Background/Purpose: Expression of HLA-B27 and human beta 2-microglobulin (hb₂m) in rats induces a spontaneous inflammatory disease resembling human spondyloarthritis (SpA). SpA overlaps with IBD in genetic predisposition, pathogenic mechanisms and clinical manifestations. While key components of rat SpA have been studied in great detail, a complete understanding of the associated inflammatory bowel disease (IBD) has not been established. The goal of this project is to determine how HLA-B27 alters gut transcriptome in transgenic rats that develop SpA-like disease. HLA-B27 and hb₂m transgenic (TG) rats (33-3 transgene locus) on a Lewis (LEW), Fischer (F344), and Dark Agouti (DA) background from two different animal facilities are being studied along with strain-specific controls. TG LEW and F344 rats develop SpA beginning with colitis at about 8 weeks of age, while TG DA rats are disease free. Arthritis develops later in LEW and F344 TG animals, and is more variable.

Methods: To assess colitis, tissue samples from colon were assessed by H&E staining and scored for histological differences. Samples were also assessed for transcriptome differences using RNA-Seq. Total poly (A) enriched RNA was reverse transcribed into double stranded complementary DNA (dscDNA) and sequenced on Illumina HiSeq 2000. Raw reads were mapped to Rat rn5 genome using Tophat (2.0.8). Transcript expression levels in Reads per Kilobase Million (RPKM) and ANOVA comparisons were calculated using Partek GS (6.6/6.14.0514). A minimum fold change of 50% (p value ≤ 0.05% and q value ≤0.2) with Max Mean ≥-3.3 was used as cutoff criteria for identifying differentially expressed genes between TG and WT animals on LEW, F344 and DA background at 2, 3 and 6 months. These genes were subjected to pathway exploration by Ingenuity Pathway Analysis (IPA) software.

Results: Inflammation in the colon was documented by histopathological analysis.  Transcriptome analysis revealed that LEW and F344 TG animals exhibit up-regulation of the genes for IFN response (e.g. Tap1, Tap2, Irf1, Cxcl10, Oas1 Gbp-2, Stat1). The Il17 pathway is highly up regulated at all age groups whereas Il23 up regulation became statistically significant at 6 months of age. Apoptotic signaling and iNos (Nos2) pathways as well as the oxidative stress (Gpx2, Nox1, Duox2) pathway in colon were up-regulated as compared to their age matched WT controls. Susceptibility genes (Card9, Nod2) as well as IBD associated genes (Tnf, Ltb, Reg3γ, Ccl2, Ccr7) were up regulated in TG F344 and LEW animals. DA background had a protective affect since TG DA did not exhibit significant gene expression changes consistent with the fact that they do not develop either SpA or IBD.

Conclusion: Transcriptome analysis of the TG inflamed colon depicts upregulation of interferon and Il23/Il17 pathways suggesting a shift in the immune microenvironment in the colon. The interferon signature contrasts results recently obtained from isolated dendritic cells, and underscores the role of interferon in this disease process. These results increase our understanding of SpA associated IBD and may lead to the identification of potential biomarkers for use in diagnosis and treatment.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/analysis-of-the-gut-transcriptome-in-hla-b27-transgenic-rats-by-rna-seq-reveals-prominent-interferon-and-il-23il-17-axis-signatures/