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Chromosome pairing constitutes an important level of genome organization, yet the mechanisms that regulate pairing in somatic cells and the impact on 3D chromatin organization are still poorly understood. Here, we address these questions in Drosophila, an organism with robust somatic pairing. In Drosophila, pairing preferentially occurs at loci consisting of numerous architectural protein binding sites (APBSs), suggesting a role of architectural proteins (APs) in pairing regulation. Amongst these, the anti-pairing function of the condensin II subunit CAP-H2 is well established. However, the factors that regulate CAP-H2 localization and action at APBSs remain largely unknown. Here, we identify two factors that control CAP-H2 occupancy at APBSs and, therefore, regulate pairing. We show that Z4, interacts with CAP-H2 and is required for its localization at APBSs. We also show that hyperosmotic cellular stress induces fast and reversible unpairing in a Z4/CAP-H2 dependent manner. Moreover, by combining the opposite effects of Z4 depletion and osmostress, we show that pairing correlates with the strength of intrachromosomal 3D interactions, such as active (A) compartment interactions, intragenic gene-loops, and polycomb (Pc)-mediated chromatin loops. Altogether, our results reveal new players in CAP-H2-mediated pairing regulation and the intimate interplay between inter-chromosomal and intra-chromosomal 3D interactions.
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Adenosina Trifosfatases , Cromatina , Pareamento Cromossômico , Proteínas de Ligação a DNA , Proteínas de Drosophila , Animais , Adenosina Trifosfatases/metabolismo , Adenosina Trifosfatases/genética , Sítios de Ligação , Cromatina/metabolismo , Proteínas Cromossômicas não Histona , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Proteínas de Drosophila/metabolismo , Proteínas de Drosophila/genética , Complexos Multiproteicos/metabolismo , Complexos Multiproteicos/genética , Pressão Osmótica , Ligação Proteica , Dedos de ZincoRESUMO
BACKGROUND: Delirium is one of the most common adverse events in older people during hospitalization, especially in the emergency department. Reliable, easy-to-use instruments are necessary to properly manage delirium in this setting. This study aims to evaluate the diagnostic validity of the Spanish version of the 4 'A's Test (4AT) in the ED. METHODS: A diagnostic accuracy study was conducted in patients over 65 years old admitted to the Emergency Department who did not have a formal diagnosis of dementia or a severe mental health disorder. Face and content validity were evaluated by an expert panel. Emergency nurses performed the evaluation with 4AT, whilst blinded and trained researchers assessed patients with the Revised Delirium Rating Scale as the gold standard. The content validity index, sensitivity, specificity, positive and negative predictive values, likelihood ratios, Youden's Index and ROC curves were calculated to evaluate the diagnostic accuracy of the instrument. RESULTS: Of 393 eligible patients, 380 were finally analyzed. Content validity yielded a median content validity index of 4 (interquartile range: 0). The Spanish 4AT sensitivity (95.83%; 95% ECI: 78.9-99.9%), specificity (92.98%; 95% CI: 89.8-95.4%), positive predictive value (47.92%) and negative predictive value (99.7%) were satisfactory. Youden's index was 0.89. Positive likelihood ratio was 13.65, and negative likelihood ratio 0.045. The area under the curve was 0.97. CONCLUSIONS: The Spanish version of the 4AT for use in the Emergency Departments is easy-to-use and applicable. The validation results indicate that it is a valid instrument with sufficient predictive validity to identify patients at risk of delirium in the Emergency Departments. Moreover, it is a tool that facilitates the management of an adverse event that is associated with increased mortality and morbidity.
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In the nucleus, chromatin is intricately structured into multiple layers of 3D organization important for genome activity. How distinct layers influence each other is not well understood. In particular, the contribution of chromosome pairing to 3D chromatin organization has been largely neglected. Here, we address this question in Drosophila, an organism that shows robust chromosome pairing in interphasic somatic cells. The extent of chromosome pairing depends on the balance between pairing and anti-pairing factors, with the anti-pairing activity of the CAP-H2 condensin II subunit being the best documented. Here, we identify the zinc-finger protein Z4 as a strong anti-pairer that interacts with and mediates the chromatin binding of CAP-H2. We also report that hyperosmotic cellular stress induces fast and reversible chromosome unpairing that depends on Z4/CAP-H2. And, most important, by combining Z4 depletion and osmostress, we show that chromosome pairing reinforces intrachromosomal 3D interactions. On the one hand, pairing facilitates RNAPII occupancy that correlates with enhanced intragenic gene-loop interactions. In addition, acting at a distance, pairing reinforces chromatin-loop interactions mediated by Polycomb (Pc). In contrast, chromosome pairing does not affect which genomic intervals segregate to active (A) and inactive (B) compartments, with only minimal effects on the strength of A-A compartmental interactions. Altogether, our results unveil the intimate interplay between inter-chromosomal and intra-chromosomal 3D interactions, unraveling the interwoven relationship between different layers of chromatin organization and the essential contribution of chromosome pairing.
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(1) Background: There is currently a global consensus that the quality of comprehensive care for acutely hospitalised elderly people should include addressing functionality and mobility, cognitive status, prevention of pressure ulcers, urinary incontinence, falls and delirium, as well as pain control and medication-related problems. The aim of this study is to develop and validate a clinical prediction rule for multimorbid patients admitted to an acute care hospital unit for any of the five adverse events included in our vulnerability pentad: falls, pressure ulcers, urinary incontinence, pain and delirium. (2) Methods: Longitudinal analytical clinimetric study, with two cohorts. The study population will consist of multimorbid patients hospitalised for acute care, referred from the Emergency Room. A clinical prediction rule will be proposed, incorporating predictive factors of these five adverse outcomes described. This study has received funding, awarded in November 2020 (PI-0107-2020), and was approved in October 2019 by the Research Ethics Committee â³Costa del Solâ³. (3) Conclusions: Preventing adverse events in hospitalised patients is particularly important for those with multimorbidity. By applying a clinical prediction rule to detect specific risks, an estimate can be obtained of their probability of occurrence.
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Delírio , Úlcera por Pressão , Incontinência Urinária , Idoso , Regras de Decisão Clínica , Delírio/diagnóstico , Hospitalização , Humanos , Multimorbidade , Dor , Úlcera por Pressão/epidemiologiaRESUMO
AHCC® is a standardized extract of cultured mushroom (Lentinula edodes) mycelia with a wide variety of therapeutic effects including anti-inflammatory, antitumor and antiviral effects. Trichinellosis, a food-borne parasitic zoonosis is caused by the nematode Trichinella spp. Infection with Trichinella is characterized by the induction of a Th1-type response at the beginning of the intestinal phase, followed by a dominant Th2-type response which is essential for parasite expulsion. The aim of this study was to evaluate the immunomodulatory effect of AHCC® in a murine model of Trichinella spiralis infection. Swiss CD1 mice were infected with T. spiralis larvae and treated with AHCC®. Standard treatment with albendazole (ABZ) was used as control in the assessment of parasite burden. The small intestine was taken out and the proximal segment was evaluated for several parameters: gene expression of immune and stress-reticulum mediators, histological damage score, goblet cell count and Mucin 2 (Muc2) gene expression. AHCC® modulated expression levels of both Th1 and Th2 cytokines and reduced histological damage score. In addition, AHCC® diminished the number of adults of T. spiralis in treated animals. AHCC® treatment anticipates T. spiralis expulsion and increases goblet cell number and Muc2 gene expression.
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Mucina-2 , Cogumelos Shiitake , Trichinella spiralis , Triquinelose , Animais , Contagem de Células , Doenças Inflamatórias Intestinais/tratamento farmacológico , Doenças Inflamatórias Intestinais/metabolismo , Doenças Inflamatórias Intestinais/parasitologia , Camundongos , Mucina-2/antagonistas & inibidores , Mucina-2/biossíntese , Cogumelos Shiitake/química , Trichinella spiralis/efeitos dos fármacos , Triquinelose/tratamento farmacológicoRESUMO
Background: Dysbiosis and mucin depletion are related with intestinal barrier dysfunction and seems to be an early pathophysiological event in inflammatory bowel disease (IBD). The objective of this work is to study these parameters in the natural history of colitis in IL-10 deficient mice (IL-10-/-). Methods: Wild type (WT) and IL-10-/-. mice were followed until sacrifice at 3, 5, 10, 20, 57, and 70 weeks. Body weight, colonic weight/length ratio and in vivo intestinal permeability were registered. Expression of inflammatory and adhesion molecules in the colon was explored by qPCR as Mucin-2 (MUC-2) and molecules involved in goblet cell maturation Interleukin-18 (IL-18) and WAP Four-Disulfide Core Domain 2 (WFDC2), the endoplasmic reticulum stress markers X-box-binding protein (Xbp-1) and Reticulon-4B (RTN-4B). Bacterial composition in feces and colonic mucosa was determined by massive sequencing of the V3-V4 regions of 16S rDNA gene. Results: IL-10-/- mice showed histological inflammation at weeks 20 and 57, but most notably the intestinal permeability was significantly higher from week 10. Concordantly, the number of goblet cells and expression of MUC-2, IL-18, WFDC2 and Xbp-1 were significantly lower in KO from week 10. Nevertheless, no significant differences were found in the mRNA expression of MUC-2 or Xbp-1 between both groups-derived colon organoids. Significant bacterial differences began at week 5, being the Akkermansia deficiency in KO the most relevant result. Conclusion: Gut microbiota alterations and mucin depletion are associated with early intestinal barrier dysfunction and precede overt gut inflammation in this animal model of IBD.
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The increased risk of cardiovascular disease (CVD) conferred by hepatitis C virus (HCV) is especially relevant after liver transplantation (LT), but its mechanism is still not well defined. This study aimed to evaluate the influence of HCV eradication in inflammatory and endothelial activation markers after LT. We evaluated inflammatory (TNF-alfa, IL-6, IL-8, and MCP-1) and endothelial activation (E-selectin, ICAM-1, VCAM-1, and MMP-9) markers before and after eradication in 45 LT recipients with HCV infection (LT+/HCV+) and 44 non-transplanted HCV-infected patients (LT-/HCV+). We also considered an additional group of 40 LT recipients without HCV infection (LT+/HCV-). LT+/HCV+ patients presented a higher endothelial activation status before eradication compared with LT+/HCV- patients. However, levels of E-selectin, ICAM-1, VCAM-1, and MMP-9 were comparable between LT+/HCV+ and LT-/HCV+ patients before eradication. HCV eradication decreased ICAM-1 (5466.55 pg/ml vs. 3354.88 pg/ml, P < 0.001) and VCAM-1 (10456.52 pg/ml vs. 6658.85 pg/ml, P < 0.001) levels in LT+/HCV+ and LT-/HCV+ patients. Remarkably, HCV eradication restored levels of endothelial activation markers of LT+/HCV+ patients compared with that of LT+/HCV- patients. HCV plays a major role in endothelial dysfunction after LT. Furthermore, HCV eradication restores endothelial activation despite the exposure to immunosuppressive therapy.
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Hepatite C Crônica , Hepatite C , Transplante de Fígado , Antivirais/uso terapêutico , Hepacivirus , Hepatite C/complicações , Hepatite C/tratamento farmacológico , Hepatite C Crônica/tratamento farmacológico , HumanosRESUMO
BACKGROUND: Liver X receptor (LXR) exerts anti-inflammatory effects in macrophages. The aim of this study was to explore the expression and function of LXR in the colonic epithelium under inflammatory conditions. METHODS: The expression of LXR was explored by Western blot and immunohistochemistry in colonic biopsies from patients diagnosed with inflammatory bowel disease (IBD) and control patients. In addition, LXR and its target gene expression were analyzed in the colon from interleukin (IL)-10-deficient (IL-10-/-) and wild-type mice. Caco-2 cells were pretreated with the synthetic LXR agonist GW3965 and further challenged with IL-1ß, the expression of IL-8 and chemokine (C-C motif) ligand (CCL)-28 chemokines, the activation of mitogen-activated protein (MAP) kinases, and the nuclear translocation of the p65 subunit of nuclear factor kappa B was evaluated. Glibenclamide was used as an ABCA1 antagonist. RESULTS: We found that LXR expression was downregulated in colonic samples from patients with IBD and IL-10-/- mice. The nuclear positivity of LXR inversely correlated with ulcerative colitis histologic activity. Colonic IL-1ß mRNA levels negatively correlated with both LXRα and LXRß in the colon of IL-10-/- mice, where a decreased mRNA expression of the LXR target genes ABCA1 and FAS was shown. In addition, IL-1ß decreased the expression of the LXR target gene ABCA1 in cultured intestinal epithelial cells. The synthetic LXR agonist GW3965 led to a decreased nuclear positivity of the p65 subunit of nuclear factor kappa B, a phosphorylation ratio of the p44-42 MAP kinase, and the expression of CCL-28 and IL-8 in IL-1ß-stimulated Caco-2 cells. The pharmacological inhibition of ABCA1 increased the phosphorylation of p44-42 after GW3965 treatment and IL-1ß stimulation. CONCLUSIONS: The LXR-ABCA1 pathway exerts anti-inflammatory effects in intestinal epithelial cells and is impaired in the colonic mucosa of patients with IBD and IL-10-/- mice.
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Colite , Doenças Inflamatórias Intestinais , Transportador 1 de Cassete de Ligação de ATP/química , Transportador 1 de Cassete de Ligação de ATP/genética , Transportador 1 de Cassete de Ligação de ATP/metabolismo , Animais , Anti-Inflamatórios , Células CACO-2 , Colite/induzido quimicamente , Colite/tratamento farmacológico , Células Epiteliais , Humanos , Doenças Inflamatórias Intestinais/tratamento farmacológico , Interleucina-10 , Interleucina-8/genética , Interleucina-8/metabolismo , Receptores X do Fígado , Camundongos , NF-kappa B , Receptores Nucleares Órfãos/genética , RNA MensageiroRESUMO
BACKGROUND: informal caregivers have a high risk of suffering from diseases derived from the chronic stress to which they are subjected for their dedication to the care of their relatives. Such stress has a direct influence on the person cared for, mainly affecting the quality of their care. Therefore, the aim of the present study is to assess the association of caregiving on physical and mental perceived health in family caregivers of dependent adults with complex chronic diseases. METHODS: a prospective longitudinal cohort study, with a follow-up period of 36 months (HUELLA cohort). The exposed cohort will be formed by family caregivers of dependent patients with complex chronic pathologies. The unexposed cohort will be taken from the general population adjusted for age, sex and health. Outcome variables will include attendance to health services, consumption of psychoactive drugs, dedication to care (only in exposed cohort), concession of the Act on Promotion of Personal Autonomy and Care for dependent persons (exposed only), perceived physical and mental health, depression level, burden level and new diagnosis of chronic pathology of the caregiver during the study. RESULTS: the expected results will be applicable and will incorporate improvements to the usual health system clinical practice, providing feedback to professionals dedicated to the provision, planning and design of services to family caregivers, as well as to groups and organizations of caregivers. CONCLUSIONS: investments in preventing low-quality informal care are key, mainly through early identification and interventions to support caregivers who suffer from stress, anxiety or depression.
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Cuidadores , Saúde Mental , Qualidade de Vida , Adolescente , Adulto , Cuidadores/psicologia , Estudos de Casos e Controles , Criança , Doença Crônica , Estudos de Coortes , Estudos Transversais , Humanos , Estudos Longitudinais , Estudos ProspectivosRESUMO
The selection of the anesthetic regime is a crucial component in many experimental animal studies. In rodent models of liver disease, the combination of ketamine and diazepam (KD), generally by the intramuscular (i.m.) route, has traditionally been the anesthesia of choice for the evaluation of systemic and hepatic hemodynamics but it presents several problems. Here, we compared the performance of inhalational sevoflurane (Sevo) against the KD combination as the anesthesia used for hemodynamic studies involving the measurement of portal pressure in normal rats (Ctrl) and rats with non-cirrhotic portal hypertension induced by partial portal vein ligation (PPVL). Compared with Ctrl rats, rats with PPVL presented characteristic alterations that were not influenced by the anesthetic regime, which included liver atrophy, splenomegaly, increased plasma fibrinogen, decreased alkaline phosphatase and glycemia, and frequent ascites. The use of the KD combination presented several disadvantages compared with the inhalational anesthesia with sevoflurane, including considerable mortality, a higher need of dose adjustments to maintain an optimal depth of anesthesia, increases of heart rate, and alteration of blood biochemical parameters such as the concentration of aspartate aminotransferase, lactate, and lactic dehydrogenase. Rats anesthetized with sevoflurane, on the other hand, presented lower respiratory rates. Importantly, the anesthetic regime did not influence the measurement of portal pressure either in Ctrl or PPVL rats, with the increase of portal pressure being similar in Sevo- and KD- anesthetized groups of PPVL rats compared with their respective control groups. Overall, our results suggest that anesthesia with sevoflurane is preferable to the combination of KD for performing systemic and hepatic hemodynamic studies in rats with non-cirrhotic portal hypertension.
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Diazepam/farmacologia , Hipertensão Portal/tratamento farmacológico , Ketamina/farmacologia , Pressão na Veia Porta/efeitos dos fármacos , Sevoflurano/farmacologia , Anestesia por Inalação , Anestesiologia , Anestésicos Dissociativos/farmacologia , Anestésicos Inalatórios/farmacologia , Animais , Modelos Animais de Doenças , Fígado/irrigação sanguínea , Masculino , Veia Porta/efeitos dos fármacos , Ratos , Ratos Sprague-DawleyRESUMO
OBJECTIVE: Non-alcoholic steatohepatitis (NASH) is characterized by a robust pro-inflammatory component at both hepatic and systemic levels together with a disease-specific gut microbiome signature. Protein tyrosine phosphatase 1 B (PTP1B) plays distinct roles in non-immune and immune cells, in the latter inhibiting pro-inflammatory signaling cascades. In this study, we have explored the role of PTP1B in the composition of gut microbiota and gut barrier dynamics in methionine and choline-deficient (MCD) diet-induced NASH in mice. METHODS: Gut features and barrier permeability were characterized in wild-type (PTP1B WT) and PTP1B-deficient knockout (PTP1B KO) mice fed a chow or methionine/choline-deficient (MCD) diet for 4 weeks. The impact of inflammation was studied in intestinal epithelial and enteroendocrine cells. The secretion of GLP-1 was evaluated in primary colonic cultures and plasma of mice. RESULTS: We found that a shift in the gut microbiota shape, disruption of gut barrier function, higher levels of serum bile acids, and decreased circulating glucagon-like peptide (GLP)-1 are features during NASH. Surprisingly, despite the pro-inflammatory phenotype of global PTP1B-deficient mice, they were partly protected against the alterations in gut microbiota composition during NASH and presented better gut barrier integrity and less permeability under this pathological condition. These effects concurred with higher colonic mucosal inflammation, decreased serum bile acids, and protection against the decrease in circulating GLP-1 levels during NASH compared with their WT counterparts together with increased expression of GLP-2-sensitive genes in the gut. At the molecular level, stimulation of enteroendocrine STC-1 cells with a pro-inflammatory conditioned medium (CM) from lipopolysaccharide (LPS)-stimulated macrophages triggered pro-inflammatory signaling cascades that were further exacerbated by a PTP1B inhibitor. Likewise, the pro-inflammatory CM induced GLP-1 secretion in primary colonic cultures, an effect augmented by PTP1B inhibition. CONCLUSION: Altogether our results have unraveled a potential role of PTP1B in the gut-liver axis during NASH, likely mediated by increased sensitivity to GLPs, with potential therapeutic value.
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Microbioma Gastrointestinal/genética , Mucosa Intestinal/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 1/deficiência , Proteína Tirosina Fosfatase não Receptora Tipo 1/genética , Animais , Deficiência de Colina/complicações , Dieta/efeitos adversos , Modelos Animais de Doenças , Expressão Gênica , Técnicas de Inativação de Genes , Peptídeo 1 Semelhante ao Glucagon/sangue , Inflamação/metabolismo , Fígado/metabolismo , Masculino , Metionina/deficiência , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/etiologia , Permeabilidade , Células RAW 264.7RESUMO
Despite being a cardinal experimental model, the induction of cirrhosis in rats by repeated exposure to carbon tetrachloride (CCl4) has low reproducibility. Here, we compared two models of cirrhosis induced by orogastric administration of CCl4 once (CCl4-1xWk) or twice a week (CCl4-2xWk) for 12 weeks in male Sprague-Dawley rats. Control rats received water instead of CCl4. Both CCl4 protocols similarly attenuated body weight gain (p < 0.01 vs. Control). Although both CCl4 protocols increased hepatic fibrosis, portal hypertension and splenomegaly, the magnitude of these alterations was higher and more consistent in CCl4-2xWk rats. Importantly, two CCl4-1xWk rats did not develop cirrhosis versus a 100% yield of cirrhosis in CCl4-2xWk rats. The CCl4-2xWk protocol consistently induced liver atrophy together with hematological, biochemical and coagulation abnormalities characteristic of advanced cirrhosis that were absent in CCl4-1xWk rats. Ascites occurred in 20% and 80% of rats in theCCl4-1xWk and CCl4-2xWk groups (p < 0.01). All rats showed normal renal function, arterial blood gases and stable systemic hemodynamics. The total dose of CCl4 and mortality rate were similar in both protocols. The CCl4-2xWk protocol, therefore, was highly reproducible and effective for the induction of experimental cirrhosis within a confined time, representing a valuable advance for liver research.
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Intoxicação por Tetracloreto de Carbono , Hipertensão Portal , Cirrose Hepática , Fígado , Esplenomegalia , Animais , Intoxicação por Tetracloreto de Carbono/metabolismo , Intoxicação por Tetracloreto de Carbono/patologia , Modelos Animais de Doenças , Hipertensão Portal/induzido quimicamente , Hipertensão Portal/metabolismo , Hipertensão Portal/patologia , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Ratos , Ratos Sprague-Dawley , Esplenomegalia/induzido quimicamente , Esplenomegalia/metabolismo , Esplenomegalia/patologiaRESUMO
BACKGROUND & AIMS: Recent studies suggest that heparins reduce liver fibrosis and the risk of decompensation of liver disease. Here, we evaluated the effects of enoxaparin in several experimental models of advanced cirrhosis. METHODS: Cirrhosis was induced in male Sprague-Dawley (SD) rats by: (i) Oral gavage with carbon tetrachloride (CCl4ORAL ), (ii) Bile duct ligation (BDL) and (iii) CCl4 inhalation (CCl4INH ). Rats received saline or enoxaparin s.c. (40 IU/Kg/d or 180 IU/Kg/d) following various protocols. Blood biochemical parameters, liver fibrosis, endothelium- and fibrosis-related genes, portal pressure, splenomegaly, bacterial translocation, systemic inflammation and survival were evaluated. Endothelial dysfunction was assessed by in situ bivascular liver perfusions. RESULTS: Enoxaparin did not ameliorate liver function, liver fibrosis, profibrogenic gene expression, portal hypertension, splenomegaly, ascites development and infection, serum IL-6 levels or survival in rats with CCl4ORAL or BDL-induced cirrhosis. Contrarily, enoxaparin worsened portal pressure in BDL rats and decreased survival in CCl4ORAL rats. In CCl4INH rats, enoxaparin had no effects on hepatic endothelial dysfunction, except for correcting the hepatic arterial dysfunction when enoxaparin was started with the CCl4 exposure. In these rats, however, enoxaparin increased liver fibrosis and the absolute values of portal venous and sinusoidal resistance. CONCLUSIONS: Our results do not support a role of enoxaparin for improving liver fibrosis, portal hypertension or endothelial dysfunction in active disease at advanced stages of cirrhosis. These disease-related factors and the possibility of a limited therapeutic window should be considered in future studies evaluating the use of anticoagulants in cirrhosis.
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Anticoagulantes/farmacologia , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Enoxaparina/farmacologia , Hipertensão Portal/prevenção & controle , Cirrose Hepática Experimental/prevenção & controle , Fígado/efeitos dos fármacos , Pressão na Veia Porta/efeitos dos fármacos , Animais , Anticoagulantes/toxicidade , Translocação Bacteriana/efeitos dos fármacos , Biomarcadores/sangue , Coagulação Sanguínea/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/sangue , Doença Hepática Induzida por Substâncias e Drogas/patologia , Doença Hepática Induzida por Substâncias e Drogas/fisiopatologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Enoxaparina/toxicidade , Hipertensão Portal/sangue , Hipertensão Portal/patologia , Hipertensão Portal/fisiopatologia , Mediadores da Inflamação/sangue , Fígado/metabolismo , Fígado/patologia , Circulação Hepática/efeitos dos fármacos , Cirrose Hepática Experimental/sangue , Cirrose Hepática Experimental/patologia , Cirrose Hepática Experimental/fisiopatologia , Masculino , Microcirculação/efeitos dos fármacos , Ratos Sprague-DawleyRESUMO
This investigation evaluated, in broiler chickens Pectoralis and Gastrocnemius muscles, the effect of the dietary supplementation with sodium selenite (0.3 ppm) versus selenomethionine (0.3 ppm), on the fatty acids composition, lipids indices, and enzymes indexes for desaturase, elongase, and thioesterase. The selenium reduced, in both muscles, the content of atherogenic fatty acids, C14:0 and C16:0, while it increased the C18:1 level. On the other hand, selenium increased, in both muscles, the content of C18:3n3 and EPA, but not DPA and DHA. No selenium effect was detected for PUFA/SFA, n-6, n-3, n-6/n-3, and atherogenic and thrombogenic indices. As for the enzyme indexes, a selenium effect is only detected for thioesterase. Taken together, the results highlight the potential effect of dietary selenium, mainly selenomethionine, in the modulation of the composition of fatty acids in chicken meat, in particular, reducing the content of atherogenic fatty acids and increasing the health promoting n-3 PUFA.
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Citoesqueleto de Actina/metabolismo , Microtúbulos/metabolismo , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/metabolismo , Proteínas Nogo/fisiologia , Acetilação , Citoesqueleto de Actina/ultraestrutura , Actinas/análise , Animais , Aorta , Movimento Celular , Tamanho Celular , Células Cultivadas , Quimiotaxia , Citoesqueleto/metabolismo , Citoesqueleto/ultraestrutura , Retículo Endoplasmático/química , Adesões Focais , Desacetilase 6 de Histona , Histona Desacetilases/metabolismo , Masculino , Miócitos de Músculo Liso/ultraestrutura , Proteínas Nogo/antagonistas & inibidores , Proteínas Nogo/genética , Processamento de Proteína Pós-Traducional , Interferência de RNA , Ratos , Ratos Sprague-Dawley , Tubulina (Proteína)/metabolismo , Proteínas rho de Ligação ao GTP/metabolismoRESUMO
BACKGROUND & AIMS: Different mechanisms including portal hypertension and hypersplenism have been involved in the development of thrombocytopenia in cirrhosis. However, the relative contribution of each one is unknown. The aim was to evaluate simultaneously different mechanisms that determine platelet count in cirrhosis. METHODS: Cross-sectional study including cirrhotics (n = 120) with hepatic venous pressure gradient (HVPG) measurement. Samples were obtained from peripheral (P) veins to evaluate thrombopoietin (TPO), stem cell factor, hepatocyte growth factor (HGF), tumour necrosis factor, interleukin-(IL6) and (IL11) and from hepatic (H) veins to evaluate TPO. A subgroup (n = 72) had spleen volume estimation. H and P-TPO were also measured in non-cirrhotic patients (n = 15). RESULTS: Patients (Child A: 55, B: 43, C: 22) had a median platelet count of 81 000/mm(3) (IQR 60 500, 110 750), which correlated with spleen volume (r = -0.38, P < 0.001). Platelets were associated also to HVPG (r = -0.47, P = 0.004) and P-TPO (r = 0.31, P = 0.050) only in compensated patients. H-TPO decreased, and the proportion of patients with P-TPO > H-TPO increased, with the presence and the severity of liver disease. H-TPO was correlated with liver function (bilirubin r = -0.350, P < 0.001 and international normalized ratio r = -0.227, P = 0.011). Patients with H-TPO < P-TPO had higher levels of IL-11 and HGF. CONCLUSION: Platelet count in cirrhosis is associated mainly to spleen volume, although portal hypertension as estimated by HVPG and liver function plays a significant role in compensated patients. H-TPO and the proportion of patients with P-TPO > H-TPO were associated to the presence and severity of liver disease.
Assuntos
Hiperesplenismo/sangue , Hipertensão Portal/sangue , Cirrose Hepática , Baço/patologia , Trombocitopenia , Estudos Transversais , Feminino , Humanos , Hiperesplenismo/diagnóstico , Hiperesplenismo/etiologia , Hipertensão Portal/diagnóstico , Hipertensão Portal/etiologia , Coeficiente Internacional Normatizado , Fígado/patologia , Cirrose Hepática/sangue , Cirrose Hepática/complicações , Cirrose Hepática/diagnóstico , Testes de Função Hepática/métodos , Masculino , Pessoa de Meia-Idade , Gravidade do Paciente , Contagem de Plaquetas/métodos , Índice de Gravidade de Doença , Estatística como Assunto , Trombocitopenia/sangue , Trombocitopenia/diagnóstico , Trombocitopenia/etiologiaRESUMO
Inflammatory bowel disease (IBD) is characterized by an impaired intestinal barrier function. We aimed to investigate the role of reticulon-4B (RTN-4B/NOGO-B), a structural protein of the endoplasmic reticulum, in intestinal barrier function and IBD. We used immunohistochemistry, confocal microscopy, real-time PCR, and Western blotting to study tissue distribution and expression levels of RTN-4B/NOGO-B in control and IBD samples from mouse and humans. We also targeted RTN-4B/NOGO-B using siRNAs in cultured human intestinal epithelial cell (IECs). Epithelial barrier permeability was assessed by transepithelial electrical resistance (TEER) measurement. RTN-4B/NOGO-B is expressed in the intestine mainly by IECs. Confocal microscopy revealed a colocalization of RTN-4B, E-cadherin, and polymerized actin fibers in tissue and cultured IECs. RTN-4B mRNA and protein expression were lower in the colon of IL-10(-/-) compared with wild-type mice. Colocalization of RTN-4B/E-cadherin/actin was reduced in the colon of IL-10(-/-) mice. Analysis of endoscopic biopsies from IBD patients showed a significant reduction of RTN-4B/NOGO-B expression in inflamed mucosa compared with control. Treatment of IECs with H2O2 reduced TEER values and triggered phosphorylation of RTN-4B in serine 107 residues as well as downregulation of RTN-4B expression. Acute RTN-4B/NOGO-B knockdown by siRNAs resulted in a decreased TEER values and reduction of E-cadherin and α-catenin expression and in the amount of F-actin-rich filaments in IECs. Epithelial RTN-4B/NOGO-B was downregulated in human and experimental IBD. RTN-4B participates in the intestinal epithelial barrier function, most likely via its involvement in E-cadherin, α-catenin expression, and actin cytoskeleton organization at sites of cell-to-cell contacts.
Assuntos
Doenças Inflamatórias Intestinais/metabolismo , Proteínas da Mielina/metabolismo , Junções Íntimas/metabolismo , Actinas/metabolismo , Animais , Caderinas/metabolismo , Células Cultivadas , Células Epiteliais/metabolismo , Humanos , Peróxido de Hidrogênio/farmacologia , Doenças Inflamatórias Intestinais/patologia , Interleucina-10/deficiência , Interleucina-10/metabolismo , Mucosa Intestinal/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas da Mielina/genética , Proteínas Nogo , Reação em Cadeia da Polimerase em Tempo Real/métodosRESUMO
AIM: To evaluate the expression of serum fibrosis markers in liver transplantation (LT) recipients on everolimus monotherapy compared to patients on an anti-calcineurin regimen. METHODS: This cross-sectional case-control study included LT patients on everolimus monotherapy (cases) (E) (n = 30) and matched controls on an anti-calcineurin regimen (calcineurin inhibitors, CNI), paired by etiology of liver disease and time since LT (n = 30). Clinical characteristics, blood tests and elastography were collected. Serum levels of transforming growth factor-ß (TGF-ß), angiopoietin-1, tumor necrosis factor (TNF), platelet derived growth factor, amino-terminal propeptide of type III procollagen (PIIINP), hyaluronic acid (HA), VCM-1 (ng/mL), interleukin (IL)-10, interferon-inducible protein 10 (IP-10), vascular endothelial growth factor and hepatocyte growth factor (HGF) (pg/mL) were determined by enzyme-linked immunosorbent assay. Expression of these markers between E and CNI was compared. Stratified analysis was done according to factors that may influence liver fibrosis. Variables are described with medians (interquartillic range) or percentages. RESULTS: A total of 60 patients [age: 59 (49-64), hepatitis C virus (HCV): n = 21 (35%), time from LT: 73 mo (16-105)] were included. Patients had been on everolimus for a median of 15 mo. No differences in inflammatory activity, APRI test or liver elastography were found between the groups. No significant differences were observed between the groups in serum levels of PIIINP, metalloproteinase type = 1, angiopoietin, HGF, IP-10, TNF-α, IL-10 and vascular cell adhesion molecule. Patients on E had a lower expression of TGF-ß [E: 12.7 (3.7-133.6), CNI: 152.5 (14.4-333.2), P = 0.009] and HA [E: 702.89 (329.4-838.2), CNI: 1513.6 (691.9-1951.4), P = 0.001] than those on CNI. This difference was maintained in the stratified analysis when recipient age is more than 50 years (TFG-ß1: P = 0.06; HA: P = 0.005), in patients without active neoplasia (TFG-ß1, P = 0.009; HA: P = 0.01), according to time since LT (> than 5 years, TFG-ß1: P = 0.001; HA: P = 0.002), related to previous history of biliary complications (HA: P = 0.01) and HCV recurrence (HA: P = 0.004). Liver transplant recipients with everolimus monotherapy had less serum expression of TGF-ß y HA than matched patients with anti-calcineurins. This difference remains when classifying patients according to donor age and time since LT. Due to the small sample size, when examining patients with a prior history of biliary complications or recurrent HCV, the difference was non-significant but trends towards the lower expression of TFG-ß1 in the everolimus group. Mammalian target of rapamycin (mTOR) plays a role in the transformation of quiescent hepatocellular stellate cell to their active profibrotic state, and experimental models have demonstrated the potential activity of mTOR inhibition in attenuating fibrogenesis. CONCLUSION: This study supports a possible role of everolimus in liver fibrosis modulation after LT in a clinical setting and suggests that tailoring immunosuppression could avoid fibrosis progression in the allograft.
RESUMO
BACKGROUND & AIMS: The modulation of gut flora constitutes a therapeutic tool in patients with liver disease, but some of its modalities require further investigation. Here, we evaluated the effects of probiotics on the hepatic and systemic haemodynamic alterations of advanced liver disease. METHODS: Seventeen patients with cirrhosis and ascites were prospectively included, five of whom abandoned this study prematurely. Hepatic and systemic haemodynamic evaluations were performed at baseline and after 6 weeks of receiving an oral VSL#3 probiotic preparation. Peripheral blood analyses included the evaluation of cytokines (TNF-alpha, IL-1beta, IL-6), bacterial translocation [bacterial DNA and lipopolysaccharide-binding protein (LBP)] and nitric oxide end-products (NOx). RESULTS: In 12 patients completing this study, the oral administration of VSL#3 resulted in reductions of the hepatic venous pressure gradient (HVPG, P < 0.001), cardiac index and heart rate (both P < 0.01) and in increases of the systemic vascular resistance (P < 0.05) and mean arterial pressure (P = 0.06). HVPG decreased at least 10% from baseline in eight patients (67%). Serum sodium increased in most patients (P < 0.01). All these changes were unrelated to the detection of bacterial DNA or to the levels of LBP, pro-inflammatory cytokines or NOx. No significant adverse effects were observed. CONCLUSION: Administration of the probiotic mixture VSL#3 improved the hepatic and systemic haemodynamics and serum sodium levels in patients with cirrhosis. Our results identify major effects of probiotics in liver disease and provide the rationale for assessing their therapeutic potential against the progression of portal hypertension and its complications in future clinical trials.