RESUMEN
It is well known that maternal diabetes causes various congenital malformations. Although there are many reports that folic acid (FA) administration in pregnancy reduces the risk of birth defects including neural tube defects (NTDs), a precise analysis on the preventive effect of FA against diabetic embryopathy has not been done yet. In this study, we analyzed the preventive effects of FA on congenital malformations including NTDs, cardiovascular, and skeletal malformations using a diabetic mouse model. Female mice were rendered hyperglycemic by streptozotocin and then mated. Pregnant diabetic mice were treated daily with FA (3 mg/kg body weight) or saline between gestational days (GD) 6 and 10. On GD 18, fetuses were examined for congenital malformations. FA did not affect plasma glucose levels. In the DM control group, the incidence of NTDs, cardiovascular, and skeletal malformations was 28.4%, 28.5%, and 29.7%, respectively. In the FA-treated group, the corresponding proportions reduced to 6.0%, 2.5% and 12.5%, respectively. A whole-mount TUNEL revealed an increased apoptosis in the hindbrain region of embryos from DM control group on day 9.5, and the apoptosis was decreased by FA treatment. Maternal plasma homocysteine levels on GD 9.5 were significantly lowered in DM control group compared with those in non-DM group, and FA treatment did not show a significant effect. These results indicate that FA is effective for the prevention of various diabetic embryopathy including NTDs, cardiovascular, and skeletal malformations, and suggested that this effect is independent from homocysteine metabolism and possibly mediated by decreasing the abnormal apoptosis during organogenesis.
Asunto(s)
Anomalías Congénitas/prevención & control , Diabetes Mellitus Experimental/tratamiento farmacológico , Ácido Fólico/uso terapéutico , Embarazo en Diabéticas , Animales , Glucemia/análisis , Peso Corporal , Anomalías Congénitas/etiología , Diabetes Mellitus Experimental/sangre , Diabetes Mellitus Experimental/complicaciones , Evaluación Preclínica de Medicamentos , Embrión de Mamíferos , Femenino , Peso Fetal/efectos de los fármacos , Masculino , Intercambio Materno-Fetal/efectos de los fármacos , Ratones , Ratones Endogámicos ICR , Embarazo , Resultado del Embarazo , Embarazo en Diabéticas/sangre , Embarazo en Diabéticas/patología , Embarazo en Diabéticas/veterinaria , EstreptozocinaRESUMEN
OBJECTIVE: ZAKI-4 was identified as a thyroid hormone-responsive gene in cultured human fibroblasts. A single ZAKI-4 gene encodes two isoforms, ZAKI-4 alpha and beta, both inhibiting calcineurin activity. ZAKI-4 alpha and beta differ at their N termini, and show distinct distribution profiles in human tissues. The aim of this study was to elucidate the organization of the mouse ZAKI-4 gene and to determine the effect of thyroid hormone on the expression of ZAKI-4 isoforms in vivo. DESIGN: We cloned mouse homologues of human ZAKI-4 alpha and beta cDNA. Fluorescence in situ hybridization and bioinformatics analysis were employed to determine the gene organization. The effect of thyroid hormone on the expression of ZAKI-4 isoforms in mouse brain and heart was also studied. METHODS: Total RNA extracted from mouse cerebellum was used to clone ZAKI-4 alpha and beta cDNAs by RT-PCR followed by rapid amplification of cDNA ends. Mice were rendered hypothyroid by feeding a low iodine diet supplemented with propylthiouracil for 2 weeks. In one group (hyperthyroid) L-T(3) was injected i.p. for the last 4 days whereas another group (hypothyroid) received vehicle only. Non-treated mice were controls. RESULTS AND CONCLUSION: Mouse ZAKI-4 alpha and beta cDNAs were highly homologous to the human isoforms. The gene was mapped on chromosome 17qC, syntenic to human chromosome 6 where the human ZAKI-4 gene is located. As observed in human, ZAKI-4 alpha mRNA was expressed only in brain whereas beta mRNA was distributed in other tissues as well, such as heart and skeletal muscle. ZAKI-4 alpha mRNA was lower in the cerebral cortex of hypothyroid mice. Injection of L-T(3) caused an increase in ZAKI-4 beta mRNA in heart; however, expression of neither ZAKI-4 alpha nor beta mRNA was influenced by thyroid status in other tissues. These results indicate that expression of ZAKI-4 alpha and beta isoforms is regulated by thyroid hormone in vivo, and the regulation is isoform- and tissue-specific.
Asunto(s)
Regulación de la Expresión Génica/fisiología , Hipotiroidismo/genética , Proteínas Musculares/genética , Proteínas , Triyodotironina/farmacología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Encéfalo/metabolismo , Encéfalo/fisiología , Inhibidores de la Calcineurina , Mapeo Cromosómico , Regulación de la Expresión Génica/efectos de los fármacos , Hipotiroidismo/metabolismo , Hibridación Fluorescente in Situ , Péptidos y Proteínas de Señalización Intracelular , Masculino , Ratones , Ratones Endogámicos ICR , Datos de Secuencia Molecular , Proteínas Musculares/biosíntesis , Miocardio/metabolismo , Isoformas de Proteínas , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Alineación de Secuencia , Distribución TisularRESUMEN
The house musk shrew Suncus murinus (Insectivora: Soricidae) has been reported as having low thyroxine to 3,3'5-triiodothyronine (T(3)) converting activity in liver and kidney homogenates and was assumed to be type 1 iodothyronine deiodinase (D1)-deficient. To study whether this is due to structural abnormality of shrew D1, we cloned the cDNA and characterized the enzyme. The deduced amino acid sequence of shrew D1 was found to be highly homologous to other known D1s and the enzyme itself to have similar catalytic activity. However, unlike in other species, the D1 activity was detected only in liver. Moreover, the D1 activity in liver of the shrew was less than half of that in rat liver and its expression was not up-regulated by T(3). In contrast, a very high activity of D2 was demonstrated in brain and brown adipose tissue. The present study also revealed that the serum level of T(3) in the shrew was in the same range as these in other mammals. These results suggest that D2 contributes to the production and maintenance of T(3) levels in the house musk shrew.
Asunto(s)
Clonación Molecular , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Yoduro Peroxidasa/genética , Musarañas/genética , Triyodotironina/farmacología , Tejido Adiposo Pardo/enzimología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Núcleo Celular/química , Corteza Cerebral/enzimología , ADN Complementario , Evolución Molecular , Yoduro Peroxidasa/análisis , Yoduro Peroxidasa/química , Riñón/enzimología , Hígado/enzimología , Hígado/ultraestructura , Datos de Secuencia Molecular , Hipófisis/enzimología , ARN Mensajero/análisis , Receptores de Hormona Tiroidea/análisis , Musarañas/metabolismo , Glándula Tiroides/enzimología , Tirotropina/sangre , Tiroxina/sangre , Distribución Tisular , Triyodotironina/sangre , Triyodotironina Inversa/sangreRESUMEN
Thyroid hormone exerts its biological effect by binding to a TR. Both liganded and unliganded TRs regulate the transcription of T(3)-responsive genes. Cofactors with activating or repressing function modulate the transcriptional regulation by TRs. We showed that steroid receptor coactivator 1 (SRC-1)-deficient mice (SRC-1(-/-)) exhibit partial resistance to thyroid hormone at the level of the pituitary thyrotrophs. To determine whether SRC-1 deficiency affects globally T(3)-dependent transcriptional regulation, we studied the effects of thyroid hormone deprivation and replacement on the expression of several genes in different tissues of SRC-1(-/-) and wild-type mice (SRC-1(+/+)). Thyroid hormone deficiency was induced by a low iodine diet (LoI) supplemented with propylthiouracil (PTU) for 2 wk. L-T(3) was injected ip for the last 4 d in one group (PTU+T(3) group), and another group (PTU group) received only vehicle. Levels of mRNAs for T(3)-responsive genes were determined by Northern blotting: GH and TSH beta in pituitary; type 1 iodothyronine 5'-deiodinase, spot 14 (S14), and malic enzyme in liver; and sarcoplasmic reticulum calcium adenosine triphosphatase 2 and myosin heavy chain alpha and beta in heart. Serum parameters, TSH, total cholesterol, creatine kinase, and alkaline phosphatase (AP), were also measured. Hypothyroidism produced a comparable increase in TSH beta mRNA in both genotypes, but its suppression by L-T(3) was attenuated in SRC-1(-/-) mice. In contrast, hypothyroidism failed to reduce S14 mRNA levels in SRC-1(-/-) mice. As a consequence, the response to L-T(3) was not observed in these mice. SRC-1 deficiency had no effect on the expression of the rest of the T(3)-responsive genes examined. Of the four serum parameters, the T(3)-mediated decrease in TSH and changes in AP were attenuated in SRC-1(-/-) mice. We conclude that SRC-1 deficiency altered the expression of only some of the T(3)-responsive genes. SRC-1 appears to be involved not only in transcriptional activation by liganded TRs, but also in the suppression by liganded or unliganded TRs. Some of the effects of SRC-1 may be TR isoform specific.