ASU Scholarship Showcase

Although insulin resistance in skeletal muscle is well-characterized, the role of circulating whole blood in the metabolic syndrome phenotype is not well understood. We set out to test the hypothesis that genes involved in inflammation, insulin signaling and mitochondrial function would be altered in expression in the whole blood of individuals with metabolic syndrome. We further wanted to examine whether similar relationships that we have found previously in skeletal muscle exist in peripheral whole blood cells. All subjects (n=184) were Latino descent from the Arizona Insulin Resistance registry. Subjects were classified based on the metabolic syndrome phenotype according to the National Cholesterol Education Program’s Adult Treatment Panel III. Of the 184 Latino subjects in the study, 74 were classified with the metabolic syndrome and 110 were without. Whole blood gene expression profiling was performed using the Agilent 4x44K Whole Human Genome Microarray. Whole blood microarray analysis identified 1,432 probes that were altered in expression ≥1.2 fold and P<0.05 after Benjamini-Hochberg in the metabolic syndrome subjects. KEGG pathway analysis revealed significant enrichment for pathways including ribosome, oxidative phosphorylation and MAPK signaling (all Benjamini-Hochberg P<0.05). Whole blood mRNA expression changes observed in the microarray data were confirmed by quantitative RT-PCR. Transcription factor binding motif enrichment analysis revealed E2F1, ELK1, NF-kappaB, STAT1 and STAT3 significantly enriched after Bonferroni correction (all P<0.05). The results of the present study demonstrate that whole blood is a useful tissue for studying the metabolic syndrome and its underlying insulin resistance although the relationship between blood and skeletal muscle differs.

In spite of well-documented health benefits of vegetarian diets, less is known regarding the effects of these diets on athletic performance. In this cross-sectional study, we compared elite vegetarian and omnivore adult endurance athletes for maximal oxygen uptake (VO2 max) and strength. Twenty-seven vegetarian (VEG) and 43 omnivore (OMN) athletes were evaluated using VO2 max testing on the treadmill, and strength assessment using a dynamometer to determine peak torque for leg extensions. Dietary data were assessed using detailed seven-day food logs. Although total protein intake was lower among vegetarians in comparison to omnivores, protein intake as a function of body mass did not differ by group (1.2 ± 0.3 and 1.4 ± 0.5 g/kg body mass for VEG and OMN respectively, p = 0.220). VO2 max differed for females by diet group (53.0 ± 6.9 and 47.1 ± 8.6 mL/kg/min for VEG and OMN respectively, p < 0.05) but not for males (62.6 ± 15.4 and 55.7 ± 8.4 mL/kg/min respectively). Peak torque did not differ significantly between diet groups. Results from this study indicate that vegetarian endurance athletes’ cardiorespiratory fitness was greater than that for their omnivorous counterparts, but that peak torque did not differ between diet groups. These data suggest that vegetarian diets do not compromise performance outcomes and may facilitate aerobic capacity in athletes.

In vitro models that mimic in vivo host-pathogen interactions are needed to evaluate candidate drugs that inhibit bacterial virulence traits. We established a new approach to study Pseudomonas aeruginosa biofilm susceptibility on biotic surfaces, using a three-dimensional (3-D) lung epithelial cell model. P. aeruginosa formed antibiotic resistant biofilms on 3-D cells without affecting cell viability. The biofilm-inhibitory activity of antibiotics and/or the anti-biofilm peptide DJK-5 were evaluated on 3-D cells compared to a plastic surface, in medium with and without fetal bovine serum (FBS). In both media, aminoglycosides were more efficacious in the 3-D cell model. In serum-free medium, most antibiotics (except polymyxins) showed enhanced efficacy when 3-D cells were present. In medium with FBS, colistin was less efficacious in the 3-D cell model. DJK-5 exerted potent inhibition of P. aeruginosa association with both substrates, only in serum-free medium. DJK-5 showed stronger inhibitory activity against P. aeruginosa associated with plastic compared to 3-D cells. The combined addition of tobramycin and DJK-5 exhibited more potent ability to inhibit P. aeruginosa association with both substrates. In conclusion, lung epithelial cells influence the efficacy of most antimicrobials against P. aeruginosa biofilm formation, which in turn depends on the presence or absence of FBS.

Background: Peanut consumption favorably influences satiety. This study examined the acute effect of peanut versus grain bar preloads on postmeal satiety and glycemia in healthy adults and the long-term effect of these meal preloads on body mass in healthy overweight adults.

Methods: In the acute crossover trial (n = 15; 28.4 ± 2.9 y; 23.1 ± 0.9 kg/m²), the preload (isoenergetic peanut or grain bar with water, or water alone) was followed after 60 min with ingestion of a standardized glycemic test meal. Satiety and blood glucose were assessed immediately prior to the preload and to the test meal, and for two hours postmeal at 30-min intervals. In the parallel-arm, randomized trial (n = 44; 40.5 ± 1.6 y, 31.8 ± 0.9 kg/m²), the peanut or grain bar preload was consumed one hour prior to the evening meal for eight weeks. Body mass was measured at 2-week intervals, and secondary endpoints included blood hemoglobin A1c and energy intake as assessed by 3-d diet records collected at pre-trial and trial weeks 1 and 8.

Results: Satiety was elevated in the postprandial period following grain bar ingestion in comparison to peanut or water ingestion (p = 0.001, repeated-measures ANOVA). Blood glucose was elevated one hour after ingestion of the grain bar as compared to the peanut or water treatments; yet, total glycemia did not vary between treatments in the two hour postprandial period. In the 8-week trial, body mass was reduced for the grain bar versus peanut groups after eight weeks (−1.3 ± 0.4 kg versus −0.2 ± 0.3 kg, p = 0.033, analysis of covariance). Energy intake was reduced by 458 kcal/d in the first week of the trial for the grain bar group as compared to the peanut group (p = 0.118). Hemoglobin A1c changed significantly between groups during the trial (−0.25 ± 0.07% and −0.18 ± 0.12% for the grain bar and peanut groups respectively, p = 0.001).

Conclusions: Compared to an isoenergetic peanut preload, consumption of a grain bar preload one hour prior to a standardized meal significantly raised postmeal satiety. Moreover, consumption of the grain bar prior to the evening meal was associated with significant weight loss over time suggesting that glycemic carbohydrate ingestion prior to meals may be a weight management strategy.

Background: Height is an important health assessment measure with many applications. In the medical practice and in research settings, height is typically measured with a stadiometer. Although lasers are commonly used by health professionals for measurement including facial imaging, corneal thickness, and limb length, it has not been utilized for measuring height. The purpose of this feasibility study was to examine the ease and accuracy of a laser device for measuring height in children and adults.

Findings: In immediate succession, participant height was measured in triplicate using a stadiometer followed by the laser device. Measurement error for the laser device was significantly higher than that for the stadiometer (0.35 and 0.20 cm respectively). However, the measurement techniques were highly correlated (r² = 0.998 and 0.990 for the younger [<12 y, n = 25] and older [≥12 y, n = 100] participants respectively), and the estimated reliability between measurement techniques was 0.999 (ICC; 95 % CI: 0.998,1.000) and 0.995 (ICC; 95 % CI: 0.993,0.997) for the younger and older groups respectively. The average differences between the two styles of measurement (e.g., stadiometer minus laser) were significantly different from zero: +0.93 and +0.45 cm for the younger and older groups respectively.

Conclusions: These data demonstrate that laser technology can be adapted to measure height in children and adults. Although refinement is needed, the laser device for measuring height merits further development.

Background: Immunomodulatory drugs (IMiDs), such as lenalidomide, are therapeutically active compounds that bind and modulate the E3 ubiquitin ligase substrate recruiter cereblon, thereby affect steady-state levels of cereblon and cereblon binding partners, such as ikaros and aiolos, and induce many cellular responses, including cytotoxicity to multiple myeloma (MM) cells. Nevertheless, it takes many days for MM cells to die after IMiD induced depletion of ikaros and aiolos and thus we searched for other cereblon binding partners that participate in IMiD cytotoxicity.

Methods: Cereblon binding partners were identified from a MM cell line expressing histidine-tagged cereblon by pulling down cereblon and its binding partners and verified by co-immunoprecipitation. IMiD effects were determined by western blot analysis, cell viability assay, microRNA array and apoptosis analysis.

Results: We identified argonaute 2 (AGO2) as a cereblon binding partner and found that the steady-state levels of AGO2 were regulated by cereblon. Upon treatment of IMiD-sensitive MM cells with lenalidomide, the steady-state levels of cereblon were significantly increased, whereas levels of AGO2 were significantly decreased. It has been reported that AGO2 plays a pivotal role in microRNA maturation and function. Interestingly, upon treatment of MM cells with lenalidomide, the steady-state levels of microRNAs were significantly altered. In addition, silencing of AGO2 in MM cells, regardless of sensitivity to IMiDs, significantly decreased the levels of AGO2 and microRNAs and massively induced cell death.

Conclusion: These results support the notion that the cereblon binding partner AGO2 plays an important role in regulating MM cell growth and survival and AGO2 could be considered as a novel drug target for overcoming IMiD resistance in MM cells.

In eutherians, the placenta acts as a barrier and conduit at the maternal-fetal interface. Syncytiotrophoblasts, the multinucleated cells that cover the placental villous tree surfaces of the human placenta, are directly bathed in maternal blood and are formed by the fusion of progenitor cytotrophoblasts that underlie them. Despite their crucial role in fetal protection, many of the events that govern trophoblast fusion and protection from microbial infection are unknown. We describe a three-dimensional (3D)–based culture model using human JEG-3 trophoblast cells that develop syncytiotrophoblast phenotypes when cocultured with human microvascular endothelial cells. JEG-3 cells cultured in this system exhibit enhanced fusogenic activity and morphological and secretory activities strikingly similar to those of primary human syncytiotrophoblasts. RNASeq analyses extend the observed functional similarities to the transcriptome, where we observed significant overlap between syncytiotrophoblast-specific genes and 3D JEG-3 cultures. Furthermore, JEG-3 cells cultured in 3D are resistant to infection by viruses and Toxoplasma gondii, which mimics the high resistance of syncytiotrophoblasts to microbial infections in vivo. Given that this system is genetically manipulatable, it provides a new platform to dissect the mechanisms involved in syncytiotrophoblast development and microbial resistance.

Despite serving as the primary entry portal for coxsackievirus B (CVB), little is known about CVB infection of the intestinal epithelium, owing at least in part to the lack of suitable in vivo models and the inability of cultured cells to recapitulate the complexity and structure associated with the gastrointestinal (GI) tract. Here, we report on the development of a three-dimensional (3-D) organotypic cell culture model of Caco-2 cells to model CVB infection of the gastrointestinal epithelium. We show that Caco-2 cells grown in 3-D using the rotating wall vessel (RWV) bioreactor recapitulate many of the properties of the intestinal epithelium, including the formation of well-developed tight junctions, apical-basolateral polarity, brush borders, and multicellular complexity. In addition, transcriptome analyses using transcriptome sequencing (RNA-Seq) revealed the induction of a number of genes associated with intestinal epithelial differentiation and/or intestinal processes in vivo when Caco-2 cells were cultured in 3-D. Applying this model to CVB infection, we found that although the levels of intracellular virus production were similar in two-dimensional (2-D) and 3-D Caco-2 cell cultures, the release of infectious CVB was enhanced in 3-D cultures at early stages of infection. Unlike CVB, the replication of poliovirus (PV) was significantly reduced in 3-D Caco-2 cell cultures. Collectively, our studies show that Caco-2 cells grown in 3-D using the RWV bioreactor provide a cell culture model that structurally and transcriptionally represents key aspects of cells in the human GI tract and can thus be used to expand our understanding of enterovirus-host interactions in intestinal epithelial cells.

The probiotic effects of Lactobacillus reuteri have been speculated to partly depend on its capacity to produce the antimicrobial substance reuterin during the reduction of glycerol in the gut. In this study, the potential of this process to protect human intestinal epithelial cells against infection with Salmonella enterica serovar Typhimurium was investigated. We used a three-dimensional (3-D) organotypic model of human colonic epithelium that was previously validated and applied to study interactions between S. Typhimurium and the intestinal epithelium that lead to enteric salmonellosis. Using this model system, we show that L. reuteri protects the intestinal cells against the early stages of Salmonella infection and that this effect is significantly increased when L. reuteri is stimulated to produce reuterin from glycerol. More specifically, the reuterin-containing ferment of L. reuteri caused a reduction in Salmonella adherence and invasion (1 log unit), and intracellular survival (2 log units). In contrast, the L. reuteri ferment without reuterin stimulated growth of the intracellular Salmonella population with 1 log unit. The short-term exposure to reuterin or the reuterin-containing ferment had no observed negative impact on intestinal epithelial cell health. However, long-term exposure (24 h) induced a complete loss of cell-cell contact within the epithelial aggregates and compromised cell viability. Collectively, these results shed light on a potential role for reuterin in inhibiting Salmonella-induced intestinal infections and may support the combined application of glycerol and L. reuteri. While future in vitro and in vivo studies of reuterin on intestinal health should fine-tune our understanding of the mechanistic effects, in particular in the presence of a complex gut microbiota, this the first report of a reuterin effect on the enteric infection process in any mammalian cell type.

Extra-intestinal pathogenic E. coli (ExPEC), including avian pathogenic E. coli (APEC), pose a considerable threat to both human and animal health, with illness causing substantial economic loss. APEC strain χ7122 (O78∶K80∶H9), containing three large plasmids [pChi7122-1 (IncFIB/FIIA-FIC), pChi7122-2 (IncFII), and pChi7122-3 (IncI₂)]; and a small plasmid pChi7122-4 (ColE2-like), has been used for many years as a model strain to study the molecular mechanisms of ExPEC pathogenicity and zoonotic potential. We previously sequenced and characterized the plasmid pChi7122-1 and determined its importance in systemic APEC infection; however the roles of the other pChi7122 plasmids were still ambiguous. Herein we present the sequence of the remaining pChi7122 plasmids, confirming that pChi7122-2 and pChi7122-3 encode an ABC iron transport system (eitABCD) and a putative type IV fimbriae respectively, whereas pChi7122-4 is a cryptic plasmid. New features were also identified, including a gene cluster on pChi7122-2 that is not present in other E. coli strains but is found in Salmonella serovars and is predicted to encode the sugars catabolic pathways. In vitro evaluation of the APEC χ7122 derivative strains with the three large plasmids, either individually or in combinations, provided new insights into the role of plasmids in biofilm formation, bile and acid tolerance, and the interaction of E. coli strains with 3-D cultures of intestinal epithelial cells. In this study, we show that the nature and combinations of plasmids, as well as the background of the host strains, have an effect on these phenomena. Our data reveal new insights into the role of extra-chromosomal sequences in fitness and diversity of ExPEC in their phenotypes.