Modulation of adhesion molecules expression on the surface of cord blood (CB) CD34(+) cells may assist in overcoming the delay in cord blood engraftment. Likewise, utilization of diverse growth factors such as neuropeptides could also be helpful. Therefore, we aimed to assess the role of Substance P (SP) along with a cytokine cocktail on CB CD34(+) adhesion molecule expression. CB CD34(+) cells were cultured in a serum-free media containing different concentrations of SP in combination with a cytokine cocktail (SCF, FL, TPO, IL-3, and IL-6). Expression of adhesion molecules CXCR4, CD44, CD49e, and CD62L was analyzed after 7 and/or 11 days of cell cultivation. Additionally, the colonogenic capacity of cells was analyzed by colony formation unit assay. Our results show an enhanced percentage of CD34(+)cells with CXCR4, CD44, and CD62L on day 7, as compared with control. Furthermore, an increase in frequency was observed for CD49e(+) CD34(+)cells by day 7 in both test and control groups compared with day 0. Colonogenic assays show occurrence of more total colony formation and immature progenitor cells in SP-treated cells. Our study indicates that SP could act as an effective modulator for expression of cell adhesion molecules.