Lack of genotoxicity of rhubarb (rhizome) in the Ames and micronucleus in vitro tests.

Hydroxyanthracene derivatives are widely distributed in the plant kingdom, mainly in botanicals such as the Hypericum, Rheum, Rhamnus and Aloe genera. For centuries, plants containing hydroxyanthracene derivatives have been used as herbal remedies, mainly as laxatives. The root and underground stem (rhizome) are used to make medicine, primarily for digestive complaints including constipation, diarrhoea, heartburn, stomach pain, gastrointestinal bleeding, and preparation for certain gastrointestinal diagnostic procedures. The use of hydroxyanthracene-containing botanicals has raised the attention of European Food Safety Authority (EFSA) for the potential genotoxicity activity, that in 2018 concluded "[.] and that there is a safety concern for extracts containing hydroxyanthracene derivatives although uncertainty persists". No genotoxic activity has been reported with other constituents such as rhein, physcion and chrysophanol. In the present study, Rhubarb ethanolic extract of ground rhubarb rhizome (hydroxyanthracene total content 1.39 %) was tested in the Ames Assay in Salmonella typhimurium and Escherichia coli, up to 5000 µg/plate and up to 5000 µg/mL in human lymphocytes Micronucleus Test (OECD 471 and 487 respectively) in vitro mutagenic and genotoxic effects. Under the experimental conditions used, the rhubarb rhizome extract showed no genotoxic activity.

Lack of in vivo genotoxic effect of dried whole Aloe ferox juice.

Aloe ferox Mill is widely used as a traditional herbal medicine for the treatment of a broad spectrum of illnesses given its laxative, anti-inflammatory, bitter tonic, anti-oxidant, antimicrobial and anti-cancer properties. Using the in vivo alkaline comet assay in animals (OECD 489), this study investigated the potential in vivo genotoxicity of dried Aloe ferox juice at dose levels of 500, 1000, and 2000 mg/kg/day in mice. Aloe ferox showed no genotoxic activity in preparations of single cells from the colon of the treated Hsd:ICR (CD-1) male mice. No statistically significant increase in DNA migration over the negative control was observed by analysis of variance for both comet parameters, tail moment and tail intensity, apart from the positive control ethyl methanesulphonate that induced clear and statistically significant increases in DNA migration parameters over the concurrent controls. The new reported scientific evidence unequivocally demonstrates that dried Aloe ferox juice containing hydroxyanthracene derivatives does not induce DNA damage in preparations of single cells from colon in in vivo comet genotoxicity studies. This suggests that the hyperplastic changes and mucosal hyperplasia observed after long-term administration of Aloe vera non-decolourised whole leaf extract may be attributed to an epigenetic effect of the material under investigation.

Rhus coriaria L. Fruit Extract Prevents UV-A-Induced Genotoxicity and Oxidative Injury in Human Microvascular Endothelial Cells.

Rhus coriaria L. (sumac) is a small plant widely diffused in the Mediterranean region. Its fruit are often consumed as a spice but are also present in traditional medicine of several countries. Recently, interest in this plant has increased and many scientific works reported its beneficial effects including antioxidant and anti-inflammatory properties. Plant extracts can be successfully used against ultraviolet rays, which are able to reach and damage the human skin; however, sumac extracts were never applied to this usage. Thus, in this study, we used a macerated ethanol extract of Rhus coriaria L. dried fruit (mERC) to demonstrate its preventive role against the damage induced by ultraviolet-A rays (UV-A) on microvascular endothelial cells (HMEC-1). In vitro effects of the extract pre-treatment and UV-A exposure were evaluated in detail. The antioxidant capacity was assessed by reactive oxygen species (ROS) formation and cellular antioxidant activity measurement. Genoprotective effects of mERC were investigated as well. Our findings indicate that the extract acts as a cell cycle inhibitor or apoptosis inducer, according to the level of damage. The present work provides new insights into the usage of Rhus coriaria extracts against skin injuries.

Effects of Vitis vinifera L. leaves extract on UV radiation damage in human keratinocytes (HaCaT).

Vitis vinifera L. water extract from red grapevine leaves contains high levels of polyphenols in quantities similar to those found in red grape and grape seeds. Phenolic compounds are the largest group of natural antioxidants with also an anti-inflammatory activity, widely demonstrated both in vitro and in vivo. Interestingly, their antioxidant effect relies not only on the direct radical scavenging activity but also on their ability in modulating cellular signalling transduction pathways. UV radiation exerts multiple effects on skin cells inducing apoptosis, senescence and carcinogenesis. The aim of this study was to investigate the antioxidant and the DNA protective potentials of Vitis vinifera L. water extract against UV-A and UV-B radiation in HaCaT cells, a human keratinocytes cell line. Comet and É£H2AX assays were used to assess DNA damage in UV irradiated cells pre-treated or not with the extract (100 µg/mL). For UV-B, DNA damage resulted significantly increased at 40 mJ/cm2 dose determining cell cycle arrest and apoptosis. For UV-A, DNA damage was significant at 10 J/cm2 while cell cycle arrest and apoptosis were evident only at 25 J/cm2. The extract (1h of pre-treatment) highlights the antioxidant and scavenger activity on the UV-A, while the maintenance of the apoptosis with both UV-A and UV-B must be interpreted as an anti-mutagenic effect.

Effect of plant extracts on the genotoxicity of 1'-hydroxy alkenylbenzenes.

Food-borne alkenylbenzenes are potential risks for human health because they are known to induce liver tumors in rodent bioassays at high dose levels. This carcinogenicity is ascribed to the conversion of their 1'-hydroxymetabolites to the ultimate DNA reactive and carcinogenic 1'-sulfoxymetabolites. The aim of this study was to investigate the in vitro genotoxicity of some botanical extracts used as Plant Food Supplements (PFS) and to compare it with the individual substances, estragole, safrole and their 1'-hydroxy-derivative activity. The genotoxicity of the PFSs was evaluated in HepG2 cell line by comet and micronucleus assays. Unlike the 1'-hydroxy derivatives, PFS extracts and parent alkenylbenzenes did not show genotoxicity at any of the tested concentrations. The sulfotransferase inhibitor pentachlorophenol (PCP) reduced the 1'-hydroxy compound-induced response in the comet and micronucleus assays, thus confirming that the formation of sulfoxy-metabolites is essential for inducing genotoxic effects. When the cells were treated with hydroxylated alkenylbenzenes in the presence of PFSs, a reduction in genotoxic activity of synthetic compounds was observed.

Optimization of the THP-1 activation assay to detect pharmaceuticals with potential to cause immune mediated drug reactions.

Despite important impacts of systemic hypersensitivity induced by pharmaceuticals, for such endpoint no reliable preclinical approaches are available. We previously established an in vitro test to identify contact and respiratory allergens based on interleukin-8 (IL-8) production in THP-1 cells. Here, we challenged it for identification of pharmaceuticals associated with systemic hypersensitivity reactions, with the idea that drug sensitizers share common mechanisms of cell activation. Cells were exposed to drugs associated with systemic hypersensitivity reactions (streptozotocin, sulfamethoxazole, neomycin, probenecid, clonidine, procainamide, ofloxacin, methyl salicylate), while metformin was used as negative drug. Differently to chemicals, drugs tested were well tolerated, except clonidine and probenecid, with no signs of cytotoxicity up to 1-2mg/ml. THP-1 activation assay was adjusted, and conditions, that allow identification of all sensitizing drugs tested, were established. Next, using streptozotocin and selective inhibitors of PKC-ß and p38 MAPK, two pathways involved in chemical allergen-induced cell activation, we tested the hypothesis that similar pathways were also involved in drug-induced IL-8 production and CD86 upregulation. Results indicated that drugs and chemical allergens share similar activation pathways. Finally, we made a structure-activity hypothesis related to hypersensitivity reactions, trying to individuate structural requisite that can be involved in immune mediated adverse reactions.

High interleukin-10 production is associated with low antibody response to influenza vaccination in the elderly.

The present study was designed to determine the correlation among dehydroepiandrosterone (DHEA), cortisol plasma levels, and immune functionality at the time of vaccination with antibody response to influenza vaccination in young and old, healthy volunteers. Fifty-two elderly subjects, ages 63-85 years, and 14 young subjects, ages 26-41 years, entered the study. Plasma levels of DHEA and cortisol and in vitro cytokine production in response to lipopolysaccharide (LPS) and phytohaemagglutinin (PHA) by peripheral blood leukocytes were assessed at the time of vaccination, and antibody titer was measured before and 18 days after influenza virus vaccination. Elderly subjects were characterized by an increase in the cortisol:DHEA ratio, mainly as a result of a decrease in DHEA. A decrease in LPS-induced tumor necrosis factor alpha (TNF-alpha), increased PHA-induced interleukin-10 (IL-10) release, and similar PHA-induced interferon-gamma production were observed in elderly subjects compared with young volunteers. Lower antibody titer to influenza A virus was observed in elderly individuals, and the seroconversion factor was found to be correlated inversely with IL-10 production and correlated directly with TNF-alpha production and to a lesser extent, with the plasma level of DHEA. These results suggest that altered cytokine production in elderly subjects at the moment of vaccination can be predictive of a low response to influenza vaccination and warrant the study of strategies to improve protection afforded by the use of vaccines.