Fibronectin containing extra domain A (EDA(+) FN), a functional glycoprotein participating in several cellular processes, correlates with chronic liver disease. Herein, we aim to investigate the expression and secretion of EDA(+) FN from hepatocytes in nonalcoholic fatty liver disease (NAFLD) and the underlying mechanisms. Circulating levels of EDA(+) FN were determined by ELISA in clinical samples. Western blotting and flow cytometry were performed on L02 and HepG2 cell lines to analyze whether the levels of EDA(+) FN were associated with endoplasmic reticulum (ER) stress-related cell death. Circulating levels of EDA(+) FN in NAFLD patients were significantly higher than those in control subjects, and positively related with severity of ultrasonographic steatosis score. In cultured hepatocytes, palmitate up-regulated the expression of EDA(+) FN in a dose-dependent manner. Conversely, when the cells were pretreated with 4-phenylbutyrate, a specific inhibitor of ER stress, up-regulation of EDA(+) FN could be abrogated. Moreover, silencing CHOP by shRNA enhanced the release of EDA(+) FN from hepatocytes following palmitate treatment, which was involved in ER stress-related cell damage. These findings suggest that the up-regulated level of EDA(+) FN is associated with liver damage in NAFLD, and ER stress-mediated cell damage contributes to the release of EDA(+) FN from hepatocytes.