Immunohistomorphometric analysis of transplanted umbilical cord blood-derived mesenchymal stem cells and the resulting anti-inflammatory effects on nerve regeneration of injured canine spinal cord

In order to investigate the beneficial mechanism of transplanted canine umbilical cord blood-derived mesenchymal stem cells (UC-MSCs) on spinal cord injury (SCI), we evaluated recovery in a canine SCI model by examining neurological function and nerve regeneration. Recovery was assessed by clinical observation and by examining regenerated nerve cells by immuno-histomorphometric analysis and by assessing anti-inflammation by measuring mRNA expression of inflammation-related cytokines. Extradural compression of the spinal cord in 21 dogs (4.6±0.4 kg, ∼2-3 yrs) was performed using a balloon catheter for 12 hours. All of the dogs showed hind limb paralysis after compression. Functional recovery of the hind limbs was evaluated by the Olby score and Revised modified tarlov scale. Experimental dogs were divided into cUCB and control groups. In the cUCB group (n=12), UC-MSCs were infected with a lentivirus-vector labeled GFP gene and injected into the SCI site. In the control group (n=9), only PBS was injected into the SCI site. Seven dogs (control = 3, cUCB = 4) were euthanized and their injured spinal cords were collected at 1, 4 and 8 weeks after transplantation. Nerve regeneration was assessed on longitudinal sections at the epicenter. After transplantation of UC-MSCs, functional improvement up to 5 points of the Olby score and 4 points of Revised modified tarlov scale were observed. Compared to the control group, immuno-histomorphometrical analyses showed that cells labeled with GFAP were significantly reduced and cells labeled with Tuj1 and NF160 were increased in the cUCB group (p < 0.05). In western blot analysis, total gliosis in the cUCB group was reduced by 35% compared to controls and surviving nerve cells in the SCI lesion were increased by more than 50 % compared to controls at 8 weeks after transplantation. However, it must be noted that a small number of nerve cells were derived from the transplanted UC-MSCs. In addition, the expression of COX2, IL1, IL6, TNF and TGF-β, in the cUCB group were down regulated. These findings suggest that improvements of neurological function seen after transplantation of UC-MSCs into injured spinal cord might be due primarily to reduced gliosis by anti-inflammation, increased survival of endogenous nerve cells and enhanced function of survived endogenous nerve cells by engrafted cells.