Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 15 de 15
Filtrar
Más filtros










Base de datos
Intervalo de año de publicación
1.
Genetics ; 2021 Nov 22.
Artículo en Inglés | MEDLINE | ID: mdl-34850872

RESUMEN

Primary cilia are sensory and signaling hubs with a protein composition that is distinct from the rest of the cell due to the barrier function of the transition zone (TZ) at the base of the cilium. Protein transport across the TZ is mediated in part by the BBSome, and mutations disrupting TZ and BBSome proteins cause human ciliopathy syndromes. Ciliopathies have phenotypic variability even among patients with identical genetic variants, suggesting a role for modifier loci. To identify potential ciliopathy modifiers, we performed a mutagenesis screen on nphp-4 mutant Caenorhabditis elegans and uncovered a novel allele of bbs-5. Nphp-4;bbs-5 double mutant worms have phenotypes not observed in either individual mutant strain. To test whether this genetic interaction is conserved, we also analyzed zebrafish and mouse mutants. While Nphp4 mutant zebrafish appeared overtly normal, Bbs5 mutants exhibited scoliosis. When combined, Nphp4;Bbs5 double mutant zebrafish did not exhibit synergistic effects, but the lack of a phenotype in Nphp4 mutants makes interpreting these data difficult. In contrast, Nphp4;Bbs5 double mutant mice were not viable and there were fewer mice than expected carrying three mutant alleles. In addition, postnatal loss of Bbs5 in mice using a conditional allele compromised survival when combined with an Nphp4 allele. As cilia are still formed in the double mutant mice, the exacerbated phenotype is likely a consequence of disrupted ciliary signaling. Collectively, these data support an evolutionarily conserved genetic interaction between Bbs5 and Nphp4 alleles that may contribute to the variability in ciliopathy phenotypes.

2.
Front Cell Dev Biol ; 9: 705182, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34970537

RESUMEN

Atxn10 is a gene known for its role in cytokinesis and is associated with spinocerebellar ataxia (SCA10), a slowly progressing cerebellar syndrome caused by an intragenic pentanucleotide repeat expansion. Atxn10 is also implicated in the ciliopathy syndromes nephronophthisis (NPHP) and Joubert syndrome (JBTS), which are caused by the disruption of cilia function leading to nephron loss, impaired renal function, and cerebellar hypoplasia. How Atxn10 disruption contributes to these disorders remains unknown. Here, we generated Atxn10 congenital and conditional mutant mouse models. Our data indicate that while ATXN10 protein can be detected around the base of the cilium as well as in the cytosol, its loss does not cause overt changes in cilia formation or morphology. Congenital loss of Atxn10 results in embryonic lethality around E10.5 associated with pericardial effusion and loss of trabeculation. Similarly, tissue-specific loss of ATXN10 in the developing endothelium (Tie2-Cre) and myocardium (cTnT-Cre) also results in embryonic lethality with severe cardiac malformations occurring in the latter. Using an inducible Cagg-CreER to disrupt ATXN10 systemically at postnatal stages, we show that ATXN10 is also required for survival in adult mice. Loss of ATXN10 results in severe pancreatic and renal abnormalities leading to lethality within a few weeks post ATXN10 deletion in adult mice. Evaluation of these phenotypes further identified rapid epithelial-to-mesenchymal transition (EMT) in these tissues. In the pancreas, the phenotype includes signs of both acinar to ductal metaplasia and EMT with aberrant cilia formation and severe defects in glucose homeostasis related to pancreatic insufficiency or defects in feeding or nutrient intake. Collectively, this study identifies ATXN10 as an essential protein for survival.

4.
Am J Physiol Renal Physiol ; 321(2): F162-F169, 2021 08 01.
Artículo en Inglés | MEDLINE | ID: mdl-34180717

RESUMEN

Kidney resident macrophages (KRMs) are involved in maintaining renal homeostasis and in controlling the pathological outcome of acute kidney injury and cystic kidney disease in mice. In adult mice, KRMs maintain their population through self-renewal with little or no input from the peripheral blood. Despite recent data suggesting that a transcriptionally similar population of KRM-like cells is present across species, the idea that they are self-renewing and minimally dependent on peripheral blood input in other species has yet to be proven due to the lack of an appropriate model and cross-species expression markers. In this study, we used our recently identified cross-species KRM cell surface markers and parabiosis surgery in inbred Lewis rats to determine if rat KRMs are maintained independent of peripheral blood input, similar to their mouse counterparts. Flow cytometry analysis indicated that parabiosis surgery in the rat results in the establishment of chimerism of T/B cells, neutrophils, and monocyte-derived infiltrating macrophages in the blood, spleen, and kidney 3 wk after parabiosis surgery. Analysis of KRMs using the cell surface markers CD81 and C1q indicated that these cells have minimal chimerism and, therefore, receive little input from the peripheral blood. These data indicate that KRM properties are conserved in at least two different species.NEW & NOTEWORTHY In this report, we performed parabiosis surgery on inbred Lewis rats and showed that rat kidney resident macrophages (KRMs), identified using our novel cross-species markers, are minimally dependent on peripheral blood input. Thus, for the first time, to our knowledge, we confirm that a hallmark of mouse KRMs is also present in KRMs isolated from another species.


Asunto(s)
Riñón/citología , Macrófagos/citología , Monocitos/citología , Animales , Femenino , Masculino , Parabiosis , Ratas , Ratas Endogámicas Lew , Bazo/citología
5.
Dev Dyn ; 2021 Mar 16.
Artículo en Inglés | MEDLINE | ID: mdl-33728725

RESUMEN

BACKGROUND: Genetic tools to study gene function and the fate of cells in the anterior limb bud are very limited. RESULTS: We describe a transgenic mouse line expressing CreERT2 from the Aristaless-like 4 (Alx4) promoter that induces recombination in the anterior limb. Cre induction at embryonic day 8.5 revealed that Alx4-CreERT2 labeled cells using the mTmG Cre reporter contributed to anterior digits I to III as well as the radius of the forelimb. Cre activity is expanded further along the AP axis in the hindlimb than in the forelimb resulting in some Cre reporter cells contributing to digit IV. Induction at later time points labeled cells that become progressively restricted to more anterior digits and proximal structures. Comparison of Cre expression from the Alx4 promoter transgene with endogenous Alx4 expression reveals Cre expression is slightly expanded posteriorly relative to the endogenous Alx4 expression. Using Alx4-CreERT2 to induce loss of intraflagellar transport 88 (Ift88), a gene required for ciliogenesis, hedgehog signaling, and limb patterning, did not cause overt skeletal malformations. However, the efficiency of deletion, time needed for Ift88 protein turnover, and for cilia to regress may hinder using this approach to analyze cilia in the limb. Alx4-CreERT2 is also active in the mesonephros and nephric duct that contribute to the collecting tubules and ducts of the adult nephron. Embryonic activation of the Alx4-CreERT2 in the Ift88 conditional line results in cyst formation in the collecting tubules/ducts. CONCLUSION: Overall, the Alx4-CreERT2 line will be a new tool to assess cell fates and analyze gene function in the anterior limb, mesonephros, and nephric duct.

6.
Hum Mol Genet ; 30(3-4): 234-246, 2021 04 26.
Artículo en Inglés | MEDLINE | ID: mdl-33560420

RESUMEN

Primary cilia are critical sensory and signaling compartments present on most mammalian cell types. These specialized structures require a unique signaling protein composition relative to the rest of the cell to carry out their functions. Defects in ciliary structure and signaling result in a broad group of disorders collectively known as ciliopathies. One ciliopathy, Bardet-Biedl syndrome (BBS; OMIM 209900), presents with diverse clinical features, many of which are attributed to defects in ciliary signaling during both embryonic development and postnatal life. For example, patients exhibit obesity, polydactyly, hypogonadism, developmental delay and skeletal abnormalities along with sensory and cognitive deficits, but for many of these phenotypes it is uncertain, which are developmental in origin. A subset of BBS proteins assembles into the core BBSome complex, which is responsible for mediating transport of membrane proteins into and out of the cilium, establishing it as a sensory and signaling hub. Here, we describe two new mouse models for BBS resulting from a targeted LacZ gene trap allele (Bbs5-/-) that is a predicted congenital null mutation and conditional (Bbs5flox/flox) allele of Bbs5. Bbs5-/- mice develop a complex phenotype consisting of increased pre-weaning lethality craniofacial and skeletal defects, ventriculomegaly, infertility and pituitary anomalies. Utilizing the conditional allele, we show that the male fertility defects, ventriculomegaly and pituitary abnormalities are only present when Bbs5 is disrupted prior to postnatal day 7, indicating a developmental origin. In contrast, mutation of Bbs5 results in obesity, independent of the age of Bbs5 loss.


Asunto(s)
Síndrome de Bardet-Biedl/metabolismo , Proteínas del Citoesqueleto/genética , Modelos Animales de Enfermedad , Mutación , Proteínas de Unión a Fosfato/genética , Hipófisis/anomalías , Animales , Síndrome de Bardet-Biedl/genética , Síndrome de Bardet-Biedl/patología , Síndrome de Bardet-Biedl/fisiopatología , Proteínas del Citoesqueleto/metabolismo , Masculino , Ratones , Fenotipo , Proteínas de Unión a Fosfato/metabolismo , Hipófisis/crecimiento & desarrollo , Hipófisis/metabolismo
7.
Invest Ophthalmol Vis Sci ; 61(10): 17, 2020 08 03.
Artículo en Inglés | MEDLINE | ID: mdl-32776140

RESUMEN

Purpose: To identify the role of the BBSome protein Bardet-Biedl syndrome 5 (BBS5) in photoreceptor function, protein trafficking, and structure using a congenital mutant mouse model. Methods: Bbs5-/- mice (2 and 9 months old) were used to assess retinal function and morphology. Hematoxylin and eosin staining of retinal sections was performed to visualize histology. Electroretinography was used to analyze rod and cone photoreceptor function. Retinal protein localization was visualized using immunofluorescence (IF) within retinal cryosections. TUNEL staining was used to quantify cell death. Transmission electron microscopy (TEM) was used to examine retinal ultrastructure. Results: In the Bbs5-/- retina, there was a significant loss of nuclei in the outer nuclear layer accompanied by an increase in cell death. Through electroretinography, Bbs5-/- mice showed complete loss of cone photoreceptor function. IF revealed mislocalization of the cone-specific proteins M- and S-opsins, arrestin-4, CNGA3, and GNAT2, as well as a light-dependent arrestin-1 mislocalization, although perpherin-2 was properly localized. TEM revealed abnormal outer segment disk orientation in Bbs5-/-. Conclusions: Collectively, these data suggest that, although BBS5 is a core BBSome component expressed in all ciliated cells, its role within the retina mediates specific photoreceptor protein cargo transport. In the absence of BBS5, cone-specific protein mislocalization and a loss of cone photoreceptor function occur.


Asunto(s)
Proteínas Portadoras/metabolismo , Proteínas del Citoesqueleto/fisiología , Proteínas de Unión a Fosfato/fisiología , Células Fotorreceptoras Retinianas Conos/metabolismo , Degeneración Retiniana/metabolismo , Segmento Externo de las Células Fotorreceptoras Retinianas/metabolismo , Animales , Western Blotting , Proteínas del Citoesqueleto/metabolismo , Modelos Animales de Enfermedad , Electrorretinografía , Etiquetado Corte-Fin in Situ , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microscopía Electrónica de Transmisión , Microscopía Fluorescente , Opsinas/metabolismo , Proteínas de Unión a Fosfato/metabolismo , Transporte de Proteínas , Células Fotorreceptoras Retinianas Conos/ultraestructura , Degeneración Retiniana/patología , Segmento Externo de las Células Fotorreceptoras Retinianas/ultraestructura , Células Fotorreceptoras Retinianas Bastones/metabolismo , Células Fotorreceptoras Retinianas Bastones/ultraestructura
8.
Am J Physiol Renal Physiol ; 316(3): F463-F472, 2019 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-30600684

RESUMEN

Heterozygosity for human polycystic kidney and hepatic disease 1 ( PKHD1) mutations was recently associated with cystic liver disease and radiographic findings resembling medullary sponge kidney (MSK). However, the relevance of these associations has been tempered by a lack of cystic liver or renal disease in heterozygous mice carrying Pkhd1 gene trap or exon deletions. To determine whether heterozygosity for a smaller Pkhd1 defect can trigger cystic renal disease in mice, we generated and characterized mice with the predicted truncating Pkhd1C642* mutation in a region corresponding to the middle of exon 20 cluster of five truncating human mutations (between PKHD1G617fs and PKHD1G644*). Mouse heterozygotes or homozygotes for the Pkhd1C642* mutation did not have noticeable liver or renal abnormalities on magnetic resonance images during their first weeks of life. However, when aged to ~1.5 yr, the Pkhd1C642* heterozygotes developed prominent cystic liver changes; tissue analyses revealed biliary cysts and increased number of bile ducts without signs of congenital hepatic fibrosis-like portal field inflammation and fibrosis that was seen in Pkhd1C642* homozygotes. Interestingly, aged female Pkhd1C642* heterozygotes, as well as homozygotes, developed radiographic changes resembling MSK. However, these changes correspond to proximal tubule ectasia, not an MSK-associated collecting duct ectasia. In summary, by demonstrating that cystic liver and kidney abnormalities are triggered by heterozygosity for the Pkhd1C642* mutation, we provide important validation for relevant human association studies. Together, these investigations indicate that PKHD1 mutation heterozygosity (predicted frequency 1 in 70 individuals) is an important underlying cause of cystic liver disorders and MSK-like manifestations in a human population.


Asunto(s)
Quistes/diagnóstico por imagen , Enfermedades Renales/diagnóstico por imagen , Túbulos Renales Proximales/diagnóstico por imagen , Hepatopatías/diagnóstico por imagen , Riñón Esponjoso Medular/diagnóstico por imagen , Receptores de Superficie Celular/metabolismo , Animales , Quistes/genética , Quistes/metabolismo , Diagnóstico Diferencial , Dilatación Patológica/diagnóstico por imagen , Dilatación Patológica/genética , Dilatación Patológica/metabolismo , Modelos Animales de Enfermedad , Enfermedades Renales/genética , Enfermedades Renales/metabolismo , Túbulos Renales Proximales/metabolismo , Hepatopatías/genética , Hepatopatías/metabolismo , Imagen por Resonancia Magnética , Riñón Esponjoso Medular/genética , Riñón Esponjoso Medular/metabolismo , Ratones , Ratones Noqueados , Receptores de Superficie Celular/genética
9.
FASEB J ; 33(1): 1440-1455, 2019 01.
Artículo en Inglés | MEDLINE | ID: mdl-30133325

RESUMEN

The transition zone (TZ) is a domain at the base of the cilium that is involved in maintaining ciliary compartment-specific sensory and signaling activity by regulating cilia protein composition. Mutations in TZ proteins result in cilia dysfunction, often causing pleiotropic effects observed in a group of human diseases classified as ciliopathies. The purpose of this study is to describe the importance of the TZ component Meckel-Grüber syndrome 6 ( Mks6) in several organ systems and tissues regarding ciliogenesis and cilia maintenance using congenital and conditional mutant mouse models. Similar to MKS, congenital loss of Mks6 is embryonic lethal, displaying cilia loss and altered cytoskeletal microtubule modifications but only in specific cell types. Conditional Mks6 mutants have a variable cystic kidney phenotype along with severe retinal degeneration with mislocalization of phototransduction cascade proteins. However, other phenotypes, such as anosmia and obesity, which are typically associated with cilia and TZ dysfunction, were not evident. These data indicate that despite Mks6 being a core TZ component, it has tissue- or cell type-specific functions important for cilia formation and cilia sensory and signaling activities. Lewis, W. R., Bales, K. L., Revell, D. Z., Croyle, M. J., Engle, S. E., Song, C. J., Malarkey, E. B., Uytingco, C. R., Shan, D., Antonellis, P. J., Nagy, T. R., Kesterson, R. A., Mrug, M. M., Martens, J. R., Berbari, N. F., Gross, A. K., Yoder, B. K. Mks6 mutations reveal tissue- and cell type-specific roles for the cilia transition zone.


Asunto(s)
Cilios/metabolismo , Proteínas del Citoesqueleto/genética , Mutación , Acetilación , Animales , Trastornos de la Motilidad Ciliar/genética , Citoplasma/metabolismo , Encefalocele/genética , Femenino , Genes Letales , Enfermedades Renales Quísticas/genética , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Trastornos del Olfato/genética , Fenotipo , Enfermedades Renales Poliquísticas/genética , Degeneración Retiniana/genética , Retinitis Pigmentosa/genética , Tubulina (Proteína)/metabolismo , Aumento de Peso/genética
10.
PLoS One ; 9(9): e106576, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-25184295

RESUMEN

It has been known for decades that neurons throughout the brain possess solitary, immotile, microtubule based appendages called primary cilia. Only recently have studies tried to address the functions of these cilia and our current understanding remains poor. To determine if neuronal cilia have a role in behavior we specifically disrupted ciliogenesis in the cortex and hippocampus of mice through conditional deletion of the Intraflagellar Transport 88 (Ift88) gene. The effects on learning and memory were analyzed using both Morris Water Maze and fear conditioning paradigms. In comparison to wild type controls, cilia mutants displayed deficits in aversive learning and memory and novel object recognition. Furthermore, hippocampal neurons from mutants displayed an altered paired-pulse response, suggesting that loss of IFT88 can alter synaptic properties. A variety of other behavioral tests showed no significant differences between conditional cilia mutants and controls. This type of conditional allele approach could be used to distinguish which behavioral features of ciliopathies arise due to defects in neural development and which result from altered cell physiology. Ultimately, this could lead to an improved understanding of the basis for the cognitive deficits associated with human cilia disorders such as Bardet-Biedl syndrome, and possibly more common ailments including depression and schizophrenia.


Asunto(s)
Cilios/metabolismo , Miedo , Aprendizaje por Laberinto , Neurogénesis/genética , Animales , Síndrome de Bardet-Biedl/genética , Síndrome de Bardet-Biedl/patología , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Cilios/genética , Depresión/genética , Depresión/patología , Hipocampo/crecimiento & desarrollo , Hipocampo/metabolismo , Hipocampo/patología , Humanos , Ratones , Ratones Noqueados , Neuronas/metabolismo , Neuronas/patología , Esquizofrenia/genética , Esquizofrenia/patología , Proteínas Supresoras de Tumor/genética
11.
Cilia ; 2(1): 8, 2013 Jul 03.
Artículo en Inglés | MEDLINE | ID: mdl-23819925

RESUMEN

BACKGROUND: Cilia are found on nearly every cell type in the mammalian body, and have been historically classified as either motile or immotile. Motile cilia are important for fluid and cellular movement; however, the roles of non-motile or primary cilia in most tissues remain unknown. Several genetic syndromes, called the ciliopathies, are associated with defects in cilia structure or function and have a wide range of clinical presentations. Much of what we know about the formation and maintenance of cilia comes from model systems like C. elegans and Chalmydomonas. Studies of mammalian cilia in live tissues have been hampered by difficulty visualizing them. RESULTS: To facilitate analyses of mammalian cilia function we generated an inducible CiliaGFP mouse by targeting mouse cDNA encoding a cilia-localized protein somatostatin receptor 3 fused to GFP (Sstr3::GFP) into the ROSA26 locus. In this system, Sstr3::GFP is expressed from the ubiquitous ROSA26 promoter after Cre mediated deletion of an upstream Neo cassette flanked by lox P sites. Fluorescent cilia labeling was observed in a variety of live tissues and after fixation. Both cell-type specific and temporally regulated cilia labeling were obtained using multiple Cre lines. The analysis of renal cilia in anesthetized live mice demonstrates that cilia commonly lay nearly parallel to the apical surface of the tubule. In contrast, in more deeply anesthetized mice the cilia display a synchronized, repetitive oscillation that ceases upon death, suggesting a relationship to heart beat, blood pressure or glomerular filtration. CONCLUSIONS: The ability to visualize cilia in live samples within the CiliaGFP mouse will greatly aid studies of ciliary function. This mouse will be useful for in vivo genetic and pharmacological screens to assess pathways regulating cilia motility, signaling, assembly, trafficking, resorption and length control and to study cilia regulated physiology in relation to ciliopathy phenotypes.

12.
Development ; 138(9): 1675-85, 2011 May.
Artículo en Inglés | MEDLINE | ID: mdl-21429982

RESUMEN

Skin and hair follicle morphogenesis and homeostasis require the integration of multiple signaling pathways, including Hedgehog (Hh) and Wingless (Wnt), and oriented cell divisions, all of which have been associated with primary cilia. Although studies have shown that disrupting dermal cilia causes follicular arrest and attenuated Hh signaling, little is known about the role of epidermal cilia. Here, epidermal cilia function was analyzed using conditional alleles of the ciliogenic genes Ift88 and Kif3a. At birth, epidermal cilia mutants appeared normal, but developed basaloid hyperplasia and ingrowths into the dermis of the ventrum with age. In addition, follicles in the tail were disorganized and had excess sebaceous gland lobules. Epidermal cilia mutants displayed fewer long-term label-retaining cells, suggesting altered stem cell homeostasis. Abnormal proliferation and differentiation were evident from lineage-tracing studies and showed an expansion of follicular cells into the interfollicular epidermis, as is seen during wound repair. These phenotypes were not associated with changes in canonical Wnt activity or oriented cell division. However, nuclear accumulation of the ΔNp63 transcription factor, which is involved in stratification, keratinocyte differentiation and wound repair, was increased, whereas the Hh pathway was repressed. Intriguingly, the phenotypes were not typical of those associated with loss of Hh signaling but exhibited similarities with those of mice in which ΔNp63 is overexpressed in the epidermis. Collectively, these data indicate that epidermal primary cilia may function in stress responses and epidermal homeostasis involving pathways other than those typically associated with primary cilia.


Asunto(s)
Cilios/fisiología , Células Epidérmicas , Folículo Piloso/fisiología , Homeostasis/fisiología , Fenómenos Fisiológicos de la Piel , Animales , Animales Recién Nacidos , Cilios/genética , Cilios/metabolismo , Epidermis/metabolismo , Epidermis/fisiología , Regulación del Desarrollo de la Expresión Génica , Folículo Piloso/citología , Folículo Piloso/metabolismo , Homeostasis/genética , Integrasas/genética , Integrasas/metabolismo , /metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Fenotipo , Fenómenos Fisiológicos de la Piel/genética , Transgenes/genética , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo
13.
Curr Biol ; 17(18): 1586-94, 2007 Sep 18.
Artículo en Inglés | MEDLINE | ID: mdl-17825558

RESUMEN

The assembly of primary cilia is dependent on intraflagellar transport (IFT), which mediates the bidirectional movement of proteins between the base and tip of the cilium. In mice, congenic mutations disrupting genes required for IFT (e.g., Tg737 or the IFT kinesin Kif3a) are embryonic lethal, whereas kidney-specific disruption of IFT results in severe, rapidly progressing cystic pathology. Although the function of primary cilia in most tissues is unknown, in the kidney they are mechanosenstive organelles that detect fluid flow through the tubule lumen. The loss of this flow-induced signaling pathway is thought to be a major contributing factor to cyst formation. Recent data also suggest that there is a connection between ciliary dysfunction and obesity as evidenced by the discovery that proteins associated with human obesity syndromes such as Alström and Bardet-Biedl localize to this organelle. To more directly assess the importance of cilia in postnatal life, we utilized conditional alleles of two ciliogenic genes (Tg737 and Kif3a) to systemically induce cilia loss in adults. Surprisingly, the cystic kidney pathology in these mutants is dependent on the time at which cilia loss was induced, suggesting that cyst formation is not simply caused by impaired mechanosensation. In addition to the cystic pathology, the conditional cilia mutant mice become obese, are hyperphagic, and have elevated levels of serum insulin, glucose, and leptin. We further defined where in the body cilia are required for normal energy homeostasis by disrupting cilia on neurons throughout the central nervous system and on pro-opiomelanocortin-expressing cells in the hypothalamus, both of which resulted in obesity. These data establish that neuronal cilia function in a pathway regulating satiety responses.


Asunto(s)
Cilios/metabolismo , Flagelos/metabolismo , Enfermedades Renales Quísticas/metabolismo , Riñón/metabolismo , Obesidad/metabolismo , Adiposidad , Animales , Cilios/genética , Femenino , Riñón/patología , Enfermedades Renales Quísticas/etiología , Enfermedades Renales Quísticas/patología , Hígado/metabolismo , Hígado/patología , Masculino , Mecanotransducción Celular , Ratones , Mutación , Obesidad/etiología , Obesidad/patología , Páncreas/metabolismo , Páncreas/patología , Transporte de Proteínas/genética , Proteínas Supresoras de Tumor/genética
14.
Am J Physiol Cell Physiol ; 290(4): C952-63, 2006 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-16207792

RESUMEN

The Tg737 degrees (rpk) autosomal recessive polycystic kidney disease (ARPKD) mouse carries a hypomorphic mutation in the Tg737 gene. Because of the absence of its protein product Polaris, the nonmotile primary monocilium central to the luminal membrane of ductal epithelia, such as the cortical collecting duct (CCD) principal cell (PC), is malformed. Although the functions of the renal monocilium remain elusive, primary monocilia or flagella on neurons act as sensory organelles. Thus we hypothesized that the PC monocilium functions as a cellular sensor. To test this hypothesis, we assessed the contribution of Polaris and cilium structure and function to renal epithelial ion transport electrophysiology. Properties of Tg737 degrees (rpk) mutant CCD PC clones were compared with clones genetically rescued with wild-type Tg737 cDNA. All cells were grown as polarized cell monolayers with similarly high transepithelial resistance on permeable filter supports. Three- to fourfold elevated transepithelial voltage (V(te)) and short-circuit current (I(sc)) were measured in mutant orpk monolayers vs. rescued controls. Pharmacological and cell biological examination of this enhanced electrical end point in mutant monolayers revealed that epithelial Na(+) channels (ENaCs) were upregulated. Amiloride, ENaC-selective amiloride analogs (benzamil and phenamil), and protease inhibitors (aprotinin and leupeptin) attenuated heightened V(te) and I(sc). Higher concentrations of additional amiloride analogs (ethylisopropylamiloride and dimethylamiloride) also revealed inhibition of V(te). Cell culture requirements and manipulations were also consistent with heightened ENaC expression and function. Together, these data suggest that ENaC expression and/or function are upregulated in the luminal membrane of mutant, cilium-deficient orpk CCD PC monolayers vs. cilium-competent controls. When the genetic lesion causes loss or malformation of the monocilium, ENaC-driven Na(+) hyperabsorption may explain the rapid emergence of severe hypertension in a majority of patients with ARPKD.


Asunto(s)
Cilios/metabolismo , Células Epiteliales , Riñón , Riñón Poliquístico Autosómico Recesivo/metabolismo , Canales de Sodio/metabolismo , Sodio/metabolismo , Amilorida/análogos & derivados , Amilorida/metabolismo , Animales , Células Cultivadas , Modelos Animales de Enfermedad , Electrofisiología , Células Epiteliales/citología , Células Epiteliales/metabolismo , Canales Epiteliales de Sodio , Riñón/citología , Riñón/metabolismo , Ratones , Péptido Hidrolasas/metabolismo , Riñón Poliquístico Autosómico Recesivo/genética , Bloqueadores de los Canales de Sodio/metabolismo , Canales de Sodio/genética , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo
15.
Lab Invest ; 85(1): 45-64, 2005 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-15580285

RESUMEN

While relatively ignored for years as vestigial, cilia have recently become the focus of intense interest as organelles that result in severe pathologies when disrupted. Here, we further establish a connection between cilia dysfunction and disease by showing that loss of polaris (Tg737), an intraflagellar transport (IFT) protein required for ciliogenesis, causes abnormalities in the exocrine and endocrine pancreas of the Tg737(orpk) mouse. Pathology is evident late in gestation as dilatations of the pancreatic ducts that continue to expand postnatally. Shortly after birth, the acini become disorganized, undergo apoptosis, and are largely ablated in late stage pathology. In addition, serum amylase levels are elevated and carboxypeptidase is abnormally activated within the pancreas. Ultrastructural analysis reveals that the acini undergo extensive vacuolization and have numerous 'halo-granules' similar to that seen in induced models of pancreatitis resulting from duct obstruction. Intriguingly, although the acini are severely affected in Tg737(orpk) mutants, cilia and Tg737 expression are restricted to the ducts and islets and are not detected on acinar cells. Analysis of the endocrine pancreas in Tg737(orpk) mutants revealed normal differentiation and distribution of cell types in the islets. However, after fasting, mutant blood glucose levels are significantly lower than controls and when challenged in glucose tolerance tests, Tg737(orpk) mutants exhibited defects in glucose uptake. These findings are interesting in light of the recently proposed role for polaris, the protein encoded by the Tg737 gene, in the hedgehog pathway and hedgehog signaling in insulin production and glucose homeostasis.


Asunto(s)
Páncreas/anomalías , Páncreas/patología , Proteínas Supresoras de Tumor , Animales , Animales Recién Nacidos , Apoptosis , Proliferación Celular , Cilios/ultraestructura , Modelos Animales de Enfermedad , Femenino , Técnica del Anticuerpo Fluorescente Indirecta , Prueba de Tolerancia a la Glucosa , Islotes Pancreáticos/anomalías , Islotes Pancreáticos/metabolismo , Islotes Pancreáticos/patología , Masculino , Ratones , Ratones Mutantes , Ratones Transgénicos , Páncreas/enzimología , Páncreas Exocrino/anomalías , Páncreas Exocrino/metabolismo , Páncreas Exocrino/patología , Conductos Pancreáticos/anomalías , Conductos Pancreáticos/metabolismo , Conductos Pancreáticos/patología , Proteínas Supresoras de Tumor/deficiencia , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , beta-Galactosidasa/metabolismo
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA
...