RESUMEN
BACKGROUND: The purpose of this study was to determine what was "best achievable practice" with the implementation of a novel enteral feeding protocol (Enhanced Protein-Energy Provision via the Enteral Route Feeding Protocol [PEP uP protocol]). METHODS: This study was a multicenter quality improvement collaborative wherein we describe nutrition practices and outcomes within PEP uP sites. We report the minimum, average, and maximal site-level performance on aspects related to nutrition practices and outcomes. RESULTS: In 2014, 7 intensive care units (ICUs) in the United States implemented the PEP uP protocol. On average, over the first 5 ICU days, patients received 35% (site range, 26%-53%) of their prescribed energy requirements and 42% (site range, 29%-66%) of their prescribed protein requirements from enteral nutrition. In PEP uP sites, 71% (site range, 58%-95%) of patients received a semidigested formula within 72 hours of admission to the ICU, 72% had a volume-based goal as the initial feeding strategy (site range, 47%-100%), 56% had prophylactic protein supplements (site range, 0%-100%), and 19% received prophylactic motility agents (site range, 0%-85%). CONCLUSIONS: There was variable success with the implementation of the different components of the PEP uP protocol. Improving the implementation of the various components may further increase nutrition delivery.
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Cuidados Críticos/métodos , Proteínas en la Dieta/administración & dosificación , Ingestión de Energía , Nutrición Enteral/métodos , Nutrición Enteral/normas , Mejoramiento de la Calidad , Protocolos Clínicos , Conducta Cooperativa , Cuidados Críticos/normas , Femenino , Humanos , Unidades de Cuidados Intensivos , Masculino , Persona de Mediana Edad , Necesidades Nutricionales , Estudios Prospectivos , Estados UnidosRESUMEN
Since 2015, Society of Critical Care Medicine/American Society for Parenteral and Enteral Nutrition and Canadian critical care nutrition support guidelines have both been updated. Despite a similar evidentiary basis, there remain key differences between guideline recommendations. These differences in recommendations may pose confusion for the clinician and may encumber widespread applicability. The aim of this review was to enhance practitioner confidence in applying critical care nutrition support guidelines to patient care in their settings by outlining the similarities and differences between the American and Canadian methods for guideline development and describing the key differences and reasons behind the differences.
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Cuidados Críticos , Apoyo Nutricional , Guías de Práctica Clínica como Asunto , Canadá , Cuidados Críticos/tendencias , Medicina Basada en la Evidencia/tendencias , Humanos , Apoyo Nutricional/efectos adversos , Apoyo Nutricional/tendencias , Medicina de Precisión/tendencias , Sociedades Científicas , Estados UnidosRESUMEN
Nutrition support is a necessary therapy for critically ill cardiac surgery patients. However, conclusive evidence for this population, consisting of well-conducted clinical trials is lacking. To clarify optimal strategies to improve outcomes, an international multidisciplinary group of 25 experts from different clinical specialties from Germany, Canada, Greece, USA and Russia discussed potential approaches to identify patients who may benefit from nutrition support, when best to initiate nutrition support, and the potential use of pharmaco-nutrition to modulate the inflammatory response to cardiopulmonary bypass. Despite conspicuous knowledge and evidence gaps, a rational nutritional support therapy is presented to benefit patients undergoing cardiac surgery.
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Procedimientos Quirúrgicos Cardíacos/métodos , Enfermedades Cardiovasculares/dietoterapia , Consenso , Apoyo Nutricional/tendencias , Adulto , Humanos , Comunicación Interdisciplinaria , Internacionalidad , Metabolismo/fisiología , Estado NutricionalRESUMEN
BACKGROUND: Critically ill patients are at increased risk of developing malnutrition-related complications because of physiological changes, suboptimal delivery, and reduced intake. Strategies to improve nutrition during critical illness recovery are required to prevent iatrogenic underfeeding and risk of malnutrition. The purpose of this study was to assess the feasibility and acceptability of a novel family-centered intervention to improve nutrition in critically ill patients. MATERIALS AND METHODS: A 3-phase, prospective cohort feasibility study was conducted in 4 intensive care units (ICUs) across 2 countries. Intervention feasibility was determined by patient eligibility, recruitment, and retention rates. The acceptability of the intervention was assessed by participant perspectives collected through surveys. Participants included family members of the critically ill patients and ICU and ward healthcare professionals (HCPs). RESULTS: A total of 75 patients and family members, as well as 56 HCPs, were enrolled. The consent rate was 66.4%, and 63 of 75 (84%) of family participants completed the study. Most family members (53/55; 98.1%) would recommend the nutrition education program to others and reported improved ability to ask questions about nutrition (16/20; 80.0%). Family members viewed nutrition care more positively in the ICU. HCPs agreed that families should partner with HCPs to achieve optimal nutrition in the ICU and the wards. Health literacy was identified as a potential barrier to family participation. CONCLUSION: The intervention was feasible and acceptable to families of critically ill patients and HCPs. Further research to evaluate intervention impact on nutrition intake and patient-centered outcomes is required.
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Enfermedad Crítica/terapia , Familia , Desnutrición/prevención & control , Apoyo Nutricional , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Australia , Canadá , Dieta , Estudios de Factibilidad , Femenino , Estudios de Seguimiento , Educación en Salud/economía , Humanos , Unidades de Cuidados Intensivos , Masculino , Persona de Mediana Edad , Estado Nutricional , Nutricionistas , Estudios Prospectivos , Factores de Riesgo , Adulto JovenRESUMEN
BACKGROUND: Selenium (Se) is an essential trace element with antioxidant, anti-inflammatory, and immunomodulatory effects. So far, several randomized clinical trials (RCTs) have demonstrated that parenteral Se may improve clinical outcomes in intensive care unit (ICU) patients. Since publication of our previous systematic review and meta-analysis on antioxidants in the ICU, reports of several trials have been published, including the largest RCT on Se therapy. The purpose of the present systematic review was to update our previous data on intravenous (IV) Se in the critically ill. METHODS: We searched MEDLINE, Embase, and the Cochrane Central Register of Controlled Trials. We included RCTs with parallel groups comparing parenteral Se as single or combined therapy with placebo. Potential trials were evaluated according to specific eligibility criteria, and two reviewers abstracted data from original trials in duplicate independently. Overall mortality was the primary outcome; secondary outcomes were infections, ICU length of stay (LOS), hospital LOS, ventilator days, and new renal dysfunction. RESULTS: A total of 21 RCTs met our inclusion criteria. When the data from these trials were aggregated, IV Se had no effect on mortality (risk ratio [RR] 0.98, 95 % CI 0.90-1.08, P = 0.72, heterogeneity I 2 = 0 %). In addition, when the results of ten trials in which researchers reported on infections were statistically aggregated, there was no significant treatment effect of parenteral Se (RR 0.95, 95 % CI 0.88-1.02, P = 0.15, I 2 = 0 %). There was no positive or negative effect of Se therapy on ICU and hospital LOS, renal function, or ventilator days. CONCLUSIONS: In critically ill patients, IV Se as monotherapy does not improve clinical outcomes.
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Antioxidantes/administración & dosificación , Enfermedad Crítica/mortalidad , Enfermedad Crítica/terapia , Selenio/administración & dosificación , Relación Dosis-Respuesta a Droga , Humanos , Infusiones Intravenosas , Mortalidad/tendencias , Ensayos Clínicos Controlados Aleatorios como Asunto/métodos , Resultado del TratamientoRESUMEN
Critical illness is hypothesized to associate with loss of "health-promoting" commensal microbes and overgrowth of pathogenic bacteria (dysbiosis). This dysbiosis is believed to increase susceptibility to nosocomial infections, sepsis, and organ failure. A trial with prospective monitoring of the intensive care unit (ICU) patient microbiome using culture-independent techniques to confirm and characterize this dysbiosis is thus urgently needed. Characterizing ICU patient microbiome changes may provide first steps toward the development of diagnostic and therapeutic interventions using microbiome signatures. To characterize the ICU patient microbiome, we collected fecal, oral, and skin samples from 115 mixed ICU patients across four centers in the United States and Canada. Samples were collected at two time points: within 48 h of ICU admission, and at ICU discharge or on ICU day 10. Sample collection and processing were performed according to Earth Microbiome Project protocols. We applied SourceTracker to assess the source composition of ICU patient samples by using Qiita, including samples from the American Gut Project (AGP), mammalian corpse decomposition samples, childhood (Global Gut study), and house surfaces. Our results demonstrate that critical illness leads to significant and rapid dysbiosis. Many taxons significantly depleted from ICU patients versus AGP healthy controls are key "health-promoting" organisms, and overgrowth of known pathogens was frequent. Source compositions of ICU patient samples are largely uncharacteristic of the expected community type. Between time points and within a patient, the source composition changed dramatically. Our initial results show great promise for microbiome signatures as diagnostic markers and guides to therapeutic interventions in the ICU to repopulate the normal, "health-promoting" microbiome and thereby improve patient outcomes. IMPORTANCE Critical illness may be associated with the loss of normal, "health promoting" bacteria, allowing overgrowth of disease-promoting pathogenic bacteria (dysbiosis), which, in turn, makes patients susceptible to hospital-acquired infections, sepsis, and organ failure. This has significant world health implications, because sepsis is becoming a leading cause of death worldwide, and hospital-acquired infections contribute to significant illness and increased costs. Thus, a trial that monitors the ICU patient microbiome to confirm and characterize this hypothesis is urgently needed. Our study analyzed the microbiomes of 115 critically ill subjects and demonstrated rapid dysbiosis from unexpected environmental sources after ICU admission. These data may provide the first steps toward defining targeted therapies that correct potentially "illness-promoting" dysbiosis with probiotics or with targeted, multimicrobe synthetic "stool pills" that restore a healthy microbiome in the ICU setting to improve patient outcomes.
RESUMEN
BACKGROUND: Critical illness is characterized by a loss of commensal flora and an overgrowth of potentially pathogenic bacteria, leading to a high susceptibility to nosocomial infections. Probiotics are living non-pathogenic microorganisms, which may protect the gut barrier, attenuate pathogen overgrowth, decrease bacterial translocation and prevent infection. The purpose of this updated systematic review is to evaluate the overall efficacy of probiotics and synbiotic mixtures on clinical outcomes in critical illness. METHODS: Computerized databases from 1980 to 2016 were searched. Randomized controlled trials (RCT) evaluating clinical outcomes associated with probiotic therapy as a single strategy or in combination with prebiotic fiber (synbiotics). Overall number of new infections was the primary outcome; secondary outcomes included mortality, ICU and hospital length of stay (LOS), and diarrhea. Subgroup analyses were performed to elucidate the role of other key factors such as probiotic type and patient mortality risk on the effect of probiotics on outcomes. RESULTS: Thirty trials that enrolled 2972 patients were identified for analysis. Probiotics were associated with a significant reduction in infections (risk ratio 0.80, 95 % confidence interval (CI) 0.68, 0.95, P = 0.009; heterogeneity I (2) = 36 %, P = 0.09). Further, a significant reduction in the incidence of ventilator-associated pneumonia (VAP) was found (risk ratio 0.74, 95 % CI 0.61, 0. 90, P = 0.002; I (2) = 19 %). No effect on mortality, LOS or diarrhea was observed. Subgroup analysis indicated that the greatest improvement in the outcome of infections was in critically ill patients receiving probiotics alone versus synbiotic mixtures, although limited synbiotic trial data currently exists. CONCLUSION: Probiotics show promise in reducing infections, including VAP in critical illness. Currently, clinical heterogeneity and potential publication bias reduce strong clinical recommendations and indicate further high quality clinical trials are needed to conclusively prove these benefits.
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Infecciones Bacterianas/prevención & control , Enfermedad Crítica , Infección Hospitalaria/prevención & control , Probióticos/uso terapéutico , Simbióticos , Humanos , Neumonía Asociada al Ventilador/prevención & control , Ensayos Clínicos Controlados Aleatorios como Asunto , Resultado del TratamientoRESUMEN
BACKGROUND: Enteral nutrition (EN) is recommended as the preferred route for early nutrition therapy in critically ill adults over parenteral nutrition (PN). A recent large randomized controlled trial (RCT) showed no outcome differences between the two routes. The objective of this systematic review was to evaluate the effect of the route of nutrition (EN versus PN) on clinical outcomes of critically ill patients. METHODS: An electronic search from 1980 to 2016 was performed identifying relevant RCTs. Individual trial data were abstracted and methodological quality of included trials scored independently by two reviewers. The primary outcome was overall mortality and secondary outcomes included infectious complications, length of stay (LOS) and mechanical ventilation. Subgroup analyses were performed to examine the treatment effect by dissimilar caloric intakes, year of publication and trial methodology. We performed a test of asymmetry to assess for the presence of publication bias. RESULTS: A total of 18 RCTs studying 3347 patients met inclusion criteria. Median methodological score was 7 (range, 2-12). No effect on overall mortality was found (1.04, 95 % CI 0.82, 1.33, P = 0.75, heterogeneity I(2) = 11 %). EN compared to PN was associated with a significant reduction in infectious complications (RR 0.64, 95 % CI 0.48, 0.87, P = 0.004, I(2) = 47 %). This was more pronounced in the subgroup of RCTs where the PN group received significantly more calories (RR 0.55, 95 % CI 0.37, 0.82, P = 0.003, I(2) = 0 %), while no effect was seen in trials where EN and PN groups had a similar caloric intake (RR 0.94, 95 % CI 0.80, 1.10, P = 0.44, I(2) = 0 %; test for subgroup differences, P = 0.003). Year of publication and methodological quality did not influence these findings; however, a publication bias may be present as the test of asymmetry was significant (P = 0.003). EN was associated with significant reduction in ICU LOS (weighted mean difference [WMD] -0.80, 95 % CI -1.23, -0.37, P = 0.0003, I(2) = 0 %) while no significant differences in hospital LOS and mechanical ventilation were observed. CONCLUSIONS: In critically ill patients, the use of EN as compared to PN has no effect on overall mortality but decreases infectious complications and ICU LOS. This may be explained by the benefit of reduced macronutrient intake rather than the enteral route itself.
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Enfermedad Crítica/enfermería , Nutrición Enteral/enfermería , Estado Nutricional/fisiología , Nutrición Parenteral/enfermería , Ensayos Clínicos Controlados Aleatorios como Asunto , Adulto , Enfermedad Crítica/epidemiología , Nutrición Enteral/métodos , Humanos , Unidades de Cuidados Intensivos , Nutrición Parenteral/métodosAsunto(s)
Enfermedad Crítica/terapia , Emulsiones Grasas Intravenosas/uso terapéutico , Aceites de Pescado/administración & dosificación , Emulsiones Grasas Intravenosas/química , Humanos , Unidades de Cuidados Intensivos , Tiempo de Internación , Metaanálisis como Asunto , Nutrición Parenteral , Ensayos Clínicos Controlados Aleatorios como AsuntoRESUMEN
INTRODUCTION: Intravenous fish oil (FO) lipid emulsions (LEs) are rich in ω-3 polyunsaturated fatty acids, which exhibit anti-inflammatory and immunomodulatory effects. We previously demonstrated that FO-containing LEs may be able to decrease mortality and ventilation days in patients who are critically ill. Since 2014, several additional randomized controlled trials (RCTs) of FO-containing LEs have been published. Therefore, the purpose of this systematic review was to update our previous systematic review with the aim of elucidating the efficacy of FO-containing LEs on clinical outcomes of patients who are critically ill. METHODS: We searched electronic databases from 1980 to 2014. We included four new RCTs conducted in critically ill adult patients in which researchers evaluated FO-containing LEs in parenterally or enterally fed patients. RESULTS: A total of 10 RCTs (n = 733) met inclusion criteria. The mean methodological score was 8 (range, 3 to 12). No effect on overall mortality was found. When we aggregated the results of five RCTs in which infections were reported, we found that FO-containing LEs significantly reduced infections (risk ratio (RR) = 0.64; 95% confidence interval (CI), 0.44 to 0.92; P = 0.02; heterogeneity I (2) = 0%). Subgroup analysis demonstrated that predominantly enteral nutrition-based trials showed a tendency toward a reduction in mortality (RR = 0.69; 95% CI, 0.40 to 1.18; P =0.18; heterogeneity I (2) =35%). High-quality trials showed a significant reduction in hospital length of stay (LOS) (weighted mean difference = -7.42; 95% CI, -11.89 to -2.94; P = 0.001), whereas low-quality trials had no effect (P = 0.45). The results of the test for subgroup differences in hospital LOS was significant (P = 0.001). CONCLUSION: FO-containing LEs may be associated with a reduction in infections and also could be associated with a reduction in duration of ventilation and hospital LOS. Further large-scale RCTs are warranted and should be aimed at consolidating potential positive treatment effects.
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Enfermedad Crítica/terapia , Emulsiones Grasas Intravenosas/uso terapéutico , Aceites de Pescado/uso terapéutico , Enfermedad Crítica/mortalidad , Aceites de Pescado/efectos adversos , Humanos , Unidades de Cuidados Intensivos , Tiempo de Internación , Nutrición Parenteral/métodosRESUMEN
BACKGROUND: Previous studies have documented widespread iatrogenic underfeeding in intensive care unit (ICU) patients. In an experimental setting, we demonstrated the safety and efficacy of a novel enteral feeding protocol designed to overcome the main barriers to adequate delivery of enteral nutrition (EN), the Enhanced Protein-Energy Provision via the Enteral Route Feeding Protocol (PEP uP protocol). The purpose of this article is to describe our experience with implementing this feeding protocol under "real-world" settings in Canada. MATERIALS AND METHODS: This study is a multicenter quality improvement initiative with a concurrent control group. Selected ICUs implemented the PEP uP protocol, and nutrition practices and outcomes were compared with a concurrent control group of ICUs. RESULTS: In 2013, of the 24 ICUs from Canada that participated in the International Nutrition Survey, 8 implemented the PEP uP protocol and the remaining 16 served as concurrent control sites. Patients at PEP uP sites received 60.1% of their prescribed energy requirements from EN compared with 49.9% in patients from control hospitals (P = .02). In addition, patients in PEP uP protocol sites received more protein from EN (61.0% vs 49.7% of prescribed amounts; P = .01), were more likely to receive protein supplements (71.8% vs 47.7%; P = .01), and were more likely to receive >80% of their protein requirements by day 3 (46.1% vs 29.3%; P = .05) compared with patients in control hospitals. CONCLUSIONS: In the real-life setting, the PEP uP protocol can improve the delivery of EN to critically ill patients.
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Protocolos Clínicos , Cuidados Críticos/métodos , Enfermedad Crítica/terapia , Nutrición Enteral/métodos , Unidades de Cuidados Intensivos , Anciano , Anciano de 80 o más Años , Canadá , Proteínas en la Dieta/administración & dosificación , Ingestión de Energía , Femenino , Humanos , Masculino , Persona de Mediana Edad , Necesidades Nutricionales , Mejoramiento de la CalidadRESUMEN
Clinical practice guidelines (CPGs) are systematically developed statements to assist practitioners and patient decisions about appropriate healthcare for specific clinical circumstances, and are designed to minimize practice variation, improve costs, and improve clinical outcomes. The Canadian Critical Care Practice Guidelines (CCPGs) were first published in 2003 and most recently updated in 2013. A total of 68 new randomized controlled trials were identified since the last version in 2009, 50 of them published between 2009 and 2013. The remaining articles were trials published before 2009 but were not identified in previous iterations of the CCPGs. For clinical practice guidelines to be useful to practitioners, they need to be up-to-date and be reflective of the current body of evidence. Herein we describe the process by which the CCPGs were updated. This process resulted in 10 new sections or clinical topics. Of the old clinical topics, 3 recommendations were upgraded, 4 were downgraded, and 27 remained the same. To influence decision making at the bedside, these updated guidelines need to be accompanied by active guideline implementation strategies. Optimal implementation strategies should be guided by local contextual factors including barriers and facilitators to best practice recommendations. Moreover, evaluating and monitoring performance, such as participating in the International Nutrition Survey of practice, should be part of any intensive care unit's performance improvement strategy. The active implementation of the updated CCPGs may lead to better nutrition care and improved patient outcomes in the critical care setting.
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Cuidados Críticos/métodos , Nutrición Enteral/métodos , Política Nutricional , Guías de Práctica Clínica como Asunto , Canadá , Medicina Basada en la Evidencia , Adhesión a Directriz , Humanos , Ensayos Clínicos Controlados Aleatorios como AsuntoRESUMEN
Hox genes encode transcription factors that play essential roles during embryo morphogenesis and organogenesis. Expression of several Hox members persists at the adult age, indicating a wide spectrum of action from embryonic to postnatal life. In the present study, we reported that in adult mice, the Hoxa5 gene shows a dynamic expression profile in the ovary that depends on the estrous cycle, the gestational status, and the age of the female, suggesting that Hoxa5 may have distinct physiological functions in the ovary. Consistent with a role for Hoxa5 in ovarian function, Hoxa5(-/-) nulliparous females exhibit precocious puberty and an early onset of estrous acyclicity. They show a prolonged estrous cycle with increased metestrus-diestrus length, a phenotype that worsens with age. Older mutant females also develop ovarian epithelial inclusion cysts reminiscent of human endosalpingiosis. Immunolabeling studies suggest that these cysts originate from the ovarian surface epithelium, a source of epithelial ovarian carcinomas. Staining of the Hoxa5(-/-) ovarian cysts by the ovarian cancer markers paired box gene 8 (PAX8) and Wilms' tumor 1 (WT1) further strengthens the notion that these cysts may constitute preneoplastic lesions. Moreover, the deregulation of the estrous cycle and the presence of ovarian epithelial cysts in Hoxa5(-/-) older females correlate with a reduced expression of specific epidermal growth factor receptor signaling components, namely Egfr, Areg, and Btc. Altogether, our data unveil that Hoxa5, a stroma-specific gene, plays a significant role in ovarian biology and may be involved in ovarian cancer predisposition.
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Células Epiteliales/citología , Estro/fisiología , Regulación de la Expresión Génica , Proteínas de Homeodominio/metabolismo , Quistes Ováricos/metabolismo , Ovario/metabolismo , Fosfoproteínas/metabolismo , Animales , Femenino , Perfilación de la Expresión Génica , Genes Homeobox , Predisposición Genética a la Enfermedad , Genotipo , Inmunohistoquímica/métodos , Hibridación in Situ , Ratones , Ratones Transgénicos , Modelos Genéticos , Mutación , Neoplasias Ováricas/genética , Factor de Transcripción PAX8 , Factores de Transcripción Paired Box/metabolismo , Fenotipo , Factores de TranscripciónRESUMEN
Cre-expressing mouse lines constitute an important asset to mammalian genetics, allowing the deletion of genes in a spatio-temporal specific manner. Our study on Hox gene function in lung development has led us to use a lung endoderm-specific deletion with the Sftpc-cre mouse line expressing the Cre recombinase gene under the control of human surfactant protein C regulatory sequences. In control experiments, the Cre recombinase faithfully activated the Rosa26-lacZ reporter gene in lung epithelium. However as early as e15.5, lungs from Sftp-Cre(+) embryos showed abnormal dilated cysts. This unexpected phenotype was also observed in mice carrying the conditional lung epithelial Hoxa5 deletion, indicating some bias due to Cre deleterious effects. Excessive apoptosis, likely due to Cre toxicity, could explain the abnormal cysts. Our findings illustrate the need for appropriate control experiments and careful interpretation of data to discriminate between the phenotype due to the targeted mutation and the confounding effects of the Cre recombinase.
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Eliminación de Gen , Proteínas de Homeodominio/genética , Integrasas/genética , Pulmón/citología , Fosfoproteínas/genética , Animales , Línea Celular , Regulación del Desarrollo de la Expresión Génica , Marcación de Gen , Genes Reporteros , Genotipo , Humanos , Ratones , Ratones Transgénicos , Mutagénesis Sitio-Dirigida , Fenotipo , Recombinación Genética , Factores de TranscripciónRESUMEN
BACKGROUND: The genomic organization of Hox clusters is fundamental for the precise spatio-temporal regulation and the function of each Hox gene, and hence for correct embryo patterning. Multiple overlapping transcriptional units exist at the Hoxa5 locus reflecting the complexity of Hox clustering: a major form of 1.8 kb corresponding to the two characterized exons of the gene and polyadenylated RNA species of 5.0, 9.5 and 11.0 kb. This transcriptional intricacy raises the question of the involvement of the larger transcripts in Hox function and regulation. METHODOLOGY/PRINCIPAL FINDINGS: We have undertaken the molecular characterization of the Hoxa5 larger transcripts. They initiate from two highly conserved distal promoters, one corresponding to the putative Hoxa6 promoter, and a second located nearby Hoxa7. Alternative splicing is also involved in the generation of the different transcripts. No functional polyadenylation sequence was found at the Hoxa6 locus and all larger transcripts use the polyadenylation site of the Hoxa5 gene. Some larger transcripts are potential Hoxa6/Hoxa5 bicistronic units. However, even though all transcripts could produce the genuine 270 a.a. HOXA5 protein, only the 1.8 kb form is translated into the protein, indicative of its essential role in Hoxa5 gene function. The Hoxa6 mutation disrupts the larger transcripts without major phenotypic impact on axial specification in their expression domain. However, Hoxa5-like skeletal anomalies are observed in Hoxa6 mutants and these defects can be explained by the loss of expression of the 1.8 kb transcript. Our data raise the possibility that the larger transcripts may be involved in Hoxa5 gene regulation. SIGNIFICANCE: Our observation that the Hoxa5 larger transcripts possess a developmentally-regulated expression combined to the increasing sum of data on the role of long noncoding RNAs in transcriptional regulation suggest that the Hoxa5 larger transcripts may participate in the control of Hox gene expression.
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Empalme Alternativo/genética , Embrión de Mamíferos/metabolismo , Proteínas de Homeodominio/genética , Fosfoproteínas/genética , Regiones Promotoras Genéticas , Transcripción Genética , Animales , Animales Recién Nacidos , Secuencia de Bases , Huesos/anomalías , Huesos/patología , Secuencia Conservada , ADN Intergénico/genética , Embrión de Mamíferos/anomalías , Embrión de Mamíferos/patología , Evolución Molecular , Regulación del Desarrollo de la Expresión Génica , Heterocigoto , Proteínas de Homeodominio/metabolismo , Ratones , Ratones Mutantes , Datos de Secuencia Molecular , Proteínas de Neoplasias/genética , Fosfoproteínas/metabolismo , Biosíntesis de Proteínas , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores de TranscripciónRESUMEN
Hox genes encode transcription factors of crucial importance in the pattern formation of a large spectrum of species. Several studies have now proposed a role for these developmental genes in cancer biology. It has been suggested that HOXA5 possesses growth-suppressive properties through activation of p53 expression in human breast tissue. To assess the genetic cooperation that may exist between Hoxa5 and p53 in tumorigenesis, we generated Hoxa5/p53 compound mutant mice. The presence of Hoxa5 null alleles increased the susceptibility of p53(-/-) mice to develop tumors with a high prevalence for thymic lymphoma, suggesting that the loss of function of the two genes collaborate in tumor formation. To extend our analysis to mammary tumorigenesis, we performed Hoxa5/p53 whole mammary gland transplantations into wild-type hosts. In the p53(-/-) background, the presence of one Hoxa5 mutant allele had no impact on mammary tumor formation. In contrast, the complete loss of Hoxa5 function influenced the tumorigenic outcome of p53(+/-) mammary glands. However, the collaborative nature of this interaction did not depend on the transcriptional regulation of p53 by Hoxa5. Altogether, our data establish that Hoxa5 and p53 cooperate in mammary tumorigenesis in vivo.
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Carcinoma/mortalidad , Genes p53/fisiología , Proteínas de Homeodominio/fisiología , Neoplasias Mamarias Animales/mortalidad , Fosfoproteínas/fisiología , Animales , Carcinoma/genética , Femenino , Predisposición Genética a la Enfermedad , Proteínas de Homeodominio/genética , Linfoma/genética , Linfoma/mortalidad , Linfoma/patología , Neoplasias Mamarias Animales/genética , Ratones , Ratones Noqueados , Trasplante de Neoplasias , Neoplasias Primarias Múltiples/genética , Neoplasias Primarias Múltiples/mortalidad , Evaluación de Resultado en la Atención de Salud , Fosfoproteínas/genética , Análisis de Supervivencia , Neoplasias del Timo/genética , Neoplasias del Timo/mortalidad , Neoplasias del Timo/patología , Factores de TranscripciónRESUMEN
Analysis of the Hoxa5(-/-) mutants has revealed the critical role of Hoxa5 in survival, specification of axial identity, and ontogeny of organs, including the respiratory tract. The presence of the selection cassette in the original Hoxa5(-/-) mutation may interfere with the interpretation of the phenotypes. To circumvent this aspect and to bypass the lethality of the Hoxa5 mutation, we have designed a conditional approach and generated Hoxa5 allelic variants. The conditional allele (Hoxa5(floxed)) behaves as a wild-type allele. In contrast, both the Hoxa5(Delta) and the Hoxa5(floxneo) alleles are characterized by the loss of the functional transcript and protein, the lethality due to lung defects and the skeletal homeotic transformations similar to those of the Hoxa5(-/-) mutants. Analysis of neighboring Hox gene expression patterns in the Hoxa5 mutants produced further confirmed that the Hoxa5 allelic variants are true null alleles.
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Proteínas de Homeodominio/genética , Mutación , Fosfoproteínas/genética , Alelos , Animales , Células Cultivadas , Cruzamientos Genéticos , Células Madre Embrionarias/metabolismo , Regulación del Desarrollo de la Expresión Génica , Genes Homeobox , Genes Letales/fisiología , Ingeniería Genética , Pulmón/citología , Pulmón/embriología , Ratones , Ratones Endogámicos , Ratones Noqueados , Tórax/embriología , Factores de TranscripciónRESUMEN
Chromosomal translocation is a common cause of leukaemia and the most common chromosome translocations found in leukaemia patients involve the mixed lineage leukaemia (MLL) gene. AF10 is one of more than 30 MLL fusion partners in leukaemia. We have recently demonstrated that the H3K79 methyltransferase hDOT1L contributes to MLL-AF10-mediated leukaemogenesis through its interaction with AF10 (ref. 5). In addition to MLL, AF10 has also been reported to fuse to CALM (clathrin-assembly protein-like lymphoid-myeloid) in patients with T-cell acute lymphoblastic leukaemia (T-ALL) and acute myeloid leukaemia (AML). Here, we analysed the molecular mechanism of leukaemogenesis by CALM-AF10. We demonstrate that CALM-AF10 fusion is both necessary and sufficient for leukaemic transformation. Additionally, we provide evidence that hDOT1L has an important role in the transformation process. hDOT1L contributes to CALM-AF10-mediated leukaemic transformation by preventing nuclear export of CALM-AF10 and by upregulating the Hoxa5 gene through H3K79 methylation. Thus, our study establishes CALM-AF10 fusion as a cause of leukaemia and reveals that mistargeting of hDOT1L and upregulation of Hoxa5 through H3K79 methylation is the underlying mechanism behind leukaemia caused by CALM-AF10 fusion.
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Transformación Celular Neoplásica/patología , Proteínas de Homeodominio/biosíntesis , Leucemia Experimental/patología , Metiltransferasas/fisiología , Proteínas de Ensamble de Clatrina Monoméricas/fisiología , Proteínas de Fusión Oncogénica/fisiología , Factores de Transcripción/fisiología , Animales , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/patología , Proliferación Celular , Transformación Celular Neoplásica/metabolismo , Células Cultivadas , N-Metiltransferasa de Histona-Lisina , Humanos , Leucemia Experimental/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones SCID , Fosfoproteínas/biosíntesis , Células U937 , Regulación hacia ArribaRESUMEN
Recent progress has enlightened the involvement of Hox genes in organogenesis. Several Hox genes are expressed in normal and neoplastic mammary glands. Using Hoxa5 mutant mice, we showed that Hoxa5-/- females present nursing defects. Characterization of the Hoxa5-/- mammary gland phenotype reveals changes in proliferation and differentiation of the epithelium of nulliparous and pregnant Hoxa5-/- females that precede the abnormal secretory activity at parturition. These defects likely underlie the incapacity of Hoxa5-/- dams to properly feed their pups. Hoxa5 expression is restricted to the mammary stroma at specific stages of mammary gland development. The loss of Hoxa5 function causes accelerated lobuloalveolar epithelium development, a phenotype that can be rescued upon grafting of mutant mammary epithelium into wild-type fat pads. Conversely, reciprocal grafting experiments demonstrate that Hoxa5-/- stroma cannot support normal proliferation of wild-type mammary epithelium. These data establish the essential contribution of Hoxa5 to mammary epithelium instruction by means of mesenchymal-epithelial crosstalk.