Limits...
Spatial transcription of CYP1A in fish liver.

Olsvik PA, Lie KK, Saele Ø, Sanden M - BMC Physiol. (2007)

Bottom Line: Transcription of CYP1A and GST was higher in the middle section of the liver compared to the distal and proximal parts of the organ.Overall, the qRT-PCR and ISH results reported here suggest that gene expression analysis should be performed on as pure cell populations as possible.If bulk tissue samples are to be used, one should always check how evenly the target genes are expressed in tissue sections and organs in every study.

View Article: PubMed Central - HTML - PubMed

Affiliation: National Institute of Nutrition and Seafood Research, N-5817 Bergen, Norway. pal.olsvik@nifes.no

ABSTRACT

Background: The aim of this work was to study how evenly detoxifying genes are transcribed spatially in liver tissue of fish. Ten Atlantic salmon Salmo salar were intraperitoneally injected with 50 mg/kg of the strong CYP1A inducer beta-naphthoflavone and liver tissue harvested seven days later. The liver from 10 control and 10 exposed fish were split into eight sections, RNA extracted and three reference (beta-actin, elongation factor 1AB (EF1AB)) and two detoxifying genes (CYP1A and GST) quantified with real-time RT-PCR. The cellular localization of the EF1AB and CYP1A mRNA in the liver of control and beta-naphthoflavone treated fish was then determined by in situ hybridization (ISH) using EF1AB and CYP1A biotinylated oligonucleotide probes.

Results: The study shows that genes encoding phase I and phase II conjugating enzymes are unevenly transcribed in different parts of the liver of Atlantic salmon seven days after a single-dose of beta-naphthoflavone exposure. Transcription of CYP1A and GST was higher in the middle section of the liver compared to the distal and proximal parts of the organ. The ISH data suggest that CYP1A transcription happens mainly in hepatocyte cells in the liver, and that hepatocytes in the vicinity of blood vessels respond stronger to beta-naphthoflavone than cells further away from the blood supply.

Conclusion: Overall, the qRT-PCR and ISH results reported here suggest that gene expression analysis should be performed on as pure cell populations as possible. If bulk tissue samples are to be used, one should always check how evenly the target genes are expressed in tissue sections and organs in every study.

Show MeSH

Related in: MedlinePlus

Atlantic salmon Salmo salar liver partitioned into eight transversal sections for transcriptional analysis and three transversal cross-sections for in situ hybridization. P = Proximal, M = Mid, D = Distal. Scale bar = cm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2238752&req=5

Figure 1: Atlantic salmon Salmo salar liver partitioned into eight transversal sections for transcriptional analysis and three transversal cross-sections for in situ hybridization. P = Proximal, M = Mid, D = Distal. Scale bar = cm.

Mentions: In this study the goal was to examine the macroscopic distribution and cellular localization of two detoxifying genes and of three reference genes to evaluate if these are evenly expressed throughout the different parts of the Atlantic salmon liver. For this reason, the liver was cut transversally into eight parts (Fig. 1), and RNA extracted from each part for quantitative qRT-PCR analysis in control fish as well as in fish exposed to the strong CYP1A inducer BNF. We also sliced off three cross-sections of the liver for histological examination, to evaluate if two of the studied genes are evenly expressed in different cell types. The three sections were cut transversally from the proximal, mid and distal regions of the liver. In situ hybridization was used to examine the spatial patterns of gene expression of one detoxifying gene (CYP1A) and one reference gene (elongation factor 1AB in anatomically different areas of the liver.


Spatial transcription of CYP1A in fish liver.

Olsvik PA, Lie KK, Saele Ø, Sanden M - BMC Physiol. (2007)

Atlantic salmon Salmo salar liver partitioned into eight transversal sections for transcriptional analysis and three transversal cross-sections for in situ hybridization. P = Proximal, M = Mid, D = Distal. Scale bar = cm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2238752&req=5

Figure 1: Atlantic salmon Salmo salar liver partitioned into eight transversal sections for transcriptional analysis and three transversal cross-sections for in situ hybridization. P = Proximal, M = Mid, D = Distal. Scale bar = cm.
Mentions: In this study the goal was to examine the macroscopic distribution and cellular localization of two detoxifying genes and of three reference genes to evaluate if these are evenly expressed throughout the different parts of the Atlantic salmon liver. For this reason, the liver was cut transversally into eight parts (Fig. 1), and RNA extracted from each part for quantitative qRT-PCR analysis in control fish as well as in fish exposed to the strong CYP1A inducer BNF. We also sliced off three cross-sections of the liver for histological examination, to evaluate if two of the studied genes are evenly expressed in different cell types. The three sections were cut transversally from the proximal, mid and distal regions of the liver. In situ hybridization was used to examine the spatial patterns of gene expression of one detoxifying gene (CYP1A) and one reference gene (elongation factor 1AB in anatomically different areas of the liver.

Bottom Line: Transcription of CYP1A and GST was higher in the middle section of the liver compared to the distal and proximal parts of the organ.Overall, the qRT-PCR and ISH results reported here suggest that gene expression analysis should be performed on as pure cell populations as possible.If bulk tissue samples are to be used, one should always check how evenly the target genes are expressed in tissue sections and organs in every study.

View Article: PubMed Central - HTML - PubMed

Affiliation: National Institute of Nutrition and Seafood Research, N-5817 Bergen, Norway. pal.olsvik@nifes.no

ABSTRACT

Background: The aim of this work was to study how evenly detoxifying genes are transcribed spatially in liver tissue of fish. Ten Atlantic salmon Salmo salar were intraperitoneally injected with 50 mg/kg of the strong CYP1A inducer beta-naphthoflavone and liver tissue harvested seven days later. The liver from 10 control and 10 exposed fish were split into eight sections, RNA extracted and three reference (beta-actin, elongation factor 1AB (EF1AB)) and two detoxifying genes (CYP1A and GST) quantified with real-time RT-PCR. The cellular localization of the EF1AB and CYP1A mRNA in the liver of control and beta-naphthoflavone treated fish was then determined by in situ hybridization (ISH) using EF1AB and CYP1A biotinylated oligonucleotide probes.

Results: The study shows that genes encoding phase I and phase II conjugating enzymes are unevenly transcribed in different parts of the liver of Atlantic salmon seven days after a single-dose of beta-naphthoflavone exposure. Transcription of CYP1A and GST was higher in the middle section of the liver compared to the distal and proximal parts of the organ. The ISH data suggest that CYP1A transcription happens mainly in hepatocyte cells in the liver, and that hepatocytes in the vicinity of blood vessels respond stronger to beta-naphthoflavone than cells further away from the blood supply.

Conclusion: Overall, the qRT-PCR and ISH results reported here suggest that gene expression analysis should be performed on as pure cell populations as possible. If bulk tissue samples are to be used, one should always check how evenly the target genes are expressed in tissue sections and organs in every study.

Show MeSH
Related in: MedlinePlus