Project description:Background- Resistant starch is a prebiotic metabolized by the gut bacteria. It has been shown to attenuate chronic kidney disease (CKD) progression in rats. Previous studies employed taxonomic analysis using 16S rRNA sequencing and untargeted metabolomics profiling. Here we expand these studies by metaproteomics, gaining new insight into the host-microbiome interaction. Methods- Differences between cecum contents in CKD rats fed a diet containing resistant starch with those fed a diet containing digestible starch were examined by comparative metaproteomics analysis. Taxonomic information was obtained using unique protein sequences. Our methodology results in quantitative data covering both host and bacterial proteins. Results - 5,834 proteins were quantified, with 947 proteins originating from the host organism. Taxonomic information derived from metaproteomics data surpassed previous 16S RNA analysis, and reached species resolutions for moderately abundant taxonomic groups. In particular, the Ruminococcaceae family becomes well resolved – with butyrate producers and amylolytic species such as R. bromii clearly visible and significantly higher while fibrolytic species such as R. flavefaciens are significantly lower with resistant starch feeding. The observed changes in protein patterns are consistent with fiber-associated improvement in CKD phenotype. Several known host CKD-associated proteins and biomarkers of impaired kidney function were significantly reduced with resistant starch supplementation. Conclusions- Metaproteomics analysis of cecum contents of CKD rats with and without resistant starch supplementation reveals changes within gut microbiota at unprecedented resolution, providing both functional and taxonomic information. Proteins and organisms differentially abundant with RS supplementation point toward a shift from mucin degraders to butyrate producers.
Project description:This phase II, randomized pilot trial studies the effect of the consumption of foods made with resistant starch compared to foods made with corn starch on biomarkers that may be related to colorectal cancer progression in stage I-III colorectal cancer survivors. Foods made with resistant starch may beneficially influence markers of inflammation, insulin resistance, and the composition of gut bacteria in colorectal cancer survivors.
Project description:Resistant starches (RS) are dietary compounds processed by the gut microbiota into metabolites, such as butyrate, that are beneficial to the host. The production of butyrate by the microbiome appears to be affected by the plant source and type of RS as well as the individual’s microbiota. In this study, we used in vitro culture and metaproteomic methods to explore the consistency and variations in individual microbiome's functional responses to three types of RS - RS2(Hi Maize 260), RS3(Novelose 330) and RS4(Fibersym RW). Results showed that RS2 and RS3 significantly altered the levels of protein expression in the individual gut microbiomes, while RS4 did not result in significant protein changes. Significantly elevated protein groups were enriched in carbohydrate metabolism and transport functions of families Eubacteriaceae, Lachnospiraceae and Ruminococcaceae. In addition, Bifidobacteriaceae was significantly increased in response to RS3. We also observed taxon-specific enrichments of starch metabolism and pentose phosphate pathways corresponding to this family. Functions related to starch utilization, ABC transporters and pyruvate metabolism pathways were consistently increased in the individual microbiomes in response to RS2 and RS3; in contrast, the downstream butyrate producing pathway response varied. Our study confirm that different types of RS have markedly variable functional effects on the human gut microbiome, and also found considerable inter-individual differences in microbiome pathway responses.
Project description:Background. Resistant Starch (RS) improves CKD outcomes. In this report we study how RS modulates host-microbiome interactions in CKD by measuring changes in abundance of proteins and bacteria in the gut. In addition, we demonstrate RS-mediated reduction in CKD-induced kidney damage. Methods. Eight mice underwent 5/6 nephrectomy to induce CKD and 8 served as healthy controls. CKD and Healthy (H) groups were further split into those receiving RS (CKDRS, n=4; HRS, n=4) and those on normal diet (CKD, n=4, H, n=4). Kidney injury was evaluated by measuring BUN/creatinine and by most-mortem histopathological evaluation. Cecal contents were analyzed mass spectrometry-based metaproteomics. PEAKS Studio was used to identify peptides via de novo sequencing. A set of R/Bioconductor packages and in house written scripts was further used to infer bacteria present, to evaluate changes in proteins and bacterial abundances, and to perform statistical analysis and hierarchical clustering. Results. The 5/6 nephrectomy compromised kidney function as seen by an increase in creatinine and BUN. Representative photomicrographs of trichrome-stained kidney sections showed reduced tubulointerstitial injury in CKD mice fed HAM-RS2 diet comparing to CKD mice fed normal diet. Identified organisms and proteins point toward a higher population of butyrate-producing bacteria, and reduced abundance of mucin-degrading bacteria. Conclusion. Resistant starch slows the progression of chronic kidney disease. Gut barrier function through maintenance of the mucin barrier plays a role in RS-associated improvements in CKD phenotype. Resistant starch supplementation leads to the active bacterial proliferation and the reduction of harmful bacterial metabolites.
Project description:Background Lytic polysaccharide monooxygenases (LPMOs) are often studied in simple models involving activity measurements of a single LPMO or a blend thereof with hydrolytic enzymes towards an insoluble substrate. However, the contribution of LPMOs to polysaccharide breakdown in complex cocktails of hydrolytic and oxidative enzymes, similar to fungal secretomes, remains elusive. Typically, two starch-specific AA13 LPMOs are encoded by mainly Ascomycota genomes. Here, we investigate the impact of LPMO loss on the growth and degradation of starches of varying resistance to amylolytic hydrolases by Aspergillus nidulans. Results Deletion of the genes encoding AnAA13A that possesses a CBM20 starch-binding module, AnAA13B (lacking a CBM20) or both AA13s genes resulted in reduction in growth on solid media with resistant, but not soluble processed potato starch. Larger size and amount of residual starch granules were observed for the AA13 KO strains as compared to the reference and the impairment of granular starch degradation was more severe for the AnAA13A KO based on a microscopic analysis. After five days of growth on raw potato starch in liquid media, the mount of residual starch was about 5-fold higher for the AA13 KO strains compared to the reference, which underscores the importance of LPMOs for degradation of especially resistant starches. Proteomic analyses revealed substantial changes in the secretomes of the AA13 double KO, followed by the AnAA13A deficient strains, whereas no significant changes in the proteome were observed for the AnAA13B deficient strain. Conclusions This study shows that the loss of AA13, especially the starch-binding AnAA13A, impairs degradation of resistant potato starch, but has limited impact less-resistant wheat starch and has no impact at all on processed solubilised starch. The effects of LPMO loss are more pronounced at the later stages of fungal growth, when the less-accessible regions of the substrate accumulate. The striking impact of the loss of a single LPMO against a whole secretome offers insight into the crucial role played by AA13 in the degradation of resistant starch and presents a methodological framework to analyse the contribution of distinct LPMOs towards complex substrates under in vivo conditions.
Project description:RNA-seq based transcriptome analysis was employed to understand the genome-wide expression patterns under cultivation with resistant starch (RS). To identify differentially expressed genes, we compared RS-induced transcriptome to non-RS transcriptome as sole carbon source.
Project description:The starch, acting as the major energy-producing component of the daily diet, is the main carbohydrate in mammal nutrition. However, the nutritional value of starch can vary widely depending upon its source and site of digestion. The distinct physiological responses were previously observed both in human and other mammals, but still little is known about the underlying mechanisms regarding the metabolic shifts due to the intake of various dietary starches. Here, we assessed the overall metabolic changes in weaned pigs induced by different dietary starch sources at the transcriptome level. Sixteen weaned pigs (DurocÃLandraceÃYorkshire) were selected and randomly allotted to diets containing either wheat (WH) or cassava (CA) starch as the energy source (n=8). We measured serum metabolites and hormones and generated transcriptional profiles of liver. 648 genes in liver were differentially expressed in response to dietary starch sources. Pathway analysis indicated that dietary starch sources altered both carbohydrate and lipid metabolism in liver. In contrast, CA may be more healthful as dietary energy source than WH by down-regulating lipogenesis and steroidogenesis in liver. Sixteen weaned pigs (DurocÃLandraceÃYorkshire) with an average initial body weight of 7.37±0.25 kg were selected and randomly allotted to two dietary treatments (either wheat or cassava starch as the energy source) for 21 d. At the end of the trial, the liver tissue were collected for transcriptome analysis using Agilent porcine microarrays.
Project description:The purpose of this study is to determine if consumption of bagels made with resistant starch for 8 weeks can improve markers of type 2 diabetes, colon cancer and satiety in adults.