseases. The situation becomes still more complex when several risk factors and prevalent diseases need to be considered at the same time as it happens in multivariate analyses. Because this situation represents a bias, and not confounding or effect modification, controlling for the effect of prevalent diseases is not appropriate.     

双酚A雄性生殖毒性的体内外实验研究

目的探讨双酚A对雄性动物生殖机能的影响。方法将双酚A混入饲料(0、1和5g／kg)连续饲喂成年32只SD大鼠14d,放免法测定睾酮和雌二醇并进行右睾组织形态分析;对原代培养的支持细胞染毒(0、10^-7、10^-6、10^-5、10^-4moL／L)。结果5g／kg的双酚A组的右睾平均重量1．53g显著低于对照组1．62g,但1g／kg组与对照组差异无统计学意义。形态观察发现,2个双酚A组中的曲细精管基底膜均与生精细胞分离;部分生精细胞和支持细胞发生核固缩和空泡变性;同时,双酚A处理使黏附于支持细胞的生精细胞平均数量由对照组的7．94个分别减少为4．13和3．04个。此外,双酚A在体内外实验中均抑制睾丸支持细胞的波形蛋白表达,阻碍细胞骨架和胞间联结的形成,使支持细胞在体外培养中的形态变得异常细长。但双酚A对血清雌二醇和睾酮浓度的影响没有统计学意义。结论双酚A可能通过破坏支持细胞骨架和改变支持细胞形态而损害雄性生殖功能。     

The toxicity of organoarsenic-based warfare agents: In vitro and in vivo studies

The four arsenic-containing chemical warfare agents (CWA) Adamsite (technical, 10-chloro-9–10-dihydrophenarsazine), Clark 1 (Diphenyl arsine chloride), Clark 2 (Diphenyl arsine cyanide), and Lewisite (2-chloro-ethenyl dichloro arsine) were evaluated toxicologically using an in vitro and an in vivo model. The CWA were tested in vitro, using human leucocytes, in order to evaluate their effects on cell proliferation and cell cycle kinetics assayed by flow cytometry. The concentration for total inhibition of cell proliferation for the CWAs ranged from 0.3 g/mL (Lewisite) to 15 g/mL (Clark 1). The results showed no differences in cell proliferation between the different stages of the S-phase either at no-effect or at effect concentrations when samples treated with the CWAs were compared with untreated controls. As₂O₅ was used as a reference, and the arsenic concentration required for total inhibition of the cell proliferation was 1.7 mg/mL. A lower arsenic concentration, 0.83 g/mL, showed a decreasing proliferation ratio in the different parts of the S-phase with S_early and S_late having the highest and lowest ratios, respectively. In addition, there was a positive correlation between the unlabeled S-phase cell ratio and the arsenic concentration, indicating cessation of DNA-synthesis. Conclusively, the examined CWAs exerts a toxicity more potent than arsenic, with respect to cell proliferation. This might indicate that the toxicity of these arsenic-containing CWAs only to a minor extent can be explained by their arsenic content. Because of their relatively good water solubility and ability to easily degrade to arsenic acid, organo-arsenics, e.g., Lewisite, pose less of an environmental threat than organo-arsenics that are sparingly soluble and chemically persistent, like Adamsite. The four CWAs were also tested in vivo in a dietary exposure study, using juvenile three-spined stickleback (Gasterosteus aculeatus L.) as test organism, with measurement of EROD-activity and studies of liver morphology. The results from the in vivo study indicated small effects, suggesting that a 10-week period of dietary exposure at tested dose levels (1 and 100 ng/ml) affects juvenile three-spined stickleback only to a minor extent.     

A review of depleted uranium biological effects: in vitro and in vivo studies

The use of depleted uranium in armor-penetrating munitions remains a source of controversy because of the numerous unanswered questions about its long-term health effects. Although no conclusive epidemiologic data have correlated DU exposure to specific health effects, studies using cultured cells and laboratory rodents continue to suggest the possibility of leukemogenic, genetic, reproductive, and neurological effects from chronic exposure. Until issues of concern are resolved with further research, the use of depleted uranium by the military will continue to be controversial.     

Cereal fructans: in vitro and in vivo studies on availability in rats and humans

The bioavailability of cereal fructans (fructooligosaccharides) was investigated both in vitro and in vivo. In vitro studies indicated very slow hydrolysis by human gastric juice and by homogenate of the intestinal mucosa (rat). After intubation of fructans into the stomachs of rats, the recovery of fructans in the small intestine and colon was approximately the same as that of an unabsorbed marker (polyethylene glycol), indicating no or very low disappearance of fructans in the small intestine. In vivo studies of the small intestine in rats showed that the rate of disappearance of fructans was lower than that of mannose, which is known to be absorbed through passive diffusion. In addition the cariogenic effect of cereal fructans was compared to that of glucose. Acid formation from low molecular-weight fructans was found in human dental plaque in vitro. A mouth rinse with unfractionated fructans, containing some quantities of sucrose, fructose and glucose, resulted in relatively low pH values in human plaque in vivo, even if the decrease in pH was somewhat less pronounced when compared with a mouth rinse with glucose.     

Lipid nanoparticles for transdermal delivery of flurbiprofen: formulation, in vitro, ex vivo and in vivo studies

The aim of the study is to prepare aqueous dispersions of lipid nanoparticles – flurbiprofen solid lipid nanoparticles (FLUSLN) and flurbiprofen nanostructured lipid carriers (FLUNLC) by hot homogenization followed by sonication technique and then incorporated into the freshly prepared hydrogels for transdermal delivery. They are characterized for particle size, for all the formulations, more than 50% of the particles were below 300 nm after 90 days of storage at RT. DSC analyses were performed to characterize the state of drug and lipid modification. Shape and surface morphology were determined by TEM which revealed fairly spherical shape of the formulations. Further they were evaluated for in vitro drug release characteristics, rheological behaviour, pharmacokinetic and pharmacodynamic studies. The pharmacokinetics of flurbiprofen in rats following application of SLN gel (A1) and NLC gel (B1) for 24 h were evaluated. The C_max of the B1 formulation was 38.67 ± 2.77 μg/ml, which was significantly higher than the A1 formulation (C_max = 21.79 ± 2.96 μg/ml). The C_max and AUC of the B1 formulation were 1.8 and 2.5 times higher than the A1 gel formulation respectively. The bioavailability of flurbiprofen with reference to oral administration was found to increase by 4.4 times when gel formulations were applied. Anti-inflammatory effect in the Carrageenan-induced paw edema in rat was significantly higher for B1 and A1 formulation than the orally administered flurbiprofen. Both the SLN and NLC dispersions and gels enriched with SLN and NLC possessed a sustained drug release over period of 24 h but the sustained effect was more pronounced with the SLN and NLC gel     

Adjustment of prognostic effects in prevalent case-control studies on genotype

Since genotypes are unchangeable, adjustment of prognostic effects in prevalent case-control studies may produce an unbiased estimate of odds ratio (OR) for disease occurrence. In this paper, the prognostic effects on OR is demonstrated, then three approaches to examine and/or adjust the OR are presented. The demonstration shows that the prognostic effects are larger in diseases with poor prognosis than in those with better prognosis. Genotypes increasing disease risk and fatality rate are underestimated, while those increasing the risk and improving prognosis are overestimated. The simplest approach to examine the OR derived from prevalent case-control studies is to conduct stratified analysis according to the interval between diagnosis and study enrollment. When the stratified analysis finds no substantial difference in the estimate, the OR reflects mainly the relative risk for disease occurrence. The proportion of genotype among putative cases at diagnosis can be estimated from prevalent cases by a logistic model, producing the OR adjusted for the interval from diagnosis. An incomplete-data case-control design is also applicable to adjust the prognostic effects. An actual prevalent case-control study on breast cancer is used to demonstrate the three approaches. They are useful to compensate the disadvantage of prevalent case-control studies.     

Modulators of cellular senescence: mechanisms,promises, and challenges from in vitro studies with dietary bioactive compounds

Cellular senescence is considered an important mechanism to prevent malignant transformation of potentially mutated cells but, persistence of senescent cells within tissues alters microenvironment in ways that can promote cancer and aging phenotype thus underlining pathophysiologic processes of different age-related diseases. Coincident with this increased knowledge, understanding and finding modulators of the dynamics that control senescent-cell formation, fate and subsequent effect on tissue function has gained critical interest in experimental gerontology and cancer research. The purpose of this review is to discuss the evidence that various dietary bioactive compounds can modulate cellular senescence in vitro and to summarize findings and mechanisms that might be useful for the development of health-promoting nutraceuticals. An overview of cellular senescence and its impact in aging and cancer is described along with the strategies and pathways that are currently being investigated to target cellular senescence. Particular emphasis is given to the mechanisms by which bioactive dietary factors (i.e. most polyphenols) can delay or induce cellular senescence in vitro and how this knowledge could be used to explain the opposite effects shown in cancer lines and primary cells by some of these compounds. In addition, the problems to translate findings from modulation of cellular senescence in vitro into experimental treatments or clinical trials able to prevent or counteract age-related diseases are briefly described. The information herein provided might be useful to design further research in the field as well as to develop new nutraceuticals to be tested in experimental models and clinical trials.