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PubMed2002

Evaluation of the potential effects of ingredients added to cigarettes. Part 3: in vitro genotoxicity and cytotoxicity.

Cigarette mainstream smoke from blended cigarettes with and without the addition of ingredients was assayed for its cytotoxicity and genotoxicity. In total, 333 ingredients commonly used in cigarette manufacturing were assigned to three different groups. Each group of ingredients was added at a low and a high level to the test cigarettes. The mutagenicity of the particulate phase of the resulting cigarette smoke was assayed in the Salmonella plate incorporation (Ames) assay with tester strains TA98, TA100, TA102, TA1535 and TA1537. The cytotoxicity of the gas/vapor phase and the particulate phase was determined in the neutral red uptake assay with mouse embryo BALB/c 3T3 cells. Within the sensitivity and specificity of the test systems, the in vitro mutagenicity and cytotoxicity of the cigarette smoke were not increased by the addition of the ingredients.

3T3 Cells↗

PubMed1981

[Dynamics of the secretory and excretory functions of the stomach and activity of the kallikrein-kinin system following removal of the parotid glands].

After removal of parotid glands in 104 white rats, the gastric acidity increased and excretion of neutral red occurred. Simultaneously the kallikrein activity and kallikreinogen concentration as well as concentration of kininogen became enhanced whereas the bradykinin of plasma decreased. The data suggest that the mechanism of activation of the stomach secretory and excretory functions after removal of parotid glands involves inactivation of the blood kallikrein system.

Animals↗

PubMed1977

"Dye/light" Dye-induced photosensitization of herpes virus. A clinical trial on humans.

A series of 21 patients with herpetic corneal ulcers was treated by photodynamic inactivation (using a single application of neutral red 1 per cent solution followed by a 15-min exposure to a 40 watt light of 440--550 nm), carbolization, or idoxuridine (IDU) ointment. No significant difference was found in the number that healed, the mean healing time, or the number of recurrences after any of the three treatments.

Clinical Trials as Topic↗

PubMed2006

Bioavailability and toxicity of pentachlorophenol in contaminated soil evaluated on coelomocytes of Eisenia andrei (Annelida: Lumbricidae).

Pentachlorophenol (PCP) is widely distributed and highly persistent in soil, and represents a threat to the health of ecosystems. The present study aimed to assess the toxicity and bioavailability of PCP in soils as a function of different aging periods with the attempt to select a good toxicological assay for Eisenia andrei Bouché (Annelida: Lumbricidae). The experiments were performed on soil contaminated with PCP at 15 and 150ppm. After different aging periods (20, 60 and 120 days from spiking), bioavailability and toxicity were evaluated on E. andrei kept for 7 and 14 days in treated soils. The actual bioavailability decreased in relation to the aging for both PCP concentrations. No membrane damage was observed on coelomocytes collected by ethanol extrusion. Modifications in distribution of coelomocyte subpopulations were detected by flow cytometry on samples aged for 60 and 120 days at 150ppm PCP contamination. The reduction of lysosomal membrane stability, measured by neutral red retention time, was observed in all treatments. Worm mortality increased with aging in soils spiked with 150ppm of PCP. In conclusion, aging did not seem to reduce PCP cytotoxicity. This is the first report on in vivo toxicity of PCP evaluated on coelomocytes of E. andrei using different assays.

Animals↗

PubMed1985

Identification and purification of NK cells with lysosomotropic vital stains: correlation of lysosome content with NK activity.

The lysosome content of lymphocytes has been analyzed with lysosomotropic vital stains and the fluorescence-activated cell sorter (FACS). Large granular lymphocytes (LGL), which account for virtually all natural killing activity in peripheral blood, are quantitatively different from small lymphocytes (SL) in this respect. LGL obtained by Percoll gradient density centrifugation accumulate more of the lysosomotropic vital dyes than SL do, staining with either neutral red or mepacrine (quinacrine). Furthermore among the LGL-rich, low density lymphocyte population highly, granulated cells can be separated from less granulated ones by mepacrine staining and FACS. Thus, separated highly granulated LGL express very high natural killing, whereas the less granulated low density large lymphocytes do not kill.

Cell Separation↗

PubMed1978

Simultaneous monitoring by optical techniques of respiratory chain and intracellular pH in toad ventricle strip.

Intracellular pH and oxidative metabolism can be measured in toad ventricle strips simultaneously by the use of the pH indicator dye, neutral red, and a rapid scanning spectrophotometer. The effects of hypoxia and acidification on mechanical function are approximately additive. The decrease in tension due to slight acidification is probably through an effect on the portion of the twitch tension supported by anaerobic metabolism.

Anaerobiosis↗

PubMed1982

A rapid method for staining inclusions of Chlamydia psittaci and Chlamydia trachomatis.

A new staining method was developed for the detection of inclusions of Chlamydia psittaci and Chlamydia trachomatis inclusions in cell cultures. Using a combination of methyl green and neutral red stains and washing at pH 5.0, inclusions were stained red while cell cytoplasm was pale pink and cell nuclei were pale green. The method was significantly better than Giemsa staining and comparable to immunofluorescence for detecting C psittaci inclusions. Its sensitivity for detection C trachomatis inclusions by dark field microscopy was similar to that of Giemsa staining.

Animals↗

PubMed1997

Evaluation of the toxicity of Uncaria tomentosa by bioassays in vitro.

Aqueous extracts of Uncaria tomentosa (Willdenow ex Roemer and Schultes) DC. (Rubiaceae) ('Uña de gato'), were analyzed for the presence of toxic compounds in Chinese hamster ovary cells (CHO) and bacterial cells (Photobacterium phosphoreum). Toxicity was evaluated by four systems: Neutral red assay (NR), total protein content (KB), tetrazolium assay (MTT) and Microtox test. The extracts of U. tomentosa did not show toxicity in vitro at the concentrations tested. Testing in vitro could be a valuable tool for evaluating toxicity of medicinal plants.

Analysis of Variance↗

PubMedDate not supplied

An evaluation of three in vitro cytotoxicity assays.

A number of methods for determining the general toxic effects of test chemicals on cells in culture are now at the validation stage. Three such methods based on measurement of total cell protein or neutral red uptake and on the detection of morphological effects have been compared. The cell line used was 3T3-L1 (a continuous fibroblast cell line derived from mouse embryos). The results obtained in a blind trial with 30 coded chemicals indicated a close correlation between the relative cytotoxicities of chemicals tested by all three methods.

Cell Survival↗

PubMed1999

Chlorate as an inorganic disinfection by product in swimming pools.

Chlorate and chlorite concentrations were determined in water samples taken from 33 swimming pools. In the pools under investigation, disinfection of the water is carried out either by gaseous chlorine (n = 14) or hypochlorite solution in conjunction with flocculation and sand filtration. A number of the pools also use ozone treatment to augment the disinfection process. Chlorite was not detectable in any of the samples (detection limit 1 mg/l). High concentrations of chlorate were detected in samples from a number of the pools; in one case as high as 40 mg/l. Higher chlorate concentrations were found to be associated with those pools using hypochlorite solution as a disinfecting agent. In contrast, relatively low chlorate concentrations were found in pools treated with gaseous chlorine. In order to elucidate any relationship between the chlorate content of pool water and that of the respective hypochlorite stock solution, chlorate and bromate concentrations were determined in the hypochlorite stock solutions of nine pools. Bromate concentration in the stock solutions were not found to exceed 1.2 g/l, chlorate was measured in concentrations of up to 44.5 g/l. The additional use of ozone as part of the water purification process appears to have no significant influence on chlorate concentration. Chlorate has no bactericidal properties and does not interfere with the measurement of certain parameters relevant to hygiene in swimming pools such as free and combined chlorine, pH or redox potential. At present, the effects of high chlorate concentrations in swimming pool water are unclear. Our initial investigations indicate that chlorate has no cytotoxic (Neutral-Red assay) or irritating properties (HET-CAM assay). However, both chlorate and chlorite are known to interfere with the haematopoetic system. In Germany, the MCL for chlorite in drinking water is 0.2 mg/l. It is therefore strongly recommended that measures should be taken to reduce chlorate concentrations in swimming pool water.

Animals↗

PubMed1976

[Morphofunctional express-diagnosis of liver conditons].

Methods of simultaneous detection of the vitally-essential indices of the hepatic tissue status by injecting neutral red vital dye into the portal vein are described. The liver tissue morphological status, the hepatic cell absorbing capacity, the blood microcirculatory efficacy and the pH shift were assessed on a histological preparation from the liver slice. The dye was extracted from the other portion of the same liver slice and the total amount of the dye absorbed was estimated.

Animals↗

PubMed1998

Change in colony morphology influences the virulence as well as the biochemical properties of the Mycobacterium avium complex.

Factors that influence colony morphology are of crucial importance for drug development as well as for understanding the virulence of Mycobacterium avium complex (MAC) strains. The MAC 101 strain used in the present study grows as smooth transparent (SmT) colonies that tend to become opaque and pigmented when incubated for long periods of time. However, when MAC was passaged in animals, two types of colonies were recovered. The new rough transparent (RgT) colony morphology appeared more flat and transparent, having a central spot, irregular edges at times, and a dry, granular appearance like that of the rough mutants. In animal studies, the RgT bacilli multiplied at a much faster rate than that of the SmT bacilli, causing 60-80% mortality compared with the 10% mortality observed in mice infected with SmT. In vitro studies indicated that the SmT MAC did not grow and multiply as well in resident peritoneal macrophages as the RgT MAC did. The two morphotypes did not differ in their growth ratesin vitro but the RgT MAC failed to reduce dimethylthiazol-diphenyltetrazolium bromide (MTT), alamar blue and neutral red, suggesting that there might be significant changes in the cell wall or elsewhere causing changes in cellular permeability. These two morphotypes could serve as models for studying the biochemical markers or the identification of factors responsible for the virulence of the MAC.

Animals↗

PubMed1995

Development in one dimension: the rapid differentiation of Dictyostelium discoideum in glass capillaries.

When Dictyostelium discoideum cells are drawn into a fine glass capillary, they rapidly begin the first steps toward the formation of prestalk and prespore zones. Some of the events occur within a minute or two, whereas others follow later. The cells in the front segment are actively motile and those in the hind segment are passive. The volumes of the segments are proportional for different-sized cell masses, and those proportions are the same as those found in normal slugs. When the cells are stained with the vital dye neutral red, the anterior zone becomes darker simultaneously with the formation of the division line. Green fluorescent protein expressed from a stalk-specific promoter is synthesized mostly in the anterior end. Later, this capillary prestalk zone shows a sharp increase in alkaline phosphatase activity, which is known to be characteristic of prestalk cells.

Alkaline Phosphatase↗

PubMed1983

Toxic effects of some antibiotics on rabbit kidney cells.

The cellular toxicity of some antibiotics (cephaloridine, cefazolin, cefoperazone and gentamicin) was studied in vitro using rabbit kidney cell cultures. In order to evaluate the importance of the different factors causing toxic effects, the authors studied the correlations between toxicity and antibiotic concentration and between toxicity and period of contact. The toxic effects were measured by LDH release and by neutral red uptake inhibition. In both tests, gentamicin clearly showed curvilinear dose-toxicity and time-toxicity correlation but the cephalosporins did not do so in either test. However the antibiotics tested all showed various degrees of toxicity, suggesting that their specific toxic activity on renal cells was greater than that of aspecific variables linked to pharmacokinetic behaviour.

Animals↗

PubMed2006

Cytotoxicity study of plasma-sprayed hydroxyapatite coating on high nitrogen austenitic stainless steels.

Stainless steel has been frequently used for temporary implants but its use as permanent implants is restricted due to its low pitting corrosion resistance. Nitrogen additions to these steels improve both mechanical properties and corrosion resistance, particularly the pitting and crevice corrosion resistance. Many reports concerning allergic reactions caused by nickel led to the development of nickel free stainless steel; it has excellent mechanical properties and very high corrosion resistance. On the other hand, stainless steels are biologically tolerated and no chemical bonds are formed between the steel and the bone tissue. Hydroxyapatite coatings deposited on stainless steels improve osseointegration, due their capacity to form chemical bonds (bioactive fixation) with the bone tissue. In this work hydroxyapatite coatings were plasma-sprayed on three austenitic stainless steels: ASTM-F138, ASTM-F1586 and the nickel-free Böhler-P558. The coatings were analyzed by SEM and XDR. The cytotoxicity of the coatings/steels was studied using the neutral red uptake method by quantitative evaluation of cell viability. The three uncoated stainless steels and the hydroxyapatite coated Böhler-P558 did not have any toxic effect on the cell culture. The hydroxyapatite coated ASTM-F138 and ASTM-F1586 stainless steels presented cytotoxicity indexes (IC50%) lower than 50% and high nickel contents in the extracts.

Animals↗

PubMed1991

In vitro metabolism and toxicity assessment of N-methylcarbazole in primary cultured rat hepatocytes.

N-Methycarbazole (NMC), a carcinogen and mutagen in tobacco smoke, was converted to two major metabolites by primary cultured rat hepatocytes as measured by high performance liquid chromatography (HPLC): N-hydroxymethylcarbazole (NHMC) and carbazole. These two metabolites had comparable retention times and identical ultraviolet spectra as those of reference standards. Identical retention times and mass spectra were also observed as detected by gas chromatography-mass spectroscopy (GC-MS) for NHMC and its reference standard. The toxicities of NMC and its two metabolites were assessed by lactate dehydrogenase (LDH) leakage and neutral red (NR) uptake. The rank order of cytotoxicity of NMC and its metabolites was found to be: NHMC greater than NMC greater than carbazole. Thus, we conclude that the hydroxylation of NMC to NHMC may represent a toxification step, while the further dealkylation to carbazole is most likely a detoxication process.

Animals↗

PubMedDate not supplied

Cultured porcine urinary bladder epithelial cells as a screening model for genotoxic effects of aromatic amines: characterisation and application of the cell culture model.

Isolated epithelial cells from porcine urinary bladders were maintained in dividing long-term monolayer cultures, and were used as a model system for the urinary bladder in toxicological studies in vitro. To examine the state of differentiation during the culture period, the culture system was characterised morphologically by light and transmission electron microscopy and by immune fluorescence labelling with antibodies against cytokeratins 7,13 and pan. The cultured cells were identified as urothelial epithelium by their polarised structure, and by their expression of several uroepithelial specific morphological features, such as fusiform vesicles, tight junctions and an asymmetric apical cell membrane. Additionally, the cells were labelled with anti-cytokeratin 7,13 and pan antibodies, and negatively with anti-vimentin antibodies. The maintenance of suitable culture conditions was shown by the stable enzyme activities of (gamma-glutamyltranspeptidase, alkaline phosphatase and acid phosphatase over a culture period of 4 weeks. A good viability of the cultured cells under the chosen culture conditions was shown by the presence of low amounts of lactate dehydrogenase (< of = 5%) in the culture medium. The activities of the chosen marker enzymes for cell differentiation (gamma-glutamyltranspeptidase), lysosomes (acid phosphatase) and luminal membranes (alkaline phosphatase) were relatively stable over the observed culture period. Enzyme activities involved in metabolism of xenobiotics were determined, to define the ability for metabolism in cultured cells compared with bladder tissue in situ. Several constitutive phase I and II enzyme activities were found to be stable during the culture period, indicating that the cultured cells should be able to metabolise xenobiotics in a comparable manner to the urothelium in vivo. The cytotoxic effects of xenobiotics were investigated and IC50 values were determined by means of lactate dehydrogenase leakage and inhibition of neutral red uptake. The induction of sister chromatid exchanges was used as a parameter for the genotoxic effects of several xenobiotics. This cell culture system was found to be a very good screening system for the testing of substances that affect the bladder, especially aromatic amines.

Amines↗

PubMedDate not supplied

Toxicological characterisation of sludge from sewage treatment plants using toxicity identification evaluation protocols based on in vitro toxicity tests.

The ecotoxicological characterisation of complex mixtures, such as sludge from sewage treatment plants, is complex. Toxicity identification evaluation (TIE) protocols, developed by the United States Environmental Protection Agency (US EPA); to identify toxic pollutants in complex effluents, are useful tools in this context; to solve the difficulties in assessing unknown organic pollutants by analytical methods, the usefulness of bioassays to detect the relevant (toxic) organic compounds present in complex samples, and the possibilities of in vitro cytotoxicity tests as screening tools, offers a profitable combination. The sludge obtained from a sewage treatment plant was extracted by acetonitrile using a microwave extractor and fractionated in an HPLC system. The toxicity of every fraction was assayed using a RTG-2 cytotoxicty test, based on the fibroblastic RTG-2 fish cell line (ATCC, CCL N. 55). At exponential growth, three endpoints, beta-galactosidase activity, culture viability assayed by the neutral red assay (NR) and inhibition of growth rate using the FRAME KB protein assay (KBP), were used. By plotting the toxicity of each fraction vs elution time, the corresponding "toxicograms" were built. The UV and fluorescence chromatograms are compared to the three toxicograms (one for each toxicity endpoint).

Animal Testing Alternatives↗