Objective To determine whether serum deprivation can induce transdifferentiation of cardiac fibroblasts into endothelial cells in the ischemic and hypoxic microenvironment of acute myocardial infarction. Methods Fibroblasts were separated and cultured from hearts of new born C57BL/6 mice, and leukemia inhibitory factor (LIF) was used to accelerate the self-renewal, keep plasticity and inhibit differentiation to the terminal fates. The fibroblasts were exposed to five different intervention conditions:control group (DMEM+10% fetal bovine serum[FBS]) and serum deprivation (DMEM without FBS) for 24 h, 48 h and 72 h groups. qPCR was used to detect the expression of cell lineage specific genes; the in vitro angiogenesis test and Dil-Ac-LDL phagocytic function test were used to observe function of fibroblasts; and ELISA assay was used to examine the level of vascular endothelial growth factor (VEGF) secreted by fibroblasts. Results Compared with the control group, the number of capillaries-like bifurcation formed by fibroblasts was significantly increased at different serum deprivation time points (P<0.05), but no induced engulfment of Dil-Ac-LDL was noticed. Compared with the control group, the expressions of endothelial specific genes CD31 and VE-herin in fibroblasts were significantly increased at 48 h of serum deprivation (P<0.05); the expression of CD31 at 48 h of serum deprivation was 13.7 times that of the control group (P<0.05), and at 72 h of serum deprivation it was two times that of the control group (P<0.05). ELISA results showed that VEGF level at 48 h of serum deprivation was seven times that of the control group, and at 72 h of serum deprivation it was 3.7 times (P<0.05). Conclusion The serum deprivation can stimulate the transdifferentiation of cardiac fibroblasts into endothelial cell lineages.