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1.
Braz. j. med. biol. res ; 54(2): e10394, 2021. tab, graf
Article in English | LILACS | ID: biblio-1153512

ABSTRACT

MicroRNAs (miRNAs) have been indicated to be frequently dysregulated in various cancers and promising biomarkers for colon cancer. The present study aimed to assess the prognostic significance and biological function of miR-1273a in colon cancer. The expression levels of miR-1273a was estimated using quantitative real-time polymerase chain reaction. Kaplan-Meier survival curves and Cox regression analysis were used to evaluate the prognostic value of miR-1273a in patients of colon cancer. The effects of miR-1273a on cell proliferation, migration, and invasion were investigated by cell experiments. The expression of miR-1273a was downregulated in colon cancer tissues and tumor cell lines compared with the normal controls (all P<0.001). The aberrant expression of miR-1273a was associated with vascular invasion (P=0.005), differentiation (P=0.023), lymph node metastasis (P=0.021), and TNM stage (P=0.004). The patients with low miR-1273a expression had low overall survival compared with the patients with high miR-1273a expression (log-rank P=0.002). miR-1273a was detected to be an independent prognostic biomarker for patients. Furthermore, the results of cell experiments revealed that miR-1273a downregulation promoted, while miR-1273a upregulation suppressed the cell proliferation, migration, and invasion. In conclusion, all data indicated that a downregulated expression of miR-1273a predicted poor prognosis for colon cancer and enhanced tumor cell proliferation, migration, and invasion. Thus, we suggest that methods to promote miR-1273a expression may serve as novel therapeutic strategies in colon cancer.


Subject(s)
Humans , Male , Female , Middle Aged , Colonic Neoplasms/diagnosis , MicroRNAs/genetics , Biomarkers, Tumor/genetics , Cell Movement/genetics , Colonic Neoplasms/genetics , Cell Proliferation/genetics , Neoplasm Invasiveness
2.
Braz. j. med. biol. res ; 54(8): e10940, 2021. graf
Article in English | LILACS | ID: biblio-1285675

ABSTRACT

Recently, an increasing number of studies have reported that dysregulation of circular RNA (circRNA) expression plays critical roles in the progression of several cancers, including colorectal cancer (CRC). However, the detailed molecular mechanisms of circRNAs involvement in CRC remain largely unknown. Here, we confirmed that the level of circEGFR was significantly increased in CRC tissues compared to matched adjacent non-tumor tissues, and a high level of circEGFR was correlated with poor clinicopathological characteristics and poor prognosis in patients with CRC. Moreover, increased circEGFR expression promoted CRC cell proliferation, migration, and invasion in vitro. Mechanistically, circEGFR acted as a ceRNA for miR-106a-5p to relieve the repressive effect of miR-106a-5p on DDX5 mRNA. Moreover, circEGFR enhanced DDX5 expression, thereby upregulating p-AKT levels. Together, these findings showed that circEGFR promoted CRC cell proliferation, migration, and invasion through the miR-106a-5p/DDX5/AKT axis, and may serve as a promising diagnostic marker and therapeutic target for CRC patients.


Subject(s)
Humans , Colorectal Neoplasms/genetics , MicroRNAs/genetics , Gene Expression Regulation, Neoplastic , Cell Movement/genetics , Cell Line, Tumor , Cell Proliferation/genetics , DEAD-box RNA Helicases , RNA, Circular
3.
Braz. j. med. biol. res ; 54(6): e10474, 2021. tab, graf
Article in English | LILACS | ID: biblio-1285667

ABSTRACT

Osteosarcoma is a highly malignant tumor that occurs in the bone. Previous studies have shown that multiple microRNAs (miRNAs) regulate the development of osteosarcoma. This study aimed to explore the role of miR-629-5p and its target gene, caveolin 1 (CAV1), in osteosarcoma development. To analyze the expression of miR-629-5p and CAV1 mRNA in osteosarcoma tissues and cell lines, qRT-PCR analysis was performed. Dual-luciferase reporter experiments were subsequently performed to validate the relationship between CAV1 and miR-629-5p. CCK8 assay was used to measure osteosarcoma cell proliferation, and wound-healing assay was performed to study their migratory phenotype. Our findings revealed that miR-629-5p was overexpressed in osteosarcoma tissues and cells, and thereby enhanced cell proliferation and migration. Further, we validated that miR-629-5p targets CAV1 mRNA directly. CAV1 expression, which was negatively correlated with miR-629-5p expression, was found to be downregulated in osteosarcoma tissue samples. Moreover, our data showed that an increase in CAV1 level led to a decline in osteosarcoma cell proliferation and migration, which could be rescued by miR-629-5p upregulation. Overall, our study confirmed that miR-629-5p promoted osteosarcoma proliferation and migration by directly inhibiting CAV1.


Subject(s)
Humans , Bone Neoplasms/genetics , Osteosarcoma/genetics , MicroRNAs/genetics , Gene Expression Regulation, Neoplastic , Cell Movement/genetics , Cell Line, Tumor , Cell Proliferation/genetics , Caveolin 1/genetics
4.
Braz. j. med. biol. res ; 54(4): e10117, 2021. tab, graf
Article in English | LILACS | ID: biblio-1153531

ABSTRACT

The long noncoding RNA (lncRNA) H19 is involved in the pathogenesis of endometriosis by modulating the proliferation and invasion of ectopic endometrial cells in vitro, but related in vivo studies are rare. This study aimed to investigate the role of lncRNA H19 in a nude mouse model of endometriosis. Ectopic endometrial stromal cells (ecESCs) were isolated from ectopic endometrium of patients with endometriosis and infected with lentiviruses expressing short hairpin RNA (shRNA) negative control (LV-NC-shRNA) or lncRNA-H19 shRNA (LV-H19-shRNA). The ecESCs infected with LV-NC-shRNA and LV-H19-shRNA were subcutaneously implanted into forty 6- to 8-week-old female nude mice. The size and weight of the endometriotic implants were measured at 1, 2, 3, and 4 weeks after implantation and compared, and lncRNA H19 levels in endometriotic implants were evaluated using real-time polymerase chain reaction (RT-PCR). All nude mice survived the experimental period, and no significant differences in body weight were observed between the experimental group and the control group. All nude mice developed histologically confirmed subcutaneous endometriotic lesions with glandular structures and stroma after 1 week of implantation. The subcutaneous lesions in the LV-NC-shRNA group after 1, 2, 3, and 4 weeks of implantation were larger than those in the LV-H19-shRNA group, and lncRNA H19 levels in subcutaneous lesions in the LV-NC-shRNA group were significantly higher than those in the LV-H19-shRNA group. Knockdown of lncRNA H19 suppresses endometriosis in vivo. Further study is required to explore the underlying mechanism in the future.


Subject(s)
Humans , Animals , Female , Rabbits , Endometriosis/genetics , RNA, Long Noncoding/genetics , RNA, Small Interfering/genetics , Cell Proliferation/genetics , Endometrium , Mice, Nude
5.
Clinics ; 76: e2175, 2021. tab, graf
Article in English | LILACS | ID: biblio-1249578

ABSTRACT

OBJECTIVE: The long non-coding RNA (lncRNA) KCNQ1 overlapping transcript 1 (KCNQ1OT1) exerts vital regulatory functions in diverse tumors. However, the biological function of KCNQ1OT1 in esophageal squamous cell carcinoma (ESCC) remains unclear. METHODS: KCNQ1OT1 expression was detected in ESCC tissues using quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation, apoptosis, migration, and invasion were detected by the CCK-8 assay, EdU assay, flow cytometry analysis, and Transwell experiments, respectively. Bioinformatics analysis, luciferase reporter experiments, and RNA immunoprecipitation assays were used to predict and validate the regulatory relationships between KCNQ1OT1, microRNA-133b (miR-133b) and epidermal growth factor receptor (EGFR). RESULTS: KCNQ1OT1 expression was remarkably upregulated in ESCC tissues and cell lines. Overexpression of KCNQ1OT1 markedly promoted ESCC cell proliferation, migration, and invasion and enhanced the expression of N-cadherin, MMP-2, and MMP-9, but inhibited apoptosis and E-cadherin expression in ESCC cell lines; KCNQ1OT1 knockdown exerted the opposite effects. KCNQ1OT1 could directly bind to miR-133b and suppress its expression, and miR-133b reversed the effects of KCNQ1OT1 overexpression in ESCC cells. MiR-133b reduced the expression of epidermal growth factor receptor (EGFR); further, KCNQ1OT1 activated the phosphatidylinositol 3-kinase/AKT serine/threonine kinase 1 (PI3K/AKT) signaling pathway by repressing miR-133b repression and indirectly upregulating EGFR. KCNQ1OT1 expression was positively correlated with EGFR mRNA expression and negatively correlated with miR-133b expression. CONCLUSION: KCNQ1OT1 facilitates ESCC progression by sponging miR-133b and activating the EGFR/PI3K/AKT pathway.


Subject(s)
Humans , Esophageal Neoplasms/genetics , MicroRNAs/genetics , RNA, Long Noncoding/genetics , Esophageal Squamous Cell Carcinoma/genetics , Phosphatidylinositol 3-Kinases , Cell Proliferation/genetics , KCNQ1 Potassium Channel/genetics
6.
Chinese Medical Journal ; (24): 2091-2101, 2021.
Article in English | WPRIM | ID: wpr-887631

ABSTRACT

BACKGROUND@#Long non-coding RNA (lncRNA) actin filament-associated protein 1 antisense RNA 1 (AFAP1-AS1) functions as a competing endogenous RNA to regulate target genes expression by sponging microRNAs (miRs) to play cancer-promoting roles in cancer stem cells. However, the regulatory mechanism of AFAP1-AS1 in cervical cancer (CC) stem cells is unknown. The present study aimed to provide a new therapeutic target for the clinical treatment of CC.@*METHODS@#Hyaluronic acid receptor cluster of differentiation 44 variant exon 6 (CD44v6)(+) CC cells were isolated by flow cytometry (FCM). Small interfering RNAs of AFAP1-AS1 (siAFAP1-AS1) were transfected into the (CD44v6)(+) cells. The levels of AFAP1-AS1 were measured by quantitative real-time PCR (qRT-PCR). Sphere formation assay, cell cycle analysis, and Western blotting were used to detect the effect of siAFAP1-AS1. RNA pull-down and luciferase reporter assay were used to verify the relationship between miR-27b-3p and AFAP1-AS1 or vascular endothelial growth factor (VEGF)-C.@*RESULTS@#CD44v6(+) CC cells had remarkable stemness and a high level of AFAP1-AS1. However, AFAP1-AS1 knockdown with siAFAP1-AS1 suppressed the cell cycle transition of G(1)/S phase and inhibited self-renewal of CD44v6(+) CC cells, the levels of the stemness markers octamer-binding transcription factor 4 (OCT4), osteopontin (OPN), and cluster of differentiation 133 (CD133), and the epithelial-mesenchymal transition (EMT)-related proteins Twist1, matrix metalloprotease (MMP)-9, and VEGF-C. In the mechanism study, miR-27b-3p/VEGF-C signaling was demonstrated to be a key downstream of AFAP1-AS1 in the CD44v6(+) CC cells.@*CONCLUSIONS@#LncRNA AFAP1-AS1 knockdown inhibits the CC cell stemness by upregulating miR-27b-3p to suppress VEGF-C.


Subject(s)
Cell Line, Tumor , Cell Proliferation/genetics , Female , Gene Expression Regulation, Neoplastic , Humans , MicroRNAs/genetics , RNA, Long Noncoding/genetics , Uterine Cervical Neoplasms/genetics , Vascular Endothelial Growth Factor A/genetics , Vascular Endothelial Growth Factor C
7.
Article in Chinese | WPRIM | ID: wpr-879535

ABSTRACT

OBJECTIVE@#To compare the mRNA level of cell proliferation-related genes Twist1, SIRT1, FGF2 and TGF-β3 in placenta mesenchymal stem cells (PA-MSCs), umbilical cord mensenchymals (UC-MSCs) and dental pulp mesenchymal stem cells (DP-MSCs).@*METHODS@#The morphology of various passages of PA-MSCs, UC-MSCs and DP-MSCs were observed by microscopy. Proliferation and promoting ability of the three cell lines were detected with the MTT method. Real-time PCR (RT-PCR) was used to determine the mRNA levels of Twist1, SIRT1, FGF2, TGF-β3.@*RESULTS@#The morphology of UC-MSCs and DP-MSCs was different from that of PA-MSCs. Proliferation ability and promoting ability of the PA-MSCs was superior to that of UC-MSCs and DP-MSCs. In PA-MSCs, expression level of Twist1 and TGF-β3 was the highest and FGF2 was the lowest. SIRT1 was highly expressed in UC-MSCs. With the cell subcultured, different expression levels of Twist1, SIRT1, FGF2, TGF-β3 was observed in PA-MSCs, UC-MSCs and DP-MSCs.@*CONCLUSION@#Up-regulated expression of the Twist1, SIRT1 and TGF-β3 genes can promote proliferation of PA-MSCs, UC-MSCs and DP-MSCs, whilst TGF-β3 may inhibit these. The regulatory effect of Twist1, SIRT1, FGF2 and TGF-β3 genes on PA-MSCs, UC-MSCs and DP-MSCs are different.


Subject(s)
Cell Differentiation , Cell Proliferation/genetics , Cells, Cultured , Dental Pulp/cytology , Female , Fibroblast Growth Factor 2/genetics , Humans , Mesenchymal Stem Cells/cytology , Nuclear Proteins/genetics , Placenta/cytology , Pregnancy , Sirtuin 1/genetics , Transforming Growth Factor beta3/genetics , Twist-Related Protein 1/genetics , Umbilical Cord/cytology
8.
Article in Chinese | WPRIM | ID: wpr-921928

ABSTRACT

OBJECTIVE@#To explore the effects of siRNA hsa-circ-0000885 modified bone marrow mesenchymal stem cells (BMSCs) on osteogenic differentiation, cell proliferation and apoptosis in order to provide new ideas and methods for the clinical treatment of osteoporosis (OP).@*METHODS@#From September 2018 to February 2020, 13 patients with osteoporosis admitted to our hospital were selected as the research objects, including 11 females and 2 males, with an age of (65.45±10.77) years old. After obtaining the informed consent of patients, peripheral blood tissues were extracted. Then the expression level of cir-cRNA in peripheral blood mononuclear cells(PBMC) was detected by circ RNA chip. The expression of circ RNA was silenced by siRNA technology. The BMSCs were transfected with lentivirus. According to the siRNA interference plasmid hsa-circ-0000885, the cells were divided into the blank group, the empty vector group and the siRNA interference group. After 72 hours of treatment, the cell cycle was detected by flow cytometry, the apoptosis level was detected by AV-PI kit, and the osteogenic differentiation ability of BMSCs was detected by ALP staining.@*RESULTS@#The expression of hsa-circ-0000885 in PBMC of patients with osteoporosis was significantly higher than that of healthy controls (@*CONCLUSION@#The lentivirus mediated siRNA hsa-circ-0000885 plasmid transfected into BMSCs and osteoclast co culture system can promote cell proliferation, inhibit apoptosis and promote osteogenic differentiation of BMSCs, which can be used as a potential therapeutic target for OP patients.


Subject(s)
Aged , Apoptosis/genetics , Cell Differentiation , Cell Proliferation/genetics , Cells, Cultured , Coculture Techniques , Female , Humans , Lentivirus , Leukocytes, Mononuclear , Mesenchymal Stem Cells , Middle Aged , Osteoclasts , Osteogenesis/genetics , RNA, Small Interfering/genetics , Transfection
9.
Chinese Medical Journal ; (24): 1345-1355, 2021.
Article in English | WPRIM | ID: wpr-878149

ABSTRACT

BACKGROUND@#Although increasing abnormal expression of circular RNAs (circRNAs) has been revealed in various cancers, there were a small number of studies about circRNAs in gastric cancer (GC). Here, we explored the expression and function of a novel circRNA, circ_0049447, in GC.@*METHODS@#A total of 80 GC tissues and non-tumorous tissues were collected from the First Affiliated Hospital of China Medical University. And all cells were cultured with 10% fetal bovine serum and incubated at 37°C and 5% CO2. The expression of circ_0049447 was quantified by real-time polymerase chain reaction. The biological function of circ_0049447 on proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) was evaluated by cell counting kit-8 (CCK-8), colony formation assay, transwell migration and invasion assay, and Western blotting. Luciferase report assay was used to verify the direct binding between circ_0049447 and predicted microRNA (miRNA). Furthermore, a xenograft mouse model was used to validate the function of circ_0049447 in vivo.@*RESULTS@#We demonstrated that circ_0049447 was downregulated in GC (P < 0.001). The area under the receiver operating characteristic curve reached 0.838, while sensitivity was 82.3% and specificity was 77.2%. CCK-8 and colony formation assay showed that overexpression of circ_0049447 could inhibit the proliferation (P < 0.05). Transwell migration and invasion assay showed upregulated circ_0049447 could impede migration in GC cells (P < 0.05). In addition, overexpression of circ_0049447 could impede GC cell EMT. Upregulation of miR-324-5p in GC specimens and direct binding between miR-324-5p with circ_0049447 proven by luciferase reporter assay indicated that circ_0049447 may inhibit GC by sponging certain miRNA.@*CONCLUSION@#Circ_0049447 acts as a tumor suppressor in GC through reducing proliferation, migration, invasion, and EMT, and it is a promising biomarker for diagnosis.


Subject(s)
Animals , Cell Line, Tumor , Cell Proliferation/genetics , China , Epithelial-Mesenchymal Transition/genetics , Mice , Stomach Neoplasms/genetics
10.
Chinese Medical Journal ; (24): 564-572, 2021.
Article in English | WPRIM | ID: wpr-878081

ABSTRACT

BACKGROUND@#The pathogenesis of osteosarcoma (OS) is still unclear, and it is still necessary to find new targets and drugs for anti-OS. This study aimed to investigate the role and mechanism of the anti-OS effects of miR-296-5p.@*METHODS@#We measured the expression of miR-296-5p in human OS cell lines and tissues. The effect of miR-296-5p and its target gene staphylococcal nuclease and tudor domain containing 1 on proliferation, migration, and invasion of human OS lines was examined. The Student's t test was used for statistical analysis.@*RESULTS@#We found that microRNA (miR)-296-5p was significantly downregulated in OS cell lines and tissues (control vs. OS, 1.802 ± 0.313 vs. 0.618 ± 0.235, t = 6.402, P < 0.01). Overexpression of miR-296-5p suppressed proliferation, migration, and invasion of OA cells. SND1 was identified as a target of miR-296-5p by bioinformatic analysis and dual-luciferase reporter assay. Overexpression of SND1 abrogated the effects induced by miR-296-5p upregulation (miRNA-296-5p vs. miRNA-296-5p + SND1, 0.294 ± 0.159 vs. 2.300 ± 0.277, t = 12.68, P = 0.003).@*CONCLUSION@#Our study indicates that miR-296-5p may function as a tumor suppressor by targeting SND1 in OS.


Subject(s)
Bone Neoplasms/genetics , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Endonucleases/genetics , Gene Expression Regulation, Neoplastic , Genes, Tumor Suppressor , Humans , MicroRNAs/genetics , Osteosarcoma/genetics
11.
Article in Chinese | WPRIM | ID: wpr-921525

ABSTRACT

Objective To explore the effect of miR-145-5p on the proliferation and apoptosis of human ovarian cancer cells and the possible molecular mechanisms involved.Methods Real-time quantitative PCR was performed to detect the expression of miR-145-5p in ovarian epithelial cells and ovarian cancer cells.CCK-8 and flow cytometry were used to detect the effects of miR-145-5p overexpression on the proliferation and apoptosis of ovarian cancer cells.TargetScan was employed to predict the target genes of miR-145-5p.Western blotting,dual luciferase reporter assay and rescue experiment were employed to predict and verify the underlying molecular mechanism of miR-145-5p function.Results The expression of miR-145-5p in ovarian cancer cells was significantly lower than that in normal ovarian epithelial cells(


Subject(s)
Apoptosis/genetics , Carcinoma, Ovarian Epithelial/genetics , Cell Line, Tumor , Cell Proliferation/genetics , Female , Gene Expression Regulation, Neoplastic , Humans , MicroRNAs/genetics , Ovarian Neoplasms/genetics
12.
Chinese Medical Journal ; (24): 2999-3008, 2021.
Article in English | WPRIM | ID: wpr-921262

ABSTRACT

BACKGROUND@#The association between miR-532-3p and tongue squamous cell carcinoma (TSCC) has been examined in the literature to improve the survival rate of patients with this tumor. However, further studies are needed to confirm the regulatory roles of this microRNA (miRNA) in TSCC. The objective of this study was to investigate the roles played by and the underlying mechanism used by the miR-532-3p/podoplanin (PDPN) axis in TSCC development.@*METHODS@#Western blotting and quantitative real-time reverse transcription-polymerase chain reaction (RT-qPCR) were performed to evaluate the PDPN expression level in TSCC tissues and cells. The proliferative, adhesive, and migratory capabilities of TSCC cells (CAL-27 and CTSC-3) were examined using cell counting kit-8 (CCK-8), cell adhesion, and wound-healing assays, respectively. The dual-luciferase reporter (DLR) assay was later conducted to confirm the relationship between miR-532-3p and PDPN.@*RESULTS@#The results indicated that PDPN expression was enriched in TSCC tissues and cells, and that the expression of PDPN was associated with some clinicopathological parameters of TSCC, including lymph node metastasis (P = 0.001), tumor-node-metastasis (TNM) staging (P = 0.010), and grading (P = 0.010). Further analysis also showed that PDPN knockdown inhibited the viability, adhesive ability, and migratory capacity of CAL-27 and CTSC-3 cells, effects that could be reversed by the application of a miR-532-3p inhibitor. Additionally, PDPN was found to be a direct target of miR-532-3p.@*CONCLUSIONS@#This research suggested that by targeting PDPN, miR-532-3p could inhibit cell proliferation viability, adhesion, and migration in TSCC. Findings also revealed that the miR-532-3p/PDPN axis might provide more insights into the prognosis and treatment of TSCC.


Subject(s)
Carcinoma, Squamous Cell/genetics , Cell Line, Tumor , Cell Proliferation/genetics , Gene Expression Regulation, Neoplastic , Humans , Membrane Glycoproteins , MicroRNAs/genetics , Tongue Neoplasms/genetics
13.
Chinese Medical Journal ; (24): 2619-2628, 2021.
Article in English | WPRIM | ID: wpr-921210

ABSTRACT

BACKGROUND@#Microribose nucleic acids (miRNAs) are implicated in the progression of lung adenocarcinoma. MicroRNA-345-5p (miR-345-5p) is a recently identified anti-oncogene in some human cancers, but its functional role and possible molecular mechanism in lung adenocarcinoma remain unknown. This study aimed to identify the biological function and underlying mechanism of miR-345-5p in lung adenocarcinoma cells.@*METHODS@#In this study, lung adenocarcinoma tissues and adjacent tissues were collected in the First Affiliated Hospital of Anhui Medical University between April 2016 and February 2017. The expression of miR-345-5p and ras homolog family member A (RhoA) in lung adenocarcinoma tissues and human lung adenocarcinoma cell lines (A549, H1650, PC-9, and H441) was detected by reverse transcription quantitative polymerase chain reaction analysis. Functional assays including colony formation, flow cytometry analysis, wound healing, and transwell assays were performed to assess the proliferation, apoptosis, migration, and invasion of lung adenocarcinoma cells. In addition, RNA pulldown and luciferase reporter assays were conducted to evaluate the relationship between miR-345-5p and RhoA. Difference between the two groups was analyzed with Student's t test, while that among multiple groups was analyzed with one-way analysis of variance.@*RESULTS@#MiR-345-5p expression displayed lower level in lung adenocarcinoma tissues (0.241 ± 0.095 vs.1.000 ± 0.233, t = 19.247, P < 0.001) and cell lines (F = 56.992, P < 0.001) than control tissues and cells. Functional experiments demonstrated that upregulation of miR-345-5p inhibited the malignant phenotypes of lung adenocarcinoma cells via suppressing cell proliferation, migration, invasion, and facilitating cell apoptosis. Additionally, RhoA was verified to be the downstream target of miR-345-5p. Expression of RhoA was downregulated by overexpression of miR-345-5p in PC-9 (0.321 ± 0.047 vs. 1.000 ± 0.127, t = 8.536, P < 0.001) and H1650 (0.398 ± 0.054 vs. 1.000 ± 0.156, t = 4.429, P = 0.011) cells. Rescue assays revealed that overexpression of RhoA rescued the suppressive effects of miR-345-5p upregulation on proliferation, migration, and invasion of lung adenocarcinoma cells. Further, miR-345-5p was found to regulate the Rho/Rho-associated protein kinase (ROCK) signaling pathway by downregulation of RhoA in lung adenocarcinoma cells.@*CONCLUSIONS@#MiR-345-5p plays a tumor suppressor role in lung adenocarcinoma cells by downregulating RhoA to inactivate the Rho/ROCK pathway.


Subject(s)
Adenocarcinoma of Lung/genetics , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Gene Expression Regulation, Neoplastic , Humans , Lung Neoplasms/genetics , MicroRNAs/genetics , Up-Regulation/genetics , rho-Associated Kinases/genetics , rhoA GTP-Binding Protein/genetics
14.
Chinese Medical Journal ; (24): 2721-2729, 2021.
Article in English | WPRIM | ID: wpr-921206

ABSTRACT

BACKGROUND@#The chaperonin containing t-complex (CCT) proteins play an important role in cell cycle-related protein degradation in yeast and mammals. The role of the chaperonin containing t-complex 4 (CCT4), one subtype of CCT proteins, in the progress of hepatocellular carcinoma (HCC) was not fully elucidated. Here, we aimed to explore the mechanisms of CCT4 in HCC.@*METHODS@#In this study, we used the UALCAN platform to analyze the relationship between CCT4 and HCC, and the association of CCT4 with the overall survival (OS) of HCC patients was also analyzed. CCT4 expression in HCC tumor tissues and normal tissues was also determined by western blot (WB) assay. Lentivirus vector was used to knock down the CCT4 expression, and quantitative polymerase chain reaction and WB were used to determine the level of CCT4 in HCC cell lines. Cell counting kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) assays were used to detect the cell proliferation, and flow cytometry (FCM) was performed to evaluate the effect of CCT4 on the apoptosis of HCC cells. Co-immunoprecipitation (co-IP) assay and WB were used to explore the mechanisms of CCT4 regulating the growth of HCC. Data were calculated from at least three replicate experiments and expressed as mean ± standard deviation. Student's t test, paired t test, and Kaplan-Meier analysis were used to compare across different groups.@*RESULTS@#We found CCT4 was upregulated in HCC tissues compared with normal tissues, and its high expression was associated with poor prognosis (P < 0.001). CCT4 was significantly increased in HCC tumor tissues compared with normal tissues (0.98 ± 0.12 vs. 0.23 ± 0.05, t = 7.73, P < 0.001). After being transfected with CCT4 short-hairpin RNA (shRNA), CCT4 was decreased in mRNA level and protein level in both Huh7 (mRNA level: 0.41 ± 0.07 vs. 1.01 ± 0.11, t = 8.09, P = 0.001; protein level: 0.61 ± 0.03 vs. 0.93 ± 0.07, t = 7.19, P = 0.002) and Hep3b cells (mRNA level: 0.55 ± 0.11 vs. 1.04 ± 0.15, t = 4.51, P = 0.011; protein level: 0.64 ± 0.10 vs. 0.95 ± 0.08, t = 4.32, P = 0.012). CCK8 assay indicated that CCT4 knockdown inhibited cell proliferation in both Huh7 (OD value of 3 days: 0.60 ± 0.14 vs. 0.97 ± 0.16, t = 3.13, P = 0.036; OD value of 4 days: 1.03 ± 0.07 vs. 1.50 ± 0.12, t = 5.97, P = 0.004) and Hep3b (OD value of 3 days: 0.69 ± 0.14 vs. 1.10 ± 0.11, t = 3.91, P = 0.017; OD value of 4 days: 1.12 ± 0.12 vs. 1.48 ± 0.13, t = 3.55, P = 0.024) cells. EdU assay showed that CCT4 knockdown inhibited the cell proliferation in both Huh7 (EdU positive rate: [31.25 ± 3.41]% vs. [58.72 ± 3.78]%, t = 9.34, P = 0.001) and Hep3b cells (EdU positive rate: [44.13 ± 7.02]% vs. [61.79 ± 3.96]%, t = 3.79, P = 0.019). FCM assay suggested that CCT4 knockdown induced apoptosis in HCC cells (apoptosis rate of Huh7: [9.10 ± 0.80]% vs. [3.66 ± 0.64]%, t = -9.18, P = 0.001; apoptosis rate of Hep3b: [6.69 ± 0.72]% vs. [4.20 ± 0.86]%, t = -3.84, P = 0.018). We also found that CCT4 could regulate anaphase-promoting complex (APC)Cdc20 activity via interacting with Cdc20. Furthermore, CCT4 knockdown induced securin (0.65 ± 0.06 vs. 0.44 ± 0.05, t = -4.69, P = 0.009) and B-cell lymphoma-2 (Bcl-2) interacting mediator of cell death (Bim; 0.96 ± 0.06 vs. 0.61 ± 0.09, t = -5.65, P = 0.005) accumulation. The upregulation of securin inhibited cell growth by downregulating cyclin D1 (0.65 ± 0.05 vs. 1.04 ± 0.07, t = 8.12, P = 0.001), and the accumulation of Bim inhibited Bcl-2 (0.77 ± 0.04 vs. 0.87 ± 0.04, t = 3.00, P = 0.040) and activated caspase 9 (caspase 9: 0.77 ± 0.04 vs. 0.84 ± 0.05, t = 1.81, P = 0.145; cleaved caspase 9: 0.64 ± 0.06 vs. 0.16 ± 0.07, t = 1.81, P = 0.001), which led to elevated apoptosis.@*CONCLUSIONS@#Overall, these results showed that CCT4 played an important role in HCC pathogenesis through, at least partly, interacting with Cdc20.


Subject(s)
Animals , Apoptosis , Carcinoma, Hepatocellular/genetics , Cdc20 Proteins , Cell Line, Tumor , Cell Proliferation/genetics , Gene Expression Regulation, Neoplastic , Humans , Liver Neoplasms/genetics
15.
Chinese Medical Journal ; (24): 2465-2474, 2021.
Article in English | WPRIM | ID: wpr-921151

ABSTRACT

BACKGROUND@#Ubiquitin-conjugating enzyme E2C (UBE2C) has been shown to be associated with the occurrence of various cancers and involved in many tumorigenic processes. This study aimed to investigate the specific molecular mechanism through which UBE2C affects breast cancer (BC) proliferation.@*METHODS@#BC-related datasets were screened according to filter criteria in the Gene Expression Omnibus (GEO) database and The Cancer Genome Atlas (TCGA) database. Then differentially expressed genes (DEGs) were identified using Venn diagram analysis. By using DEGs, we conducted the following analyses including Gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), protein-protein interaction (PPI), and survival analysis, and then validated the function of the hub gene UBE2C using quantitative reverse transcription-polymerase chain reaction (RT-qPCR), cell counting kit-8 (CCK-8) assay, transwell assay, and Western blot assay.@*RESULTS@#In total, 151 DEGs were identified from the GEO and TCGA databases. The results of GO analysis demonstrated that the DEGs were significantly enriched with mitotic nuclear division, lipid droplet, and organic acid-binding. KEGG analysis showed that the peroxisome proliferators-activated receptor (PPAR) signaling pathway, regulation of lipolysis in adipocytes, and proximal tubule bicarbonate reclamation were significantly enriched in the signal transduction pathway category. The top three hub genes that resulted from the PPI network were FOXM1, UBE2C, and CDKN3. The results of survival analysis showed a close relationship between UBE2C and BC. The results of CCK-8 and transwell assays suggested that the proliferation and invasion of UBE2C knockdown cells were significantly inhibited (P < 0.050). The results of Western blot assay showed that the level of phosphorylated phosphatase and tensin homology deleted on chromosome 10 (p-PTEN) was obviously increased (P < 0.050), whereas the levels of phosphorylated protein kinase B (p-AKT), phosphorylated mammalian target of rapamycin (p-mTOR), and hypoxia-inducible factor-1 alpha (HIF-1α) were dramatically decreased (P < 0.050) in the UBE2C knockdown cell.@*CONCLUSION@#UBE2C can promote BC proliferation by activating the AKT/mTOR signaling pathway.


Subject(s)
Biomarkers, Tumor , Breast Neoplasms/pathology , Cell Proliferation/genetics , Computational Biology , Female , Gene Expression Regulation, Neoplastic , Humans , Proto-Oncogene Proteins c-akt/genetics , Signal Transduction/genetics , TOR Serine-Threonine Kinases/genetics , Ubiquitin-Conjugating Enzymes/metabolism
16.
Chinese Medical Journal ; (24): 2611-2618, 2021.
Article in English | WPRIM | ID: wpr-921137

ABSTRACT

BACKGROUND@#Nucleolar protein 6 (NOL6) is a nucleolar RNA-associated protein that is highly conserved between species. It has been proved to be associated with the prognosis of liver cancer. However, the underlying mechanism has not been fully established. This study aimed to assess the relationship between NOL6 and liver cancer prognosis.@*METHODS@#We constructed an NOL6-short hairpin RNA (shRNA)-expressing lentivirus. Through viral transfection, cell growth assay and fluorescence-activated cell sorting, we evaluated the effect of shRNA-mediated NOL6 knockdown on the proliferation, colony formation, and apoptosis of hepatocellular carcinoma (HCC) cells. The relationship between NOL6 expression and HCC patient survival has been established through bioinformatics analysis. We also explored the downstream molecular regulatory network of NOL6 in HCC by performing an Ingenuity Pathway Analysis in the database.@*RESULTS@#Increased NOL6 expression was detected in HCC cells compared to normal controls; HCC patients with high NOL6 expression had poorer prognoses than those with low expression. NOL6 knockdown inhibited HCC cell proliferation, apoptosis, and colony formation. Also, MAPK8, CEBPA, and FOSL1 were selected as potential downstream genes of NOL6.@*CONCLUSIONS@#NOL6 up-regulates HCC cell proliferation and affects downstream expression of related genes. Moreover, NOL6 is considered to be associated with poor prognosis in HCC patients.


Subject(s)
Apoptosis/genetics , Carcinoma, Hepatocellular/genetics , Cell Line, Tumor , Cell Proliferation/genetics , Gene Expression Regulation, Neoplastic , Humans , Liver Neoplasms/genetics , Nuclear Proteins , Prognosis
17.
Acta Physiologica Sinica ; (6): 491-500, 2021.
Article in Chinese | WPRIM | ID: wpr-887684

ABSTRACT

Many studies have shown that circular RNAs (circRNAs) play a key regulatory role in the whole biological process of tumors. The purpose of this study was to explore the biological function and molecular mechanism of circ_0001666 in non-small cell lung cancer (NSCLC), so as to provide new targets for the diagnosis and treatment of NSCLC. Gene expression profiles were downloaded from Gene Expression Omnibus (GEO, GSE101586) and the differential genes were obtained by using GEO2R analysis. The quantitative real time polymerase chain reaction (qRT-PCR) was used to detect the expression level of circ_0001666 in NSCLC cells. Cell counting kit-8 (CCK-8) and Annexin V-FITC apoptosis detection kit were respectively used to assess the cell proliferation and apoptosis, where circ_0001666 was knockdown in NSCLC cells. The targeted relationship among mircoRNA 330-5p (miR-330-5p), circ_0001666, and high mobility group A2 protein (HMGA2) was verified by bioinformatics prediction, dual-luciferase reporter gene, RNA immunoprecipitation (RIP) and RNA pull down assay. The results showed that the expression of circ_0001666 in NSCLC cells was significantly up-regulated than that in normal lung epithelial cells. Circ_0001666 knockdown reduced the cell viability and promoted the apoptosis of NSCLC cells, which could be reversed by miR-330-5p inhibitors. MiR-330-5p is the downstream target of circ_0001666 and can be adsorbed by circ_0001666. HMGA2 is a target gene of miR-330-5p, which can be indirectly regulated by circ_0001666. The results suggest that circ_0001666 promotes the proliferation and inhibits apoptosis of NSCLC cells via miR-330-5p/HMGA2 axis.


Subject(s)
Apoptosis/genetics , Carcinoma, Non-Small-Cell Lung/genetics , Cell Proliferation/genetics , HMGA2 Protein , Humans , Lung Neoplasms/genetics , MicroRNAs/genetics , RNA, Circular
18.
Article in Chinese | WPRIM | ID: wpr-880841

ABSTRACT

OBJECTIVE@#To investigate the effects of overexpression of long noncoding RNA (lncRNA) MEG3 on the proliferation and invasion of glioblastoma U251 cells by suppressing the expression of hypoxia inducible factor 1@*METHODS@#The expression of lncRNA MEG3 and HIF1@*RESULTS@#The expression of MEG3 was significantly lower and HIF1@*CONCLUSIONS@#MEG3 overexpression inhibits the proliferation and invasion of U251 cells through suppressing the expression of HIF1


Subject(s)
Apoptosis , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Gene Expression Regulation, Neoplastic , Glioblastoma/genetics , Humans , MicroRNAs , RNA, Long Noncoding/genetics
19.
Braz. j. med. biol. res ; 53(1): e8883, Jan. 2020. tab, graf
Article in English | LILACS | ID: biblio-1055486

ABSTRACT

Opa-interacting protein 5 antisense transcript 1 (OIP5-AS1) is one kind of cytoplasmic long non-coding RNA (lncRNA), which has been demonstrated to play a critical function in multiple cancers. However, the detailed mechanism of OIP5-AS1 in the regulation of cervical cancer progression is still obscure. Here, we demonstrated that lncRNA OIP5-AS1 was upregulated in cervical cancer and was correlated with poor prognosis by bioinformatics studies. OIP5-AS1 depletion inhibited cell proliferation and promoted cell apoptosis in cervical cancer cells. Furthermore, we clarified that ROCK1 was the downstream effector of OIP5-AS1 and OIP5-AS1 acted as a molecular sponge of miR-143-3p. Finally, we verified that OIP5-AS1 exerted its function in the regulation of cervical cancer progression via interacting with miR-143-3p to regulate ROCK1 expression. Our study revealed novel mechanisms about how lncRNA OIP5-AS1 executed its function in cervical cancer and thus provided potential therapeutic targets for the disease.


Subject(s)
Humans , Female , Uterine Cervical Neoplasms/pathology , Apoptosis/physiology , MicroRNAs/metabolism , Cell Proliferation/physiology , rho-Associated Kinases/metabolism , RNA, Long Noncoding/metabolism , Gene Expression Regulation, Neoplastic , Up-Regulation , Uterine Cervical Neoplasms/metabolism , Blotting, Western , Apoptosis/genetics , Reverse Transcriptase Polymerase Chain Reaction , MicroRNAs/genetics , Cell Line, Tumor , Cell Proliferation/genetics , rho-Associated Kinases/genetics , RNA, Long Noncoding/genetics
20.
Braz. j. med. biol. res ; 53(4): e8882, 2020. tab, graf
Article in English | LILACS | ID: biblio-1100927

ABSTRACT

Hepatocellular carcinoma (HCC) is one of the most common primary malignant tumors of the liver worldwide. Liver resection and transplantation are currently the only effective treatments; however, recurrence and metastasis rates are still high. Previous studies have shown that the epithelial-mesenchymal transition (EMT) is a key step in HCC invasion and metastasis. Inhibition of EMT has become a new therapeutic strategy for tumors. Recently, puerarin, a well-characterized component of traditional Chinese medicine, has been isolated from Pueraria radix and exerts positive effects on many diseases, particularly cancers. In this study, CCK-8, EdU immunofluorescence, colony formation, wound healing, and migration assays were used to detect the effects of puerarin on HCC cells. We further analyzed the relationship between puerarin and miR-21/PTEN/EMT markers in HCC cell lines. Our results showed that HCC cell proliferation, migration, invasion, tumor formation, and metastasis were reduced by puerarin in vitro and in vivo. Additionally, puerarin inhibited the EMT process of HCC by affecting the expression of Slug and Snail. Moreover, oncogenic miR-21 was inhibited by puerarin, coupled with an increase in the tumor suppressor gene PTEN. Increasing miR-21 expression or decreasing PTEN expression reversed the inhibition effects of puerarin in HCC. These data confirmed that puerarin affects HCC through the miR-21/PTEN/EMT regulatory axis. Overall, puerarin may represent a chemopreventive and/or chemotherapeutic agent for HCC treatment.


Subject(s)
Animals , Male , Carcinoma, Hepatocellular/drug therapy , PTEN Phosphohydrolase/genetics , Epithelial-Mesenchymal Transition/drug effects , Isoflavones/therapeutic use , Liver Neoplasms/drug therapy , Antineoplastic Agents/therapeutic use , Pyrroles , Drug Screening Assays, Antitumor , Carcinoma, Hepatocellular/genetics , MicroRNAs/genetics , Cell Proliferation/drug effects , Cell Proliferation/genetics , Disease Models, Animal , Isoflavones/pharmacology , Liver Neoplasms/genetics , Neoplasm Invasiveness , Antineoplastic Agents/pharmacology
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