RESUMEN
Progressive remodeling of the heart, resulting in cardiomyocyte (CM) loss and increased inflammation, fibrosis, and a progressive decrease in cardiac function, are hallmarks of myocardial infarction (MI)-induced heart failure. We show that MCB-613, a potent small molecule stimulator of steroid receptor coactivators (SRCs) attenuates pathological remodeling post-MI. MCB-613 decreases infarct size, apoptosis, hypertrophy, and fibrosis while maintaining significant cardiac function. MCB-613, when given within hours post MI, induces lasting protection from adverse remodeling concomitant with: 1) inhibition of macrophage inflammatory signaling and interleukin 1 (IL-1) signaling, which attenuates the acute inflammatory response, 2) attenuation of fibroblast differentiation, and 3) promotion of Tsc22d3-expressing macrophages-all of which may limit inflammatory damage. SRC stimulation with MCB-613 (and derivatives) is a potential therapeutic approach for inhibiting cardiac dysfunction after MI.
Asunto(s)
Ciclohexanonas/farmacología , Infarto del Miocardio/fisiopatología , Piridinas/farmacología , Receptores de Esteroides/metabolismo , Remodelación Ventricular/efectos de los fármacos , Animales , Diferenciación Celular/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Fibroblastos/patología , Fibrosis , Pruebas de Función Cardíaca , Inflamación/patología , Macrófagos/efectos de los fármacos , Macrófagos/patología , Ratones , Infarto del Miocardio/genética , Infarto del Miocardio/patología , Células RAW 264.7 , ARN/genética , ARN/metabolismo , Transcripción Genética/efectos de los fármacosRESUMEN
Ovulation and luteinization are initiated in preovulatory follicles by the luteinizing hormone (LH) surge; however, the signaling events that mediate LH actions in these follicles remain incompletely defined. Two key transcription factors that are targets of LH surge are C/EBPalpha and C/EBPbeta, and their depletion in granulosa cells results in complete infertility. Microarray analyses of these mutant mice revealed altered expression of a number of genes, including growth arrest specific-1 (Gas1). To investigate functions of Gas1 in ovulation- and luteinization-related processes, we crossed Cyp19a1-Cre and Gas1(flox/flox) mice to conditionally delete Gas1 in granulosa and cumulus cells. While expression of Gas1 is dramatically increased in granulosa and cumulus cells around 12-16 h post-human chorionic gonadotropin (hCG) stimulation in wild-type mice, this increase is abolished in Cebpa/b double mutant and in Gas1 mutant mice. GAS1 is also dynamically expressed in stromal cells of the ovary independent of C/EBPalpha/beta. Female Gas1 mutant mice are fertile, exhibit enhanced rates of ovulation, increased fertility, and higher levels of Areg and Lhcgr mRNA in granulosa cells. The morphological appearance and vascularization of corpora lutea appeared normal in these mutant females. Interestingly, levels of mRNA for a number of genes (Cyp11a1, Star, Wnt4, Prlr, Cd52, and Sema3a) associated with luteinization are decreased in corpora lutea of Gas1 mutant mice as compared with controls at 24 h post-hCG; these differences were no longer detectable by 48 h post-hCG. The C/EBP target Gas1 is induced in granulosa cells and is associated with ovulation and luteinization.
Asunto(s)
Proteína beta Potenciadora de Unión a CCAAT/genética , Proteínas Potenciadoras de Unión a CCAAT/genética , Proteínas de Ciclo Celular/genética , Cuerpo Lúteo/metabolismo , Ovulación/genética , Animales , Proteína beta Potenciadora de Unión a CCAAT/metabolismo , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Proteínas de Ciclo Celular/metabolismo , Células del Cúmulo/metabolismo , Femenino , Proteínas Ligadas a GPI/genética , Proteínas Ligadas a GPI/metabolismo , Células de la Granulosa/metabolismo , Luteinización/genética , Luteinización/metabolismo , Ratones , Ratones Noqueados , Ovulación/metabolismoRESUMEN
Background: Recent work has shown that the NLR-family-pyrin-domain-containing 3 (NLRP3) inflammasome is expressed in cardiomyocytes and when specifically activated causes atrial electrical remodeling and arrhythmogenicity. Whether the NLRP3-inflammasome system is functionally important in cardiac fibroblasts (FBs) remains controversial. In this study, we sought to uncover the potential contribution of FB NLRP3-inflammasome signaling to the control of cardiac function and arrhythmogenesis. Methods: Digital-PCR was performed to determine the expression of NLRP3-pathway components in FBs isolated from human biopsy samples of AF and sinus rhythm patients. NLRP3-system protein expression was determined by immunoblotting in atria of canines with electrically maintained AF. Using the inducible, resident fibroblast (FB)-specific Tcf21-promoter-Cre system (Tcf21iCre as control), we established a FB-specific knockin (FB-KI) mouse model with FB-restricted expression of constitutively active NLRP3. Cardiac function and arrhythmia susceptibility in mice were assessed by echocardiography, programmed electrical stimulation, and optical mapping studies. Results: NLRP3 and IL1B were upregulated in atrial FBs of patients with persistent AF. Protein levels of NLRP3, ASC, and pro-Interleukin-1ß were increased in atrial FBs of a canine AF model. Compared with the control mice, FB-KI mice exhibited enlarged left atria (LA) and reduced LA contractility, a common determinant of AF. The FBs from FB-KI mice were more transdifferentiated, migratory, and proliferative compared to the FBs from control mice. FB-KI mice showed increased cardiac fibrosis, atrial gap junction remodeling, and reduced conduction velocity, along with increased AF susceptibility. These phenotypic changes were supported by single nuclei (sn)RNA-seq analysis, which revealed enhanced extracellular matrix remodeling, impaired communication among cardiomyocytes, and altered metabolic pathways across multiple cell types. Conclusions: Our results show that the FB-restricted activation of the NLRP3-inflammasome system leads to fibrosis, atrial cardiomyopathy, and AF. Activation of NLRP3-inflammasome in resident FBs exhibits cell-autonomous function by increasing the activity of cardiac FBs, fibrosis, and connexin remodeling. This study establishes the NLRP3-inflammasome as a novel FB-signaling pathway contributing to AF pathogenesis.
RESUMEN
Multicellular organisms have evolved sophisticated mechanisms to recover and maintain original tissue functions following injury. Injury responses require a robust transcriptomic response associated with cellular reprogramming involving complex gene expression programs critical for effective tissue repair following injury. Steroid receptor coactivators (SRCs) are master transcriptional regulators of cell-cell signaling that is integral for embryogenesis, reproduction, normal physiological function, and tissue repair following injury. Effective therapeutic approaches for facilitating improved tissue regeneration and repair will likely involve temporal and combinatorial manipulation of cell-intrinsic and cell-extrinsic factors. Pleiotropic actions of SRCs that are critical for wound healing range from immune regulation and angiogenesis to maintenance of metabolic regulation in diverse organ systems. Recent evidence derived from studies of model organisms during different developmental stages indicates the importance of the interplay of immune cells and stromal cells to wound healing. With SRCs being the master regulators of cell-cell signaling integral to physiologic changes necessary for wound repair, it is becoming clear that therapeutic targeting of SRCs provides a unique opportunity for drug development in wound healing. This review will provide an overview of wound healing-related functions of SRCs with a special focus on cellular and molecular interactions important for limiting tissue damage after injury. Finally, we review recent findings showing stimulation of SRCs following cardiac injury with the SRC small molecule stimulator MCB-613 can promote cardiac protection and inhibit pathologic remodeling after myocardial infarction.
Asunto(s)
Coactivadores de Receptor Nuclear/fisiología , Cicatrización de Heridas/genética , Animales , Regulación de la Expresión Génica , Humanos , Familia de Multigenes/fisiología , Neovascularización Fisiológica/genética , Coactivador 1 de Receptor Nuclear/fisiología , Coactivador 2 del Receptor Nuclear/fisiología , Coactivador 3 de Receptor Nuclear/fisiología , Transducción de Señal/genéticaRESUMEN
Cyclin D1 is a cell cycle control protein that plays an important role in regenerating liver and many types of cancer. Previous reports have shown that cyclin D1 can directly enhance estrogen receptor activity and inhibit androgen receptor activity in a ligand-independent manner and thus may play an important role in hormone-responsive malignancies. In this study, we examine a distinct mechanism by which cyclin D1 regulates sex steroid signaling, via altered metabolism of these hormones at the tissue and cellular level. In male mouse liver, ectopic expression of cyclin D1 regulated genes involved in the synthesis and degradation of sex steroid hormones in a pattern that would predict increased estrogen and decreased androgen levels. Indeed, hepatic expression of cyclin D1 led to increased serum estradiol levels, increased estrogen-responsive gene expression, and decreased androgen-responsive gene expression. Cyclin D1 also regulated the activity of several key enzymatic reactions in the liver, including increased oxidation of testosterone to androstenedione and decreased conversion of estradiol to estrone. Similar findings were seen in the setting of physiological cyclin D1 expression in regenerating liver. Knockdown of cyclin D1 in HuH7 cells produced reciprocal changes in steroid metabolism genes compared with cyclin D1 overexpression in mouse liver. In conclusion, these studies establish a novel link between the cell cycle machinery and sex steroid metabolism and provide a distinct mechanism by which cyclin D1 may regulate hormone signaling. Furthermore, these results suggest that increased cyclin D1 expression, which occurs in liver regeneration and liver diseases, may contribute to the feminization seen in these settings.
Asunto(s)
Andrógenos/biosíntesis , Ciclina D1/metabolismo , Estrógenos/biosíntesis , Hígado/metabolismo , Animales , Inhibidores de la Aromatasa/farmacología , Línea Celular Tumoral , Ciclina D1/genética , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/fisiología , Humanos , Letrozol , Regeneración Hepática , Masculino , Ratones , Ratones Endogámicos BALB C , Nitrilos/farmacología , Triazoles/farmacologíaRESUMEN
Human progesterone receptors (PR) are phosphorylated by cyclin-dependent protein kinase 2 (CDK2) at multiple sites, including Ser400. Herein, we have addressed the significance of phosphorylation of this residue. PR phospho-Ser400-specific antibodies revealed regulated phosphorylation of Ser400 in response to progestins and mitogens, and this correlated with increased CDK2 levels and activity. Expression of cyclin E elevated CDK2 activity and downregulated PR independently of ligand. Similarly, overexpression of activated mutant CDK2 increased PR transcriptional activity in the absence and presence of progestin. Mutation of PR Ser400 to alanine (S400A) blocked CDK2-induced PR activity in the absence, but not in the presence, of progestin. PR was unresponsive to activated CDK2 in breast cancer cells with elevated p27, and RNA interference knock-down of p27 partially restored CDK2-induced ligand-independent PR activation. Similarly, in p27(-/-) mouse embryonic fibroblasts, elevated CDK2 activity increased wild-type (wt) but not S400A PR transcriptional activity in the absence of progestin. CDK2 induced nuclear localization of unliganded wt but not S400A PR; liganded S400A PR exhibited delayed nuclear accumulation. These studies demonstrate that CDK2 regulates PR in the absence of progestins via phosphorylation of Ser400, thus revealing a novel mechanism for upregulated PR transcriptional activity in human breast cancer cells expressing altered cell cycle regulatory molecules.
Asunto(s)
Quinasas CDC2-CDC28/metabolismo , Receptores de Progesterona/química , Receptores de Progesterona/metabolismo , Serina/metabolismo , Transcripción Genética , Alanina/metabolismo , Sustitución de Aminoácidos , Animales , Western Blotting , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Quinasas CDC2-CDC28/análisis , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Células Cultivadas , Ciclina E/metabolismo , Quinasa 2 Dependiente de la Ciclina , Inhibidor p27 de las Quinasas Dependientes de la Ciclina , Regulación hacia Abajo , Embrión de Mamíferos/citología , Activación Enzimática , Fibroblastos/metabolismo , Eliminación de Gen , Células HeLa , Humanos , Ligandos , Ratones , Mitógenos/metabolismo , Fosforilación , Progestinas/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Interferencia de ARN , Receptores de Progesterona/genética , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismoRESUMEN
Mutations in the tumor protein p53 (TP53) are the most frequently occurring genetic events in high-grade ovarian cancers, especially the prevalence of the Trp53(R172H)-mutant allele. In this study, we investigated the impact of the Trp53(R172H)-mutant allele on epithelial ovarian cancer (EOC) in vivo We used the Pten/Kras(G12D)-mutant mouse strain that develops serous EOC with 100% penetrance to introduce the mutant Trp53(R172H) allele (homolog for human Trp53(R172H)). We demonstrate that the Trp53(R172H) mutation promoted EOC but had differential effects on disease features and progression depending on the presence or absence of the wild-type (WT) TP53 allele. Heterozygous WT/Trp53(R172H) alleles facilitated invasion into the ovarian stroma, accelerated intraperitoneal metastasis, and reduced TP53 transactivation activity but retained responsiveness to nutlin-3a, an activator of WT TP53. Moreover, high levels of estrogen receptor α in these tumors enhanced the growth of both primary and metastatic tumors in response to estradiol. Ovarian tumors homozygous for Trp53(R172H) mutation were undifferentiated and highly metastatic, exhibited minimal TP53 transactivation activity, and expressed genes with potential regulatory functions in EOC development. Notably, heterozygous WT/Trp53(R172H) mice also presented mucinous cystadenocarcinomas at 12 weeks of age, recapitulating human mucinous ovarian tumors, which also exhibit heterozygous TP53 mutations (â¼50%-60%) and KRAS mutations. Therefore, we present the first mouse model of mucinous tumor formation from ovarian cells and supporting evidence that mutant TP53 is a key regulator of EOC progression, differentiation, and responsiveness to steroid hormones. Cancer Res; 76(8); 2206-18. ©2016 AACR.
Asunto(s)
Diferenciación Celular/genética , Estradiol/fisiología , Genes p53 , Metástasis de la Neoplasia/genética , Neoplasias Glandulares y Epiteliales/patología , Neoplasias Ováricas/patología , Animales , Carcinoma Epitelial de Ovario , Progresión de la Enfermedad , Heterocigoto , Ratones , Neoplasias Glandulares y Epiteliales/genética , Neoplasias Ováricas/genéticaRESUMEN
Cyclin D1 is a cell cycle protein that promotes proliferation by mediating progression through key checkpoints in G1 phase. It is also a proto-oncogene that is commonly overexpressed in human cancers. In addition to its canonical role in controlling cell cycle progression, cyclin D1 affects other aspects of cell physiology, in part through transcriptional regulation. In this study, we find that cyclin D1 inhibits the activity of a key metabolic transcription factor, peroxisome proliferator-activated receptor α (PPARα), a member of nuclear receptor family that induces fatty acid oxidation and may play an anti-neoplastic role. In primary hepatocytes, cyclin D1 inhibits PPARα transcriptional activity and target gene expression in a cdk4-independent manner. In liver and breast cancer cells, knockdown of cyclin D1 leads to increased PPARα transcriptional activity, expression of PPARα target genes, and fatty acid oxidation. Similarly, cyclin D1 depletion enhances binding of PPARα to target sequences by chromatin immunoprecipitation. In proliferating hepatocytes and regenerating liver in vivo, induction of endogenous cyclin D1 is associated with diminished PPARα activity. Cyclin D1 expression is both necessary and sufficient for growth factor-mediated repression of fatty acid oxidation in proliferating hepatocytes. These studies indicate that in addition to playing a pivotal role in cell cycle progression, cyclin D1 represses PPARα activity and inhibits fatty acid oxidation. Our findings establish a new link between cyclin D1 and metabolism in both tumor cells and physiologic hepatocyte proliferation.
Asunto(s)
Ciclina D1/metabolismo , Ácidos Grasos/metabolismo , Neoplasias Hepáticas Experimentales/metabolismo , PPAR alfa/metabolismo , Animales , Línea Celular Tumoral , Células Hep G2 , Humanos , Metabolismo de los Lípidos , Hígado/metabolismo , Hígado/patología , Neoplasias Hepáticas Experimentales/patología , Masculino , Ratones , Ratones Endogámicos BALB C , Oxidación-Reducción , Proto-Oncogenes Mas , Ratas , Ratas Sprague-Dawley , TransfecciónRESUMEN
Atopy is a persistent, aberrant humoral response to certain classes of proteins (allergens) characterized by the presence of allergen-specific IgE. Yet, in both atopic and non-atopic individuals, allergen-specific responses involving the IgA and IgG subclasses have been observed, which evidence does not support models suggesting inherited differences in sensitivity to certain protein classes. Using the major ragweed component Amb a 1 as a model allergen, we assessed the humoral responses in three groups of unrelated donors: (A) atopic, ragweed sensitive; (B) atopic, but not ragweed sensitive; (C) non-atopic. As expected, Amb a 1-specific IgE was present in group A only. However, there were essentially no differences in the relative proportions of Amb a 1-specific IgA(1,2) and IgG(1-4) among the groups. We also determined the Amb a 1 binding affinities for IgG(1) and IgG(4) in the three groups, and compared these to Amb a 1-specific IgE binding affinities in group A. Group A donors' Amb a 1-IgE had extremely high affinities (10(8) to 10(11)M(-1)), but their Amb a 1-IgG(1) and Amb a 1-IgG(4) affinities were significantly lower (10(7) to 10(10)M(-1)). The average IgG(4) binding affinities in groups B and C were slightly higher than that of IgG(4) in group A, although not statistically significant. However, the IgG(1) affinity for Amb a 1 among group C, non-atopic donors was significantly elevated and comparable to the IgE affinity observed in group A, ragweed atopics. Inhibition studies with allergen-specific IgE-free serum showed that all isotypes recognized the major epitopes seen by IgE. These results suggest that there may be a "selective competition" among isotypes for allergens that is driven by the ability to produce high affinity, allergen-specific immunoglobulins.
Asunto(s)
Alérgenos/inmunología , Hipersensibilidad/inmunología , Adolescente , Adulto , Afinidad de Anticuerpos , Reacciones Antígeno-Anticuerpo , Antígenos de Plantas , Unión Competitiva , Niño , Femenino , Humanos , Inmunoglobulina E/inmunología , Inmunoglobulina G/análisis , Inmunoglobulina G/clasificación , Isotipos de Inmunoglobulinas/análisis , Masculino , Persona de Mediana Edad , Proteínas de Plantas/inmunología , Receptores de Antígenos de Linfocitos B/fisiologíaRESUMEN
Evolutionary Action analyses of The Cancer Gene Atlas data sets show that many specific p53 missense and gain-of-function mutations are selectively overrepresented and functional in high-grade serous ovarian cancer (HGSC). As homozygous alleles, p53 mutants are differentially associated with specific loss of heterozygosity (R273; chromosome 17); copy number variation (R175H; chromosome 9); and up-stream, cancer-related regulatory pathways. The expression of immune-related cytokines was selectively related to p53 status, showing for the first time that specific p53 mutants impact, and are related to, the immune subtype of ovarian cancer. Although the majority (31%) of HGSCs exhibit loss of heterozygosity, a significant number (24%) maintain a wild-type (WT) allele and represent another HGSC subtype that is not well defined. Using human and mouse cell lines, we show that specific p53 mutants differentially alter endogenous WT p53 activity; target gene expression; and responses to nutlin-3a, a small molecular that activates WT p53 leading to apoptosis, providing "proof of principle" that ovarian cancer cells expressing WT and mutant alleles represent a distinct ovarian cancer subtype. We also show that siRNA knock down of endogenous p53 in cells expressing homozygous mutant alleles causes apoptosis, whereas cells expressing WT p53 (or are heterozygous for WT and mutant p53 alleles) are highly resistant. Therefore, despite different gene regulatory pathways associated with specific p53 mutants, silencing mutant p53 might be a suitable, powerful, global strategy for blocking ovarian cancer growth in those tumors that rely on mutant p53 functions for survival. Knowing p53 mutational status in HGSC should permit new strategies tailored to control this disease.
Asunto(s)
Apoptosis , Variaciones en el Número de Copia de ADN , Imidazoles/metabolismo , Mutación/genética , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Piperazinas/metabolismo , Proteína p53 Supresora de Tumor/genética , Animales , Supervivencia Celular , Femenino , Humanos , Ratones , Neoplasias Ováricas/metabolismo , ARN Interferente Pequeño/genética , Células Tumorales Cultivadas , Proteína p53 Supresora de Tumor/antagonistas & inhibidoresRESUMEN
Epithelial ovarian cancer is a diverse molecular and clinical disease, yet standard treatment is the same for all subtypes. TP53 mutations represent a node of divergence in epithelial ovarian cancer histologic subtypes and may represent a therapeutic opportunity in subtypes expressing wild type, including most low-grade ovarian serous carcinomas, ovarian clear cell carcinomas and ovarian endometrioid carcinomas, which represent approximately 25% of all epithelial ovarian cancer. We therefore sought to investigate Nutlin-3a--a therapeutic which inhibits MDM2, activates wild-type p53, and induces apoptosis--as a therapeutic compound for TP53 wild-type ovarian carcinomas. Fifteen epithelial ovarian cancer cell lines of varying histologic subtypes were treated with Nutlin-3a with determination of IC50 values. Western Blot (WB) and quantitative real-time polymerase chain reaction (qRT-PCR) analyses quantified MDM2, p53, and p21 expression after Nutlin-3a treatment. DNA from 15 cell lines was then sequenced for TP53 mutations in exons 2-11 including intron-exon boundaries. Responses to Nutlin-3a were dependent upon TP53 mutation status. By qRT-PCR and WB, levels of MDM2 and p21 were upregulated in wild-type TP53 sensitive cell lines, and p21 induction was reduced or absent in mutant cell lines. Annexin V assays demonstrated apoptosis in sensitive cell lines treated with Nutlin-3a. Thus, Nutlin-3a could be a potential therapeutic agent for ovarian carcinomas expressing wild-type TP53 and warrants further investigation.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica , Imidazoles/farmacología , Ovario/efectos de los fármacos , Piperazinas/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/agonistas , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Exones , Femenino , Humanos , Concentración 50 Inhibidora , Intrones , Mutación , Ovario/metabolismo , Ovario/patología , Proteínas Proto-Oncogénicas c-mdm2/genética , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Análisis de Secuencia de ADN , Transducción de Señal , Proteína p53 Supresora de Tumor/agonistas , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
We evaluated atopy-associated parameters in 1,099 people (aged 6-84 years) from families with history for atopy. All were tested for serum total immunoglobulin E (IgE) and allergen sensitivity by skin prick test. Specific IgE tests were done in randomly selected families. There was a decline with age in serum total IgE values, and relative atopy "incidence rates" were slightly lower among those older than 60 years. However, there was no change with age in sensitivity or severity of atopy. Among those sensitized to ragweed (Ambrosia artemisilfolia), there was no age-associated change in IgE levels specific to Amb a 1, a major allergen extracted from ragweed, and no change in the binding affinity of IgE for the Amb a 1 allergen. Among families with atopic histories, the underlying atopic mechanisms are particularly robust, and the atopic propensity remains into advanced age. In addition, established atopic responses may be focused in an immune system compartment either independent of or minimally influenced by T-cell activity.
Asunto(s)
Susceptibilidad a Enfermedades/epidemiología , Hipersensibilidad/epidemiología , Hipersensibilidad/inmunología , Inmunoglobulina E/inmunología , Linfocitos T/inmunología , Adolescente , Adulto , Distribución por Edad , Anciano , Anciano de 80 o más Años , Alérgenos/farmacología , Niño , Estudios de Cohortes , Dermatitis Atópica/diagnóstico , Dermatitis Atópica/epidemiología , Dermatitis Atópica/inmunología , Familia , Femenino , Humanos , Hipersensibilidad/diagnóstico , Inmunoglobulina E/análisis , Incidencia , Masculino , Persona de Mediana Edad , Pruebas del Parche , Medición de Riesgo , Muestreo , Índice de Severidad de la Enfermedad , Distribución por Sexo , Linfocitos T/fisiologíaRESUMEN
Breast cancers often have increased mitogen-activated protein kinase (MAPK) activity; this pathway influences breast cancer cell growth in part by targeting steroid hormone receptors. Activation of p42 and p44 MAPKs increases progesterone receptor (PR) transcriptional activity in the presence of progestins, and triggers their rapid down-regulation by the ubiquitin-proteasome pathway. In turn, progestins increase the expression of type I growth factor receptor tyrosine kinases that feed into MAPK activation. Most recently, progestins have been shown to activate the p42/p44 MAPK module in a progesterone receptor (PR) dependent manner, but independently of their function as transcription factors. Indeed, mechanisms of bi-directional cross-talk between these two pathways are becoming well-documented. In this reveiw we provide an overview of the primary ways in which steroid hormone receptor and growth factor cross-talk occurs, using examples from our work and others with human PR as a model receptor. We highlight the regulation of PR by phosphorylation and the role of intracellular protein kinases as key mediators of PR action. Cross-talk between growth factor and PR-mediated signaling events is an important means by which growth regulatory genes may be coordinately regulated, and may contribute to the growth and development of hormonally responsive normal breast tissue and to breast cancer progression.
RESUMEN
The functional status of the tumor repressor protein (TP53 or TRP53) is a defining feature of ovarian cancer. Mutant or null alleles of TP53 are expressed in greater than 90% of all high-grade serous adenocarcinomas. Wild-type TP53 is elevated in low-grade serous adenocarcinomas in women and in our Pten;Kras;Amhr2-Cre mutant mouse model. Disruption of the Trp53 gene in this mouse model did not lead to high-grade ovarian cancer but did increase expression of estrogen receptor α (ESR1) and markedly enhanced the responsiveness of these cells to estrogen. Specifically, when Trp53-positive and Trp53 null mutant mice were treated with estradiol or vehicle, only the Trp53 null and Esr1-positive tumors respond vigorously to estradiol in vivo and exhibit features characteristic of high-grade type ovarian cancer: invasive growth into the ovarian stroma, rampant metastases to the peritoneal cavity, and nuclear atypia. Estrogen promoted and progesterone suppressed the growth of Trp53 null ovarian tumors and tumor cells injected ip, sc, or when grown in matrigel. Exposure of the Trp53 depleted cells to estrogen also has a profound impact on the tumor microenvironment and immune-related events. These results led to the new paradigm that TRP53 status is related to the susceptibility of transformed ovarian surface epithelial cells to estradiol-induced metastases and nuclear atypia via increased levels of estradiol receptor α.
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Adenocarcinoma/metabolismo , Estradiol/fisiología , Neoplasias Ováricas/metabolismo , Proteína p53 Supresora de Tumor/fisiología , Adenocarcinoma/inmunología , Adenocarcinoma/secundario , Animales , Línea Celular Tumoral , Proliferación Celular , Receptor alfa de Estrógeno/metabolismo , Femenino , Humanos , Ratones , Ratones Endogámicos NOD , Ratones Transgénicos , Trasplante de Neoplasias , Neoplasias Ováricas/inmunología , Neoplasias Ováricas/patología , Ovario/patología , Progesterona/fisiologíaRESUMEN
Ovarian cancer in women is a complex and deadly disease, where the molecular events that initiate and control tumor formation remain poorly defined. Therefore, mouse models provide one approach for determining the mechanisms by which specific oncogenic factors cause ovarian surface epithelial cell and granulosa cell transformation. This minireview summarizes the phenotypes of current mouse models that have been generated and some of the underlying mechanisms they have provided.
Asunto(s)
Modelos Animales de Enfermedad , Neoplasias Ováricas/patología , Neoplasias Ováricas/fisiopatología , Animales , Carcinoma Epitelial de Ovario , Transformación Celular Neoplásica/patología , Femenino , Tumor de Células de la Granulosa/patología , Tumor de Células de la Granulosa/fisiopatología , Ratones , Neoplasias Glandulares y Epiteliales/patología , Neoplasias Glandulares y Epiteliales/fisiopatología , Fenotipo , Proteína p53 Supresora de Tumor/fisiologíaRESUMEN
Cumulus cell-oocyte complex (COC) expansion is obligatory for LH-induced ovulation and is initiated by LH induction of the epidermal growth factor (EGF)-like factors that mediate the synthesis of the hyaluronan-rich matrix and hyaluronan-stabilizing factors. COC expansion also involves the movement of cumulus cells within the matrix by mechanisms that have not been characterized. We document herein that two proteases, calpain 2 and to a lesser extent calpain 1, are expressed in cumulus cells and that the proteolytic activity of these enzymes is rapidly and significantly increased in COC isolated from human chorionic gonadotropin-induced ovulatory follicles in vivo. Stimulation of calpain activity was associated with proteolytic degradation of paxillin and talin (two components of focal adhesion complexes), cell detachment, and the formation of cell surface bleb-like protrusions. Injection of a calpain inhibitor in vivo reduced 1) human chorionic gonadotropin-stimulated calpain enzyme activity, 2) cell detachment, 3) membrane protrusion formation, and 4) COC expansion by mechanisms that did not alter Has2 expression. During EGF-like factor induction of COC expansion in culture, calpain activity was increased by ERK1/2 and intracellular Ca(2+) signaling pathways. Inhibition of calpain activity in cultured COC blocked cumulus cell detachment, protrusion formation, and the vigorous movement of cumulus cells. As a consequence, COC expansion was impaired. Collectively, these results show that two highly coordinated processes control COC expansion. One process involves the synthesis of the hyaluronan matrix, and the other mediates cumulus cell detachment and movement. The latter are controlled by calpain activation downstream of the EGF receptor activation of the Ca(2+) pathway and ERK1/2 pathways.
Asunto(s)
Calpaína/metabolismo , Células del Cúmulo/metabolismo , Factor de Crecimiento Epidérmico/farmacología , Oocitos/metabolismo , Animales , Western Blotting , Calpaína/antagonistas & inhibidores , Movimiento Celular/efectos de los fármacos , Movimiento Celular/fisiología , Células Cultivadas , Gonadotropina Coriónica/farmacología , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Ratones , Ratones Endogámicos C57BL , Paxillin/metabolismo , Reacción en Cadena de la PolimerasaRESUMEN
This review summarizes studies providing evidence (1) that endogenous RAS activation regulates important physiological events during ovulation and luteinization (2) that expression of the mutant, active KRAS(G12D) in granulosa cells in vivo causes abnormal follicle growth arrest leading to premature ovarian failure and (3) that KRAS(G12D) expression in ovarian surface epithelial (OSE) cells renders them susceptible to the pathological outcome of transformation and tumor formation. These diverse effects of RAS highlight how critical its activation is linked to cell- and stage-specific events in the ovary that control normal processes and that can also lead to altered granulosa cell and OSE cell fates.
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Activación Enzimática , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Ovario/enzimología , Proteínas ras/metabolismo , Animales , Carcinoma Epitelial de Ovario , Femenino , Células de la Granulosa/enzimología , Humanos , Mutación Missense , Neoplasias Glandulares y Epiteliales/enzimología , Neoplasias Glandulares y Epiteliales/genética , Neoplasias Ováricas/enzimología , Neoplasias Ováricas/genética , Ovulación , Proteínas ras/genética , Proteínas ras/fisiologíaRESUMEN
Loss of Pten in the Kras(G12D);Amhr2-Cre mutant mice leads to the transformation of ovarian surface epithelial (OSE) cells and rapid development of low-grade, invasive serous adenocarcinomas. Tumors occur with 100% penetrance and express elevated levels of wild-type tumor repressor protein 53 (TRP53). To test the functions of TRP53 in the Pten;Kras (Trp53+) mice, we disrupted the Trp53 gene yielding Pten;Kras(Trp53-) mice. By comparing morphology and gene expression profiles in the Trp53+ and Trp53- OSE cells from these mice, we document that wild-type TRP53 acts as a major promoter of OSE cell survival and differentiation: cells lacking Trp53 are transformed yet are less adherent, migratory, and invasive and exhibit a gene expression profile more like normal OSE cells. These results provide a new paradigm: wild-type TRP53 does not preferentially induce apoptotic or senescent related genes in the Pten;Kras(Trp53+) cancer cells but rather increases genes regulating DNA repair, cell cycle progression, and proliferation and decreases putative tumor suppressor genes. However, if TRP53 activity is forced higher by exposure to nutlin-3a (a mouse double minute-2 antagonist), TRP53 suppresses DNA repair genes and induces the expression of genes that control cell cycle arrest and apoptosis. Thus, in the Pten;Kras(Trp53+) mutant mouse OSE cells and likely in human TP53+ low-grade ovarian cancer cells, wild-type TRP53 controls global molecular changes that are dependent on its activation status. These results suggest that activation of TP53 may provide a promising new therapy for managing low-grade ovarian cancer and other cancers in humans in which wild-type TP53 is expressed.
Asunto(s)
Adenocarcinoma/patología , Adenocarcinoma/fisiopatología , Neoplasias Ováricas/patología , Neoplasias Ováricas/fisiopatología , Proteína p53 Supresora de Tumor/fisiología , Animales , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Ciclo Celular/fisiología , Diferenciación Celular/fisiología , Línea Celular Tumoral , Proliferación Celular , Supervivencia Celular/fisiología , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica/fisiología , Humanos , Imidazoles/farmacología , Ratones , Ratones Noqueados , Piperazinas/farmacología , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Following acute hepatic injury, the metabolic capacity of the liver is altered during the process of compensatory hepatocyte proliferation by undefined mechanisms. In this study, we examined the regulation of de novo lipogenesis by cyclin D1, a key mediator of hepatocyte cell cycle progression. In primary hepatocytes, cyclin D1 significantly impaired lipogenesis in response to glucose stimulation. Cyclin D1 inhibited the glucose-mediated induction of key lipogenic genes, and similar effects were seen using a mutant (D1-KE) that does not activate cdk4 or induce cell cycle progression. Cyclin D1 (but not D1-KE) inhibited the activity of the carbohydrate response element-binding protein (ChREBP) by regulating the glucose-sensing motif of this transcription factor. Because changes in ChREBP activity could not fully explain the effect of cyclin D1, we examined hepatocyte nuclear factor 4α (HNF4α), which regulates numerous differentiated functions in the liver including lipid metabolism. We found that both cyclins D1 and D1-KE bound to HNF4α and significantly inhibited its recruitment to the promoter region of lipogenic genes in hepatocytes. Conversely, knockdown of cyclin D1 in the AML12 hepatocyte cell line promoted HNF4α activity and lipogenesis. In mouse liver, HNF4α bound to a central domain of cyclin D1 involved in transcriptional repression. Cyclin D1 inhibited lipogenic gene expression in the liver following carbohydrate feeding. Similar findings were observed in the setting of physiologic cyclin D1 expression in the regenerating liver. In conclusion, these studies demonstrate that cyclin D1 represses ChREBP and HNF4α function in hepatocytes via Cdk4-dependent and -independent mechanisms. These findings provide a direct link between the cell cycle machinery and the transcriptional control of metabolic function of the liver.
Asunto(s)
Ciclina D1/metabolismo , Glucosa/farmacología , Factor Nuclear 4 del Hepatocito/metabolismo , Lipogénesis/efectos de los fármacos , Proteínas Nucleares/metabolismo , Factores de Transcripción/metabolismo , Animales , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice , Células Cultivadas , Ciclina D1/antagonistas & inhibidores , Ciclina D1/genética , Quinasa 4 Dependiente de la Ciclina/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Hepatocitos/citología , Hepatocitos/metabolismo , Metabolismo de los Lípidos/efectos de los fármacos , Ratones , Proteínas Nucleares/genética , Unión Proteica , Estructura Terciaria de Proteína , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Factores de Transcripción/genéticaRESUMEN
Cdk2 was once believed to play an essential role in cell cycle progression, but cdk2(-/-) mice have minimal phenotypic abnormalities. In this study, we examined the role of cdk2 in hepatocyte proliferation, centrosome duplication and survival. Cdk2(-/-) hepatocytes underwent mitosis and had normal centrosome content after mitogen stimulation. Unlike wild-type cells, cdk2(-/-) liver cells failed to undergo centrosome overduplication in response to ectopic cyclin D1 expression. After mitogen stimulation in culture or partial hepatectomy in vivo, cdk2(-/-) hepatocytes demonstrated diminished proliferation. Cyclin D1 is a key mediator of cell cycle progression in hepatocytes, and transient expression of this protein is sufficient to promote robust proliferation of these cells in vivo. In cdk2(-/-) mice and animals treated with the cdk2 inhibitor seliciclib, cyclin D1 failed to induce hepatocyte cell cycle progression. Surprisingly, cdk2 ablation or inhibition led to massive hepatocyte and animal death following cyclin D1 transfection. In a transgenic model of chronic hepatic cyclin D1 expression, seliciclib induced hepatocyte injury and animal death, suggesting that cdk2 is required for survival of cyclin D1-expressing cells even in the absence of substantial proliferation. In conclusion, our studies demonstrate that cdk2 plays a role in liver regeneration. Furthermore, it is essential for centrosome overduplication, proliferation and survival of hepatocytes that aberrantly express cyclin D1 in vivo. These studies suggest that cdk2 may warrant further investigation as a target for therapy of liver tumors with constitutive cyclin D1 expression.