Biomedical subjects

Shioko Kimura

Publications and source records attributed to Shioko Kimura.

At least 19 recordsLinked to original sources

PubMed2007

GATA and Nkx factors synergistically regulate tissue-specific gene expression and development in vivo.

In vitro studies have suggested that members of the GATA and Nkx transcription factor families physically interact, and synergistically activate pulmonary epithelial- and cardiac-gene promoters. However, the relevance of this synergy has not been demonstrated in vivo. We show that Gata6-Titf1 (Gata6-Nkx2.1) double heterozygous (G6-Nkx DH) embryos and mice have severe defects in pulmonary epithelial differentiation and distal airway development, as well as reduced phospholipid production. The defects in G6-Nkx DH embryos and mice are similar to those observed in human neonates with respiratory distress syndromes, including bronchopulmonary dysplasia, and differential gene expression analysis reveals essential developmental pathways requiring synergistic regulation by both Gata6 and Titf1 (Nkx2.1). These studies indicate that Gata6 and Nkx2.1 act in a synergistic manner to direct pulmonary epithelial differentiation and development in vivo, providing direct evidence that interactions between these two transcription factor families are crucial for the development of the tissues in which they are co-expressed.

Alleles↗

PubMed2006

Deletion of the Ttf1 gene in differentiated neurons disrupts female reproduction without impairing basal ganglia function.

Thyroid transcription factor 1 (TTF1) [also known as Nkx2.1 (related to the NK-2 class of homeobox genes) and T/ebp (thyroid-specific enhancer-binding protein)], a homeodomain gene required for basal forebrain morphogenesis, remains expressed in the hypothalamus after birth, suggesting a role in neuroendocrine function. Here, we show an involvement of TTF1 in the control of mammalian puberty and adult reproductive function. Gene expression profiling of the nonhuman primate hypothalamus revealed that TTF1 expression increases at puberty. Mice in which the Ttf1 gene was ablated from differentiated neurons grew normally and had normal basal ganglia/hypothalamic morphology but exhibited delayed puberty, reduced reproductive capacity, and a short reproductive span. These defects were associated with reduced hypothalamic expression of genes required for sexual development and deregulation of a gene involved in restraining puberty. No extrapyramidal impairments associated with basal ganglia dysfunction were apparent. Thus, although TTF1 appears to fulfill only a morphogenic function in the ventral telencephalon, once this function is satisfied in the hypothalamus, TTF1 remains active as part of the transcriptional machinery controlling female sexual development.

Animals↗

PubMed2006

Identification and expression of alternative splice variants of the mouse Ppp1r3b gene in lung epithelial cells.

Hepatic glycogen-targeting subunit GL of protein phosphatase-1 (PP1), which is encoded by the gene Ppp1r3b, is one of a family of proteins that target PP1 to glycogen and regulate its activity on enzymes of glycogen metabolism. GL is primarily expressed in the liver where it promotes hepatic glycogen synthesis in response to insulin. Here, we show that GL is highly expressed in embryonic mouse lungs. GL utilizes two alternative promoters and 5' non-coding exons, which produce at least three alternatively spliced transcripts that encode identical proteins. Expression of GL in the lung is detectable as early as embryonic day (E) 12.5 and increases by E16.5, however, it declines before birth. In situ hybridization of mouse embryonic lung revealed that GL is expressed in bronchial epithelial cells. Thus, the temporal and spatial expression of GL in the lung suggests that it has a potential role in glycogen metabolism during lung development.

Alternative Splicing↗

PubMed2006

Peroxisome proliferator-activated receptor alpha activation during pregnancy severely impairs mammary lobuloalveolar development in mice.

To identify the potential functions of peroxisome proliferator-activated receptor alpha (PPARalpha) in skin development, transgenic mice were generated to target constitutively activated PPARalpha (VP16PPARalpha) to the stratified epithelia by use of the keratin K5 promoter. In addition to marked alterations in epidermal development, the transgenic mice had a severe defect in lactation during pregnancy resulting in 100% pup mortality. In this study, the alteration of mammary gland development in these transgenic mice was investigated. The results showed that expression of the VP16PPARalpha transgene during pregnancy resulted in impaired development of lobuloalveoli, which is associated with reduced proliferation and increased apoptosis of mammary epithelia. Mammary epithelia from transgenic mice also showed a significant reduction in the expression of beta-catenin and a down-regulation of one of its target genes, cyclin D1, which is thought to be required for lobuloalveolar development. Furthermore, upon PPARalpha ligand treatment, similar effects on lobuloalveolar development were observed in wild-type mice, but not in PPARalpha-null mice. These findings suggest that PPARalpha activation has a marked influence in mammary lobuloalveolar development.

Animals↗

PubMed2006

Regulation of mouse hepatic alpha-amino-beta-carboxymuconate-epsilon-semialdehyde decarboxylase, a key enzyme in the tryptophan-nicotinamide adenine dinucleotide pathway, by hepatocyte nuclear factor 4alpha and peroxisome proliferator-activated receptor alpha.

Nicotinamide adenine dinucleotide (NAD) plays a critical role in the maintenance of cellular energy homeostasis. alpha-Amino-beta-carboxymuconate-epsilon-semialdehyde decarboxylase (ACMSD) is the key enzyme regulating de novo synthesis of NAD from l-tryptophan (Trp), designated the Trp-NAD pathway. Acmsd gene expression was found to be under the control of both hepatocyte nuclear factor 4alpha (HNF4alpha) and peroxisome proliferator-activated receptor alpha (PPARalpha). Constitutive expression of ACMSD mRNA levels were governed by HNF4alpha and downregulated by activation of PPARalpha by the ligand Wy-14,643 ([4-chloro-6-(2,3-xylidino)-2-pyrimidinylthio]acetic acid]), as revealed by studies with hepatic HNF4alpha-null mice and PPARalpha-null mice, respectively. Transient transfection and electrophoretic mobility shift analyses showed an HNF4alpha binding site in the Acmsd gene promoter that directed transactivation of reporter gene constructs by HNF4alpha. The Acmsd promoter was not responsive to PPARalpha in transactivation assays. Wy-14,643 treatment decreased HNF4alpha protein levels in wild-type, but not PPARalpha-null, mouse livers, with no changes in HNF4alpha mRNA. These results show that Wy-14,643, through PPARalpha, post-transcriptionally down-regulates HNF4alpha protein levels, leading to reduced expression of the HNF4alpha target gene Acmsd.

Animals↗

PubMed2006

Thyroid-specific enhancer-binding protein/NKX2.1 is required for the maintenance of ordered architecture and function of the differentiated thyroid.

Thyroid-specific enhancer-binding protein (T/ebp)/Nkx2.1-null mouse thyroids degenerate by embryonic day (E) 12-13 through apoptosis whereas T/ebp/Nkx2.1-heterogyzgous mice exhibit hypothyroidism with elevated TSH levels. To understand the role of T/ebp/Nkx2.1 in the adult thyroid, a thyroid follicular cell-specific conditional knockout (KO) mouse line, T/ebp(fl/fl);TPO-Cre, was established that expresses Cre recombinase under the human thyroid peroxidase (TPO) gene promoter. These mice appeared to be healthy and exhibited loss of T/ebp/Nkx2.1 expression in many, but not all, thyroid follicular cells as determined by immunohistochemistry and real-time PCR, thus presenting a T/ebp-thyroid-conditional hypomorphic mice. Detailed analysis of the thyroids from T/ebp(fl/fl), T/ebp(fl/fl);TPO-Cre, and T/ebp(fl/ko) mice, where the latter mouse line is derived from crosses with the original T/ebp/Nkx2.1-heterozygous mice, revealed that T/ebp(fl/fl);TPO-Cre mice can be classified into two groups with different phenotypes: one having atrophic/degenerative thyroid follicles with frequent presence of adenomas and extremely high serum TSH levels, and the other having an altered thyroid structure with reduced numbers of extraordinary dilated follicles consisting of excessive numbers of follicular cells as compared with those usually found in the normal thyroid. The latter phenotype was also observed in aged T/ebp(fl/ko) mouse thyroids. In vitro three-dimensional thyroid primary cultures using thyroids from T/ebp(fl/fl);TPO-Cre, T/ebp(fl/ko), and T/ebp(fl/fl) mice, and the latter treated with recombinant adenovirus with and without Cre expression, demonstrated that only cells from T/ebp(fl/fl) mice without adeno-Cre treatment formed follicular structures. Taken together, these results suggest that T/ebp/Nkx2.1 is required for maintenance of the normal architecture and function of differentiated thyroids.

Alleles↗

PubMed2006

Hop functions downstream of Nkx2.1 and GATA6 to mediate HDAC-dependent negative regulation of pulmonary gene expression.

Hop is an unusual homeodomain protein that was first identified in the developing heart where it functions downstream of Nkx2.5 to modulate cardiac gene expression. Hop functions through interactions with histone deacetylase (HDAC) 2 to mediate repression of cardiac-specific genes, and recent studies show that HDAC activity and HDAC2 expression are decreased in people with chronic obstructive pulmonary disease. Here, we show that Hop is expressed in airway epithelium coincident with HDAC2, and expression is induced by the combination of dexamethasone and cAMP in parallel with induction of surfactant protein gene expression. Hop functions in the developing pulmonary airway, acting downstream of Nkx2.1 and GATA6, to negatively regulate surfactant protein expression. Loss of Hop expression in vivo results in defective type 2 pneumocyte development with increased surfactant production and disrupted alveolar formation. Thus Hop represents a novel regulator of pulmonary maturation that is induced by glucocorticoids to mediate functionally important HDAC-dependent negative feedback regulation.

Animals↗

PubMed2006

A common regulatory region functions bidirectionally in transcriptional activation of the human CYP1A1 and CYP1A2 genes.

The human CYP1A1 and CYP1A2 genes on chromosome 15 are orientated head-to-head and are separated by a 23-kilobase (kb) intergenic spacer region. Thus, the possibility exists for sharing common regulatory elements contained in the spacer region responsible for transcriptional activation and regulation of the CYP1A1 and CYP1A2 genes. In the present study, a reporter gene construct containing -22.4 kb of the 5'-flanking region of the CYP1A2 gene was found to support beta-naphthoflavone (BNF) and 3-methylchoranthrene (3-MC)-mediated transcriptional activation. The responsive region was also functional in directing activation of the CYP1A1 promoter, indicating that the region works bidirectionally to govern transcriptional activation of both CYP1A1 and CYP1A2. To simultaneously evaluate transcriptional activation of both genes, a dual reporter vector was developed in which the spacer region was inserted between two different reporter genes, firefly luciferase and secreted alkaline phosphatase. Transient transfection of the dual reporter vector in HepG2 cells revealed increases in both reporter activities after exposure of the cells to BNF and 3-MC. Deletion studies of the spacer region indicated that a region from -464 to -1829 of the CYP1A1 gene works bidirectionally to enhance the transcriptional activation of not only CYP1A1 but also CYP1A2. In addition, a negative bidirectional regulatory region was found to exist from -18,989 to -21,992 of the CYP1A1 gene. These data established that induction of human CYP1A1 and CYP1A2 is simultaneously controlled through bidirectional and common regulatory elements.

5' Flanking Region↗

PubMed2006

Uteroglobin-related protein 1 expression suppresses allergic airway inflammation in mice.

RATIONALE: Uteroglobin-related protein (UGRP) 1, which is highly expressed in the epithelial cells of the airways, has been suggested to play a role in lung inflammation. OBJECTIVES: The aim of study was to understand the effect of overexpressed UGRP1 on lung inflammation in a mouse model of allergic airway inflammation. METHODS: Ovalbumin-sensitized and -challenged mice, a model for allergic airway inflammation, were used in conjunction with recombinant adenovirus expressing UGRP1. MEASUREMENTS AND MAIN RESULTS: We demonstrated that intranasal administration of adeno-UGRP1 successfully delivered UGRP1 to the epithelial cells of airways and markedly reduced the number of infiltrating inflammatory cells, particularly eosinophils, in lung tissue as well as the level of proinflammatory cytokines such as interleukin (IL)-4, IL-5, and IL-13 in bronchoalveolar lavage fluids. The healed phase of inflammation was clearly seen in the peripheral areas of adeno-UGRP1-treated mouse lungs. CONCLUSION: These results demonstrate that UGRP1 can suppress inflammation in the mouse model of allergic airway inflammation. Based on this result, we propose UGRP1 as a novel therapeutic candidate for treating lung inflammation such as is found in asthma.

Adenoviridae↗

PubMed2006

Origin of the ultimobranchial body cyst: T/ebp/Nkx2.1 expression is required for development and fusion of the ultimobranchial body to the thyroid.

The ultimobranchial body (UBB) is an outpocketing of the fourth pharyngeal pouch that fuses with the thyroid diverticulum, giving rise to calcitonin-producing C-cells. In this study, we demonstrate that the UBB is composed of two types of cells: one expressing T/ebp/Nkx2.1 and the other expressing p63. The former cell type, accounting for a majority of the UBB, requires T/ebp/Nkx2.1 for their survival. In contrast, the p63-positive cells, even in the absence of T/ebp/Nkx2.1 expression, can proliferate and give rise to a vesicular structure that is lined by a monolayer of p63-negative cells, surrounded by a cluster and/or single layer of p63-positive cells, displaying the basal/stem cell phenotype. T/ebp/Nkx2.1 haploinsufficiency causes abnormal fusion of the UBB with the thyroid diverticulum, which stays as a cluster of C-cells around the vesicular structure, similar to the one observed in mice null for T/ebp/Nkx2.1 expression. These results demonstrate that T/ebp/Nkx2.1 plays a role in the survival of UBB cells, their dissemination into the thyroid diverticulum, and the formation of UBB-derived vesicular structure.

Animals↗

PubMed2006

Alterations in skin and stratified epithelia by constitutively activated PPARalpha.

Peroxisome proliferator-activated receptor (PPAR)alpha is a pleiotropic regulator in many cell types and has recently been implicated in skin homeostasis. To determine the role of PPARalpha in skin physiology, transgenic mice were generated using the tetracycline Tet-off regulatory system to target constitutively activated PPARalpha to the epidermis and other stratified epithelia by the bovine keratin K5 promoter. Expression of the transgene during early development resulted in postnatal lethality within 2 days after birth. A thin epidermis, few hair follicles, and abnormal development of the tongue were observed in neonatal transgenic mice. Early mortality was not observed when transgenic PPARalpha expression was diminished by administration of doxycycline (dox) to the mothers. The alterations noted in neonatal mice were not observed in adult mice upon re-expression of the PPARalpha transgene by withdrawing dox. Attenuated hyperplasia of interfollicular epidermis after topical application of the tumor promotor 12-O-tetradecanoyl-phorbol-13-acetate (TPA) was observed in adult mice expressing the PPARalpha transgene. In addition, expression of the PPARalpha transgene in mammary gland during pregnancy resulted in abnormal development of this organ and impaired lactation. Further investigations using primary keratinocytes revealed that expression of the transgene in keratinocytes resulted in increased differentiation and decreased proliferation, which may contribute to the observed phenotype in the transgenic mice. Thus, these results indicate that PPARalpha plays an important role in the development of stratified epithelia including skin, tongue, and mammary gland.

Animals↗

PubMed2005

Induction of uteroglobin-related protein 2 (Ugrp2) gene expression by the Th2 cytokines IL-4 and IL-13.

Uteroglobin-related proteins 1 and 2 (UGRP1 and -2) are thought to play important roles in inflammation and immunologic responses in the lung. In this study we demonstrate that IL-4 and IL-13 enhance Ugrp2 gene expression in the mouse transformed Clara cell line, mtCC, in a time- and dose-dependent manner. Addition of actinomycin D abrogated the IL-4- and IL-13-induced increase of Ugrp2 expression, demonstrating that this increase occurs at the transcriptional level. When mtCC cells were pretreated with IFN-gamma before the addition of IL-4 or IL-13, IL-4- and 13-induced Ugrp2 mRNA increase was markedly decreased. IL-4 and IL-13 induced phosphorylation of STAT6 in mtCC cells, which binds to the proximal STAT-binding element (SBE) in the Ugrp2 gene promoter, leading to transcriptional activation of this gene. Mutations of the proximal SBE abrogated the binding of activated STAT6 to this site and the IL-4-induced increase in Ugrp2 gene promoter activity. IFN-gamma-activated STAT1 binds to the same SBE in the Ugrp2 gene promoter to which STAT6 binds and decreases the binding of STAT6 to this site. Furthermore, an IL-4-induced increase in Ugrp2 expression was not observed in primary cultures of lung cells derived from STAT6-deficient mice. These results indicate that Ugrp2 expression is enhanced by IL-4 and IL-13 through STAT6 binding to the proximal SBE located in the Ugrp2 gene promoter.

Animals↗

PubMed2005

A mouse model demonstrates a multigenic origin of congenital hypothyroidism.

Congenital hypothyroidism with thyroid dysgenesis (TD) is a frequent human condition characterized by elevated levels of TSH in response to reduced thyroid hormone levels. Congenital hypothyroidism is a genetically heterogeneous disease. In the majority of cases studied, no causative mutations have been identified and very often the disease does not show a Mendelian transmission. However, in approximately 5% of cases, it can be a consequence of mutations in genes encoding the TSH receptor or the transcription factors TITF1, FOXE1, or PAX8. We report here that in mouse models, the combination of partial deficiencies in the Titf1 and Pax8 genes results in an overt TD phenotype that is absent in either of the singly deficient, heterozygous mice. The disease is characterized by a small thyroid gland, elevated levels of TSH, reduced thyroglobulin biosynthesis, and high occurrence of hemiagenesis. The observed phenotype is strain specific, and the pattern of transmission indicates that at least two other genes, in addition to Titf1 and Pax8, are necessary to generate the condition. These results show that TD can be of multigenic origin in mice and strongly suggest that a similar pathogenic mechanism may be observed in humans.

Animals↗

PubMed2005

Ventromedial hypothalamic nucleus-specific enhancer of Ad4BP/SF-1 gene.

Ad4BP/SF-1 [Ad4 binding protein/steroidogenic factor-1 (designated NR5A1)] is a transcription factor essential for animal reproduction. Based on the phenotypes observed in gene-disrupted mice, Ad4BP/SF-1 is thought to be involved in establishment of the hypothalamic-pituitary-gonadal axis. However, the mechanisms underlying tissue-specific expression of Ad4BP/SF-1 are largely unknown. Here, we investigated the cis-regulatory regions of the mouse Ad4BP/SF-1 gene by transgenic mouse assays, and identified a ventromedial hypothalamic nucleus (VMH)-specific enhancer. The enhancer localized in intron 6 is highly conserved between mouse, human, and chick. The enhancer has the potential to reproduce endogenous gene expression from the fetal ventromedial diencephalon to the adult VMH. The VMH enhancer was characterized by the presence of suppressive and activating elements. Mutation of the former element resulted in ectopic lacZ reporter gene expression in an area dorsal to the intrinsic expression domain and in the ventricular zone, whereas mutations in the latter containing ATTA motifs led to the disappearance of the reporter gene expression, suggesting the involvement of homeobox proteins. Using nuclear extracts prepared from the adult hypothalami, EMSAs identified specific protein binding to the activating elements but not to the suppressive element.

Animals↗

PubMed2005

Induction of uteroglobin-related protein 2 (Ugrp2) expression by EGF and TGFalpha.

Uteroglobin-related protein 2 (UGRP2) is thought to play a role in inflammation and/or epithelial cell differentiation in the lung. Induction of Ugrp2 mRNA expression by epidermal growth factor (EGF) and transforming growth factor alpha was examined using mouse transformed lung Clara cell-derived mtCC cells. The EGF-induced increase of Ugrp2 occurred at the transcriptional level that required the EGF receptor and the activation of the ERK-MAPK and phosphoinositide-3 kinase pathways.

Animals↗

PubMed2005

Interleukin-5 reduces the expression of uteroglobin-related protein (UGRP) 1 gene in allergic airway inflammation.

Airway inflammation is thought to play a major role in the pathogenesis of bronchial asthma. The precise role of individual inflammatory cells, mediator and asthma related genes in allergic lung diseases is not completely understood. The uteroglobin-related protein (UGRP) 1 was proposed to be an asthma candidate gene and play a role in regulating lung inflammation, however its precise function in the airways remains obscure. In this investigation, we used a mouse model of allergic airway inflammation to establish a relationship between UGRP 1 and IL-5 in airway inflammation. Ovalbumin (OVA) challenged mice demonstrate eosinophilia in airway tissues and high levels of IL-5 in bronchoalveolar lavage (BAL) fluid analogous to that found in bronchial asthma. Interestingly, these "OVA-challenged" mice show down-regulation of Ugrp1 expression as compared with the control group. Regression analysis further demonstrates a significant negative correlation between Ugrp1 mRNA expression in the lung and IL-5 levels in BAL fluid with r = 0.948 and P < 0.0001 when IL-5 levels were normalized by log transformation. Intranasal instillation of IL-5 to mice revealed an inhibitory effect of IL-5 on the expression of Ugrp1 mRNA. Together, these results indicate an involvement of IL-5 in the down-regulation of Ugrp1 expression in airway inflammation such as allergic asthma disease.

Administration, Intranasal↗

PubMed2005

Differential metabolism of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in mice humanized for CYP1A1 and CYP1A2.

The procarcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is the most abundant heterocyclic amine formed during the cooking of foods. Metabolism of PhIP by CYP1A2 differs substantially between humans and rodents, with more N2-hydroxylation (activation) and less 4'-hydroxylation (detoxication) in humans. Therefore, the human response to PhIP and other heterocyclic amine exposure may not be accurately reflected in the laboratory rodent. By generating mouse models expressing the human genes, species differences in heterocyclic amine metabolism can be addressed. Two transgenic mouse lines were developed, one expressing the human CYP1A1 CYP1A2 transgene in a mouse Cyp1a1-null background (hCYP1A1) and another expressing human CYP1A1 CYP1A2 in a mouse Cyp1a2-null background (hCYP1A2). Expression of human CYP1A2 protein was detected in the liver and also at considerably lower levels in extrahepatic tissues such as lung, kidney, colon, and heart. In the hCYP1A1 and hCYP1A2 mice, 3-methylcholanthrene (3-MC) induced both human CYP1A1 and CYP1A2 protein in the liver. Differences in the metabolism of the heterocyclic amine PhIP were observed between wild-type and hCYP1A2 mice. PhIP was preferentially metabolized by N2-hydroxylation in hCYP1A2 mice, whereas in wild-type mice, 4'-hydroxylation was the predominant pathway. Since the N2-hydroxylation pathway for PhIP metabolism has been reported to be predominant in humans, these results illustrate the potential effectiveness of using these transgenic, humanized mice as models for determining human health risks to PhIP and other heterocyclic amines instead of wild-type mice.

Animals↗

PubMed2005

Directed differentiation of neural cells to hypothalamic dopaminergic neurons.

Hypothalamic neurons play a key role in homeostasis, yet little is known about their differentiation. Here, we demonstrate that Shh and Bmp7 from the adjacent prechordal mesoderm govern hypothalamic neural fate, their sequential action controlling hypothalamic dopaminergic neuron generation in a Six3-dependent manner. Our data suggest a temporal distinction in the requirement for the two signals. Shh acts early to specify dopaminergic neurotransmitter phenotype. Subsequently, Bmp7 acts on cells that are ventralised by Shh, establishing aspects of hypothalamic regional identity in late-differentiating/postmitotic cells. The concerted actions of Shh and Bmp7 can direct mouse embryonic stem cell-derived neural progenitor cells to a hypothalamic dopaminergic fate ex vivo.

Animals↗