Comprehensive Gene Analysis of IgG4-Related Ophthalmic Disease Using RNA Sequencing

• Main Authors: Masaki Asakage, Yoshihiko Usui, Naoya Nezu, Hiroyuki Shimizu, Kinya Tsubota, Kazuhiko Umazume, Naoyuki Yamakawa, Tomohiro Umezu, Hirotsugu Suwanai, Masahiko Kuroda, Hiroshi Goto
• Format: Article
• Language: English
• Published: MDPI AG 2020-10-01
• Series: Journal of Clinical Medicine
• Subjects: IgG4-related disease; IgG4-related ophthalmic disease; RNA sequencing; MMP12; SPP1; orbital lymphoproliferative disorders
• Online Access: https://www.mdpi.com/2077-0383/9/11/3458

High-throughput RNA sequencing (RNA-seq) uses massive parallel sequencing technology, allowing the unbiased analysis of genome-wide transcription levels and tumor mutation status. Immunoglobulin G4-related ophthalmic disease (IgG4-ROD) is a fibroinflammatory disease characterized by the enlargement of the ocular adnexal tissues. We analyzed RNA expression levels via RNA-seq in the biopsy specimens of three patients diagnosed with IgG4-ROD. Mucosa-associated lymphoid tissue (MALT) lymphoma, reactive lymphoid hyperplasia (RLH), normal lacrimal gland tissue, and adjacent adipose tissue were used as the controls (<i>n</i> = 3 each). RNA-seq was performed using the NextSeq 500 system, and genes with |fold change| ≥ 2 and <i>p</i> < 0.05 relative to the controls were defined as differentially expressed genes (DEGs) in IgG4-ROD. To validate the results of RNA-seq, real-time polymerase chain reaction (PCR) was performed in 30 IgG4-ROD and 30 orbital MALT lymphoma tissue samples. RNA-seq identified 35 up-regulated genes, including matrix metallopeptidase 12 (MMP12) and secreted phosphoprotein 1 (SPP1), in IgG4-ROD tissues when compared to all the controls. Many pathways related to the immune system were included when compared to all the controls. Expressions of MMP12 and SPP1 in IgG4-ROD tissues were confirmed by real-time PCR and immunohistochemistry. In conclusion, we identified novel DEGs, including those associated with extracellular matrix degradation, fibrosis, and inflammation, in IgG4-ROD biopsy specimens. These data provide new insights into molecular pathogenetic mechanisms and may contribute to the development of new biomarkers for diagnosis and molecular targeted drugs.