RESUMEN
Gastroenteropancreatic (GEP) neuroendocrine neoplasm (NEN) that consists of neuroendocrine tumor and neuroendocrine carcinoma (NEC) is a lethal but under-investigated disease owing to its rarity. To fill the scarcity of clinically relevant models of GEP-NEN, we here established 25 lines of NEN organoids and performed their comprehensive molecular characterization. GEP-NEN organoids recapitulated pathohistological and functional phenotypes of the original tumors. Whole-genome sequencing revealed frequent genetic alterations in TP53 and RB1 in GEP-NECs, and characteristic chromosome-wide loss of heterozygosity in GEP-NENs. Transcriptome analysis identified molecular subtypes that are distinguished by the expression of distinct transcription factors. GEP-NEN organoids gained independence from the stem cell niche irrespective of genetic mutations. Compound knockout of TP53 and RB1, together with overexpression of key transcription factors, conferred on the normal colonic epithelium phenotypes that are compatible with GEP-NEN biology. Altogether, our study not only provides genetic understanding of GEP-NEN, but also connects its genetics and biological phenotypes.
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Bancos de Muestras Biológicas , Tumores Neuroendocrinos/patología , Organoides/patología , Animales , Cromosomas Humanos/genética , Genotipo , Humanos , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Neoplasias Intestinales/genética , Neoplasias Intestinales/patología , Masculino , Ratones , Modelos Genéticos , Mutación/genética , Tumores Neuroendocrinos/genética , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patología , Fenotipo , Neoplasias Gástricas/genética , Neoplasias Gástricas/patología , Transcriptoma/genética , Secuenciación Completa del GenomaRESUMEN
With ageing, normal human tissues experience an expansion of somatic clones that carry cancer mutations1-7. However, whether such clonal expansion exists in the non-neoplastic intestine remains unknown. Here, using whole-exome sequencing data from 76 clonal human colon organoids, we identify a unique pattern of somatic mutagenesis in the inflamed epithelium of patients with ulcerative colitis. The affected epithelium accumulates somatic mutations in multiple genes that are related to IL-17 signalling-including NFKBIZ, ZC3H12A and PIGR, which are genes that are rarely affected in colon cancer. Targeted sequencing validates the pervasive spread of mutations that are related to IL-17 signalling. Unbiased CRISPR-based knockout screening in colon organoids reveals that the mutations confer resistance to the pro-apoptotic response that is induced by IL-17A. Some of these genetic mutations are known to exacerbate experimental colitis in mice8-11, and somatic mutagenesis in human colon epithelium may be causally linked to the inflammatory process. Our findings highlight a genetic landscape that adapts to a hostile microenvironment, and demonstrate its potential contribution to the pathogenesis of ulcerative colitis.
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Colitis Ulcerosa/genética , Epitelio/metabolismo , Interleucina-17/genética , Mutación , Colitis Ulcerosa/metabolismo , Humanos , Interleucina-17/metabolismo , Fenotipo , Transducción de SeñalRESUMEN
Sphingosine 1-phosphate (S1P) has been implicated in brown adipose tissue (BAT) formation and energy consumption; however, the mechanistic role of sphingolipids, including S1P, in BAT remains unclear. Here, we showed that, in mice, BAT activation by cold exposure upregulated mRNA and protein expression of the S1P-synthesizing enzyme sphingosine kinase 1 (SphK1) and S1P production in BAT. Treatment of wild-type brown adipocytes with exogenous S1P or S1P receptor subtype-selective agonists stimulated triglyceride (TG) breakdown only marginally, compared with noradrenaline. However, genetic deletion of Sphk1 resulted in hypothermia and diminished body weight loss upon cold exposure, suggesting that SphK1 is involved in thermogenesis through mechanisms different from receptor-mediated, extracellular action of S1P. In BAT of wild-type mice, SphK1 was localized largely in the lysosomes of brown adipocytes. In the brown adipocytes of Sphk1-/- mice, the number of lysosomes was reduced and lysosomal function, including proteolytic activity, acid esterase activity, and motility, was impaired. Concordantly, nuclear translocation of transcription factor EB, a master transcriptional regulator of lysosome biogenesis, was reduced, leading to decreased mRNA expression of the lysosome-related genes in Sphk1-/- BAT. Moreover, BAT of Sphk1-/- mice showed greater TG accumulation with dominant larger lipid droplets in brown adipocytes. Inhibition of lysosomes with chloroquine resulted in a less extent of triglyceride accumulation in Sphk1-/- brown adipocytes compared with wild-type brown adipocytes, suggesting a reduced lysosome-mediated TG breakdown in Sphk1-/- mice. Our results indicate a novel role of SphK1 in lysosomal integrity, which is required for TG breakdown and thermogenesis in BAT.
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Adipocitos Marrones , Transducción de Señal , Ratones , Animales , Adipocitos Marrones/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Esfingosina/metabolismo , Tejido Adiposo Pardo/metabolismo , ARN Mensajero/metabolismo , Lisofosfolípidos/metabolismo , Triglicéridos/metabolismoRESUMEN
BACKGROUND & AIMS: In the mouse intestinal epithelium, Lgr5+ stem cells are vulnerable to injury, owing to their predominantly cycling nature, and their progenies de-differentiate to replenish the stem cell pool. However, how human colonic stem cells behave in homeostasis and during regeneration remains unknown. METHODS: Transcriptional heterogeneity among colonic epithelial cells was analyzed by means of single-cell RNA sequencing analysis of human and mouse colonic epithelial cells. To trace the fate of human colonic stem or differentiated cells, we generated LGR5-tdTomato, LGR5-iCasase9-tdTomato, LGR5-split-Cre, and KRT20-ERCreER knock-in human colon organoids via genome engineering. p27+ dormant cells were further visualized with the p27-mVenus reporter. To analyze the dynamics of human colonic stem cells in vivo, we orthotopically xenotransplanted fluorescence-labeled human colon organoids into immune-deficient mice. The cell cycle dynamics in xenograft cells were evaluated using 5-ethynyl-2'-deoxyuridine pulse-chase analysis. The clonogenic capacity of slow-cycling human stem cells or differentiated cells was analyzed in the context of homeostasis, LGR5 ablation, and 5-fluorouracil-induced mucosal injury. RESULTS: Single-cell RNA sequencing analysis illuminated the presence of nondividing LGR5+ stem cells in the human colon. Visualization and lineage tracing of slow-cycling LGR5+p27+ cells and orthotopic xenotransplantation validated their homeostatic lineage-forming capability in vivo, which was augmented by 5-FU-induced mucosal damage. Transforming growth factor-ß signaling regulated the quiescent state of LGR5+ cells. Despite the plasticity of differentiated KRT20+ cells, they did not display clonal growth after 5-FU-induced injury, suggesting that occupation of the niche environment by LGR5+p27+ cells prevented neighboring differentiated cells from de-differentiating. CONCLUSIONS: Our results highlight the quiescent nature of human LGR5+ colonic stem cells and their contribution to post-injury regeneration.
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Receptores Acoplados a Proteínas G , Células Madre , Humanos , Ratones , Animales , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Células Madre/metabolismo , Colon/metabolismo , Mucosa Intestinal/metabolismo , Fluorouracilo , Factores de Crecimiento Transformadores/metabolismoRESUMEN
BACKGROUND AND OBJECTIVES: An optimal postoperative surveillance protocol for colorectal cancer (CRC) is dependent on understanding the time line of recurrence. By hazard function analysis, this study aimed at evaluating the time of occurrence of metastasis. METHODS: A total of 21,671 Stage I-III colon cancer patients were retrospectively included from the Japanese study group for postoperative follow-up of colorectal cancer database. RESULTS: The 5-year incidence by metastasized organ was 6.3% for liver (right:left = 5.5%:7.0%, p = .0067), 6.0% for lung (right:left:rectum = 3.7%:4.4%:8.8%, p = 7.05E-45), and 2.0% for peritoneal (right:left:rectum = 3.1%:2.0%:1.2%, p = 1.29E-12). The peak of liver metastasis hazard rate (HR) (0.67 years) was earlier and higher than those of other metastases. The peak HR tended to be delayed in early stage CRCs (0.91, 0.76, and 0.52 years; for Stages I, II, and III, respectively). When analyzed as per the primary tumor location (right-sided, left-sided, and rectum), the peak HR for lung metastasis was twice as high for rectal cancer than for colon cancer, and peritoneal metastasis had a high HR in right-sided colon cancers. CONCLUSION: The time course for the risk of recurrence in various metastatic organs based on the primary tumor site was clearly visualized in this study. This will aid in individualizing postoperative surveillance schedules.
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Neoplasias Colorrectales/patología , Cirugía Colorrectal/mortalidad , Neoplasias Hepáticas/secundario , Recurrencia Local de Neoplasia/patología , Anciano , Neoplasias Colorrectales/epidemiología , Neoplasias Colorrectales/cirugía , Femenino , Estudios de Seguimiento , Humanos , Japón/epidemiología , Funciones de Verosimilitud , Neoplasias Hepáticas/epidemiología , Neoplasias Hepáticas/cirugía , Masculino , Recurrencia Local de Neoplasia/epidemiología , Recurrencia Local de Neoplasia/cirugía , Pronóstico , Estudios Retrospectivos , Tasa de SupervivenciaRESUMEN
PURPOSE: Prognosis after peritoneal metastases in colorectal cancer is worse than that after lung or liver metastases. Previously, we demonstrated the safety of intraperitoneal (ip) administration of paclitaxel (PTX) combined with mFOLFOX6/CapeOX plus bevacizumab for colorectal cancer with peritoneal metastasis in a phase-I trial. Here, we evaluated the efficacy of this chemotherapy. METHODS: We enrolled six patients with histologically confirmed peritoneal metastases secondary to colorectal cancer. PTX was administered through a peritoneal access port, in combination with oxaliplatin-based systematic chemotherapy. Response rate, progression-free survival, 1-year survival rate, frequency of improvement in peritoneal cancer index (PCI), and cytology in peritoneal lavage were evaluated. This study was registered in the University Hospital Medical Information Network Clinical Trial Registry on July 1, 2016 (UNIN000022924). RESULTS: Three patients received the mFOLFOX6-bevacizumab regimen, whereas the other three received the CapeOX-bevacizumab regimen. The response rate was 25%. PCI score improved in 50% of the cases. Peritoneal lavage cytology that was positive in five patients before initiating the chemotherapy turned negative during chemotherapy in all patients. One-year survival rate was 100%, progression-free survival was 8.8 months (range, 6.8-12 months), and median survival time was 29.3 months. CONCLUSION: The ip administration of PTX with systemic chemotherapy can potentially control peritoneal metastases in colorectal cancer.
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Neoplasias Colorrectales , Neoplasias Peritoneales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias Colorrectales/tratamiento farmacológico , Humanos , Oxaliplatino , Paclitaxel/uso terapéutico , Neoplasias Peritoneales/tratamiento farmacológicoRESUMEN
Phosphatidylinositol 3-phosphate (PI(3)P) is the predominant phosphoinositide species in early endosomes and autophagosomes, in which PI(3)P dictates traffic of these organelles. Phosphoinositide levels are tightly regulated by lipid-kinases and -phosphatases; however, a phosphatase that converts PI(3)P back to phosphatidylinositol in the endosomal and autophagosomal compartments is not fully understood. We investigated the subcellular distribution and functions of myotubularin-related protein-4 (MTMR4), which is distinct among other MTMRs in that it possesses a PI(3)P-binding FYVE domain, in lung alveolar epithelium-derived A549 cells. MTMR4 was localized mainly in late endosomes and autophagosomes. MTMR4 knockdown markedly suppressed the motility, fusion, and fission of PI(3)P-enriched structures, resulting in decreases in late endosomes, autophagosomes, and lysosomes, and enlargement of PI(3)P-enriched early and late endosomes. In amino acid- and serum-starved cells, MTMR4 knockdown decreased both autophagosomes and autolysosomes and markedly increased PI(3)P-containing autophagosomes and late endosomes, suggesting that the fusion with lysosomes of autophagosomes and late endosomes might be impaired. Notably, MTMR4 knockdown inhibited the nuclear translocation of starvation stress responsive transcription factor-EB (TFEB) with reduced expression of lysosome-related genes in starved cells. These findings indicate that MTMR4 is essential for the integrity of endocytic and autophagic pathways.
RESUMEN
Benign multicystic peritoneal mesothelioma (BMPM) is a rare condition, particularly in men, and the preoperative diagnosis poses a challenge. Here, we present a case involving single-incision laparoscopic surgery (SILS) for BMPM in a 24-year-old man with a pelvic mass and a history of ulcerative colitis. Pelvic imaging revealed multifocal cysts, prompting the performance of SILS. The tumor was successfully resected with no residual lesions, and pathology confirmed the diagnosis of BMPM. This case represents the first documented instance of SILS being employed for BMPM in a man. BMPM, characterized by pelvic multifocal cysts, is a differential diagnosis, and SILS emerges as a viable option for both diagnosis and treatment.
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Laparoscopía , Mesotelioma Quístico , Neoplasias Peritoneales , Humanos , Masculino , Laparoscopía/métodos , Neoplasias Peritoneales/cirugía , Neoplasias Peritoneales/diagnóstico , Neoplasias Peritoneales/patología , Neoplasias Peritoneales/diagnóstico por imagen , Mesotelioma Quístico/cirugía , Mesotelioma Quístico/patología , Mesotelioma Quístico/diagnóstico , Mesotelioma Quístico/diagnóstico por imagen , Adulto JovenRESUMEN
UNLABELLED: Sinusoidal vasoconstriction, in which hepatic stellate cells operate as contractile machinery, has been suggested to play a pivotal role in the pathophysiology of portal hypertension. We investigated whether sphingosine 1-phosphate (S1P) stimulates contractility of those cells and enhances portal vein pressure in isolated perfused rat livers with Rho activation by way of S1P receptor 2 (S1P(2) ). Rho and its effector, Rho kinase, reportedly contribute to the pathophysiology of portal hypertension. Thus, a potential effect of S1P(2) antagonism on portal hypertension was examined. Intravenous infusion of the S1P(2) antagonist, JTE-013, at 1 mg/kg body weight reduced portal vein pressure by 24% without affecting mean arterial pressure in cirrhotic rats induced by bile duct ligation at 4 weeks after the operation, whereas the same amount of S1P(2) antagonist did not alter portal vein pressure and mean arterial pressure in control sham-operated rats. Rho kinase activity in the livers was enhanced in bile duct-ligated rats compared to sham-operated rats, and this enhanced Rho kinase activity in bile duct-ligated livers was reduced after infusion of the S1P(2) antagonist. S1P(2) messenger RNA (mRNA) expression, but not S1P(1) or S1P(3) , was increased in bile duct-ligated livers of rats and mice and also in culture-activated rat hepatic stellate cells. S1P(2) expression, determined in S1P 2LacZ/+ mice, was highly increased in hepatic stellate cells of bile duct-ligated livers. Furthermore, the increase of Rho kinase activity in bile duct-ligated livers was observed as early as 7 days after the operation in wildtype mice, but was less in S1P 2-/- mice. CONCLUSION: S1P may play an important role in the pathophysiology of portal hypertension with Rho kinase activation by way of S1P(2) . The S1P(2) antagonist merits consideration as a novel therapeutic agent for portal hypertension.
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Hemodinámica/efectos de los fármacos , Hipertensión Portal/tratamiento farmacológico , Pirazoles/farmacología , Piridinas/farmacología , Receptores de Lisoesfingolípidos/antagonistas & inhibidores , Quinasas Asociadas a rho/metabolismo , Animales , Conductos Biliares/cirugía , Células Cultivadas/efectos de los fármacos , Modelos Animales de Enfermedad , Activación Enzimática/efectos de los fármacos , Activación Enzimática/genética , Regulación de la Expresión Génica , Hemodinámica/fisiología , Células Estrelladas Hepáticas/efectos de los fármacos , Células Estrelladas Hepáticas/fisiología , Hipertensión Portal/fisiopatología , Immunoblotting , Inmunohistoquímica , Infusiones Intravenosas , Ligadura , Masculino , Ratones , Ratones Transgénicos , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores de Lisoesfingolípidos/efectos de los fármacos , Receptores de Lisoesfingolípidos/genética , Valores de Referencia , Sensibilidad y Especificidad , Quinasas Asociadas a rho/efectos de los fármacosRESUMEN
INTRODUCTION: At our institute, we usually perform robot-assisted surgery for rectal cancer as minimally invasive surgery. It is necessary to recognize the tumor edge accurately when deciding where to place the distal cutting line of the rectum. In this article, with video presentation, we demonstrate the usefulness of intraoperative sonography (IOUS) for detecting the rectal tumor site in robotic surgery. This is the first report to discuss the IOUS image of rectal cancer. MATERIALS AND SURGICAL TECHNIQUE: After mobilization of the rectum in robotic procedure, the rectum should be straightened. Drop the laparoscopic ultrasonography probe through the 12-mm assistant port and place it at the anterior wall of the rectum. By presenting operative and ultrasound findings simultaneously on a single monitor, the operator can recognize the tumor location accurately and decide on the cutting line. We report three cases in the supporting video presentation. DISCUSSION: Rectal tumors can be detected by IOUS, and this modality is effective for determining the cutting line of the rectum. Real-time navigation by IOUS can be performed noninvasively and easily, so it is expected to be helpful in cases of robotic rectal cancer resection.
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Laparoscopía , Neoplasias del Recto , Procedimientos Quirúrgicos Robotizados , Robótica , Humanos , Recto/cirugía , Ultrasonografía , Neoplasias del Recto/diagnóstico por imagen , Neoplasias del Recto/cirugía , Neoplasias del Recto/patología , Procedimientos Quirúrgicos Robotizados/métodos , Laparoscopía/métodosRESUMEN
BACKGROUND: Although, endothelial nitric oxide (NO) synthase (eNOS) is believed to antagonize vascular remodeling induced by the angiotensin II (AngII) type-1 receptor, the exact signaling mechanism remains unclear. METHODS AND RESULTS: By expressing eNOS to vascular smooth muscle cells (VSMCs) via adenovirus, we investigated a signal transduction mechanism of the eNOS gene transfer in preventing vascular remodeling induced by AngII. We found marked inhibition of AngII-induced Rho/Rho-kinase activation and subsequent VSMC migration by eNOS gene transfer whereas G(q)-dependent transactivation of the epidermal growth factor receptor by AngII remains intact. This could be explained by the specific inhibition of G(12/13) activation by eNOS-mediated G(12/13) phosphorylation. CONCLUSIONS: The eNOS/NO cascade specifically targets the Rho/Rho-kinase system via inhibition of G(12/13) to prevent vascular migration induced by AngII, representing a novel signal cross-talk in cardiovascular protection by NO.
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Movimiento Celular , Subunidades alfa de la Proteína de Unión al GTP G12-G13/metabolismo , Músculo Liso Vascular/enzimología , Miocitos del Músculo Liso/enzimología , Óxido Nítrico Sintasa de Tipo III/metabolismo , Receptor de Angiotensina Tipo 1/metabolismo , Quinasas Asociadas a rho/metabolismo , Adenoviridae/genética , Angiotensina II/metabolismo , Animales , Bovinos , Células Cultivadas , GMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de GMP Cíclico/metabolismo , Células Endoteliales/enzimología , Receptores ErbB/metabolismo , Vectores Genéticos , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo III/genética , Fosforilación , Proteína Fosfatasa 1/metabolismo , Ratas , Transducción de Señal , Factores de Tiempo , Transducción GenéticaRESUMEN
Class II phosphatidylinositol 3-kinases (PI3K), PI3K-C2α and PI3K-C2ß, are involved in cellular processes including endocytosis, cilia formation and autophagy. However, the role of PI3K-C2α and PI3K-C2ß at the organismal level is not well understood. We found that double knockout (KO) mice with both smooth muscle-specific KO of PI3K-C2α and global PI3K-C2ß KO, but not single KO mice of either PI3K-C2α or PI3K-C2ß, exhibited reductions in arterial blood pressure and substantial attenuation of contractile responses of isolated aortic rings. In wild-type vascular smooth muscle cells, double knockdown of PI3K-C2α and PI3K-C2ß but not single knockdown of either PI3K markedly inhibited contraction with reduced phosphorylation of 20-kDa myosin light chain and MYPT1 and Rho activation, but without inhibition of the intracellular Ca2+ mobilization. These data indicate that PI3K-C2α and PI3K-C2ß play the redundant but essential role for vascular smooth muscle contraction and blood pressure regulation mainly through their involvement in Rho activation.
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Calcio/metabolismo , Fosfatidilinositol 3-Quinasas Clase II/metabolismo , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/fisiología , Proteínas de Unión al GTP rho/metabolismo , Animales , Presión Sanguínea/fisiología , Células Cultivadas , Fosfatidilinositol 3-Quinasas Clase II/genética , Modelos Animales de Enfermedad , Isoenzimas , Ratones , Ratones Noqueados , Contracción Muscular/fisiología , Músculo Liso Vascular/citología , Músculo Liso Vascular/enzimología , Proteínas de Unión al GTP rho/genéticaRESUMEN
Pinocytosis is an important fundamental cellular process that is used by the cell to transport fluid and solutes. Phosphoinositide 3-kinases (PI3Ks) regulate a diverse array of dynamic membrane events. However, it is not well-understood which PI3K isoforms are involved in specific mechanisms of pinocytosis. We performed knockdown studies of endogenous PI3K isoforms and clathrin heavy chain (CHC) mediated by small interfering RNA (siRNA). The results demonstrated that the class II PI3K PI3K-C2α and PI3K-C2ß, but not the class I or III PI3K, were required for pinocytosis, based on an evaluation of fluorescein-5-isothiocyanate (FITC)-dextran uptake in endothelial cells. Pinocytosis was partially dependent on both clathrin and dynamin, and both PI3K-C2α and PI3K-C2ß were required for clathrin-mediated-but not clathrin-non-mediated-FITC-dextran uptake at the step leading up to its delivery to early endosomes. Both PI3K-C2α and PI3K-C2ß were co-localized with clathrin-coated pits and vesicles. However, PI3K-C2ß, but not PI3K-C2α, was highly co-localized with actin filament-associated clathrin-coated structures and required for actin filament formation at the clathrin-coated structures. These results indicate that PI3K-C2α and PI3K-C2ß play differential, indispensable roles in clathrin-mediated pinocytosis.
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Fosfatidilinositol 3-Quinasas Clase II/metabolismo , Clatrina/metabolismo , Células Endoteliales/metabolismo , Células Endoteliales/fisiología , Fosfatidilinositol 3-Quinasas/metabolismo , Pinocitosis/fisiología , Citoesqueleto de Actina/metabolismo , Animales , Células Cultivadas , Células Endoteliales de la Vena Umbilical Humana , Humanos , ARN Interferente Pequeño/metabolismoRESUMEN
Class II phosphoinositide 3-kinases (PI3Ks), PI3K-C2α and PI3K-C2ß, are highly homologous and distinct from class I and class III PI3Ks in catalytic products and domain structures. In contrast to class I and class III PI3Ks, physiological roles of PI3K-C2α and PI3K-C2ß are not fully understood. Because we previously demonstrated that PI3K-C2α is involved in vascular smooth muscle contraction, we studied the phenotypes of smooth muscle-specific knockout (KO) mice of PI3K-C2α and PI3K-C2ß. The pup numbers born from single PI3K-C2α-KO and single PI3K-C2ß-KO mothers were similar to those of control mothers, but those from double KO (DKO) mothers were smaller compared with control mice. However, the number of intrauterine fetuses in pregnant DKO mothers was similar to that in control mice. Both spontaneous and oxytocin-induced contraction of isolated uterine smooth muscle (USM) strips was diminished in DKO mice but not in either of the single KO mice, compared with control mice. Furthermore, contraction of USM of DKO mice was less sensitive to a Rho kinase inhibitor. Mechanistically, the extent of oxytocin-induced myosin light chain phosphorylation was greatly reduced in USM from DKO mice compared with control mice. The oxytocin-induced rise in the intracellular Ca2+ concentration in USM was similar in DKO and control mice. However, Rho activation in the intracellular compartment was substantially attenuated in DKO mice compared with control mice, as evaluated by fluorescence resonance energy transfer imaging technique. These data indicate that both PI3K-C2α and PI3K-C2ß are required for normal USM contraction and parturition mainly through their involvement in Rho activation.
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Fosfatidilinositol 3-Quinasas Clase II/metabolismo , Músculo Liso Vascular/enzimología , Parto , Fosfatidilinositol 3-Quinasas/metabolismo , Contracción Uterina , Útero/enzimología , Proteína de Unión al GTP rhoA/metabolismo , Animales , Fosfatidilinositol 3-Quinasas Clase II/genética , Femenino , Ratones , Ratones Noqueados , Contracción Muscular , Músculo Liso Vascular/fisiología , Cadenas Ligeras de Miosina , Fosfatidilinositol 3-Quinasas/genética , Fosforilación , Útero/fisiología , Proteína de Unión al GTP rhoA/genéticaRESUMEN
Atherosclerosis is the major cause of ischemic coronary heart diseases and characterized by the infiltration of cholesterol-accumulating macrophages in the vascular wall. Although sphingolipids are implicated in atherosclerosis as both membrane components and lipid mediators, the precise role of sphingolipids in atherosclerosis remains elusive. Here, we found that genetic deficiency of sphingosine kinase-2 (SphK2) but not SphK1 aggravates the formation of atherosclerotic lesions in mice with ApoE deficiency. Bone marrow chimaera experiments show the involvement of SphK2 expressed in bone marrow-derived cells. In macrophages, deficiency of SphK2, a major SphK isoform in this cell type, results in increases in cellular sphingosine and ceramides. SphK2-deficient macrophages have increases in lipid droplet-containing autophagosomes and autolysosomes and defective lysosomal degradation of lipid droplets via autophagy with an impaired luminal acidic environment and proteolytic activity in the lysosomes. Transgenic overexpression of SphK1 in SphK2-deficient mice rescued aggravation of atherosclerosis and abnormalities of autophagosomes and lysosomes in macrophages with reductions of sphingosine, suggesting at least partial overlapping actions of two SphKs. Taken together, these results indicate that SphK2 is required for autophagosome- and lysosome-mediated catabolism of intracellular lipid droplets to impede the development of atherosclerosis; therefore, SphK2 may be a novel target for treating atherosclerosis.
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Apolipoproteínas E/genética , Aterosclerosis/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Animales , Aterosclerosis/metabolismo , Aterosclerosis/patología , Células de la Médula Ósea/metabolismo , Colesterol/metabolismo , Modelos Animales de Enfermedad , Humanos , Metabolismo de los Lípidos/genética , Lípidos/genética , Macrófagos/metabolismo , Macrófagos/patología , Ratones , Esfingolípidos/genética , Esfingolípidos/metabolismo , Esfingosina/metabolismoRESUMEN
PURPOSE: Peritoneal carcinomatosis of colorectal cancer origin is associated with poor prognosis. With regard to ovarian, gastric, and pancreatic cancer, the safety and efficacy of intraperitoneal administration of paclitaxel (ip PTX) has been demonstrated. This drug can be administered easily and repeatedly through a catheter into the peritoneal cavity. In this phase I study, we evaluated the safety of ip PTX combined with 5-fluorouracil, folinic acid, oxaliplatin, and bevacizumab (mFOLFOX6-bevacizumab) or capecitabine, oxaliplatin, and bevacizumab (CapeOX-bevacizumab) for colorectal cancer with peritoneal metastasis. METHODS: Colorectal cancer patients with histologically confirmed peritoneal carcinomatosis were enrolled. After the implantation of a peritoneal access port, 20 mg/m2 of ip PTX was administered weekly, in combination with mFOLFOX6-bevacizumab or CapeOX-bevacizumab. Primary endpoint was the safety of the combination chemotherapy. RESULTS: Among the six patients enrolled, three received the mFOLFOX6-bevacizumab plus ip PTX regimen and three received the CapeOX-bevacizumab plus ip PTX regimen. Dose-limiting toxicity was not observed. Overall, grade 3 adverse events, such as leukopenia and neutropenia, were observed in two of three patients (66.7%) for each chemotherapeutic regimen, but no grade 4 adverse events were observed. Moreover, adverse events associated with the peritoneal access port, such as infection or occlusion of the catheter, were not observed. CONCLUSIONS: The adverse events of mFOLFOX6-bevacizumab or CapeOX-bevacizumab in combination with ip PTX were considered similar to those described in previous studies of oxaliplatin-based treatment alone. 1 year after the start of chemotherapy, the efficacy of ip PTX will be evaluated as a secondary outcome.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Peritoneales/tratamiento farmacológico , Adulto , Anciano , Anciano de 80 o más Años , Bevacizumab/administración & dosificación , Capecitabina/administración & dosificación , Neoplasias Colorrectales/patología , Femenino , Fluorouracilo/administración & dosificación , Estudios de Seguimiento , Humanos , Leucovorina/administración & dosificación , Masculino , Persona de Mediana Edad , Oxaliplatino/administración & dosificación , Paclitaxel/administración & dosificación , Seguridad del Paciente , Neoplasias Peritoneales/secundario , Pronóstico , Adulto JovenRESUMEN
Idiopathic pulmonary fibrosis is a devastating disease with poor prognosis. The pathogenic role of the lysophospholipid mediator sphingosine-1-phosphate and its receptor S1PR2 in lung fibrosis is unknown. We show here that genetic deletion of S1pr2 strikingly attenuated lung fibrosis induced by repeated injections of bleomycin in mice. We observed by using S1pr2LacZ/+ mice that S1PR2 was expressed in alveolar macrophages, vascular endothelial cells and alveolar epithelial cells in the lung and that S1PR2-expressing cells accumulated in the fibrotic legions. Bone marrow chimera experiments suggested that S1PR2 in bone marrow-derived cells contributes to the development of lung fibrosis. Depletion of macrophages greatly attenuated lung fibrosis. Bleomycin administration stimulated the mRNA expression of the profibrotic cytokines IL-13 and IL-4 and the M2 markers including arginase 1, Fizz1/Retnla, Ccl17 and Ccl24 in cells collected from broncho-alveolar lavage fluids (BALF), and S1pr2 deletion markedly diminished the stimulated expression of these genes. BALF cells from bleomycin-administered wild-type mice showed a marked increase in phosphorylation of STAT6, a transcription factor which is activated downstream of IL-13, compared with saline-administered wild-type mice. Interestingly, in bleomycin-administered S1pr2-/- mice, STAT6 phosphorylation in BALF cells was substantially diminished compared with wild-type mice. Finally, pharmacological S1PR2 blockade in S1pr2+/+ mice alleviated bleomycin-induced lung fibrosis. Thus, S1PR2 facilitates lung fibrosis through the mechanisms involving augmentation of IL-13 expression and its signaling in BALF cells, and represents a novel target for treating lung fibrosis.
Asunto(s)
Fibrosis Pulmonar Idiopática/etiología , Interleucina-13/metabolismo , Macrófagos/metabolismo , Receptores de Lisoesfingolípidos/metabolismo , Animales , Bleomicina/toxicidad , Líquido del Lavado Bronquioalveolar/química , Líquido del Lavado Bronquioalveolar/citología , Modelos Animales de Enfermedad , Fibrosis Pulmonar Idiopática/metabolismo , Fibrosis Pulmonar Idiopática/patología , Interleucina-13/genética , Macrófagos/efectos de los fármacos , Macrófagos/patología , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores de Lisoesfingolípidos/deficiencia , Receptores de Lisoesfingolípidos/genética , Factor de Transcripción STAT6/metabolismo , Transducción de Señal , Receptores de Esfingosina-1-Fosfato , Quimera por Trasplante/genética , Quimera por Trasplante/metabolismo , Regulación hacia ArribaRESUMEN
Inflammation and apoptosis develop in skeletal muscle after major trauma, including burn injury, and play a pivotal role in insulin resistance and muscle wasting. We and others have shown that inducible nitric oxide synthase (iNOS), a major mediator of inflammation, plays an important role in stress (e.g., burn)-induced insulin resistance. However, it remains to be determined how iNOS induces insulin resistance. Moreover, the interrelation between inflammatory response and apoptosis is poorly understood, although they often develop simultaneously. Nuclear factor (NF)-κB and p53 are key regulators of inflammation and apoptosis, respectively. Sirt1 inhibits p65 NF-κB and p53 by deacetylating these transcription factors. Recently, we have shown that iNOS induces S-nitrosylation of Sirt1, which inactivates Sirt1 and thereby increases acetylation and activity of p65 NF-κB and p53 in various cell types, including skeletal muscle cells. Here, we show that iNOS enhances burn-induced inflammatory response and apoptotic change in mouse skeletal muscle along with S-nitrosylation of Sirt1. Burn injury induced robust expression of iNOS in skeletal muscle and gene disruption of iNOS significantly inhibited burn-induced increases in inflammatory gene expression and apoptotic change. In parallel, burn increased Sirt1 S-nitrosylation and acetylation and DNA-binding capacity of p65 NF-κB and p53, all of which were reversed or ameliorated by iNOS deficiency. These results indicate that iNOS functions not only as a downstream effector but also as an upstream enhancer of burn-induced inflammatory response, at least in part, by Sirt1 S-nitrosylation-dependent activation (acetylation) of p65 NF-κB. Our data suggest that Sirt1 S-nitrosylation may play a role in iNOS-mediated enhanced inflammatory response and apoptotic change, which, in turn, contribute to muscle wasting and supposedly to insulin resistance after burn injury.
Asunto(s)
Apoptosis/fisiología , Quemaduras/patología , Inflamación/patología , Músculo Esquelético/patología , Óxido Nítrico Sintasa de Tipo II/metabolismo , Sirtuina 1/metabolismo , Factor de Transcripción ReIA/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Acetilación , Animales , Proteínas de Unión al ADN/metabolismo , Activación Enzimática , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Óxido Nítrico Sintasa de Tipo II/genéticaRESUMEN
Pneumatosis cystoides intestinalis (PCI) is a rare gastrointestinal complication of systemic sclerosis (SSc) characterized by intramural accumulation of gas within thin-walled cysts. We report the case of an 82-year-old female patient with pneumoperitoneum due to PCI associated with SSc and review the features of the 39 Japanese cases. The median patient age was 57 years (range 24-83 years) and the male/female ratio was 1 : 12. In the recent decade, 14 out of 15 cases (93.3%) evaluated with CT scans were diagnosed with PCI. The results suggest that CT scan may be a useful diagnostic tool for detecting PCI. PCI in patients with SSc is usually benign and requires only conservative therapy. However, two patients (5.1%) with signs of peritoneal irritation required surgery. When peritoneal irritation secondary to additional pathology is observed, surgical treatment may be warranted; a precise diagnosis for this condition is therefore essential.
RESUMEN
The aim of the present study was to evaluate the effect of underweight status on the survival of elderly patients undergoing surgery for colorectal cancer (CRC). A total of 113 patients aged ≥75 years who underwent curative surgery for CRC were included. In addition to standard perioperative variables, body mass index (BMI) was assessed. The patients were categorized as underweight (BMI<18.5 kg/m2) or non-underweight (BMI≥18.5 kg/m2). The 3-year overall survival (OS) and cancer-specific survival (CSS) were analyzed. Of the 113 patients, 24 (21%) were underweight. The two groups were well-balanced regarding all factors evaluated. In the multivariate analysis, underweight status was an independent indicator of lower 3-year OS [hazard ratio (HR)=2.65; 95% confidence interval (CI): 1.08-6.50; P=0.033] and CSS (HR=3.51, 95% CI: 1.16-10.60; P=0.025) rates. Compared with the non-underweight group, the underweight group had significantly worse 3-year OS (66.7 vs. 86.5%, respectively; P=0.017) and CSS (74.1 vs. 90.9%, respectively; P=0.025) rates. Therefore, underweight status was a significant risk factor for poor survival in elderly CRC patients. The development of effective nutritional interventions may improve the prognosis of such patients.