Background <p>Chronic anal fissures in patients with Crohn’s disease (CD) remain a significant therapeutic challenge, particularly when linked to active perianal disease. Conventional treatments often fail, highlighting the need for alternative approaches. This study explores the efficacy and safety of freshly collected autologous adipose tissue injection (AATI) for treating chronic fissures in patients with CD.</p> Methods <p>Nine patients with CD with anal fissures were included. The primary outcome was complete healing (CH) at 3&#xa0;months after last AATI, defined as full fissure re-epithelialization and complete pain relief. Secondary outcomes included changes in defecation pain (visual analog scale [VAS]), anal discomfort (VAS), Perianal Disease Activity Index (PDAI), and St. Mark’s Incontinence Score (SMIS).</p> Results <p>Five patients (56%) achieved CH after one (<i>n</i> = 4) or two (<i>n</i> = 1) AATI. Partial healing was observed in four patients (44%). Defecation pain improved from a VAS score of 7.5 (IQR 5.0–8.5) to 2.75 (0.0–4.5; <i>p</i> = 0.009), anal discomfort from VAS score of median 5.0 (2.5–6.5) to 1.0 (0.0–2.5; <i>p</i> = 0.014), PDAI from 5.0 (3.0–6.0) to 1.0 (1.0–2.0; <i>p</i> = 0.022), and SMIS from 7.0 (4.0–9.0) to 4.0 (0.0–4.0; <i>p</i> = 0.041). No treatment-related complications occurred.</p> Conclusion <p>AATI may be a promising new treatment of chronic anal fissures in patients with CD. Effects of AATI should be explored further in controlled trials.</p>