Genevieve M. Forster

Liposome-siRNA-Peptide Complexes Cross the Blood-Brain Barrier and Significantly Decrease PrPC on Neuronal Cells and PrPRES in Infected Cell Cultures

Background Recent advances toward an effective therapy for prion diseases employ RNA interference to suppress PrPC expression and subsequent prion neuropathology, exploiting the phenomenon that disease severity and progression correlate with host PrPC expression levels. However, delivery of lentivirus encoding PrP shRNA has demonstrated only modest efficacy in vivo. Methodology/Principal Findings Here we describe a new siRNA delivery system incorporating a small peptide that binds siRNA and acetylcholine receptors (AchRs), acting as a molecular messenger for delivery to neurons, and cationic liposomes that protect siRNA-peptide complexes from serum degradation. Conclusions/Significance Liposome-siRNA-peptide complexes (LSPCs) delivered PrP siRNA specifically to AchR-expressing cells, suppressed PrPC expression and eliminated PrPRES formation in vitro. LSPCs injected intravenously into mice resisted serum degradation and delivered PrP siRNA throughout the brain to AchR and PrPC-expressing neurons. These data promote LSPCs as effective vehicles for delivery of PrP and other siRNAs specifically to neurons to treat prion and other neuropathological diseases.

Rice varietal differences in bioactive bran components for inhibition of colorectal cancer cell growth

Rice bran chemical profiles differ across rice varieties and have not yet been analysed for differential chemopreventive bioactivity. A diverse panel of seven rice bran varieties was analysed for growth inhibition of human colorectal cancer (CRC) cells. Inhibition varied from 0% to 99%, depending on the variety of bran used. Across varieties, total lipid content ranged 5-16%, individual fatty acids had 1.4- to 1.9-fold differences, vitamin E isoforms (α-, γ-, δ-tocotrienols, and tocopherols) showed 1.3- to 15.2-fold differences, and differences in γ-oryzanol and total phenolics ranged between 100-275ng/mg and 57-146ngGAE/mg, respectively. Spearman correlation analysis was used to identify bioactive compounds implicated in CRC cell growth inhibitory activity. Total phenolics and γ-tocotrienol were positively correlated with reduced CRC cell growth (p<0.05). Stoichiometric variation in rice bran components and differential effects on CRC viability merit further evaluation elucidate their role in dietary CRC chemoprevention.

Dietary rice bran promotes resistance to Salmonella enterica serovar Typhimurium colonization in mice

BackgroundDietary rice bran consists of many bioactive components with disease fighting properties; including the capacity to modulate the gut microbiota. Studies point to the important roles of the gut microbiota and the mucosal epithelium in the establishment of protection against enteric pathogens, such as Salmonella. The ability of rice bran to reduce the susceptibility of mice to a Salmonella infection has not been previously investigated. Therefore, we hypothesized that the incorporation of rice bran into the diet would inhibit the colonization of Salmonella in mice through the induction of protective mucosal responses.ResultsMice were fed diets containing 0%, 10% and 20% rice bran for one week prior to being orally infected with Salmonella enterica serovar Typhimurium. We found that mice consuming the 10 and 20% rice bran diets exhibited a reduction in Salmonella fecal shedding for up to nine days post-infection as compared to control diet fed animals (p < 0.05). In addition, we observed decreased concentrations of the pro-inflammatory cytokines, TNF-alpha, IFN-gamma, and IL-12 (p < 0.05) as well as increased colonization of native Lactobacillus spp. in rice bran fed mice (p < 0.05). Furthermore, in vitro experiments revealed the ability of rice bran extracts to reduce Salmonella entry into mouse small intestinal epithelial cells.ConclusionsIncreasing rice bran consumption represents a novel dietary means for reducing susceptibility to enteric infection with Salmonella and potentially via induction of native Lactobacillus spp.

Effects of dietary cooked navy bean on the fecal microbiome of healthy companion dogs.

Background Cooked bean powders are a promising novel protein and fiber source for dogs, which have demonstrated potential to alter microbial composition and function for chronic disease control and prevention. This study aimed to determine the impact of cooked navy bean powder fed as a staple food ingredient on the fecal microbiome of healthy adult pet dogs. Methodology/Principal Findings Fecal samples from healthy dogs prior to dietary control and after 4 wk of dietary treatment with macro- and micronutrient matched diets containing either 0 or 25% cooked navy beans (n = 11 and n = 10, respectively) were analyzed by 454-pyrosequencing of the 16S rRNA gene. There were few differences between dogs fed the control and navy bean diets after 4 wk of treatment. These data indicate that there were no major effects of navy bean inclusion on microbial populations. However, significant differences due to dietary intervention onto both research diets were observed (i.e., microbial populations at baseline versus 4 wk of intervention with 0 or 25% navy bean diets). After 4 wk of dietary intervention on either control or navy bean diet, the Phylum Firmicutes was increased and the Phyla Actinobacteria and Fusobacteria were decreased compared to baseline. Conclusions No negative alterations of microbial populations occurred following cooked navy bean intake in dogs, indicating that bean powders may be a viable protein and fiber source for commercial pet foods. The highly variable microbial populations observed in these healthy adult pet dogs at baseline is one potential reason for the difficulty to detect alterations in microbial populations following dietary changes. Given the potential physiological benefits of bean intake in humans and dogs, further evaluation of the impacts of cooked navy bean intake on fecal microbial populations with higher power or more sensitive methods are warranted.

Effects of cooked navy bean powder on apparent total tract nutrient digestibility and safety in healthy adult dogs

Dry beans (Phaseolus vulgaris L., Fabaceae) are a low glycemic index food containing protein, fiber, minerals, essential vitamins, and bioactive compounds and have not been evaluated for inclusion in commercial canine diets. The objective of this study was to establish the apparent total tract digestibility and safety of cooked navy bean powder when incorporated into a canine diet formulation at 25% (wt/wt) compared with a macro- and micro-nutrient matched control. Twenty-one healthy, free-living, male and female adult dogs of different breeds were used in a randomized, blinded, placebo controlled, 28-d dietary intervention study. Apparent total tract energy and nutrient digestibility of the navy bean powder diet were compared with the control diet. Digestibilities and ME content were 68.58 and 68.89% DM, 78.22 and 79.49% CP, 77.57 and 74.91% OM, 94.49 and 93.85% acid hydrolyzed fat, and 3,313 and 3,195 kcal ME/kg for the navy bean diet and control diet, respectively. No differences were observed between the groups. No increased flatulence or major change in fecal consistency was observed. Navy bean powder at 25% (wt/wt) of total diet was determined to be palatable (on the basis of intake and observation) and digestible in a variety of dog breeds. No changes were detected in clinical laboratory values, including complete blood counts, blood biochemical profiles, and urinalysis in either the bean or control diet groups. These results indicate that cooked navy bean powder can be safely included as a major food ingredient in canine diet formulations and provide a novel quality protein source, and its use warrants further investigation as a functional food for chronic disease control and prevention.

Nutritional Weight Loss Therapy with Cooked Bean Powders Regulates Serum Lipids and Biochemical Analytes in Overweight and Obese Dogs

Background: Emerging evidence supports that dry bean (Phaseolus vulgaris L.,) intake promotes weight loss and regulates blood lipids. Overweight and obese dogs represent a novel translational model for diet controlled evaluation of dry beans and for assessing the effects of bean intake on serum biomarkers of weight loss. Our objective was to evaluate changes in serum biomarkers associated with weight loss after four weeks of cooked navy bean or black bean powder intake (25% weight/weight) compared to an isocaloric, macro and micronutrient matched control diet in overweight/obese dogs. Methods: Thirty client-owned, adult dogs of diverse breeds were randomized to 1 of 3 dietary study groups at the Colorado State University Veterinary Teaching Hospital. Body weights were measured weekly and a blood serum chemistry panel was performed at baseline and 2 and 4 weeks post intervention. Results: Average percent weight lost after 4 weeks for dogs consuming the control diet was 4.20% (± 0.88), 5.22% (± 0.91) for dogs consuming the black bean diet, and 6.52% (± 0.95) for dogs consuming the navy bean diet. Serum cholesterol decreased by an average of 17 mg/ml (P<0.02) in the control group, 40 mg/dl (P<0.001) in black bean group, and 54 mg/dl (P<0.001) in the navy bean group. Triglycerides, high-density lipoprotein, and low- density lipoprotein were also changed in bean groups compared to control. Furthermore, serum blood urea nitrogen was decreased in the navy bean group, creatinine was increased in both bean groups, alkaline phosphatase was decreased in the black bean group, and total protein, aspartate aminotransferase, and total bilirubin were decreased in the control group at 4 weeks compared to baseline. Conclusion: Overweight and obese canines represent an advanced translational model and dietary bean intake regulates lipid metabolism in overweight and obese dogs.

Multiresidue Analysis of Pesticides in Urine of Healthy Adult Companion Dogs

The objective of this study was to determine the background exposures to pesticides as detected in urine from 21 healthy companion dogs in Northern Colorado. A panel of 301 pesticides was used to screen urine samples collected from dogs using an established ultraperformance liquid chromatography-mass spectrometry (UPLC-MS/MS) platform. Canine food intakes were controlled for one month on diets that were also screened for pesticide contents. Fifteen distinct pesticides were detected in urine. The most frequently detected compounds in canine urine samples collected over a 1 month period were atrazine, fuberidazole, imidacloprid, terbumeton, and clopyralid. Fuberidazole was the only pesticide detected in both the diets and urine. Companion dogs develop many similar chronic diseases as humans and represent a relevant model for biomonitoring combinations of environmental pesticide exposures, as well as for evaluating the potential relationships between environmental exposures and disease risk.

A Comparative Study of Serum Biochemistry, Metabolome and Microbiome Parameters of Clinically Healthy, Normal Weight, Overweight, and Obese Companion Dogs

ABSTRACT The aim of this study was to compare fecal microbiome, plasma, fecal and urine metabolomes, and serum biochemistry of adult companion dogs according to body condition scores. Blood, serum/plasma, urine, and fecal samples were collected from 66 clinically healthy, adult companion dogs of either normal weight (NW), overweight (OW), or obese dogs (OB). analyses included fecal microbiome analyses via 16S ribosomal RNA gene amplicon; sequencing, nontargeted plasma, fecal, and urine metabolomics using liquid chromatography/gas chromatography‐mass; spectrometry, and serum biochemistry for each dog. Few significant differences in serum biochemistry and fecal microbiome Operational Taxonomic Unit (OTU) were found between weight groups and there was high OTU variation between individual dogs. NW dogs had higher relative abundance of the genus Eubacterium (log‐fold change 4.3, adjusted P value=.003) and lower relative abundance of the family Bifidobacteriaceae (log‐fold change −3.6, adjusted P value=.02) compared to OB dogs. The microbiome of NW dogs had higher OTU richness compared with OB dogs. Metabolome analysis showed 185 plasma, 37 fecal, and 45 urine metabolites that significantly differed between NW and OW or OB dogs. There were notable significant differences in relative abundance of several plasma phospholipid moieties and fecal volatile fatty acids between weight phenotypes. The combinations of host and gut microbiota and metabolic shifts suggest a pattern that could help detection of early metabolic changes in overweight dogs before the development of obesity related disease. The results of this study support the need for continued investigation into sensitive measures of metabolic aberrancies in overweight dogs.

Fecal Microbial Communities of Overweight and Obese Client-Owned Dogs Fed Cooked Bean Powders as Assessed by 454-Pyrosequencing

Dry beans are consumed around the world and contain a multitude of health benefits, some of which may be related to the gut microbiome. The objective of this study was to evaluate the effects of feeding a 25% cooked Navy Bean (NB) or Black Bean (BB) powder on the fecal microbiota of overweight and obese companion dogs undergoing calorically restricted weight loss, compared to dogs fed an iso-nutrient control diet using 454 pyrosequencing. A double-blinded, placebo-controlled, three-arm clinical trial was conducted. Thirty client-owned, clinically healthy, overweight or obese, adult, male and female dogs of diverse breeds were randomized to one of the three isocaloric, nutritionally complete weight loss diets containing either 0% bean powder (placebo-control); 25% cooked BB powder; or 25% cooked NB powder and calorically restricted to achieve weight loss of up to 2% body weight/wk. for 4 wks. Fresh fecal samples were collected from each dog immediately after completing the 4-wk diet intervention and weight loss phase. Fecal genomic DNA was extracted and used to create 16S rRNA gene amplicons, which were subjected to 454-pyrosequencing. Predominant bacterial phyla present in all dogs included Firmicutes, Fusobacteria, Actinobacteria, Proteobacteria, and Bacteroidetes. Predominant fecal bacterial genera included Clostridium, Blautia, Fusobacterium, and undefined Lachnospiraceae. Fecal undefined Ruminococcus were greater (P<0.05) in dogs fed BB compared to dogs fed the control diet. Client-owned dogs with various dietary and environmental exposures, ages, and breeds were evaluated for fecal microbiota changes during short-term weight loss on different diets. A high variability in fecal microbiota was observed in this free living dog population, leading to few differences among treatments. The gut microbiota may be an area for investigation during long-term weight loss in companion animals, but such studies require a higher level of dietary control and larger sample sizes.