Tuina Massage Improves Cognitive Functions of Hypoxic-Ischemic Neonatal Rats by Regulating Genome-Wide DNA Hydroxymethylation Levels

In addition to abnormalities of motor and posture, children with cerebral palsy (CP) often have intellectual disability. As a complementary and alternative traditional Chinese medicine (TCM) therapy, Chinese Tuina massage, also called Tuina in China, has been widely applied in clinical treatment for CP in China for a long time. However, the molecular basis for this still remains largely unknown. Recently, DNA hydroxymethylation has been shown to be sensitive to environment and plays critical roles in some neurological disorders, whereas the research focusing on the relationship between 5 hmC and Tuina therapy for cerebral palsy is deficient. In our study, we first observed that Tuina improved learning and memory functions of hypoxic-ischemic (HI) rat pups. Meanwhile, 5 hmC level of the temporal lobe cortex in the HI neonatal rat model is decreased significantly compared to that of the rats in control and Tuina groups. Then, we used the hMeDIP-Seq method to explore whether and how DNA hydroxymethylation is involved in Tuina therapy for cerebral palsy. Genomic annotation of DhMRs of HI group's hypo-hydroxymethylation to genes revealed enrichment in multiple neurodevelopmental signaling pathways. Moreover, we found the depletion of 5 hmC modifications in genes associated with neuronal development was accompanied by reduced mRNA levels of these genes. Taken together, our results indicate that Tuina may regulate the expression of neurodevelopment-related genes by changing the status of DNA hydroxymethylation, thereby improving learning and memory functions of cerebral palsy.