Background Mesenchymal stem cells (MSCs) have been broadly used experimentally in various clinical contexts. The addition of MSCs to initial steroid therapy for acute graft-versus-host disease (aGVHD) may improve patient outcomes. However, investigations regarding prognostic factors affecting the efficacy of MSC therapy for steroid-refractory aGVHD remain controversial. We thus conducted a systematic review and meta-analysis of published clinical trials to determine possible prognostic factors affecting the efficacy of MSCs in treating steroid-refractory aGVHD. Methods and Findings Clinical trials using MSC therapy for steroid-refractory aGVHD were identified by searching PubMed and EMBASE databases. A total of 6,963 citations were reviewed, and 13 studies met the inclusion criteria. A total of 301 patients from thirteen studies were included. Of these, 136 patients showed a complete response (CR), and 69 patients displayed a partial (PR) or mixed response (MR). In total, 205 patients exhibited overall response (ORR). Patients with skin steroid-refractory aGVHD showed a better clinical response than gastrointestinal (CR: odds ratio [OR] = 1.93, 95% confidence interval [95% CI]: 1.05-3.57, p < 0.05) and liver (CR: OR = 2.30, 95% CI: 1.12-4.69, p < 0.05, and ORR: OR = 2.93, 95% CI: 1.06-8.08, p < 0.05) steroid-refractory aGVHD. Those with grade II steroid-refractory aGVHD exhibited a better clinical response following MSC therapy than recipients with grade III-IV (CR: OR = 3.22, 95% CI: 1.24-8.34, p < 0.05). Completion therapy may improve the CR but reduce ORR compared with induction therapy (CR: OR = 0.20, 95% CI: 0.09-0.44, p < 0.05; ORR: OR = 2.18, 95% CI: 1.17-4.05, p = 0.01). There was also a trend towards a better clinical response in children compared with adults (CR: OR = 2.41, 95% CI: 1.01-5.73, p = 0.05). Conclusions Age, skin involvement, lower aGVHD grade, and the number of infusions are the main prognostic factors affecting the efficacy of MSC therapy for steroid-refractory aGVHD.