Icaritin Inhibits Lung Cancer Through Promoting MiR-370 To Down-regulating PIM1

BACKGROUND: miRNAs play an important role in the development of various tumors.Understanding the relationship between lung cancer and miRNAs will be critical to the study of lung cancer in depth and will provide potential new targets and new methods for the treatment of lung cancer.Icaritin is the main active ingredient of traditional Chinese medicine epimedium.Modern pharmacological studies have confirmed that icaritin has anti-tumor effect.However,the effect of icaritin on lung cancer and its mechanism is still unknown.Methods: The miR-370 level was detected in patients with lung cancer,and the survival time of the patients was followed up.The relationship between miR-370 and PIM1 in lung cancer patients was analyzed.The miR-370 matched protein was predicted by micro RNA database and function analysis software.And the binding of miR-370 to PIM1 protein was confirmed by double luciferase assay.The transcription of mRNA was detected by quantitative PCR and the expression of protein was detected by western-blot in lung cancer cells as well as in normal lung cell.The viability of lung cancer cells was determined by CCK-8 method.The growth of lung cancer cells was detected by clonal formation assay.Annexin V-FITC/PI flow cytometry and the expression of caspase-3were used to detect the apoptosis of the cells.Wound healing was used to detect the migration of the cells and transwell was used to detect the invasive of the cells.MiR-370 lentivirus,miR-370 overexpression plasmid,PIM1 lentivirus and PIM1 overexpression plasmids were constructed.MiR-370 and PIM1 were silenced and overexpressed to analyze the role of miR-370 and PIM1 in the cells.To observe the effect of icaritin on the body weight and tumor volume,nude mice with transplanted lung tumor was raised.The effect of icaritin on the apoptosis in nude mice was detected by tunel method and CAS-3immunohistochemical method.PCR and western-blot were used to detect the expression of miR-370 and PIM1 in nude mice.The effect of icaritin on the viabilty,growth,apoptosis,migration and invasion were also investigated in lung cancer cells.Result: The expression of miR-370 in lung cancer tissue was lower than that innormal lung tissue,and the prognosis of the patients with higher miR-370 expression was much better than with lower miR-370 expression.The expression of miR-370 in lung cancer tissues was negatively related to the expression of PIM1.Micro RNA database and function analysis software showed that miR-370 could match the PIM1 protein,and this result was further confirmed by double fluorescein reporter assay.The expression of miR-370 in normal lung cells was also higher than that in the two lung cancer cells.The miR-370 could inhibit the viability,growth,migration and invasion as well as enhance the apoptosis in the lung cancer cells by down-regulating the transcription and expression of PIM1.Icaritin could enhance the expression of miR-370 and suppress the expression of PIM1 to inhibit the growth of transplanted lung tumor and promote the apoptosis in nude mice.And icaritin could significantly inhibit the viability,growth,migration and invasion as well as promote the apoptosis in lung cancer cells.The effect of icaritin was related to the enhancement of miR-370 and down-regulation of PIM1.Conclusion: miR-370 could down-regulate PIM1 to inhibit lung cancer,and the anti-tumor effect of icaritin was related to the enhancement of miR-370 and down-regulation of PIM1.