Background MicroRNAs （miRNAs） contribute to tumorigenesis by acting as either oncogenes or tumor suppressor genes. In this study, we investigated the role of miR-145 in the pathogenesis of uveal melanoma. Methods Ex...Background MicroRNAs （miRNAs） contribute to tumorigenesis by acting as either oncogenes or tumor suppressor genes. In this study, we investigated the role of miR-145 in the pathogenesis of uveal melanoma. Methods Expression profiles of miRNAs in uveal melanoma were performed using Agilent miRNA array. Quantitative real-time polymerase chain reaction was used to screen the expression levels of miR-145 in normal uveal tissue, uveal melanoma tissue, and uveal melanoma cell lines. Lenti-virus expression system was used to construct MUM-2B and OCM-1 cell lines with stable overexpression of miR-145. Cell proliferation, cell cycle, and cell apoptosis of these miR-145 overexpression cell lines were examined by MTT assay and flow cytometry respectively. The target genes of miR-145 were predicted by bioinformatics and confirmed using a luciferase reporter assay. The expression of insulin-like growth factor-1 receptor （IGF-1R）, insulin receptor substrate-1 （IRS-1） proteins was determined by Western blotting analysis. IRS- 1 was knocked down in OCM-1 cells. TUNEL, BrdU, and flow cytometry assay were performed in IRS-1 knocked down OCM-1 cell lines to analyze its function. Results Forty-seven miRNAs were up regulated in uveal melanoma and 61 were down regulated, miR-145 expression was significantly lower in uveal melanoma sample and the cell lines were compared with normal uveal sample. Overexpression of miR-145 suppressed cell proliferation by blocking the G1 phase entering S phase in uveal melanoma cells, and promoted uveal melanoma cell apoptosis. IRS-1 was identified as a potential target of miR-145 by dual luciferase reporter assay. Knocking down of IRS-1 had similar effect as overexpression of miR-145. Conclusion miR-145 might act as a tumor suppressor in uveal melanoma, and downregulation of the target IRS-1 might be a potential mechanism.展开更多
Objective: The present study aimed to investigate circular RNA(circRNA) expression in uveal melanoma(UM).Methods: First,we used microarray to compare the expression profiles of circRNA in five UM samples and five norm...Objective: The present study aimed to investigate circular RNA(circRNA) expression in uveal melanoma(UM).Methods: First,we used microarray to compare the expression profiles of circRNA in five UM samples and five normal uvea tissues.Next,bioinformatics analyses,including gene ontology(GO) analysis and pathway analysis,were applied to study these differentially expressed circRNAs to predict pathogenic pathways that may be involved.Quantitative real-time polymerase chain reaction(qRT-PCR) in 20 UM samples and 20 normal uvea samples was used to confirm the circRNA expression profiles obtained from the microarray data.Finally,we analyzed the interaction between validated circRNAs and their potential cancer-associated miRNA targets.Results: In total,50,579 circRNAs [fold change(FC) ≥2.0; P<0.05],including 20,654 up-regulated and 29,925 down-regulated circRNAs,were identified as differentially expressed between UM tissues and normal uvea tissues.We used qRT-PCR to verify seven dysregulated circRNAs indicated by the microarray data,including hsacirc0119873,hsacirc0128533,hsacirc0047924,hsacirc0103232,hsa-circRNA10628-6,hsacirc0032148 and hsacirc0133460,which may be promising candidates to study future molecular mechanisms.Conclusions: This study explored,for the first time,the abnormal expression of circRNAs in UM and described the expression profile of circRNAs,providing a new potential target for the mechanism of UM and future treatment of UM.展开更多
AIM： To reveal the insight mechanism of liver metastasis in uveal melanoma, we investigated cell functions of microRNA-21 in three different uveal melanoma cell lines and analyze the relationship of target gene p53 a...AIM： To reveal the insight mechanism of liver metastasis in uveal melanoma, we investigated cell functions of microRNA-21 in three different uveal melanoma cell lines and analyze the relationship of target gene p53 and its downstream targets which been found significant expression in our previous study.METHODS： Quantitative real-time polymerase chain reaction （qRT-PCR） was used to detect microRNA-21 expression in normal uveal tissue and uveal melanoma cell lines. Lenti-virus expression system was used to construct OCM-1, MuM-2B and M619 cell line with stable overexpression and inhibition of microRNA-21. In vitro cell function tests such as cell proliferation, cell apoptosis, cell circle and abilities of migration and invasion were examined by MTT, BrdU assay, flow cytometry, transwell assay and Matrigel invasion assay respectively. The target gene was predicted by bioinformatics and confirmed by using a dual luciferase reporter assay. The expression of p53 and its suspected downstream targets LIM and SH3 protein 1 （LASP1） and Glutathione S Transferase pi （GST-Pi） were determined by qRT-PCR in mRNA level and western blotting analysis in protein level. Finally, the effect of microRNA-21 in a xenograft tumor model was assessed in four-week-old BALB/c nude mice. RESULTS： Compared to normal uveal melanoma, expressions of microRNA-21 were significantly higher in uveal melanoma cell lines. Overexpression of microRNA-21 promoted proliferation, migration, and invasion of OCM-1, M619 and MuM-2B cells, while inhibition of microRNA-21 reveal opposite effects. Wild type p53 was identified as a target gene of microRNA-21-3p, and proved by dual luciferase reporter assay. Up-regulated microRNA-21 inhibited the expression of wild type p53 gene, and the increased expression of LASP1 in mRNA level and protein level, while down-regulated microRNA-21 presented opposite way. However, GST-pi showed the potential pattern as expected, but relative mRNA level showed no statistically significant difference in OCM-1 cells.展开更多
AIM: To explore the effect of parthenolide(PTL) on human uveal melanoma(UM) cells(C918 and SP6.5 cells) and its molecular mechanism. METHODS: Carboxyfluorescein succinimidyl amino ester(CFSE) assays and cell counting ...AIM: To explore the effect of parthenolide(PTL) on human uveal melanoma(UM) cells(C918 and SP6.5 cells) and its molecular mechanism. METHODS: Carboxyfluorescein succinimidyl amino ester(CFSE) assays and cell counting kit-8(CCK-8) were performed to detect the cell viability. Flow cytometry was used to analyze cell cycle and apoptosis. Quantitative realtime polymerase chain reaction(qRT-PCR) and Western blot assays were performed to measure proliferation-related and apoptosis-related factors.RESULTS: Firstly, PTL decreased the viability of C918 and SP6.5 cells in a dose-dependent manner, and the effect of PTL on C918 cells was stronger than on SP6.5;however, it did not affect normal cells. Secondly, PTL increased the proportion of cell number at cell cycle G1 phase in C918 cells, and decreased the proportion of cell number at S phase, but the proportion did not change at G2 phase. In addition, PTL induced the apoptosis of C918 cells, and decreased the expressions of Cyclin D1, B-cell lymphoma-2(Bcl-2) and B-cell lymphoma-extra large(Bcl-XL). Also, PTL increased Cyclin inhibition protein 1(P21), Bcl-2-associated X protein(Bax), Cysteinyl aspartate specific proteinas-3(Caspase-3) and Caspase-9 expression. However, the expression of Caspase-8 was not changed. CONCLUSION: PTL inhibites proliferation and induces apoptosis in UM cells by arresting G1 phase and regulating mitochondrial pathway, however, it does not affect normal cells.展开更多
BACKGROUND:Metastatic liver melanoma is a rare event in the Chinese population with extremely poor prognosis.Any treatment that controls a metastatic hepatic lesion potentially prolongs survival.This study aimed to ev...BACKGROUND:Metastatic liver melanoma is a rare event in the Chinese population with extremely poor prognosis.Any treatment that controls a metastatic hepatic lesion potentially prolongs survival.This study aimed to evaluate the survival of patients with isolated liver metastases from uveal melanoma treated with partial hepatectomy or non-surgical management and to find the best therapeutic modality for these patients.METHODS:From January 1996 to September 2008,eight patients with liver metastases secondary to uveal melanoma were admitted to our hospital.Five patients underwent partial hepatectomy and 3 received other treatments(TACE,RFA,PEI).Their medical records were reviewed and overall survival was analyzed.RESULTS:The patients comprised 3 men and 5 women,with a median age of 44 years.Six patients presented with liver metastases at the time the primary tumor was diagnosed.The interval from the diagnosis of uveal melanoma to liver metastasis in the remaining 2 patients was 9.5 and 32.5 months,respectively.The median survival after the treatment of liver metastasis was 11.5 and 7.5 months in the surgical and nonsurgical groups,respectively.There was no procedure-related mortality in the whole study cohort.CONCLUSIONS:Partial hepatectomy or other therapies were safe and feasible for isolated liver metastases from uveal mela-noma.Aggressive treatment with multidisciplinary modalities may result in prolonged survival.展开更多
Objective： The objective of the study was to summarize the role of DNA methylation in the development and metastasis of uveal melanoma （UM）. Data Sources： The relevant studies in MEDLINE were searched. Study Selec...Objective： The objective of the study was to summarize the role of DNA methylation in the development and metastasis of uveal melanoma （UM）. Data Sources： The relevant studies in MEDLINE were searched. Study Selection： In this review, we performed a comprehensive literature search in MEDLINE using ＂uveal melanoma＂ AND （＂DNA methylation＂ OR ＂epigenetics＂） for original research/review articles published before February 2018 on the relationship between DNA methylation and UM. References of the retrieved studies were also examined to search for potentially relevant papers. Results： Previous studies on the relationship between DNA methylation and UM covered many genes including tumor suppressor genes （TSGs）, cyclin-dependent kinase genes, and other genes. Among them, the TSG genes such as RASSFIA and p16INK4a, which encodes a cyclin-dependent kinase inhibitor, are relatively well-studied genes. Specifically, a high percentage of promoter methylation of RASSF1A was observed in UM cell lines and/or patients with UM. Promoter methylation of RASSFIA was also associated with the development of metastasis. Similarly, a high percentage of promoter hypernlethylation of p16INK4a was found in UM cell lines. DNA promoter methylation can control the expression of p16INK4a, which affect cell growth, migration, and invasion in UM. Many other genes might also be involved in the pathogenesis of UM such as tile Ras and EF-hand domain containing （RASEF） gene, RAB31, hTERT, embryonal fyn-associated substrate, and deleted in split-hand/split-foot 1. Conclusions： Our review reveals the complex mechanisms underlying the tumorigenesis of UM and highlights the great needs of future studies to discover more genes/5＇-C-phosphate-G-3＇ sites contributing to the development/metastasis of U M and explore the mechanisms through which epigenetic changes exert their function in UM.展开更多
基金This study was supported by grants from the National Natural Science Foundation of China （No. 81272981） and Natural Sciences Fundation of Beijing, China （No. 7112031）.
文摘Background MicroRNAs （miRNAs） contribute to tumorigenesis by acting as either oncogenes or tumor suppressor genes. In this study, we investigated the role of miR-145 in the pathogenesis of uveal melanoma. Methods Expression profiles of miRNAs in uveal melanoma were performed using Agilent miRNA array. Quantitative real-time polymerase chain reaction was used to screen the expression levels of miR-145 in normal uveal tissue, uveal melanoma tissue, and uveal melanoma cell lines. Lenti-virus expression system was used to construct MUM-2B and OCM-1 cell lines with stable overexpression of miR-145. Cell proliferation, cell cycle, and cell apoptosis of these miR-145 overexpression cell lines were examined by MTT assay and flow cytometry respectively. The target genes of miR-145 were predicted by bioinformatics and confirmed using a luciferase reporter assay. The expression of insulin-like growth factor-1 receptor （IGF-1R）, insulin receptor substrate-1 （IRS-1） proteins was determined by Western blotting analysis. IRS- 1 was knocked down in OCM-1 cells. TUNEL, BrdU, and flow cytometry assay were performed in IRS-1 knocked down OCM-1 cell lines to analyze its function. Results Forty-seven miRNAs were up regulated in uveal melanoma and 61 were down regulated, miR-145 expression was significantly lower in uveal melanoma sample and the cell lines were compared with normal uveal sample. Overexpression of miR-145 suppressed cell proliferation by blocking the G1 phase entering S phase in uveal melanoma cells, and promoted uveal melanoma cell apoptosis. IRS-1 was identified as a potential target of miR-145 by dual luciferase reporter assay. Knocking down of IRS-1 had similar effect as overexpression of miR-145. Conclusion miR-145 might act as a tumor suppressor in uveal melanoma, and downregulation of the target IRS-1 might be a potential mechanism.
基金supported by Beijing Municipal Administration of Hospitals’ Ascent Plan(No.DFL20150201)the National Natural Science Foundation of China(No.81570891)+2 种基金Beijing Natural Science Foundation(No.7151003)Advanced Health Care Professionals Development Project of Beijing Municipal Health Bureau(No.2014-2-003)The Capital Health Research and Development of Special(No.2016-1-2051)
文摘Objective: The present study aimed to investigate circular RNA(circRNA) expression in uveal melanoma(UM).Methods: First,we used microarray to compare the expression profiles of circRNA in five UM samples and five normal uvea tissues.Next,bioinformatics analyses,including gene ontology(GO) analysis and pathway analysis,were applied to study these differentially expressed circRNAs to predict pathogenic pathways that may be involved.Quantitative real-time polymerase chain reaction(qRT-PCR) in 20 UM samples and 20 normal uvea samples was used to confirm the circRNA expression profiles obtained from the microarray data.Finally,we analyzed the interaction between validated circRNAs and their potential cancer-associated miRNA targets.Results: In total,50,579 circRNAs [fold change(FC) ≥2.0; P<0.05],including 20,654 up-regulated and 29,925 down-regulated circRNAs,were identified as differentially expressed between UM tissues and normal uvea tissues.We used qRT-PCR to verify seven dysregulated circRNAs indicated by the microarray data,including hsacirc0119873,hsacirc0128533,hsacirc0047924,hsacirc0103232,hsa-circRNA10628-6,hsacirc0032148 and hsacirc0133460,which may be promising candidates to study future molecular mechanisms.Conclusions: This study explored,for the first time,the abnormal expression of circRNAs in UM and described the expression profile of circRNAs,providing a new potential target for the mechanism of UM and future treatment of UM.
基金Supported by the National Natural Science Foundation of China (No.81570891 No.81272981)+3 种基金Beijing Natural Science Foundation (No.7151003)Advanced Health Care Professionals Development Project of Beijing Municipal Health Bureau (No.2014-2-003)the Capital Health Research and Development of Special (No.2016-1-2051)Hospitals’ Ascent Plan (No.DFL20150201)
文摘AIM： To reveal the insight mechanism of liver metastasis in uveal melanoma, we investigated cell functions of microRNA-21 in three different uveal melanoma cell lines and analyze the relationship of target gene p53 and its downstream targets which been found significant expression in our previous study.METHODS： Quantitative real-time polymerase chain reaction （qRT-PCR） was used to detect microRNA-21 expression in normal uveal tissue and uveal melanoma cell lines. Lenti-virus expression system was used to construct OCM-1, MuM-2B and M619 cell line with stable overexpression and inhibition of microRNA-21. In vitro cell function tests such as cell proliferation, cell apoptosis, cell circle and abilities of migration and invasion were examined by MTT, BrdU assay, flow cytometry, transwell assay and Matrigel invasion assay respectively. The target gene was predicted by bioinformatics and confirmed by using a dual luciferase reporter assay. The expression of p53 and its suspected downstream targets LIM and SH3 protein 1 （LASP1） and Glutathione S Transferase pi （GST-Pi） were determined by qRT-PCR in mRNA level and western blotting analysis in protein level. Finally, the effect of microRNA-21 in a xenograft tumor model was assessed in four-week-old BALB/c nude mice. RESULTS： Compared to normal uveal melanoma, expressions of microRNA-21 were significantly higher in uveal melanoma cell lines. Overexpression of microRNA-21 promoted proliferation, migration, and invasion of OCM-1, M619 and MuM-2B cells, while inhibition of microRNA-21 reveal opposite effects. Wild type p53 was identified as a target gene of microRNA-21-3p, and proved by dual luciferase reporter assay. Up-regulated microRNA-21 inhibited the expression of wild type p53 gene, and the increased expression of LASP1 in mRNA level and protein level, while down-regulated microRNA-21 presented opposite way. However, GST-pi showed the potential pattern as expected, but relative mRNA level showed no statistically significant difference in OCM-1 cells.
基金Supported by the Health Special Project of Jilin Province Department of Finance (No.3D5177883429)
文摘AIM: To explore the effect of parthenolide(PTL) on human uveal melanoma(UM) cells(C918 and SP6.5 cells) and its molecular mechanism. METHODS: Carboxyfluorescein succinimidyl amino ester(CFSE) assays and cell counting kit-8(CCK-8) were performed to detect the cell viability. Flow cytometry was used to analyze cell cycle and apoptosis. Quantitative realtime polymerase chain reaction(qRT-PCR) and Western blot assays were performed to measure proliferation-related and apoptosis-related factors.RESULTS: Firstly, PTL decreased the viability of C918 and SP6.5 cells in a dose-dependent manner, and the effect of PTL on C918 cells was stronger than on SP6.5;however, it did not affect normal cells. Secondly, PTL increased the proportion of cell number at cell cycle G1 phase in C918 cells, and decreased the proportion of cell number at S phase, but the proportion did not change at G2 phase. In addition, PTL induced the apoptosis of C918 cells, and decreased the expressions of Cyclin D1, B-cell lymphoma-2(Bcl-2) and B-cell lymphoma-extra large(Bcl-XL). Also, PTL increased Cyclin inhibition protein 1(P21), Bcl-2-associated X protein(Bax), Cysteinyl aspartate specific proteinas-3(Caspase-3) and Caspase-9 expression. However, the expression of Caspase-8 was not changed. CONCLUSION: PTL inhibites proliferation and induces apoptosis in UM cells by arresting G1 phase and regulating mitochondrial pathway, however, it does not affect normal cells.
基金supported by a grant from the Chinese Key Project for Infectious Diseases(2008ZX10002-025)
文摘BACKGROUND:Metastatic liver melanoma is a rare event in the Chinese population with extremely poor prognosis.Any treatment that controls a metastatic hepatic lesion potentially prolongs survival.This study aimed to evaluate the survival of patients with isolated liver metastases from uveal melanoma treated with partial hepatectomy or non-surgical management and to find the best therapeutic modality for these patients.METHODS:From January 1996 to September 2008,eight patients with liver metastases secondary to uveal melanoma were admitted to our hospital.Five patients underwent partial hepatectomy and 3 received other treatments(TACE,RFA,PEI).Their medical records were reviewed and overall survival was analyzed.RESULTS:The patients comprised 3 men and 5 women,with a median age of 44 years.Six patients presented with liver metastases at the time the primary tumor was diagnosed.The interval from the diagnosis of uveal melanoma to liver metastasis in the remaining 2 patients was 9.5 and 32.5 months,respectively.The median survival after the treatment of liver metastasis was 11.5 and 7.5 months in the surgical and nonsurgical groups,respectively.There was no procedure-related mortality in the whole study cohort.CONCLUSIONS:Partial hepatectomy or other therapies were safe and feasible for isolated liver metastases from uveal mela-noma.Aggressive treatment with multidisciplinary modalities may result in prolonged survival.
文摘Objective： The objective of the study was to summarize the role of DNA methylation in the development and metastasis of uveal melanoma （UM）. Data Sources： The relevant studies in MEDLINE were searched. Study Selection： In this review, we performed a comprehensive literature search in MEDLINE using ＂uveal melanoma＂ AND （＂DNA methylation＂ OR ＂epigenetics＂） for original research/review articles published before February 2018 on the relationship between DNA methylation and UM. References of the retrieved studies were also examined to search for potentially relevant papers. Results： Previous studies on the relationship between DNA methylation and UM covered many genes including tumor suppressor genes （TSGs）, cyclin-dependent kinase genes, and other genes. Among them, the TSG genes such as RASSFIA and p16INK4a, which encodes a cyclin-dependent kinase inhibitor, are relatively well-studied genes. Specifically, a high percentage of promoter methylation of RASSF1A was observed in UM cell lines and/or patients with UM. Promoter methylation of RASSFIA was also associated with the development of metastasis. Similarly, a high percentage of promoter hypernlethylation of p16INK4a was found in UM cell lines. DNA promoter methylation can control the expression of p16INK4a, which affect cell growth, migration, and invasion in UM. Many other genes might also be involved in the pathogenesis of UM such as tile Ras and EF-hand domain containing （RASEF） gene, RAB31, hTERT, embryonal fyn-associated substrate, and deleted in split-hand/split-foot 1. Conclusions： Our review reveals the complex mechanisms underlying the tumorigenesis of UM and highlights the great needs of future studies to discover more genes/5＇-C-phosphate-G-3＇ sites contributing to the development/metastasis of U M and explore the mechanisms through which epigenetic changes exert their function in UM.
