Bariatric surgery treatment has increased worldwide, being the most efficient procedure for the treatment of severe obesity. The underlying beneficial metabolic effects go beyond weight loss, which has led to the consideration of RYGB for diabetic patients with milder forms of obesity. This is the first randomized controlled clinical trial that simultaneously explored clinical and microbiota changes in diabetic patients with class-1 obesity after RYGB versus standard medical therapy. We found that both RYGB and standard medical therapy groups improved anthropometric outcomes at 1 year of follow-up, with RYGB patients having significantly higher improvements. However, only RYGB patients achieved improvement/remission in diabetes status (n = 7, 87.5%) and, significantly improved anthropometric and glycaemic profiles, independently and progressively, during the first year of follow-up and simultaneously had gut microbiota changes.
Gut microbiota plays a relevant role in the complex causes of obesity and T2D, and is hypothesised to be involved in the modulation of metabolic status after RYGB. Microbial richness is a simple descriptive parameter of the microbiome. A healthy microbial ecosystem is usually characterised by an elevated level of microbial richness (21, 22). In addition, one of the key features of the microbiome that characterises obesity is a low level of microbial richness, which is correlated with metabolic disorders, such as low-grade inflammation, insulin resistance, and adipocyte size (21, 31). Here, microbial richness was measured at the genus level. Despite a trend of lower baseline genus richness in patients in the surgical arm, we observed a continuous increase after RYGB during the 12 months of follow-up. Our results confirm previous observations that RYGB increases microbiome richness, not only in patients with morbid obesity, but also for a broader BMI range, including patients with a BMI 30–35 Kg/m2 (29).
As expected, there was a strong inverse association between increase in microbial richness and improvement of clinical phenotype, including anthropometric, metabolic (waist circumference, diastolic blood pressure, A1c), and inflammatory (hsCRP) biomarkers. These results suggest that systemic and anatomical changes induced by RYGB can restore a putative loss of microbial richness with an improvement of metabolic profile. On the contrary, genus richness did not change in the standard medical therapy arm and ended-up being significantly lower at M12, with no differences in glycaemic profile, comparing to baseline. This suggests that standard medical therapy optimisation does not target the gut microbiota, which reinforces the hypothesis that modulation of gut microbiota by pre or probiotics could be a complementary strategy for improving glycaemic status in this context.
Across enterotypes, there were no significant differences in genus richness, nor in clinical variables at baseline. We hypothesise that as the sequencing depth was relatively low in this study, it provided a low number of observed genera, which significantly influenced the enterotyping outcome. Indeed, the evaluation of the Bray-Curtis distance is mostly driven by the most abundant genera, which affects the sensitivity of the analyses. Finally, the number of individuals in each group (four for the baseline K4 therapy, and five for the baseline K4 bypass) decreased the statistical power, making the interpretation difficult. However, we observe a significantly higher abundance of Bacteroides genus in the RYGB group in comparison with the medical group in baseline. This bacterial genus has been associated to a dysbiotic microbiome composition associated to low microbial diversity, low microbial cell density and enriched in several pathologies like Crohn disease, primary sclerosing cholangitis and Inflammatory Bowel Disease (32, 33). This dysbiotic microbiome composition is also enriched in severe obese patients under RYGB before surgical intervention, decreasing progressively after bariatric surgery in parallel with improvements in microbial richness, clinical conditions and weight loss (34).
One year after surgery, we observed a significant decrease in three bacterial taxa belonging to the Firmicutes phylum (Ruminococcus, Lachnospiraceae_unclassified and Faecalibacterium), which are recognised as having anti-inflammatory properties with a beneficial impact on metabolic health (14, 29). The decrease of Firmicutes lineages after bariatric surgery has been reported in different studies (35–37), whereas contradictory results are observed in diffferent studies related to the presence of Faecalibacterium prausnitzii (38–40). This leads us to hypothesise that, despite RYGB improving the metabolic status in T2D patients with mild obesity, RYGB did not have the ability to restore a healthy microbiome composition, especially if it started with a highly dysbiotic microbiome state. We also observed an increase in abundance of bacteria of the phylum Proteobacteria (Klebsiella, Gammaproteobacteria, Enterobacter, Gammaproteobacteria_unclassified), one year after the surgical intervention, as well as of Veillonellaceae_unclassified (Firmicutes phyla). Increases in Veillonella and other oral bacterial lineages after RYGB surgery has been reported in other studies associated to decrease of acid secretions consequence to the stomach size reduction, which could facilitate the intestinal colonization of oral bacteria (36, 40), whereas an increase in gamma-proteobacteria after bariatric surgery is a common finding - both in humans and in mice (30, 41, 42). In our study, we also observed an increase of the Proteobacteria/Firmicutes ratio after RYGB. The increase of bacteria belonging to the phylum Proteobacteria was associated with the improvement of metabolic and inflammatory parameters after bariatric surgery (43). In an animal model, the increase of proteobacteria was also accompanied by a reduction in inflammatory response and glucose homeostasis improvement (44). The phylum Proteobacteria is composed of facultative anaerobes, and consequently oxygen increase (45) combined with higher pH after RYGB (in the gut) could contribute to an increase of these bacteria in parallel with improvement s t on metabolic health. If this increase in proteobacterial lineages after bariatric surgery have a direct contribution to improvement of health status of severe obese patients or is a response to the drastic anatomical changes in the gut environment consequence of the surgical procedure (high oxygen availability, higher pH, higher amounts of undigested nutrients in parallel to caloric restriction) would require further experiments with animal models in parallel with more precise characterization of enterobacterial lineages increased after bariatric surgery at strain level with shotgun metagenomics data and de-novo sequence assembly for better understanding of its functional role.
On the other hand, we did not observe any significant differences with regards to ratio Firmicutes/Bacteroidetes (rFB). The concept that obesity is associated with a lower percentage of Bacteroidetes and a higher percentage of Firmicutes in obese individuals is contradicted by several studies, which demonstrate that there is no difference in relative abundance of Firmicutes and/or Bacteroides and no association of weight loss with the rFB (46, 47) in obese individuals. Furthermore, T2D has been linked to a decreased abundance of Firmicutes and an increase in bacteria belonging to Bacteroidetes and Proteobacteria, when compared to obese patients (48). However, it is difficult to validate these links from the results of our study, which included T2D patients with mild obesity, knowing the Firmicutes phylum and Bacteroides contain at least 250 and 20 genera, respectively. Higher taxonomic levels may not necessarily reflect specific bacteria changes, however, the results of our study are in line with those of Campisciano et al., as they corroborate that rFB is not a predictive biomarker of the outcome for metabolic surgery (49).
6.1. Strengths and Limitations
This study assessed patients’ evolution at different points in time over the first year of follow-up, including the first and third months after interventions, contrary to the majority of published studies which lack comprehensive data regarding the first six months following baseline and through a 12-month period. These intermediate timepoint assessments allowed us to monitor a clear evolution of the clinical profile.
The sample size could have impaired to a degree the multivariate analysis - however, the results are consistent throughout the timepoints as well as with other studies. Of note is the fact that our observations can be limited by different technical aspects of microbiome analysis, including the collection, generation, and quantification of the abundance profiles. In addition, heterogeneity in dietary profiles and physical activity can also explain part of phenotypic outcomes during the treatment. Dietary data were mainly recorded for the medical arm, but not for the surgical arm throughout the different time-points, which limits our ability to control for food intake in the changes in microbiome profiles. Consequently, dietary analyses were not shown.
In conclusion, our research suggests that there is a remarkable phenotypic improvement after metabolic surgery which occurs simultaneously with gut microbiota changes. Nevertheless, gut microbiome changes alone cannot explain the beneficial metabolic health impact of RYGB. Other mechanisms such as diet, hormonal changes, bile acids metabolism, and physical activity need to be further explored in this equation in order to better explain the metabolic improvement of T2D patients with mild obesity after RYGB.