Hypoxia Imaging in Hepatic Fibrosis: A Preliminary Application of CAIX‐Targeted MRI Probe in Rats

This study aimed to validate the feasibility of hypoxia-targeted magnetic resonance imaging (MRI) molecular probe (CAIX-Gd) for evaluating hypoxia status in rats with hepatic fibrosis. A macrocyclic ligand compound (Compound 3) was synthesized using the carbonic anhydrase IX (CAIX) inhibitor acetazolamide as the targeting moiety, and its structure was confirmed by ¹H-NMR spectroscopy. CAIX-Gd was then prepared by chelating gadolinium, with free gadolinium content quantified via the xylenol orange assay. In biocompatibility tests, CAIX-Gd administration showed no significant histopathological changes compared with saline controls. For hypoxia detection, rats were divided into three groups: (A) hepatic fibrosis (HF) with CAIX-Gd, (B) HF with DTPA-Gd, and (C) normal controls (NC) with CAIX-Gd. The difference in T1 relaxation time (∆T1) pre- and post-contrast was measured in regions of interest (ROI). HF rats showed increased collagen deposition (Masson staining), α-SMA activation, and elevated hypoxia markers (PIMO/HIF-1α) compared to controls (all p < 0.05). In group C, mild ∆T1 shortening was seen at 10 min post-injection. While ∆T1 continuously decreased in group B, the shortening was significantly greater in group A (p < 0.05), peaking at 10 min. In conclusion, the CAIX-Gd probe was successfully synthesized and demonstrated potential for the non-invasive MRI detection of hypoxia in hepatic fibrosis.