Assay of intestinal disaccharidases
Abstract
A simplified method for the assay of small intestine disaccharidase activity is described which is suitable for use both on the “standard” scale (final volume 3.2 ml) and on the “ultramicro” scale (final volume 320 μl).
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When digestive physiology doesn't match “diet”: Lumpenus sagitta (Stichaeidae) is an “omnivore” with a carnivorous gut
2023, Comparative Biochemistry and Physiology -Part A : Molecular and Integrative PhysiologyWhat an animal ingests and what it digests can be different. Thus, we examined the nutritional physiology of Lumpenus sagitta, a member of the family Stichaeidae, to better understand whether it could digest algal components like its better studied algivorous relatives. Although L. sagitta ingests considerable algal content, we found little evidence of algal digestion. This fish species has a short gut that doesn't show positive allometry with body size, low amylolytic activity that actually decreases as the fish grow, no ontogenetic changes in digestive enzyme gene expression, elevated N-acetyl-glucosaminidase activity (indicative of chitin breakdown), and an enteric microbial community that is consistent with carnivory and differs from members of its family that consume and digest algae. Hence, we are left concluding that L. sagitta is not capable of digesting the algae it consumes, and instead, are likely targeting epibionts on the algae itself, and other invertebrates consumed with the algae. Our study expands the coverage of dietary and digestive information for the family Stichaeidae, which is becoming a model for fish digestive physiology and genomics, and shows the power of moving beyond gut content analyses to better understand what an animal can actually digest and use metabolically.
An increased weaning age and liquid feed enhances weight gain compared to piglets fed dry feed pre-weaning
2023, AnimalIncreasing age and providing liquid creep feed could potentially increase the solid feed intake in pre-weaning piglets, which may in turn promote gut maturation and post-weaning feed intake, possibly lessening the severity of the growth-check associated with the suckling-to-weaning transition. Therefore, this study aimed to investigate if feeding dry- versus liquid creep feed (DF vs. LF) and weaning in week 4 or 5 (4W or 5W) could accelerate maturational changes to the small intestines of pre-weaning piglets by increasing digestive and absorptive capacity. In a 2 × 2 factorial study the effect of weaning age (WA) and feeding strategy (FS) on weaning weight, pre-weaning accumulated gain (AG), and average daily gain was measured for 12 923 piglets. A subpopulation of 15 piglets from each treatment group (4WDF, 4WLF, 5WDF and 5WLF; n = 60) were sacrificed to assess the effects of WA and FS on weight of digestive organs, activity of maltase, lactase and sucrase, and gene expression level of sodium-glucose linked transporter 1 (SGLT-1), glucose transporter 2 (GLUT2) and peptide transporter 1 (PepT1) in the proximal part of the small intestine (SI). No interactions were found but average weaning weight was affected by WA (P < 0.001) and FS (P < 0.001), where 5W were heavier than 4W and LF were heavier than DF. Correspondingly, the average daily gain (ADG) was affected by both WA (P = 0.003) and FS (P < 0.001). Only WA affected the relative weight of the digestive organs, where stomach weight, weight of SI and colon weight were heavier in 5W piglets compared to 4W. Lactase activity tended to decrease with age (P = 0.061), but there was no difference in the activity of maltase or sucrase between any of the treatment groups. Similarly, there was no differences in gene expression level of SGLT1, GLUT2 or PepT1 between neither the two ages nor feeding strategies. In conclusion, both WA and FS affect weaning weight and weight gain of piglets in the pre-weaning period.
Ergosterol fraction from Agaricus bisporus modulates adipogenesis and skeletal glucose uptake in high fat diet induced obese C57BL/6 mice
2023, Life SciencesThe study aimed to optimize a method of extracting ergosterol rich concentrate (ECF) and to evaluate its significant impact on adipogenesis and associated complications in high-fat diet (HFD) induced obese mice.
A comparative analysis (soxhlet and ultra sound assisted extraction) was done to obtain the highest yield of ergosterol from Agaricus bisporus. The ECF was evaluated for the biological effect on 3T3-L1 pre-adipocytes in-vitro and in male C57BL/6 mice model in-vivo.
Ultra sound assisted extraction method using the solvent n-hexane resulted in highest ergosterol yield. ECF treatment significantly reduced the differentiation and lipid accumulation in pre-adipocyte cells without any cytotoxicity. In-vivo study illustrated beneficial impact on cholesterol metabolism by down regulating the hepatic gene expression of LXR-α, HMG-CoR and up-regulating LDL-R expression. Significant increase in fecal excretion of cholesterol and bile acids have also been observed among the ECF treated animals compared to high fat diet (HFD) fed mice. ECF had an anti-adipogenic activity in-vivo mainly by inhibiting the activity of PPAR-γ, C/EBP-α and SREBP-1c. The results also depicted the improvement of obesity associated insulin resistance by ECF treatment manly via decrease in plasma resistin and up-regulation in skeletal GLUT4 protein expression.
Our study illustrated diverse activity of ECF in the therapeutic management of obesity associated metabolic complications mainly by reducing adipogenesis and improving glucose uptake in skeletal muscle in conjunction with improved cholesterol metabolism.
Sustained changes in digestive physiology and microbiome across sequential generations of zebrafish fed different diets
2022, Comparative Biochemistry and Physiology -Part A : Molecular and Integrative PhysiologyAlterations to ratios of protein and fiber in an organism's diet have been shown to structurally and functionally alter its individual digestive physiology. However, it is unclear how these dietary changes may affect phenotypic changes across generations. We utilized feeding trials, morphological analyses, enzyme activities, and 16S rRNA sequencing of the gut microbiome of zebrafish (Danio rerio) to determine how variations to fiber and protein concentrations, kept consistent across sequential generations, affect phenotypic changes. Our results show that Parental (P) and first generation (F1) fish did not differ from each other in terms of their intestine length, intestine mass, enzyme activity levels, and microbial community composition for any of the three experimental diets (high-protein/low-fiber, moderate-protein/fiber, and low-protein/high-fiber). However, each of the three experimental diets for the P and F1 fish, as well as the ancestral diet fish, did have distinct microbial community structure from one another. This indicates that there is a strong dietary effect on digestive physiology and gut microbial community and that these effects are consistent when the diet is kept homogenous across generations.
Ultrasonication of Milk Decreases the Content of Exosomes and MicroRNAs in an Exosome-Defined Rodent Diet
2022, Journal of NutritionBovine milk exosomes (BMEs) harbor regulatory proteins, lipids, and microRNAs. Consumption of an exosome- and RNA-depleted (ERD) diet elicited phenotypes compared with controls fed an exosome- and RNA-sufficient (ERS) diet in mice. All other ingredients were identical in the diets. ERD and ERS diets were prepared by substituting ultrasonicated and nonultrasonicated milk, respectively, for casein in the AIN-93G formulation.
The objective of this study was to assess the effect of ultrasonication of milk on exosome content and bioavailability, and cargo content.
Bovine milk was ultrasonicated and exosomes were isolated by ultracentrifugation [ultrasonicated exosomes (USEs)]; controls were not ultrasonicated [nonultrasonicated exosomes (NSEs)]. Exosome count, size, and morphology were assessed using a nanoparticle tracker and electron microscopy. RNAs, lipids, and proteins were analyzed by RNA sequencing and MS. Intestinal transport, bioavailability, and distribution were measured by using fluorophore-labeled USEs and NSEs in Caco-2 cells, FHs 74 Int cells, and C57BL/6J mice (n = 3; age: 6–8 wk).
The exosome count was 76% ± 22% lower in USEs than in NSEs (P < 0.05). Ultrasonication caused a degradation of ≤100% of microRNAs. USEs and NSEs contained 145 and 332 unique lipid signatures, respectively (P < 0.05). We detected a total of 525 and 484 proteins in USEs and NSEs, respectively. The uptake of USEs decreased by 46% ± 30% and 40% ± 27% compared with NSEs in Caco-2 and FHs 74 Int cells, respectively (P < 0.05). The hepatic accumulation of USEs was 48% ± 28% lower than the accumulation of NSEs in mice (P < 0.05).
Ultrasonication of milk depletes bioavailable BMEs in studies of Caco-2 cells, FHs 74 Int cells, and C57BL/6J mice and causes a near-complete degradation of microRNA cargos.
Digestive and immune functions in the intestine of wild Ballan wrasse (Labrus bergylta)
2021, Comparative Biochemistry and Physiology -Part A : Molecular and Integrative PhysiologyThis study was carried out to profile key characteristics of intestinal functions and health in wild-caught Ballan wrasse. To describe functional variation along the intestine, samples were collected from four intestinal segments, named from the proximal to the distal segment: IN1, IN2, IN3 and IN4. The sections showed quite similar structure, i.e. regarding mucosal fold height and branching, lamina propria and submucosal width and cellular composition and thickness of the muscle layers. Leucine aminopeptidase and maltase capacity decreased from IN1 to IN4, suggesting a predominant role of IN1 in digestion. Gene expression levels of vitamin C transporter (slc23a1) and fatty acid transporters (cd36 and fabp2) were higher in IN1 than in IN4, indicating a more important role of the proximal intestine regarding transport of vitamins and fatty acids. Higher expression of the gene coding for IgM heavy chain constant region (ighm) was found in IN4 than in IN1, suggesting an important immune function of the distal intestine. Other immune related genes il1b, il6, cd40, showed similar expression in the proximal and the distal part of the intestine. Parasite infection, especially the myxozoan parasite Enteromyxum leei, coincided with infiltration of lymphocytic and eosinophilic granular cells in the submucosa and lamina propria. The present study established reference information necessary for interpretation of results of studies of intestinal functions and health in cultured Ballan wrasse.