transforming-growth-factor-beta and Abnormalities--Drug-Induced

N629D KCNH2 is a human missense long-QT2 mutation. Previously, we reported that the N629D/N629D mutation embryos disrupted cardiac looping, right ventricle development, and ablated IKr activity at E9.5. The present study evaluates the role of KCNH2 in vasculogenesis.. N629D/N629D yolk sac vessels and aorta consist of sinusoids without normal arborization. Isolated E9.5 +/+ first branchial arches showed normal outgrowth of mouse ERG-positive/α-smooth muscle actin coimmunolocalized cells; however, outgrowth was grossly reduced in N629D/N629D. N629D/N629D aortas showed fewer α-smooth muscle actin positive cells that were not coimmunolocalized with mouse ERG cells. Transforming growth factor-β treatment of isolated N629D/N629D embryoid bodies partially rescued this phenotype. Cultured N629D/N629D embryos recapitulate the same cardiovascular phenotypes as seen in vivo. Transforming growth factor-β treatment significantly rescued these embryonic phenotypes. Both in vivo and in vitro, dofetilide treatment, over a narrow window of time, entirely recapitulated the N629D/N629D fetal phenotypes. Exogenous transforming growth factor-β treatment also rescued the dofetilide-induced phenotype toward normal.. Loss of function of KCNH2 mutations results in defects in cardiogenesis and vasculogenesis. Because many medications inadvertently block the KCNH2 potassium current, these novel findings seem to have clinical relevance.

Topics: Abnormalities, Drug-Induced; Animals; Cells, Cultured; Embryo Culture Techniques; Embryonic Stem Cells; ERG1 Potassium Channel; Ether-A-Go-Go Potassium Channels; Fetal Death; Gene Expression Regulation, Developmental; Genotype; Heart Defects, Congenital; Humans; Mice, 129 Strain; Mice, Transgenic; Morphogenesis; Mutation, Missense; Neovascularization, Physiologic; Phenethylamines; Phenotype; Potassium Channel Blockers; Signal Transduction; Sulfonamides; Transforming Growth Factor beta; Vascular Malformations

Topics: Abnormalities, Drug-Induced; Animals; Embryonic Stem Cells; ERG1 Potassium Channel; Ether-A-Go-Go Potassium Channels; Fetal Death; Heart Defects, Congenital; Humans; Mutation, Missense; Neovascularization, Physiologic; Phenethylamines; Potassium Channel Blockers; Sulfonamides; Transforming Growth Factor beta; Vascular Malformations

Retinoic acid (RA) is a vitamin A derivative that participates in patterning and regulation of inner ear development. Either excess RA or RA deficiency during a critical stage of inner ear development can produce teratogenic effects. Previous studies have shown that in utero exposure of the developing mouse inner ear to a high dose of all-trans RA (atRA) results in severe malformations of the inner ear that are associated with diminished levels of endogenous transforming growth factor-beta1 (TGF-beta(1)) protein.. In this study, the effects of a teratogenic level of atRA on levels and patterns of expression of TGFbeta receptor II (TGFbetaRII) and Smad2, a downstream component of the TGFbeta signal transduction pathway, are investigated in the developing mouse inner ear. The expression pattern of endogenous RA receptor alpha (RARalpha) and the ability of an RARalpha(1)-specific antisense oligonucleotide (AS) to modulate otic capsule chondrogenesis are demonstrated in the inner ear and in culture.. Endogenous TGFbetaRII and Smad2 are downregulated in the inner ear following in utero atRA treatment. In addition, a reduction in endogenous TGFbeta(1) and a marked suppression of chondrogenesis occur in RARalpha(1) AS-treated cultures in comparison to untreated or oligonucleotide-treated control cultures. This chondrogenic suppression can be partially overcome by supplementation of RARalpha(1) AS-treated cultures with exogenous TGFbeta(1) protein.. Our findings support a role for TGFbeta in the physiological and pathological effects of RA on inner ear development.

Topics: Abnormalities, Drug-Induced; Animals; Chondrogenesis; DNA-Binding Proteins; Down-Regulation; Ear, Inner; Epithelium; Female; Gene Expression; Male; Mesoderm; Mice; Oligonucleotides, Antisense; Pregnancy; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Retinoic Acid; Receptors, Transforming Growth Factor beta; Retinoic Acid Receptor alpha; Signal Transduction; Smad2 Protein; Trans-Activators; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tretinoin

To investigate the morphologic changes of embryonic palatal development exposed to retinoic acid (RA) in mouse, and to detect the significance of the expression of TGFbeta1, TGFbeta3, EGF and BCL2.. The stage of palatal development was examined by light microscopy. S-P immunohistochemistry and in-situ hybridization was used to detect spatio-temporal patterns of expression of TGFbeta1, TGFbeta3, EGF and BCL2 in embryonic palate.. The fetus exposed to RA resulted in formation of small palatal shelves without contact and fusion of each other to form and intact palate. RA can regulate the embryonic palatal expression of genes involved in RA-induced cleft palate.. RA can inhibit the proliferation of MEPM cell to form small palatal shelves and induce abnormal differentiation of MEE cell causing the bi-palatal shelves no contact and fuse with each other, then induce the formation of cleft palate. RA can regulate the spatio-temporal patterns of expression of TGFbeta1, TGFbeta3 and EGF in embryonic palatal processes and the change of special expression of these genes in embryonic palatal processes are involved in RA-induced cleft palate.

Topics: Abnormalities, Drug-Induced; Animals; Cleft Palate; Embryo, Mammalian; Epidermal Growth Factor; Female; Mice; Mice, Inbred C57BL; Palate; Transforming Growth Factor beta; Tretinoin

For unknown reasons, activation of the maternal immune system in mice reduces morphologic defects caused by diverse teratogenic agents. Such immune stimulation of the maternal animal has been correlated with altered cytokine mRNA transcripts in the placenta (e.g., TGFbeta2) as well as in fetal target tissues of the teratogen (e.g., TNFalpha in fetal heads of cyclophosphamide-exposed pregnant mice). The teratogen urethane was reported to down-regulate cell cycle and apoptotic regulatory genes in fetal mouse heads that displayed cleft palate, an effect that was also reversed by maternal immune stimulation. The molecular mediators of the above phenomena have not been identified, however proteins synthesized and released by activated maternal immune cells have been suggested. The present studies therefore evaluated the effects of maternal immune stimulation in urethane-exposed mice on thymus and spleen leukocyte populations, in an attempt to identify events that may correlate with protection against birth defects. Immune stimulation did not change the hypocellularity of the thymus nor the altered T cell differentiation caused by urethane. A limited and transient increase in splenic leukocyte number, including increased T and B lymphocytes and macrophages, was caused by immune stimulation and was not felt to play a significant role in reduced morphologic defects. Urethane treatment caused down-regulated expression of numerous genes involved in cell-cycle control, while maternal immune stimulation caused comparative up-regulation of many of these genes. Coordinate shifts in gene expression by treatment were evaluated using principal component analysis, which identified several growth factor genes that were differentially expressed in mice receiving urethane alone as compared to urethane plus immune stimulation. Up-regulated expression of TGFbeta3 and GM-CSF genes, in particular, was observed in leukocytes of urethane-exposed mice receiving immunostimulation. Interestingly, the cytokine products of these two genes were recently suggested as growth factors that may be related to reduction of fetal defects caused by teratogens. Genes for growth factors IGF-I, IGF-II and IL-2 were also identified as differentially expressed in urethane vs. urethane+immune stimulation mice, suggesting that these proteins should be considered for a potential contributing effect to reduced birth defects caused by immunostimulation.

Topics: Abnormalities, Drug-Induced; Adjuvants, Immunologic; Animals; Antigens, Surface; Cytokines; Female; Gene Expression Regulation; Granulocyte-Macrophage Colony-Stimulating Factor; Lymphocyte Count; Male; Mice; Mice, Inbred ICR; Oligonucleotide Array Sequence Analysis; Pregnancy; Pregnancy, Animal; Thymus Gland; Transforming Growth Factor beta; Transforming Growth Factor beta3; Urethane

In female rats, in uteroexposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) during critical periods of organogenesis causes a permanent thread of tissue, consisting of a core of mesenchyme surrounded by keratinized epithelia, across the vaginal opening. The objective of the current study was to determine the earliest time after exposure to TCDD during fetal development that morphological changes in the development of the lower reproductive tract could be detected. In addition, the spatio-temporal expression of several growth factors within the developing reproductive tract was investigated to provide insight into the mechanism of action involved in TCDD-induced vaginal thread formation. Pregnant rats received a single oral dose of 1.0 microg TCDD/kg on gestation day (GD) 15. Dams were sacrificed on GD 17, 18, 19, and 21 and individual reproductive tracts were isolated from female fetuses. As early as GD 18, TCDD produced distinct abnormalities in the female reproductive tract. The width of mesenchyme separating the Mullerian ducts was significantly greater in TCDD-exposed female GD 18 and 19 fetuses and the zone of unfused Mullerian ducts was substantially increased on GD 19 and 21. TCDD induced alterations within the developing reproductive tract in the subcellular and temporal expression of transforming growth factor-beta3 (TGF-beta3) and epidermal growth factor receptor (EGFR). DNA array analysis suggested effects on several genes expressed on GD 18 and 19.

Topics: Abnormalities, Drug-Induced; Animals; Embryonic and Fetal Development; Female; Gene Expression Regulation, Developmental; Genes, erbB-1; Genitalia, Female; Immunohistochemistry; Laminin; Maternal Exposure; Mesoderm; Mesonephros; Mullerian Ducts; Polychlorinated Dibenzodioxins; Pregnancy; Rats; Rats, Long-Evans; Time Factors; Transforming Growth Factor beta; Transforming Growth Factor beta3; Urethra

TGFbetas are among the main immunoregulatory molecules contributing to successful embryonic development. Besides, our and other studies revealed that maternal immunopotentiation has a potential to increase the resistance of the embryo to the teratogenic insult. This work was designed to evaluate: (1) whether the formation of teratogen-induced anomalies is accompanied by an altered pattern of TGFbeta2 expression in embryonic cells and (2) whether maternal immunopotentiation modifies the pattern of TGFbeta2 expression in embryos responding to the teratogenic insult.. Experiments were performed in embryos of ICR mice exposed to 15 and 40 mg/kg of a reference teratogen, cyclophosphamide (CP) on day 12 of gestation. A group of mice was immunopotentiated with xenogeneic rat splenocytes 21 hr before the beginning of mating. Embryos were examined for the occurrence of gross structural anomalies 24 and 72 hr after CP treatment. Then, immunohistohemistry and in situ hybrydization assays were used to evaluate the expression of TGFbeta2 protein and mRNA in the brain, face, limbs and liver of these embryos.. No external anomalies were observed in embryos examined 24 hr after CP treatment. Embryos examined 72 hr after CP treatment at 40 mg/kg exhibited agnathia, micrognathia, kinky tail, phocomelia, but no signs of dismorphogenesis were observed in the liver at the organ level. A significant increase in the expression of TGFbeta2 mRNA was observed in cells, residing in the brain, face and limbs but not in the liver of CP-exposed embryos tested 24 hr after CP injection in both doses. The level of TGFbeta2 protein in these embryos did not differ from that of controls. In embryos tested 72 hr after CP injection in the high dose both TGFbeta2 protein and mRNA expression were found to be elevated. Maternal immunopotentiation while enhancing the embryo's resistance to CP practically abolished an elevated expression of the TGFbeta2 mRNA detected in tested organ structures of embryos of non-immunopotentiated CP treated mice 24 hr after CP injection in both the low and the high doses. Also, a significant decrease in the level of TGFbeta2 mRNA expression was observed in embryos of immunopotentiated mice examined 72 hr after CP treatment.. The results of this work show a possible involvement of TGFbeta2 in the formation of teratogen-induced structural anomalies and suggest that the stimulation of the maternal immune system may realize its protective effect by normalizing the level of TGFbeta2 expression in teratogen-targeted embryonic structures.

Topics: Abnormalities, Drug-Induced; Adoptive Transfer; Animals; Apoptosis; Cyclophosphamide; Male; Mice; Mice, Inbred ICR; Rats; Rats, Long-Evans; RNA, Messenger; Transforming Growth Factor beta; Transplantation, Heterologous