Abstract: Objective To study the change of immune status of umbilical cord mesenchymal stem cells (UCMSC) stimulated by toll like receptor 7 (TLR7) agonist CL264. Methods TLR7 specific ligand CL264 was used to stimulate the UCMSC. Flow cytometry was conducted to assay the expression of co-stimulators 〔human leukocyte antigen（HLA）-E, CD80 and CD86〕and surface markers of stem cells (CD29, CD59 and CD90). Quantitative PCR was applied to measure the expression variation of immune-related molecules and stem cell markers. Cell differentiation experiment was used to study the change of differentiation ability of UCMSC upon CL264 stimulation. Peripheral blood mononuclear cells (PBMC) were isolated from healthy human and then co-cultured with UCMSC in the presence of CL264. Cytotoxicity assay was used to measure the attack of PBMC to UCMSC. Results Expression of co-stimulatory molecules CD86 and HLA-E were enhanced in UCMSC upon CL264 stimulation. Real-time PCR indicated that many pro-inflammatory molecules 〔interleukin (\IL) -1β, (\IL)-6, (\IL)-8, (\IL)-10, interferon (\IFN) -β, \IFN-γ, nuclear factor-κB (\NF-κB), transforming growth factor-β （\TGF-β) 〕 were induced in the presence of CL264 while the expression of stem cells markers were inhibited 〔Kruppel-like factor-4 (\Klf4), Nestin, SRY-related high-mobility-group-box protein-2 (\Sox2), \Lin28〕. Activation of TLR7 also increased the immune attack of PBMC on UCMSC. Our study also indicated that the treatment of CL264 did not influence the differentiation ability of UCMSC. Conclusion TLR7 agonist CL264 could increase the immunogenicity of UCMSC.