n-3 polyunsaturated fatty acid (n-3 PUFA) are considered to be associated with liver regeneration. We investigated the effects of n-3 PUFA on hepatic tight junction (TJs) and liver regeneration after 70% partial hepatectomy (PH) in rats. Male Sprague-Dawley rats were divided into four groups: sham group; control group, fish oil (FO; 1 mL/kg), and the FO (2 mL/kg) group. We examined changes in expression of hepatic TJs by at confocal microscopy in liver regeneration by routine clinical chemistry methods for hepatic function, and in activation of liver kinase B1–adenosine monophosphate–activated protein kinase (AMPK) signaling pathway. Using Western blot analysis. After PH survival was higher in the FO than the control group. We observed treatment with n-3 PUFA to activated the LKB1-AMPK signaling pathway as well as to earlier, stronger and prolonged of the expression of Occludin, Claudin-3, zonula occludens-1, and proliferating cell nuclear antigen proteins. In addition, hepatic TJ structures and the level of liver function were protected after n-3 PUFA treatment. After PH in rats, n-3 PUFA enhanced expression and protected the structure of hepatic TJs via the LKB1-AMPK signaling pathway. Moreover, it may promote liver regeneration partly via the LKB1-AMPK signaling pathway. It protected postoperative hepatic function and may be a liver protective agent against liver failure.