Abstract: Mechanism of Lithium Chloride in Inhibiting Invasion of Breast CancerMDA-MB-231 CellsLihongWANG, Qinghua LI, JianWANG,Wei GAO, Ya'ni LIN, Huawen LI,Weina JIN, Guoqiang CHANG, Tianxiang PANGCorrespondence to: Tianxiang PANG, E-mail: pang@ihcams.ac.cnNational Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy ofMedical Sciences and Peking Union Medical College, Tianjin 300020, ChinaGrant support: This study was supported by Natural Science Key Program of Science and Technology Commission Foundation of Tian-jin (No. 08JCZDJC19100 and 09JCZDJC17300)Abstract Objective: To investigate the effect of lithium chloride (LiCl) treatment on the invasion of breast cancer cell lineMDA-MB-231, and its corresponding control on the expression of neutrophil gelatinase associated lipocalin ( NGAL ), a key moleculeresponsible for the regulation of breast tumor cell metastasis. Methods: After LiCl treatment at various concentrations, the cell viabilityof MDA-MB-231 was determined by MTT. Meanwhile, the cell pseudopod formation was observed through inverted optical micro-scope, and changes in the capacity of cell invasion was analyzed by Transwell method, with pretreatment by a layer of Matrigel. The ex-pression of NGAL mRNA and protein was detected by quantitative real time PCR and Western blot, respectively. Finally, small pharma-cological molecules FH535 ( inhibitor for Wnt/β-catenin signaling pathway ) and Bay117082 ( inhibitor for NF-κB signaling pathway )were used to analyze the important signaling pathways involved in the regulation of NGAL expression. Results: The effect of LiCl onthe cell viability of MDA-MB-231 cell line was not statistically significant when the LiCl concentration was lower than 10 mmol/L.Representative pictures from the inverted microscope showed that cell pseudopod formation was inhibited after treatment with 5 mmol/L or 10 mmol/L LiCl for 24h. However, this effect was somewhat reversed by another selective inhibitor Cariporide for Na+/H+ ex-changer 1 ( NHE1 ). The analysis of Transwell indicated that the capacity of cell invasion was apparently inhibited by LiCl, and theNGAL mRNA and protein expression was reduced by the same treatment. LiCl could regulate the NGAL expression after the transcrip-tion. Treatment with inhibitors for Wnt/β-catenin and NF-κB signing pathways for 24h respectively dramatically downregulated the ex-pression of NGAL protein. The results indicated that both of Wnt/β-catenin and NF-κB signaling pathways were involved in regulatingthe expression of NGAL. Conclusion: LiCl can inhibit the pseudopodium formation and invasion of MDA-MB-231 cells by inhibitingNF-κB to regulate NGAL expression.Keywords Lithium chloride ( LiCl ); Breast cancer; MDA-MB-231 cell; Neutrophil gelatinase-associated lipocalin; Invasion