Microbiome-Gut Barrier Signatures in ALS — Experiment Design

SUMMARY

# Microbiome-Gut Barrier Signatures in ALS — Experiment Design ## Background and Rationale This prospective longitudinal clinical study addresses the critical knowledge gap regarding microbiome-gut barrier dysfunction in ALS pathogenesis and progression. Growing evidence suggests that gut dysbiosis and intestinal barrier compromise may contribute to neuroinflammation and motor neuron degeneration through the gut-brain axis, but causal relationships remain unclear. The study employs a comprehensi

METHODOLOGY NOTES

Phase 1 (Months 1-6): Recruit participants through ALS clinics and neurology departments. Obtain informed consent and baseline assessments including ALSFRS-R, demographic data, medication history, and dietary questionnaires. Collect baseline biosamples: fecal samples for microbiome analysis, blood for gut barrier markers (zonulin, LPS, I-FABP), and serum for cytokine profiling (IL-1β, TNF-α, IL-6, IL-10). Phase 2 (Months 3-24): Conduct longitudinal follow-up visits every 3 months. Repeat clinical assessments and biosample collection at each visit. Perform 16S rRNA sequencing using V3-V4 primers on Illumina MiSeq platform. Conduct shotgun metagenomic sequencing on subset of samples (n=60 per group) using Illumina NovaSeq. Measure gut permeability markers using ELISA kits: serum zonulin (Immundiagnostik), LPS (Hycult Biotech), and intestinal fatty acid binding protein. Analyze fecal and serum metabolites using LC-MS/MS targeting short-chain fatty acids, bile acids, and tryptophan metabol