Prenatal nicotine exposure increases heart susceptibility to ischemia/reperfusion injury in adult offspring

Abstract

In the present study we tested the hypothesis that prenatal nicotine exposure increases heart susceptibility to ischemia/reperfusion (I/R) injury in adult offspring. Nicotine was administered to pregnant rats via subcutaneous osmotic minipumps throughout gestation. Nicotine treatment resulted in a rapid and transient decrease in food-intake and a moderate decrease in maternal body weight gain. Hearts were isolated from adult male and female offspring and subjected to I/R in a Langendorff preparation. Nicotine significantly attenuated left ventricle (LV) developed pressure, heart rate, and coronary flow rate in female but not male hearts at baseline. Additionally, nicotine significantly increased LV infarct size and attenuated postischemic recovery of LV function in both male and female offspring with more pronounced effects in females. In female but not male hearts, nicotine significantly decreased the postischemic coronary flow rate. However, coronary nitric oxide release was decreased in male but not female hearts. Caspase-3, -8, and -9 levels were not significantly changed in either female or male hearts. However, nicotine caused a significant decrease in protein levels of protein kinase (PK) Cepsilon in both male and female hearts and a decrease in PKCdelta levels in female hearts only. Control studies of maternal food restriction showed that a moderate decrease in maternal body weight gain had no effect on female hearts but significantly improved postischemic recovery of LV function in male hearts. The results suggest that prenatal nicotine exposure causes in utero programming of the PKC isozyme gene expression pattern in the developing heart and increases heart susceptibility to I/R injury in adult offspring.

Fig. 1

Effect of nicotine administration on maternal food intake and body weight gain. Nicotine was administrated to pregnant rats through osmotic pumps, as described under Materials and Methods. Control rats received saline. Maternal food intake and body weight gain were measured daily. In separate studies, pregnant rats were divided into two groups and were fed with food equivalent to that determined in saline control and nicotine-treated animals, respectively. Data are means ± S.E.M. n = 4 to 6.

Fig. 2

Effect of maternal nicotine administration and food restriction on neonatal body weight. Nicotine was administrated to pregnant rats through osmotic pumps, as described under Materials and Methods. Control rats received saline. In separate studies, pregnant rats were divided into two groups and were fed with equivalent food to that determined in saline control and nicotine-treated animals, respectively. Body weight was measured in offspring from 1 to 30 days of age. Data are means ± S.E.M. Data were analyzed by two-way ANOVA with age as one factor and nicotine treatment as the other.*, P < 0.05 versus nicotine treatment. n = 4 to 6 litters.

Fig. 3

Effect of maternal nicotine administration on postischemic recovery of LV function in adult male offspring. Nicotine was administrated to pregnant rats through osmotic pumps, as described under Materials and Methods. Control rats received saline. Hearts were obtained from 3-month-old male offspring and were subjected to 25 min of ischemia and 60 min of reperfusion in a Langendorff preparation. Data are means ± S.E.M. Data were analyzed by two-way ANOVA with ischemia-reperfusion as one factor and nicotine treatment as the other.*, P < 0.05, versus saline control for the entire curve. n = 11.

Fig. 4

Effect of maternal nicotine administration on postischemic recovery of LV function in adult female offspring. Nicotine was administrated to pregnant rats through osmotic pumps, as described under Materials and Methods. Control rats received saline. Hearts were obtained from 3-month-old female offspring and were subjected to 25 min of ischemia and 60 min of reperfusion in a Langendorff preparation. Data are means ± S.E.M. Data were analyzed by two-way ANOVA with ischemia-reperfusion as one factor and nicotine treatment as the other.*, P < 0.05, versus saline control for the entire curve. n = 9 to 11.

Fig. 5

Effect of maternal nicotine administration on I/R-induced myocardial infarction in adult offspring. Nicotine was administrated to pregnant rats through osmotic pumps, as described under Materials and Methods. Control rats received saline. Hearts were obtained from 3-month-old male and female offspring and were subjected to 25 min of ischemia and 60 min of reperfusion in a Langendorff preparation. Left ventricles were collected at the end of reperfusion, and myocardial infarct size was determined with 1% triphenyltetrazolium chloride staining and expressed as a percentage of the total left ventricular weight. Data are means ± S.E.M. a, P < 0.05 versus saline control; b, P < 0.05 versus male. n = 9 to 11.

Fig. 6

Effect of maternal nicotine administration on coronary flow rate in adult offspring. Nicotine was administrated to pregnant rats through osmotic pumps, as described under Materials and Methods. Control rats received saline. Hearts were obtained from 3-month-old male and female offspring and were subjected to 25 min of ischemia and 60 min of reperfusion in a Langendorff preparation. Pulmonary artery effluent was collected as an index of coronary flow (milliliters per minute per gram of heart wet weight). Data are means ± S.E.M. Data were analyzed by two-way ANOVA with ischemia-reperfusion as one factor and nicotine treatment as the other. *, P < 0.05 versus saline control for the entire curve. n = 9 to 11.

Fig. 7

Effect of maternal nicotine administration on coronary NO release in adult offspring. Nicotine was administrated to pregnant rats through osmotic pumps, as described under Materials and Methods. Control rats received saline. Hearts were obtained from 3-month-old male and female offspring and were subjected to 25 min of ischemia and 60 min of reperfusion in a Langendorff preparation. Pulmonary artery effluent was collected and NO (nanomoles per minute per gram of heart wet weight) was measured using the chemiluminescence method, as described under Materials and Methods. Data are means ± S.E.M. Data were analyzed by two-way ANOVA with ischemia-reperfusion as one factor and nicotine treatment as the other. *P < 0.05, versus saline control for the entire curve. n = 9 to 11.

Fig. 8

Effect of maternal nicotine administration on caspase protein levels in the hearts of adult offspring. Nicotine was administrated to pregnant rats through osmotic pumps, as described under Materials and Methods. Control rats received saline. Hearts were obtained from 3-month-old male and female offspring, and caspase-3, -8, and -9 protein levels were determined using Western blot analyses. Data are means ± S.E.M. a, P < 0.05 versus other caspases in the same sex; b, P < 0.05 versus the same caspase in male hearts. n = 4.

Fig. 9

Effect of maternal nicotine administration on PKCε and PKCδ protein levels in the hearts of adult offspring. Nicotine was administrated to pregnant rats through osmotic pumps, as described under Materials and Methods. Control rats received saline. Hearts were obtained from 3-month-old male and female offspring and protein levels of PKCε and PKCδ were determined using Western blot analyses. Data are means ± S.E.M. a, P < 0.05 versus saline control; b, P < 0.05 versus male. n = 4.

Fig. 10

Effect of maternal food restriction on postischemic recovery of LV function in adult offspring. Pregnant rats were divided into two groups and were fed with equivalent food to that determined in saline control and nicotine-treated animals, respectively. Hearts were obtained from 3-month-old male and female offspring and were subjected to 25 min of ischemia and 60 min of reperfusion in a Langendorff preparation. Data are means ± S.E.M. Data were analyzed by two-way ANOVA with ischemia-reperfusion as one factor and maternal food restriction as the other. *, P < 0.05 versus control for the entire curve. n = 5.