A semiparametric model for between-subject attributes: Applications to beta-diversity of microbiome data.

The human microbiome plays an important role in our health and identifying factors associated with microbiome composition provides insights into inherent disease mechanisms. By amplifying and sequencing the marker genes in high-throughput sequencing, with highly similar sequences binned together, we obtain operational taxonomic units (OTUs) profiles for each subject. Due to the high-dimensionality and nonnormality features of the OTUs, the measure of diversity is introduced as a summarization at the microbial community level, including the distance-based beta-diversity between individuals. Analyses of such between-subject attributes are not amenable to the predominant within-subject-based statistical paradigm, such as t-tests and linear regression. In this paper, we propose a new approach to model beta-diversity as a response within a regression setting by utilizing the functional response models (FRMs), a class of semiparametric models for between- as well as within-subject attributes. The new approach not only addresses limitations of current methods for beta-diversity with cross-sectional data, but also provides a premise for extending the approach to longitudinal and other clustered data in the future. The proposed approach is illustrated with both real and simulated data.

Indoles: metabolites produced by intestinal bacteria capable of controlling liver disease manifestation.

Alterations in the bacteria that reside in our gastrointestinal tract play a role in the pathogenesis and progression of many disorders including liver and gastrointestinal diseases. Both qualitative (composition) and quantitative (amount) changes in gut microbes are associated with increased susceptibility to liver disease. Importantly, the intestinal microbiota is involved in the regulation of many host signalling pathways via the generation of different metabolites. Hence, dysbiosis influences disease development and progression by directly affecting the host-bacteria metabolic interaction. Microbe-derived harmful metabolites can translocate to distant organs due to increased intestinal permeability as observed during dysbiosis. Contrary, certain bacterial metabolites such as tryptophan metabolites contribute to intestinal and systemic homeostasis. Here, we provide an overview of current evidence describing to what extent microbial metabolites modulate the development of chronic liver diseases such as alcoholic steatohepatitis and nonalcoholic fatty liver disease with a special emphasis on indoles.

Current Concepts, Opportunities, and Challenges of Gut Microbiome-Based Personalized Medicine in Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NALFD) is now a leading cause of chronic liver disease worldwide, in part, as a consequence of rapidly rising levels of obesity and metabolic syndrome and is a major risk factor for cirrhosis, hepatocellular carcinoma, and liver-related mortality. From NAFLD stems a myriad of clinical challenges related to both diagnosis and management. A growing body of evidence suggests an intricate linkage between the gut microbiome and the pathogenesis of NAFLD. We highlight how our current knowledge of the gut-liver axis in NAFLD may be leveraged to develop gut microbiome-based personalized approaches for disease management, including its use as a non-invasive biomarker for diagnosis and staging, as a target for therapeutic modulation, and as a marker of drug response. We will also discuss current limitations of these microbiome-based approaches. Ultimately, a better understanding of microbiota-host interactions in NAFLD will inform the development of novel preventative strategies and precise therapeutic targets.

Results of allergen-specific immunotherapy in 117 dogs with atopic dermatitis.

The success of the treatment of 117 dogs with atopic dermatitis with allergen-specific immunotherapy for up to 48 months was assessed. An excellent response (remission with exclusive immunotherapy) was recorded in 18 of the dogs, a good response (more than 50 per cent reduction in medication and improvement of clinical signs) was recorded in 57, a moderate response was recorded in 24 and a poor response in 18. The mould antigens in the allergen extract were stored in a separate vial before administration and the success rate of the immunotherapy including mould antigens was much higher than in an earlier study in which mould and pollen antigens had been stored in one vial. The success rate was not affected significantly by the age of the dogs when the disease developed, or by their age or the period for which they had shown clinical signs when the treatment began; it was also unaffected by whether pollens, moulds or dust mites were used as antigens, or by whether the offending allergens had been identified by intradermal testing or by serum testing for allergen-specific immunoglobulin E.

Different effects of a CD14 gene polymorphism on disease outcome in patients with alcoholic liver disease and chronic hepatitis C infection.

AIM: Clinical and experimental data suggest that gut-derived endotoxins are an important pathogenic factors for progression of chronic liver disease. Recently, a C-T (-159) polymorphism in the promoter region of the CD14 gene was detected and found to confer increased CD14 expression and to be associated with advanced alcoholic liver damage. Here, we investigated this polymorphism in patients with less advanced alcoholic liver disease (ALD) and chronic hepatitis C virus (HCV) infection. METHODS: CD14 genotyping was performed by PCR-RFLP analysis in (a) 121 HCV patients, (b) 62 patients with alcohol-associated cirrhosis (Alc-Ci), (c) 118 individuals with heavy alcohol abuse without evidence of advanced liver damage (Alc-w/o Ci), and (d) 247 healthy controls. Furthermore, serum levels of soluble CD14 (sCD14) and transaminases were determined. RESULTS: The TT genotype was significantly more frequent in Alc-Ci compared to Alc-w/o Ci or controls (40.3% vs 23.7% or 24.0%, respectively). In Alc-w/o Ci, serum levels of transaminases did not differ significantly between patients with different CD14 genotypes. In HCV patients, TT-homozygotes had significantly higher sCD14 levels and sCD14 serum levels were significantly higher in patients with advanced fibrosis or cirrhosis. However, no association was found between CD14 genotypes and histological staging or grading. CONCLUSION: Considering serum transaminases as surrogate markers for alcoholic liver damage, the CD14 polymorphism seems to exhibit different effects during the course of ALD. Differences in genotype distribution between cirrhotic HCV patients and alcoholics and the known functional impact of this polymorphism on CD14 expression levels further indicate differences in the pathophysiological role of CD14 and CD14-mediated lipopolysaccharides signal transduction with regard to the stage as well as the type of the underlying liver disease.

Oral selamectin in the treatment of canine generalised demodicosis.

The success rates and adverse effects of a selamectin spot-on preparation, given orally weekly or every other week, against canine generalised demodicosis were evaluated in 44 dogs, 39 with juvenile-onset demodicosis and five with adult-onset demodicosis. The dogs received selamectin at a dose of 24 to 48 mg/kg once a week, or the same dose every two weeks. Thirty-eight dogs completed the study, four dogs were lost to follow-up and two dogs died. Nine dogs went into remission: all had the juvenile-onset form of the disease. There was no difference between the two treatment groups, but dogs with clinical scores of 65 or less responded significantly better than those with a score of over 65 (P=0.0015). The most frequent adverse effects were vomiting and diarrhoea. Two dogs exhibited mild reversible neurological side effects, which resolved with cessation of treatment. Difficulties in oral administration were observed in several dogs.

CD40 activates NF-kappa B and c-Jun N-terminal kinase and enhances chemokine secretion on activated human hepatic stellate cells.

Activated hepatic stellate cells (HSCs) are the main producers of extracellular matrix in the fibrotic liver and contribute to hepatic inflammation through the secretion of chemokines and the recruitment of leukocytes. This study assesses the function of CD40 on human HSCS: Activated human HSCs express CD40 in culture and in fibrotic liver, as determined by flow cytometry, RT-PCR, and immunohistochemistry. CD40 expression is strongly enhanced by IFN-gamma. Stimulation of CD40 with CD40 ligand (CD40L)-transfected baby hamster kidney cells induces NF-kappaB, as demonstrated by the activation of I-kappaB kinase (IKK), increased NF-kappaB DNA binding, and p65 nuclear translocation. CD40-activated IKK also phosphorylates a GST-p65 substrate at serine 536 in the transactivation domain 1. Concomitant with the activation of IKK, CD40L-transfected baby hamster kidney cell treatment strongly activates c-Jun N-terminal kinase. CD40 activation increases the secretion of IL-8 and monocyte chemoattractant protein-1 by HSCs 10- and 2-fold, respectively. Adenovirally delivered dominant negative (dn) IKK2 and TNFR-associated factor 2dn inhibit IKK-mediated GST-I-kappaB and GST-p65 phosphorylation, NF-kappaB binding, and IL-8 secretion, whereas IKK1dn and NF-kappaB-inducing kinase dominant negative do not have inhibitory effects. We conclude that the CD40-CD40L receptor-ligand pair is involved in a cross-talk between HSCs and immune effector cells that contributes to the perpetuation of HSC activation in liver fibrosis through TNFR-associated factor 2- and IKK2-dependent pathways.

The role of Smad3 in mediating mouse hepatic stellate cell activation.

Transforming growth factor beta (TGF-beta) is the most potent profibrogenic mediator in liver fibrosis. Although Smad proteins have been identified as intracellular mediators in the TGF-beta signaling pathway, the function of individual Smad proteins remains poorly understood. The aim of this study was to explore the contribution of Smad3 in mediating TGF-beta responses in a model of acute liver injury in vivo and in culture-activated hepatic stellate cells (HSCs). Wild-type, Smad3 heterozygous or Smad3 homozygous knockout mice were treated with a single intragastric administration of CCl(4). After 72 hours, the induction of hepatic collagen alpha1(I) and alpha2(I) messenger RNA (mRNA) levels in Smad3 knockout mice was only 42% and 64%, respectively, of the levels induced in wild-type mice. However, smooth muscle alpha-actin (alpha-SMA) was expressed at a slightly higher level in livers from knockout mice compared with wild-type mice. In culture-activated HSCs from Smad3 knockout mice, collagen alpha1(I) mRNA was 73% of wild-type HSCs, but alpha-SMA expression was the same. HSCs from knockout mice showed a higher proliferation rate than wild-type HSCs. Smad3-deficient HSCs did not form TGF-beta1-induced Smad-containing DNA-binding complexes. In conclusion, (1) maximal expression of collagen type I in activated HSCs requires Smad3 in vivo and in culture; (2) Smad3 is not necessary for HSC activation as assessed by alpha-SMA expression; (3) Smad3 is necessary for inhibition of proliferation of HSCs, which might be TGF-beta-dependent; and (4) Smad3 is required for TGF-beta1-mediated Smad-containing DNA-binding complex formation in cultured HSCs.

TAK1/JNK and p38 have opposite effects on rat hepatic stellate cells.

After liver injury, hepatic stellate cells (HSCs) undergo a process of activation with expression of smooth muscle alpha-actin (alpha-SMA), an increased proliferation rate, and a dramatic increase in synthesis of type I collagen. The intracellular signaling mechanisms of activation and perpetuation of the activated phenotype in HSCs are largely unknown. In this study the role of the stress-activated protein kinases, c-Jun N-terminal kinase (JNK) and p38, were evaluated in primary cultures of rat HSCs. The effect of JNK was assessed by using an adenovirus expressing a dominant negative form of transforming growth factor beta (TGF-beta)-activated kinase 1 (TAK1) (Ad5dnTAK1) and a new selective pharmacologic inhibitor SP600125. The effect of p38 was assessed with the selective pharmacologic inhibitor SB203580. These kinases were inhibited starting either in quiescent HSCs (culture day 1) or in activated HSCs (culture day 5). Although blocking TAK1/JNK and p38 decreased the expression of alpha-SMA protein in early stages of HSC activation, no effect was observed when TAK1/JNK or p38 were inhibited in activated HSCs. JNK inhibition increased and p38 inhibition decreased collagen alpha1(I) mRNA level as measured by RNase protection assays, with maximal effects observed in early stages of HSC activation. Furthermore, TAK1/JNK inhibition decreased HSC proliferation, whereas p38 inhibition led to an increased proliferation rate of HSCs, independently of its activation status. These results show novel roles for the TAK1/JNK pathway and p38 during HSC activation in culture. Despite similar activators of TAK1/JNK and p38, their functions in HSCs are distinct and opposed.