The influence of Wharton's jelly-derived mesenchymal stromal cells on T regulatory cells in patients with autism spectrum disorder

Increasing evidence supports the role of immune system in autism spectrum disorders (ASD). A substantial number of individuals was found to have altered immune response or dysregulated immunophenotype including systemic deficit of T regulatory cells (Tregs), which are known as modulators of immune homeostasis. Mesenchymal stromal cells (MSC) could potentially exert their immunomodulatory effects by skewing lymphocytes towards Tregs. After approval of local bioethical committee, we have obtained peripheral blood from 23 ASD children and 12 typically developing (TD) controls aged between 2 and 6 years. All children were examined by a trained psychologist with Autism Spectrum Disorder Rating Scale (ASRS) either to rule out developmental disabilities or confirm ASD diagnosis according to DSM-V criteria and assess intensity of autistic traits. Wharton's jelly-derived MSC (WJ-MSC) from third party donors were seeded on a 6-well plates 24 hours prior to blood sampling. Peripheral blood mononuclear cells (PBMNC) were separated using density gradient centrifugation. A co-culture model was established based on a direct cell-to-cell interactions or paracrine mechanism with the use of cell culture inserts. Tregs were defined as CD4+CD25+CD127low and immunolabelled with adequate antibodies at the starting point and subsequently at 24 and 72 hours of culture. There were no differences in Treg number and percentge between ASD children and TD controls. Treg percentage was significantly different during culture in both study (p < 0.0001) and control groups (p = 0.03). Post-hoc analysis in study group revealed significant increase of Tregs in insert culture system after 24 (p = 0.0047, median 8.35%, 95%CI 7.6–10.77%) and 72 hours (p = 0.0008, median 8.96%, 95%CI 7.08–10.83%) of culture in comparison to PBMNC without WJ-MSC (24 hours: median 7.34, 95%CI 6.41–8.27%; 48 hours: median 5.88, 95%CI 4.89–7.39%). Post-hoc analysis in control group revealed no differences, which could be attributed to small sample size. Psychological studies revealed that Treg percentage correlated negatively with attention deficits (p = 0.04, cor = −0.35) and sensory impairment (p = 0.05, cor = −0.33) and with social relationships (p = 0.06, cor = −0.33) on the verge of statistical significance. The diagnosis of ASD did not influence results of co-culture studies. The up-regulation of Treg percentage during culture with WJ-MSC and cell inserts suggests a major role of paracrine mechanisms in Treg induction.