Effect of Lipopolysaccharide on the Progression of Non-Alcoholic Fatty Liver Disease in High Caloric Diet-Fed Mice.

The incidence of non-alcoholic steatohepatitis (NASH) is increasing. Because gut microbiota have been highlighted as one of the key factors in the pathogenesis of metabolic syndrome, we investigated the involvement of the bacterial component in the progression of non-alcoholic fatty liver (NAFL) to NASH. C57BL/6 mice were fed with maintenance food (MF, groups A and B) or a high caloric diet (HCD, groups C and D) for 1 month. Mice were then divided into four groups: Groups A and C were inoculated with PBS, while groups B and D were inoculated with lipopolysaccharide (LPS) plus complete Freund's adjuvant (CFA). The inoculations were performed a total of 3 times over 3 months. At 6 months, while hepatic steatosis was observed in groups C and D, cellular infiltration and fibrosis were less evident in group C than in group D. Inflammatory cytokines were upregulated in groups B and D. 16S rRNA pyrosequencing of whole colon homogenates containing faeces showed that certain bacterial groups, such as Bacteroidaceae, Peptostreptococcaceae and Erysipelotrichaceae, were increased in groups C and D. Although loading of bacterial components (LPS) resulted in hepatic inflammation in both MF- and HCD-fed mice, HCD feeding was more crucial in the progression of NAFL during the triggering phase.

Differing effects of liraglutide on gastric emptying in Japanese patients with type 2 diabetes.

This study was performed to clarify the influence of liraglutide on gastric emptying in Japanese patients with type 2 diabetes. In 16 patients, the [(13) C]-acetate breath test was performed to compare gastric emptying before and after liraglutide treatment. We found two patterns of response, with gastric emptying being delayed by liraglutide in seven patients (delayers) and not delayed in nine patients (non-delayers). The mean increase of the maximum gastric emptying time was 31 ± 4 min (p < 0.01 vs. baseline) in the delayers, while it was only 2 ± 3 min (p = 0.60 vs. baseline) in the non-delayers. The delayers showed a greater early decrease of AUC-PG from 0 to 60 min, despite no increase of the plasma insulin level compared with non-delayers. In conclusion, the effect of liraglutide treatment on gastric emptying shows heterogeneity, and patients can be classified as delayers or non-delayers.

A new flexible regression model with application to recovery probability Covid-19 patients.

The aim of this study is to propose a generalized odd log-logistic Maxwell mixture model to analyze the effect of gender and age groups on lifetimes and on the recovery probabilities of Chinese individuals with COVID-19. We add new properties of the generalized Maxwell model. The coefficients of the regression and the recovered fraction are estimated by maximum likelihood and Bayesian methods. Further, some simulation studies are done to compare the regressions for different scenarios. Model-checking techniques based on the quantile residuals are addressed. The estimated survival functions for the patients are reported by age range and sex. The simulation study showed that mean squared errors decay toward zero and the average estimates converge to the true parameters when sample size increases. According to the fitted model, there is a significant difference only in the age group on the lifetime of individuals with COVID-19. Women have higher probability of recovering than men and individuals aged ≥60 years have lower recovered probabilities than those who aged <60 years. The findings suggest that the proposed model could be a good alternative to analyze censored lifetime of individuals with COVID-19.

The re-parameterized inverse Gaussian regression to model length of stay of COVID-19 patients in the public health care system of Piracicaba, Brazil.

Among the models applied to analyze survival data, a standout is the inverse Gaussian distribution, which belongs to the class of models to analyze positive asymmetric data. However, the variance of this distribution depends on two parameters, which prevents establishing a functional relation with a linear predictor when the assumption of constant variance does not hold. In this context, the aim of this paper is to re-parameterize the inverse Gaussian distribution to enable establishing an association between a linear predictor and the variance. We propose deviance residuals to verify the model assumptions. Some simulations indicate that the distribution of these residuals approaches the standard normal distribution and the mean squared errors of the estimators are small for large samples. Further, we fit the new model to hospitalization times of COVID-19 patients in Piracicaba (Brazil) which indicates that men spend more time hospitalized than women, and this pattern is more pronounced for individuals older than 60 years. The re-parameterized inverse Gaussian model proved to be a good alternative to analyze censored data with non-constant variance.

A new regression model for rates and proportions data with applications.

We propose a new continuous distribution in the interval ( 0 , 1 ) based on the generalized odd log-logistic-G family, whose density function can be symmetrical, asymmetric, unimodal and bimodal. The new model is implemented using the gamlss packages in R. We propose an extended regression based on this distribution which includes as sub-models some important regressions. We employ a frequentist and Bayesian analysis to estimate the parameters and adopt the non-parametric and parametric bootstrap methods to obtain better efficiency of the estimators. Some simulations are conducted to verify the empirical distribution of the maximum likelihood estimators. We compare the empirical distribution of the quantile residuals with the standard normal distribution. The extended regression can give more realistic fits than other regressions in the analysis of proportional data.

The multinomial logistic regression model for predicting the discharge status after liver transplantation: estimation and diagnostics analysis.

The multinomial logistic regression model (MLRM) can be interpreted as a natural extension of the binomial model with logit link function to situations where the response variable can have three or more possible outcomes. In addition, when the categories of the response variable are nominal, the MLRM can be expressed in terms of two or more logistic models and analyzed in both frequentist and Bayesian approaches. However, few discussions about post modeling in categorical data models are found in the literature, and they mainly use Bayesian inference. The objective of this work is to present classic and Bayesian diagnostic measures for categorical data models. These measures are applied to a dataset (status) of patients undergoing kidney transplantation.

Olanzapine potentiates neuronal survival and neural stem cell differentiation: regulation of endoplasmic reticulum stress response proteins.

Recent clinical neuroimaging studies have suggested that morphological brain changes occur and progress in the course of schizophrenia. Although the neurogenetic and neurotrophic effects of antipsychotics are considered to contribute to the prevention of reduction in brain volume, the cellular molecular mechanisms of action of antipsychotics have not yet been elucidated. We examined the effects of antipsychotics on the endoplasmic reticulum (ER) stress-induced damages of neurons and neural stem cells (NSCs) using cultured cells. In the neuronal cultures, the atypical antipsychotic olanzapine protected neurons from thapsigargin (1 microM)-induced injury. It was observed that a low concentration of thapsigargin (10 nM) that did not affect the neuronal survival could reduce neuronal differentiation of cultured NSCs, suggesting a role of ER stress in the differentiation function of NSCs. Treatment with olanzapine increased the neuronal differentiation suppressed by the exposure to thapsigargin (10 nM). The thapsigargin-induced ER chaperones, GRP78, which indicate the ER stress condition of the cell, were decreased by the treatment with the atypical antipsychotics olanzapine and quetiapine but not by the typical antipsychotic haloperidol. These results indicate that the amelioration of ER-stress might be involved in the cellular mechanisms of atypical antipsychotics to produce neuroprotective and neurogenetic actions in neurons and NSCs, suggesting potential roles of these drugs for treatment of schizophrenia.

Neural stem cell transplantation in a model of fetal alcohol effects.

Neural stem cell (NSC) transplantation has been investigated and developed in areas such as brain injury, stroke and neurodegenerative diseases. Recently, emerging evidence suggest that many of clinical symptoms observed in psychiatric disease are likely related to neural network disruptions including neurogenesis dysfunction. In the present study, we transplanted NSCs into a model of fetal alchol effects (FAE) for the purpose of investigating the possibility of regenerative therapy for the FAE. We labeled NSCs with fluorescent dye and radioisotope which were transplanted into FAE rats by intravenous injection. The transplanted cells were detected in wide areas of brain and were greater in number in the brains of the FAE group compared to the control group. Furthermore NSC transplantation attenuated behavioral abnormalities in FAE animals. These results suggest NSC transplantation as a potental new therapy for human FAE.

Quantitative reduction of type I adenylyl cyclase in human alcoholics.

The amounts of adenylyl cyclase type I (AC I) were examined in various parts of the postmortem brains from alcoholics who prior to death had been abstinent from alcohol for at least 6 months and compared with controls using immunoblot analysis with anti-AC I specific antibody. It was revealed that a significant reduction of AC I was observed in both frontal and temporal cortices. On the other hand, in other areas (occipital cortex, caudate nucleus, putamen, and hippocampus) the amounts were comparable between alcoholics and controls. In the next step, we examined two subtypes of human AC mRNA levels (AC I and AC VIII) in blood cells by quantitative RT-PCR using [alpha-32P]dCTP with two sets of the synthetic oligonucleotide primers based on the DNA sequences reported elsewhere (Villacres, E.C. et al., Genomics 16 (1993) 473-478; J. Parma et al., Biochem. Biophys. Res. Commun. 179 (1991) 455-462). The amounts of amplified DNAs of both AC I and AC VIII were significantly smaller in alcoholics than in controls. On the other hand, the amounts of amplified DNA of beta-actin DNA were almost equal between alcoholics and controls. It appears from these results that a reduction in the amount of AC subtypes may be a biological marker for alcoholics.

Phosphorylation of liver gap junction protein by protein kinase C.

The 27 kDa protein, a major component of rat liver gap junctions, was shown to be phosphorylated in vitro by protein kinase C. The stoichiometry of the phosphorylation indicated that approx. 0.33 mol phosphate was incorporated per mol 27 kDa protein. Phosphorylation was entirely dependent on the presence of calcium and was virtually specific for serine residues. For comparison, the gap junction protein was also examined for its phosphorylation by cAMP-dependent protein kinase, the extent of phosphorylation being one-tenth that exerted by protein kinase C.

Effect of ionic strength on production of cAMP- and Ca2+-independent protein kinase from rat liver plasma membrane.

Production of cAMP- and Ca2+-independent protein kinase was stimulated when rat liver plasma membrane was incubated with increasing concentrations of NaCl. This protein kinase release was diminished by addition of protease inhibitor. The molecular mass of this enzyme was approx. 50 kDa and a high concentration of Mg2+ was required for whole histone phosphorylation. These properties are similar to those of the protease-activated form of protein kinase C. The NaCl effect could be replaced by other salts such as LiCl and NaHCO3. These results suggest that membrane-bound protein kinase C is activated by limited proteolysis corresponding to an increase in ionic strength.

Identification of the phosphorylation sites of H2B histone by a catalytic fragment of p72syk from porcine spleen.

Phosphorylated sites of calf thymus H2B histone were investigated with a catalytic fragment of 72 kDa protein-tyrosine kinase (p72syk). Three of five tyrosine residues in H2B histone can be phosphorylated by this kinase. In this analysis, H2B histone was thoroughly phosphorylated in vitro with [gamma-32P]ATP and the kinase, and then digested with a lysylendopeptidase. The resulting radioactive phosphopeptides were separated by a reverse-phase column on high performance liquid chromatography. Subsequent sequential Edman degradation of the purified phosphopeptides revealed that 40Y, 83Y and 121Y were phosphorylated. 121Y is the major phosphorylated residue in H2B histone. No phosphorylation was detected in 37Y and 42Y. Although the consensus sequence was not defined from these analyses, our data suggest that higher-order structure(s) in addition to primary one may participate in recognition of H2B histone by this protein kinase.

Calcium--calmodulin-dependent activation of porcine liver phosphorylase kinase.

Porcine liver phosphorylase kinase was activated about 1.5-fold by calmodulin in a calcium-dependent manner. Half-maximal stimulation was observed at about 80 nM calmodulin and the activation was almost pH-independent. The specific binding of porcine liver phosphorylase kinase to calmodulin--Sepharose affinity column exhibited an absolute dependence upon the presence of calcium. The physiological role of the calmodulin-dependent activation for liver phosphorylase kinase is discussed.

Possible role of Na+ influx in phorbol ester-induced down-regulation of protein kinase C in HL60 cells.

Amiloride, an inhibitor of Na+/H+ exchange, inhibited down-regulation of protein kinase C in HL60 cells induced by tumor-promoting phorbol ester in dose-dependent manner judging from immunoblot analysis. This inhibition was observed with regard to type I (gamma), type II (beta), and type III (alpha) isozymes of protein kinase C. On the other hand, monensin, a Na+ ionophore, accelerated the down-regulation of protein kinase C induced by phorbol ester. When we examined 22Na+ uptake by HL60 cells, the higher uptake was observed after stimulation with phorbol ester compared to the control cells and this 22Na+ uptake was strongly inhibited by the addition of amiloride. However, monensin further stimulated the 22Na+ uptake observed in phorbol ester-treated cells. These data suggest that the increase in intracellular Na+ concentration may be one of the triggers for the induction of down regulation of protein kinase C.

Biosynthetic processing and quaternary interactions of proprotein convertase SPC4 (PACE4).

SPC4 (PACE4), a member of the eukaryotic family of subtilisin-like proprotein convertases, is synthesized as a proenzyme (proSPC4) which undergoes proteolytic removal of N-terminal propeptide during transit through the secretory pathway. As this propeptide processing seems to be a key event in the functional expression of SPC4, we have investigated its mechanism and the intracellular site where it occurs. In transfected fibroblast cells, the 110-kDa proSPC4 undergoes slow cleavage to generate a 103-kDa mature enzyme in the endoplasmic reticulum (ER). Site-directed mutagenesis studies demonstrate that the proteolytic activation of SPC4 occurs mainly through a unimolecular autocatalytic process and propeptide cleavage is a prerequisite for its export from the ER. Sedimentation velocity and chemical cross-linking analysis demonstrate that the precursor protein in the cells exists as both a monomer and a dimer-sized complex whereas mature SPC4 exists only as a monomer. These results suggest that the cleavage of the N-terminal propeptide of SPC4 plays a regulatory role in its activation and secretion through the change in its oligomeric state.

(-)-Epigallocatechin gallate inhibits mos activation-mediated xenopus oocyte maturation induced by progesterone.

(-)-Epigallocatechin gallate (EGCG), one of the constituents of green tea known to have a tumor preventing effect, inhibited maturation of Xenopus laevis oocytes induced by progesterone when this polyphenol was microinjected into oocytes at a final concentration of about 1 mM. Western blot and activity measurement analyses showed that Mos translation and the subsequent activations of mitogen-activated protein kinase and p90(rsk), probably by protein phosphorylation, seemed to have been inhibited by the microinjection of EGCG. These results suggest that EGCG may have the ability to control Xenopus oocyte maturation at least during the stage of Mos activation.

Hypoxic induction of adrenomedullin in cultured human umbilical vein endothelial cells.

OBJECTIVES: The current study evaluated the hypoxic induction of adrenomedullin gene expression and secretion, and its mechanism in cultured human umbilical vein endothelial cells (HUVEC). METHODS: HUVEC were exposed to hypoxia or normoxia as controls for 1 to 24 h. Using Northern blot analysis and a radioimmunoassay, we evaluated adrenomedullin expression in HUVEC. The transcriptional component of adrenomedullin gene regulation was assessed by nuclear run-off experiments, and adrenomedullin mRNA half-life was measured by actinomycin D experiments. RESULTS: We found that hypoxic conditions (1-3% oxygen) significantly increased adrenomedullin mRNA and protein in HUVEC. This increase was inversely proportional to oxygen tension and was reversible upon re-exposure to a 21% oxygen environment Nuclear run-off experiments revealed the enhanced transcriptional rate of adrenomedullin gene. Next, actinomycin D experiments revealed the enhanced adrenomedullin mRNA stability. CONCLUSIONS: These results indicate that hypoxia increases adrenomedullin gene expression and secretion in HUVEC by transcriptional and post-transcriptional mechanisms. Hypoxic induction of adrenomedullin may play a pathophysiological role in the vascular systems.

Classification of hepatic venous outflow obstruction: ambiguous terminology of the Budd-Chiari syndrome.

Severe hepatic venous outflow obstruction and its manifestations often are recorded under the label "Budd-Chiari syndrome." Unfortunately, this label is ambiguous; it does not clearly identify the site of the lesion (hepatic veins versus inferior vena cava), its morphologic features (thrombotic versus nonthrombotic), or its cause. In the literature, implied or expressed definitions vary. Use of a standardized topographic and pathogenetic classification of hepatic venous outflow obstruction would enable investigators to group patients with comparable conditions, as required for therapeutic trials, prognostic evaluations, and studies of pathogenetic pathways. Review of our own cases revealed that hepatic venous outflow obstruction involving large hepatic veins is usually thrombotic and that isolated obstruction of the inferior vena cava or of small hepatic veins is usually nonthrombotic. Application of such a classification seems feasible and may yield useful results.

Immunohistochemical characterization of hepatic lymphocytes in primary biliary cirrhosis in comparison with primary sclerosing cholangitis and autoimmune chronic active hepatitis.

We analyzed the immunophenotypes of hepatic cellular infiltrates by quantitative immunohistochemical methods in biopsy specimens from 20 patients with primary biliary cirrhosis (PBC), 19 with primary sclerosing cholangitis, and 11 with autoimmune chronic active hepatitis. Specifically, we sought to identify activated T cells, interferon-gamma-producing cells, and natural killer cells. The portal cellular infiltrate in PBC contained a preponderance of CD4 cells in comparison with CD8 cells, with a CD4/CD8 ratio of 2.45:1. The cellular infiltrate in areas of piecemeal necrosis contained mostly CD8 cells. Infiltrating CD8 cells in PBC had the surface phenotype of cytotoxic (CD8-positive, CD11b-negative) cells. Approximately 4% of T cells expressed interleukin 2 receptors. Interferon-gamma-staining cells were rarely identified (in less than 2%). Cells that expressed the natural killer cell markers CD16, CD56, or CD57 were infrequent, constituting approximately 5% of the cellular infiltrate. The composition of the infiltrates was similar in patients with PBC and chronic active hepatitis. Natural killer cells were twice as common in patients with primary sclerosing cholangitis (P < 0.05) as in those with PBC. The inflammatory infiltrates in areas of piecemeal necrosis were similar in the three diseases and differed from those found within the portal area, in that CD8 cells were preponderant. In all three liver diseases, almost 90% of bile ducts expressed class II HLA antigens. These findings support the hypothesis that cytotoxic T cells of either the CD4 or CD8 immunophenotype but not natural killer cells may be involved in the pathogenesis of PBC and chronic active hepatitis.(ABSTRACT TRUNCATED AT 250 WORDS)

Hepatoportal arteriovenous fistula: morphologic features studied after orthotopic liver transplantation.

A 51-year-old woman underwent orthotopic liver transplantation because of a small-duct primary sclerosing cholangitis associated with chronic ulcerative colitis and a hepatoportal arteriovenous fistula. Arteriograms before liver transplantation and specimen arteriograms revealed a convolution of arteries in the right hepatic lobe which communicated with a massively dilated and partly thrombosed portal vein branch. The lesion was probably the result of a previous liver biopsy. Microscopic examination of the fistula and of specimens taken at a distance from the fistula showed prominent intimal fibroplasia of portal vein branches. The vascular changes were so severe that hepatic artery and portal vein branches closely resembled each other on routine sections. We are unaware of such a finding in other conditions and therefore believe that recognition of the described vascular abnormalities in liver biopsy specimens should lead pathologists to comment that an arterioportal fistula might be present.