Advancing brain barriers RNA sequencing: guidelines from experimental design to publication

Area of Science:

• Neuroscience
• Genomics
• Bioinformatics

Background:

• RNA sequencing (RNA-Seq) is crucial for analyzing gene expression and characterizing tissues.
• Its application in brain barrier research has yielded valuable bulk and single-cell RNA-Seq datasets.
• Understanding brain barrier molecular mechanisms requires well-executed RNA-Seq studies, which involve complex workflows.

Purpose of the Study:

• To provide comprehensive guidelines for RNA-Seq studies in brain barrier research.
• To address challenges identified through interdisciplinary collaboration.
• To facilitate the design, execution, and interpretation of RNA-Seq experiments for brain barriers.

Main Methods:

• The study integrates experience from the European PhD Training Network BtRAIN, fostering collaboration between bioinformaticians and brain barrier researchers.
• It offers a step-by-step workflow for RNA-Seq studies, focusing on vertebrate blood-brain barrier (BBB) and blood-cerebrospinal-fluid barrier (BCSFB).
• Guidelines cover experimental design, workflow decisions, and data interpretation, highlighting strengths and weaknesses of choices.

Main Results:

• RNA-Seq studies offer powerful datasets for deepening knowledge of brain barrier molecular mechanisms.
• The guidelines address obstacles in RNA-Seq data analysis for brain barriers.
• Recommendations are provided for accurate data interpretation and publication to ensure data accessibility and reproducibility.

Conclusions:

• Next-generation transcriptomic profiling of brain barriers is essential for understanding CNS homeostasis and disease.
• Advancements in RNA-Seq necessitate interdisciplinary approaches between brain barrier researchers and bioinformaticians.
• These guidelines aim to enhance collaboration and advance RNA-Seq study design within the brain barrier research community.