Cerebral ischaemia reperfusion (CIR) affects microRNA (miR) expression and causes substantial inflammation. Here, we investigated the influence and underlying mechanism of miR‐27a‐3p in rats with CIR. First, biliverdin treatment relieved cerebral infarction and decreased the levels of serum interleukin (IL)‐1β, IL‐6, and TNF‐α. Through our previous study, we found key miR‐27a‐3p and its targeted gene LITAF might involve in the molecular mechanism of CIR. Then, the regulation between miR‐27a‐3p and LITAF was verified by the temporal miR‐27a‐3p and LITAF expression profiles and luciferase assay. Moreover, intracerebroventricular injection of the miR‐27a‐3p mimic significantly decreased the LITAF, TLR4, NF‐κB, and IL‐6 levels at 24 h post‐surgery, whereas miR‐27a‐3p inhibitor reversed these effects. Furthermore, miR‐27a‐3p mimic could relieve cerebral infarct and neurologic deficit after CIR. In addition, injection of miR‐27a‐3p mimic decreased neuronal damage induced by CIR. Taken together, our results suggest that miR‐27a‐3p protects against CIR by relieving inflammation, neuronal damage, and neurologic deficit via regulating LITAF and the TLR4/NF‐κB pathway. Intracerebroventricular injection of the miR‐27a‐3p mimic significantly decreased the Litaf, IL‐6, TLR4, and NF‐κB levels and the double‐labelled cell count 24 h post‐surgery, whereas miR‐27a‐3p inhibitor reversed these effects. miR‐27a‐3p mimic could relieve cerebral infarct and neurologic deficit after CIR. Increasing miR‐27‐3p levels protect against CIR by relieving inflammation, neuronal damage, and neurologic deficit by inhibiting LITAF and the TLR4/NF‐κB pathway.