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Fetal programming of blood pressure in a transgenic mouse model of altered intrauterine environment
S34 SMFM Abstracts 75
DEVELOPMENTAL ENHANCEMENT IN THE NEONATAL PERIOD IN HEALTHY MICE LAURA TOSO1, IRENE CAMERONI2, DANIEL ABEBE1, CATHERINE SPONG1, 1Unit on Perinatal and Developmental Neurobiology, NICHD&NIAAA, National Institute of Health (NIH), Bethesda, Maryland, 2University of Milano-Bicocca, Obstetrics and Gynecology, Monza, Milan, Italy OBJECTIVE: In a previous study we showed that either prenatal or postnatal administration of neurotrophic peptides (NAPCSAL) enhanced learning in adult and aged healthy mice (Toso L, 2006). Prenatal treatment also prevented alcohol-induced neonatal developmental deficits in a model for fetal alcohol syndrome (Endres M, 2005). In a model for Down Syndrome (DS) prenatal treatment prevented developmental deficits in neonates with DS. Since only 1⁄4 of the progeny inherits the trisomy, 3/4 of the litter treated were healthy wild type. We were interested in looking at the peptides’ effect on development in this group. STUDY DESIGN: Ts65Dn females were mated with wild-type (B6C3HF1) male mice. Pregnant females were treated either with peptides (NAPCSAL 20mgC20mg, i.p.) or placebo on gestational days 8-12. Offspring were tested for developmental milestones (7 motor, 6 sensory) on postnatal days 5-21 by investigators blinded to treatment and genotype. Determination of genotype for DS or wild type (control)was by rt-PCR for amyloid precursor protein. Genotype was determined after completion of all studies. Statistical analysis included ANOVA with post-hoc Bonferroni-Dunn with P!0.05 considered significant. RESULTS: 17 controlCplacebo pups from 6 litters and 9 controlC NAPCSAL pups from 6 litters were tested. Treatment with NAPCSAL resulted in enhancement in achieving developmental milestones in 5/7 (71%) motor (cliff aversion, negative geotaxis, righting, screen climbing and vibrissa placing) and 2/6 (33%) sensory (ear twitch and weak tactile stimulation) milestones, all p!0.05.
77
MULTI-DRUG RESISTANCE RECEPTORS REGULATE TRANSMISSION OF THERAPEUTIC AGENTS ACROSS THE PLACENTA SAMUEL PARRY1, JIAN ZHANG1, 1University of Pennsylvania, Obstetrics and Gynecology, Philadelphia, Pennsylvania OBJECTIVE: Multi-drug resistance (MDR) proteins are efflux transporters that are expressed in the human placenta and are thought to protect the fetus from harmful drug exposures. We studied the role of MDR proteins in transplacental transmission of potentially therapeutic agents, including protease inhibitors and corticosteroids. STUDY DESIGN: We used quantitative PCR and FACS analyses to measure MDR-1 and MRP-1 mRNA and protein expression in transformed extravillous trophoblast (HTR-8/SVneo) cells and villous cytotrophoblast (BeWo) cells before and after treatment with saquinavir, which is an MDR protein substrate. We measured H3-labeled dexamethasone and H3-labeled ritonavir levels in BeWo cells before and after treatment with anti-MDR-1 antibodies or cyclosporine, which inhibits MDR proteins in other cells. All experiments were performed in triplicate and repeated three times. RESULTS: Compared to baseline expression levels, MDR-1 and MRP-1 mRNA levels were increased 2.0-5.6 fold in HTR-8/SVneo and BeWo cells after treatment with saquinavir. GAPDH mRNA levels (control) were unaffected by treatment with saquinavir. The MDR proteins were expressed on 18.2-34.0% of BeWo cells, and protein expression was significantly increased after treatment with saquinavir. H3-dexamethasone levels increased 2.0 fold in BeWo cells after treatment with anti-MDR-1 antibodies (p!0.0001), while H3-ritonavir levels increased 1.6 fold after treatment with anti-MDR-1 antibodies (p!0.0001). Similar results were observed when MDR protein function was blocked by treatment with cyclosporine. Doseresponse and time-course relationships were established. CONCLUSION: MDR proteins regulate drug levels in trophoblast cells and may mediate transmission of potentially therapeutic agents across the placenta. Further studies are warranted to determine the effects of individual variations in MDR protein expression and the potential therapeutic benefits of agents that block MDR protein function in the placenta. 0002-9378/$ - see front matter doi:10.1016/j.ajog.2006.10.090
NAPCSAL enhanced development CONCLUSION: Prenatal treatment with neurotrophic peptides NAPCSAL enhanced neonatal development in normal mice. These findings highlight a new possibility for the prevention of pathologic conditions that affect the neonatal period. 0002-9378/$ - see front matter doi:10.1016/j.ajog.2006.10.087 76
FETAL PROGRAMMING OF BLOOD PRESSURE IN A TRANSGENIC MOUSE MODEL OF ALTERED INTRAUTERINE ENVIRONMENT MONICA LONGO1, FANGXIAN LU1, PHYLLIS GAMBLE1, GARLAND D. ANDERSON1, ESTHER TAMAYO1, GARY D. V. HANKINS1, GEORGE SAADE1, 1University of Texas Medical Branch, Obstetrics & Gynecology/Maternal Fetal Medicine, Galveston, Texas OBJECTIVE: Nitric oxide is involved in the vascular adaptation to pregnancy, and knockout (KO) mice lacking nitric oxide synthase (NOS3) have abnormal utero-placental perfusion. Using these transgenic animals, we previously showed that the adverse intrauterine environment alters fetal programming vascular reactivity in the adult offspring. Our aim in this study was to determine if the altered vascular programming noted in vitro is associated with altered blood pressure (BP) in vivo. STUDY DESIGN: Homozygous NOS3 knockout (C57BL/6J-Nos3tm1Unc, NOS3ÿ/ÿKO) and wild-type controls (NOS3C/CWT) were cross-bred to obtain heterozygous offspring that developed in KO mothers (NOS3C/ÿMat) versus wild-type control mothers (NOS3C/ÿPat). At 21 weeks of age, the NOS3C/ÿMat and NOS3C/ÿPat heterozygous male offspring were fitted with catheters inserted through the carotid artery into the aortic arch and connected to a telemetric transmitter for BP measurement. After recovery, the BP was recorded continuously for 3 days in the conscious unrestrained animals. Daily mean BP, as well as mean BP during daytime (8 am to 8 pm) and nighttime (8 pm to 8 am) were calculated. Student t-test was used for statistical analysis (significance: p!0.05). RESULTS: Daily mean BP was significantly higher in the NOS3C/ÿMat compared with NOS3C/ÿPat offspring throughout the 3 day recording (118.04G1.09 versus 109.3G0.7 mmHg; p!0.05). Daytime and nighttime mean BP remained higher in the the NOS3C/ÿMat compared with NOS3C/ÿPat offspring. However, diurnal variation of BP was maintained within each group, with lower mean BP in nighttime versus daytime. CONCLUSION: Despite being genomically-similar, blood pressure in adult offspring that developed in a uterine environment lacking NOS3 is higher than the offspring that developed in a normal uterine environment. These findings support that the fetal vascular programming previously reported in vitro is associated with altered blood pressure in vivo, and highlight the role of the intrauterine environment in determining health and disease in the adult. 0002-9378/$ - see front matter doi:10.1016/j.ajog.2006.10.089
78
DURATION OF LACTATION AND INCIDENCE OF MYOCARDIAL INFARCTION ALISON STUEBE1, KARIN MICHELS1, WALTER WILLETT2, JOANN MANSON3, JANET RICH-EDWARDS3, 1Brigham and Women’s Hospital, Harvard Medical School, Department of Obstetrics and Gynecology, Boston, Massachusetts, 2 Harvard School of Public Health, Boston, Massachusetts, 3Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts OBJECTIVE: Lactation is associated with favorable maternal glucose and lipid metabolism, as well as lower blood pressures. Mothers who breastfeed have a lower risk of type 2 diabetes. We hypothesized that lactation may also be associated with a lower risk of coronary heart disease. STUDY DESIGN: We conducted a prospective observational cohort study of 92,648 parous women in the Nurses Health Study. Our primary outcome was incident myocardial infarction or sudden death from 1986 to 2002. RESULTS: During the study period, 2923 cases of coronary heart disease were diagnosed during 1,442,735 person-years of follow-up. Women who had breastfed for more than 23 months had 34% lower risk of coronary heart disease (95% CI 20-45%, p for trend !0.0001) than women who never breastfed, controlling for age, parity and history of stillbirth. Adjusting for covariates, women who breastfed for more than 23 months had a 19% lower coronary heart disease risk (95% CI 2-33%, p for trend = 0.027) than women who never breastfed, controlling for early adult adiposity, family history, smoking, diet, exercise, and use of aspirin, postmenopausal hormones and multivitamins (see Table). CONCLUSION: In a large, prospective cohort study, longer duration of breastfeeding was associated with a reduced risk of coronary heart disease. Incident Coronary Heart Disease and Duration of Lactation in Months. HR (95% CI) None
O0 to 3
O3 to 6
O6 to 11
O11 to 23
O23
Cases P-Y Parity, ageadjusted
1202 531413 1.0 (ref)
Covariateadjusted
1.0 (ref)
721 329240 0.90 (0.82 0.98) 1.00 (0.91 1.09)
341 172215 0.89 (0.79 – 1.00) 0.97 (0.86 1.10)
259 139429 0.90 (0.79 1.03) 1.03 (0.90 1.18)
271 174092 0.77 (0.67 0.88) 0.91 (0.80 1.04)
129 96346 0.66 (0.55 0.80) 0.81 (0.67 0.98)
0002-9378/$ - see front matter doi:10.1016/j.ajog.2006.10.091
DEVELOPMENTAL ENHANCEMENT IN THE NEONATAL PERIOD IN HEALTHY MICE LAURA TOSO1, IRENE CAMERONI2, DANIEL ABEBE1, CATHERINE SPONG1, 1Unit on Perinatal and Developmental Neurobiology, NICHD&NIAAA, National Institute of Health (NIH), Bethesda, Maryland, 2University of Milano-Bicocca, Obstetrics and Gynecology, Monza, Milan, Italy OBJECTIVE: In a previous study we showed that either prenatal or postnatal administration of neurotrophic peptides (NAPCSAL) enhanced learning in adult and aged healthy mice (Toso L, 2006). Prenatal treatment also prevented alcohol-induced neonatal developmental deficits in a model for fetal alcohol syndrome (Endres M, 2005). In a model for Down Syndrome (DS) prenatal treatment prevented developmental deficits in neonates with DS. Since only 1⁄4 of the progeny inherits the trisomy, 3/4 of the litter treated were healthy wild type. We were interested in looking at the peptides’ effect on development in this group. STUDY DESIGN: Ts65Dn females were mated with wild-type (B6C3HF1) male mice. Pregnant females were treated either with peptides (NAPCSAL 20mgC20mg, i.p.) or placebo on gestational days 8-12. Offspring were tested for developmental milestones (7 motor, 6 sensory) on postnatal days 5-21 by investigators blinded to treatment and genotype. Determination of genotype for DS or wild type (control)was by rt-PCR for amyloid precursor protein. Genotype was determined after completion of all studies. Statistical analysis included ANOVA with post-hoc Bonferroni-Dunn with P!0.05 considered significant. RESULTS: 17 controlCplacebo pups from 6 litters and 9 controlC NAPCSAL pups from 6 litters were tested. Treatment with NAPCSAL resulted in enhancement in achieving developmental milestones in 5/7 (71%) motor (cliff aversion, negative geotaxis, righting, screen climbing and vibrissa placing) and 2/6 (33%) sensory (ear twitch and weak tactile stimulation) milestones, all p!0.05.
77
MULTI-DRUG RESISTANCE RECEPTORS REGULATE TRANSMISSION OF THERAPEUTIC AGENTS ACROSS THE PLACENTA SAMUEL PARRY1, JIAN ZHANG1, 1University of Pennsylvania, Obstetrics and Gynecology, Philadelphia, Pennsylvania OBJECTIVE: Multi-drug resistance (MDR) proteins are efflux transporters that are expressed in the human placenta and are thought to protect the fetus from harmful drug exposures. We studied the role of MDR proteins in transplacental transmission of potentially therapeutic agents, including protease inhibitors and corticosteroids. STUDY DESIGN: We used quantitative PCR and FACS analyses to measure MDR-1 and MRP-1 mRNA and protein expression in transformed extravillous trophoblast (HTR-8/SVneo) cells and villous cytotrophoblast (BeWo) cells before and after treatment with saquinavir, which is an MDR protein substrate. We measured H3-labeled dexamethasone and H3-labeled ritonavir levels in BeWo cells before and after treatment with anti-MDR-1 antibodies or cyclosporine, which inhibits MDR proteins in other cells. All experiments were performed in triplicate and repeated three times. RESULTS: Compared to baseline expression levels, MDR-1 and MRP-1 mRNA levels were increased 2.0-5.6 fold in HTR-8/SVneo and BeWo cells after treatment with saquinavir. GAPDH mRNA levels (control) were unaffected by treatment with saquinavir. The MDR proteins were expressed on 18.2-34.0% of BeWo cells, and protein expression was significantly increased after treatment with saquinavir. H3-dexamethasone levels increased 2.0 fold in BeWo cells after treatment with anti-MDR-1 antibodies (p!0.0001), while H3-ritonavir levels increased 1.6 fold after treatment with anti-MDR-1 antibodies (p!0.0001). Similar results were observed when MDR protein function was blocked by treatment with cyclosporine. Doseresponse and time-course relationships were established. CONCLUSION: MDR proteins regulate drug levels in trophoblast cells and may mediate transmission of potentially therapeutic agents across the placenta. Further studies are warranted to determine the effects of individual variations in MDR protein expression and the potential therapeutic benefits of agents that block MDR protein function in the placenta. 0002-9378/$ - see front matter doi:10.1016/j.ajog.2006.10.090
NAPCSAL enhanced development CONCLUSION: Prenatal treatment with neurotrophic peptides NAPCSAL enhanced neonatal development in normal mice. These findings highlight a new possibility for the prevention of pathologic conditions that affect the neonatal period. 0002-9378/$ - see front matter doi:10.1016/j.ajog.2006.10.087 76
FETAL PROGRAMMING OF BLOOD PRESSURE IN A TRANSGENIC MOUSE MODEL OF ALTERED INTRAUTERINE ENVIRONMENT MONICA LONGO1, FANGXIAN LU1, PHYLLIS GAMBLE1, GARLAND D. ANDERSON1, ESTHER TAMAYO1, GARY D. V. HANKINS1, GEORGE SAADE1, 1University of Texas Medical Branch, Obstetrics & Gynecology/Maternal Fetal Medicine, Galveston, Texas OBJECTIVE: Nitric oxide is involved in the vascular adaptation to pregnancy, and knockout (KO) mice lacking nitric oxide synthase (NOS3) have abnormal utero-placental perfusion. Using these transgenic animals, we previously showed that the adverse intrauterine environment alters fetal programming vascular reactivity in the adult offspring. Our aim in this study was to determine if the altered vascular programming noted in vitro is associated with altered blood pressure (BP) in vivo. STUDY DESIGN: Homozygous NOS3 knockout (C57BL/6J-Nos3tm1Unc, NOS3ÿ/ÿKO) and wild-type controls (NOS3C/CWT) were cross-bred to obtain heterozygous offspring that developed in KO mothers (NOS3C/ÿMat) versus wild-type control mothers (NOS3C/ÿPat). At 21 weeks of age, the NOS3C/ÿMat and NOS3C/ÿPat heterozygous male offspring were fitted with catheters inserted through the carotid artery into the aortic arch and connected to a telemetric transmitter for BP measurement. After recovery, the BP was recorded continuously for 3 days in the conscious unrestrained animals. Daily mean BP, as well as mean BP during daytime (8 am to 8 pm) and nighttime (8 pm to 8 am) were calculated. Student t-test was used for statistical analysis (significance: p!0.05). RESULTS: Daily mean BP was significantly higher in the NOS3C/ÿMat compared with NOS3C/ÿPat offspring throughout the 3 day recording (118.04G1.09 versus 109.3G0.7 mmHg; p!0.05). Daytime and nighttime mean BP remained higher in the the NOS3C/ÿMat compared with NOS3C/ÿPat offspring. However, diurnal variation of BP was maintained within each group, with lower mean BP in nighttime versus daytime. CONCLUSION: Despite being genomically-similar, blood pressure in adult offspring that developed in a uterine environment lacking NOS3 is higher than the offspring that developed in a normal uterine environment. These findings support that the fetal vascular programming previously reported in vitro is associated with altered blood pressure in vivo, and highlight the role of the intrauterine environment in determining health and disease in the adult. 0002-9378/$ - see front matter doi:10.1016/j.ajog.2006.10.089
78
DURATION OF LACTATION AND INCIDENCE OF MYOCARDIAL INFARCTION ALISON STUEBE1, KARIN MICHELS1, WALTER WILLETT2, JOANN MANSON3, JANET RICH-EDWARDS3, 1Brigham and Women’s Hospital, Harvard Medical School, Department of Obstetrics and Gynecology, Boston, Massachusetts, 2 Harvard School of Public Health, Boston, Massachusetts, 3Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts OBJECTIVE: Lactation is associated with favorable maternal glucose and lipid metabolism, as well as lower blood pressures. Mothers who breastfeed have a lower risk of type 2 diabetes. We hypothesized that lactation may also be associated with a lower risk of coronary heart disease. STUDY DESIGN: We conducted a prospective observational cohort study of 92,648 parous women in the Nurses Health Study. Our primary outcome was incident myocardial infarction or sudden death from 1986 to 2002. RESULTS: During the study period, 2923 cases of coronary heart disease were diagnosed during 1,442,735 person-years of follow-up. Women who had breastfed for more than 23 months had 34% lower risk of coronary heart disease (95% CI 20-45%, p for trend !0.0001) than women who never breastfed, controlling for age, parity and history of stillbirth. Adjusting for covariates, women who breastfed for more than 23 months had a 19% lower coronary heart disease risk (95% CI 2-33%, p for trend = 0.027) than women who never breastfed, controlling for early adult adiposity, family history, smoking, diet, exercise, and use of aspirin, postmenopausal hormones and multivitamins (see Table). CONCLUSION: In a large, prospective cohort study, longer duration of breastfeeding was associated with a reduced risk of coronary heart disease. Incident Coronary Heart Disease and Duration of Lactation in Months. HR (95% CI) None
O0 to 3
O3 to 6
O6 to 11
O11 to 23
O23
Cases P-Y Parity, ageadjusted
1202 531413 1.0 (ref)
Covariateadjusted
1.0 (ref)
721 329240 0.90 (0.82 0.98) 1.00 (0.91 1.09)
341 172215 0.89 (0.79 – 1.00) 0.97 (0.86 1.10)
259 139429 0.90 (0.79 1.03) 1.03 (0.90 1.18)
271 174092 0.77 (0.67 0.88) 0.91 (0.80 1.04)
129 96346 0.66 (0.55 0.80) 0.81 (0.67 0.98)
0002-9378/$ - see front matter doi:10.1016/j.ajog.2006.10.091