Abstract: PURPOSE: CD133^+/- cells were isolated and purified from primary oral squamous cell carcinoma(OSCC) to explore the effects of different culture conditions on the maintenance and biological characteristics of CD133⁺ primary OSCC. METHODS: CCK-8 was used to detect the ability of proliferation and cisplatin resistance between CD133^+/- cell subsets. Transwell assay was used to compare the invasive ability of two cell subsets under the action of cisplatin. Flow cytometry was used to detect the proportion of CD133⁺ cells cultured by serum free medium(SFM) (with or without leukemia inhibitory factor, LIF) or serum supplied medium (SSM). Subcutaneous tumor model in nude mice was used to verify the difference in tumorigenicity of CD133^+/- cell subsets. The transplanted tumor was removed for H-E staining and immunohistochemistry (IHC). SPSS 25.0 software package was used for statistical analysis. RESULTS: Compared with CD133^- cell subsets, CD133⁺ cell subsets had stronger ability of proliferation in vitro(P<0.05) and cisplatin tolerance(P<0.001). Cisplatin had a stronger effect on the invasive ability of CD133^- cell subsets than CD133⁺ cell subsets (P<0.01). No significant difference in the proportion of CD133⁺ cell between LIF-SFM and no-LIF-SFM was found (P>0.05); but compared with SSM culture method, SFM culture method could maintain the proportion of CD133⁺ cell better(P<0.05). CD133⁺ cell subsets showed stronger tumorigenic ability with fewer cells than CD133^- cell subsets in nude mice(P<0.05). CONCLUSIONS: Serum free culture method can better maintain the characteristics of primary OSCC stem cells, but the addition of LIF has no significant effect on the maintenance of stemness of primary OSCC cells.

Key words: Serum free medium method, Leukemia inhibitory factor, Carcinoma stem cells, CD133, Oral squamous cell carcinoma