Abstract Background and Aims The long-term safety and efficacy of allogeneic bone marrow-derived mesenchymal stromal cell bmMSC therapy in perianal Crohn’s disease CD fistulas is unknown. We aimed to provide a 4-year clinical evaluation of allogeneic bmMSC treatment of perianal CD fistulas. Methods A double-blind dose-finding study for local bmMSC therapy in 21 patients with refractory perianal fistulising Crohn’s disease was performed at the Leiden University Medical Center in 2012–2014. All patients treated with bmMSCs 1 x 107 bmMSCs cohort 1, n = 5; 3 × 107 bmMSCs cohort 2, n = 5; 9 × 107 bmMSCs cohort 3, n = 5 were invited for a 4-year evaluation. Clinical events were registered, fistula closure was evaluated, and anti-human leukocyte antigen HLA antibodies were assessed. Patients were also asked to undergo a pelvic magnetic resonance imaging MRI and rectoscopy. Results Thirteen out of 15 patients 87% treated with bmMSCs were available for long-term follow-up. Two non-MSC related malignancies were observed. No serious adverse events thought to be related to bmMSC therapy were found. In cohort 2 n = 4, all fistulas were closed 4 years after bmMSC therapy. In cohort 1 n = 4 63%, and in cohort 3 n = 5 43%, of the fistulas were closed, respectively. In none of the patients anti-HLA antibodies could be detected 24 weeks and 4 years after therapy. Pelvic MRI showed significantly smaller fistula tracts after 4 years. Conclusions Allogeneic bmMSC therapy for CD-associated perianal fistulas is also in the long-term a safe therapy. In bmMSC-treated patients, fistulas with closure at Week 24 were still closed after 4 years.