OBJECTIVE Several studies in animal models have shown the safety and effectiveness of mesenchymal stem cell conditioned medium (MSC-CM) in inflammatory lesions involving muscles and joints. METHODS In this report, we retrospectively evaluated 16 patients who received local administration of the human adipose-derived mesenchymal stem cells conditioned medium (hAMSC-CM) for musculoskeletal chronic pain. Overall, 27 body locations expressing pain have been treated. The local administrated dose was 5 ml in the joint cavity and/or 2 ml in the other locations. The patients were asked to conduct self-evaluation of the degree of pain using a numeric rating scale (NRS) questionnaire and record the severity of pain before administration and at 15 min, 1 day, 1 week, and 4 weeks after administration. A second administration has been performed in 7 locations. The analysis was done considering two conditions: the "current pain status" and the "worst pain status in a week." RESULTS The results showed statistically significant differences between before and after administration at each time point for "current pain status" and at 1-week and 4-week time points for "worst pain status in a week" after first administration (Tukey-Kramer test). After second administration, significant differences were found at 1-week and 4-week time points for "current pain status". No serious adverse effect was found. CONCLUSIONS It was concluded that local administration of hAMSC-CM appears to be safe and could be expected to have effective therapeutic value against musculoskeletal chronic pain. Further studies are needed to clarify analgesic effects of hAMSC-CM and its underlying mechanism(s).