The Kaohsiung Journal of Medical Sciences

Extracellular vesicles of Bifidobacterium longum reverse the acquired carboplatin resistance in ovarian cancer cells via p53 phosphorylation on Ser15

AbstractWe previously found that the relative abundance of Bifidobacterium was increased after chemotherapy; however, the role of Bifidobacterium longum in chemotherapeutic drug resistance in ovarian cancer (OVC) remains unclear. This study aimed to understand the potential effects and mechanism of B. longum extracellular vesicles (B. longum‐EVs) on carboplatin (CBP) resistance in OVC. Eight normal and 11 ovarian tissues were collected and the expression of B. longum genomic DNA and its association with acquired CBP resistance in OVC patients was determined. After isolating EVs by ultracentrifugation from B. longum (ATCC 15707), CBP‐resistant A2780 cells were treated with PBS, CBP, B. longum‐EVs, or CBP + B. longum‐EVs, and subsequently analyzed by CCK‐8, Edu staining, Annexin V/PI double staining, wound healing, and Transwell assays to detect cell viability, proliferation, apoptosis, migration, and invasion, respectively. MRP1, ATP7A, ATP7B, and p53 expression as well as p53 phosphorylation were measured by western blot analysis. S15A mutation of p53 was assessed to examine the potential role of p53 Ser15 phosphorylation in CBP‐resistant OVC. B. longum levels were elevated and positively associated with CBP resistance in OVC patients. Only high concentrations of B. longum‐EVs attenuated A2780 cell proliferation, apoptosis, migration, and invasion. B. longum‐EVs exposure significantly enhanced the sensitivity of CBP‐resistant A2780 cells to CBP and decreased the expression of drug resistance‐related proteins. The effect of B. longum‐EVs on reversing CBP resistance was completely inhibited by S15A mutation of p53. B. longum‐EVs enhanced the sensitivity of OVC cells to CBP through p53 phosphorylation on Ser15.

lncRNA LINC00958 Activates NOTCH3 by Competitively Inhibiting miR‐129‐2‐3p to Exacerbate the Malignant Biological Behaviors of Endometrial Cancer Cells

ABSTRACT Endometrial cancer (EC) is one of the most prevalent gynecologic malignancies, with increasing incidence worldwide. Recent studies have highlighted the critical roles of long noncoding RNAs (lncRNAs) in regulating tumor progression. This study investigated the oncogenic function of lncRNA LINC00958 in EC and its underlying mechanism involving the miR‐129‐2‐3p/NOTCH3 signaling axis. Experimental results showed that LINC00958 was significantly upregulated in EC cell lines. Overexpression of LINC00958 enhanced cell proliferation, migration, and invasion, while inhibiting apoptosis, as evidenced by increased Bcl‐2 and decreased cleaved caspase‐3 and Bax expression. Conversely, silencing LINC00958 suppressed these malignant behaviors. Mechanistically, LINC00958 acted as a competing endogenous RNA (ceRNA), directly binding to miR‐129‐2‐3p and thereby relieving its suppression on NOTCH3. Dual‐luciferase reporter assays confirmed the direct interaction between LINC00958 and miR‐129‐2‐3p, as well as between miR‐129‐2‐3p and NOTCH3. Immunofluorescence analysis further demonstrated enhanced nuclear translocation of NOTCH3 following LINC00958 overexpression. Functional rescue experiments showed that miR‐129‐2‐3p overexpression or NOTCH3 knockdown effectively counteracted the tumor‐promoting effects of LINC00958. In vivo xenograft experiments using Ishikawa cells supported the in vitro findings, confirming that LINC00958 promotes tumor growth by modulating the miR‐129‐2‐3p/NOTCH3 axis. Overall, this study identifies LINC00958 as a novel oncogenic lncRNA in EC, which facilitates tumor progression by sponging miR‐129‐2‐3p and enhancing NOTCH3 signaling. These findings provide new insights into the molecular mechanisms of EC and suggest that targeting the LINC00958/miR‐129‐2‐3p/NOTCH3 axis may represent a promising therapeutic strategy.

Long non‐coding RNA VPS9D1‐AS1 enhances proliferation, invasion, and epithelial‐mesenchymal transition in endometrial cancer via miR‐377‐3p/SGK1

AbstractEndometrial cancer (EC) is a kind of gynecologic malignancy with a rising incidence rate. This study aimed to explore the role of VPS9D1 antisense RNA1 (VPS9D1‐AS1) in EC. The expression of VPS9D1‐AS1, microRNA (miR)‐377‐3p, and serum and glucocorticoid‐regulated kinase 1 (SGK1) was detected by Quantitative Real‐Time PCR (qRT‐PCR). Cell proliferation, invasion and epithelial‐mesenchymal transition (EMT) were determined by cell counting kit‐8 (CCK‐8), 5‐Ethynyl‐2′‐Deoxyuridine (EdU) transwell, and western bolt. VPS9D1‐AS1 was predicted to sponge miR‐377‐3p via Starbase, and verified by luciferase reporter, RNA binding protein immunoprecipitation (RIP), and RNA pull‐down experiments. The clinical characteristics of VPS9D1‐AS1, miR‐377‐3p, and SGK1 were analyzed. The role of VPS9D1‐AS1 on EC tumorigenesis was assessed in xenografted nude mice. VPS9D1‐AS1 was upregulated in EC cells and tissues. Interference of VPS9D1‐AS1 inhibited growth, invasion, and EMT of EC cells. Mechanically, VPS9D1‐AS1 was a molecular sponge of miR‐377‐3p, and overexpression of miR‐377‐3p reversed VPS9D1‐AS1‐induced EC cells proliferation, invasion, and EMT. Moreover, SGK1 was confirmed to bind with miR‐377‐3p. Furthermore, overexpression of SGK1 alleviated sh‐VPS9D1‐AS1‐caused effects on EC cells. High level of VPS9D1‐AS1 and SGK1, or low miR‐377‐3p expression predicted a poor prognosis. The expression of the three genes was correlated with lymph node metastasis, pathological stage, and International Federation of Gynecology and Obstetrics (FIGO) stage, but not associated with age, ER, and PR expression. Interestingly, knockdown of VPS9D1‐AS1 suppressed EC tumor growth in mice. VPS9D1‐AS1 promoted cell invasion, proliferation, and EMT via modulating miR‐377‐3p/SGK1 axis, which provided new options for therapeutic strategies of EC.

miR‐335 modulates Numb alternative splicing via targeting RBM10 in endometrial cancer

AbstractNumb is a conserved protein plays important roles in the development of cancer. Two Numb isoforms have been found produced by alternative splicing and play contrast roles in regulating cellular functions. It is reported that the expression of Numb long isoform (Numb‐L) was increased in various kinds of cancers, but in endometrial cancer, the condition is still unknown. The level of two Numb transcripts and protein isoforms were detected by semiquantitative polymerase chain reaction and immunoblotting in 47 paired endometrial tumor and adjacent non‐tumor control tissues. The level of three alternative splicing related proteins: RBM5, RBM6, and RBM10 was determined by immunoblotting. MiRNAs targeting RBM10 were predicted by bioinformatics tools and their interaction with RBM10 was confirmed by luciferase assay and immunoblotting. The function of miR‐335 in endometrial cancer was examined in xenograft mouse model. Numb‐L level was increased in tumors and negatively correlated with RBM10 protein level. RBM10 mRNA level was not significantly altered in endometrial tumors suggesting its expression may regulated by post transcriptional regulators such as miRNAs. We identified miR‐133a, miR‐133b, and miR‐335 directly target RBM10, but only miR‐335 level increased in tumors and negatively correlated with RBM10 protein level. miR‐335 overexpression promoted tumor growth by downregulating RBM10 and upregulating Numb‐L level in xenograft mouse model. miR‐335 overexpression promoted Numb‐L expression via targeting RBM10 in endometrial cancer, which may provide new biomarkers for EC diagnosis.

Alternative splicing of VEGFA is regulated by RBM10 in endometrial cancer

AbstractVascular endothelial growth factor A (VEGFA) gene has three alternative exons which results in multiple isoforms. VEGFA has been found overexpressed in patients with endometrial cancer, but the VEGFA expression pattern and how it is regulated are still unknown. The level of VEGFA transcripts and protein isoforms were detected by semi‐quantitative Polymerase chain reaction (PCR) and immunoblotting in 29 paired endometrial tumor and adjacent nontumor control tissues. The level of three alternative splicing related proteins: RBM5, RBM6, and RBM10 was determined by immunoblotting. The H3K27Ac level in RBM10 promoter region was detected by ChIP‐PCR. The RBM10 promoter region methylation level were quantified by methylation‐sensitive high resolution melting. VEGFA165a was overexpressed and VEGFA165b level was reduced in tumors. RBM10 level was reduced in tumors. RBM10 level was negatively correlated with VEGFA165a level and positively correlated with VEGFA165b level in tumors. Using HEC‐1‐A and RL95‐2 cells, we confirmed that VEGFA165a/b expressed pattern was controlled by RBM10. MALAT1 level was increased in tumors but not involved in VEGFA alternative splicing. Reduced H3K27Ac level and increased DNA methylation in the promoter region controlled RBM10 expression in tumors. VEGFA alternative splicing in endometrial cancer was regulated by RBM10, the expression of which was controlled by histone acetylation and DNA methylation.

Differential expression of host oncogenes in human papillomavirus‐associated nasopharyngeal and cervical epithelial cancers

AbstractHuman papillomavirus (HPV)‐related cervical and nasopharyngeal cancers differ in molecular mechanisms underlying the oncogenic processes. The disparity may be attributed to differential expression of oncoproteins. The current study investigated the host oncogenes expression pattern in HPV‐associated cervical and nasopharyngeal cancer. Formalin‐fixed paraffin‐embedded tissues originating from the nasopharyngeal and cervical regions were screened using Hematoxylin and Eosin staining. Genomic DNA and total RNA were extracted from confirmed cancer biopsies and non‐cancer tissues (NC). HPV was detected by PCR using MY09/GP5+/6+ primers. Protein expression levels of AKT, IQGAP1, and MMP16 in HPV‐infected cancers and controls were determined by immunohistochemistry. RT‐qPCR was used to profile mRNAs of the oncogenes. AKT and IQGAP1 proteins were highly expressed in the epithelial cancers compared with the non‐cancer tissues (p < 0.05). IQGAP1 and MMP16 mRNAs level was significantly higher in the cancers than in the NC (p < 0.05), but not AKT mRNA levels. MMP16 protein was ubiquitously expressed in all tissues. AKT mRNA level was significantly elevated in CC compared with NPC (p < 0.001). However, the difference in AKT, IQGAP1 and MMP16 proteins level between CC and NPC was not significant (p > 0.05). The oncoproteins expression level between the HPV‐positive and HPV‐negative cancer biopsies showed no significant difference (p < 0.05). Current study reports AKT but not IQGAP1 and MMP16 mRNAs differentially expression in cervical and nasopharyngeal cancers, independent of HPV infection status.

Long Noncoding RNA TOB1‐AS1 Represses Cervical Cancer Cell Proliferation, Invasion, and Migration via the MicroRNA‐27a‐3p/Thioredoxin‐Interacting Protein Molecular Axis

ABSTRACT Cervical cancer (CC) remains a major global health concern, particularly due to its aggressive nature and limited treatment options in advanced stages. Long noncoding RNA (lncRNA) TOB1‐AS1 has been proposed as a tumor suppressor, yet its regulatory mechanism in CC remains unclear. This study aimed to elucidate the role of TOB1‐AS1 in CC progression through the miR‐27a‐3p/thioredoxin‐interacting protein (TXNIP) molecular axis. Functional gain‐ and loss‐of‐function assays were conducted to assess the effects of TOB1‐AS1, miR‐27a‐3p, and TXNIP on cell proliferation, invasion, migration, and apoptosis. RT‐qPCR, Western blotting, dual‐luciferase reporter assays, and in vivo xenograft models were used to validate interactions and phenotypic outcomes. TOB1‐AS1 was found to be downregulated in CC cells. Its overexpression suppressed proliferation, invasion, and migration, while enhancing apoptosis. Mechanistically, TOB1‐AS1 functioned as a competing endogenous RNA (ceRNA) by sponging miR‐27a‐3p, thereby restoring TXNIP expression. Modulating miR‐27a‐3p or TXNIP levels partially reversed the effects of TOB1‐AS1. In vivo, TOB1‐AS1 overexpression significantly inhibited tumor growth and altered miR‐27a‐3p and TXNIP expression profiles. These findings suggest that lncRNA TOB1‐AS1 acted as a ceRNA of miR‐27a‐3p to upregulate TXNIP, thereby suppressing CC cell proliferation, invasion and migration.

MYB Activates the Hedgehog Signaling Pathway to Repress Natural Killer Cytotoxicity in Cervical Cancer

ABSTRACT Natural killer (NK) cells present in the tumor microenvironment serve as a critical line of defense against various malignancies, including cervical cancer. While MYB is known to drive malignancy progression, its influence on NK cell activity remains poorly understood. This study aimed to elucidate the role of MYB in regulating NK cell cytotoxicity and its underlying mechanism in cervical cancer cells. MYB expression in cervical cancer tissues and cells was analyzed using bioinformatics and qRT‐PCR. Cell viability was assessed via CCK‐8 assay, while NK cell‐mediated killing of cervical cancer cells was evaluated through cytotoxicity assays. The expression levels of cytotoxic factors (IFN‐ γ and TNF‐ α ) were measured by ELISA, whereas perforin and granzyme B were detected via immunofluorescence. Apoptosis was analyzed using flow cytometry. To investigate the impact of MYB on the hedgehog signaling pathway, the expression levels of related factors (PTCH1, Gli1, and Gli2) were assessed using qRT‐PCR and Western blot. Bioinformatics and qRT‐PCR analyses revealed MYB overexpression in cervical cancer. Signaling pathway prediction indicated MYB enrichment in cytotoxic signaling pathways. Functional experiments demonstrated that MYB overexpression activated the hedgehog signaling pathway, thereby suppressing NK cell cytotoxicity in cervical cancer. Rescue experiments using the hedgehog signaling inhibitor GANT58 attenuated the suppressive effect of MYB overexpression on NK cytotoxicity. In summary, MYB inhibited NK cell cytotoxicity by activating the hedgehog signaling pathway in cervical cancer, suggesting its potential as a novel diagnostic marker and immunotherapeutic target.

LINC01871 facilitates cervical cancer cell migration and immune escape by targeting miR‐873‐3p/MAP3K2 axis

AbstractCervical cancer (CC) poses a significant threat to women's health and lives worldwide. Emerging evidence indicates that long noncoding RNA LINC01871 is closely associated with immune regulation in CC patients. However, its specific role and underlying mechanisms in CC remain poorly understood. In this study, we examined the expression levels and subcellular localization of LINC01871 in CC cell lines. Functional assays, including transwell migration and invasion assays as well as macrophage phagocytosis assays, were conducted to estimate CC cell invasiveness, migration, and immune response. Western blotting was performed to assess the protein expression of markers associated with epithelial–mesenchymal transition (EMT), immunity, and MAPK signaling. A luciferase reporter assay was used to confirm the interactions between LINC01871, miR‐873‐3p, and MAP3K2. Additionally, a xenograft mouse model was employed to investigate the in vivo effects of LINC01871 on CC progression. Our results revealed that LINC01871 is highly expressed and predominantly localized in the cytoplasm of CC cell lines. LINC01871 knockdown significantly suppressed CC cell invasion, migration, EMT, and immune escape in vitro and reduced tumor growth in vivo. Mechanistically, LINC01871 was found to interact with miR‐873‐3p, leading to the upregulation of MAP3K2 and subsequent activation of MAPK signaling. Furthermore, MAP3K2 overexpression rescued the inhibitory effects of LINC01871 silencing on the malignant behaviors of CC cells. In conclusion, LINC01871 facilitates CC metastasis by driving EMT and modulating the immune response via the miR‐873‐3p/MAP3K2/MAPK signaling pathway.

LncRNA GAS5 inhibits the proliferation and invasion of ovarian clear cell carcinoma via the miR‐31‐5p/ARID1A axis

AbstractTo investigate the role of the lncRNA growth arrest special 5 (GAS5) in ovarian clear cell carcinoma (OCCC), we measured the expression of GAS5 and miR‐31‐5p in OCCC tissue samples and OCCC cell lines using RT‐qPCR. MTT and colony formation assays were used to measure cell viability and colony formation ability. Cell invasion was determined by Transwell assays. The binding between GAS5 and miR‐31‐5p as well as miR‐31‐5p and ARID1A was determined by dual‐luciferase reporter assays. The ARID1A protein levels were detected using western blotting. Kaplan–Meier curves were used for the analysis of the 5‐year survival rate of patients with OCCC. GAS5 and ARID1A levels were significantly decreased, while miR‐31‐5p levels were strongly elevated in the OCCC tissues and cell lines. Patients with lower GAS5/ARID1A levels had shorter overall survival times. Overexpression of GAS5 or inhibition of miR‐31‐5p suppressed cell viability and invasion of OCCC cells and upregulated the protein levels of ARID1A. Moreover, overexpression of miR‐31‐5p reversed the effects of overexpression of GAS5. Cotransfection with pcDNA3.1‐GAS5 and miR‐31‐5p inhibitor led to the lowest cell viability and cell invasion rates. A dual‐luciferase reporter assay was performed to confirm the target relationship between GAS5 and miR‐31‐5p, as well as between miR‐31‐5p and ARID1A. LncRNA GAS5 inhibited cell viability and invasion of OCCC through activation of ARID1A by sponging miR‐31‐5p.

Hypoxia Triggers ALYREF ‐Mediated m 5 C Methylation of KIF20A to Activate KIF20A / BUB1 for Generating Ferroptosis Resistance in Cervical Cancer Cells

ABSTRACT Ferroptosis resistance is a key player in cervical cancer (CC) development. Hypoxia is a negative factor affecting CC treatment by inducing ferroptosis resistance. Our study aimed to investigate the detailed mechanisms of hypoxia‐induced ferroptosis resistance in CC cells. The mRNA and protein levels were assessed using RT‐qPCR and western blot. Immunofluorescence staining was used to analyze the co‐localization of Budding uninhibited by benzimidazole 1 (BUB1) and Kinesin family member 20A (KIF20A) in CC cells. Fe 2+ , MDA, and GSH levels were measured by commercial kits. Cell viability was determined by CCK‐8. The molecular interactions were analyzed by RIP or Co‐IP. MeRIP was adopted to measure the m 5 C level of KIF20A. We found that hypoxia facilitated ferroptosis resistance in CC cells. Hypoxia led to the upregulation of KIF20A, and KIF20A knockdown weakened hypoxia‐mediated ferroptosis resistance in CC cells. Mechanistically, Aly/REF export factor (ALYREF) stabilized KIF20A mRNA stability via m 5 C modification. In addition, KIF20A upregulated BUB1 in CC cells by directly interacting with BUB1. Rescue experiments indicated that BUB1 overexpression partially reversed the inhibitory effect of KIF20A knockdown on hypoxia‐mediated ferroptosis resistance in CC cells. In conclusion, hypoxia triggers ALYREF‐mediated m 5 C methylation of KIF20A to activate the KIF20A/BUB1 axis, thereby inducing ferroptosis resistance in CC cells.

Repressing IRS1/2 by NT157 inhibits the malignant behaviors of ovarian cancer through inactivating PI3K/AKT/mTOR pathway and inducing autophagy

AbstractInsulin receptor substrate 1 and 2 (IRS1/2) have been found involved in many cancers development and their inhibitors exert significant tumor‐suppressive effects. Here, we tried to explore the function of NT157, an IGF1R‐IRS1/2 inhibitor, in ovarian cancer. We treated ovarian cancer cells with varying doses of NT157. The MTT assay was employed to evaluate cell proliferation and colony formation assay was used for detecting colony‐forming ability. TUNEL assay was adopted to test cell apoptosis. Cell invasion was checked by the Transwell assay. The expression of apoptosis‐related proteins, autophagy markers, IRS1/2, and PI3K/AKT/mTOR pathway was compared by Western blot, immunofluorescence, or qRT‐PCR. As indicated by the data, NT157 abated the viability, proliferation, and induced autophagy of ovarian cancer cells. Overexpressing IRS1/2 attenuated the tumor‐suppressive effect of NT157 and heightened the PI3K/AKT/mTOR pathway activation. Inhibition of the PI3K/AKT/mTOR pathway enhanced the tumor‐suppressive effect of NT157 and facilitated NT157‐mediated autophagy. However, the autophagy inhibitor 3‐MA partly reversed NT‐157‐mediated antitumor effects. In conclusion, this study disclosed that NT157 suppressed the malignant phenotypes of ovarian cancer cells by inducing autophagy and hampering the expression of IRS1/2 and PI3K/AKT/mTOR pathway.

LDLR promotes autophagy‐mediated cisplatin resistance in ovarian cancer associated with the PI3K/AKT/mTOR signaling pathway

AbstractAutophagy is one of the underlying causes of resistance to many antitumor drugs, including cisplatin (DDP). The low‐density lipoprotein receptor (LDLR) is a regulator of ovarian cancer (OC) progression. However, whether LDLR regulates DDP resistance in OC via autophagy‐related pathways remains unclear. LDLR expression was measured by quantitative real‐time PCR, western blot (WB) and IHC staining. A Cell Counting Kit 8 assay was employed to evaluate DDP resistance and cell viability, and flow cytometry was used to assess apoptosis. WB analysis was employed to evaluate the expression of autophagy‐related proteins and PI3K/AKT/mTOR signaling pathway proteins. The autophagolysosomes and the fluorescence intensity of LC3 were observed by transmission electron microscopy and immunofluorescence staining, respectively. A xenograft tumor model was established to explore the role of LDLR in vivo. LDLR was highly expressed in OC cells, which was correlated with disease progression. In DDP‐resistant OC cells, high LDLR expression was related to DDP resistance and autophagy. Downregulation of LDLR repressed autophagy and growth in DDP‐resistant OC cell lines by activating the PI3K/AKT/mTOR pathway, and these effects were eliminated by an mTOR inhibitor. In addition, LDLR knockdown also reduced OC tumor growth by suppressing autophagy associated with the PI3K/AKT/mTOR pathway. LDLR promoted autophagy‐mediated DDP resistance in OC associated with the PI3K/AKT/mTOR pathway, indicating that LDLR might be a new target to prevent DDP resistance in OC patients.

Promotive role of eukaryotic translation initiation factor 4A isoform 3 in ovarian cancer cell growth and aerobic glycolysis through the pyruvate dehydrogenase kinase 4 signaling

AbstractOvarian cancer (OC) represents one of the most detrimental gynecological malignancies. RNA‐binding protein eukaryotic translation initiation factor 4A isoform 3 (EIF4A3) is well‐regarded as a definitive oncogene that contributes to the development of multiple malignant tumors. This study sought to elucidate the molecular mechanism of EIF4A3 in OC growth and aerobic glycolysis by regulation of pyruvate dehydrogenase kinase 4 (PDK4) mRNA stability. We determined the EIF4A3 and PDK4 expression levels in OC cell lines and normal ovarian epithelial cells, and subsequently evaluated the cell viability and colony formation by cell counting kit‐8 and colony formation assays. The degree of cell aerobic glycolysis was evaluated by measurements of lactic acid production, glucose intake, adenosine triphosphate level, extracellular oxygen consumption, and protein levels of pyruvate kinase isozymes M2 and hexokinase‐2. Afterwards, we verified the binding of EIF4A3 and PDK4 mRNA via RNA immunoprecipitation, and determined the mRNA stability after actinomycin D treatment. Finally, a series of rescue experiments was performed with pcDNA3.1‐PDK4. EIF4A3 and PDK4 were upregulated in OC cells. Silencing EIF4A3 obstructed cell proliferation and aerobic glycolysis, while the same was annulled by EIF4A3 overexpression. Mechanically, EIF4A3 could bind to PDK4 mRNA to stabilize its mRNA and upregulate its protein levels. PDK4 overexpression inverted the inhibitory role of silencing EIF4A3 in proliferation and aerobic glycolysis. Overall, our findings highlighted that EIF4A3 induced OC progression by stabilizing PDK4 mRNA.

Immature renal tissue occurring in cystic teratoma of ovary

Exosomal circNFIX promotes angiogenesis in ovarian cancer via miR‐518a‐3p/TRIM44 axis

AbstractOvarian cancer (OC) is a gynecological cancer with high mortality. OC‐derived exosomal circRNAs can regulate angiogenesis. This study aims to explore the role and mechanism of exosomal circRNA nuclear factor I X (CircNFIX) derived from OC cells in angiogenesis. Quantitative real‐time polymerase chain reaction was employed to evaluate the levels of circNFIX, miR‐518a‐3p, and tripartite motif protein 44 (TRIM44) in OC and adjacent tissues. Exosomes from the ovarian surface epithelial cell (HOSEpiC) and OC cells (SKOV3 or OVCAR3) were isolated by differential centrifugation. Exosomes were cocultured with the human umbilical vein endothelial cells (HUVECs). The angiogenesis capacity was analyzed by Tube formation assay. 3‐(4,5‐dimethyl‐2‐thiazolyl)‐2,5‐diphenyl‐2‐H‐tetrazolium bromide (MTT) and Transwell assays were used to determine the cell viability and migration ability. The dual‐luciferase report, RNA immunoprecipitation (RIP), and RNA pull‐down assays were applied to validate the gene's interaction. CircNFIX and TRIM44 expression were higher and miR‐518a‐3p was lower in OC tissues than in the adjacent tissues. Upregulated circNFIX and TRIM44 were significantly correlated with the tumor size and International Federation of Gynecology and Obstetrics (FIGO) stage of OC patients. HUVECs treated OC‐derived exosomes had higher proliferation, migration, and angiogenesis capacities than the control group. While OC‐derived exosomal circNFIX silencing restrained HUVECs' proliferation, migration, and angiogenesis, compared with the OC‐derived exosomes group. OC‐derived exosomal circNFIX positively regulated TRIM44 expression by targeting miR‐518a‐3p in HUVECs. OC‐derived exosomal circNFIX promoted angiogenesis by regulating the Janus‐activated kinase/signal transducer and activator of transcription 1 (JAK/STAT1) pathway via miR‐518a‐3p/TRIM44 axis in HUVECs.

MiR‐10a‐5p restrains the aggressive phenotypes of ovarian cancer cells by inhibiting HOXA1

AbstractMicroRNAs (miRNAs) are dysregulated in human ovarian carcinoma (OC). But the mechanism underlying miR‐10a‐5p in regulating the progression of OC need deeply explored. In the current study, we observed that miR‐10a‐5p was down‐expressed in OC samples and OC cell lines. In addition, miR‐10a‐5p restrained the viability, colony formation, migration ability and invasiveness of OC cells. We further ascertained Homeobox A1 (HOXA1) was a downstream gene of miR‐10a‐5p. Furthermore, HOXA1 was distinctly upregulated in OC samples. Finally, upregulation of HOXA1 abolished the suppressive effects of miR‐10a‐5p on OC cells. These observations suggested that miR‐10a‐5p suppressed the aggressive phenotypes of OC cells via regulating HOXA1.

SP1‐induced lncRNA DANCR contributes to proliferation and invasion of ovarian cancer

AbstractTranscription factor SP1 could manipulate pathways involved in ovarian cancer progression. LncRNAs are involved in SP1‐mediated tumorigenesis. LncRNA DANCR could promote metastasis of ovarian cancer. However, the regulatory function and involvement of SP1‐induced lncRNA DANCR in ovarian cancer remain elusive. Data from this study showed that SP1 was up‐regulated in ovarian cancer tissues and cells (CAOV3, SKOV3, A2780), and SP1 could bind to the promoter region of DANCR through chromatin immunoprecipitation and leuciferase activity assays. Therefore, DANCR was transcriptionally induced by SP1 in ovarian cancer tissues and cells (CAOV3, SKOV3, A2780). Functionally, reduced expression of DANCR suppressed cell viability, migration and invasion of CAOV3, while enhanced DANCR expression contributed to SKOV3 growth. Over‐expression of SP1 reversed the suppressive effects of DANCR interference on ovarian cancer progression. In conclusion, SP1‐induced DANCR contributed to oncogenic potential of ovarian cancer, suggesting a promising therapeutic target for ovarian cancer.

Long noncoding RNA HOTTIP promotes the metastatic potential of ovarian cancer through the regulation of the miR‐615‐3p/SMARCE1 pathway

AbstractUpregulation of lncRNA HOXA transcript at the distal tip (HOTTIP) plays important roles in cancer progression. Nevertheless, its functions in the growth and metastasis of ovarian carcinoma are unknown. In this study, we demonstrated overexpression of HOTTIP in ovarian cancer cell lines and clinical tissues. Further, we showed that higher level of HOTTIP was associated with poor survival of ovarian cancer patients. Notably, HOTTIP silencing restrained proliferation, migration, and invasiveness of ovarian carcinoma cells. On the other hand, upregulation of HOTTIP remarkably exacerbated the aggressive traits of ovarian carcinoma cells. In addition, HOTTIP served as a sponge of miR‐615‐3p to upregulate SMARCE1 level. Further, upregulation of miR‐615‐3p or downregulation of SMARCE1 reversed the carcinogenic impacts of HOTTIP in ovarian cancer. HOTTIP and miR‐615‐3p expression levels in ovarian cancer cells were negatively correlated, whereas HOTTIP and SMARCE1 expression levels were positively correlated. In nude mice, downregulation of HOTTIP reduced cell growth in vivo. In summary, lncRNA HOTTIP promotes the growth and metastatic phenotypes of ovarian cancer via regulating miR‐615‐3p/SMARCE1 pathway.

miR‐744‐5p inhibits cellular proliferation and invasion via targeting ARF1 in epithelial ovarian cancer

AbstractmiR‐744‐5p has been demonstrated to play an important role in cancer progression. However, the functions of miR‐744‐5p in epithelial ovarian cancer (EOC) are not well clarified. In this study, our aim was to investigate the role of miR‐744‐5p and its underlying molecular mechanism in cell progression of EOC. EOC clinical tissues and matched adjacent ovarian epithelial tissues were collected from 18 patients. Tissues and cell lines were analyzed by qPCR or Western blot to investigate the expression of miR‐744‐5p and ARF1 in EOC. Cell proliferative capacity was assessed by CCK8 and colony formation assays. Wound healing and transwell assays were performed to evaluate cell migration and invasion. The potential binding relation between miR‐744‐5p and IRF1 was demonstrated by dual luciferase report assay. The results showed that expression of miR‐744‐5p was low in EOC clinical tissues and cells. Overexpression of miR‐744‐5p inhibited proliferation, migration, and invasion of EOC cells. Further mechanistic study identified that ARF1 is a target of miR‐744‐5p, which is negatively correlated with the expression of miR‐744‐5p, and overexpression of ARF1 could reverse the inhibition of miR‐744‐5p on the proliferation, migration, and invasion of EOC cells. Taken together, our results indicated that miR‐744‐5p attenuated EOC progression via targeting IRF1, providing potential guidance for the clinical treatment of ovarian cancer.

Collision of sclerosing stromal tumor and signet‐ring stromal tumor of the ovary: Report of the first case

Silencing of SBF2‐AS1 inhibits cell growth and invasion by sponging microRNA‐338‐3p in serous ovarian carcinoma

AbstractLong noncoding RNA SET‐binding factor 2 (SBF2) antisense RNA 1 (AS1) is associated with the growth and metastasis of multiple cancer types, but its biological roles in serous ovarian carcinoma (SOC) remain unclear. In this study, the aberrant upregulation of SBF2‐AS1 is detected in SOC after analysis of differentially expressed genes between SOC tissues and normal fallopian tubes from the public Gene Expression Omnibus (GEO) database. We determine that knockdown of SBF2‐AS1 inhibits SOC cell proliferation and invasion by sponging miR‐338‐3p. MiR‐338‐3p acts as a tumor suppressor in SOC, and E26 transformation specific‐1 (ETS1) is identified as a potential target of miR‐338‐3p regulation. Furthermore, SBF2‐AS1 could modulate ETS1 by operating as a competing endogenous RNA for miR‐338‐3p. This finding elucidates a new mechanism for SBF2‐AS1 in SOC development and provides a potential target for SOC therapeutic intervention.

Long non‐coding RNA‐H19 promotes ovarian cancer cell proliferation and migration via the microRNA‐140/Wnt1 axis

AbstractTo explore the effect and underlying molecular mechanism of long non‐coding RNA (lncRNA)‐H19 on ovarian cancer (OC) cells, a total of 41 cases of OC and adjacent normal tissues were collected. H19 and microRNA (miR)‐140 expressions in OC tissues and cells were detected using quantitative real‐time polymerase chain reaction (qRT‐RCR). The correlation between H19 expression and prognosis of OC patient was analyzed. siRNA (si)‐H19 and si‐negative control (NC) were transfected into OC cells. Cell proliferation was checked by cell counting kit‐8 assay and colony formation assay, and cell migration and invasion were analyzed via Transwell assay. The targeted binding relationship between H19 and miR‐140 was predicted and verified, miR‐140 downstream gene was predicted and Wnt1 was screened out. The impact of in‐miR‐140 on the si‐H19‐induced decreased OC cell proliferation and migration was evaluated. H19 expression was upregulated in OC tissues and cells, and its overexpression was associated with a poor prognosis of OC. si‐H19 remarkably reduced OC cell proliferation and migration. H19 upregulated Wnt1 expression through targeting miR‐140 in OC cells. Altogether, miR‐140 was notably downregulated in OC, and in‐miR‐140 partially inhibited the si‐H19‐induced decrease of OC cell proliferation and migration. H19 competitively bound to miR‐140 to upregulate Wnt1, thereby promoting OC cell proliferation and migration.

CircNFIX stimulates the proliferation, invasion, and stemness properties of ovarian cancer cells by enhancing SH3RF3 mRNA stability via binding LIN28B

AbstractWe aimed to study the regulatory roles and mechanism of circular nuclear factor IX (circNFIX) in cancer growth and stemness properties of ovarian cancer (OC). CircNFIX and SH3RF3 levels in OC tissues and cells were tested by quantitative real‐time PCR. RNase R treatment quantified circNFIX RNA stability. Molecular interaction among circNFIX, LIN28B, and SH3RF3 was predicted by bioinformatics software and validated through RNA immunoprecipitation (RIP) assay. The gain‐ or loss‐experiments of circNFIX on capabilities of metastasis and stemness in vitro were assessed using Cell Counting Kit‐8, Transwell, western blot, and sphere‐formation assays. CircNFIX and SH3RF3 were markedly elevated in OC tissues and OC cells. Knocking down circNFIX repressed the proliferation, migration, invasion, and stemness properties of A2780 and SKOV3 cells. The RIP assay verified the direct binding relationship between LIN28B, circNFIX, and SH3RF3. Additionally, overexpression of circNFIX elevated the SH3RF3 expression, while this effect was reversed by LIN28B silence. Rescue experiments demonstrated that the overexpression of SH3RF3 reversed the knockdown of circNFIX on OC cells' proliferation, metastasis, and stemness properties. CircNFIX improved the mRNA stability and translation of SH3RF3 via recruiting LIN28B, thus promoting the proliferation, invasion, and stemness properties of OC cells in vitro.

MFAP2 aggravates tumor progression through activating FOXM1/β‐catenin‐mediated glycolysis in ovarian cancer

AbstractOvarian cancer is one of the most common gynecological tumors that seriously endanger the health and quality of life of women. Microfibril‐associated protein 2 (MFAP2) has been demonstrated to play crucial roles in the development of multiple tumors. However, the function of MFAP2 in ovarian cancer remains unclear. In this study, we found that MFAP2 was upregulated in ovarian cancer and cells and was positively correlated with FOXM1 and glycolysis‐related genes. The results of Cell Count Kit‐8, colony formation, and flow cytometry assays indicated that MFAP2 promoted cell proliferation. In addition, MFAP2 promotes cell proliferation, glucose uptake, lactate production; increases ATP levels, extracellular acidification ratio, and oxygen consumption ratio in ovarian cancer cells and increases the expression of glycolytic proteins. Further mechanistic analysis suggests that MFAP2 promotes FOXM1/β‐catenin‐mediated glycolysis signaling in ovarian cancer cells. Knockdown of MFAP2 inhibits ovarian cancer xenograft tumor growth and expression of Ki‐67, MFAP2, FOXM1, GLUT1, HK2, and β‐catenin in mice. In conclusion, MFAP2 promotes cell proliferation and glycolysis by modulating the FOXM1/β‐catenin signaling pathway in ovarian cancer, which may offer a fresh insight into the treatment of ovarian cancer in the glycolysis pathway.

MicroRNA‐324‐5p affects the radiotherapy response of cervical cancer via targeting ELAV‐like RNA binding protein 1

AbstractCervical cancer (CC) seriously threatens the health of women. Radiation therapy (RT) is the major treatment for CC. However, the recurrent CC can acquire resistance to RT. Thus, it is necessary to find a new method for reversing RT resistance in CC. It has been reported that miR‐324‐5p can suppress the progression of multiple cancers. However, whether it can reverse resistance to RT in CC remains unclear. qRT‐PCR and Western blotting were used to detect gene and protein expression in CC cells, respectively. Cell proliferation was tested by CCK‐8 assay and colony formation assay. In addition, cell apoptosis was detected by flow cytometry. Transwell assays were performed to detect cell migration. Dual luciferase reporter assay and TargetScan were used to explore the targets of microRNA‐324‐5p (miR‐324‐5p). MiR‐324‐5p was downregulated in CC cells. Overexpression of miR‐324‐5p sensitized CC cells to RT. In addition, miR‐324‐5p mimics significantly induced apoptosis and inhibits the migration of CC cells in the presence of 137Cs ionizing radiation. Furthermore, miR‐324‐5p sensitized CC cells to ionizing radiation by targeting ELAV‐like RNA binding protein 1 (ELAVL1). MiR‐324‐5p overexpression affects the radiotherapy response of CC by targeting ELAVL1, which may serve as a new target for the treatment of CC.

Tumor suppressor miR‐192‐5p targets TRPM7 and inhibits proliferation and invasion in cervical cancer

AbstractCervical cancer is the fourth highest mortality cancer among women worldwide. Many researchers have discovered the major anticancer role of miR‐192‐5p. However, no study has revealed the effect of miR‐192‐5p on cervical cancer and its molecular mechanism. Therefore, in this study, we aimed to explore the role of miR‐192‐5p in proliferation, invasion of cervical cancer, and its regulatory mechanism. Firstly, the expression level of miR‐192‐5p was examined by real‐time quantitative polymerase chain reaction. Cell counting kit‐8 analysis was applied to detect the proliferation of transfected Caski and SiHa cells. Flow cytometry assay was applied to detect the apoptosis of transfected Caski and SiHa cells. Our result showed that miR‐192‐5p restrained cervical cancer cell proliferation and induced apoptosis. Then we employed wound healing and transwell assays to analyze the migration and invasion abilities of Caski and SiHa cells in vitro. The results showed that miR‐192‐5p had an inhibitory effect on cervical cancer migration and invasion. The results of in vivo experiment demonstrated that miR‐192‐5p also inhibited tumor development in nude mice. We further detected that the binding of transient receptor potential melastatin‐subfamily member 7 (TRPM7) to miR‐192‐5p using bioinformatic methods and dual‐luciferase reporter assay. Finally, we found that TRPM7 overexpression reversed the inhibitory effects of miR‐192‐5p on proliferation, migration, and invasion on cervical cancer cells. In conclusion, the findings of the present study revealed that miR‐192‐5p performs an inhibitory role in cervical cancer proliferation and invasion by targeting TRPM7.

Inhibiting microRNA‐301b suppresses cell growth and targets RNF38 in cervical carcinoma

AbstractIt has been reported microRNA‐301b (miR‐301b) was involved in the tumorigenesis of some cancers, but it has not been investigated in cervical carcinoma yet. In this study, miR‐301b was found significantly upregulated in cervical carcinoma, and patients with high miR‐301b expression had a shorter overall survival. When miR‐301b was knocked down in cervical carcinoma cells, the cell growth could be significantly abolished. Our further studies showed miR‐301b targeted RNF38, and inhibited its expression in cervical carcinoma cells. Moreover, RNF38 was found downregulated in cervical carcinoma, and miR‐301b expression in cervical tissues was found negatively correlated with RNF38 expression. In addition, overexpression of RNF38 significantly inhibited cervical carcinoma cell growth, but overexpression of miR‐301b suppressed RNF38‐induced cell growth inhibition in cervical carcinoma. Collectively, this study suggested miR‐301b could be a novel target for cervical carcinoma treatment.

Inhibiting ubiquitin conjugating enzyme E2 N by microRNA‐590‐3p reduced cell growth of cervical carcinoma

AbstractThe ubiquitin conjugating enzyme E2 N (UBE2N) has been reported to be involved in the tumorigenesis of several tumors, but its function in cervical carcinoma has not been investigated yet. In the present study, UBE2N was found elevated in cervical carcinoma, and patients with high UBE2N had a shorter overall survival than patients with low expression. Additionally, knockdown of UBE2N decreased the activation of MEK1/2 and p38 in cervical carcinoma cells, and UBE2N knockdown also markedly inhibited cervical carcinoma cell growth. Our further studies found that microRNA‐590‐3p (miR‐590‐3p) was significantly decreased in cervical carcinoma, and patients with high miR‐590‐3p had a longer overall survival than patients with low expression. Moreover, miR‐590‐3p expression was found negatively correlated with UBE2N expression in cervical carcinoma, and our further studies showed that miR‐590‐3p targeted UBE2N and inhibited its expression in cervical carcinoma. Overexpression of miR‐590‐3p could inhibit cervical carcinoma cell growth, but enhanced UBE2N could rescue miR‐590‐3p‐induced cell growth inhibition in cervical carcinoma. This study indicated that targeting miR‐590‐3p/UBE2N axis could be a potential strategy for the treatment of cervical carcinoma.

miR‐636 represses cell survival by targeting CDK6/Bcl‐2 in cervical cancer

AbstractCervical cancer is widely known as one of the most common types of cancer diagnosed in women, and microRNAs (miRNAs) has been characterized as an important regulator in tumor progression, such as cervical cancer. MiR‐636 was found to play a tumor suppressor role in hepatocellular carcinoma tumorigenesis. However, the tumorigenic mechanism of miR‐636 on cervical cancer has not yet been found. In the present study, we first found that miR‐636 was significantly downregulated in cervical cancer tissues and cell lines. in vitro gain‐ and loss‐of‐function assays revealed that overexpression of miR‐636 inhibited cell proliferation and induced cell apoptosis, while knockdown of miR‐636 reversed the effect on cervical tumorigenesis. Furthermore, cyclin‐dependent kinase 6 (CDK6) and B‐cell lymphoma 2 (Bcl‐2) were characterized as targets of miR‐636. Notably, overexpression of CDK6 or Bcl‐2 could reverse the inhibitory effect of miR‐636 on cervical cancer progression. Mechanistically, miR‐636 repressed cell survival by targeting CDK6/Bcl‐2 in cervical cancer, which may be the underlying mechanism of miR‐636‐inhibited cervical progression. In conclusion, our findings clarified the biologic significance of miR‐636/CDK6/Bcl‐2 axis in cervical cancer progression and suggested the potential therapeutic target ability of miR‐636 in treatment of cervical cancer.

Significance of desmoglein‐2 on cell malignant behaviors via mediating MAPK signaling in cervical cancer

AbstractDesmoglein‐2 (DSG2) is an integral component of desmosomes, maintaining cell‐cell adhension in multiple cancers. It has been well studied in epithelial cells, cardiomyocytes and primary prostate cancer, colon cancer, skin squamous cell carcinoma, except for cervical cancer. Hence, we performed this study to examine the function of DSG2 in cervical cancer. We used TCGA and Oncomine databases to assess the expression level of DSG2 in cervical cancer cases. Kaplan‐Meier method with log‐rank test was utilized to plot overall survival (OS) curve. The reverse transcription‐quantitative polymerase chain reaction (qRT‐PCR) and western blotting were performed to detect the expression of DSG2 in cells. Cell Counting Kit‐8 (CCK‐8), wound‐healing analysis, and transwell assay were carried out to examine proliferation, migration, and invasion of cells. A higher level of DSG2 in cervical cancer was associated with lower OS rate. Knockdown of DSG2 inhibited cervical cancer cell proliferation, migration, and invasion, while DSG2 enhancement promoted cell proliferation, migration, and invasion. Moreover, the proteins expression of p‐MEK and p‐ERK that are required for mitogen‐activated protein kinases (MAPK) pathway were downregulated after reducing DSG2. In conclusion, these findings illustrated the importance of DSG2 in cervical cancer development and cell behaviors by mediating MAPK signaling pathway, suggesting DSG2 maybe a novel therapeutic target in control of cervical cancer.

PAX9 functions as a tumor suppressor gene for cervical cancer via modulating cell proliferation and apoptosis

AbstractTo investigate the effect of PAX9 on the progression of cervical cancer (CC). PAX9 expression was quantified in CC tissues and adjacent normal tissues, as well as human CC cell lines and human cervical epithelial cells (HCerEpiC). PAX9‐overexpression lentiviral vectors were transfected into CC cell lines, followed by the measurement of proliferation and apoptosis and the quantification of apoptosis‐related proteins. In vivo, mice were subcutaneously injected with CaSki cells transfected with PAX9‐overexpression lentiviral vectors and control vectors. Then, the volume and weight of tumors were measured followed by hematoxylin and eosin (HE) staining, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining, and immunohistochemistry. PAX9 expression in the CC tissues was lower than that in the adjacent normal tissues, which was correlated with the FIGO stage, tumor size, infiltration depth, parametrium invasion, lympho‐vascular space invasion tumor‐positive lymph nodes, and prognosis. Furthermore, PAX9 in CC cell lines was also lower than in HCerEpiC. PAX9 inhibits the CC cell proliferation and promotes the apoptosis, with the up‐regulations of caspase‐3, poly(ADP‐ribose) polymerase (PARP), and Bax and the down‐regulation of Bcl‐2. In vivo experiments demonstrated that in the PAX9 group, the tumor weight and volume were lower than those in the vector group accompanying the decreased Ki‐67, cleaved‐caspase‐3, and Bax expressions and the increased TUNEL and Bcl‐2 expression. PAX9 was lowly expressed in the CC tissues and associated with the clinicopathological characteristics and prognosis. PAX9 could inhibit proliferation of CC cell lines and promote the apoptosis, thus suppressing the tumor growth in vivo, indicating its potential therapeutic role for CC treatment.

Circular RNA circ_0023404 serves as a miR‐636 sponge to promote malignant behaviors in cervical cancer cells through upregulation of CYP2S1

AbstractCervical cancer is the most common malignant gynecological tumor. Circular RNA (circRNA) circ_0023404 is reported to be upregulated in cervical cancer cells. This aim is to explore the role and mechanism of circ_0023404 in cervical cancer. circ_0023404, microRNA‐636 (miR‐636), and cytochrome P450 2S1 (CYP2S1) levels were detected by real‐time quantitative polymerase chain reaction (RT‐qPCR). Cell proliferation, migration, invasion, and apoptosis were detected by 3‐(4,5‐dimethyl‐2‐thiazolyl)‐2,5‐diphenyl‐2‐H‐tetrazolium bromide (MTT) assay, 5‐ethynyl‐2′‐deoxyuridine (EDU) assay, colony formation assay, transwell assay, and cytometry assay. Protein levels of cyclin D1, matrix metallopeptidase 9 (MMP9), Bcl‐2‐associated X protein (Bax), and CYP2S1 were examined by western blot assay. The binding relationship between miR‐636 and circ_0023404 or CYP2S1 was predicted by Circinteractome or targetscan, and then verified by a dual‐luciferase reporter assay and RNA pull‐down assay. circ_0023404 and CYP2S1 expression were increased, and miR‐636 was decreased in cervical cancer tissues and cells. Moreover, circ_0023404 knockdown could repress proliferation, migration, invasion, and promote apoptosis of cervical cancer cells in vitro. Mechanically, circ_0023404 could regulate CYP2S1 expression by sponging miR‐636. circ_0023404 silencing could attenuate the progression of cervical cancer cells partly by targeting the miR‐636/CYP2S1 axis, hinting at a promising therapeutic target for cervical cancer.

Circular RNA hsa_circ_0000730 restrains cell proliferation, migration, and invasion in cervical cancer through miR‐942‐5p/PTEN axis

AbstractCircular RNAs (circRNAs) play prominent roles in regulating the progression of cancers. This study is aimed to decipher the role of hsa_circ_0000730 in cervical cancer (CC).The differentially expressed circRNAs of CC were screened out from the Gene Expression Omnibus database. qRT‐PCR was used to detect circ_0000730 expression in CC tissues and cell lines, and the Kaplan–Meier curve was adopted to figure out the relationship between circ_000730 expression and the overall survival time of CC patients. BrdU assay and Tanswell assay were utilized to examine the proliferation, migration, and invasion of CC cells. Western blot was adopted to detect PTEN protein expression. Bioinformatics analysis and dual‐luciferase reporter assay were used to examine the target relationship between miR‐942‐5p and circ_0000730 or PTEN, respectively.Circ_0000730 was among the differentially expressed circRNAs in CC. Circ_0000730 was significantly down‐regulated in the cancer tissues of 50 CC patients and CC cell lines. Additionally, underexpression of circ_0000730 was associated with the shorter survival time of CC patients. Gain‐ and loss‐of‐function assays highlighted that circ_0000730 significantly inhibited the proliferation, migration, and invasion of CC cells. Mechanistically, miR‐942‐5p was identified as a downstream target of circ_0000730, and circ_0000730 could positively regulate PTEN expression via repressing miR‐942‐5p in CC cells.Circ_0000730 inhibits the proliferation, migration, and invasion of CC cells via regulating miR‐942‐5p/PTEN axis. Circ_0000730 probably acts as a tumor suppressor in CC, and it may be a candidate target for the treatment of CC.

Circular RNA circ_0011385 promotes cervical cancer progression through competitively binding to miR‐149‐5p and up‐regulating SOX4 expression

AbstractCircular RNAs (circRNAs), emerging as a new type of non‐coding RNAs, play important roles in cancers. Instead, the functions and mechanisms of circ_0011385 in cervical cancer (CC) are still inconclusive. Microarray data GSE102686 was downloaded from Gene Expression Omnibus (GEO) database, and were utilized to screen out circRNAs differently expressed in CC tissues. Circ_0011385, miR‐149‐5p, SRY‐box transcription factor 4 (SOX4) mRNA expressions in CC tissues and cells were probed by quantitative real‐time PCR (qRT‐PCR). CC cell lines with circ_0011385 knockdown were constructed, and he multiplication, migration, invasion, and apoptosis of CC cells were evaluated by cell counting kit‐8 (CCK‐8) method, transwell assay, and flow cytometry. In addition, the targeting relationships between miR‐149‐5p and circ_0011385 or SOX4 mRNA 3′UTR were probed by dual‐luciferase reporter gene assay and RNA pull‐down assay. The regulatory function of circ_0011385 and miR‐149‐5p on SOX4 expression was studied with western blot. Expressions of circ_0011385 and SOX4 mRNA were raised in CC tissues and cells, while miR‐149‐5p expression was decreased. Knocking down circ_0011385 restrained the multiplication, migration, and invasion of CC cells and induced the apoptosis. Circ_0011385 directly targeted miR‐149‐5p, and SOX4 was the target of miR‐149‐5p, which could be positively regulated by circ_0011385. Circ_0011385 elevates SOX4 expression by targeting miR‐149‐5p, thus participating in promoting the malignant biological behaviors of CC cells.

Circular RNA circRNA_101996 promoted cervical cancer development by regulating miR‐1236‐3p/TRIM37 axis

AbstractCircular RNAs (circRNAs) appear to be significant modulators in various physiological processes. Recently, it is found that circRNA_101996 exerts important roles in various cancers. Our previous studies showed that circRNA_101996 promoted cervical cancer growth and metastasis by regulating miR‐8075/TPX2. However, the potential regulatory role of circRNA_101996 in cervical cancer still needs further investigation. Our results in this study suggested that circRNA_101996 was over‐expressed in cervical cancer patients. circRNA_101996 up‐regulation remarkably assisted cell proliferation, cell cycle progression, and cell migration in cervical cancer, while circRNA_101996 knockdown exerted the inverse effects. The molecular investigations indicated that circRNA_101996 could increase the expression level of miR‐1236‐3p, tripartite motif‐containing 37 (TRIM37), through binding to miR‐1236‐3p and reducing its expression. Moreover, in vivo results demonstrated that circRNA_101996 shRNA can function as a tumor suppressor through down‐regulating TRIM37 in cervical cancer. In conclusion, our data indicated that circRNA_101996/miR‐1236‐3p/TRIM37 axis accelerated cervical cancer development, providing novel insights into cervical cancer diagnosis and treatment.

Apigenin suppresses proliferation, invasion, and epithelial–mesenchymal transition of cervical carcinoma cells by regulation of miR‐152/BRD4 axis

AbstractThe present study aimed to investigate the role of apigenin and the molecular mechanism of miR‐152‐5p and bromodomain containing 4 (BRD4) in the proliferation, invasion, and epithelial–mesenchymal transition (EMT) of cervical carcinoma cells. Quantitative real‐time PCR was used to detect the transfection efficiency and the expression of miR‐152‐5p and BRD4. Western blotting was conducted to evaluate the protein level of BRD4, E‐cadherin, N‐cadherin, and MMP9. Luciferase reporter assay was performed to confirm whether miR‐152‐5p bound to BRD4. MTT and Transwell invasion assay were applied to determine the cell proliferation and invasion, respectively. MiR‐152‐5p was downregulated and BRD4 was upregulated in cervical carcinoma tissue. Besides, miR‐152‐5p could directly bind to BRD4 in Hela and CaSki cells. In addition, apigenin inhibited proliferation, invasion, and EMT of Hela and CaSki cells by regulating miR‐152‐5p/BRD4 axis. Apigenin suppresses proliferation, invasion, and induced EMT of cervical carcinoma cells by regulation of miR‐152‐5p/BRD4 axis.

Oncogenic miRNA‐1908 targets HDAC10 and promotes the aggressive phenotype of cervical cancer cell

AbstractMicroRNAs (miRNAs) have vital functions in tumorigenesis and cancer progression. The significance of miR‐1908 in cervical cancer has not been determined. We revealed that miR‐1908 was notably upregulated in cervical cancer. Upregulation of miR‐1908 increased cervical carcinoma cell growth and invasion. Downregulation of miR‐1908 caused the opposite effects. We confirmed that histone deacetylase 10 (HDAC10) was a potential target of miR‐1908 using bioinformatics analysis and luciferase reporter gene assays. Western blot analysis showed that miR‐1908 regulated the expression of HDAC10 by binding its 3′‐UTR. In addition, ectopic expression of HDAC10 partially reversed the promoting effects of miR‐1908. In conclusion, our findings indicated that miR‐1908 targets HDAC10 in cervical cancer and regulates aggressive cervical cancer cell phenotypes.

LncRNA AATBCindicates development and facilitates cell growth and metastasis of cervical cancer as a sponge ofmiR‐1245b‐5p

AbstractWith the increasing incidence and mortality rate, cervical cancer has been considered one of the most frequent malignant tumors in females. Exploration of tumor progression‐related biomarkers could facilitate the identification of novel and targeted therapy strategies. To assess the significance of lncRNA AATBC (AATBC) and its potential regulatory mechanism in cervical cancer, and to identify a potential biomarker, this study enrolled 123 patients with cervical cancer. Paired tissue samples were collected. The expression levels of AATBC and miR‐1245b‐5p were analyzed by RT‐qPCR and their significance in the development and prognosis of cervical cancer was evaluated using chi‐square and Cox analyses. In vitro, the regulatory effect of AATBC on the cellular processes of cervical cancer was estimated by CCK8 and Transwell assay. The interaction between ATTBC and miR‐1245b‐5p was assessed by luciferase reporter assay. Significant upregulation of AATBC and reduced miR‐1245b‐5p level in cervical cancer were observed, which showed a negative correlation between their expression levels. Close relationships of AATBC and miR‐1245b‐5p with the FIGO stage and lymph node metastasis were revealed. AATBC showed a significant prognostic value and miR‐1245b‐5p was found to mediate the tumor inhibitory effect of AATBC knockdown, which is speculated to be the underlying molecular mechanism of AATBC in cervical cancer development. Upregulation of AATBC indicted the malignant development and adverse prognosis of cervical cancer. AATBC served as a tumor promoter of cervical cancer by modulating miR‐1245b‐5p.

miR‐96‐5p regulates cervical cancer cell resistance to cisplatin by inhibiting lncRNA TRIM52‐AS1 and promoting IGF2BP2

AbstractMicroRNA (miRNA) and long noncoding RNA (lncRNA) are both regulators of cancer progression. This study sought to discuss the functional mechanism of miR‐96‐5p/lncRNA TRIM52 antisense RNA 1 (head‐to‐head; TRIM52‐AS1) in cervical cancer (CC) cell resistance to cisplatin (DDP). DDP‐resistant CC cell line was established using increasing concentrations of DDP, followed by transfection with miR‐96‐5p inhibitor, or si‐TRIM52‐AS1, or insulin‐like growth factor 2 mRNA binding protein 2 (IGF2BP2) overexpression vector. Expression levels of miR‐96‐5p, TRIM52‐AS1, and IGF2BP2 were determined. Changes in IC50 value to DDP, cell proliferation, and apoptosis rate were evaluated by cell‐counting kit‐8 assay, colony formation, and flow cytometry. The bindings of miR‐96‐5p to IGF2BP2 and TRIM52‐AS1 to IGF2BP2 were verified by dual‐luciferase or RNA pull‐down assays. These experiments revealed an up‐expression of miR‐96‐5p and IGF2BP2 while an under‐expression of TRIM52‐AS1 in CC cells. After DDP treatment, miR‐96‐5p inhibition increased apoptosis and decreased proliferation and DDP resistance. miR‐96‐5p bound to TRIM52‐AS1 and downregulated TRIM52‐AS1 expression, and TRIM52‐AS1 bound to IGF2BP2 to inhibit IGF2BP2 expression. TRIM52‐AS1 inhibition or IGF2BP2 overexpression neutralized the inhibition of silencing miR‐96‐5p on CC cell resistance to DDP. Overall, miR‐96‐5p improved CC cell resistance to DDP by inhibiting TRIM52‐AS1 and promoting IGF2BP2.