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Serotonin potentiates transforming growth factor-beta3 induced biomechanical remodeling in avian embryonic atrioventricular valves.

Buskohl PR, Sun MJ, Sun ML, Thompson RP, Butcher JT - PLoS ONE (2012)

Bottom Line: Blockade of TGFβ type I receptors via SB431542 inhibited the TGFβ3 effects.Elevated 5-HT in ovo resulted in elevated remodeling gene expression and increased TGFβ signaling activity, supporting our ex-vivo findings.Collectively, these results highlight TGFβ/5-HT signaling as a potent mechanism for control of biomechanical remodeling of AV cushions during development.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical and Aerospace Engineering, Cornell University, Ithaca, New York, United States of America.

ABSTRACT
Embryonic heart valve primordia (cushions) maintain unidirectional blood flow during development despite an increasingly demanding mechanical environment. Recent studies demonstrate that atrioventricular (AV) cushions stiffen over gestation, but the molecular mechanisms of this process are unknown. Transforming growth factor-beta (TGFβ) and serotonin (5-HT) signaling modulate tissue biomechanics of postnatal valves, but less is known of their role in the biomechanical remodeling of embryonic valves. In this study, we demonstrate that exogenous TGFβ3 increases AV cushion biomechanical stiffness and residual stress, but paradoxically reduces matrix compaction. We then show that TGFβ3 induces contractile gene expression (RhoA, aSMA) and extracellular matrix expression (col1α2) in cushion mesenchyme, while simultaneously stimulating a two-fold increase in proliferation. Local compaction increased due to an elevated contractile phenotype, but global compaction appeared reduced due to proliferation and ECM synthesis. Blockade of TGFβ type I receptors via SB431542 inhibited the TGFβ3 effects. We next showed that exogenous 5-HT does not influence cushion stiffness by itself, but synergistically increases cushion stiffness with TGFβ3 co-treatment. 5-HT increased TGFβ3 gene expression and also potentiated TGFβ3 induced gene expression in a dose-dependent manner. Blockade of the 5HT2b receptor, but not 5-HT2a receptor or serotonin transporter (SERT), resulted in complete cessation of TGFβ3 induced mechanical strengthening. Finally, systemic 5-HT administration in ovo induced cushion remodeling related defects, including thinned/atretic AV valves, ventricular septal defects, and outflow rotation defects. Elevated 5-HT in ovo resulted in elevated remodeling gene expression and increased TGFβ signaling activity, supporting our ex-vivo findings. Collectively, these results highlight TGFβ/5-HT signaling as a potent mechanism for control of biomechanical remodeling of AV cushions during development.

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5-HT treatment modulates TGFβ3 mediated gene expression.A) TGFβ3 mRNA transcripts increase with physiological 5-HT (470 nM, 5-HT), but decrease at high dose (47 µM, 5-HT+). αSMA, RhoA, and col1α2, were not affected by physiological 5-HT dose, but were significantly downregulated with high 5-HT treatment. B) High 5-HT treatment mitigates exogenous TGFβ3 induced contractile gene expression, while TGFβ3 induced proliferation was independent of 5-HT dose. mean ± SEM, n = 3–5 pooled samples of 8–10 cushions, *p<0.05 via ANOVA comparisons with controls.
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pone-0042527-g005: 5-HT treatment modulates TGFβ3 mediated gene expression.A) TGFβ3 mRNA transcripts increase with physiological 5-HT (470 nM, 5-HT), but decrease at high dose (47 µM, 5-HT+). αSMA, RhoA, and col1α2, were not affected by physiological 5-HT dose, but were significantly downregulated with high 5-HT treatment. B) High 5-HT treatment mitigates exogenous TGFβ3 induced contractile gene expression, while TGFβ3 induced proliferation was independent of 5-HT dose. mean ± SEM, n = 3–5 pooled samples of 8–10 cushions, *p<0.05 via ANOVA comparisons with controls.

Mentions: Exogenous 5-HT administration potentiated remodeling-relevant gene expression in organ cultured AV cushion mesenchyme. TGFβ3 mRNA transitioned from 1.9±0.1 fold upregulation over controls at physiological 5-HT to 0.40±0.16 downregulation at high 5-HT dose (Figure 5A). The physiological 5-HT dose had no statistically significant effect on αSMA, col1α2, cyclin b2, and RhoA expression. In contrast, high 5-HT significantly decreased transcription of αSMA (0.18±0.09), collagen1α2 (0.22±0.07), and RhoA (0.46±0.11 Figure 5A). No effect on cyclin b2 expression was observed at either dose, suggesting proliferation was not directly regulated by 5-HT. Physiological 5-HT did not affect TGFβ3 induced gene expression (Figure 5B), but high dose 5-HT markedly reduced gene expression of TGFβ3 (0.86±0.20 vs. 2.2±0.6), αSMA (1.4±0.4 vs. 5.3±0.4), collagen1α2 (1.3±0.3 vs. 3.8±0.9), and RhoA (1.3±0.2 vs. 2.1±0.3) (Figure 5B). Proliferation-related gene cyclin b2 was not significantly affected by 5-HT in combination with TGFβ3. These results suggest that exogenous 5-HT potentiates TGFβ3 more likely through interaction with upstream activation points and/or TGFβ3 synthesis, rather than by interacting with TGFβ3 downstream targets directly.


Serotonin potentiates transforming growth factor-beta3 induced biomechanical remodeling in avian embryonic atrioventricular valves.

Buskohl PR, Sun MJ, Sun ML, Thompson RP, Butcher JT - PLoS ONE (2012)

5-HT treatment modulates TGFβ3 mediated gene expression.A) TGFβ3 mRNA transcripts increase with physiological 5-HT (470 nM, 5-HT), but decrease at high dose (47 µM, 5-HT+). αSMA, RhoA, and col1α2, were not affected by physiological 5-HT dose, but were significantly downregulated with high 5-HT treatment. B) High 5-HT treatment mitigates exogenous TGFβ3 induced contractile gene expression, while TGFβ3 induced proliferation was independent of 5-HT dose. mean ± SEM, n = 3–5 pooled samples of 8–10 cushions, *p<0.05 via ANOVA comparisons with controls.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3412853&req=5

pone-0042527-g005: 5-HT treatment modulates TGFβ3 mediated gene expression.A) TGFβ3 mRNA transcripts increase with physiological 5-HT (470 nM, 5-HT), but decrease at high dose (47 µM, 5-HT+). αSMA, RhoA, and col1α2, were not affected by physiological 5-HT dose, but were significantly downregulated with high 5-HT treatment. B) High 5-HT treatment mitigates exogenous TGFβ3 induced contractile gene expression, while TGFβ3 induced proliferation was independent of 5-HT dose. mean ± SEM, n = 3–5 pooled samples of 8–10 cushions, *p<0.05 via ANOVA comparisons with controls.
Mentions: Exogenous 5-HT administration potentiated remodeling-relevant gene expression in organ cultured AV cushion mesenchyme. TGFβ3 mRNA transitioned from 1.9±0.1 fold upregulation over controls at physiological 5-HT to 0.40±0.16 downregulation at high 5-HT dose (Figure 5A). The physiological 5-HT dose had no statistically significant effect on αSMA, col1α2, cyclin b2, and RhoA expression. In contrast, high 5-HT significantly decreased transcription of αSMA (0.18±0.09), collagen1α2 (0.22±0.07), and RhoA (0.46±0.11 Figure 5A). No effect on cyclin b2 expression was observed at either dose, suggesting proliferation was not directly regulated by 5-HT. Physiological 5-HT did not affect TGFβ3 induced gene expression (Figure 5B), but high dose 5-HT markedly reduced gene expression of TGFβ3 (0.86±0.20 vs. 2.2±0.6), αSMA (1.4±0.4 vs. 5.3±0.4), collagen1α2 (1.3±0.3 vs. 3.8±0.9), and RhoA (1.3±0.2 vs. 2.1±0.3) (Figure 5B). Proliferation-related gene cyclin b2 was not significantly affected by 5-HT in combination with TGFβ3. These results suggest that exogenous 5-HT potentiates TGFβ3 more likely through interaction with upstream activation points and/or TGFβ3 synthesis, rather than by interacting with TGFβ3 downstream targets directly.

Bottom Line: Blockade of TGFβ type I receptors via SB431542 inhibited the TGFβ3 effects.Elevated 5-HT in ovo resulted in elevated remodeling gene expression and increased TGFβ signaling activity, supporting our ex-vivo findings.Collectively, these results highlight TGFβ/5-HT signaling as a potent mechanism for control of biomechanical remodeling of AV cushions during development.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical and Aerospace Engineering, Cornell University, Ithaca, New York, United States of America.

ABSTRACT
Embryonic heart valve primordia (cushions) maintain unidirectional blood flow during development despite an increasingly demanding mechanical environment. Recent studies demonstrate that atrioventricular (AV) cushions stiffen over gestation, but the molecular mechanisms of this process are unknown. Transforming growth factor-beta (TGFβ) and serotonin (5-HT) signaling modulate tissue biomechanics of postnatal valves, but less is known of their role in the biomechanical remodeling of embryonic valves. In this study, we demonstrate that exogenous TGFβ3 increases AV cushion biomechanical stiffness and residual stress, but paradoxically reduces matrix compaction. We then show that TGFβ3 induces contractile gene expression (RhoA, aSMA) and extracellular matrix expression (col1α2) in cushion mesenchyme, while simultaneously stimulating a two-fold increase in proliferation. Local compaction increased due to an elevated contractile phenotype, but global compaction appeared reduced due to proliferation and ECM synthesis. Blockade of TGFβ type I receptors via SB431542 inhibited the TGFβ3 effects. We next showed that exogenous 5-HT does not influence cushion stiffness by itself, but synergistically increases cushion stiffness with TGFβ3 co-treatment. 5-HT increased TGFβ3 gene expression and also potentiated TGFβ3 induced gene expression in a dose-dependent manner. Blockade of the 5HT2b receptor, but not 5-HT2a receptor or serotonin transporter (SERT), resulted in complete cessation of TGFβ3 induced mechanical strengthening. Finally, systemic 5-HT administration in ovo induced cushion remodeling related defects, including thinned/atretic AV valves, ventricular septal defects, and outflow rotation defects. Elevated 5-HT in ovo resulted in elevated remodeling gene expression and increased TGFβ signaling activity, supporting our ex-vivo findings. Collectively, these results highlight TGFβ/5-HT signaling as a potent mechanism for control of biomechanical remodeling of AV cushions during development.

Show MeSH
Related in: MedlinePlus