The Investigation Of The Roles And Underlying Mechanism Of Urothelial Carcinoma Associated 1(UCA1) In Lung Cancer

Part I:Study on the expression of UCAl/microRNA-193a/HMGB1 pathway in blood and cancer and adjacent normal tissue samples from clinical lung cancer patientsBackground:Lung cancer is a leading cause of death worldwide.Nowadays,many types of therapy for lung cancer were tried and achieved some advancements,there are still no significant improvements in patients’ outcomes.Thus,we need more uniformly effective and complementary option in the treatment of lung cancer.Long non-coding RNAs have been documented aberrantly expressed and exerted crucial role including affecting the cell growth,cell migration,cell invasion and apoptosis in a variety of cancers by affecting gene expression at various levels including transcription,RNA processing and transport,and translation.Human urothelial carcinoma associated 1(UCA1)is a potential new type of biomarker for tumor diagnosis and exerts oncogenic effect on various human cancers.However,the mechanism of oncogenic role of UCA1 in lung cancer remains unclear.We will study how UCA1 exerts an oncogenic role in tumor proliferation and metastasis and serves as a novel biomarker for diagnosis,therapy and prognosis of cancers.Purpose:Measure the expression levels of UCA1,microRNA-193a and HMGB1 in plasma and tissue samples of lung cancer patients and normal subjects were examined,and their correlation and clinical significance were analyzed.Methods:1)QPCR analysis of the expressions of UCA 1,miR-193a and HMGB 1 in plasma samples from the lung cancer patients and healthy people;2)QPCR analysis of the expressions of UCA 1,miR-193 a and HMGB 1 in cancer tissues and adjacent normal tissues from the lung cancer patients;3)Immunohistochemistry analysis of the expression of HMGB 1 in cancer tissues and adjacent normal tissues from the lung cancer patients;4)Western blots analysis of the expression of HMGB1 in cancer tissues and adjacent normal tissues from the lung cancer patients.Results:1)UCA1 was up-regulated in the plasma samples from the lung cancer patients than healthy people;miR-193a was down-regulated in the plasma samples from the lung cancer patients than healthy people;HMGB1 was up-regulated in the plasma samples from the lung cancer patients than healthy people;2)Increased UCA1 was found in cancer tissues in 14 of 15 patient samples as well as HMGB1 in 13 of 15 patient samples and conversely,decreased miR-193a was found in 13 of 15 patient samples,compared with those in paired normal tissues;3)Increased HMGB1 protein was found in cancer tissues in 13 of 15 patient samples,compared with those in adjacent normal tissues;4)Increased HMGB 1 was found in cancer tissues in 13 of 15 patient samples,compared with those in paired normal tissues.Conclusion:The relationship between the expression levels of the UCA1/microRNA-193a/HMGB1 pathway components may have important clinical application value.Part Ⅱ:The effect and underlying mechanism of UCA1 on the biological behavior of lung cancer cellsBackground:Lung cancer is a leading cause of death worldwide.Long non-coding RNAs have been documented aberrantly expressed and exerted crucial role in variety of cancers.Urothelial carcinoma associated 1(UCA1)is a potential new type of biomarker for tumor diagnosis and exerts oncogenic effects on various human cancers.However,the mechanism of oncogenic role of UCA1 in lung cancer remains unclear.Purpose:1)Study the role of UCA1 in the cell growth and migration of lung cancer cells;2)Answer the expression and regulation of the UCA1/miR-193a/HMGB1 pathway;3)Dissect the underlying mechanism of the role of UCA1 in the cell growth and migration of lung cancer cells.Methods:1)The small interfering RNA specific to UCA1 was transfected into lung cancer cell lines SKMES-1 and H520,then the expression of UCA1,miR-193a and HMGB1 was detected by qPCR assay;the expression of HMGB1 was tested by Western blots;the change of cell viability was measured by CCK8 assay.Migration chamber assay to detect changes in cell migration capacity;2)Dual fluorescein plum reporter gene detects binding between UCA1 and miR-193a,miR-193a and HMGB1;3)Transfection of miR-193a mimics and inhibitors in lung cancer cell lines SKMES-1 and H520,then detected the expression of HMGB1 by qPCR and western blots;CCK8 assay to detect changes in cell viability;migration chamber assay to detect changes in cell migration capacity;4)Lung cancer cell lines SKMES-1 and H520 transfected with small interfering RNA of HMGB 1,then detected the expression of HMGB 1 by qPCR and western blots;CCK8 assay to detect changes in cell viability;migration chamber assay to detect changes in cell migration ability;5)In lung cancer cell lines SKMES-1 and H520 transfection interfered with UCA1/miR-193a/HMGB1 pathway,and then detected HMGB1 expression by qPCR and western blots;CCK8 assay detected cell viability;Cell migration assay changed in cell migration.Results:1)The small interfering RNA specific to UCA1 could repress the UCA1 expression and the suppression of UCA1 inhibited the cell growth and migration of lung cancer cells;2)Dual-luciferase reporter analysis found that there were binding role between the UCA1 and miR-193a,miR-193a and HMGB1;3)The suppression of UCA1 promoted the miR-193a expression and the miR-193a mimics inhibitd the cell growth and migration and the miR-193a inhibitor promoted that of lung cancer cells;4)QPCR and western blots analysis demonstrated that the miR-193a regulated the expression of HMGB1;5)CCK8 and Transwell analysis displayed that miR-193a regulated the expression of HMGB1 and the modulation affected the role of miR-193a.Conclusion:Our study demonstrated that UCA1 exerts oncogenes activity in lung cancer,acting mechanistically by upregulating HMGB 1 expression through ’sponging’ miR-193a.