Critical Steps for Human Gut Exfoliome RNA Profiling Analysis Using Non-Invasive Stool Samples.

INTRODUCTION: Dietary exposure and drug treatments influence gut cellular pathways and hence growth and potentially even the gut-brain-microbiome axis. Since eukaryotic mRNA presents poly-A sequence that distinguishes them from the prokaryotes mRNA, we could analyze the gene expression of human gut cells using exfoliated gut cells available in stool samples. However, the impact of the critical steps of these non-invasive methods must be analyzed. METHODS: We tested prokaryote contamination in all the steps of different procedures to analyze human exfoliome by microarrays and the influence of the fecal sampling collection process. RESULTS: The least bacterial contamination was found using RNA amplified with oligo dT from the GeneChip 3' IVT Pico Reagent Kit or using RNA purified by both Oligotex® + oligo dT. RNAlater® collection of feces affects the microarray results compared to directly frozen fecal samples, although both methods produce similar cDNA quality. CONCLUSION: This technique is a potential non-invasive diagnostic tool that can be applied to larger studies to quantify intestinal gene expression in humans with non-invasive samples, but samples should always be collected and analyzed under the same procedure.

Changes from 1986 to 2018 in the prevalence of obesity and overweight, metabolic control and treatment in children with type 1 diabetes mellitus in a Mediterranean area of Southeast Spain.

BACKGROUND: In recent decades, a global increase in the prevalence of childhood overweight and obesity has been observed in children and adolescents with type 1 diabetes. METHODS: This retrospective, cross-sectional, population study examined three groups (1986, 2007, and 2018) of children and adolescents aged < 16 years diagnosed with type 1 diabetes. Overweight and obesity were defined according to the World Health Organization recommendations. RESULTS: The prevalence of overweight and obesity in diabetic children and adolescents was 30.2% (95% CI: 23.1-38.3). There was a significant increase from 1986 to 2007 (11.9% to 41.7%, p = 0.002) and from 1986 to 2018 (11.9% to 34.8%, p = 0.012), but no significant differences were found from 2007 to 2018 (41.7% to 34.8%, p = 0.492). The age at diagnosis was lower in the group with excess body mass (p = 0.037). No significant differences were observed in age (p = 0.690), duration of diabetes (p = 0.163), distribution according to sex (p = 0.452), metabolic control (HbA1c, p = 0.909), or insulin units kg/day (p = 0.566), between diabetic patients with overweight or obesity and those with normal weight. From 2007 to 2018, the use of insulin analogs (p = 0.009) and a higher number of insulin doses (p = 0.007) increased significantly, with no increase in the prevalence of overweight and obesity. CONCLUSIONS: The prevalence of overweight and obesity in diabetic children and adolescents increased in the 1990s and the beginning of the twenty-first century, with stabilization in the last decade. Metabolic control and DM1 treatment showed no association with this trend.