Investigation of two different anoxia models by 2-dimensional gel electrophoresis

TY - ABST

T1 - Investigation of two different anoxia models by 2-dimensional gel electrophoresis

AU - Wulff, Tune

AU - Jessen, Flemming

AU - Hoffmann, Else Kay

N1 - Udgivelsesdato: March Volumne: 20

PY - 2006

Y1 - 2006

N2 - This study presents a broad investigation of the effects of different types of anoxia in the Rainbow Trout Hypodermal Fibroblasts cell line, including a comparison of two widely used anoxia models. Anoxia was obtained either by flushing with N2 (FN) or by adding NaN3 to the medium, since chemical anoxia obtained by NaN3 is a widely used model for simulating anoxia (Ossum et al., 2004). The effects of anoxia were studied by protein expression analysis using 2-dimensional gel electrophoresis followed by MS/MS. In this way we were able to separate more than 1500 protein spots with an apparent range of molecular masses from 10 to 120 kDa and pI values from 4 to 7. In response to 24 h of anoxia obtained by FN we found that the sexpression of more than 30 proteins changed, including proteins involved in cell metabolism as well as protective proteins. Chemical anoxia for 30 minutes and subsequent reoxygenation overnight (CO) induced changes in the expression of more than 40 proteins. Two hours of anoxia obtained by FN followed by reoxygenation overnight (AO), resulted in changes in the abundance of more than 30 proteins. Surprisingly the response to AO was very different to the response to CO, actually only the levels of five proteins changed similarly. In summery we found almost 100 different proteins to be involved in the different aspects of anoxia. We also found a remarkable difference between the effects induced by CO compared to AO, showing the existence of great differences in expression patterns depending on the nature of the stimuli. This calls to circumspection when results obtained by different types of anoxia are compared. We thank Fishnet.dk for their financial support.

AB - This study presents a broad investigation of the effects of different types of anoxia in the Rainbow Trout Hypodermal Fibroblasts cell line, including a comparison of two widely used anoxia models. Anoxia was obtained either by flushing with N2 (FN) or by adding NaN3 to the medium, since chemical anoxia obtained by NaN3 is a widely used model for simulating anoxia (Ossum et al., 2004). The effects of anoxia were studied by protein expression analysis using 2-dimensional gel electrophoresis followed by MS/MS. In this way we were able to separate more than 1500 protein spots with an apparent range of molecular masses from 10 to 120 kDa and pI values from 4 to 7. In response to 24 h of anoxia obtained by FN we found that the sexpression of more than 30 proteins changed, including proteins involved in cell metabolism as well as protective proteins. Chemical anoxia for 30 minutes and subsequent reoxygenation overnight (CO) induced changes in the expression of more than 40 proteins. Two hours of anoxia obtained by FN followed by reoxygenation overnight (AO), resulted in changes in the abundance of more than 30 proteins. Surprisingly the response to AO was very different to the response to CO, actually only the levels of five proteins changed similarly. In summery we found almost 100 different proteins to be involved in the different aspects of anoxia. We also found a remarkable difference between the effects induced by CO compared to AO, showing the existence of great differences in expression patterns depending on the nature of the stimuli. This calls to circumspection when results obtained by different types of anoxia are compared. We thank Fishnet.dk for their financial support.

M3 - Conference abstract in journal

SN - 0892-6638

SP - A1433

JO - F A S E B Journal

JF - F A S E B Journal

IS - 5

T2 - Investigation of two different anoxia models by 2-dimensional gel electrophoresis

Y2 - 29 November 2010

ER -