Cryopreservation and storage of mussel (Mytilus spp.) haemocytes for latent analysis by the Comet assay

Kwok, Anthony; Lyons, Brett P.; Hodges, Nikolas J.; Bean, Tim P.

doi:10.1016/j.mrgentox.2012.09.010

Cited by 16 publications

(8 citation statements)

References 35 publications

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“…BaP) was consistent with previous studies (e.g. Banni et al 2010;Di et al 2011;Kwok et al 2013;Mitchelmore et al 1998). The degree of damage observed in mussel haemocytes was a little less than that observed by Lacaze et al (2014) for in vitro exposures to N-Ad in fish hepatocytes and was not significantly different to controls at the 5 % probability level but was at the more precautionary 10 % level (Fig.…”

Section: Resultssupporting

confidence: 93%

Diamondoid naphthenic acids cause in vivo genetic damage in gills and haemocytes of marine mussels

Dissanayake

Scarlett

Jha

2016

Environ Sci Pollut Res

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Diamondoids are polycyclic saturated hydrocarbons that possess a cage-like carbon skeleton approaching that of diamond. These 'nano-diamonds' are used in a range of industries including nanotechnologies and biomedicine. Diamondoids were thought to be highly resistant to degradation, but their presumed degradation acid products have now been found in oil sands process-affected waters (OSPW) and numerous crude oils. Recently, a diamondoid-related structure, 3-noradamantane carboxylic acid, was reported to cause genetic damage in trout hepatocytes under in vitro conditions. This particular compound has never been reported in the environment but led us to hypothesise that other more environmentally relevant diamondoid acids could also be genotoxic. We carried out in vivo exposures (3 days, semi-static) of marine mussels to two environmentally relevant diamondoid acids, 1-adamantane carboxylic acid and 3,5-dimethyladamantane carboxylic acid plus 3-noradamantane carboxylic acid with genotoxic damage assessed using the Comet assay. An initial screening test confirmed that these acids displayed varying degrees of genotoxicity to haemocytes (increased DNA damage above that of controls) when exposed in vivo to a concentration of 30 μmol L(-1). In a further test focused on 1-adamantane carboxylic acid with varying concentrations (0.6, 6 and 30 μmol L(-1)), significant (P < 0.05%) DNA damage was observed in different target cells (viz. gills and haemocytes) at 0.6 μmol L(-1). Such a level of induced genetic damage was similar to that observed following exposure to a known genotoxin, benzo(a)pyrene (exposure concentration, 0.8 μmol L(-1)). These findings may have implications for a range of worldwide industries including oil extraction, nanotechnology and biomedicine.

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Section: Resultssupporting

confidence: 93%

Diamondoid naphthenic acids cause in vivo genetic damage in gills and haemocytes of marine mussels

Dissanayake

Scarlett

Jha

2016

Environ Sci Pollut Res

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“…The molluscan digestive gland, the analogous of the vertebrate liver and therefore of high relevance in toxicological studies, was shown to yield levels of single strand breakage likely too high (from autolytic processes) for a valid application of the Comet Assay without proper cell sorting and viability check (refer to Raimundo et al, 2010 , in a study with the cephalopod Octopus vulgaris and Hartl et al, 2004 with the clam Ruditapes philippinarum ). Recent advances have also shown the feasibility of obtaining adequate cultures of molluscan cells for in vitro studies using the Comet Assay (Michel and Vincent-Hubert, 2012 ) and even the possibility to cryopreserve mussel haemocytes (Kwok et al, 2013 ). Altogether, these advances certainly contribute to standardize the Comet Assay in biomonitoring and genotoxicity testing with bivalves and other molluscs.…”

Section: A Methodological Overviewmentioning

confidence: 99%

The Comet Assay and its applications in the field of ecotoxicology: a mature tool that continues to expand its perspectives

et al. 2015

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Since Singh and colleagues, in 1988, launched to the scientific community the alkaline Single Cell Gel Electrophoresis (SCGE) protocol, or Comet Assay, its uses and applications has been increasing. The thematic areas of its current employment in the evaluation of genetic toxicity are vast, either in vitro or in vivo, both in the laboratory and in the environment, terrestrial or aquatic. It has been applied to a wide range of experimental models: bacteria, fungi, cells culture, arthropods, fishes, amphibians, reptiles, mammals, and humans. This document is intended to be a comprehensive review of what has been published to date on the field of ecotoxicology, aiming at the following main aspects: (i) to show the most relevant experimental models used as bioindicators both in the laboratory and in the field. Fishes are clearly the most adopted group, reflecting their popularity as bioindicator models, as well as a primary concern over the aquatic environment health. Amphibians are among the most sensitive organisms to environmental changes, mainly due to an early aquatic-dependent development stage and a highly permeable skin. Moreover, in the terrestrial approach, earthworms, plants or mammalians are excellent organisms to be used as experimental models for genotoxic evaluation of pollutants, complex mix of pollutants and chemicals, in both laboratory and natural environment. (ii) To review the development and modifications of the protocols used and the cell types (or tissues) used. The most recent developments concern the adoption of the enzyme linked assay (digestion with lesion-specific repair endonucleases) and prediction of the ability to repair of oxidative DNA damage, which is becoming a widespread approach, albeit challenging. For practical/technical reasons, blood is the most common choice but tissues/cells like gills, sperm cells, early larval stages, coelomocytes, liver or kidney have been also used. (iii) To highlight correlations with other biomarkers. (iv) To build a constructive criticism and summarize the needs for protocol improvements for future test applications within the field of ecotoxicology. The Comet Assay is still developing and its potential is yet underexploited in experimental models, mesocosmos or natural ecosystems.

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“…In partic ular, mollusk cells have been used to study biomineral ization (Sud et al, 2001;Barik et al, 2004;Endoh, Hasegawa, 2006;Gong et al, 2008aGong et al, , 2008bAuzoux Bordenave, Domart Coulon, 2010), embryonic cell differentiation (Odintsova et al, 2010), and infection by parasitic organisms (Villena, 2003) and to test the toxicity of the environment (Domart Coulon et al, 2000;Le Pennec, Le Pennec, 2001, 2003Pennec et al, 2002;Latire et al, 2012;Kwok et al, 2013).…”

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confidence: 99%

Culture of mussel (Mytiuls edulis L.) mantle cells

Daugavet

Блинова

2015

Cell Tiss. Biol.

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To date, cell lines derived from marine invertebrates have not been available. Hence primary cell cultures serve as model systems for various experiments. In the present study, we established a primary culture of mussel Mytilus edulis L. mantle cells. Cells were isolated by means of explant culture and enzymatic disso ciation of mantle tissue. They maintained viability up to 22 months regardless of the initiation method. In course of culturing, cells that were transferred onto new plates successfully attached to a new surface. The physiological activity of cultured cells was also confirmed by formation of crystals, which appeared after 4-6 months. After a continuous time of culturing, mantle cells may be successfully cryopreserved. We used 5% DMSO with postfreezing survival up to 50%. These results demonstrate that M. edulis mantle cells can main tain viability and physiological activity for an exceptionally long time and can be cryopreserved for further examination.

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Cryopreservation and storage of mussel (Mytilus spp.) haemocytes for latent analysis by the Comet assay

Cited by 16 publications

References 35 publications

Diamondoid naphthenic acids cause in vivo genetic damage in gills and haemocytes of marine mussels

Diamondoid naphthenic acids cause in vivo genetic damage in gills and haemocytes of marine mussels

The Comet Assay and its applications in the field of ecotoxicology: a mature tool that continues to expand its perspectives

Culture of mussel (Mytiuls edulis L.) mantle cells

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