Advanced

Comparative sensitivity of the harbor seal (Phoca vitulina) 11B7501 cell line to natural and anthropogenic environmental contaminants

Kleinert, Christine (2012) BIOP14 20112
Degree Projects in Biology
Abstract
Popular science summary:

Effects of xenobiotics on the harbor seal’s immune system

Naturally synthesized toxins and also synthetic substances have both been suspected to cause mass mortality events in marine mammals. Top-predators at the end of the food chain, like harbor seals (Phoca vitulina), accumulate particularly high concentrations of contaminants. Certain substances have been proven to have a negative influence on various parameters of the immune system. To assess the immunotoxic potential of selected compounds in harbor seals, a 11B7501 B lymphoma cell line was used.

In this study, the adverse effect of nanoparticles (zinc oxide, ZnO NP) and phycotoxins (decarbamoyl-saxitoxin, gonyautoxin-2 and -3, domoic acid) on the... (More)
Popular science summary:

Effects of xenobiotics on the harbor seal’s immune system

Naturally synthesized toxins and also synthetic substances have both been suspected to cause mass mortality events in marine mammals. Top-predators at the end of the food chain, like harbor seals (Phoca vitulina), accumulate particularly high concentrations of contaminants. Certain substances have been proven to have a negative influence on various parameters of the immune system. To assess the immunotoxic potential of selected compounds in harbor seals, a 11B7501 B lymphoma cell line was used.

In this study, the adverse effect of nanoparticles (zinc oxide, ZnO NP) and phycotoxins (decarbamoyl-saxitoxin, gonyautoxin-2 and -3, domoic acid) on the immune system was assessed. Therefore, the viability of cells, phagocytosis, lymphocyte proliferation, apoptosis and the cell cycle was analysed.

Material and Methods
The main method used in this study, was flow cytometry. The light scatter of cells in a suspension is read and the information about size (FSC) and complexity (SSC) is collected in a dot plot. Cell populations can be electronically gated and cellular activity is measured. For cell cycle, apoptosis and viability measurement propidium iodide was used; an agent that intercalates with the DNA and RNA of dead cells. The phagocytosis was detected with fluorescent latex beads that cells with phagocytic activity were able to engulf. The lymphocyte proliferation was measured with liquid scintillation – a method where the radioactive base methyl-3H-thymidine is incorporated into newly synthesized DNA.

Results and Discussion
An immunotoxic effect on phagocytosis was seen above 12,5 μg/ml for ZnO hydrate. Viability was affected for both ZnO hydrate and ZnO NP(UNF). On lymphocyte proliferation and the cell cycle and apoptosis an effect was found for the same compounds above 25 μg/ml and 12,5 μg/ml, respectively. Since ZnO NP(UNF) and ZnO hydrate (microscale) showed similar effects, the toxicity of these compounds was not accentuated by the nanoparticle scale. ZnO NP(F) did not show adverse effects, and it has yet to be verified if the filtration method did not possibly influence the concentration of the solution.

None of the tested phycotoxins showed an adverse effect on the immune cells in any of the assays. However, in previous studies the neurotoxic potential of phycotoxins was already proven. Domoic acid, a glutamate receptor antagonist, caused lesions in the brain areas where glutamergic pathways were concentrated, and saxitoxins have been shown to cause respiratory paralysis due to neuronal cell death that is induced by high calcium levels in the cells. In mass mortality events of various marine mammal species, both compound classes have been found in organs and serum.

Advisor: Berglund, Olof
Supervisor: Fournier, Michel (INRS-Institut Armand-Frappier)
Master´s Degree Project 60 credits in Ecotoxicology from September 2011 – May 2012
Department of Biology, Lund University (Less)
Please use this url to cite or link to this publication:
author
Kleinert, Christine
supervisor
organization
course
BIOP14 20112
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
3633644
date added to LUP
2013-04-16 16:20:43
date last changed
2013-04-16 16:20:43
@misc{3633644,
  abstract     = {Popular science summary:

Effects of xenobiotics on the harbor seal’s immune system

Naturally synthesized toxins and also synthetic substances have both been suspected to cause mass mortality events in marine mammals. Top-predators at the end of the food chain, like harbor seals (Phoca vitulina), accumulate particularly high concentrations of contaminants. Certain substances have been proven to have a negative influence on various parameters of the immune system. To assess the immunotoxic potential of selected compounds in harbor seals, a 11B7501 B lymphoma cell line was used.

In this study, the adverse effect of nanoparticles (zinc oxide, ZnO NP) and phycotoxins (decarbamoyl-saxitoxin, gonyautoxin-2 and -3, domoic acid) on the immune system was assessed. Therefore, the viability of cells, phagocytosis, lymphocyte proliferation, apoptosis and the cell cycle was analysed. 

Material and Methods
The main method used in this study, was flow cytometry. The light scatter of cells in a suspension is read and the information about size (FSC) and complexity (SSC) is collected in a dot plot. Cell populations can be electronically gated and cellular activity is measured. For cell cycle, apoptosis and viability measurement propidium iodide was used; an agent that intercalates with the DNA and RNA of dead cells. The phagocytosis was detected with fluorescent latex beads that cells with phagocytic activity were able to engulf. The lymphocyte proliferation was measured with liquid scintillation – a method where the radioactive base methyl-3H-thymidine is incorporated into newly synthesized DNA.

Results and Discussion
An immunotoxic effect on phagocytosis was seen above 12,5 μg/ml for ZnO hydrate. Viability was affected for both ZnO hydrate and ZnO NP(UNF). On lymphocyte proliferation and the cell cycle and apoptosis an effect was found for the same compounds above 25 μg/ml and 12,5 μg/ml, respectively. Since ZnO NP(UNF) and ZnO hydrate (microscale) showed similar effects, the toxicity of these compounds was not accentuated by the nanoparticle scale. ZnO NP(F) did not show adverse effects, and it has yet to be verified if the filtration method did not possibly influence the concentration of the solution.

None of the tested phycotoxins showed an adverse effect on the immune cells in any of the assays. However, in previous studies the neurotoxic potential of phycotoxins was already proven. Domoic acid, a glutamate receptor antagonist, caused lesions in the brain areas where glutamergic pathways were concentrated, and saxitoxins have been shown to cause respiratory paralysis due to neuronal cell death that is induced by high calcium levels in the cells. In mass mortality events of various marine mammal species, both compound classes have been found in organs and serum.

Advisor: Berglund, Olof
Supervisor: Fournier, Michel (INRS-Institut Armand-Frappier)
Master´s Degree Project 60 credits in Ecotoxicology from September 2011 – May 2012
Department of Biology, Lund University},
  author       = {Kleinert, Christine},
  language     = {eng},
  note         = {Student Paper},
  title        = {Comparative sensitivity of the harbor seal (Phoca vitulina) 11B7501 cell line to natural and anthropogenic environmental contaminants},
  year         = {2012},
}