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Little is known regarding whether the NFE2/miR-423–5p and FAM3A-ATP-Akt pathway in liver mediates exercise allured alleviation of insulin resistance connected with diet-induced obesity. This research inquired the influence of exercise on liver insulin sensitivity and whole body insulin resistance in high-fat diet fed rats.Forty male Sprague-Dawley rats at seven-week-old were assigned to four groups at random: standard diet as normal control group (NC, n=10), high-fat diet group (HFD, n=10), high-fat diet with chronic exercise intervention group (HFD-CE, n=10) and high-fat diet with acute exercise intervention group (HFD-AE, n=10).Compared with rats fed with a standard diet, eight-week high-fat diet feeding lead to elevated body weight, visceral fat content and serum FFAs, and decreased insulin sensitivity index. Moreover, high-fat diet enhanced NFE2 protein expression and miR-423–5p level, decreased FAM3A mRNA and protein expression, ATP level and Akt phosphorylation in liver. In contrast, physical exercise, both chronic and acute exercise alleviated whole body insulin resistance, reduced hepatic NFE2 and miR-423–5p expression, and serum FFAs level, meanwhile enhanced FAM3A mRNA and protein expression, ATP level and Akt phosphorylation in liver. The current findings indicated that exercise in diet-induced obesity, both chronic and acute, induce a momentous regulation in NFE2/miR-423–5p and FAM3A-ATP-Akt pathway in liver, and improve hepatic insulin sensitivity and whole body insulin resistance.All these results supply crucial evidence in our comprehending of the molecular mechanism that connected exercise to an alleviation of insulin resistance.