Assessment of XCI skewing and demonstration of XCI escape region based on single-cell RNA sequencing: comparison between female Grave's disease and control.

Background: The reactivation and loss of mosaicism hypothesis due to X chromosome inactivation (XCI) skewing and escape could influence gender differences in autoimmune diseases. XCI selectively inactivates one of the two X chromosomes in females.

Methods: To estimate XCI skewing and the occurrence of XCI escape, we conducted a normal female (NF) without a history of autoimmune thyroid disease (AITD) and a patient with Grave's disease (GD) based on a thyroid diagnosis. After single-cell RNA sequencing, heterozygous variants were converted and transformed. XCI skewing was calculated using the formula and the skewing degree was defined. NF/GD genes were compared using correction methods. Positions are heterozygous within a single cell as indicated by a unique barcode.

Results: XCI skewing showed 45.8%/48.9% relatively random, 29.4%/27.0% skewing, 24.6%/23.7% severe skewing, and 0.2%/0.4% extreme severe skewing. 24.8%/24.1% in NF/GD exhibited severe skewing or higher. A total of 13 genes were significantly associated with XCI skewing ratios in NF/GD cells. In total, 371/250 nucleotide positions with only one barcode (representing a unique cell) were identified for XCI escape. A total of 143/52 nucleotide positions spanned 20/6 genes, and 12/1 genes were identified as XCI escapes.

Conclusions: These results could aid in understanding the immunogenetics of gender differences in various autoimmune disease pathophysiologies.