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Transcriptional response of rat frontal cortex following acute in vivo exposure to the pyrethroid insecticides permethrin and deltamethrin.

Harrill JA, Li Z, Wright FA, Radio NM, Mundy WR, Tornero-Velez R, Crofton KM - BMC Genomics (2008)

Bottom Line: In the present study, pyrethroids induced changes in gene expression in the frontal cortex near the threshold for decreases in ambulatory motor activity in vivo.Finally, SAFE analysis of gene expression data identified branching morphogenesis as a biological process sensitive to pyrethroids and subsequent in vitro experiments confirmed this predicted effect.The novel findings regarding pyrethroid effects on branching morphogenesis indicate these compounds may act as developmental neurotoxicants that affect normal neuronal morphology.

View Article: PubMed Central - HTML - PubMed

Affiliation: Curriculum in Toxicology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA. harrill.josh@epa.gov

ABSTRACT

Background: Pyrethroids are neurotoxic pesticides that interact with membrane bound ion channels in neurons and disrupt nerve function. The purpose of this study was to characterize and explore changes in gene expression that occur in the rat frontal cortex, an area of CNS affected by pyrethroids, following an acute low-dose exposure.

Results: Rats were acutely exposed to either deltamethrin (0.3 - 3 mg/kg) or permethrin (1 - 100 mg/kg) followed by collection of cortical tissue at 6 hours. The doses used range from those that cause minimal signs of intoxication at the behavioral level to doses well below apparent no effect levels in the whole animal. A statistical framework based on parallel linear (SAM) and isotonic regression (PIR) methods identified 95 and 53 probe sets as dose-responsive. The PIR analysis was most sensitive for detecting transcripts with changes in expression at the NOAEL dose. A sub-set of genes (Camk1g, Ddc, Gpd3, c-fos and Egr1) was then confirmed by qRT-PCR and examined in a time course study. Changes in mRNA levels were typically less than 3-fold in magnitude across all components of the study. The responses observed are consistent with pyrethroids producing increased neuronal excitation in the cortex following a low-dose in vivo exposure. In addition, Significance Analysis of Function and Expression (SAFE) identified significantly enriched gene categories common for both pyrethroids, including some relating to branching morphogenesis. Exposure of primary cortical cell cultures to both compounds resulted in an increase (approximately 25%) in the number of neurite branch points, supporting the results of the SAFE analysis.

Conclusion: In the present study, pyrethroids induced changes in gene expression in the frontal cortex near the threshold for decreases in ambulatory motor activity in vivo. The penalized regression methods performed similarly in detecting dose-dependent changes in gene transcription. Finally, SAFE analysis of gene expression data identified branching morphogenesis as a biological process sensitive to pyrethroids and subsequent in vitro experiments confirmed this predicted effect. The novel findings regarding pyrethroid effects on branching morphogenesis indicate these compounds may act as developmental neurotoxicants that affect normal neuronal morphology.

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Comparison of PIR and SAM regression methods. Panels A & B plot the penalized isotonic regression (PIR) test statistic (Mi, x-axis) against the penalized linear regression (SAM) test statistic (di, y-axis) for deltamethrin and permethrin, respectively. All 31,042 probe sets present on the Affymetrix Rat 230 2.0 GeneChip® are shown. Data points in green have an empirical p-value < 0.01 for both the PIR and SAM methods. Data points in blue have an empirical p-value < 0.01 for the PIR regression only. Data points in orange have an empirical p-value < 0.01 for the SAM regression only. In the deltamethrin and permethrin analyses, 49.5% and 53.7% of all probe sets identified by either the PIR or SAM method had p < 0.01 for both methods. Data points circled in red have a q-value < 0.10 in permutation-based FDR calculations employed in the SAM algorithm. Note that the rank order of statistical significance was similar between the two methods in that probe sets commonly identified using the PIR or SAM method tend to appear in the upper-right and lower left hand corners of the scatterplots (green points).
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Figure 1: Comparison of PIR and SAM regression methods. Panels A & B plot the penalized isotonic regression (PIR) test statistic (Mi, x-axis) against the penalized linear regression (SAM) test statistic (di, y-axis) for deltamethrin and permethrin, respectively. All 31,042 probe sets present on the Affymetrix Rat 230 2.0 GeneChip® are shown. Data points in green have an empirical p-value < 0.01 for both the PIR and SAM methods. Data points in blue have an empirical p-value < 0.01 for the PIR regression only. Data points in orange have an empirical p-value < 0.01 for the SAM regression only. In the deltamethrin and permethrin analyses, 49.5% and 53.7% of all probe sets identified by either the PIR or SAM method had p < 0.01 for both methods. Data points circled in red have a q-value < 0.10 in permutation-based FDR calculations employed in the SAM algorithm. Note that the rank order of statistical significance was similar between the two methods in that probe sets commonly identified using the PIR or SAM method tend to appear in the upper-right and lower left hand corners of the scatterplots (green points).

Mentions: Both the PIR (isotonic) and SAM (linear) penalized regression methods identified dose-dependent increases and decreases in mRNA expression in the frontal cortex 6 h after an acute, oral exposure to both deltamethrin and permethrin. A comparison of the PIR and SAM regression models demonstrate that the two methods yield similar results in terms of identifying dose-responsive probe sets for both deltamethrin and permethrin (Figure 1A &1B). SAM analyses identified a small number of probe sets with dose-dependent increases in expression following either deltamethrin (n = 7) or permethrin (n = 10) exposure using the permutation-based FDR values as the significance criteria (q < 0.10, see Figure 1A &1B). The PIR analyses did not identify any probe sets for either pyrethroid with dose-dependent changes in expression at q < 0.10. A less statistically conservative method of identifying dose-related changes in probe set expression identified a larger number of significantly altered probe sets than that observed using the FDR criteria. Using a screening threshold of p < 0.01 the SAM analysis identified 70 and 61 probe sets with dose-dependent changes in expression for deltamethrin and permethrin, respectively, while the PIR analysis identified 93 and 85, respectively (Figure 1A–B). The overlap between probe sets identified as dose-responsive using the empirical p-value thresholds is considerable but incomplete. Overall, these parallel methods yield comparable results in that a rank-ordered list of dose-dependent changes in expression constructed using either the PIR or SAM test-statistics identifies the same groups of probe sets as being the most significantly changed within both the deltamethrin and permethrin test cohorts


Transcriptional response of rat frontal cortex following acute in vivo exposure to the pyrethroid insecticides permethrin and deltamethrin.

Harrill JA, Li Z, Wright FA, Radio NM, Mundy WR, Tornero-Velez R, Crofton KM - BMC Genomics (2008)

Comparison of PIR and SAM regression methods. Panels A & B plot the penalized isotonic regression (PIR) test statistic (Mi, x-axis) against the penalized linear regression (SAM) test statistic (di, y-axis) for deltamethrin and permethrin, respectively. All 31,042 probe sets present on the Affymetrix Rat 230 2.0 GeneChip® are shown. Data points in green have an empirical p-value < 0.01 for both the PIR and SAM methods. Data points in blue have an empirical p-value < 0.01 for the PIR regression only. Data points in orange have an empirical p-value < 0.01 for the SAM regression only. In the deltamethrin and permethrin analyses, 49.5% and 53.7% of all probe sets identified by either the PIR or SAM method had p < 0.01 for both methods. Data points circled in red have a q-value < 0.10 in permutation-based FDR calculations employed in the SAM algorithm. Note that the rank order of statistical significance was similar between the two methods in that probe sets commonly identified using the PIR or SAM method tend to appear in the upper-right and lower left hand corners of the scatterplots (green points).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2626604&req=5

Figure 1: Comparison of PIR and SAM regression methods. Panels A & B plot the penalized isotonic regression (PIR) test statistic (Mi, x-axis) against the penalized linear regression (SAM) test statistic (di, y-axis) for deltamethrin and permethrin, respectively. All 31,042 probe sets present on the Affymetrix Rat 230 2.0 GeneChip® are shown. Data points in green have an empirical p-value < 0.01 for both the PIR and SAM methods. Data points in blue have an empirical p-value < 0.01 for the PIR regression only. Data points in orange have an empirical p-value < 0.01 for the SAM regression only. In the deltamethrin and permethrin analyses, 49.5% and 53.7% of all probe sets identified by either the PIR or SAM method had p < 0.01 for both methods. Data points circled in red have a q-value < 0.10 in permutation-based FDR calculations employed in the SAM algorithm. Note that the rank order of statistical significance was similar between the two methods in that probe sets commonly identified using the PIR or SAM method tend to appear in the upper-right and lower left hand corners of the scatterplots (green points).
Mentions: Both the PIR (isotonic) and SAM (linear) penalized regression methods identified dose-dependent increases and decreases in mRNA expression in the frontal cortex 6 h after an acute, oral exposure to both deltamethrin and permethrin. A comparison of the PIR and SAM regression models demonstrate that the two methods yield similar results in terms of identifying dose-responsive probe sets for both deltamethrin and permethrin (Figure 1A &1B). SAM analyses identified a small number of probe sets with dose-dependent increases in expression following either deltamethrin (n = 7) or permethrin (n = 10) exposure using the permutation-based FDR values as the significance criteria (q < 0.10, see Figure 1A &1B). The PIR analyses did not identify any probe sets for either pyrethroid with dose-dependent changes in expression at q < 0.10. A less statistically conservative method of identifying dose-related changes in probe set expression identified a larger number of significantly altered probe sets than that observed using the FDR criteria. Using a screening threshold of p < 0.01 the SAM analysis identified 70 and 61 probe sets with dose-dependent changes in expression for deltamethrin and permethrin, respectively, while the PIR analysis identified 93 and 85, respectively (Figure 1A–B). The overlap between probe sets identified as dose-responsive using the empirical p-value thresholds is considerable but incomplete. Overall, these parallel methods yield comparable results in that a rank-ordered list of dose-dependent changes in expression constructed using either the PIR or SAM test-statistics identifies the same groups of probe sets as being the most significantly changed within both the deltamethrin and permethrin test cohorts

Bottom Line: In the present study, pyrethroids induced changes in gene expression in the frontal cortex near the threshold for decreases in ambulatory motor activity in vivo.Finally, SAFE analysis of gene expression data identified branching morphogenesis as a biological process sensitive to pyrethroids and subsequent in vitro experiments confirmed this predicted effect.The novel findings regarding pyrethroid effects on branching morphogenesis indicate these compounds may act as developmental neurotoxicants that affect normal neuronal morphology.

View Article: PubMed Central - HTML - PubMed

Affiliation: Curriculum in Toxicology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA. harrill.josh@epa.gov

ABSTRACT

Background: Pyrethroids are neurotoxic pesticides that interact with membrane bound ion channels in neurons and disrupt nerve function. The purpose of this study was to characterize and explore changes in gene expression that occur in the rat frontal cortex, an area of CNS affected by pyrethroids, following an acute low-dose exposure.

Results: Rats were acutely exposed to either deltamethrin (0.3 - 3 mg/kg) or permethrin (1 - 100 mg/kg) followed by collection of cortical tissue at 6 hours. The doses used range from those that cause minimal signs of intoxication at the behavioral level to doses well below apparent no effect levels in the whole animal. A statistical framework based on parallel linear (SAM) and isotonic regression (PIR) methods identified 95 and 53 probe sets as dose-responsive. The PIR analysis was most sensitive for detecting transcripts with changes in expression at the NOAEL dose. A sub-set of genes (Camk1g, Ddc, Gpd3, c-fos and Egr1) was then confirmed by qRT-PCR and examined in a time course study. Changes in mRNA levels were typically less than 3-fold in magnitude across all components of the study. The responses observed are consistent with pyrethroids producing increased neuronal excitation in the cortex following a low-dose in vivo exposure. In addition, Significance Analysis of Function and Expression (SAFE) identified significantly enriched gene categories common for both pyrethroids, including some relating to branching morphogenesis. Exposure of primary cortical cell cultures to both compounds resulted in an increase (approximately 25%) in the number of neurite branch points, supporting the results of the SAFE analysis.

Conclusion: In the present study, pyrethroids induced changes in gene expression in the frontal cortex near the threshold for decreases in ambulatory motor activity in vivo. The penalized regression methods performed similarly in detecting dose-dependent changes in gene transcription. Finally, SAFE analysis of gene expression data identified branching morphogenesis as a biological process sensitive to pyrethroids and subsequent in vitro experiments confirmed this predicted effect. The novel findings regarding pyrethroid effects on branching morphogenesis indicate these compounds may act as developmental neurotoxicants that affect normal neuronal morphology.

Show MeSH
Related in: MedlinePlus