| Objective To detect the expression of The dual specificity tyrosine-phosphorylation-regulated kinase(DYRK)1B(DYRK1B)in breast cancer tissues,and analyze the correlation between DYRK1 B and the clinical pathological parameters of breast cancer.To investigate the effect of DYRK1 B on the biological behavior of breast cancer cell and its regulation mechanism.Ultimately,define the effect of DYRK1 B on the proliferation and apoptosis of breast cancer.Methods(1)Tissue level: Firstly,the expression of DYRK1 B in eight pairs of breast cancer and adjacent non-tumor tissues were detected by Western Blot.Secondly,immunohistochemistry was used to detect the expression of DYRK1 B and Ki-67 in 120 cases of breast cancer tissue specimens.The correlation between DYRK1 B and the clinicopathological parameters of breast cancer patients was analyzed,including age,tumor size,tissue type,grade,axillary lymph node status,estrogen receptor(ER),progesterone receptor(PR),HER-2,Ki-67 and the other clinicopathological parameters.Finally,analysis of the relationship between DYRK1 B and the prognosis of breast cancer patients was detected by Kaplan-Merier and multivariate analyses.(2)Cellular level: Firstly,western Blot was performed to evaluate the expression of DYRK1 B in normal breast cell and breast cancer cell lines.Secondly,the interference plasmid of DYRK1B(DYRK1B-siRNA)were transfected into breast cancer cells.And detected the influence of DYRK1 B on breast cancer cells proliferation by cell proliferation assay(CCK-8 and clone formation assay)and apoptosis assay by flow cytometer.Finally,the effect of forkhead box O1(FoxO1)on apoptosis of breast cancer cells induced by DYRK1 B was explored by Western Blot,flow cytometry and immunofluorescence.Results(1)Tissue level: Finally,the results showed that the expression of DYRK1 B in breast cancer tissues was significantly higher than that of corresponding adjacent non-tumor tissues by Western Blot.And the protein level of DYRK1 B was consistent with that of PCNA.Secondly,the results of clinical statistics analysis showed that DYRK1 B was positively correlated with tumor size(P = 0.012),histological grade(P = 0.002),ER(P = 0.025)and Ki-67(P < 0.001)of breast cancer patients.Finally,Kaplan-Meier survival analysis revealed that the high expression of DYRK1 B was significantly associated with a decrease in overall survival of breast cancer patients.And multivariate analyses confirmed that DYRK1 B could be used as an independent prognostic indicator for patients’ overall survival(P < 0.05).(2)Cellular level: Finally,western Blot showed that DYRK1 B was over-expressed in breast cancer cell lines,compared to normal breast cells.Secondly,CCK-8,clone formation and flow cytometer indicated that down-regulation of DYRK1 B could inhibit the proliferation and promote apoptosis of breast cancer cells.And the expression of PCNA and Cyclin D1 decreased and the expression of cleaved caspase-3 and cleaved PARP-1(two apoptosis markers)increased after DYRK1 B knockdown.Finally,the expression of phosphorylated FoxO1 was reduced in Dyrk1B-siRNA-treated breast cancer cells.And combined siRNAs of Dyrk1 B with Fox O1 led to fewer apoptotic cells compared to Dyrk1 B siRNA alone in breast cancer by Western blot and flow cytometry analysis.At the same time,immunofluorescent staining showed that knockdown Dyrk1 B induced nuclear accumulation of FoxO1 in breast cancer cells.Conclusions(1)DYRK1B is highly expressed in breast cancer and closely related to the poor prognosis of breast cancer patients.These results suggest that DYRK1 B may be closely related to the development of breast cancer and can be used as one of the prognostic indicators of breast cancer.(2)The depletion of DYRK1 B significantly inhibits proliferation and induces apoptosis in breast cancer cells.DYRK1 B may be a potential therapeutic target for breast cancer.(3)Fox O1 may be involved in Dyrk1B-mediated survival of breast cancer cells. |
PDF Full Text Request