Publications by authors named "Yallampalli Chandra"

66 Publications

Progesterone receptor isoform B regulates the -- pathway to suppress uterine contractility.

Proc Natl Acad Sci U S A 2021 Mar;118(11)

Reproductive and Developmental Biology Laboratory, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709;

Uterine contractile dysfunction leads to pregnancy complications such as preterm birth and labor dystocia. In humans, it is hypothesized that progesterone receptor isoform PGR-B promotes a relaxed state of the myometrium, and PGR-A facilitates uterine contraction. This hypothesis was tested in vivo using transgenic mouse models that overexpress PGR-A or PGR-B in smooth muscle cells. Elevated PGR-B abundance results in a marked increase in gestational length compared to control mice (21.1 versus 19.1 d respectively, < 0.05). In both ex vivo and in vivo experiments, PGR-B overexpression leads to prolonged labor, a significant decrease in uterine contractility, and a high incidence of labor dystocia. Conversely, PGR-A overexpression leads to an increase in uterine contractility without a change in gestational length. Uterine RNA sequencing at midpregnancy identified 1,174 isoform-specific downstream targets and 424 genes that are commonly regulated by both PGR isoforms. Gene signature analyses further reveal PGR-B for muscle relaxation and PGR-A being proinflammatory. Elevated PGR-B abundance reduces and and increases expression, which manifests a genetic profile of compromised oxytocin signaling. Functionally, both endogenous PLCL2 and its paralog PLCL1 can attenuate uterine muscle cell contraction in a CRISPRa-based assay system. These findings provide in vivo support that PGR isoform levels determine distinct transcriptomic landscapes and pathways in myometrial function and labor, which may help further the understanding of abnormal uterine function in the clinical setting.
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http://dx.doi.org/10.1073/pnas.2011643118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7980420PMC
March 2021

Soluble fms-like tyrosine kinase-1 and angiotensin2 target calcitonin gene-related peptide family peptides in maternal vascular smooth muscle cells in pregnancy†.

Biol Reprod 2021 May;104(5):1071-1083

Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, TX, USA.

Calcitonin gene-related peptide (CALCB), adrenomedullin (ADM), and adrenomedullin2 (ADM2) are hypotensive peptides that belong to CALCB family of peptides. Goal of this study was to identify the effect of fms-like tyrosine kinase (sFLT-1) and angiotensin2 (Ang2) on the function of these peptides in OA smooth muscle cells (OASMC) and assess the sensitivity of OA for these peptides in preeclampsia (PE) and normotensive pregnancy.

Methods: Peptide function was assessed by Cyclic adenosine monophosphate (cAMP) assays and wire myograph; mRNA expression by Polymerase chain reaction (PCR) and protein-protein interaction by proximity ligation assay and co-immunoprecipitation.

Findings: All three peptides increased cAMP synthesis in the order of efficacy CALCB > ADM = ADM2 and vascular endothelial growth factor (VEGF) mRNA in OASMC (P < 0.05); sFLT-1 mediated decrease in cAMP synthesis (P < 0.05) is differentially rescued by all three CALCB family peptides in OASMC (P < 0.005); sFLT-1 decreased receptor activity-modifying protein (RAMP)1 and RAMP2 mRNA expression (P < 0.05); Ang2 decreased the expression of calcitonin-receptor-like receptor and RAMP1 mRNA and desensitized CALCB and ADM2 receptors in OASMC (P < 0.05); sFLT-1 increased RAMP1and Ang2 type 1 receptor (AT1R) interaction in OASMC which is inhibited in presence of all three peptides; and all three peptides relax OA in PE with enhanced ADM2 response (P < 0.05).

Conclusion: sFLT-1 and Ang2 impair OASMC mediated functional responses of CALCB family peptides which can be inhibited by respective peptide treatment. The sensitivity of OA for CALCB, ADM, and ADM2-mediated relaxation is retained in PE.
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http://dx.doi.org/10.1093/biolre/ioab026DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8111240PMC
May 2021

Modulation of Autophagy Through Regulation of 5'-AMP-Activated Protein Kinase Affects Mitophagy and Mitochondrial Function in Primary Human Trophoblasts.

Reprod Sci 2021 Aug 22;28(8):2314-2322. Epub 2021 Feb 22.

Department of Obstetrics and Gynecology, Beijing Chaoyang Hospital, Capital Medical University, Beijing, 100020, China.

The placenta is important for pregnancy maintenance, and autophagy is documented to be essential for placental development. Autophagy is responsible for degrading and recycling cellular misfolded proteins and damaged organelles. Mitophagy is a selective type of autophagy, where the autophagic machinery engulfs the damaged mitochondria for degradation, and there is reciprocal crosstalk between autophagy and mitochondria. Within these processes, 5'-AMP-activated protein kinase (AMPK) plays an important role. However, the role of AMPK regulation in both autophagy and mitochondria in primary human trophoblasts is unknown. In this study, we address this question by investigating changes in mRNA expression and the abundance of autophagy- and mitochondria-related proteins in isolated human trophoblasts after treatment with AMPK agonists and antagonists. We found that compared to the control group, autophagy was slightly suppressed in the AMPK agonist group and significantly enhanced autophagy in the AMPK antagonist group. However, the expressions of genes related to autophagosome-lysosome fusion were reduced, while genes related to lysosomal function were unchanged in both groups. Furthermore, mitophagy and mitochondrial fusion/fission were both impaired in the AMPK agonist and antagonist groups. Although mitochondrial biogenesis was enhanced in both groups, the function of mitochondrial fatty acid oxidation was increased in the AMPK agonist group but decreased in the AMPK antagonist group. Overall, our study demonstrates that AMPK regulation negatively modulates autophagy and consequently affects mitophagy, mitochondrial fusion/fission, and function in primary human trophoblasts.
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http://dx.doi.org/10.1007/s43032-021-00495-5DOI Listing
August 2021

In utero low-protein-diet-programmed type 2 diabetes in adult offspring is mediated by sex hormones in rats†.

Biol Reprod 2020 10;103(5):1110-1120

Basic Sciences Perinatology Research Laboratories, Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, Texas, USA.

Sex steroids regulate insulin sensitivity and glucose metabolism. We had characterized a lean type 2 diabetes (T2D) rat model using gestational low-protein (LP) diet programming. Our objective was to identify if endocrine dysfunction leading to decreased sex hormone levels will precede the development of T2D and if steroid replacement will prevent the onset of the disease. Pregnant rats were fed control or isocaloric LP diet from gestational day 4 until delivery. Normal diet was given to all mothers after delivery and to pups after weaning. LP offspring developed glucose intolerance and insulin resistance at 4 months. We measured sex steroid hormone profiles and expression of key genes involved in steroidogenesis in testis and ovary. Furthermore, one-month old rats were implanted with 90-day slow release T and E2 pellets for males and females, respectively. Glucose tolerance test (GTT) and euglycemic hyperinsulinemic clamp was performed at 4 months. LP-programmed T2D males had low T levels and females had low E2 levels due to dysregulated gene expression during steroidogenesis in gonads. GTT and euglycemic hyperinsulinemic clamp showed that LP males and females were glucose intolerant and insulin resistant; however, steroid supplementation prevented the onset of glucose intolerance and insulin resistance. Rats that developed T2D by LP programming have compromised gonadal steroidogenesis leading to low T and E2 in males and females, respectively. Sex steroid supplementation prevented the onset of glucose intolerance and insulin resistance indicating low sex steroid levels could cause compromised glucose metabolism ultimately leading to T2D.
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http://dx.doi.org/10.1093/biolre/ioaa133DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7609843PMC
October 2020

Complement inhibitor Crry expression in mouse placenta is essential for maintaining normal blood pressure and fetal growth.

PLoS One 2020 3;15(8):e0236968. Epub 2020 Aug 3.

Department of Obstetrics & Gynecology, Baylor College of Medicine, Houston, Texas, United States of America.

Many circumstantial evidences from human and animal studies suggest that complement cascade dysregulation may play an important role in pregnancy associated complications including preeclampsia. Deletion of rodent specific complement inhibitor gene, Complement Receptor 1-related Gene/Protein y (Crry) produces embryonic lethal phenotype due to complement activation. It is not clear if decreased expression of Crry during pregnancy produces hypertensive phenotype. We downregulated Crry in placenta by injecting inducible lentivialshRNA vectors into uterine horn of pregnant C57BL/6 mice at the time of blastocyst hatching. Placenta specific downregulation of Crry without significant loss of embryos was achieved upon induction of shRNA using an optimal doxycycline dose at mid gestation. Crry downregulation resulted in placental complement deposition. Late-gestation measurements showed that fetal weights were reduced and blood pressure increased in pregnant mice upon downregulation of Crry suggesting a critical role for Crry in fetal growth and blood pressure regulation.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0236968PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7398533PMC
October 2020

Fetal macrosomia in a Hispanic/Latinx predominant cohort and altered expressions of genes related to placental lipid transport and metabolism.

Int J Obes (Lond) 2020 08 3;44(8):1743-1752. Epub 2020 Jun 3.

Department of Obstetrics and Gynecology, Howard University College of Medicine, Washington, DC, 20059, USA.

Introduction: Fetal overgrowth, termed fetal macrosomia when birth weight is >4000 g, is the major concern in the treatment of gestational diabetes mellitus (GDM). However, to date, the underlying mechanisms of fetal macrosomia have not been understood completely. Placental lipid metabolism is emerging as a critical player in fetal growth. In this study, we hypothesized that fatty-acid transport and metabolism in the placental tissue is impaired in GDM women, dependent on fetal sex.

Methods: To test this hypothesis, we analyzed the incidence of GDM, fetal macrosomia, and obesity in a large cohort consisting of 17,995 pregnant subjects and majority of subjects being Hispanic/Latinx, and investigated expression of genes related to lipid transport and metabolism in placentas from obese women with or without GDM, and with or without fetal macrosomia.

Results: The main findings include: (1) there was a higher incidence of GDM and obesity in Hispanic subjects compared with non-Hispanic subjects, but not fetal macrosomia; (2) expressions of most of genes related to placental lipid transport and metabolism were not altered by the presence of GDM, fetal macrosomia, or fetal sex; (3) expression of FABP4 was increased in obese women with GDM and fetal macrosomia, and this occurred in male placentas; (4) expression of LPL was decreased in obese women with GDM despite fetal macrosomia, and this occurred in male placentas; (5) expression of ANGPTL3 was decreased in obese women with GDM and fetal macrosomia, but was not altered when fetal sex was included in the analysis.

Conclusions: This study indicates that there is race disparity in GDM with higher incidence of GDM in obese Hispanic women, although fetal macrosomia disparity is not present. Moreover, altered placental lipid transport may contribute to fetal overgrowth in obese women with GDM.
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http://dx.doi.org/10.1038/s41366-020-0610-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7387181PMC
August 2020

Common variants of fetal and maternal complement genes in preeclampsia: pregnancy specific complotype.

Sci Rep 2020 03 16;10(1):4811. Epub 2020 Mar 16.

Department of Obstetrics & Gynecology, Baylor College of Medicine, Houston, Texas, USA.

Preeclampsia (PE) is a pregnancy specific hypertensive disorder. If untreated PE leads to life threatening condition, eclampsia. Systemic complement activation levels are increased during pregnancy compared to non-pregnant women of childbearing age. In PE, systemic complement levels are further increased, and higher complement deposition has been observed on placentas. We hypothesize that combinations of common SNPs in maternal and fetal complement genes constitute pregnancy specific complotypes and predispose women to PE. In this study, we sequenced two maternal (factor H and C3) and one fetal (CD46) complement genes and identified a total of 9 common SNPs. Minor allele frequencies of two fetal CD46 SNPs were significantly higher in PE. Further, complotypes consisting of fetal CD46 variants and maternal CFH/C3 variants were highly prevalent in PE patients compared to normotensive pregnancies. Placental complement deposition and maternal alternative pathway 50 (AP50) values were higher in PE pregnancies. Irrespective of disease status, two CD46 variants were associated with reduced placental CD46 expression and one CFH variant was associated with increased maternal AP50 values.
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http://dx.doi.org/10.1038/s41598-020-60539-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7076030PMC
March 2020

Sex Dependent Dysregulation of Hepatic Glucose Production in Lean Type 2 Diabetic Rats.

Front Endocrinol (Lausanne) 2019 6;10:538. Epub 2019 Aug 6.

Basic Sciences Perinatology Research Laboratories, Department of Obstetrics and Gynecology, Houston, TX, United States.

We have characterized a lean type 2 diabetic rat model by gestational low protein programming. We aimed to identify if the regulation of hepatic glucose production (HGP) via gluconeogenesis and glycogenolysis is affected and if there are any sex differences. Fasting (6-7 months old) type 2 diabetic rats received HO followed by a primed constant rate infusion of [6,6-H] glucose. Blood samples were drawn during steady states after 4 h of fasting and following a euglycemic hyperinsulinemic clamp. HGP and the fraction of glucose derived from gluconeogenesis under fasting and euglycemic states were measured from steady state glucose enrichments after the infusion of [6,6-H]glucose and HO tracers. Glycogenolysis was determined by calculating the difference between total HGP and gluconeogenesis rates. Hepatic gene expression of enzymes involved in HGP were quantified using qPCR. HGP rates was similar during fasting in both groups and sexes. However, under simulated fed condition, HGP rate was suppressed in controls but not in type 2 diabetic rats. They also showed inefficient HGP suppression in a simulated fed state. Differential analysis showed that suppression of both gluconeogenesis and glycogenolysis under simulated fed state was affected in these low protein programmed type 2 diabetic rats. These effects were greater in females when compared to males. Further, key genes involved in these processes like G6Pase, Pepck, pyruvate carboxylase, and glycogen phosphorylase in liver were dysregulated. Our data shows impaired suppression of HGP via gluconeogenesis and glycogenolysis in type 2 diabetic rats with greater effects on females.
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http://dx.doi.org/10.3389/fendo.2019.00538DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6691354PMC
August 2019

Impact of adrenomedullin blockage on lipid metabolism in female mice exposed to high-fat diet.

Endocrine 2019 08 25;65(2):278-285. Epub 2019 Apr 25.

Department of Obstetrics and Gynecology, Baylor College of Medicine/Texas Children's Hospital, Houston, TX, 77030, USA.

Purpose: Adrenomedullin (ADM) levels are elevated in gestational and type 2 diabetic patients. ADM also stimulates lipolysis in vitro. Disturbed lipid metabolism has been implicated in the pathogenesis of diabetes. Here, we explore whether blockade of ADM is beneficial for metabolic homeostasis in a diabetic mouse model.

Methods: C57BL/6J female mice were placed on either a control or a high fat high sucrose (HFHS) diet for 8 weeks. At week 4, osmotic mini-pumps were implanted for constant infusion of either saline or ADM antagonist, ADM. Glucose tolerance tests were performed prior to infusion and 4 weeks after infusion began. Animals were then sacrificed and visceral adipose tissue collected for further analysis.

Results: Mice fed HFHS displayed glucose intolerance, increased mRNA expressions in VAT for Adm and its receptor components, Crlr. HFHS fed mice also had increased basal and isoprenaline-induced glycerol release by VAT explants. ADM did not significantly affect glucose intolerance. ADM did suppress basal and isoprenaline-induced glycerol release by VAT explants. This alteration was associated with enhanced mRNA expression of insulin signaling factors Insr and Glut4, and adipogenic factor Pck1.

Conclusions: HFHS diet induces glucose intolerance and enhances ADM and its receptor expressions in VAT in female mice. ADM treatment did not affect glucose intolerance in HFHS mice, but reduced both basal and isoprenaline-induced lipolysis, which is associated with enhanced expression of genes involved in adipogenesis. These results warrant further research on the effects of ADM blockade in improving lipid homeostasis in diabetic patients.
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http://dx.doi.org/10.1007/s12020-019-01927-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6901288PMC
August 2019

Preovulatory exposure to a protein-restricted diet disrupts amino acid kinetics and alters mitochondrial structure and function in the rat oocyte and is partially rescued by folic acid.

Reprod Biol Endocrinol 2019 Jan 17;17(1):12. Epub 2019 Jan 17.

Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, TX, USA.

Background: Detrimental exposures during pregnancy have been implicated in programming offspring to develop permanent changes in physiology and metabolism, increasing the risk for developing diseases in adulthood such as hypertension, diabetes, heart disease and obesity. This study investigated the effects of protein restriction on the metabolism of amino acids within the oocyte, liver, and whole organism in a rat model as well as effects on mitochondrial ultrastructure and function in the cumulus oocyte complex.

Methods: Wistar outbred female rats 8-11 weeks of age (n = 24) were assigned to three isocaloric dietary groups, including control (C), low protein (LP) and low protein supplemented with folate (LPF). Animals were superovulated and 48 h later underwent central catheterization. Isotopic tracers of 1-C-5CH-methionine, H-cysteine, U-C-cysteine and U-C-serine were administered by a 4 h prime-constant rate infusion. After sacrifice, oocytes were denuded of cumulus cells and liver specimens were obtained.

Results: Oocytes demonstrated reduced serine flux in LP vs. LPF (p < 0.05), reduced cysteine flux in LP and LPF vs. C (p < 0.05), and a trend toward reduced transsulfuration in LP vs. C and LPF. Folic acid supplementation reversed observed effects on serine flux and transsulfuration. Preovulatory protein restriction increased whole-body methionine transmethylation, methionine transsulfuration and the flux of serine in LP and LPF vs. C (p = 0.003, p = 0.002, p = 0.005). The concentration of glutathione was increased in erythrocytes and liver in LP and LPF vs. C (p = 0.003 and p = 0.0003). Oocyte mitochondrial ultrastructure in LP and LPF had increased proportions of abnormal mitochondria vs. C (p < 0.01 and p < 0.05). Cumulus cell mitochondrial ultrastructure in LP and LPF groups had increased proportions of abnormal mitochondria vs. C (p < 0.001 and p < 0.05). Preovulatory protein restriction altered oocyte expression of Drp1, Opa-1, Mfn1/2, Parl and Ndufb6 (p < 0.05) and Hk2 (p < 0.01), which are genes involved in mitochondrial fission (division) and fusion, mitochondrial apoptotic mechanisms, respiratory electron transport and glucose metabolism.

Conclusions: Preovulatory protein restriction resulted in altered amino acid metabolism, abnormal cumulus oocyte complex mitochondrial ultrastructure and differential oocyte expression of genes related to mitochondrial biogenesis.
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http://dx.doi.org/10.1186/s12958-019-0458-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6337842PMC
January 2019

Circulating Adrenomedullin Is Elevated in Gestational Diabetes and Its Role in Impaired Insulin Production by β-Cells.

J Clin Endocrinol Metab 2019 03;104(3):697-706

Department of Obstetrics and Gynecology, Baylor College of Medicine/Texas Children's Hospital, Houston, Texas.

Context: Defective pancreatic β-cell adaptation in pregnancy plays an important role in the pathophysiology of gestational diabetes mellitus (GDM), but the molecular basis remains unclear. Objectives of this study were to determine if circulating levels of adrenomedullin (ADM) in women with GDM are elevated and to assess the effects of ADM on insulin synthesis and secretion by human pancreatic β-cells.

Design: A stable gene product of ADM precursor, midregional pro-adrenomedullin (MR-proADM), was measured in plasma of pregnant women with normal glucose tolerance (NGT, n = 10) or GDM (n = 11). The β-Lox5 cell line, derived from human pancreatic β-cells, was transduced with homeodomain transcription factor pancreatic-duodenal homeobox (PDX) factor 1 (PDX1) encoding lentiviral vector and treated with different doses of ADM. mRNA for insulin, ADM, and its receptor components in β-Lox5 cells and insulin in media were measured.

Results: Plasma MR-proADM levels were significantly higher in GDM compared with patients with NGT. Pancreatic β-Lox5 cells express mRNA for insulin, ADM, and its receptor components. PDX1 transduction and cell-cell contact synergistically promote β-Lox5 cells insulin mRNA and secretion. Furthermore, ADM dose-dependently inhibited mRNA and secretion of insulin in β-Lox5 cell aggregates. These inhibitory effects were blocked by ADM antagonist ADM22-52, cAMP-dependent protein kinase A inhibitor KT5720, and Erk inhibitor PD98059, but not by PI-3K the inhibitor wortmannin.

Conclusions: Circulating ADM concentrations were elevated in pregnant women with GDM. ADM suppresses insulin synthesis and secretion by pancreatic β-cells in vitro. Thus, increased circulating ADM may contribute to the defective adaptation of β-cells in diabetic pregnancies, and blockade of ADM actions with its antagonists may improve β-cell functions.
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http://dx.doi.org/10.1210/jc.2018-01119DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6338207PMC
March 2019

Upregulation and release of soluble fms-like tyrosine kinase receptor 1 mediated by complement activation in human syncytiotrophoblast cells.

Am J Reprod Immunol 2018 11 12;80(5):e13033. Epub 2018 Aug 12.

Basic Sciences Perinatology Research Laboratories, Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, Texas.

Problem: Antiangiogenic molecule soluble fms-like tyrosine kinase receptor 1 (sFLT1) released from trophoblast cells is associated with pregnancy-specific hypertensive disorder pre-eclampsia. Cause of elevated sFLT1 in pre-eclampsia patients is not well understood. Despite evidence of excess systemic and placental complement activation in pre-eclampsia patients, its role in pathophysiology is not clear. If the complement activation plays a role in upregulation and secretion of sFLT1 is not known.

Method Of Study: Human trophoblast cells were isolated from term placentas and allowed to syncytialize. Complement was activated in vitro at sublethal levels on syncytiotrophoblast cells. Effect of complement activation on expression and release of sFLT1 was assessed by comparing its levels in these cells with and without complement activation.

Results: Sublethal level of complement activation on syncytialized human trophoblast cells induced upregulation of sFLT1 mRNA and protein. Complement also induced secretion of sFLT1 in a manner depending on degree of activation. Anaphylatoxins C3a induced upregulation but not the release of sFLT1. Release of terminal membrane attack complex (MAC) was associated with sFLT1 secretion.

Conclusion: Complement activation plays a major role in both the expression and secretion of sFLT1 from syncytial trophoblast cells. The terminal MAC complex is involved in its secretion. Increased levels of sFLT1 in pre-eclampsia patients may be due to complement-induced upregulation and secretion.
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http://dx.doi.org/10.1111/aji.13033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6202180PMC
November 2018

Adipose Tissue Inflammation and Adrenomedullin Overexpression Contribute to Lipid Dysregulation in Diabetic Pregnancies.

J Clin Endocrinol Metab 2018 10;103(10):3810-3818

Department of Obstetrics and Gynecology, Baylor College of Medicine/Texas Children's Hospital, Houston, Texas.

Context: Impaired maternal lipid metabolism in gestational diabetes mellitus (GDM) has detrimental effects on maternal health and fetal growth. We previously reported the excessive expression of adrenomedullin (ADM) and its receptors in GDM adipose tissues compared with normal glucose-tolerant pregnancies. In the present study, we determined the mechanisms underlying enhanced expression of ADM and its receptors.

Design: Omental adipose tissue (OAT) samples were collected from women during cesarian section of term pregnancy with nonoverweight (NOW; n = 9), overweight (OW; n = 8), obese (OBS; n = 10), and GDM (n = 10) status.

Results: The expression of ADM and its receptors was greater in OATs from GDM than from women who were NOW, OW, and OBS. The expression of adipokines, leptin, and resistin were significantly increased, but adiponectin was decreased in OATs from patients with GDM compared with those without GDM. Macrophage infiltration and TNF-α expression were greater in OAT from pregnant women with GDM than in pregnant women without GDM. Furthermore, TNF-α dose dependently increased mRNA for ADM and its receptor components calcitonin receptor-like receptor and receptor activity-modifying proteins 2 and 3 in OAT explants from women who were NOW. Human adipocytes treated with ADM significantly increased glycerol release in culture medium, and the increases of glycerol in culture medium of OAT from women with GDM were attenuated by ADM antagonists, ADM22-52.

Conclusions: Increased macrophage infiltration and TNF-α expression in adipose tissue from GDM, but not from OBS, tissues stimulate ADM and its receptor overexpression, leading to enhanced lipolysis and hyperlipidemia. This might contribute to fetal macrosomia and adiposity in diabetic pregnancies.
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http://dx.doi.org/10.1210/jc.2018-00905DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6456971PMC
October 2018

Folate treatment partially reverses gestational low-protein diet-induced glucose intolerance and the magnitude of reversal is age and sex dependent.

Nutrition 2018 05 13;49:81-89. Epub 2017 Nov 13.

Basic Sciences Perinatology Research Laboratories, Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, Texas, USA. Electronic address:

Objectives: Gestational low-protein (LP) programming causes glucose intolerance (GI) and insulin resistance (IR) in adult offspring. Folate supplementation has been shown to rescue the offspring from various programming effects. The aim of this study was to investigate whether folate supplementation during pregnancy reverses LP-induced GI and IR.

Methods: Pregnant rats were fed control (20% protein), isocaloric low-protein (LP, 6%) or LP with 5 mg/kg folate (LPF) diets from gestational day 4 to delivery. The control diet was given during lactation and to pups after weaning. Glucose tolerance test was done at 1, 2, and 3 mo of age followed by euglycemic-hyperinsulinemic clamp at 4 mo. Rats were sacrificed at 4 mo and their gonadal, renal, inguinal, brown fat, and pancreas were weighed and expressed relative to their body weight.

Results: LP- and LPF-fed dams showed similar weight loss during late pregnancy after decreased feed intake. Both LP and LPF pups were smaller at birth but their weights caught up like that of controls by 3 mo. In males, folate supplementation reduced LP-induced GI at 2 mo (glucose area under the curve [AUC]: 1940 mmol/L × 180 min in LP, 1629 mmol/L × 180 min in LPF, and 1653 mmol/L × 180 min in controls; P <0.05, LP versus control and P <0.01, LP versus LPF) but the effect diminished at 3 mo. In females, folate reduced GI at 1 mo (glucose AUC: 1406 mmol/L × 180 min in LP, 1264 mmol/L × 180 min in LPF, and 1281 mmol/L × 180 min in controls; P <0.05, LP versus control and LP versus LPF) but had no effect at 2 and 3 mo. Interestingly, the LPF group had higher pancreatic weights than other groups, suggesting that folate helps in pancreatic development enabling the LPF rats to produce/secrete more insulin to maintain euglycemia. Euglycemic-hyperinsulinemic clamp shows both LP and LPF are insulin resistant compared with controls by 4 mo with LPF more severe than LP in males. Interestingly, females were more insulin resistant than males.

Conclusions: Folate treatment partially reverses LP-induced GI and the magnitude of reversal is age and sex dependent. Furthermore, folate treatment does not reverse IR in either sex but makes it worse in males at 4 mo. The present study demonstrated that folate treatment is not sufficient to rescue the LP programming effects.
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http://dx.doi.org/10.1016/j.nut.2017.10.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6918946PMC
May 2018

Decreased insulin secretion in pregnant rats fed a low protein diet.

Biol Reprod 2017 Oct;97(4):627-635

Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, Texas, USA.

Low protein (LP) diet during pregnancy leads to reduced plasma insulin levels in rodents, but the underlying mechanisms remain unclear. Glucose is the primary insulin secretagogue, and enhanced glucose-stimulated insulin secretion (GSIS) in beta cells contributes to compensation for insulin resistance and maintenance of glucose homeostasis during pregnancy. In this study, we hypothesized that plasma insulin levels in pregnant rats fed LP diet are reduced due to disrupted GSIS of pancreatic islets. We first confirmed reduced plasma insulin levels, then investigated in vivo insulin secretion by glucose tolerance test and ex vivo GSIS of pancreatic islets in the presence of glucose at different doses, and KCl, glibenclamide, and L-arginine. Main findings include (1) plasma insulin levels were unaltered on day 10, but significantly reduced on days 14-22 of pregnancy in rats fed LP diet compared to those of control (CT) rats; (2) insulin sensitivity was unchanged, but glucose intolerance was more severe in pregnant rats fed LP diet; (3) GSIS in pancreatic islets was lower in LP rats compared to CT rats in the presence of glucose, KCl, and glibenclamide, and the response to L-arginine was abolished in LP rats; and (4) the total insulin content in pancreatic islets and expression of Ins2 were reduced in LP rats, but expression of Gcg was unaltered. These studies demonstrate that decreased GSIS in beta cells of LP rats contributes to reduced plasma insulin levels, which may lead to placental and fetal growth restriction and programs hypertension and other metabolic diseases in offspring.
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http://dx.doi.org/10.1093/biolre/iox100DOI Listing
October 2017

Ghrelin doesn't limit insulin release in pregnant rats fed low protein diet.

Front Biosci (Landmark Ed) 2017 06 1;22:1523-1533. Epub 2017 Jun 1.

Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston TX 77030, and Texas Children's Hospital, Houston TX 77030,

To test the hypothesis that insulin secretion in pregnant rats fed LP diet is reduced by ghrelin signaling in pancreatic islets,  we investigated plasma insulin levels, expression of ghrelin and its receptor and glucose stimulated insulin secretion (GSIS) of pancreatic islets in response to ghrelin. Plasma insulin levels were lower and ghrelin receptor abundance in islets was unaltered in LP rats. In the presence of both 2.8 and 16.7 mM glucose, GSIS was lower in LP compared to CT rats. In the presence of 2.8 mM glucose, GSIS was unaltered by ghrelin and its antagonist in both CT and LP rats. In the presence of 16.7 mM glucose, GSIS in LP rats was unchanged while in CT rats was reduced by ghrelin and reversed by ghrelin antagonist. These results indicate ghrelin signaling inhibits GSIS of pancreatic islets in pregnant rats fed CT diet, but it is blunted in pregnant rats fed LP diet and thus may not contribute to the reduction of plasma insulin and GSIS of pancreatic islets in late pregnancy.
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http://dx.doi.org/10.2741/4556DOI Listing
June 2017

Adrenomedullin2 (ADM2)/Intermedin (IMD): A Potential Role in the Pathophysiology of Preeclampsia.

J Clin Endocrinol Metab 2016 11 1;101(11):4478-4488. Epub 2016 Sep 1.

Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, Texas 77030.

Context: It is not known whether decreases in trophoblast invasion promoting the peptide, adrenomedullin2 (ADM2) system is associated with preeclampsia (PreE).

Objective: The objective of the study was to assess the changes in ADM2 levels in plasma, placenta, and amniotic fluid (AF) and its receptor components in placenta from PreE pregnancy compared with the age-matched normal and study the effect of ADM2 on the synthesis of nitric oxide (NO), endothelial nitric oxide synthase (eNOS), and matrix-metalloproteinase (MMP)-2 and MMP-9 in trophoblast cells.

Results: PreE is associated with a decreased expression of ADM2 in plasma and placenta (P < .05); ADM2 interacts with a seven-transmembrane G protein-coupled receptor, calcitonin receptor-like receptor (CRLR) in HTR-8/SVneo cells; placental expression of ADM2/CRLR complex is lower in PreE; mRNA for CRLR and receptor activity-modifying protein-3 are lower, whereas receptor activity-modifying protein-2 is higher in the PreE placenta (P < .05); ADM2 levels in the second trimester are lower in the AF from pregnant women who develop PreE later in gestation (P < .05); ADM2 is localized to the epithelium of the amnion and the ectoderm and mesoderm of the chorion in term fetal membranes; ADM2 increases NO production, eNOS, and MMP2/9-immunoreactivity, whereas ADM2 knockdown inhibits the expression of eNOS and MMP2/9 mRNA and S-nitrosylation in HTR-8/SVneo cells; and ADM2-induced increases in MMP2/9 activity is inhibited by L-nitro-arginine methyl ester in HTR-8SV/neo cells.

Conclusion: Decreases in the ADM2 system in PreE at term, in AF from pregnant women during the second trimester who develop PreE later in gestation, and ADM2-induced increases in the NO and MMP-2/9 levels in trophoblast cells suggest a potential role for ADM2 via the NO-MMP system in the pathophysiology of PreE.
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http://dx.doi.org/10.1210/jc.2016-1333DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5095259PMC
November 2016

Calcitonin Gene-Related Peptide Rescues Proximity Associations of Its Receptor Components, Calcitonin Receptor-Like Receptor and Receptor Activity-Modifying Protein 1, in Rat Uterine Artery Smooth Muscle Cells Exposed to Tumor Necrosis Factor Alpha.

Biol Reprod 2016 12 26;95(6):126. Epub 2016 Oct 26.

Department of Obstetrics and Gynecology, Baylor College of Medicine/Texas Children's Hospital, Houston, Texas

Calcitonin gene-related peptide (CALCB), adrenomedullin (ADM), and ADM2/intermedin play critical roles in vascular adaptation during pregnancy through calcitonin receptor-like receptor (CALCRL) and receptor activity-modifying proteins (RAMPs). This study was designed to assess the predominant RAMP that associates with CALCRL to form a functional receptor in the rat uterine artery smooth muscle (RUASM). We also determined if these receptor component associations are decreased by tumor necrosis factor (TNF) alpha and if CALCB, ADM, or ADM2 can rescue CALCRL/RAMP associations. Using proximity ligation assay in RUASM cells, this study shows that CALCRL predominantly associates with RAMP1 forming a CALCB receptor, and minimally with RAMP2 and RAMP3 that confer specificity for ADM and ADM2. However, knockdown of RAMP1 mRNA increases the interaction between CALCRL and RAMP3 without affecting the association of CALCRL and RAMP2. Furthermore, CALCB, ADM, and ADM2 have no effects on the associations of CALCRL with any of the RAMPs in RUASM cells. Interestingly, CALCB reverses the TNFalpha-induced decreases in CALCRL/RAMP1 associations. Furthermore, CALCB increases ERK1/2 phosphorylation in a time-dependent manner in RUASM, and the protective effect of CALCB on TNFalpha-induced inhibition of CALCRL/RAMP1 associations was significantly blocked in presence of ERK inhibitor (PD98059). In conclusion, this study demonstrates that CALCRL predominantly associates with RAMP1 forming a CALCB-specific receptor complex in RUASM cells, which is dissociated by TNFalpha. Rescue of TNFalpha-induced dissociation of CALCRL/RAMP1 complex by CALCB in RUASM cells suggests a potential use of CALCB in developing therapeutic strategies for pregnancy-related complications that are vulnerable to abnormal levels of TNFalpha, such as fetal growth restriction and preeclampsia.
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http://dx.doi.org/10.1095/biolreprod.116.143529DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5315425PMC
December 2016

Impaired Vasodilatory Responses of Omental Arteries to CGRP Family Peptides in Pregnancies Complicated by Fetal Growth Restriction.

J Clin Endocrinol Metab 2016 08 3;101(8):2984-93. Epub 2016 Jun 3.

Department of Obstetrics and Gynecology, Baylor College of Medicine/Texas Children's Hospital, Houston, Texas.

Rationale: Calcitonin gene-related peptide (CGRP), adrenomedullin (ADM), and adrenomedullin2 (ADM2)/intermedin are potent vasorelaxant peptides considered to play a role in the adaptive mechanisms in rat pregnancy through increased vasodilation in mesenteric and uterine artery.

Objective: This study was designed to demonstrate the response of omental arteries (OA) to vasoactive peptides CGRP, ADM, and ADM2 in pregnancy complications such as fetal growth restriction (FGR), and assess the changes in the expression of their receptor components in segments of OA from FGR pregnancy compared to the control.

Findings: The findings for this study are: 1) relaxation responses of OA were higher for bradykinin (78.55 ± 3.91 vs 52.67 ± 2.19; P < .05) in pregnancy with FGR compared to the normal, 2) relaxation response of OA segments to CGRP was similar with no change in the expression of G-protein couple receptor-calcitonin receptor-like receptor complex in normal healthy pregnancy and pregnancy complicated by FGR, 3) maximal relaxation response of OA were significantly (P < .05) lower for both ADM (18.2 ± 6.7 vs 38 ± 2.5) and ADM2 (26.9 ± 6.7 vs 48 ± 2.6) along with decreases in their respective ligand-receptor complex in FGR compared to the normal pregnancies, 4) expression of calcitonin receptor-like receptor mRNA was higher but its immunoreactivity was lower in OA from FGR pregnancy compared to the normal, and 5) mRNA and protein levels of RAMP1, RAMP2, and RAMP3 were lower in OA isolated from FGR pregnancies compared to the normal.

Conclusion: The current study demonstrates that FGR is associated with an increase in the sensitivity of OA to bradykinin and decreased sensitivity for ADM and ADM2 ligand-receptor system with no change in the response for CGRP compared to the normal healthy pregnancy, and suggests a potential role for ADM and ADM2 in the pathophysiology of maternal vasculature in FGR pregnancy.
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http://dx.doi.org/10.1210/jc.2016-1798DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6287517PMC
August 2016

Novel lean type 2 diabetic rat model using gestational low-protein programming.

Am J Obstet Gynecol 2016 Apr 10;214(4):540.e1-540.e7. Epub 2016 Feb 10.

Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, TX. Electronic address:

Background: Type 2 diabetes (T2D) in lean individuals is not well studied and up to 26% of diabetes occurs in these individuals. Although the cause is not well understood, it has been primarily attributed to nutritional issues during early development.

Objective: Our objective was to develop a lean T2D model using gestational low-protein (LP) programming.

Study Design: Pregnant rats were fed control (20% protein) or isocaloric LP (6%) diet from gestational day 4 until delivery. Standard diet was given to dams after delivery and to pups after weaning. Glucose tolerance test was done at 2, 4, and 6 months of age. Magnetic resonance imaging of body fat for females was done at 4 months. Rats were sacrificed at 4 and 8 months of age and their perigonadal, perirenal, inguinal, and brown fat were weighed and expressed relative to their body weight. Euglycemic-hyperinsulinemic clamp was done around 6 months of age.

Results: Male and female offspring exposed to a LP diet during gestation developed glucose intolerance and insulin resistance (IR). Further, glucose intolerance progressed with increasing age and occurred earlier and was more severe in females when compared to males. Euglycemic-hyperinsulinemic clamp showed whole body IR in both sexes, with females demonstrating increased IR compared to males. LP females showed a 4.5-fold increase in IR while males showed a 2.5-fold increase when compared to their respective controls. Data from magnetic resonance imaging on female offspring showed no difference in the subcutaneous, inguinal, and visceral fat content. We were able to validate this observation by sacrificing the rats at 4 and 8 months and measuring total body fat content. This showed no differences in body fat content between control and LP offspring in either males or females. Additionally, diabetic rats had a similar body mass index to that of the controls.

Conclusion: LP gestational programming produces a progressively worsening T2D model in rats with a lean phenotype without obesity.
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http://dx.doi.org/10.1016/j.ajog.2016.02.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4808600PMC
April 2016

Blunted hypothalamic ghrelin signaling reduces diet intake in rats fed a low-protein diet in late pregnancy.

Physiol Rep 2015 Dec;3(12)

Department of Obstetrics & Gynecology, Baylor College of Medicine, Houston, Texas

Diet intake in pregnant rats fed a low-protein (LP) diet was significantly reduced during late pregnancy despite elevated plasma levels of ghrelin. In this study, we hypothesized that ghrelin signaling in the hypothalamus is blunted under a low-protein diet condition and therefore, it does not stimulate diet intake during late pregnancy. Female Sprague-Dawley rats were fed a normal (CT) or LP diet from Day 1 of pregnancy. On Day 21, 0.5 μg ghrelin was given into the third ventricle (ICV). Diet and water intake at 30, 60, and 120 min after ICV injection was measured. Hypothalami were dissected and analyzed for expression of genes related to appetite regulation (Npy, Agrp, Pomc and Cart) and phosphorylation of AMPK and ACC proteins (downstream proteins of ghrelin receptor activation). Results include: In response to ICV injection of ghrelin, (1) diet intake was significantly lower in LP compared to CT rats; (2) water intake was not affected in LP rats; (3) expression of Npy and Agrp, but not Pomc and Cart, were higher in the hypothalamus of LP compared to CT rats; (4) the abundance of phosphorylated AMPK and the ratio of phosphorylated to total AMPK, but not the abundance of total AMPK, were lower in LP compared to CT rats; (5) the abundance of phosphorylated ACC, but not total ACC, was lower in LP rats. These findings suggest that blunted ghrelin signaling in the hypothalamus of pregnant rats fed a LP diet leads to reduced diet intake and exacerbates gestational protein insufficiency.
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http://dx.doi.org/10.14814/phy2.12629DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4760445PMC
December 2015

Complement Split Products in Amniotic Fluid in Pregnancies Subsequently Developing Early-Onset Preeclampsia.

Dis Markers 2015 18;2015:263109. Epub 2015 Oct 18.

Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, TX 77030, USA.

Objective: To determine the second-trimester amniotic fluid concentrations of complement split products in pregnancies subsequently affected by early-onset preeclampsia.

Study Design: Cohort of 731 women with singleton pregnancies undergoing second-trimester genetic amniocentesis followed up to delivery and analyzed as a nested case-control study. Cases of preeclampsia developing before 34 weeks' gestation (n = 15) were compared with 47 uncomplicated term controls. Amniotic fluid collected at amniocentesis was tested for complement split products Bb, C4a, C3a, and C5a.

Results: Women who developed early-onset preeclampsia as compared with the term pregnant controls had significantly higher (P = 0.04) median amniotic fluid C3a levels (318.7 ng/mL versus 254.5 ng/mL). Median amniotic fluid Bb levels were also significantly higher (P = 0.03) in preeclamptic women than in normal pregnant women (1127 ng/mL versus 749 ng/mL). Median levels of C4a and C5a were not significantly different between the groups.

Conclusion: Our data suggest that complement activation in early pregnancy is associated with early-onset preeclampsia. We believe this to be the first prospective study to link complement activation in amniotic fluid in early pregnancy and later development of preeclampsia. Our findings provide evidence that immune dysregulation may precede the clinical manifestations of preeclampsia and that the alternative complement pathway is principally involved.
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http://dx.doi.org/10.1155/2015/263109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4628677PMC
July 2016

Enalapril Normalizes Endothelium-Derived Hyperpolarizing Factor-Mediated Relaxation in Mesenteric Artery of Adult Hypertensive Rats Prenatally Exposed to Testosterone.

Biol Reprod 2015 Jun 13;92(6):155. Epub 2015 May 13.

Division of Reproductive Endocrinology, Department of Obstetrics and Gynecology, University of Texas Medical Branch, Galveston, Texas

Prenatal exposure to elevated testosterone levels induces adult life hypertension associated with selective impairments in endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation in mesenteric arteries. We tested whether the angiotensin-converting enzyme inhibitor enalapril restores EDHF function through regulating the activities of small (Kcnn3) and intermediate (Kcnn4) conductance calcium-activated potassium channels in mesenteric arteries. Pregnant Sprague-Dawley rats were injected subcutaneously with vehicle or testosterone propionate (0.5 mg/kg/day from Gestation Day 15 to 19), and their 6-mo-old adult male offspring were examined. A subset of rats in these two groups was given enalapril (40 mg/kg/day) for 2 wk through drinking water. Blood pressures were assessed through carotid arterial catheter and endothelium-dependent mesenteric arterial EDHF relaxation, using wire myography. Ace and Kcnn3 and Kcnn4 channel expression levels were also examined. Renal and vascular Ace expression and plasma angiotensin II levels were increased in testosterone offspring. Blood pressure levels were significantly higher in testosterone offspring than in controls, and treatment with enalapril significantly attenuated blood pressure in testosterone offspring. EDHF relaxation in testosterone offspring was reduced compared to that in controls, and it was significantly restored by enalapril treatment. Kcnn4 channel expression and function were similar between control and testosterone rats, but it was not affected by enalapril treatment. Relaxation mediated by Kcnn3 was impaired in testosterone offspring, and it was normalized by enalapril treatment. Furthermore, enalapril treatment restored expression levels of Kcnn3 channels. These findings suggest that enalapril has a positive influence on endothelial function with improvement in EDHF relaxation through normalization of Kcnn3 expression and activity.
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http://dx.doi.org/10.1095/biolreprod.115.130468DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4652613PMC
June 2015

Appetite regulation is independent of the changes in ghrelin levels in pregnant rats fed low-protein diet.

Physiol Rep 2015 Apr;3(4)

Department of Obstetrics & Gynecology, Baylor College of Medicine, Houston, Texas Texas Children's Hospital, Houston, Texas

Gestational protein restriction causes hypertension in the adult offspring. Very little is known about the food intake regulation and ghrelin signaling in pregnant dams fed a low-protein (LP) diet. We hypothesized that diet intake and ghrelin signaling are altered in pregnant rats fed the low-protein diet. Sprague-Dawley rats were fed a control (CT) or LP diet from Day 3 of pregnancy. Diet intake and body weight were monitored daily. Expression of ghrelin production-related genes in the stomach and appetite-related genes in the hypothalamus was analyzed by real-time PCR. Plasma levels of total and active ghrelin, growth hormone and leptin were measured by ELISA. Main results include: (1) Daily diet intake was greater in the LP group than in the CT group in early pregnancy, but substantially lower in late pregnancy; (2) Daily gain in body weight was substantially lower in the LP group in late pregnancy; (3) Expression of ghrelin production-related genes in the stomach and plasma total ghrelin levels were increased in LP group in late pregnancy; (4) Plasma active ghrelin levels were elevated in the LP group at mid-late pregnancy, but growth hormone and leptin levels were uncorrelated with active ghrelin in late pregnancy; and (5) Hypothalamic expression of ghrelin-stimulated genes in LP rats was unassociated with the changes in both plasma ghrelin levels and the diet intake. Taken together, the appetite in LP rats is greater in early pregnancy but reduced at late pregnancy, possibly due to ghrelin insensitivity in appetite regulation.
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http://dx.doi.org/10.14814/phy2.12368DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4425973PMC
April 2015

Prenatal testosterone exposure induces hypertension in adult females via androgen receptor-dependent protein kinase Cδ-mediated mechanism.

Hypertension 2015 Mar 8;65(3):683-690. Epub 2014 Dec 8.

Division of Reproductive Endocrinology Department of Obstetrics and Gynecology, University of Texas Medical Branch, Galveston, TX 77555.

Prenatal exposure to excess testosterone induces hyperandrogenism in adult females and predisposes them to hypertension. We tested whether androgens induce hypertension through transcriptional regulation and signaling of protein kinase C (PKC) in the mesenteric arteries. Pregnant Sprague-Dawley rats were injected with vehicle or testosterone propionate (0.5 mg/kg per day from gestation days 15 to 19, SC) and their 6-month-old adult female offspring were examined. Plasma testosterone levels (0.84±0.04 versus 0.42±0.09 ng/mL) and blood pressures (111.6±1.3 versus 104.5±2.4 mm Hg) were significantly higher in prenatal testosterone-exposed rats compared with controls. This was accompanied with enhanced expression of PKCδ mRNA (1.5-fold) and protein (1.7-fold) in the mesenteric arteries of prenatal testosterone-exposed rats. In addition, mesenteric artery contractile responses to PKC activator, phorbol-12,13-dibutyrate, was significantly greater in prenatal testosterone-exposed rats. Treatment with androgen receptor antagonist flutamide (10 mg/kg, SC, BID for 10 days) significantly attenuated hypertension, PKCδ expression, and the exaggerated vasoconstriction in prenatal testosterone-exposed rats. In vitro exposure of testosterone to cultured mesenteric artery smooth muscle cells dose dependently upregulated PKCδ expression. Analysis of PKCδ gene revealed a putative androgen responsive element in the promoter upstream to the transcription start site and an enhancer element in intron-1. Chromatin immunoprecipitation assays showed that androgen receptors bind to these elements in response to testosterone stimulation. Furthermore, luciferase reporter assays showed that the enhancer element is highly responsive to androgens and treatment with flutamide reverses reporter activity. Our studies identified a novel androgen-mediated mechanism for the control of PKCδ expression via transcriptional regulation that controls vasoconstriction and blood pressure.
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http://dx.doi.org/10.1161/HYPERTENSIONAHA.114.04521DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4326589PMC
March 2015

Adrenomedullin2 (ADM2)/intermedin (IMD) in rat ovary: changes in estrous cycle and pregnancy and its role in ovulation and steroidogenesis.

Biol Reprod 2015 Feb 13;92(2):39. Epub 2014 Nov 13.

Department of Obstetrics & Gynecology, Baylor College of Medicine, Houston, Texas.

Adrenomedullin2 (ADM2) is reported to facilitate embryo implantation and placental development. Therefore, the current study was undertaken to identify if ADM2 has a functional role in ovary to facilitate its reproductive actions. This study shows that the expression of ADM2 is differentially regulated in rat estrous cycle and that ADM2 increases the synthesis and secretion of 17beta-estradiol accompanied with an increase in the expression of steroidogenic factor 1 (Sf1), estrogen receptor Esr1, and enzymes involved in steroidogenesis in equine chorionic gonadotropin (eCG)-treated rat ovaries. In addition, inhibition of endogenous ADM2 function in eCG-treated immature rats caused impaired ovulation. Furthermore, the mRNA expression of Adm2 and receptor activity modifying protein 3 is higher in the ovary on Day 18 compared to nonpregnant and pregnant rats on Day 22. ADM2-like immunoreactivity is localized in granulosa cells, blood vessels, oocytes, cumulous oophorus, and corpus luteum of pregnant ovaries, suggesting a potential role for ADM2 in the ovary. This is supported by the presence of ADM2-like immunoreactivity in the corpus luteum during pregnancy and a decline in aromatase immunoreactivity in corpus luteum on Day 9 of gestation in rats infused with ADM2 antagonist during implantation and decidualization phase. Taken together, this study suggests a potential involvement of ADM2 in the rat ovary in regulating synthesis of estradiol to support ovulation and facilitate efficient implantation and placental development for a successful pregnancy.
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http://dx.doi.org/10.1095/biolreprod.113.112854DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4326727PMC
February 2015

Adrenomedullin promotes rat trophoblast stem cell differentiation.

Biol Reprod 2014 Sep 24;91(3):65. Epub 2014 Jul 24.

Department of Obstetrics & Gynecology, Baylor College of Medicine, Houston, Texas

Accumulating data suggest that adrenomedullin (ADM) regulates the trophoblast cell growth, migration, and invasion. However, the effect of ADM on trophoblast differentiation is poorly understood. In this study, we hypothesized that ADM promotes the differentiation of trophoblast stem cells (TSCs) into trophoblast giant cells (TGCs). Using rat TSCs, Rcho-1 cells, we investigated the effect of ADM on TSC differentiation into TGCs in differentiation or stem cell media, respectively, and explored the effect of ADM on the mechanistic target of rapamycin (MTOR) signaling in trophoblast cell differentiation. The results include: 1) in the presence of differentiation medium, 10⁻⁷ M ADM, but not lower doses, elevated (P < 0.05) Prl3b1/Esrrb (i.e., the ratio of mRNA levels) by 1.7-fold compared to that in control; 2) the supplementation of ADM antagonist, regardless of the concentration of ADM, reduced (P < 0.05) Prl3b1/Esrrb by 2-fold, compared to control group, while the supplementation of CGRP antagonist, regardless of the concentration of ADM, did not change Prl3b1/Esrrb; 3) in the presence of stem cell medium, ADM did not alter the expression of TSC and TGC marker genes, however, the ratio of Prl3b1/Esrrb was reduced (P < 0.05) by ADM antagonist compared to that in control; and 4) ADM increased (P < 0.05) phosphorylated MTOR proteins and the ratio of phosphorylated to total MTOR proteins by 2.0- and 1.7-fold, respectively. The results indicate that ADM promotes but does not induce the differentiation of TSCs to TGCs in a dose-dependent manner and MTOR signaling may play a role in this process.
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http://dx.doi.org/10.1095/biolreprod.114.120378DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4435060PMC
September 2014

Calcitonin gene related family peptides: importance in normal placental and fetal development.

Adv Exp Med Biol 2014 ;814:229-40

Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, TX, USA,

Synchronized molecular and cellular events occur between the uterus and the implanting embryo to facilitate successful pregnancy outcome. Nevertheless, the molecular signaling network that coordinates strategies for successful decidualization, placentation and fetal growth are not well understood. The discovery of calcitonin/calcitonin gene-related peptides (CT/CGRP) highlighted new signaling mediators in various physiological processes, including reproduction. It is known that CGRP family peptides including CGRP, adrenomedulin and intermedin play regulatory functions during implantation, trophoblast proliferation and invasion, and fetal organogenesis. In addition, all the CGRP family peptides and their receptor components are found to be expressed in decidual, placental and fetal tissues. Additionally, plasma levels of peptides of the CGRP family were found to fluctuate during normal gestation and to induce placental cellular differentiation, proliferation, and critical hormone signaling. Moreover, aberrant signaling of these CGRP family peptides during gestation has been associated with pregnancy disorders. It indicates the existence of a possible regulatory role for these molecules during decidualization and placentation processes, which are known to be particularly vulnerable. In this review, the influence of the CGRP family peptides in these critical processes is explored and discussed.
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http://dx.doi.org/10.1007/978-1-4939-1031-1_20DOI Listing
December 2014

Gestational exposure to elevated testosterone levels induces hypertension via heightened vascular angiotensin II type 1 receptor signaling in rats.

Biol Reprod 2014 Jul 22;91(1). Epub 2014 May 22.

Division of Reproductive Endocrinology, Department of Obstetrics and Gynecology, University of Texas Medical Branch, Galveston, Texas

Pre-eclampsia is a life-threatening pregnancy disorder whose pathogenesis remains unclear. Plasma testosterone levels are elevated in pregnant women with pre-eclampsia and polycystic ovary syndrome, who often develop gestational hypertension. We tested the hypothesis that increased gestational testosterone levels induce hypertension via heightened angiotensin II signaling. Pregnant Sprague-Dawley rats were injected with vehicle or testosterone propionate from Gestational Day 15 to 19 to induce a 2-fold increase in plasma testosterone levels, similar to levels observed in clinical conditions like pre-eclampsia. A subset of rats in these two groups was given losartan, an angiotensin II type 1 receptor antagonist by gavage during the course of testosterone exposure. Blood pressure levels were assessed through a carotid arterial catheter and endothelium-independent vascular reactivity through wire myography. Angiotensin II levels in plasma and angiotensin II type 1 receptor expression in mesenteric arteries were also examined. Blood pressure levels were significantly higher on Gestational Day 20 in testosterone-treated dams than in controls. Treatment with losartan during the course of testosterone exposure significantly attenuated testosterone-induced hypertension. Plasma angiotensin II levels were not significantly different between control and testosterone-treated rats; however, elevated testosterone levels significantly increased angiotensin II type 1 receptor protein levels in the mesenteric arteries. In testosterone-treated rats, mesenteric artery contractile responses to angiotensin II were significantly greater, whereas contractile responses to K(+) depolarization and phenylephrine were unaffected. The results demonstrate that elevated testosterone during gestation induces hypertension in pregnant rats via heightened angiotensin II type 1 receptor-mediated signaling, providing a molecular mechanism linking elevated maternal testosterone levels with gestational hypertension.
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http://dx.doi.org/10.1095/biolreprod.114.118968DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4434963PMC
July 2014

Elevated testosterone levels during rat pregnancy cause hypersensitivity to angiotensin II and attenuation of endothelium-dependent vasodilation in uterine arteries.

Hypertension 2014 Aug 19;64(2):405-14. Epub 2014 May 19.

From the Division of Reproductive Endocrinology, Department of Obstetrics and Gynecology, University of Texas Medical Branch, Galveston (V.C., K.L.V., G.R.S., G.D.H., K.S.); and Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, TX (C.S.B., C.Y.).

Elevated testosterone levels increase maternal blood pressure and decrease uterine blood flow in pregnancy, resulting in abnormal perinatal outcomes. We tested whether elevated testosterone alters uterine artery adaptations during pregnancy, and whether these alterations depend on endothelium-derived factors such as nitric oxide, endothelium-derived hyperpolarizing factor, and prostacyclin, or endothelium-independent mechanisms such as angiotensin II (Ang-II). Pregnant Sprague-Dawley rats were injected with vehicle (n=20) or testosterone propionate (0.5 mg/kg per day from gestation day 15 to 19; n=20). Plasma testosterone levels increased 2-fold in testosterone-injected rats compared with controls. Elevated testosterone significantly decreased placental and pup weights compared with controls. In endothelium-intact uterine arteries, contractile responses to thromboxane, phenylephrine, and Ang-II were greater in testosterone-treated rats compared with controls. In endothelium-denuded arteries, contractile responses to Ang-II (pD2=9.1±0.04 versus 8.7±0.04 in controls; P<0.05), but not thromboxane and phenylephrine, were greater in testosterone-treated rats. Ang-II type 1b receptor expression was increased, whereas Ang-II type 2 receptor was decreased in testosterone-exposed arteries. In endothelium-denuded arteries, relaxations to sodium nitroprusside were unaffected. Endothelium-dependent relaxation to acetylcholine was significantly lower in arteries from testosterone-treated dams (Emax=51.80±6.9% versus 91.98±1.4% in controls; P<0.05). The assessment of endothelial factors showed that nitric oxide-, endothelium-derived hyperpolarizing factor-, and prostacyclin-mediated relaxations were blunted in testosterone-treated dams. Endothelial nitric oxide synthase, small conductance calcium-activated potassium channel-3, and prostacyclin receptor expressions were significantly decreased in arteries from testosterone-treated dams. Hypoxia-inducible factor-1α, Ankrd37, and Egln were significantly increased in testosterone-exposed placentas. These results suggest that elevated maternal testosterone impairs uterine vascular function, which may lead to an increased vascular resistance and a decrease in uterine blood flow.
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http://dx.doi.org/10.1161/HYPERTENSIONAHA.114.03283DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4096063PMC
August 2014