Adaptación metabólica de la microbiota intestinal a cambios dietéticos durante la obesidad

Resumen

The human distal gut harbours a vast ensemble of microbes known gut microbiota. Recent studies show that the transfer of fecal bacteria improves metabolic markers of metabolic syndrome and insulin resistance in humans. These results raise the possibility of developing new therapeutic strategies based on the modulation of intestinal microbiota to treat or prevent obesity. Numerous studies have tried to identify and characterize specific microorganisms or a consortium of functionally related species involved in weight gain or loss. Despite great efforts, there are conflicting data because the structural and functional descriptions of the intestinal microbial communities are exclusively based on genetic data, either metagenomic and/or 16S rDNA sequences, wrongly assuming the existence of a correlation between overall microbial metabolism and community structure. To study the link between microbiota and obesity, we are exploring a different strategy, a “down-to-top” approach using metabolic activity as a means to identify the active microbial community and its dynamics in response to calorie restriction in obese. Under this premise, we initially showed that gut microbial metabolism in obese and lean was different. By analyzing the diversity of over 300 sequences of full 16S rDNA sequences (10% common species), 22 Mbps metagenomic consensus sequences (~ 70% common) and expression profiles of 613 different proteins (82% common) we showed that obese total microbiota was more abundant on the phylum Firmicutes (94.6%) compared to Bacteroidetes (3.2%), although metabolically active microbiota clearly behaves in a more homogeneus manner with both contributing equally. The lean gut showed a notable shift towards Bacteroidetes (18.9% of 16S rDNA), which became the most active fraction (81% protein). Although the two gut communities maintained largely similar gene repertoires and functional profiles, improved pili- and flagella-mediated host colonization and improved capacity for both complementary aerobic and anaerobic de novo B12 synthesis, 1,2-propanediol catabolism (most likely participating in de novo B12 synthesis) and butyrate production were observed in the obese gut, whereas bacteria from lean gut seemed to be more engaged in vitamin B6 synthesis. Interestingly, among the most abundant proteins we found two glycosyl hydrolases or glycosidases, one pectate lyase and one α-arabinofuranosidase. Glycosidases are essential enzymes required for the microbial fermentation of dietary carbohydrates polysaccharides and the production of short chain fatty acids that serve as nutrients for colonocytes, contribute to gut gluconeogenesis and activate gut-brain neural circuits. Therefore, we decided to explore the relation between microbial glycosidases with obesity. After analysing the specific activity of 23 different glycosidases, we found that the total glycosidase activity was 40 times higher in the obese and, most importantly, it positively correlated with body mass index (BMI), fasting glucose and insulin resistance (r2 ≥ 0.95 ). Once this metabolic connection between microbial functionality and host was established, we reasoned that fluorescent labelling of β-galactosidase intracellular activity would actually identify the active members within the total gut microbiota. For this purpose, fluorescently-labeled β-galactosidase-containing microorganisms were sorted and 16S rDNA gene sequencing was used to characterize diet-induced changes in microbial diversity. Our data demonstrate that the gut contains a distinctive set of anabolic microorganisms, enriched in members of Bacteroidetes, Proteobacteria and Actinobacteria, and distributed within a more even and diverse community that significantly differed from total gut microbiota. The active microbiota was less resilient to diet-induced adjustments than total community. Long-term dietary intervention increased species richness, reduced the Firmicutes:Bacteroidetes ratio and shifted the active community structure and composition to a lean microbiota. This shift did not reduce ecosystem activity because BMI determined functionality regardless of community composition. Remarkably, microbial functionality (α-glucosidase and β-galactosidase activities) positively correlated with metabolic markers, BMI and insulin resistance in obese. The set of data presented here may constitute a significant contribution to the field in several aspects. Many groups are trying dietary and therapeutic strategies targeting the intestinal microbiota to reduce obesity. Our data indicate that the change in the composition of the intestinal microbiota may not be enough to reduce its contribution to the development of metabolic obesity. Total intestinal microbiota and its active subset are significantly different. Therefore, predictions of functionality based solely on the detailed taxonomic classification for explaining metabolism are clearly unrealistic. Moreover, the strong correlation between the metabolic activity of the microbiota with BMI and insulin resistance suggests that selective inhibition of microbial glycosidases could establish potential therapy to reduce the contribution of energy extraction from diet polysaccharides. Finally, our findings points to an unknown molecular mechanism associated with subject´s BMI that modulates the anabolic capacity of the intestinal microbiota.