Role of liquid fructose/sucrose in regulating the hepatic transcriptome in a high-fat Western diet model of NAFLD.

Nonalcoholic fatty liver disease (NAFLD), which ranges from simple steatosis to nonalcoholic steatohepatitis (NASH), is the most common chronic liver disease. Yet, the molecular mechanisms for the progression of steatosis to NASH remain largely undiscovered. Thus, there is a need for identifying specific gene and pathway changes that drive the progression of NAFLD. This study uses high-fat Western diet (HFWD) together with liquid sugar [fructose and sucrose (F/S)] feeding for 12 weeks in mice to induce obesity and examine hepatic transcriptomic changes that occur in NAFLD progression. The combination of a HFWD+F/S in the drinking water exacerbated HFWD-induced obesity, hyperinsulinemia, hyperglycemia, hepatic steatosis, inflammation, and human and murine fibrosis gene set enrichment that is consistent with progression to NASH. RNAseq analysis revealed differentially expressed genes (DEGs) associated with HFWD and HFWD+F/S dietary treatments compared to Chow-fed mice. However, liquid sugar consumption resulted in a unique set of hepatic DEGs in HFWD+F/S-fed mice, which were enriched in the complement and coagulation cascades using network and biological analysis. Cluster analysis identified Orosomucoid (ORM) as a HFWD+F/S upregulated complement and coagulation cascades gene that was also upregulated in hepatocytes treated with TNFα or free fatty acids in combination with hypoxia. ORM expression was found to correlate with NAFLD parameters in obese mice. Taken together, this study examined key genes, biological processes, and pathway changes in the liver of HFWD+F/S mice in an effort to provide insight into the molecular basis for which the addition of liquid sugar promotes the progression of NAFLD.

Development and Characterization of a Novel Congenic Rat Strain for Obesity and Cancer Research.

The association between a Western Diet and colon cancer suggests that dietary factors and/or obesity may contribute to cancer progression. Our objective was to develop a new animal model of obesity and the associated pathophysiology to investigate human cancer independent of dietary components that induce obesity. A novel congenic rat strain was established by introducing the fa allele from the Zucker rat into the Rowett Nude rat to generate a "fatty nude rat". The obese phenotype was first characterized in the new model. To then examine the utility of this model, lean and obese rats were implanted with HT-29 human colon cancer xenografts and tumor growth monitored. Fatty nude rats were visibly obese and did not develop fasting hyperglycemia. Compared to lean rats, fatty nude rats developed fasting hyperinsulinemia, glucose intolerance, and insulin resistance. Colon cancer tumor growth rate and final weight were increased (P < 0.05) in fatty nude compared to lean rats. Final tumor weight was associated with p38 kinase phosphorylation (P < 0.01) in fatty nude rats. We have established a novel model of obesity and pre-type 2 diabetes that can be used to investigate human cancer and therapeutics in the context of obesity and its associated pathophysiology.

Restricted feeding for 9h in the active period partially abrogates the detrimental metabolic effects of a Western diet with liquid sugar consumption in mice.

BACKGROUND: Obesity is a major public health concern that can result from diets high in fat and sugar, including sugar sweetened beverages. A proposed treatment for dietary-induced obesity is time-restricted feeding (TRF), which restricts consumption of food to specific times of the 24-hour cycle. Although TRF shows great promise to prevent obesity and the development of chronic disease, the effects of TRF to reverse metabolic changes and the development of NAFLD in animal models of a Western diet with sugary water consumption is not known. OBJECTIVE: The objective of the current study was to evaluate the role of TRF in the treatment of obesity and NAFLD through examination of changes in metabolic and histopathologic parameters. METHODS: To better understand the role of TRF in the treatment of obesity and NAFLD, we investigated the metabolic phenotype and NAFLD parameters in a mouse model of NAFLD in which obesity and liver steatosis are induced by a Western Diet (WD): a high-fat diet of lard, milkfat and Crisco with sugary drinking water. Mice were subjected to a short-term (4-weeks) and long-term (10-weeks) TRF in which food was restricted to 9h at night. RESULTS: Prior to TRF treatment, the WD mice had increased body mass, and exhibited less activity, and higher average daytime energy expenditure (EE) than chow fed mice. Approximately 4- and 10-weeks following TFR treatment, WD-TRF had moderate but not statistically significant weight loss compared to WD-ad libitum (WD-AL) mice. There was a modest but significant reduction in the inguinal adipose tissue weight in both WD-TRF groups compared to the WD-AL groups; however, there was no difference in epididymal and retroperitoneal adipose tissue mass or adipocyte size distribution. In contrast, the diet-induced increase in normalized liver tissue weight, hepatic triglyceride, and NAFLD score was partially abrogated in the 4-week WD-TRF mice, while systemic insulin resistance was partially abrogated and glucose intolerance was completely abrogated in the 10-week WD-TRF mice. Importantly, WD-induced metabolic dysfunction (substrate utilization, energy expenditure, and activity) was partially abrogated by 4- and 10-week TRF. CONCLUSIONS: Our results support the hypothesis that TRF aids in reducing the detrimental metabolic effects of consuming a WD with sugary drinking water but does not ameliorate obesity.

High fat diet induces brain insulin resistance and cognitive impairment in mice.

High fat diet-induced obesity is associated with insulin resistance (IR) and other chronic, diet related illnesses, including dementia. Alzheimer disease is the most common form of dementia, and is characterized by the presence of amyloid plaques and neurofibrillary tangles in brain. This study was designed to determine whether diet-induced changes in peripheral insulin sensitivity could contribute to alterations in brain insulin signaling and cognitive functions. Six week old, male C57BL/6NHsd mice were randomly assigned a high fat diet (40% energy from fat) with 42g/L liquid sugar (HFS) added to the drinking water or a normal chow diet (12% energy from fat) for 14weeks. Metabolic phenotypes were characterized for energy expenditure, physical activity, and food intake, and glucose and insulin tolerance tests. In addition, we examined the changes in protein expression related to brain insulin signaling and cognitive function. Mice fed HFS exhibited a statistically significant increase in obesity, and lower glucose and insulin tolerance as compared to animals fed the normal chow diet. In brain, HFS elicited IR as evidenced by a significant decrease in tyrosine phosphorylation of insulin receptor and an increase serine phosphorylation of IRS-1. These changes were accompanied by inflammatory (NFκB, JNK) and stress responses (p38 MAPK, CHOP) in whole brain lysate. In addition, HFS mouse brain exhibited biochemical changes related to increased amyloid beta deposition and neurofibrillary tangle formation, and decreased synaptic plasticity. These results suggested changes in insulin sensitivity might contribute to cognitive impairment associated with the HFS diet in mice.

High-fat Western diet-induced obesity contributes to increased tumor growth in mouse models of human colon cancer.

Strong epidemiologic evidence links colon cancer to obesity. The increasing worldwide incidence of colon cancer has been linked to the spread of the Western lifestyle, and in particular consumption of a high-fat Western diet. In this study, our objectives were to establish mouse models to examine the effects of high-fat Western diet-induced obesity on the growth of human colon cancer tumor xenografts, and to examine potential mechanisms driving obesity-linked human colon cancer tumor growth. We hypothesize that mice rendered insulin resistant due to consumption of a high-fat Western diet will show increased and accelerated tumor growth. Homozygous Rag1tm1Mom mice were fed either a low-fat Western diet or a high-fat Western diet (HFWD), then human colon cancer xenografts were implanted subcutaneously or orthotopically. Tumors were analyzed to detect changes in receptor tyrosine kinase-mediated signaling and expression of inflammatory-associated genes in epididymal white adipose tissue. In both models, mice fed an HFWD weighed more and had increased intra-abdominal fat, and tumor weight was greater compared with in the low-fat Western diet-fed mice. They also displayed significantly higher levels of leptin; however, there was a negative correlation between leptin levels and tumor size. In the orthotopic model, tumors and adipose tissue from the HFWD group displayed significant increases in both c-Jun N-terminal kinase activation and monocyte chemoattractant protein 1 expression, respectively. In conclusion, this study suggests that human colon cancer growth is accelerated in animals that are obese and insulin resistant due to the consumption of an HFWD.

Metabolic phenotype and adipose and liver features in a high-fat Western diet-induced mouse model of obesity-linked NAFLD.

nonalcoholic fatty liver disease (NAFLD), an obesity and insulin resistance associated clinical condition - ranges from simple steatosis to nonalcoholic steatohepatitis. To model the human condition, a high-fat Western diet that includes liquid sugar consumption has been used in mice. Even though liver pathophysiology has been well characterized in the model, little is known about the metabolic phenotype (e.g., energy expenditure, activity, or food intake). Furthermore, whether the consumption of liquid sugar exacerbates the development of glucose intolerance, insulin resistance, and adipose tissue dysfunction in the model is currently in question. In our study, a high-fat Western diet (HFWD) with liquid sugar [fructose and sucrose (F/S)] induced acute hyperphagia above that observed in HFWD-fed mice, yet without changes in energy expenditure. Liquid sugar (F/S) exacerbated HFWD-induced glucose intolerance and insulin resistance and impaired the storage capacity of epididymal white adipose tissue (eWAT). Hepatic TG, plasma alanine aminotransferase, and normalized liver weight were significantly increased only in HFWD+F/S-fed mice. HFWD+F/S also resulted in increased hepatic fibrosis and elevated collagen 1a2, collagen 3a1, and TGFß gene expression. Furthermore, HWFD+F/S-fed mice developed more profound eWAT inflammation characterized by adipocyte hypertrophy, macrophage infiltration, a dramatic increase in crown-like structures, and upregulated proinflammatory gene expression. An early hypoxia response in the eWAT led to reduced vascularization and increased fibrosis gene expression in the HFWD+F/S-fed mice. Our results demonstrate that sugary water consumption induces acute hyperphagia, limits adipose tissue expansion, and exacerbates glucose intolerance and insulin resistance, which are associated with NAFLD progression.

Resistance of canine lymphoma cells to adenoviral infection due to reduced cell surface RGD binding integrins.

Recombinant adenovirus vectors (Ad) have been recognized as effective in vivo gene delivery vehicles and utilized as gene therapy agents for a number of cancers. The elucidation of viral entry mechanisms has allowed the development of recombinant vectors that exploit existing cell surface receptors to achieve entry into the cell. B lymphocytes are normally resistant to infection by adenovirus 5, likely due to the lack of the Coxsackie and Adenovirus receptor (CAR). Using reverse-transcriptase PCR and flow cytometry, the CD40 receptor has been shown to be expressed on many lymphoma cells. We exploited this finding to develop a gene therapy strategy for treatment of canine B cell lymphoma. Ad5 was targeted to cells expressing CD40 via CD40 ligand (CD40L) and was effective in infecting CD40-expressing control cells; however, both primary canine lymphoma cells and cell lines demonstrated limited evidence of transduction. Following receptor binding, adenovirus entry into cells may require interaction with α(v)ß(3/5) integrins; we demonstrate that canine lymphoma cells are deficient in these integrins. Reduced α(v)ß(3) integrin expression may render these cells incapable of internalizing Ad vectors. Thus, any viral targeting approaches for treatment of canine lymphoma must also take into account the potential lack of internalization signals.

Interferon alpha-induced inhibition of infectious bursal disease virus in chicken embryo fibroblast cultures differing in Mx genotype.

Interferon (IFN)-induced antiviral activity in cells forms an important early line of defense against viral pathogens. IFN-induced mediators are well documented in mammals, with one of the best-characterized antiviral proteins being Mx. In chickens, many alleles of Mx have been described, but functionally only the polymorphism at a site encoding residue 631 in the protein determines differential antiviral activity against vesicular stomatitis virus and influenza virus in transfection experiments. The role of chicken Mx has not been assessed with regard to infectious bursal disease virus (IBDV), an important pathogen of chickens. To examine the role of chicken IFNalpha and the Mx631 single-nucleotide polymorphism against IBDV, chicken embryo fibroblast cultures (CEF) that differed in Mx genotype (antivirally positive Mx Asn631 or antivirally negative Mx Ser631) were treated with IFNalpha and viral yield was assessed following infection with D78 vaccine-strain IBDV. IFNalpha was shown to have strong antiviral activity in this system in terms of reduced virus yield. Furthermore, the reduction in viral yield did not differ significantly among Mx genotypes, indicating that Mx Asn631 is not a pivotal determinant of resistance to this strain of IBDV in CEF.

A genetically engineered adenovirus vector targeted to CD40 mediates transduction of canine dendritic cells and promotes antigen-specific immune responses in vivo.

Targeting viral vectors encoding tumor-associated antigens to dendritic cells (DCs) in vivo is likely to enhance the effectiveness of immunotherapeutic cancer vaccines. We have previously shown that genetic modification of adenovirus (Ad) 5 to incorporate CD40 ligand (CD40L) rather than native fiber allows selective transduction and activation of DCs in vitro. Here, we examine the capacity of this targeted vector to induce immune responses to the tumor antigen CEA in a stringent in vivo canine model. CD40-targeted Ad5 transduced canine DCs via the CD40-CD40L pathway in vitro, and following vaccination of healthy dogs, CD40-targeted Ad5 induced strong anti-CEA cellular and humoral responses. These data validate the canine model for future translational studies and suggest targeting of Ad5 vectors to CD40 for in vivo delivery of tumor antigens to DCs is a feasible approach for successful cancer therapy.

The chicken BF1 (classical MHC class I) gene shows evidence of selection for diversity in expression and in promoter and signal peptide regions.

The chicken MHC B contains two classical class I genes, BF1 and BF2, with the exception of two related haplotypes lacking BF1 due to insertion/rearrangement. In light of functional specialization of BF1 and BF2 molecules, we were interested in evaluating their relative expression at the mRNA level. We evaluated several MHC haplotypes for class I gene expression by RT-quantitative PCR. BF1 transcript levels were approximately two- to fivefold lower than BF2, with the exception of one haplotype in which BF1 expression was very low. To investigate molecular explanations for differences in BF locus or allele expression, we determined nucleotide sequences of Enhancer A and proximal promoter elements of nine different BF1 alleles, as well as their signal peptide sequences. Results showed that all BF1 alleles exhibit conservation of most of the identified promoter elements, but divergence from the Enhancer A sequence identified in the more highly expressed BF2 locus. Nonetheless, extensive BF1 allelic polymorphism was found in the promoter region and in the signal peptide, with two strongly separated allelic lineages identified for both. Patterns of promoter lineages, signal peptide lineages, and exon 2 mature protein coding sequences in individual BF1 alleles suggest that recombination among these elements has contributed to diversification of BF1 alleles. Finally, identification of a novel inactivating mutation in one BF1 allele suggests past selective pressure to eliminate BF1 function.

In a time of change.

A combination of restructuring and cutbacks threatens nursing posts in Northern Ireland.