This work examines whether microglia-conditioned medium (MCM) is beneficial in stressed spinal cord cells or tissues. MCM was separated into two fractions by 50 kDa molecular cut-off centrifugation. MCM not only promoted survival of neuronal and oligodendroglial cells but effectively reduced LPS stimulation in spinal cord cultures. We further utilized the NYU weight-drop device to induce contusive spinal cord injury (SCI) in rats. Immediately after dropping the impactor from a height of 25 mm onto thoracic spinal segment, MCM was intrathecally administered. At 6 weeks post-injury, SCI rats receiving MCM > 50 kDa treatment showed significant hind-limb improvement over MCM < 50 kDa- or vehicle-treated SCI rats. Consistently, more preserved nerve fibers and fewer activated microglia were found in the injured epicenter of MCM-treated SCI rats. Taken together, secreted substances, mainly > 50 kDa, of microglia was neuroprotective against spinal cord injury