Assessment of genetic damage induced by dental bleaching agents on mouse lymphoma cells by single cell gel (comet) assay

Dental bleaching is a simple and conservative procedure for aesthetic restoration of vital discoloured teeth. However, dental bleaching agents may represent a hazard to human health, especially by causing DNA strand breaks. Genotoxicity tests form an important part of cancer research and risk assessment of potential carcinogens. In the current study, the genotoxic potential associated with exposure to dental bleaching agents was assessed by the single cell gel (comet) assay in vitro. Six commercial dental bleaching agents (Clarigel Gold - Dentsply; Whitespeed - Discus Dental; Nite White - Discus Dental; Magic Bleaching - Vigodent; Whiteness HP - FGM and Lase Peroxide - DMC) were exposed to mouse lymphoma cells in vitro. The results pointed out that all dental bleaching agents tested contributed to the DNA damage as depicted by the mean tail moment. Clear concentration-related effects were obtained for DNA damaging, being the strongest effect observed at the highest dose of the hydrogen peroxide (Whiteness HP and Lase Peroxide, at 35% concentration). on the contrary, Whitespeed (Discus Dental) induced the lowest level of DNA breakage. Taken together, these results suggest that dental bleaching agents may be a factor that increases the level of DNA damage as detected by the single cell gel (comet) assay.

Tomato-oleoresin supplement prevents doxorubicin-induced cardiac myocyte oxidative DNA damage in rats

Doxorubicin (DOX) is an efficient chemotherapeutic agent used against several types of tumors; however, its use is limited due to severe cardiotoxicity. Since it is accepted that reactive oxygen species are involved in DOX-induced cardiotoxicity, antioxidant agents have been used to attenuate its side effects. To determine tomato-oleoresin protection against cardiac oxidative DNA damage induced by DOX, we distributed Wistar male rats in control (C), lycopene (L), DOX (D) and DOX+lycopene (DL) groups. They received corn oil (C, D) or tomato-oleoresin (5 mg/kg body wt. day) (L, DL) by gavage for a 7-week period. They also received saline (C, L) or DOX (4 ma/kg body wt.) (D, DL) intraperitoneally at the 3rd, 4th, 5th, and at 6th week. Lycopene absorption was checked by HPLC. Cardiac oxidative DNA damage was evaluated by the alkaline Comet assay using formamidopyrimidine-DNA glycosylase (FPG) and endonuclease III (endo 111). Cardiomyocyte levels of SBs, SBs FPG and SBs Endo III were higher in rats from D when compared to other groups. DNA damage levels in cardiomyocytes from DL were not different when compared to C and L groups. The viability of cardiomyocytes from D or DL was lower than C or L groups (p < 0.01). Lycopene levels (mean +/- S.D. nmol/kg) in saponified hearts were similar between L (47.43 +/- 11.78) and DL (49.85 +/- 16.24) groups. Our results showed: (1) lycopene absorption was confirmed by its cardiac levels; (2) DOX-induced oxidative DNA damage in cardiomyocyte; (3) tomato-oleoresin supplementation protected against cardiomyocyte oxidative DNA damage. (c) 2007 Elsevier B.V. All rights reserved.

Modifying effect of propolis on dimethylhydrazine-induced DNA damage but not colonic aberrant crypt foci in rats

Propolis is a honeybee product with several biological and therapeutic properties, including antimutagenic and anticarcinogenic activities. The effects of an aqueous extract of propolis (AEP) were evaluated on the formation of 1,2-dimethylhydrazine (DMH)-induced aberrant crypt foci (ACF) and DNA damage in the colon of male Wistar rats by the ACF and Comet assays, respectively. AEP was administered orally at 0.01%, 0.03%, 0.1%, and 0.3% in the drinking water, which resulted in doses of approximately 12, 34, 108, and 336 mg/kg body weight/day. Animals were also given a single subcutaneous injection of 40 mg/kg DMH and sacrificed 4 hr later for evaluating DNA damage, or 4 doses of 40 mg/kg DMH, administered 2 doses/week for 2 weeks, and sacrificed 12 weeks after the last injection for evaluating ACF development in the distal colon. Administration of AEP either simultaneously with or after the DMH treatment resulted in no statistically significant reduction of ACF. In contrast, 0.01%, 0.03%, and 0.3% AEP, given simultaneously with DMH, reduced DNA damage induction in the mid and distal colon. However, 0.3% AEP alone increased DNA damage in the colon. In conclusion, AEP had no effect on the formation of DMH-induced ACF in rat colon, but it modulated DMH-induced DNA damage in colon cells. Further investigations are recommended in order to establish the conditions under which propolis produces either protective or deleterious effects. (C) 2004 Wiley-Liss, Inc.

Mate attenuates DNA damage and carcinogenesis induced by diethylnitrosamine and thermal injury in rat esophagus

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Tomato oleoresin inhibits DNA damage but not diethylnitrosamine-induced rat hepatocarcinogenesis

Various studies have shown that lycopene, a non-provitamin A carotenoid, exerts antioxidant, antimutagenic and anticarcinogenic activities in different in vitro and in vivo systems. However, the results concerning its chemopreventive potential on rat hepatocarcinogenesis are ambiguous. The aim of the present study was to investigate the antigenotoxic and anticarcinogenic effects of dietary tomato oleoresin adjusted to lycopene concentration at 30, 100 or 300ppm (administered 2 weeks before and during or 8 weeks after carcinogen exposure) on liver of male Wistar rats treated with a single intraperitoneal dose of 20 or 100 mg/kg of diethylnitrosamine (DEN), respectively. The level of DNA damage in liver cells and the development of putative preneoplastic single hepatocytes, minifoci and foci of altered hepatocytes (FHA) positive for glutathione S-transferase (GST-P) were used as endpoints. Significant reduction of DNA damage was detected when the highest lycopene concentration was administered before and during the DEN exposure (20 mg/kg). However, the results also showed that lycopene consumption did not reduce cell proliferation in normal hepatocytes or the growth of initiated hepatocytes into minifoci positive for GST-P during early regenerative response after 70% partial hepatectomy, or the number and area of GST-P positive FHA induced by DEN (100 mg/ kg) at the end of week 10. Taken together, the data suggest a chemopreventive effect of tomato oleoresin against DNA damage induced by DEN but no clear effectiveness in initiating or promoting phases of rat hepatocarcinogenesis. (c) 2008 Elsevier GmbH. All rights reserved.

Protective effects of carvedilol on systemic vascular damage induced by angiotensin II: Organ-specific effects independent of antihypertensive effects

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Histological analyses of thermal effect caused by 1.2 W diode laser irradiation at rat periodontal pockets

The aim of this in vivo study was to evaluate the thermal effects caused by 810 nm 1.2 W diode laser irradiation of periodontal tissues. Despite all data available concerning the laser application for periodontal treatment, one of the most relevant challenges is to prevent the harmful tissue heating induced by different clinical protocols. Periodontal pockets were induced at molars in 96 rats. Several irradiation powers under CW mode were investigated: 0, 400, 600, 800, 1000, 1200 mW. The pockets were irradiated using a 300 A mu m frontal illumination fiber. The animals were killed at 4 or 10 days after irradiation. The mandible was surgically removed and histologically processed. The histological sections stained with H/E demonstrated that irradiation parameters up to 1000 mW were thermally safe for the periodontal tissues. The sections stained with Brown & Brenn technique evidenced bacteria in the periodontal tissues. Consequently, the diode laser irradiation as a unique treatment was not capable to eliminate bacteria of the biofilm present in the pockets. According to the methodology used here, it was concluded that the thermal variation promoted by a diode laser can cause damage to periodontal tissues depending on the energy density used. The 1.2 W diode laser irradiation itself does not control the bacteria present in the biofilm of the periodontal pockets without mechanical action. The knowledge of proper high intensity laser parameters and methods of irradiation for periodontal protocols may prevent any undesirable thermal damage to the tissues.

Damage of the medial preoptic area impairs peripheral pilocarpine-induced salivary secretion

The existence of neural connections between the medial preoptic area (MPOA) and the salivary glands and the increase in salivation by thermal or electrical stimulation of the MPOA have suggested an important role of MPOA in the control of salivary gland function. Although direct cholinergic activation of the salivary glands induces salivation, recent studies have suggested that salivation produced by i.p. pilocarpine may also depend on the activation of central mechanisms. Therefore, in the present study, we investigated the effects of bilateral electrolytic lesions of the MPOA on the salivation induced by i.p. pilocarpine. Adult male Holtzman rats (n = 11-12/group) with bilateral sham or electrolytic lesions of the MPOA were used. One, five, and fifteen days after the brain surgery, under ketamine anesthesia, the salivation was induced by i.p. pilocarpine (1 mg/kg of body weight), and saliva was collected using preweighted small cotton balls inserted into the animal's mouth. Pilocarpine-induced salivation was reduced 1 and 5 days after MPOA lesion (341 +/- 41 and 310 +/- 35 mg/7 min, respectively, vs. sham lesions 428 +/- 32 and 495 +/- 36 mg/7 min, respectively), but it was fully recovered at the 15th day post-lesion (561 +/- 49 vs. sham lesion: 618 27 mg/7 min). Lesions of the MPOA did not affect baseline non-stimulated salivary secretion. The results confirm the importance of MPOA in the control of salivation and suggest that its integrity is necessary for the full sialogogue effect of pilocarpine. However, alternative mechanisms probably involving other central nuclei can replace MPOA function in chronically lesioned rats allowing the complete recovery of the effects of pilocarpine. (c) 2006 Published by Elsevier B.V.

Myocardial dysfunction induced by food restriction is related to morphological damage in normotensive middle-aged rats

Previous works from our laboratory have revealed that food restriction (FR) promotes discrete myocardial dysfunction in young rats. We examined the effects of FR on cardiac function, in vivo and in vitro, and ultrastructural changes in the heart of middle-aged rats. Twelve-month-old Wistar- Kyoto rats were fed a control (C) or restricted diet (daily intake reduced to 50% of the control group) for 90 days. Cardiac performance was studied by echocardiogram and in isolated left ventricular (LV) papillary muscle by isometric contraction in basal condition, after calcium chloride (5.2 mM) and beta- adrenergic stimulation with isoproterenol (10(-6) M). FR did not change left ventricular function, but increased time to peak tension, and decreased maximum rate of papillary muscle tension development. Inotropic maneuvers promoted similar effects in both groups. Ultrastructural alterations were seen in most FR rat muscle fibers and included, absence and/or disorganization of myofilaments and Z line, hyper-contracted myofibrils, polymorphic and swollen mitochondria with disorganized cristae, and a great quantity of collagen fibrils. In conclusion, cardiac muscle sensitivity to isoproterenol and elevation of extracellular calcium concentration is preserved in middle-aged FR rats. The intrinsic muscle performance depression might be related to morphological damage.

Antioxidant activity of indigo and its preventive effect against ethanol-induced DNA damage in rat gastric mucosa

Ethanol-induced oxidative damage is commonly associated with the generation of reactive oxygen molecules, leading to oxidative stress. Considering that antioxidant activity is an important mechanism of action involved in cytoprotection, the aim of this work was to evaluate the antioxidant properties of the alkaloid indigo (1) (2 mg/kg, p. o.), obtained from the leaves of Indigofera truxillensis Kunth (Fabaceae), on rat gastric mucosa submitted to ethanol-induced (100%, 1 mL, p.o.) gastric ulcer. Enzymatic assays and DNA fragmentation analysis were performed. When ethanol was administered to the control group, the sulfhydryl content (SH) and the glutathione peroxidase (GPx) activity decreased by 41% and 50%, respectively; in contrast, superoxide dismutase (SOD) and glutathione reductase (GR) activities increased by 56% and 67%, respectively. Additionally, myeloperoxidase (MPO) activity, a marker for free radical generation caused by polymorphonuclear neutrophil (PMN) tissue infiltration, also increased 4.5-fold after ethanol treatment. Rat gastric mucosa exposed to ethanol showed DNA fragmentation. Indigo alkaloid pretreatment protected rats from ethanol-induced gastric lesions. This effect was determined by the ulcerative lesion area (ULA), indicating an inhibition of around 80% at 2 mg/kg. This alkaloid also diminished GPx activity, which was higher than that observed with ethanol alone. However, this effect was counterbalanced by increased GR activity. Indigo was unable to restore alterations in SOD activity promoted by ethanol. After indigo pretreatment, SH levels and MPO activity remained normal and gastric mucosa DNA damage caused by ethanol was also partially prevented by indigo. These results suggest that the gastroprotective mechanisms of indigo include non-enzymatic antioxidant effects and the inhibition of PMN infiltration which, in combination, partially protect the gastric mucosa against ethanol-induced DNA damage.

Anti-clastogenic effect of beta-glucan extracted from barley towards chemically induced DNA damage in rodent cells

beta-Glucan (BG) was tested in vitro to determine its potential clastogenic and/or anti-clastogenic activity, and attempts were made to elucidate its possible mechanism of action by using combinations with an inhibitor of DNA polymerase. The study was carried out on cells deficient (CHO-k1) and cells proficient (HTC) in phases I and II enzymes, and the DNA damage was assessed by the chromosomal aberration assay. BG did not show a clastogenic effect, but was anti-clastogenic in both cell lines used, and at all concentrations tested (2.5, 5 and 10 mg/mL) in combination with damage inducing agents (methylmethane sulfonate in cell line CHO-k1, and methylmethane sulfonate or 2-aminoanthracene in cell line HTC). BG also showed a protective effect in the presence of a DNA polymerase beta inhibitor (cytosine arabinoside-3-phosphate, Ara-C), demonstrating that BG does not act through an anti-mutagenic mechanism of action involving DNA polymerase beta.

Protective effects of beta-glucan extracted from Agaricus brasiliensis against chemically induced DNA damage in human lymphocytes

beta-Glucans (BGs) are polysaccharides that are found in the cell walls of organisms such as bacteria, fungi, and some cereals. The objective of the present study was to investigate the genotoxic and antigenotoxic effects of BG extracted from the mushroom Agaricus brasiliensis (=Agaricus blazei Murrill ss. Heinemann). The mutagenic activity of BG was tested in single-cell gel electrophoresis assays with human peripheral lymphocytes. In addition, the protective effects against the cooked food mutagen 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) and (+/-)-anti-B[a]P-7,8-dihydrodiol-9,10-epoxide (BPDE), which is the main metabolite of B[a]P, and against ROS (H2O2)-induced DNA damage, were studied. The results showed that the compound itself was devoid of mutagenic activity, and that a significant dose-dependent protective effect against damage induced by hydrogen peroxide and Trp-P-2 occurred in the dose range 20-80 mu g/ml. To investigate the prevention of Trp-P-2-induced DNA damage, a binding assay was carried out to determine whether BG inactivates the amine via direct binding. Since no such interactions were observed, it is likely that BG interacts with enzymes involved in the metabolism of the amine.

Laser treatment of experimentally induced chronic arthritis

In this work, we investigated the effects of He-Ne laser irradiation on the inflammatory process induced in the articular cartilage of the right knee of guinea pigs. Through electron microscopy analysis it was possible to identify the induced arthritis in the articular cartilage and its modification after the laser treatment. The laser radiation promoted a reduction in the proliferation of the inflammatory cells in the damaged tissue and also induced the formation of cartilage bridges that tied the destroyed parts favoring the formation of a repaired tissue in the injured cartilage. (C) 2000 Elsevier B.V. B.V. All rights reserved.

Advantages of using nested collision induced dissociation/post-source decay with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry: sequencing of novel peptides from wasp venom

We have examined the applicability of the 'nested' collision induced dissociation/post-source decay (CID/PSD) method to the sequencing of novel peptides from solitary wasps which have neurotoxic venom for paralyzing other insects. The CID/PSD spectrum of a ladder peptide derived from an exopeptidase digest was compared with that of the intact peptide. The mass peaks observed only in the CID/PSD spectrum of a ladder peptide were extracted as C-terminal fragment ions. Assignment of C-terminal fragment ions enabled calculation of N-terminal fragment masses, leading to differentiation between N-terminal fragment ions and internal fragment ions. This methodology allowed rapid and sensitive identification by removing ambiguity in the assignment of the fragment ions, and proved useful for sequencing unknown peptides, in particular those available as natural products with a limited supply. Copyright (C) 2000 John Wiley & Sons, Ltd.

Protective effects of beta-glucan extracted from barley against benzo[a]pyrene-induced DNA damage in hepatic cell HepG2

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)