- Email: [email protected]
Effect of moderate intensity exercise on the reproductive performance of diabetic rats
Abstracts / Placenta 36 (2015) 469e521
quantified. Obese and control females were paired with control CD1 males. We evaluated pregnancy the rate and length and litter size. After delivery, pups were weighed during the lactation period. Additionally, LPS (0.05 mg/g) was administered to 15-day pregnant mice and delivery was recorded. Results: Weight-gain was higher in HFD-fed animals than in control and in CB1KO than in WT mice. We found no differences in the daily intake of kilocalories. Abdominal adipose tissue weight increased in HFD-fed animals, in both WT and CB1KO mice. Additionally, we found an increase in serum cholesterol levels in animals fed with HFD, with higher levels in CB1KO than in WT mice. No differences in the pregnancy rate and length or in litter size were found. However, pups born to obese mothers had higher weight gain during the lactation period. Almost 75% of LPS-treated WT mothers presented a defective labor in contrast to 22% in the case of CB1KO mice. Interestingly, when exposed to LPS, up to 50% of the HFD-fed CB1KO mice presented a defective labor. Conclusions: Our results suggest that ECS is involved in the development of maternal obesity, which then has consequences in pups. In addition, obesity sensitized mothers to endotoxin treatment.
PA.10. EARLY PLACENTAL ANGIOGENESIS-VASCULARIZATION AND VEGF/KDR RECEPTOR EXPRESSION DURING MOUSE ORGANOGENESIS AFTER PERIGESTATIONAL ALCOHOL CONSUMPTION M.R. Ventureira 1, C.M.A. Sobarzo 2, M. Naito 3, C. Zanuzzi 4, C. Barbeito 4, E. Reproduction and Maternal-Embryonic Cebral 1. 1 Laboratory of Physiopathology, IFIBYNE-UBA/CONICET Biodiversity and Experimental Biology Department, FCEN, University of Buenos Aires, Buenos Aires, Argentina; 2 INBIOMED-UBA/CONICET- School of Medicine, University of Buenos Aires, Buenos Aires, Argentina; 3 Department of Anatomy, Aichi Medical University, Yazakokarimata, Nagakute, Japan; 4 Histology and Embriology, Basic Sciences Department, Faculty of Veterinary Science, National University of La Plata (UNLP), Buenos Aires, Argentina The normal growth of the fetus depends on adequate placental development, in which the expression of trophoblast-decidual VEGF (vascular endothelial growth factor) conducts angiogenesis-vascularization. Previously, we found that perigestational alcohol consumption at mouse organogenesis (day 10 of gestation), causes embryo resorption, reduces decidualization and leads to histomorphological alterations of the decidual tissue. Objective: The aim was to study, after periconceptional alcohol intake, vascularization of early placentation by monitoring the number of uNK, VEGF and KDR-receptor expressions as well as KDR-activation, in murine decidua and labyrinth during organogenesis. Methods: Ethanol 10% was administered in the drinking water to CF-1 murine females (TF) for 17 days before and up to day 10 of gestation; water was administered to control females (CF). Labyrinthine development and decidual (De) vascularization (H-E, histochemistry for BSI-lectin, morphometry of decidual vascular area and labyrinth), the presence and number of uNKs (PAS and DBA-lectin) and the expression of VEGF and KDR (total and phosphorylated form) by immunohistochemistry, immunofluorescence-confocal and western blot (WB) were analyzed in the implantation sites (IS). Results: Increased percentage of TF-IS had poor labyrinthine growth and reduced De-vascular lumen (p<0.05), while significantly fewer uNKs (PASand DBA-positive cells) were found (p<0.01, vs CF). In TF, the decreased VEGF immunoexpression in trophoblast tissue, endothelial and decidual cells was coincident with its reduced VEGF expression level (p<0.001, vs CF). The immunostaining for KDR (expressed in uNK-PAS) was reduced in the TF-vascular area of decidua (p<0.05), consistent with lower KDR expression level (WB) (p<0.05 vs CF). However, the expression of phosphorylated KDR significantly increased in TF vs CF (p<0.01). Conclusions: Perigestational alcohol exposure inhibits early trophoblastdecidual vascularization by deregulating the VEGF/KDR system, suggesting that, in this mouse model, maternal alcohol intake can lead to vascular resistance and fetal-placental growth restriction at term.
491
PA.11. IMMUNE HOMEOSTASIS DURING THE DECIDUALIZATION PROGRAM: PARTICIPATION OF VIP AND DENDRITIC CELLS Esteban Grasso 1, Soledad Gori 2, Daniel Paparini 1, Lucila rez Gallino 1, Elizabeth Soczewski 1, Gabriela Salamone 2, Claudia Pe s 1, Rosanna Ramhorst 1. 1 Immunopharmacology Laboratory, School of Leiro Sciences, University of Buenos Aires, IQUIBICEN-CONICET, Argentina; 2 Immunology Laboratory, Institute of Experimental Medicine, National Academy of Medicine Buenos Aires, Argentina The decidualization program involves phenotype and functional changes in endometrial cells such as the production of different mediators that will facilitate the attachment and invasion of the blastocyst. Particularly, the vasoactive intestinal peptide (VIP) is a neuropeptide produced by endometrial stromal cells among others, which displays multiple target circuits that allow immunotolerance. Objective: We investigated VIP contribution to the decidualization program and whether this process modulates the profile of dendritic cells (DC). Methods: A human endometrial stromal cell line (hESC) was evaluated. These cells were differentiated in the presence of VIP (10-7M) or medroxyprogesterone (MPA) and dbcAMP (Positive control) for 8 days. Phenotypic markers of decidualization, VIP and their receptors were quantified by RT-PCR. DC were obtained from monocytes isolated from PBMCs from fertile women and differentiated with IL-4 and GM-CSF. VIP and IL-10 secretion were quantified by ELISA. Results: When decidualization was induced by VIP, we observed an increase in characteristic markers such as IGFBP1, PRL and the KLF13/ KLF9 ratio, IL-8 and SDF1 in a concentration-dependent manner (p<0.05). To evaluate the functional aspect of VIP-induced decidualization, an invasion assay was performed using blastocyst-like spheroids (BLS) from first trimester trophoblast cells (Swan71) cultured on hESC monolayer. BLS were able to attach and invade the hESC monolayer decidualized with VIP or MPA+dbcAMP (invasion index X±SE 0.55±0.01 0.64±0.01 0.66±0.01). When these assays were performed with conditioned media obtained from human blastocysts, we found an increase in BLS invasion to hESC decidualized with VIP (p<0.05). Finally, decidualized hESC cells increased VIP expression and secretion. Therefore we evaluated the immunomodulatory effect on DC. Decidualized hESC conditioned media induced a semi-mature profile on DC preventing the induction of HLA-DR, CD83 and CD86 expression, while inducing IL-10 secretion (p<0.05). Conclusions: Our results suggest that VIP may contribute to the decidualization process by inducing phenotypic and functional markers on hESC cells and to the maintenance of a tolerogenic DC-profile.
PA.12. EFFECT OF MODERATE INTENSITY EXERCISE ON THE REPRODUCTIVE PERFORMANCE OF DIABETIC RATS N.C.D. Macedo, A.O. Netto, F.Q. Gallego, I.M.P. Calderon, M.V.C. Rudge, G.T. Volpato, D.C. Damasceno. Laboratory of Experimental Research on Gynecology and Obstetrics, Graduate Program on Gynecology, Obstetrics and Mastology, Botucatu Medical School, Univ. Estadual Paulista_Unesp, ~o Paulo, Brazil Botucatu, Sa Background: Maternal diabetes leads to impaired reproductive outcomes. Although physical exercise is relevant to control glycemia, there are concerns related to the type and intensity of exercise indicated during pregnancy. We hypothesized that an appropriate intensity of exercise improves maternal intrauterine environment leading to favorable embryofetal development. Objective: To evaluate the effects of moderate intensity exercise on the maternal reproductive outcomes of mild diabetic rats.
492
Abstracts / Placenta 36 (2015) 469e521
Methods: Rats were randomly assigned into four groups (n¼12/group): Sedentary non-diabetic (Control¼C, glycemia<120 mg/dL); Sedentary mild diabetic (MD - Streptozotocin - 100 mg/kg b.w., sc., at birth); Exercised non-diabetic (Cex) and Exercised mild diabetic (MDex). Cex and MDex rats were subjected to a swimming program supporting a load of 4% of body weight from day 7 to 18.5 of pregnancy. On day 18.5 of pregnancy, the different experimental groups were killed for exposure of the uterine corns for the analysis of the reproductive outcomes. P<0.05 was considered as a significant statistical limit. Results: MD rats presented glucose intolerance and the area under the curve analysis showed a significant negative correlation between maternal glycemia and a decrease in the number of live fetuses. MD rats showed a lower number of live fetuses and litter weight than C rats. MDex dams presented reduced maternal weight gain, implantation number and litter weight, and increased number of live fetuses as compared with Cex. Both MD groups presented increased fetal and placental weights, and percentage of fetuses classified as large for pregnancy age in relation to control groups. Conclusion: The mild diabetic rats subjected to the swimming program presented no alterations in glucose intolerance, but the number of live fetuses and fetal weight of these rats were consistent with those in the control group, confirming beneficial effects of swimming with an appropriate intensity for the parameters analyzed. Acknowledgement: FAPESP/Brazil (Process number: 2013/23478-3).
PA.13. MATERNAL SATURATED FAT-ENRICHED DIET DETERMINES LEPTIN RESISTANCE IN THE FETAL LIVER AND PROGRAMS LIPID HOMEOSTASIS IMPAIRMENT IN THE OFFSPRING n Mazzucco, Daiana Fornes, Alicia Jawerbaum, Vero nica María Bele White. Centre from Pharmacology and Botanical Studies (CEFyBO) CONICET UBA, Buenos Aires, Argentina Maternal obesity leads to impaired metabolic programming. In a rat model of saturated fat overload in the maternal diet, we have previously found increased fetal liver lipid content. Lipid catabolism in the liver assures proper lipid homeostasis. Leptin regulates lipid catabolism in several tissues. Objective: To investigate whether an overload of saturated fat on the maternal diet impairs leptin effects on hepatic lipid catabolism in rat fetuses at term gestation and whether these anomalies are sustained in the offspring of these rats. Methods: Female Wistar rats were fed with a standard (5% fat) or with a saturated fat diet (28% fat) from 6 weeks of age (SFD rats). After 8 weeks of diet, they were mated with control males. Control and SFD rats were euthanized at 21 days of gestation and fetal livers obtained for further culture of explants (3h) with or without leptin (100 ng/ml). Another group of control and SFD rats were allowed to deliver, and their offspring euthanized at 21 or 130 days of age. Livers were analyzed for lipid accumulation (TLC) and acylCoA oxidase (ACO) expression (PCR). Results: Leptin induced a decrease in triglycerides (49%), free fatty acids (75%) and cholesteryl esters (42%) concentrations in fetal livers from control rats (p<0.05), but no changes but no changes in fetal livers from SFD rats, which showed. Leptin induced an increased ACO expression (50%, p<0.05) in control fetal livers. Fetal livers from SFD rats showed a decrease in ACO expression (25% p<0.05) and no changes with leptin additions. A decrease in ACO expression were also observed in the livers of 21- and 130day-old offspring from SFD rats (20%, p<0.05). Conclusions: Saturated fat in maternal diet induces hepatic lipid metabolism impairment in fetuses and offspring. Liver leptin resistance on lipid catabolism begins in utero and maybe responsible for the alterations on the offspring lipid metabolism.
PA.14. METABOLIC ABNORMALITIES IN THE FETAL LIVER OF RATS WITH GESTATIONAL DIABETES INDUCED BY INTRAUTERINE PROGRAMMING Daiana Fornes, Romina Higa, Ivana Linenberg, Evangelina Capobianco, Alicia Jawerbaum. Laboratory of Reproduction and Metabolism, CEFYBO-CONICET, School of Medicine, University of Buenos Aires, Buenos Aires, Argentina Gestational diabetes (GDM) is a prevalent disease with adverse consequences to both mothers and offspring. We have recently described a new GDM model obtained through adverse intrauterine programming in the offspring of diabetic rats. This model allows analyzing the regulation of lipid metabolism in the fetal liver of GDM mothers. Peroxisome proliferator activated receptors (PPARs) are nuclear receptors involved in lipid metabolic pathways, being PPARa a major regulator of lipid catabolism, while PPARg is highly involved in lipid synthesis and deposition in different tissues. Objective: To evaluate whether lipid concentrations are altered and related to impaired PPAR protein expression in the fetal liver of GDM rats. Methods: Control and GDM rats (obtained from intrauterine programming (F1) in the offspring of streptozotocin-induced diabetic rats (F0)) were evaluated on day 21 of pregnancy. Glucose and insulin concentrations were measured in maternal and fetal serum. Lipid concentrations were measured by TLC and PPARa and PPARg protein expression were evaluated through immunohistochemistry. Results: GDM rats showed increased glycemia and insulinemia only in pregnancy (p<0.01). Male and female fetuses had increased glycemia and insulinemia (p<0.01). Male fetuses from GDM rats showed increased triglycerides (33%) and cholesterol (30%) concentrations in the liver when compared to controls (p<0.05). In these GDM fetal tissues, PPARg protein expression was increased (p<0.001) whereas PPARa protein expression showed no changes compared to controls. Differently, female fetuses from GDM rats showed reduced triglycerides (34%), cholesterol (26%), phospholipids (29%) and free fatty acids (25%) in the liver, p<0.02, compared to controls. Also, in these GDM fetal tissues, both PPARg and PPARa levels were reduced (p<0.01). Conclusion: In a GDM model induced by intrauterine programming, the altered lipid deposition in the fetal liver is gender-dependent and possibly related to changes in the concentrations of the different PPAR isotypes in this fetal tissue.
PA.15. SP1 TRANSCRIPTION FACTOR IS INVOLVED IN THE REGULATION OF PLACENTAL LEPTIN EXPRESSION n Toro 1, Julieta Maymo 1, A. Pe rez-Pe rez 2, Ye sica Malena Schanton 1, Ayele nchez Margalet 2, Cecilia Gambino 1, Bernardo Maskin 3, Victor Sa gica, FCEN, UBA, IQUIBICEN, Varone 1. 1 Departamento de Química Biolo edica y CONICET, Buenos Aires, Argentina; 2 Departamento de Bioquímica M Biología Molecular, Universidad de Sevilla, Sevilla, Spain; 3 Hospital Nacional Profesor Alejandro Posadas, Buenos Aires, Argentina Objectives: Leptin is a key hormone in placental physiology. It regulates trophoblast proliferation, inhibits apoptosis, stimulates protein synthesis, and regulates fetal growth and development. The mechanisms involved in the regulation of placental leptin expression are not fully understood. Previous results from our lab demonstrated that estradiol (E2) regulates leptin expression involving genomic and non-genomic effects. Sp1 is a ubiquitous transcription factor. Several potential binding sites for Sp1 have been found in the leptin promoter. One of them is present in the region identified as PLE (enhancer of the placental leptin) involved in E2 effect. In the present study, we analyzed the effect of Sp1 in the induction of leptin expression by E2 in human placental cells.
quantified. Obese and control females were paired with control CD1 males. We evaluated pregnancy the rate and length and litter size. After delivery, pups were weighed during the lactation period. Additionally, LPS (0.05 mg/g) was administered to 15-day pregnant mice and delivery was recorded. Results: Weight-gain was higher in HFD-fed animals than in control and in CB1KO than in WT mice. We found no differences in the daily intake of kilocalories. Abdominal adipose tissue weight increased in HFD-fed animals, in both WT and CB1KO mice. Additionally, we found an increase in serum cholesterol levels in animals fed with HFD, with higher levels in CB1KO than in WT mice. No differences in the pregnancy rate and length or in litter size were found. However, pups born to obese mothers had higher weight gain during the lactation period. Almost 75% of LPS-treated WT mothers presented a defective labor in contrast to 22% in the case of CB1KO mice. Interestingly, when exposed to LPS, up to 50% of the HFD-fed CB1KO mice presented a defective labor. Conclusions: Our results suggest that ECS is involved in the development of maternal obesity, which then has consequences in pups. In addition, obesity sensitized mothers to endotoxin treatment.
PA.10. EARLY PLACENTAL ANGIOGENESIS-VASCULARIZATION AND VEGF/KDR RECEPTOR EXPRESSION DURING MOUSE ORGANOGENESIS AFTER PERIGESTATIONAL ALCOHOL CONSUMPTION M.R. Ventureira 1, C.M.A. Sobarzo 2, M. Naito 3, C. Zanuzzi 4, C. Barbeito 4, E. Reproduction and Maternal-Embryonic Cebral 1. 1 Laboratory of Physiopathology, IFIBYNE-UBA/CONICET Biodiversity and Experimental Biology Department, FCEN, University of Buenos Aires, Buenos Aires, Argentina; 2 INBIOMED-UBA/CONICET- School of Medicine, University of Buenos Aires, Buenos Aires, Argentina; 3 Department of Anatomy, Aichi Medical University, Yazakokarimata, Nagakute, Japan; 4 Histology and Embriology, Basic Sciences Department, Faculty of Veterinary Science, National University of La Plata (UNLP), Buenos Aires, Argentina The normal growth of the fetus depends on adequate placental development, in which the expression of trophoblast-decidual VEGF (vascular endothelial growth factor) conducts angiogenesis-vascularization. Previously, we found that perigestational alcohol consumption at mouse organogenesis (day 10 of gestation), causes embryo resorption, reduces decidualization and leads to histomorphological alterations of the decidual tissue. Objective: The aim was to study, after periconceptional alcohol intake, vascularization of early placentation by monitoring the number of uNK, VEGF and KDR-receptor expressions as well as KDR-activation, in murine decidua and labyrinth during organogenesis. Methods: Ethanol 10% was administered in the drinking water to CF-1 murine females (TF) for 17 days before and up to day 10 of gestation; water was administered to control females (CF). Labyrinthine development and decidual (De) vascularization (H-E, histochemistry for BSI-lectin, morphometry of decidual vascular area and labyrinth), the presence and number of uNKs (PAS and DBA-lectin) and the expression of VEGF and KDR (total and phosphorylated form) by immunohistochemistry, immunofluorescence-confocal and western blot (WB) were analyzed in the implantation sites (IS). Results: Increased percentage of TF-IS had poor labyrinthine growth and reduced De-vascular lumen (p<0.05), while significantly fewer uNKs (PASand DBA-positive cells) were found (p<0.01, vs CF). In TF, the decreased VEGF immunoexpression in trophoblast tissue, endothelial and decidual cells was coincident with its reduced VEGF expression level (p<0.001, vs CF). The immunostaining for KDR (expressed in uNK-PAS) was reduced in the TF-vascular area of decidua (p<0.05), consistent with lower KDR expression level (WB) (p<0.05 vs CF). However, the expression of phosphorylated KDR significantly increased in TF vs CF (p<0.01). Conclusions: Perigestational alcohol exposure inhibits early trophoblastdecidual vascularization by deregulating the VEGF/KDR system, suggesting that, in this mouse model, maternal alcohol intake can lead to vascular resistance and fetal-placental growth restriction at term.
491
PA.11. IMMUNE HOMEOSTASIS DURING THE DECIDUALIZATION PROGRAM: PARTICIPATION OF VIP AND DENDRITIC CELLS Esteban Grasso 1, Soledad Gori 2, Daniel Paparini 1, Lucila rez Gallino 1, Elizabeth Soczewski 1, Gabriela Salamone 2, Claudia Pe s 1, Rosanna Ramhorst 1. 1 Immunopharmacology Laboratory, School of Leiro Sciences, University of Buenos Aires, IQUIBICEN-CONICET, Argentina; 2 Immunology Laboratory, Institute of Experimental Medicine, National Academy of Medicine Buenos Aires, Argentina The decidualization program involves phenotype and functional changes in endometrial cells such as the production of different mediators that will facilitate the attachment and invasion of the blastocyst. Particularly, the vasoactive intestinal peptide (VIP) is a neuropeptide produced by endometrial stromal cells among others, which displays multiple target circuits that allow immunotolerance. Objective: We investigated VIP contribution to the decidualization program and whether this process modulates the profile of dendritic cells (DC). Methods: A human endometrial stromal cell line (hESC) was evaluated. These cells were differentiated in the presence of VIP (10-7M) or medroxyprogesterone (MPA) and dbcAMP (Positive control) for 8 days. Phenotypic markers of decidualization, VIP and their receptors were quantified by RT-PCR. DC were obtained from monocytes isolated from PBMCs from fertile women and differentiated with IL-4 and GM-CSF. VIP and IL-10 secretion were quantified by ELISA. Results: When decidualization was induced by VIP, we observed an increase in characteristic markers such as IGFBP1, PRL and the KLF13/ KLF9 ratio, IL-8 and SDF1 in a concentration-dependent manner (p<0.05). To evaluate the functional aspect of VIP-induced decidualization, an invasion assay was performed using blastocyst-like spheroids (BLS) from first trimester trophoblast cells (Swan71) cultured on hESC monolayer. BLS were able to attach and invade the hESC monolayer decidualized with VIP or MPA+dbcAMP (invasion index X±SE 0.55±0.01 0.64±0.01 0.66±0.01). When these assays were performed with conditioned media obtained from human blastocysts, we found an increase in BLS invasion to hESC decidualized with VIP (p<0.05). Finally, decidualized hESC cells increased VIP expression and secretion. Therefore we evaluated the immunomodulatory effect on DC. Decidualized hESC conditioned media induced a semi-mature profile on DC preventing the induction of HLA-DR, CD83 and CD86 expression, while inducing IL-10 secretion (p<0.05). Conclusions: Our results suggest that VIP may contribute to the decidualization process by inducing phenotypic and functional markers on hESC cells and to the maintenance of a tolerogenic DC-profile.
PA.12. EFFECT OF MODERATE INTENSITY EXERCISE ON THE REPRODUCTIVE PERFORMANCE OF DIABETIC RATS N.C.D. Macedo, A.O. Netto, F.Q. Gallego, I.M.P. Calderon, M.V.C. Rudge, G.T. Volpato, D.C. Damasceno. Laboratory of Experimental Research on Gynecology and Obstetrics, Graduate Program on Gynecology, Obstetrics and Mastology, Botucatu Medical School, Univ. Estadual Paulista_Unesp, ~o Paulo, Brazil Botucatu, Sa Background: Maternal diabetes leads to impaired reproductive outcomes. Although physical exercise is relevant to control glycemia, there are concerns related to the type and intensity of exercise indicated during pregnancy. We hypothesized that an appropriate intensity of exercise improves maternal intrauterine environment leading to favorable embryofetal development. Objective: To evaluate the effects of moderate intensity exercise on the maternal reproductive outcomes of mild diabetic rats.
492
Abstracts / Placenta 36 (2015) 469e521
Methods: Rats were randomly assigned into four groups (n¼12/group): Sedentary non-diabetic (Control¼C, glycemia<120 mg/dL); Sedentary mild diabetic (MD - Streptozotocin - 100 mg/kg b.w., sc., at birth); Exercised non-diabetic (Cex) and Exercised mild diabetic (MDex). Cex and MDex rats were subjected to a swimming program supporting a load of 4% of body weight from day 7 to 18.5 of pregnancy. On day 18.5 of pregnancy, the different experimental groups were killed for exposure of the uterine corns for the analysis of the reproductive outcomes. P<0.05 was considered as a significant statistical limit. Results: MD rats presented glucose intolerance and the area under the curve analysis showed a significant negative correlation between maternal glycemia and a decrease in the number of live fetuses. MD rats showed a lower number of live fetuses and litter weight than C rats. MDex dams presented reduced maternal weight gain, implantation number and litter weight, and increased number of live fetuses as compared with Cex. Both MD groups presented increased fetal and placental weights, and percentage of fetuses classified as large for pregnancy age in relation to control groups. Conclusion: The mild diabetic rats subjected to the swimming program presented no alterations in glucose intolerance, but the number of live fetuses and fetal weight of these rats were consistent with those in the control group, confirming beneficial effects of swimming with an appropriate intensity for the parameters analyzed. Acknowledgement: FAPESP/Brazil (Process number: 2013/23478-3).
PA.13. MATERNAL SATURATED FAT-ENRICHED DIET DETERMINES LEPTIN RESISTANCE IN THE FETAL LIVER AND PROGRAMS LIPID HOMEOSTASIS IMPAIRMENT IN THE OFFSPRING n Mazzucco, Daiana Fornes, Alicia Jawerbaum, Vero nica María Bele White. Centre from Pharmacology and Botanical Studies (CEFyBO) CONICET UBA, Buenos Aires, Argentina Maternal obesity leads to impaired metabolic programming. In a rat model of saturated fat overload in the maternal diet, we have previously found increased fetal liver lipid content. Lipid catabolism in the liver assures proper lipid homeostasis. Leptin regulates lipid catabolism in several tissues. Objective: To investigate whether an overload of saturated fat on the maternal diet impairs leptin effects on hepatic lipid catabolism in rat fetuses at term gestation and whether these anomalies are sustained in the offspring of these rats. Methods: Female Wistar rats were fed with a standard (5% fat) or with a saturated fat diet (28% fat) from 6 weeks of age (SFD rats). After 8 weeks of diet, they were mated with control males. Control and SFD rats were euthanized at 21 days of gestation and fetal livers obtained for further culture of explants (3h) with or without leptin (100 ng/ml). Another group of control and SFD rats were allowed to deliver, and their offspring euthanized at 21 or 130 days of age. Livers were analyzed for lipid accumulation (TLC) and acylCoA oxidase (ACO) expression (PCR). Results: Leptin induced a decrease in triglycerides (49%), free fatty acids (75%) and cholesteryl esters (42%) concentrations in fetal livers from control rats (p<0.05), but no changes but no changes in fetal livers from SFD rats, which showed. Leptin induced an increased ACO expression (50%, p<0.05) in control fetal livers. Fetal livers from SFD rats showed a decrease in ACO expression (25% p<0.05) and no changes with leptin additions. A decrease in ACO expression were also observed in the livers of 21- and 130day-old offspring from SFD rats (20%, p<0.05). Conclusions: Saturated fat in maternal diet induces hepatic lipid metabolism impairment in fetuses and offspring. Liver leptin resistance on lipid catabolism begins in utero and maybe responsible for the alterations on the offspring lipid metabolism.
PA.14. METABOLIC ABNORMALITIES IN THE FETAL LIVER OF RATS WITH GESTATIONAL DIABETES INDUCED BY INTRAUTERINE PROGRAMMING Daiana Fornes, Romina Higa, Ivana Linenberg, Evangelina Capobianco, Alicia Jawerbaum. Laboratory of Reproduction and Metabolism, CEFYBO-CONICET, School of Medicine, University of Buenos Aires, Buenos Aires, Argentina Gestational diabetes (GDM) is a prevalent disease with adverse consequences to both mothers and offspring. We have recently described a new GDM model obtained through adverse intrauterine programming in the offspring of diabetic rats. This model allows analyzing the regulation of lipid metabolism in the fetal liver of GDM mothers. Peroxisome proliferator activated receptors (PPARs) are nuclear receptors involved in lipid metabolic pathways, being PPARa a major regulator of lipid catabolism, while PPARg is highly involved in lipid synthesis and deposition in different tissues. Objective: To evaluate whether lipid concentrations are altered and related to impaired PPAR protein expression in the fetal liver of GDM rats. Methods: Control and GDM rats (obtained from intrauterine programming (F1) in the offspring of streptozotocin-induced diabetic rats (F0)) were evaluated on day 21 of pregnancy. Glucose and insulin concentrations were measured in maternal and fetal serum. Lipid concentrations were measured by TLC and PPARa and PPARg protein expression were evaluated through immunohistochemistry. Results: GDM rats showed increased glycemia and insulinemia only in pregnancy (p<0.01). Male and female fetuses had increased glycemia and insulinemia (p<0.01). Male fetuses from GDM rats showed increased triglycerides (33%) and cholesterol (30%) concentrations in the liver when compared to controls (p<0.05). In these GDM fetal tissues, PPARg protein expression was increased (p<0.001) whereas PPARa protein expression showed no changes compared to controls. Differently, female fetuses from GDM rats showed reduced triglycerides (34%), cholesterol (26%), phospholipids (29%) and free fatty acids (25%) in the liver, p<0.02, compared to controls. Also, in these GDM fetal tissues, both PPARg and PPARa levels were reduced (p<0.01). Conclusion: In a GDM model induced by intrauterine programming, the altered lipid deposition in the fetal liver is gender-dependent and possibly related to changes in the concentrations of the different PPAR isotypes in this fetal tissue.
PA.15. SP1 TRANSCRIPTION FACTOR IS INVOLVED IN THE REGULATION OF PLACENTAL LEPTIN EXPRESSION n Toro 1, Julieta Maymo 1, A. Pe rez-Pe rez 2, Ye sica Malena Schanton 1, Ayele nchez Margalet 2, Cecilia Gambino 1, Bernardo Maskin 3, Victor Sa gica, FCEN, UBA, IQUIBICEN, Varone 1. 1 Departamento de Química Biolo edica y CONICET, Buenos Aires, Argentina; 2 Departamento de Bioquímica M Biología Molecular, Universidad de Sevilla, Sevilla, Spain; 3 Hospital Nacional Profesor Alejandro Posadas, Buenos Aires, Argentina Objectives: Leptin is a key hormone in placental physiology. It regulates trophoblast proliferation, inhibits apoptosis, stimulates protein synthesis, and regulates fetal growth and development. The mechanisms involved in the regulation of placental leptin expression are not fully understood. Previous results from our lab demonstrated that estradiol (E2) regulates leptin expression involving genomic and non-genomic effects. Sp1 is a ubiquitous transcription factor. Several potential binding sites for Sp1 have been found in the leptin promoter. One of them is present in the region identified as PLE (enhancer of the placental leptin) involved in E2 effect. In the present study, we analyzed the effect of Sp1 in the induction of leptin expression by E2 in human placental cells.