Background: Current clinical regimen in vascularized composite allotransplantation (VCA) includes life-long immunosuppression (IS) treatment that temporarily reverses the inevitable occurrence of graft rejection and dysfunction. A hallmark of rejection is graft vasculopathy (GV), characterized by vascular inflammation and intimal hyperplasia. Mesenchymal stem cells (MSC) carry both immunomodulatory and endothelium protective properties. This study aims at attenuating GV with the use of adipose derived- MSC (AD-MSC) to achieve reduced administration of IS.
Methods: Hind limb transplantation was performed from Brown Norway to Lewis rats. Post-transplant IS was stopped after 7 days. Upon grade II rejection (swelling/redness), therapy was initiated as follows. Group1 received IS therapy with Tacrolimus and Dexamethasone (4mg/kg BW) to mimic a clinical situation. AD-MSC were administered systemically or locally in two doses of 1x10^6 cells within three days in Group2 and Group3, respectively. Group4 received saline. Upon 8days of therapy skin and femoral artery samples were stained with EVG stain to determine intimal hyperplasia, vWF as a marker for endothelial cell activation and IgM for activation of the innate immune system.
Results: At endpoint, group1 showed grade0 (1/10) and gradeII (9/10) rejection. Group2 showed gradeII (1/10) and gradeIII (9/10). Group3 had gradeI (3/10) and gradeIII (7/10). All animals in group 4 showed gradeIII rejection. Perivascular IgM deposition in skin was in line with clinical observations and lowest in group1. In the femoral artery this decrease was significant when compared to group2 and 3 (p<0.001) and group4 (p<0.0001). Intimal hyperplasia measured in the skin was also in line with clinical grading. The AD-MSC therapy showed reduced intimal hyperplasia when compared to group4. For skin arterioles >40μm (Group4 vs Group3 (p<0.005)) and in arterioles <40μm (Group4 vs Group3 (p<0.0001) and Group4 vs Group2 (p<0.05). Contrastingly, intimal hyperplasia measured in the larger femoral arteries was highest in group1 vs group2 (p=0.001), group3 (p=0.005), and group4 (p<0.001). This was supported by increased activation of endothelial cells as measured by vWF in Group1 and Group4 when compared to group 2 and 3 (p=0.0152, one-way ANOVA). vWF in skin arterioles >40 μm showed a significant increase in group1 vs group4 (p<0.05) with no differences between the groups for arterioles < 40μm.
Conclusions: The IS treatment in group1 showed no clinical grade rejection but the deeper femoral arteries revealed increased intimal hyperplasia along with endothelial cell activation, probably accounting for recurring episodes of rejection with IS treatment. AD-MSC therapy had the capacity to attenuate acute graft rejection and in addition, might elicit endothelial cell protection. Analyzing other markers for endothelial damage is warranted before AD-MSC can be used to relieve patients from long-term IS regimen and avoid repetitive rejection.