The use of 68Ga-FAPI probe, a novel way to detect the peritoneal fibrosis related to peritoneal dialysis

Peritoneal fibrosis (PF) is a primary pathological cause of peritoneal membrane failure and technique failure in patients undergoing long-term peritoneal dialysis (PD). The core mechanism involves persistent injury to the peritoneal membrane due to exposure to non-physiological dialysate factors, including high glucose concentrations, hyperosmolarity, and glucose degradation products. This injury triggers a cascade of events, notably the mesothelial-to-mesenchymal transition (MMT) of human peritoneal mesothelial cells (HPMCs), leading to the activation and proliferation of fibroblasts and excessive deposition of extracellular matrix. This ultimately results in progressive submesothelial thickening and angiogenesis. Currently, the clinical gold standard for assessing the severity and progression of PF is histologic analysis of peritoneal biopsy specimens. However, this is an invasive procedure, unsuitable for repeated monitoring and prone to sampling error. While the peritoneal equilibrium test provides an indirect measure of peritoneal function, it lacks the sensitivity for early detection of structural fibrotic changes before functional decline becomes evident. Consequently, there is a critical, unmet need for a reliable, non-invasive imaging technique capable of quantitatively evaluating the state of fibroblast activation and fibrotic burden in vivo for the early diagnosis, risk stratification, and management of PF. Fibroblast activation protein (FAP), a type II transmembrane serine protease, is highly expressed on the surface of activated fibroblasts in pathological conditions like fibrosis, but shows minimal expression in most normal adult tissues, making it an ideal target for molecular imaging. This study aimed to comprehensively investigate the potential of a FAP-targeted molecular imaging approach, using the positron emission tomography/computed tomography (PET/CT) probe 68Ga-FAPI-04, for the non-invasive detection and quantitative assessment of PD-associated peritoneal fibrosis.

We employed a multi-faceted and systematic research strategy to thoroughly evaluate the role and expression of FAP in the pathogenesis of PF.

In Vitro Experiments: Cultured human peritoneal mesothelial cells (HPMCs) were stimulated with varying concentrations of transforming growth factor-beta (TGF-β), a key pro-fibrotic cytokine, to mimic the pathological milieu of PD. We quantitatively analyzed TGF-β-induced MMT by assessing changes in the expression of classic markers (e.g., loss of E-cadherin, gain of α-smooth muscle actin [α-SMA]) using techniques such as Western blotting and immunofluorescence staining. Concurrently, we measured the corresponding protein expression levels of FAP to delineate its association with the MMT process.

Transcriptomic Data Analysis: To validate our findings in an in vivo context, we conducted a bioinformatic re-analysis of the publicly available gene expression dataset GSE162029, which contains transcriptomic profiles of mouse peritoneal tissues exposed to high-glucose conditions. We specifically mined this dataset for differential expression of the FAP gene between fibrotic and control groups, providing independent molecular evidence from a separate cohort to support our subsequent in vivo investigations.

Murine Model of PF: A progressive peritoneal fibrosis model was established in mice by daily intraperitoneal injection of 4.25% glucose dialysis solution for durations of 2 to 4 weeks. This model effectively recapitulates the key peritoneal pathological alterations observed in long-term PD patients.

In Vivo Imaging and Correlative Validation: In vivo 68Ga-FAPI-04 PET/CT imaging was performed on the modeled mice and control animals. The maximum standardized uptake value (SUVmax) of the peritoneal membrane was measured to provide a non-invasive, quantitative readout of FAP expression. Following imaging, mice were euthanized, and peritoneal tissues were harvested for extensive ex vivo analysis. This included histological staining with Hematoxylin and Eosin (H&E) to assess submesothelial thickness and Masson's Trichrome to evaluate collagen deposition. Immunohistochemical staining was performed for FAP, Fibronectin (FN), α-SMA, and E-cadherin. Peritoneal transport function was determined via a peritoneal equilibrium test. Finally, robust statistical analyses were conducted to correlate the imaging-derived SUVmax values with histological fibrosis scores, functional parameters (D/D0 glucose), and the expression levels of key molecular markers.

Stimulation with TGF-β successfully induced MMT in HPMCs and significantly upregulated FAP protein expression in a dose-dependent manner. Transcriptomic analysis confirmed a significant upregulation of FAP mRNA in the glucose-exposed murine peritoneum. In the PF mouse model, peritoneal transport function deteriorated over time. Histological examination revealed progressive submesothelial thickening, increased collagen deposition, elevated expression of FN and α-SMA, loss of E-cadherin, and increased FAP expression, all of which correlated with the duration of modeling. Crucially, 68Ga-FAPI-04 PET/CT imaging demonstrated a significantly higher peritoneal SUVmax in PF mice compared to controls. This increased radiotracer uptake showed strong positive correlations with the histological fibrosis score, FAP protein levels, the degree of functional impairment (D/D0 glucose), and the expression of the fibrotic markers FN and α-SMA.

FAP expression is significantly elevated during the development of PD-associated peritoneal fibrosis. 68Ga-FAPI-04 PET/CT enables the non-invasive and quantitative visualization of activated fibroblasts and fibrotic burden within the peritoneum, with its imaging metrics showing strong correlations with established pathological and functional markers of the disease. This positions 68Ga-FAPI-04 PET/CT as a highly promising and potentially transformative diagnostic tool for the early detection and longitudinal monitoring of PF.