Objective To study the protective effect of recombinant human erythropoietin (EPO) on brain and to explore the changes in the diversity of intestinal microbial flora in neonatal rats with hypoxic-ischemic encephalopathy (HIE) by establishing a neonatal rat model of HIE, and to provide an experimental basis for clinical application of EPO in the treatment of neonatal HIE. Methods < /b> The HIE model was established in 7-day-old neonatal SD rats. The rats were randomly divided into the HIE model group, EPO-treated group and control group. The changes of nestin expression were detected by immunohistochemistry. Feces of the rats were collected to detect the changes in intestinal microbial flora by 16s rRNA sequencing. Results The expressions of nestin at the same time point in each group were significantly different (P < 0.05). The nestin level in the control group was the lowest, that in the EPO-treated group was the highest, and the HIE model group in between. The Shannon-Wiener index of the HIE model group showed a tendency to decrease compared with the control group. Conclusions Exogenous EPO can promote the growth of neural cells in neonatal rats with HIE, indicating a certain protective effect. Meanwhile, the diversity of intestinal microbial flora of the HIE neonatal rats is also changed.