Limits...
Systematic analysis of gene expression differences between left and right atria in different mouse strains and in human atrial tissue.

Kahr PC, Piccini I, Fabritz L, Greber B, Schöler H, Scheld HH, Hoffmeier A, Brown NA, Kirchhof P - PLoS ONE (2011)

Bottom Line: Normal development of the atria requires left-right differentiation during embryonic development.These differences were validated by RT-qPCR in murine and human tissue.Western blot showed a 2-fold left-right concentration gradient in PITX2 protein in adult human atria.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology and Angiology, University Hospital Münster, Münster, Germany.

ABSTRACT

Background: Normal development of the atria requires left-right differentiation during embryonic development. Reduced expression of Pitx2c (paired-like homeodomain transcription factor 2, isoform c), a key regulator of left-right asymmetry, has recently been linked to atrial fibrillation. We therefore systematically studied the molecular composition of left and right atrial tissue in adult murine and human atria.

Methods: We compared left and right atrial gene expression in healthy, adult mice of different strains and ages by employing whole genome array analyses on freshly frozen atrial tissue. Selected genes with enriched expression in either atrium were validated by RT-qPCR and Western blot in further animals and in shock-frozen left and right atrial appendages of patients undergoing open heart surgery.

Results: We identified 77 genes with preferential expression in one atrium that were common in all strains and age groups analysed. Independent of strain and age, Pitx2c was the gene with the highest enrichment in left atrium, while Bmp10, a member of the TGFβ family, showed highest enrichment in right atrium. These differences were validated by RT-qPCR in murine and human tissue. Western blot showed a 2-fold left-right concentration gradient in PITX2 protein in adult human atria. Several of the genes and gene groups enriched in left atria have a known biological role for maintenance of healthy physiology, specifically the prevention of atrial pathologies involved in atrial fibrillation, including membrane electrophysiology, metabolic cellular function, and regulation of inflammatory processes. Comparison of the array datasets with published array analyses in heterozygous Pitx2c(+/-) atria suggested that approximately half of the genes with left-sided enrichment are regulated by Pitx2c.

Conclusions: Our study reveals systematic differences between left and right atrial gene expression and supports the hypothesis that Pitx2c has a functional role in maintaining "leftness" in the atrium in adult murine and human hearts.

Show MeSH

Related in: MedlinePlus

RT-qPCR validation of the differential expression of candidate genes in left and right atria from wild-type mice of three different strains.A Relative Log2 expression ratios between LA and RA in MF1 mice (blue, n = 4), Swiss-Agouti mice (pink, n = 4), CD1 mice (yellow, n = 4), and in wild-type mice (grey, n = 12). B Corresponding average mRNA expression in left (black) and right (white) atrial tissues of MF1, SA, and CD1 mice (n = 4 each). Gapdh was used as control. Error bars indicate standard error of the mean (SEM). Statistically significant differences as assessed by unpaired Student's t-test are represented by asterisks (*P<0.05, **P<0.01).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3198471&req=5

pone-0026389-g003: RT-qPCR validation of the differential expression of candidate genes in left and right atria from wild-type mice of three different strains.A Relative Log2 expression ratios between LA and RA in MF1 mice (blue, n = 4), Swiss-Agouti mice (pink, n = 4), CD1 mice (yellow, n = 4), and in wild-type mice (grey, n = 12). B Corresponding average mRNA expression in left (black) and right (white) atrial tissues of MF1, SA, and CD1 mice (n = 4 each). Gapdh was used as control. Error bars indicate standard error of the mean (SEM). Statistically significant differences as assessed by unpaired Student's t-test are represented by asterisks (*P<0.05, **P<0.01).

Mentions: To validate the microarray data set, we selected left (n = 5) and right (n = 4) marker candidates with different degrees of overlap between the array datasets (2 genes identified in only 1 dataset, 1 gene in 2 datasets, and 6 genes identified in 3 datasets). We assessed mRNA concentrations by RT-qPCR on additional samples of MF1 mice (n = 4, 3-month-old) and Swiss-Agouti (n = 4, 12-months-old), and on a third wild-type mouse strain (CD1, n = 4, 3.5-months-old). All genes tested by RT-qPCR were significantly differently expressed between left and right atrial tissues in at least one of the wild-type mouse strains (Figure 3A–B). For all genes, the atrium showing enriched expression was the same by array and RT-qPCR. Overall, RT-qPCR confirmed the microarray data, both in enrichment in a given tissue (right versus left atrium) and across different mouse strains (Figure 3A and 3B).


Systematic analysis of gene expression differences between left and right atria in different mouse strains and in human atrial tissue.

Kahr PC, Piccini I, Fabritz L, Greber B, Schöler H, Scheld HH, Hoffmeier A, Brown NA, Kirchhof P - PLoS ONE (2011)

RT-qPCR validation of the differential expression of candidate genes in left and right atria from wild-type mice of three different strains.A Relative Log2 expression ratios between LA and RA in MF1 mice (blue, n = 4), Swiss-Agouti mice (pink, n = 4), CD1 mice (yellow, n = 4), and in wild-type mice (grey, n = 12). B Corresponding average mRNA expression in left (black) and right (white) atrial tissues of MF1, SA, and CD1 mice (n = 4 each). Gapdh was used as control. Error bars indicate standard error of the mean (SEM). Statistically significant differences as assessed by unpaired Student's t-test are represented by asterisks (*P<0.05, **P<0.01).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3198471&req=5

pone-0026389-g003: RT-qPCR validation of the differential expression of candidate genes in left and right atria from wild-type mice of three different strains.A Relative Log2 expression ratios between LA and RA in MF1 mice (blue, n = 4), Swiss-Agouti mice (pink, n = 4), CD1 mice (yellow, n = 4), and in wild-type mice (grey, n = 12). B Corresponding average mRNA expression in left (black) and right (white) atrial tissues of MF1, SA, and CD1 mice (n = 4 each). Gapdh was used as control. Error bars indicate standard error of the mean (SEM). Statistically significant differences as assessed by unpaired Student's t-test are represented by asterisks (*P<0.05, **P<0.01).
Mentions: To validate the microarray data set, we selected left (n = 5) and right (n = 4) marker candidates with different degrees of overlap between the array datasets (2 genes identified in only 1 dataset, 1 gene in 2 datasets, and 6 genes identified in 3 datasets). We assessed mRNA concentrations by RT-qPCR on additional samples of MF1 mice (n = 4, 3-month-old) and Swiss-Agouti (n = 4, 12-months-old), and on a third wild-type mouse strain (CD1, n = 4, 3.5-months-old). All genes tested by RT-qPCR were significantly differently expressed between left and right atrial tissues in at least one of the wild-type mouse strains (Figure 3A–B). For all genes, the atrium showing enriched expression was the same by array and RT-qPCR. Overall, RT-qPCR confirmed the microarray data, both in enrichment in a given tissue (right versus left atrium) and across different mouse strains (Figure 3A and 3B).

Bottom Line: Normal development of the atria requires left-right differentiation during embryonic development.These differences were validated by RT-qPCR in murine and human tissue.Western blot showed a 2-fold left-right concentration gradient in PITX2 protein in adult human atria.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology and Angiology, University Hospital Münster, Münster, Germany.

ABSTRACT

Background: Normal development of the atria requires left-right differentiation during embryonic development. Reduced expression of Pitx2c (paired-like homeodomain transcription factor 2, isoform c), a key regulator of left-right asymmetry, has recently been linked to atrial fibrillation. We therefore systematically studied the molecular composition of left and right atrial tissue in adult murine and human atria.

Methods: We compared left and right atrial gene expression in healthy, adult mice of different strains and ages by employing whole genome array analyses on freshly frozen atrial tissue. Selected genes with enriched expression in either atrium were validated by RT-qPCR and Western blot in further animals and in shock-frozen left and right atrial appendages of patients undergoing open heart surgery.

Results: We identified 77 genes with preferential expression in one atrium that were common in all strains and age groups analysed. Independent of strain and age, Pitx2c was the gene with the highest enrichment in left atrium, while Bmp10, a member of the TGFβ family, showed highest enrichment in right atrium. These differences were validated by RT-qPCR in murine and human tissue. Western blot showed a 2-fold left-right concentration gradient in PITX2 protein in adult human atria. Several of the genes and gene groups enriched in left atria have a known biological role for maintenance of healthy physiology, specifically the prevention of atrial pathologies involved in atrial fibrillation, including membrane electrophysiology, metabolic cellular function, and regulation of inflammatory processes. Comparison of the array datasets with published array analyses in heterozygous Pitx2c(+/-) atria suggested that approximately half of the genes with left-sided enrichment are regulated by Pitx2c.

Conclusions: Our study reveals systematic differences between left and right atrial gene expression and supports the hypothesis that Pitx2c has a functional role in maintaining "leftness" in the atrium in adult murine and human hearts.

Show MeSH
Related in: MedlinePlus