FOXA1 co-activates circODC1 and ODC1 in HPV-positive cervical cancer cell growth.

As demonstrated in previous research, hsa_circ_0052602 (circODC1) is dynamically expressed in HPV-positive cervical cancer (CC). CircODC1 expression was quantified using qRT-PCR, and its role in CC cell growth was assessed via loss-of-function assays. Interactions between miR-607 and circODC1 or ODC1 were confirmed using bioinformatics and mechanistic assays. The association of FOXA1 with the circODC1 promoter was validated through ChIP and luciferase reporter assays. CircODC1 was highly expressed in HPV-positive CC cell lines, and its depletion significantly impeded malignant processes such as proliferation, migration, and invasion. We found that ODC1 also played an oncogenic role in HPV-positive CC cells. CircODC1 was shown to positively regulate ODC1 as a ceRNA, competitively binding to miR-607 to counteract its suppression of ODC1. HPV-associated FOXA1 was identified as a potential transcription factor of circODC1. Restoration experiments showed that overexpression of circODC1 could counterbalance the inhibitory effect of FOXA1 knockdown. These findings offer new insights into therapeutic strategies for HPV-positive CC patients.

M6A-induced transcription factor IRF5 contributes to the progression of cervical cancer by upregulating PPP6C.

Cervical cancer (CC) is a gynaecological malignancy tumour that seriously threatens women's health. Recent evidence has identified that interferon regulatory factor 5 (IRF5), a nucleoplasm shuttling protein, is a pivotal transcription factor regulating the growth and metastasis of various human tumours. This study aimed to investigate the function and molecular basis of IRF5 in CC development. IRF5, protein phosphatase 6 catalytic subunit (PPP6C) and methyltransferase-like 3 (METTL3) mRNA levels were evaluated by quantitative real-time (qRT)-polymerase chain reaction (PCR). IRF5, PPP6C, METTL3, B-cell lymphoma 2 and Bax protein levels were detected using western blot. Cell proliferation, migration, invasion, angiogenesis and apoptosis were determined by using colony formation, 5-ethynyl-2'-deoxyuridine (EdU), transwell, tube formation assay and flow cytometry assay, respectively. Glucose uptake and lactate production were measured using commercial kits. Xenograft tumour assay in vivo was used to explore the role of IRF5. After JASPAR predication, binding between IRF5 and PPP6C promoter was verified using chromatin immunoprecipitation and dual-luciferase reporter assays. Moreover, the interaction between METTL3 and IRF5 was verified using methylated RNA immunoprecipitation (MeRIP). IRF5, PPP6C and METTL3 were highly expressed in CC tissues and cells. IRF5 silencing significantly inhibited cell proliferation, migration, invasion, angiogenesis and glycolytic metabolism in CC cells, while induced cell apoptosis. Furthermore, the absence of IRF5 hindered tumour growth in vivo. At the molecular level, IRF5 might bind with PPP6C to positively regulate the expression of PPP6C mRNA. Meanwhile, IRF5 was identified as a downstream target of METTL3-mediated m6A modification. METTL3-mediated m6A modification of mRNA might promote CC malignant progression by regulating PPP6C, which might provide a promising therapeutic target for CC treatment.

Mesencephalic astrocyte-derived neurotrophic factor inhibits cervical cancer progression via regulating macrophage phenotype.

BACKGROUND: Cervical cancer is a common gynecologic malignant tumor, but the critical factors affecting cervical cancer progression are still not well demonstrated. Mesencephalic astrocyte-derived neurotrophic factor (MANF) has been widely recognized as an anti-inflammatory factor to regulate macrophage polarization. In this study, the effect and mechanism of MANF on cervical cancer were preliminarily explored. METHODS AND RESULTS: Kaplan-Meier curve was used to show the overall survival time of the involved cervical cancer patients with high and low MANF expression in cervical cancer tissues. MANF was highly expressed in peritumoral tissues of cervical carcinoma by using immunohistochemistry and western blot. MANF mRNA level was detected by using qRT-PCR. Dual-labeled immunofluorescence showed MANF was mainly expressed in macrophages of cervical peritumoral tissues. Moreover, MANF-silenced macrophages promoted HeLa and SiHa cells survival, migration, invasion and EMT via NF-κB signaling activation. The results of tumor formation in nude mice indicated MANF-silenced macrophages promoted cervical tumor formation in vivo. CONCLUSION: Our study reveals an inhibitory role of MANF in cervical cancer progression, indicating MANF as a new and valuable therapeutic target for cervical cancer treatment.

METTL3-mediated m6A methylation of SLC38A1 stimulates cervical cancer growth.

The objective of this study was to better characterize the role of the glutamine transporter SLC38A1 in cervical cancer and explore the underlying mechanisms. Data from public databases and clinical cervical cancer tissue samples were used to assess the expression of SLC38A1 and its prognostic significance. Immunohistochemical staining, qRT-PCR, and Western blotting were used to evaluate the expression of relevant genes and proteins. Cell viability, cell cycle, apoptosis, and intracellular glutamine content were measured using CCK-8, flow cytometry, and biochemical assays. Additionally, the RNA immunoprecipitation (RIP) assay was used to examine the impact of METTL3/IGF2BP3 on the m6A modification of the SLC38A1 3'UTR. Both cervical cancer specimens and cells showed significantly increased expression of SLC38A1 and its expression correlated with an unfavorable prognosis. Knockdown of SLC38A1 inhibited cell viability and cell cycle progression, induced apoptosis, and suppressed tumor growth in vivo. Glutaminase-1 inhibitor CB-839 reversed the effects of SLC38A1 overexpression. METTL3 promoted m6A modification of SLC38A1 and enhanced its mRNA stability through IGF2BP3 recruitment. Moreover, METTL3 silencing inhibited cell viability, cell cycle progression, intracellular glutamine content, and induced apoptosis, but these effects were reversed by SLC38A1 overexpression. In conclusion, METTL3-mediated m6A methylation of SLC38A1 stimulates cervical cancer progression. SLC38A1 inhibition is a potential therapeutic strategy for cervical cancer.

METTL14 decreases FTH1 mRNA stability via m6A methylation to promote sorafenib-induced ferroptosis of cervical cancer.

Cervical cancer (CC) is a prevalent malignancy among women worldwide. This study was designed to investigate the role of METTL14 in sorafenib-induced ferroptosis in CC. METTL14 expression and m6A methylation were determined in CC tissues, followed by analyzes correlating these factors with clinical features. Subsequently, METTL14 was knocked down in CC cell lines, and the effects on cell proliferation, mitochondrial morphology and ferroptosis were assessed using CCK-8, microscopy, and markers associated with ferroptosis, respectively. The regulatory relationship between METTL14 and FTH1 was verified using qRT-PCR and luciferase reporter assays. The functional significance of this interaction was further investigated both in vitro and in vivo by co-transfecting cells with overexpression vectors or shRNAs targeting METTL14 and FTH1 after sorafenib treatment. METTL14 expression and m6A methylation were significantly reduced in CC tissues, and lower METTL14 expression levels were associated with a poorer CC patients' prognosis. Notably, METTL14 expression increased during sorafenib-induced ferroptosis, and METTL14 knockdown attenuated the ferroptotic response induced by sorafenib in CC cells. FTH1 was identified as a direct target of METTL14, with METTL14 overexpression leading to increased m6A methylation of FTH1 mRNA, resulting in reduced stability and expression of FTH1 in CC. Furthermore, FTH1 overexpression or treatment with LY294002 partially counteracted the promotion of sorafenib-induced ferroptosis by METTL14. In vivo xenograft experiments demonstrated that inhibiting METTL14 reduced the anticancer effects of sorafenib, whereas suppression of FTH1 significantly enhanced sorafenib-induced ferroptosis and increased its anticancer efficacy. METTL14 reduces FTH1 mRNA stability through m6A methylation, thereby enhancing sorafenib-induced ferroptosis, which contributes to suppressing CC progression via the PI3K/Akt signaling pathway.

Retraction Note: BNIP1 inhibits cell proliferation, migration and invasion, and promotes apoptosis by mTOR in cervical cancer cells.

The article "BNIP1 inhibits cell proliferation, migration and invasion, and promotes apoptosis by mTOR in cervical cancer cells", by F.-H. Li, L. Xiang, L. Ran, S. Zhou, Z. Huang, M. Chen, W.-F. Yu, published in Eur Rev Med Pharmacol Sci 2019; 23 (4): 1397-1407-DOI: 10.26355/eurrev_201902_17096-PMID: 30840260 has been retracted by the Editor in Chief for the following reasons. Following some concerns raised on PubPeer regarding a possible overlap in Figure 2A, the Editor in Chief has started an investigation to assess the validity of the results as well as possible figure manipulation. The journal investigation revealed a duplication in Figure 2A between BNIP1 panels, migration and invasion, respectively and in Control and invasion panels. Consequently, the Editor in Chief mistrusts the results presented and has decided to withdraw the article. The authors have been informed about the journal's investigation but remained unresponsive. https://www.europeanreview.org/article/17096 This article has been retracted. The Publisher apologizes for any inconvenience this may cause.

Tobacco Smoke Condensate Induces Morphologic Changes in Human Papillomavirus-Positive Cervical Epithelial Cells Consistent with Epithelial to Mesenchymal Transition (EMT) with Activation of Receptor Tyrosine Kinases and Regulation of TGFB.

High-risk human papillomavirus (HR-HPV; HPV-16) and cigarette smoking are associated with cervical cancer (CC); however, the underlying mechanism(s) remain unclear. Additionally, the carcinogenic components of tobacco have been found in the cervical mucus of women smokers. Here, we determined the effects of cigarette smoke condensate (CSC; 3R4F) on human ectocervical cells (HPV-16 Ect/E6E7) exposed to CSC at various concentrations (10-6-100 µg/mL). We found CSC (10-3 or 10 µg/mL)-induced proliferation, enhanced migration, and histologic and electron microscopic changes consistent with EMT in ectocervical cells with a significant reduction in E-cadherin and an increase in the vimentin expression compared to controls at 72 h. There was increased phosphorylation of receptor tyrosine kinases (RTKs), including Eph receptors, FGFR, PDGFRA/B, and DDR2, with downstream Ras/MAPK/ERK1/2 activation and upregulation of common EMT-related genes, TGFB SNAI2, PDGFRB, and SMAD2. Our study demonstrated that CSC induces EMT in ectocervical cells with the upregulation of EMT-related genes, expression of protein biomarkers, and activation of RTKs that regulate TGFB expression, and other EMT-related genes. Understanding the molecular pathways and environmental factors that initiate EMT in ectocervical cells will help delineate molecular targets for intervention and define the role of EMT in the initiation and progression of cervical intraepithelial neoplasia and CC.

Long non-coding RNA NEAT1 promotes aerobic glycolysis and progression of cervical cancer through WNT/ß-catenin/PDK1 axis.

BACKGROUND: Cervical cancer is one of the most common gynecological cancers. Accumulated evidence shows that long non-coding RNAs (lncRNAs) play essential roles in cervical cancer occurrence and progression, but their specific functions and mechanisms remain to be further explored. METHODS: The RT-qPCR assay was used to detect the expression of NEAT1 in cervical cancer tissues and cell lines. CCK-8, colony formation, flow cytometry, western blotting, and Transwell assays were used to evaluate the impact of NEAT1 on the malignant behavior of cervical cancer cells. Glucose consumption, lactate production, ATP levels, ROS levels, MMP levels, and the mRNA expressions of glycolysis-related genes and tricarboxylic acid cycle-related genes were detected to analyze the effect of NEAT1 on metabolism reprograming in cervical cancer cells. The expressions of PDK1, ß-catenin and downstream molecules of the WNT/ß-catenin signaling pathway in cervical cancer cells and tissues were detected by western blotting, RT-qPCR, immunofluorescence and immunohistochemistry assays. RESULTS: This study investigated the role and possible molecular mechanism of lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) in cervical cancer. Our results showed that NEAT1 was highly expressed in cervical cancer tissues and cell lines. Downregulation of NEAT1 inhibited the proliferation, migration, invasion and glycolysis of cervical cancer cells, while overexpression of NEAT1 led to the opposite effects. Mechanistically, NEAT1 upregulated pyruvate dehydrogenase kinase (PDK1) through the WNT/ß-catenin signaling pathway, which enhanced glycolysis and then facilitated cervical cancer metastasis. Furthermore, NEAT1 maintained the protein stability of ß-catenin but did not affect its mRNA level. We also excluded the direct binding of NEAT1 to the ß-catenin protein via RNA pull-down assay. The suppressive impact of NEAT1 knockdown on cell proliferation, invasion, and migration was rescued by ß-catenin overexpression. The WNT inhibitor iCRT3 attenuated the carcinogenic effect induced by NEAT1 overexpression. CONCLUSION: In summary, these findings indicated that NEAT1 may contribute to the progression of cervical cancer by activating the WNT/ß-catenin/PDK1 signaling axis.

Laminin receptor 1 expression in premalignant and malignant squamous lesions of the cervix.

Laminin receptor 1 (LAMR) may have a role in the progression of premalignant squamous epithelial lesions to cervical cancer. Therefore, we aimed to investigate the expression of laminin receptor 1 (LAMR) in normal, premalignant, and malignant tissues of the uterine cervix. Paraffin blocks of 129 specimens with the diagnoses of normal cervical tissue (n = 33), cervical intraepithelial neoplasia (CIN) 1 (n = 30), CIN 2 (n = 14), CIN 3 (n = 28), and squamous cell carcinoma (n = 24) were immunohistochemically stained with LAMR antibody and its expression percentage, pattern, and intensity in these tissues were assessed. Compared to the other groups, the nonstaining with LAMR was highest in low grade squamous intraepithelial lesion (LSIL) (p < 0.0001). LAMR expression, which was positive in less than 50% of cells with weak staining, increased significantly between normal cervical epithelium and high-grade squamous intraepithelial lesion (HSIL) or invasive carcinoma, as well as between LSIL and HSIL (p < 0.0001). Between LSIL and invasive carcinoma, a significant increment was also observed for weak staining in less than 50% of cells (p < 0.001). LAMR expression, which was positive in more than 50% of cells with strong staining, was significantly higher in normal cervical tissue compared to the other groups (p < 0.0001). Disease progression related gradual increment of LAMR expression from normal cervical epithelium or LSIL towards HSIL or cervical cancer reveals that LAMR may play an important role in the transition from premalignant to malignant state in cervical lesions.

TPP1 is associated with risk of advanced precursors and cervical cancer survival.

It is unclear how telomere-binding protein TPP1 interacts with human telomerase reverse transcriptase (hTERT) and influences cervical cancer development and progression. This study included all eligible 156 cervical cancers diagnosed during 2003-2008 and followed up through 2014, 102 cervical intraepithelial neoplasia (CIN) patients, and 16 participants with normal cervix identified at the same period. Correlation of expression of TPP1 and hTERT in these lesions was assessed using Kappa statistics. TPP1 was knocked down by siRNA in three cervical cancer cell lines. We assessed mRNA expression using quantitative real-time polymerase chain reaction and protein expression using tissue microarray-based immunohistochemical staining. We further analyzed the impact of TPP1 expression on the overall survival of cervical cancer patients by calculating the hazard ratio (HR) with 95% confidence intervals (CIs) using the multivariable-adjusted Cox regression model. Compared to the normal cervix, high TPP1expression was significantly associated with CIN 3 and cervical cancers (P<0.001 for both). Expressions of TPP1 and hTERT were highly correlated in CIN 3 (Kappa statistics = 0.50, P = 0.005), squamous cell carcinoma (Kappa statistics = 0.22, P = 0.011), and adenocarcinoma/adenosquamous carcinoma (Kappa statistics = 0.77, P = 0.001). Mechanistically, knockdown of TPP1 inhibited the expression of hTERT in both mRNA and protein levels. High expression of TPP1 (HR = 2.61, 95% CI 1.23-5.51) and co-high expression of TPP1 and hTERT (HR = 2.38, 95% CI 1.28-4.43) were independently associated with worse survival in cervical cancer patients. TPP1 and hTERT expression was correlated and high expression of TPP1 was associated with high risk of CIN 3 and cervical cancer and could predict a worse survival in cervical cancer.

Emerging biomarkers and molecular targets for precision medicine in cervical cancer.

Cervical cancer remains a significant global health burden, necessitating innovative approaches for improved diagnostics and personalized treatment strategies. Precision medicine has emerged as a promising paradigm, leveraging biomarkers and molecular targets to tailor therapy to individual patients. This review explores the landscape of emerging biomarkers and molecular targets in cervical cancer, highlighting their potential implications for precision medicine. By integrating these biomarkers into comprehensive diagnostic algorithms, clinicians can identify high-risk patients at an earlier stage, enabling timely intervention and improved patient outcomes. Furthermore, the identification of specific molecular targets has paved the way for the development of targeted therapies aimed at disrupting key pathways implicated in cervical carcinogenesis. In conclusion, the evolving landscape of biomarkers and molecular targets presents exciting opportunities for advancing precision medicine in cervical cancer. By harnessing these insights, clinicians can optimize treatment selection, enhance patient outcomes, and ultimately transform the management of this devastating disease.

Cervical extracellular matrix hydrogel optimizes tumor heterogeneity of cervical squamous cell carcinoma organoids.

Cervical cancer, primarily squamous cell carcinoma, is the most prevalent gynecologic malignancy. Organoids can mimic tumor development in vitro, but current Matrigel inaccurately replicates the tissue-specific microenvironment. This limitation compromises the accurate representation of tumor heterogeneity. We collected para-cancerous cervical tissues from patients diagnosed with cervical squamous cell carcinoma (CSCC) and prepared uterine cervix extracellular matrix (UCEM) hydrogels. Proteomic analysis of UCEM identified several tissue-specific signaling pathways including human papillomavirus, phosphatidylinositol 3-kinase-AKT, and extracellular matrix receptor. Secreted proteins like FLNA, MYH9, HSPA8, and EEF1A1 were present, indicating UCEM successfully maintained cervical proteins. UCEM provided a tailored microenvironment for CSCC organoids, enabling formation and growth while preserving tumorigenic potential. RNA sequencing showed UCEM-organoids exhibited greater similarity to native CSCC and reflected tumor heterogeneity by exhibiting CSCC-associated signaling pathways including virus protein-cytokine, nuclear factor κB, tumor necrosis factor, and oncogenes EGR1, FPR1, and IFI6. Moreover, UCEM-organoids developed chemotherapy resistance. Our research provides insights into advanced organoid technology through native matrix hydrogels.

MUC1 promotes cervical squamous cell carcinoma through ERK phosphorylation-mediated regulation of ITGA2/ITGA3.

In contrast to the decreasing trends in developed countries, the incidence and mortality rates of cervical squamous cell carcinoma in China have increased significantly. The screening and identification of reliable biomarkers and candidate drug targets for cervical squamous cell carcinoma are urgently needed to improve the survival rate and quality of life of patients. In this study, we demonstrated that the expression of MUC1 was greater in neoplastic tissues than in non-neoplastic tissues of the cervix, and cervical squamous cell carcinoma patients with high MUC1 expression had significantly worse overall survival than did those with low MUC1 expression, indicating its potential for early diagnosis of cervical squamous cell carcinoma. Next, we explored the regulatory mechanism of MUC1 in cervical squamous cell carcinoma. MUC1 could upregulate ITGA2 and ITGA3 expression via ERK phosphorylation, promoting the proliferation and metastasis of cervical cancer cells. Further knockdown of ITGA2 and ITGA3 significantly inhibited the tumorigenesis of cervical cancer cells. Moreover, we designed a combination drug regimen comprising MUC1-siRNA and a novel ERK inhibitor in vivo and found that the combination of these drugs achieved better results in animals with xenografts than did MUC1 alone. Overall, we discovered a novel regulatory pathway, MUC1/ERK/ITGA2/3, in cervical squamous cell carcinoma that may serve as a potential biomarker and therapeutic target in the future.

MUC1 is overexpressed in cervical squamous cell carcinoma. MUC1 regulates ERK phosphorylation, and subsequently upregulates ITGA2 and ITGA3 expression to promote tumorigenesis in cervical squamous cell carcinoma. A combination drug regimen targeting MUC1 and ERK achieved better results compared than MUC1 alone.

Gas-Amplified Metalloimmunotherapy with Dual Activation of Pyroptosis and the STING Pathway for Remodeling the Immunosuppressive Cervical Cancer Microenvironment.

The immunosuppressive microenvironment of cervical cancer significantly hampers the effectiveness of immunotherapy. Herein, PEGylated manganese-doped calcium sulfide nanoparticles (MCSP) were developed to effectively enhance the antitumor immune response of the cervical cancer through gas-amplified metalloimmunotherapy with dual activation of pyroptosis and STING pathway. The bioactive MCSP exhibited the ability to rapidly release Ca2+, Mn2+, and H2S in response to the tumor microenvironment. H2S disrupted the calcium buffer system of cancer cells by interfering with the oxidative phosphorylation pathway, leading to calcium overload-triggered pyroptosis. On the other hand, H2S-mediated mitochondrial dysfunction further promoted the release of mitochondrial DNA (mtDNA), enhancing the activation effect of Mn2+ on the cGAS-STING signaling axis and thereby activating immunosuppressed dendritic cells. The released H2S acted as an important synergist between Mn2+ and Ca2+ by modulating dual signaling mechanisms to bridge innate and adaptive immune responses. The combination of MCSP NPs and PD-1 immunotherapy achieved synergistic antitumor effects and effectively inhibited tumor growth. This study reveals the potential collaboration between H2S gas therapy and metalloimmunotherapy and provides an idea for the design of nanoimmunomodulators for rational regulation of the immunosuppressive tumor microenvironment.

JAM3 promotes cervical cancer metastasis by activating the HIF-1α/VEGFA pathway.

Cervical cancer is the fourth most common cancer and the leading cause of mortality among women worldwide. Tumor metastasis is an important cause of poor prognosis. Determining the exact mechanisms of metastasis and potential targeted therapies is urgently needed. Junctional adhesion molecule 3 (JAM3) is an important member of the TJ tight junction (TJ) family, and its biological function in cervical cancer needs to be further clarified. We found that JAM3 was highly expressed in cervical cancer patients with lymph node metastasis and that high expression of JAM3 promoted cervical cancer cell metastasis both in vitro and in vivo. In addition, overexpression of JAM3 induces epithelial-mesenchymal transition (EMT). Moreover, silencing JAM3 suppressed cervical cancer cell migration and invasion in vitro. Finally, JAM3 overexpression activated the HIF-1α/VEGFA pathway. In conclusion, our results suggested that JAM3 promotes cervical cancer cell migration and invasion by activating the HIF-1α/VEGFA pathway. JAM3 may be a promising biomarker and effective therapeutic target for cervical cancer.

The antitumoral effect of the esculetin in HeLa cells through endoplasmic reticulum stress.

OBJECTIVE: Despite the multiple available treatment modalities, cervical cancer is one of the leading causes of mortality and morbidity among female gynecological cancers. Endoplasmic Reticulum (ER) is an effective organelle in ensuring cell homeostasis and is closely related to the development of cancer. Esculetin is a coumarin derivative that has anticancer, anti-inflammatory, antioxidant, and neuroprotective effects. Esculetin may have an anticancer effect by inducting apoptosis and ER stress. In this study, we evaluate that esculetin has an anti-tumor effect on human cervical cancer-derived (HeLa) cells via ER stress. MATERIALS AND METHODS: Esculetin was applied to the HeLa cells, and a viability test was performed using the methyl thiazolyl tetrazolium proliferation (MTT) assay. Expression levels of apoptotic genes and anti-apoptotic genes were determined by real-time polymerase chain reaction. The results were statistically evaluated. RESULTS: Analysis of the MTT assay detected that esculetin inhibited HeLa cell viability development. Based on Western blot and quantitative real-time polymerase chain reaction (qPCR) analyses, esculetin destroyed cervical cancer cells via the ER stress pathway. CONCLUSIONS: The results showed that esculetin may have a potent antitumoral effect. It can potentially be utilized in the pharmacological therapy of cervical cancer.

[Knockdown mitochondrial transcription factor A (TFAM) inhibits autophagy, proliferation, invasion and migration in human cervical cancer and osteosarcoma cells].

Objective To investigate the effects of mitochondrial transcription factor A (TFAM) on mitochondrial function, autophagy, proliferation, invasion, and migration in cervical cancer HeLa cells and osteosarcoma U2OS cells. Methods TFAM small-interfering RNA (si-TFAM) was transfected to HeLa and U2OS cells for downregulating TFAM expression. Mito-Tracker Red CMXRos staining combined with laser confocal microscopy was used to detect mitochondrial membrane potential (MMP). MitoSOXTM Red labeling was used to test mitochondrial reactive oxygen species (mtROS) levels. The expression of mitochondrial DNA (mtDNA) was detected by real-time quantitative PCR. Changes in the number of autophagosomes were detected by immunofluorescence cytochemistry. Western blot analysis was used to detect the expressions of TFAM, autophagy microtubule associated protein 1 light chain 3A/B (LC3A/B), autophagy associated protein 2A (ATG2A), ATG2B, ATG9A, zinc finger transcription factor Snail, matrix metalloproteinase 2 (MMP2) and MMP9. CCK-8 assay and plate clony formation assay were used to detect cell proliferation, while TranswellTM assay and scratch healing assay were used to detect changes in cell invasion and migration. Results The downregulation of TFAM expression resulted in a decrease in MMP and mtDNA copy number, but an increase in mtROS production. The protein content of LC3A/B decreased significantly compared to the control group and the number of autophagosomes in the cytoplasm decreased significantly. The expressions of ATG2B and ATG9A in the early stage of autophagy were significantly reduced. The expressions of Snail, MMP2 and MMP9 proteins in HeLa and U2OS cells were also decreased. The proliferation, invasion and migration ability of HeLa and U2OS cells were inhibited after being interfered with TFAM expression. Conclusion Downregulation of TFAM expression inhibits mitochondrial function, delays autophagy process and reduces the proliferation, invasion and migration ability of cervical cancer cells and osteosarcoma cells.

Phenotypic Heterogeneity of Cancer Associated Fibroblasts in Cervical Cancer Progression: FAP as a Central Activation Marker.

Cervical cancer (CC) is the fourth leading cancer among women and is one of the principal gynecological malignancies. In the tumor microenvironment, cancer-associated fibroblasts (CAFs) play a crucial role during malignant progression, exhibiting a variety of heterogeneous phenotypes. CAFs express phenotypic markers like fibroblast activation protein (FAP), vimentin, S100A4, α-smooth muscle actin (αSMA), and functional markers such as MMP9. This study aimed to evaluate the protein expression of vimentin, S100A4, αSMA, FAP, and MMP9 in mesenchymal stem cells (MSC)-CAF cells, as well as in cervical cancer samples. MSC cells were stimulated with HeLa and SiHa tumor cell supernatants, followed by protein evaluation and cytokine profile to confirm differentiation towards a CAF phenotype. In addition, automated immunohistochemistry (IHQa) was performed to evaluate the expression of these proteins in CC samples at different stages. Our findings revealed a high expression of FAP in stimulated MSC cells, accompanied by the secretion of pro/anti-inflammatory cytokines. In the other hand, CC samples were observed to have high expression of FAP, vimentin, αSMA, and MMP9. Most importantly, there was a high expression of their activation proteins αSMA and FAP during the different stages. In the early stages, a myofibroblast-like phenotype (CAFs αSMA+ FAP+), and in the late stages a protumoral phenotype (CAF αSMA- FAP+). In summary, FAP has a crucial role in the activation of CAFs during cervical cancer progression.

N6-methyladenosine methylation on FSCN1 mediated by METTL14/IGF2BP3 contributes to human papillomavirus type 16-infected cervical squamous cell carcinoma.

Human papillomavirus (HPV) infection has been reported to be associated with N6-methyladenosine (m6A) modification in cancers. However, the underlying mechanism by which m6A methylation participates in HPV-related cervical squamous cell carcinoma (CSCC) remains largely unclear. In this study, we observed that m6A regulators methyltransferase like protein (METTL14) and insulin like growth factor 2 mRNA binding protein 3 (IGF2BP3) were upregulated in HPV-positive CSCC tissues and cell lines, and their high expression predicted poor prognosis for HPV-infected CSCC patients. Cellular functional experiments verified that HPV16 oncogenes E6/E7 upregulated the expression of METTL14 and IGF2BP3 to promote cell proliferation and epithelial mesenchymal transition of CSCC cells. Next, we found that E6/E7 stabilized fascin actin-bundling protein 1 (FSCN1) mRNA and elevated FSCN1 expression in CSCC cells through upregulating METTL14/IGF2BP3-mediated m6A modification, and FSCN1 expression was also validated to be positively associated with worse outcomes of HPV-positive CSCC patients. Finally, HPV16-positive CSCC cell lines SiHa and CaSki were transfected with knockdown vector for E6/E7 or METTL14/IGF2BP3 and overexpressing vector for FSCN1, and functional verification experiments were performed through using MTT assay, flow cytometry, wound healing assay and tumour formation assay. Results indicated that knockdown of E6/E7 or METTL14/IGF2BP3 suppressed cell proliferation, migration and tumorigenesis, and accelerated cell apoptosis of HPV-positive CSCC cells. Their tumour-suppressive effects were abolished through overexpressing FSCN1. Overall, HPV E6/E7 advanced CSCC development through upregulating METTL14/IGF2BP3-mediated FSCN1 m6A modification.

Exonuclease-iii -propelled DNAzyme cascade for sensitive and reliable cervical cancer related miRNA analysis.

MicroRNAs (miRNAs) can serve as biomarkers for early-diagnosis, therapy, and postoperative care of cervical cancer. Sensitive and reliable quantification of miRNA remains a huge challenge due to its low expressing levels and background interference. Herein, we propose a novel exonuclease-III (Exo-III)-propelled DNAzyme cascade for sensitive and high-efficient miRNA analysis. This method involves the engineering of compact DNAzyme hairpin probes, including the H1 probe and H2 probe. The H1 probe is designed with exposed analyte recognition subunits that can specifically recognize target miRNA. This recognition triggers two processes: Exo-iii-assisted target regeneration and successive substrate cleavage catalyzed by DNAzyme. The unique character of Exo-III that catalyzes removal of mononucleotides from the blunt or recessed 3'-OH termini of dsDNA confers the approach with a minimal background signal. The multiple signal cycles provided an abundant signal amplification and consequently, the method exhibited a low limit of detection of 3.12 fM, and a better specificity over several homologous miRNAs. In summary, this powerful Exo-III driven DNAzyme cascaded system offers broader and more adaptable methods for comprehending the activities of miRNA in various biological occurrences.