Spatial Transcriptomics Reveals Interaction of Injury-Derived Osteopontin with Mesothelial Cells in Serosal Scarring

Abstract

Abstract Background Abdominal surgeries are often live-saving procedures. However, surgical interventions in the peritoneal cavity can cause the formation of post-surgical adhesions. These irreversible scar bands can trigger severe complications, such as chronic pelvic pain, small bowel obstruction, and infertility in women. Unfortunately, no effective treatment options for adhesion patients exist. On a cellular level, mesothelium-derived mesenchymal myofibroblasts have been identified as key pro-fibrotic players in adhesion formation. Aims Our aim is to study the molecular mechanisms of mesothelial cell recruitment in serosal repair and scarring, and thereby to identify potential targets for specific pharmacological anti-adhesion treatments. Methods We used a mesothelial cell reporter mouse system (Wt1CreERT2xRosa26tdTomato) and experimental injury models to study the differences of mesothelial cell recruitment to sites of adhesion formation in comparison to wound sites healing scar-free. To answer how injury zones affect the accumulation of mesothelial cells, we used untargeted spatial transcriptomics, identifying candidate molecules that were i) upregulated in adhesion formation sites, ii) not highly expressed in scar-free wound sites, and iii) had the potential to interact with mesothelial cells. Results We observed strong spatiotemporal correlation between accumulation of tdTomato+ cells and consecutive scarring. Spatial deconstruction of scar-specific signaling identified the candidate molecule Secreted phosphoprotein 1; an inflammatory zone-specific gene that encodes Osteopontin (OPN). After confirming increased OPN protein levels 7d post-surgery, we investigated its effect on adhesion formation using Spp1-/- mice. Indeed, Spp1-deficient mice showed significantly lower adhesion scores compared to control mice. Mechanistically, recombinant OPN had no chemotactic effect on primary human mesothelial cells, however triggered the upregulation of transcription factors associated with mesothelial-to-mesenchymal transition. Conclusion Together, these findings suggest a pivotal role for OPN in linking inflammation at injury sites with mesothelial cell recruitment in post-surgical serosal scarring. Therefore, OPN represents a potential target for novel pharmacological strategies to specifically inhibit scarring without affecting normal wound repair.