The farnesoid X receptor (FXR) plays an important role in bile acid metabolism, intestinal homeostasis, and intestinal ischemia–reperfusion (I/R) injury. We aimed to clarify the potential effects of FXR on intestinal epithelial cell tolerance to intestinal I/R injury and reveal the underlying mechanisms. An intestinal I/R injury model was established by the occlusion of the superior mesenteric artery for ischemia for 1 h, followed by reperfusion for 4 h in C57BL/6 (wild type [WT]) and FXR−/− mice. The small intestine injury was assessed by histological analysis. Diamine oxidase and TNF-α levels in the serum were measured. Expressions of Bcl-2, Bax, caspase-3, and cystathionine-γ-lyase (CSE) were determined by immunohostochemical staining. Oxygen–glucose deprivation/reperfusion (OGD/R) was used to make injury in cultured Caco-2 cells pretreated with FXR agonist (INT-747) or DL-propargylglycine (PAG) for 24 h. Cell viability and the expressions of NF-κB, TNF-α, and IL-6 were assessed. Compared with WT I/R mice, FXR knockout mice exacerbated intestinal I/R injury, intestinal epithelial apoptosis, and inflammatory response. The I/R injury in WT mice was alleviated with INT-747 pretreatment. CSE expression increased after intestinal I/R injury in WT but not in FXR−/− mice. INT-747 enhanced Caco-2 cell viability and inhibited inflammatory response by blocking the NF-κB pathway after OGD/R injury, which was diminished by a CSE-specific inhibitor (PAG). Thus, we demonstrated that FXR activation enhances intestinal epithelial cell tolerance to I/R by suppressing the inflammatory response and NF-κB pathway via CSE mediation.