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Age and ovariectomy abolish beneficial effects of female sex on rat ventricular myocytes exposed to simulated ischemia and reperfusion.

Ross JL, Howlett SE - PLoS ONE (2012)

Bottom Line: Cell shortening (edge detector) and intracellular Ca(2+) (fura-2) were measured simultaneously.However, contraction amplitudes were reduced in reperfusion in male myocytes, while contractions recovered to exceed control levels in females (62.6±5.1 vs. 140.1±15.8%; p<0.05).Age and OVX abolish these beneficial effects and induce Ca(2+) dysregulation at the level of the cardiomyocyte.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Dalhousie University, Halifax, Nova Scotia, Canada.

ABSTRACT
Sex differences in responses to myocardial ischemia have been described, but whether cardiomyocyte function is influenced by sex in the setting of ischemia and reperfusion has not been elucidated. This study compared contractions and intracellular Ca(2+) in isolated ventricular myocytes exposed to ischemia and reperfusion. Cells were isolated from anesthetized 3-month-old male and female Fischer 344 rats, paced at 4 Hz (37°C), exposed to simulated ischemia (20 mins) and reperfused. Cell shortening (edge detector) and intracellular Ca(2+) (fura-2) were measured simultaneously. Cell viability was assessed with Trypan blue. Ischemia reduced peak contractions and increased Ca(2+) levels equally in myocytes from both sexes. However, contraction amplitudes were reduced in reperfusion in male myocytes, while contractions recovered to exceed control levels in females (62.6±5.1 vs. 140.1±15.8%; p<0.05). Only 60% of male myocytes excluded trypan blue dye after ischemia and reperfusion, while all female cardiomyocytes excluded the dye (p<0.05). Parallel experiments were conducted in myocytes from ∼24-month-old female rats or 5-6-month-old rats that had an ovariectomy at 3-4 weeks of age. Beneficial effects of female sex on myocyte viability and contractile dysfunction in reperfusion were abolished in cells from 24-month-old females. Aged female myocytes also exhibited elevated intracellular Ca(2+) and alternans in ischemia. Cells from ovariectomized rats displayed increased Ca(2+) transients and spontaneous activity in ischemia compared to sham-operated controls. None of the myocytes from ovariectomized rats were viable after 15 minutes of ischemia, while 75% of sham cells remained viable at end of reperfusion (p<0.05). These findings demonstrate that cardiomyocytes from young adult females are more resistant to ischemia and reperfusion injury than cells from males. Age and OVX abolish these beneficial effects and induce Ca(2+) dysregulation at the level of the cardiomyocyte. Thus, beneficial effects of estrogen in ischemia and reperfusion are mediated, in part, by effects on cardiomyocytes.

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Aging promoted Ca2+ accumulation in ischemia and abolished beneficial effects of female sex on contractile function and cell viability.The experimental protocol is described in the legends to Figures 1 and 2. A. Mean amplitudes of contractions in young adult (filled circles) and aged female (open triangles) cells throughout exposure to simulated ischemia and reperfusion. B. Average peak Ca2+ transients recorded from young adult and aged female myocytes at 5 minute intervals throughout the experimental protocol. Mean levels of diastolic Ca2+ (C) and resting myocyte length (D) recorded at 5 minute intervals during the experiments in cells from young adult and aged females. All data were normalized to values recorded after 15 minutes of stimulation in the absence of ischemia. E. Survival curves illustrating the viability of young adult (solid line) and aged female (dashed line) cells reported at 5 minute intervals throughout the experiment. The * denotes significantly different from young adult female value (p<0.05; n = 10 young adult female cells and 8 aged female cells).
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pone-0038425-g004: Aging promoted Ca2+ accumulation in ischemia and abolished beneficial effects of female sex on contractile function and cell viability.The experimental protocol is described in the legends to Figures 1 and 2. A. Mean amplitudes of contractions in young adult (filled circles) and aged female (open triangles) cells throughout exposure to simulated ischemia and reperfusion. B. Average peak Ca2+ transients recorded from young adult and aged female myocytes at 5 minute intervals throughout the experimental protocol. Mean levels of diastolic Ca2+ (C) and resting myocyte length (D) recorded at 5 minute intervals during the experiments in cells from young adult and aged females. All data were normalized to values recorded after 15 minutes of stimulation in the absence of ischemia. E. Survival curves illustrating the viability of young adult (solid line) and aged female (dashed line) cells reported at 5 minute intervals throughout the experiment. The * denotes significantly different from young adult female value (p<0.05; n = 10 young adult female cells and 8 aged female cells).

Mentions: To determine whether ischemia and reperfusion injury in myocytes from female rats was exacerbated by aging, responses were compared in myocytes from young adult and aged female rats. Ischemia reduced peak contractions in both groups (Figure 4A). However, while contractions fully recovered upon reperfusion in the young group, contractions did not fully recover in the aged group (Figure 4A). Ca2+ transients were similar during most of ischemia and reperfusion in both groups (Figure 4B). Interestingly, aging augmented the rise in diastolic Ca2+ that occurred in ischemia (Figure 4C). The degree of hypercontracture in reperfusion was similar in the two groups (Figure 4D). However, even though all young female myocytes remained viable throughout ischemia and reperfusion, almost 30% of the aged myocytes were trypan blue positive by the end of reperfusion (p<0.05; Figure 4E). These results show that aging promoted Ca2+ loading in ischemia and abolished the beneficial effect of female sex on cell viability and contractile function in reperfusion.


Age and ovariectomy abolish beneficial effects of female sex on rat ventricular myocytes exposed to simulated ischemia and reperfusion.

Ross JL, Howlett SE - PLoS ONE (2012)

Aging promoted Ca2+ accumulation in ischemia and abolished beneficial effects of female sex on contractile function and cell viability.The experimental protocol is described in the legends to Figures 1 and 2. A. Mean amplitudes of contractions in young adult (filled circles) and aged female (open triangles) cells throughout exposure to simulated ischemia and reperfusion. B. Average peak Ca2+ transients recorded from young adult and aged female myocytes at 5 minute intervals throughout the experimental protocol. Mean levels of diastolic Ca2+ (C) and resting myocyte length (D) recorded at 5 minute intervals during the experiments in cells from young adult and aged females. All data were normalized to values recorded after 15 minutes of stimulation in the absence of ischemia. E. Survival curves illustrating the viability of young adult (solid line) and aged female (dashed line) cells reported at 5 minute intervals throughout the experiment. The * denotes significantly different from young adult female value (p<0.05; n = 10 young adult female cells and 8 aged female cells).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3368849&req=5

pone-0038425-g004: Aging promoted Ca2+ accumulation in ischemia and abolished beneficial effects of female sex on contractile function and cell viability.The experimental protocol is described in the legends to Figures 1 and 2. A. Mean amplitudes of contractions in young adult (filled circles) and aged female (open triangles) cells throughout exposure to simulated ischemia and reperfusion. B. Average peak Ca2+ transients recorded from young adult and aged female myocytes at 5 minute intervals throughout the experimental protocol. Mean levels of diastolic Ca2+ (C) and resting myocyte length (D) recorded at 5 minute intervals during the experiments in cells from young adult and aged females. All data were normalized to values recorded after 15 minutes of stimulation in the absence of ischemia. E. Survival curves illustrating the viability of young adult (solid line) and aged female (dashed line) cells reported at 5 minute intervals throughout the experiment. The * denotes significantly different from young adult female value (p<0.05; n = 10 young adult female cells and 8 aged female cells).
Mentions: To determine whether ischemia and reperfusion injury in myocytes from female rats was exacerbated by aging, responses were compared in myocytes from young adult and aged female rats. Ischemia reduced peak contractions in both groups (Figure 4A). However, while contractions fully recovered upon reperfusion in the young group, contractions did not fully recover in the aged group (Figure 4A). Ca2+ transients were similar during most of ischemia and reperfusion in both groups (Figure 4B). Interestingly, aging augmented the rise in diastolic Ca2+ that occurred in ischemia (Figure 4C). The degree of hypercontracture in reperfusion was similar in the two groups (Figure 4D). However, even though all young female myocytes remained viable throughout ischemia and reperfusion, almost 30% of the aged myocytes were trypan blue positive by the end of reperfusion (p<0.05; Figure 4E). These results show that aging promoted Ca2+ loading in ischemia and abolished the beneficial effect of female sex on cell viability and contractile function in reperfusion.

Bottom Line: Cell shortening (edge detector) and intracellular Ca(2+) (fura-2) were measured simultaneously.However, contraction amplitudes were reduced in reperfusion in male myocytes, while contractions recovered to exceed control levels in females (62.6±5.1 vs. 140.1±15.8%; p<0.05).Age and OVX abolish these beneficial effects and induce Ca(2+) dysregulation at the level of the cardiomyocyte.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Dalhousie University, Halifax, Nova Scotia, Canada.

ABSTRACT
Sex differences in responses to myocardial ischemia have been described, but whether cardiomyocyte function is influenced by sex in the setting of ischemia and reperfusion has not been elucidated. This study compared contractions and intracellular Ca(2+) in isolated ventricular myocytes exposed to ischemia and reperfusion. Cells were isolated from anesthetized 3-month-old male and female Fischer 344 rats, paced at 4 Hz (37°C), exposed to simulated ischemia (20 mins) and reperfused. Cell shortening (edge detector) and intracellular Ca(2+) (fura-2) were measured simultaneously. Cell viability was assessed with Trypan blue. Ischemia reduced peak contractions and increased Ca(2+) levels equally in myocytes from both sexes. However, contraction amplitudes were reduced in reperfusion in male myocytes, while contractions recovered to exceed control levels in females (62.6±5.1 vs. 140.1±15.8%; p<0.05). Only 60% of male myocytes excluded trypan blue dye after ischemia and reperfusion, while all female cardiomyocytes excluded the dye (p<0.05). Parallel experiments were conducted in myocytes from ∼24-month-old female rats or 5-6-month-old rats that had an ovariectomy at 3-4 weeks of age. Beneficial effects of female sex on myocyte viability and contractile dysfunction in reperfusion were abolished in cells from 24-month-old females. Aged female myocytes also exhibited elevated intracellular Ca(2+) and alternans in ischemia. Cells from ovariectomized rats displayed increased Ca(2+) transients and spontaneous activity in ischemia compared to sham-operated controls. None of the myocytes from ovariectomized rats were viable after 15 minutes of ischemia, while 75% of sham cells remained viable at end of reperfusion (p<0.05). These findings demonstrate that cardiomyocytes from young adult females are more resistant to ischemia and reperfusion injury than cells from males. Age and OVX abolish these beneficial effects and induce Ca(2+) dysregulation at the level of the cardiomyocyte. Thus, beneficial effects of estrogen in ischemia and reperfusion are mediated, in part, by effects on cardiomyocytes.

Show MeSH
Related in: MedlinePlus