Using the Quantum Cell Expansion System to Generate Mesenchymal Stromal Cells for Neuroprotection After Cardiopulmonary Bypass

Background & Aim Cardiopulmonary bypass (CPB) has been known to cause substantial systemic inflammation and trigger prolonged microglial activation in the brain. Mesenchymal stromal cells (MSCs) have significant immunomodulatory properties; and we hypothesize that MSCs would be neuroprotective for newborn babies after CPB. However, the number of MSCs required in the pre-clinical and clinical models is in excess of 20 billion. Therefore we tested whether the Quantum could generate sufficient cells for preclinical piglet models and whether the cells were safe and efficacious in vivo. Methods, Results & Conclusion Passage 5-6 MSCs were manufactured in the Quantum using a D5 media containing platelet lysate; and phenotype, trilineage potential, viability, and cell yield were assessed for each lot. To test for safety and efficacy, two-week old piglets (n=16 total) were randomly assigned to one of 3 groups: (1) Control, (2) Cardiopulmonary bypass (CPB), (3) CPB followed by MSC administration. In group 3, MSCs (10 × 10^6 per kg) were delivered through CPB during rewarming period. The brains were fixed three hours after CPB for immunohistochemistry. Microglia was identified with specific antibody to Iba1 in both cortex and white matter. Caspase activation was also evaluated in the cortex. Starting from a mean seeding amount of 17.1 × 10^6 (range: 12.5-20 × 10^6), D5-Platelet lysate media manufactured a mean 4.70 × 10^8 MSCS (range: 3.3-7.5 × 10^8) between 10-12 days at passage 4 or 5 using the Terumo Research-grade expansion set (1.7 sq meters of surface area). The mean viability of MSCs was 92.5% (range: 89.5-92.7%) and all MSCs had >95% CD73, CD90, and CD105 expression and demonstrated trilineage potential. When transferred into the piglet model, immunohistochemistry revealed that MSC treatment normalized microglia expansion and activation caused by CPB to the same levels as the no CPB control. Acute caspase activations in the cortex after CPB were also mitigated by MSC treatment. In conclusion, MSC delivery was safe in an in vivo piglet model during CPB and has the potential to reduce microglial expansion and caspase activation in children undergoing CPB. Further investigation is necessary to determine the effect of MSC delivery through CPB during pediatric cardiac surgery.