Plasma concentrations of nitric oxide and asymmetric dimethylarginine in human alcoholic cirrhosis

BACKGROUND/AIMS: The liver plays a prominent role in the metabolism of asymmetric dimethyl-l-arginine (ADMA), an endogenous inhibitor of nitric oxide (NO) synthase. This study was designed to determine whether plasma levels of ADMA and NO production are altered in patients with compensated and decompensated alcoholic cirrhosis. METHODS: Plasma levels of l-arginine, ADMA, symmetric dimethylarginine (SDMA) and NO (nitrite plus nitrate, NOx) were measured in nine patients with compensated alcoholic cirrhosis (Child-Pugh A) and 11 patients with advanced cirrhosis (Child-Pugh B-C). Seven healthy volunteers served as controls. RESULTS: ADMA and NOx concentrations in decompensated cirrhosis were higher than in the compensated group and control group (ADMA: 1.12+/-0.08 vs. 0.58+/-0.05 and 0.58+/-0.07micromol/l, respectively; P<0.05; NOx 97.90+/-10.27 vs. 37.42+/-3.91 and 40.43+/-5.30micromol/l, respectively; P<0.05). There was a positive correlation between the clinical score of the patients and concentrations of ADMA (r(2)=0.547, P<0.01) and NOx (r(2)=0.689, P<0.01). SDMA and l-arginine levels were not significantly different between the three groups. CONCLUSIONS: The results suggest that hepatocellular damage is a main determinant of elevated ADMA concentration in advanced alcoholic cirrhosis. By inhibiting NO release from vascular endothelium, ADMA might oppose the peripheral vasodilation caused by excessive NO production in severe cirrhosis.     

Podocytes are new cellular targets of haemoglobin‐mediated renal damage

Recurrent and massive intravascular haemolysis induces proteinuria, glomerulosclerosis, and progressive impairment of renal function, suggesting podocyte injury. However, the effects of haemoglobin (Hb) on podocytes remain unexplored. Our results show that cultured human podocytes or podocytes isolated from murine glomeruli bound and endocytosed Hb through the megalin–cubilin receptor system, thus resulting in increased intracellular Hb catabolism, oxidative stress, activation of the intrinsic apoptosis pathway, and altered podocyte morphology, with decreased expression of the slit diaphragm proteins nephrin and synaptopodin. Hb uptake activated nuclear factor erythroid‐2‐related factor 2 (Nrf2) and induced expression of the Nrf2‐related antioxidant proteins haem oxygenase‐1 (HO‐1) and ferritin. Nrf2 activation and Hb staining was observed in podocytes of mice with intravascular haemolysis. These mice developed proteinuria and showed podocyte injury, characterized by foot process effacement, decreased synaptopodin and nephrin expression, and podocyte apoptosis. These pathological effects were enhanced in Nrf2‐deficient mice, whereas Nrf2 activation with sulphoraphane protected podocytes against Hb toxicity both in vivo and in vitro. Supporting the translational significance of our findings, we observed podocyte damage and podocytes stained for Hb, HO‐1, ferritin and phosphorylated Nrf2 in renal sections and urinary sediments of patients with massive intravascular haemolysis, such as atypical haemolytic uraemic syndrome and paroxysmal nocturnal haemoglobinuria. In conclusion, podocytes take up Hb both in vitro and during intravascular haemolysis, promoting oxidative stress, podocyte dysfunction, and apoptosis. Nrf2 may be a potential therapeutic target to prevent loss of renal function in patients with intravascular haemolysis. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Electrocardiographic changes and conduction disturbances after transfemoral aortic valve implantation with Edwards Sapien 3 prosthesis

Objectives

The aim of this study is to describe electrocardiographic changes and conduction abnormalities in patients undergoing transcatheter aortic valve implantation (TAVI).

Immunophenotypic analysis of myelodysplastic syndromes

BACKGROUND AND OBJECTIVES: In contrast with hematologic malignancies in which the value of immunophenotypic studies is well established, information on the immunophenotypic characteristics of myelodysplastic syndromes (MDS) is scanty. The main goal of the present study was to explore the immunophenotypic differences between patients with MDS and normal individuals, including changes in distribution of cell lineages as well as phenotypic aberrations and blockades in cell maturation pathways. DESIGN AND METHODS: In MDS the proportion of bone marrow CD34+ cells was higher than in normal patients but the most immature progenitors (CD34+CD38-) were less represented. By contrast the proportion of myelomonocytic CD34+ cells was greater than in normal individuals, translating into an increased myeloid/non-myeloid CD34+ hematopoietic progenitor cell ratio. RESULTS: This suggests that in MDS, the majority of CD34+ cells are already committed to the myeloid lineage. Upon analyzing the granulo-monocytic differentiation pathway, MDS patients showed an increased proportion of monocytic cells with a decreased percentage of cells of neutrophil lineage, leading to a lower neutrophil/monocytic cell ratio. Maturational arrests in the monocytic but not in the neutrophil differentiation pathway were observed. In refractory anemia with excess blasts in transformation (RAEB-t) such blockades mainly occurred during the earliest stages of differentiation but in the other MDS subtypes they occurred in later stages. INTERPRETATION AND CONCLUSIONS: Phenotypic aberrations occurred in 90% of patients and a high proportion of cases showed >=2 aberrations. In summary, our results show that, in addition to an abnormal distribution of the bone marrow cell compartment, MDS patients frequently show aberrant phenotypes and maturational arrests. Some of these features may help in cases in which the diagnosis of MDS is questionable.     

Molecular analysis of HPRT1+ somatic cell hybrids derived from a carrier of an HPRT1 mutation responsible for Lesch‐Nyhan syndrome

Heterozygous carriers of HPRT1 mutations responsible for Lesch‐Nyhan syndrome can be detected by analysis of somatic cell hybrids derived from peripheral blood lymphocytes and Hprt1‐negative cells of rodent origin followed by selection in culture medium containing hypoxanthine, aminopterine, and thymidine (HAT). The parental origin of the X chromosome containing the normal HPRT1 allele in HPRT1⁺ hybrid cell lines can be determined by molecular haplotyping attributable to highly polymorphic X‐linked markers. We used this procedure to study a presumed carrier whose paternal active X chromosome always segregated in the cell hybrids derived from her. Conversely, her maternal X chromosome was systematically absent in most cell hybrids, or when present, it was inactive and coexisted with an active, paternal X chromosome. These results clearly demonstrated that the proband was a heterozygous carrier of a mutation responsible for HPRT1 deficiency. © 2001 Wiley‐Liss, Inc.     

Recomendaciones del Grupo Español de Trabajo en Enfermedad de Crohn y Colitis Ulcerosa (GETECCU) sobre el cribado y tratamiento de la infección tuberculosa en pacientes con enfermedad inflamatoria intestinal

There is evidence that following the recommendations on screening and treatment of tuberculosis infection does not completely prevent the onset of tuberculosis in patients with inflammatory bowel disease. This fact, and the increasing use of new biologics and immunomodulators, has led the Spanish Group Working on Crohn's Disease and Ulcerative Colitis to update their recommendations for the prevention of tuberculosis in patients with inflammatory bowel disease. Diagnostic methods for latent tuberculosis infection, different scenarios in which screening is to be performed, strategies to reduce the risk of tuberculosis once biological treatment is initiated and chemoprophylaxis guidelines for latent tuberculosis infection are reviewed, as well as the management of active tuberculosis during biological treatment. Finally, there is a summary of the current recommendations within the paper and in an algorithm.     

Hepatitis B virus genotypes and lamivudine resistance mutations in HIV/hepatitis B virus-coinfected patients

BACKGROUND: Differences in subtypes, hepatitis B early antigen (HBeAg)-negative variants, and drug resistance mutations all seem to influence the clinical and therapeutic outcome in patients with chronic hepatitis B virus (HBV) infection. Information available on the prevalence and distribution of distinct HBV variants in HIV-positive patients is scarce. METHODS: All HIV-infected patients with persistent serum hepatitis B surface antigen and detectable HBV viremia were identified in a reference HIV clinic located in Madrid, Spain. HBV load, subtypes, precore (PC) and basal core promoter (BCP) variants, and lamivudine (LAM) resistance mutations were analyzed. RESULTS: A total of 81 HBV/HIV-coinfected patients (4.1%) were identified in a population of 1968 HIV-positive patients. Plasma specimens with detectable HBV viremia could be obtained from 62 subjects, and this was the study population that underwent further virologic characterization. HBV genotype distribution was as follows: A (n = 27), D (n = 27), E (n = 1), F (n = 2), and G (n = 3). Two patients had mixed HBV genotypes (A/E and A/F). HBV subtype A was predominant (74%) among patients infected through sexual contact, whereas HBV-D was most frequent (74%) among intravenous drug users (P < 0.001). PC/BCP mutants were more frequent in patients with HBV-D than in those with HBV-A (63% vs. 18%; P < 0.01). Median time on LAM was 40 months; patients with HBV-A tended to show LAM resistance mutations more often (53% vs. 44%) and to develop them earlier (35 vs. 45 months) than patients with HBV-D. The dual L180M + M204V/I mutant was the predominant resistance pattern, although a triple rt173V + 180M + 204V, which acts as a vaccine escape mutant, was found in 1 individual. In the multivariate analysis, patients with LAM resistance mutations were significantly more frequently HBeAg-positive and older than individuals with wild-type HBV. Hepatitis-delta was recognized in 13 (21%) of these 62 HBV viremic patients, with no association with specific HBV variants. CONCLUSION: Risk transmission group, age, and positive serum HBeAg are the main determinants of distinct HBV virologic variants, including HBV genotypes and LAM-resistant mutants, in HBV/HIV-coinfected patients.     

Subcellular distribution of swine vesicular disease virus proteins and alterations induced in infected cells: a comparative study with foot-and-mouth disease virus and vesicular stomatitis virus

The intracellular distribution of swine vesicular disease virus (SVDV) proteins and the induced reorganization of endomembranes in IBRS-2 cells were analyzed. Fluorescence to new SVDV capsids appeared first upon infection, concentrated in perinuclear circular structures and colocalized to dsRNA. As in foot-and-mouth disease virus (FMDV)-infected cells, a vesicular pattern was predominantly found in later stages of SVDV capsid morphogenesis that colocalized with those of non-structural proteins 2C, 2BC and 3A. These results suggest that assembly of capsid proteins is associated to the replication complex. Confocal microscopy showed a decreased fluorescence to ER markers (calreticulin and protein disulfide isomerase), and disorganization of cis-Golgi gp74 and trans-Golgi caveolin-1 markers in SVDV- and FMDV-, but not in vesicular stomatitis virus (VSV)-infected cells. Electron microscopy of SVDV-infected cells at an early stage of infection revealed fragmented ER cisternae with expanded lumen and accumulation of large Golgi vesicles, suggesting alterations of vesicle traffic through Golgi compartments. At this early stage, FMDV induced different patterns of ER fragmentation and Golgi alterations. At later stages of SVDV cytopathology, cells showed a completely vacuolated cytoplasm containing vesicles of different sizes. Cell treatment with brefeldin A, which disrupts the Golgi complex, reduced SVDV (approximately 5 log) and VSV (approximately 4 log) titers, but did not affect FMDV growth. Thus, three viruses, which share target tissues and clinical signs in natural hosts, induce different intracellular effects in cultured cells.