Cytotoxicity assays to evaluate tannery effluents treated by photoelectrooxidation.

The advanced oxidation process (AOP) is used to increase the treatment efficiency of effluents however, it is necessary to compare the toxicity of treated and untreated effluents to evaluate if the decontamination process does not cause any biological harm. Cultured cells have been previously used to assess the genotoxic and cytotoxic potential of various compounds. Hence, the aim of this work was to assess the applicability of cytotoxicity assays to evaluate the toxicity related to the AOP treatment. Samples of an industrial effluent were collected after their treatment by a conventional method. Cytotoxicity of standard and AOP treated effluents was assessed in CRIB and HEp-2 cell line using the MTT and neutral red assays. We observed decrease at cell viability in the both assays (50% MTT and 13% NRU) when cells were exposed to the AOP treatment in the highest concentration. Thus, cytotoxic assays in cultured cells can be explored as an useful method to evaluate toxicity as well as to optimize effluents treatment process.

Cytotoxicity assays to evaluate tannery effluents treated by photoelectrooxidation / Ensaios de citotoxicidade na avaliação de efluentes de curtume tratados por fotoeletrooxidação

The advanced oxidation process (AOP) is used to increase the treatment efficiency of effluents however, it is necessary to compare the toxicity of treated and untreated effluents to evaluate if the decontamination process does not cause any biological harm. Cultured cells have been previously used to assess the genotoxic and cytotoxic potential of various compounds. Hence, the aim of this work was to assess the applicability of cytotoxicity assays to evaluate the toxicity related to the AOP treatment. Samples of an industrial effluent were collected after their treatment by a conventional method. Cytotoxicity of standard and AOP treated effluents was assessed in CRIB and HEp-2 cell line using the MTT and neutral red assays. We observed decrease at cell viability in the both assays (50% MTT and 13% NRU) when cells were exposed to the AOP treatment in the highest concentration. Thus, cytotoxic assays in cultured cells can be explored as an useful method to evaluate toxicity as well as to optimize effluents treatment process.

Resumo O processo de oxidação avançada (POA) tem sido usado para aumentar a eficiência do tratamento de efluentes; no entanto, é necessário comparar a toxicidade de efluentes tratados e não tratados para avaliar se o processo de descontaminação não é capaz de causar algum risco biológico. Cultivos celulares têm sido utilizados para avaliar o potencial genotóxico e citotóxico de vários compostos. Assim, o objetivo deste trabalho foi avaliar a aplicabilidade de ensaios de citotoxicidade para avaliar a toxicidade relacionada ao tratamento com POA. As amostras de um efluente industrial foram recolhidas após o tratamento por um método convencional. A citotoxicidade dos efluentes padrão e tratado com POA foi avaliada nas linhagens celulares CRIB e HEp-2 usando os ensaios do MTT e do vermelho neutro. Observou-se diminuição da viabilidade celular em ambos os ensaios (50% MTT e 13% VN) quando as células foram expostas à concentração mais elevada do efluente tratado com POA. Assim, os ensaios de citotoxicidade em cultivos celulares podem ser explorados como um método útil para avaliar a toxicidade, bem como para otimizar os processos de tratamento de efluentes.

Cytotoxicity assays to evaluate tannery effluents treated by photoelectrooxidation

The advanced oxidation process (AOP) is used to increase the treatment efficiency of effluents however, it is necessary to compare the toxicity of treated and untreated effluents to evaluate if the decontamination process does not cause any biological harm. Cultured cells have been previously used to assess the genotoxic and cytotoxic potential of various compounds. Hence, the aim of this work was to assess the applicability of cytotoxicity assays to evaluate the toxicity related to the AOP treatment. Samples of an industrial effluent were collected after their treatment by a conventional method. Cytotoxicity of standard and AOP treated effluents was assessed in CRIB and HEp-2 cell line using the MTT and neutral red assays. We observed decrease at cell viability in the both assays (50% MTT and 13% NRU) when cells were exposed to the AOP treatment in the highest concentration. Thus, cytotoxic assays in cultured cells can be explored as an useful method to evaluate toxicity as well as to optimize effluents treatment process.

Resumo O processo de oxidação avançada (POA) tem sido usado para aumentar a eficiência do tratamento de efluentes; no entanto, é necessário comparar a toxicidade de efluentes tratados e não tratados para avaliar se o processo de descontaminação não é capaz de causar algum risco biológico. Cultivos celulares têm sido utilizados para avaliar o potencial genotóxico e citotóxico de vários compostos. Assim, o objetivo deste trabalho foi avaliar a aplicabilidade de ensaios de citotoxicidade para avaliar a toxicidade relacionada ao tratamento com POA. As amostras de um efluente industrial foram recolhidas após o tratamento por um método convencional. A citotoxicidade dos efluentes padrão e tratado com POA foi avaliada nas linhagens celulares CRIB e HEp-2 usando os ensaios do MTT e do vermelho neutro. Observou-se diminuição da viabilidade celular em ambos os ensaios (50% MTT e 13% VN) quando as células foram expostas à concentração mais elevada do efluente tratado com POA. Assim, os ensaios de citotoxicidade em cultivos celulares podem ser explorados como um método útil para avaliar a toxicidade, bem como para otimizar os processos de tratamento de efluentes.

Evaluation of genotoxicity and cytotoxicity of water samples from the Sinos River Basin, southern Brazil.

Some water bodies in the Sinos River Basin (SRB) have been suffering the effects of pollution by residential, industrial and agroindustrial wastewater. The presence of cytotoxic and genotoxic compounds could compromise the water quality and the balance of these ecosystems. In this context, the research aimed to evaluate the genotoxicity and cytotoxicity of the water at four sites along the SRB (in the cities of Santo Antônio da Patrulha, Parobé, Campo Bom and Esteio), using bioassays in fish and cell culture. Samples of surface water were collected and evaluated in vitro using the Astyanax jacuhiensis fish species (micronucleus test and comet assay) and the Vero lineage of cells (comet assay and cytotoxicity tests, neutral red - NR and tetrazolium MTT). The micronucleus test in fish showed no significant differences between the sampling sites, and neither did the comet assay and the MTT and NR tests in Vero cells. The comet assay showed an increase in genetic damage in the fish exposed to water samples collected in the middle and lower sections of the basin (Parobé, Campo Bom and Esteio) when compared to the upper section of the basin (Santo Antônio da Patrulha). The results indicate contamination by genotoxic substances starting in the middle section of the SRB.

Cytotoxicity assays as tools to assess water quality in the Sinos River basin.

Cytotoxicity assays using cell cultures may be an alternative to assess biological toxicity of surface waters and may help to improve the control of water quality. This study compared two methods to prepare culture media for the exposure of Hep-2 cells to water samples collected from the Rolante River, an important affluent of the Sinos River. The toxicity was evaluated using the MTT and neutral red assays. Two methods were used to prepare culture media. In method 1, the sample was diluted at 1:1, 1:10, 1:100, 1:1000, 1:10.000 (v/v, sample/medium) in a standard culture medium; in method 2, water samples were used as the solvent for the culture medium, which was prepared at concentrations of 100, 80, 60, 40 and 20%. Semi-confluent cultures were then exposed to the media test for 24 hours, and cytotoxicity was determined immediately using the MTT and NR assays. Mitochondrial activity (MTT) was significantly lower at all concentrations in both methods, except at 1:1000 in method 1. However, the lysosome viability (NR) results revealed cytotoxicity only in the 1:1 sample of method 1. Both culture preparation methods were efficient and sensitive to the MTT assay, but method 2 seemed to be more adequate for the NR assay. The Rolante River has cytotoxic contaminants to Hep-2 cells, which may be one of the explanations for the poor water quality of the Sinos River basin.

Synthesis and transport of different sphingomyelin species in rat Sertoli cells.

Cellular phospholipids of Sertoli cells from immature rats were labeled with [14C]-choline. Two sphingomyelin bands (SM1 and SM2) were identified by TLC. The incorporation of [14C]-choline over a 45 h period of incubation demonstrated that there are differences in labeling kinetics between SM1 and SM2. The subcellular location of SM1 and SM2 was investigated by accessibility to bacterial sphingomyelinase. The results showed the existence of two SM pools in Sertoli cells, but an equal cellular distribution of SM1 and SM2. SM2 is characterized by a relatively high content of unsaturated fatty acids. The inhibition of vesicular flow by monensin determines a decrease of about 60-70% in incorporation into SM1 and SM2, suggesting the existence of at least two sites of sphingomyelin synthesis. Pulse-chase and time-course experiments indicated a phosphatidylcholine --> SM precursor product relationship and differences in kinetic properties between SM1 and SM2. Resynthesis experiments showed that monensin had only a partial inhibitory effect on SM1 resynthesis, and a second sphingomyelinase treatment demonstrated that the resynthesized fraction reached the outer leaflet of the plasma membrane. The 60-70% inhibition of SM synthesis by monensin showed that the trans-Golgi cisternae and the trans-Golgi network are the most likely sites of bulk SM synthesis, and that about 15% of SM was synthesized in the cis/medial Golgi apparatus. Additionally the results indicated that plasma membrane SM synthase activity could be the site of about 15% of SM synthesis in Sertoli cells.

De novo synthesis and recycling pathways of sphingomyelin in rat Sertoli cells.

Sertoli cells from 19-day-old rats have two molecular species of sphingomyelin (SM1 and SM2) with different kinetic characteristics and fatty acid composition. Here, we have studied the incorporation of [14C]-choline and [14C]-palmitic acid into SM in presence or absence of fumonisin B1, an inhibitor of ceramide synthesis, and beta-chloroalanine, an inhibitor of sphinganine synthesis. The contributions of de novo synthesis and recycling pathways were estimated by analysis of the inhibition caused by these drugs. SM1 was synthesized more by sphingosine recycling, and SM2 was synthesized principally by ceramide recycling than SM1. De novo synthesis seems to be important for the two SM types, but our results showed that this pathway is more extensively utilized by SM2. In conclusion, using Sertoli cell cultures, we have shown for the first time that in the same cell different molecular species of SM are synthesized by different pathways.