Mechanism of chromosomal mosaicism in preimplantation embryos and its effect on embryo development.

Aneuploidy is one of the main causes of miscarriage and in vitro fertilization failure. Mitotic abnormalities in preimplantation embryos are the main cause of mosaicism, which may be influenced by several endogenous factors such as relaxation of cell cycle control mechanisms, defects in chromosome cohesion, centrosome aberrations and abnormal spindle assembly, and DNA replication stress. In addition, incomplete trisomy rescue is a rare cause of mosaicism. However, there may be a self-correcting mechanism in mosaic embryos, which allows some mosaicisms to potentially develop into normal embryos. At present, it is difficult to accurately diagnose mosaicism using preimplantation genetic testing for aneuploidy. Therefore, in clinical practice, embryos diagnosed as mosaic should be considered comprehensively based on the specific situation of the patient.

Reproductive Effect by Rheumatoid Arthritis and Related Autoantibodies.

Rheumatoid arthritis (RA) is a common inflammatory arthritis in women. The effects of RA on the reproductive system are usually overlooked, as RA is not diagnosed until later in reproductive age. Whether RA itself or its related rheumatoid antibodies have an impact on female reproductive function has long been a thought-provoking issue. In brief, relevant epidemiological evidence has shown that women affected by RA are more likely to have coexisting reproductive disorders, including infertility, endometriosis, and premature ovarian insufficiency (POI), or to subsequently develop them. Furthermore, linkage between RA and pregnancy loss (PL) as well as polycystic ovary syndrome (PCOS) is also well known, albeit controversial in available evidence. RA and reproductive disorders appear to share a similar inflammatory immune response and genetic background. The stress experienced by patients with RA may affect their reproductive choices to some extent. Notably, few studies have explored the impact of rheumatoid antibodies such as rheumatoid factors (RFs) and anti-citrullinated protein antibodies (ACPAs) on reproductive disorders. Although it has been mentioned that the rate of RF and/or ACPA positivity is higher in women with a history of PL and POI, the clinical relevance of this relationship and underlying mechanisms still need to be further clarified.

Strategies and challenges of mesenchymal stem cells-derived extracellular vesicles in infertility.

Having genetically related offspring remains an unattainable dream for couples with reproductive failure. Mesenchymal stem cells (MSCs) are multipotent stromal cells derived from various human tissues and organs. As critical paracrine effectors of MSCs, extracellular vesicles (EVs) can carry and deliver bioactive content, thereby participating in intercellular communication and determining cell fate. Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) have shown promising therapeutic effects, including repairing injured endometria, restoration ovarian functions, and improving sperm quantity, morphology, and motility, owing to their regenerative potential, abundant sources, high proliferation rates, low immunogenicity, and lack of ethical issues. However, limited knowledge on purification and isolation of MSC-EVs, therapeutic effects, and unpredictable safety have caused challenges in overcoming female and male infertility. To overcome them, future studies should focus on modification/engineering of MSC-EVs with therapeutic biomolecules and combining attractive biomaterials and MSC-EVs. This review highlights the latest studies on MSC-EVs therapies in infertility and the major challenges that must be overcome before clinical translation.

GDF15 deficiency hinders human trophoblast invasion to mediate pregnancy loss through downregulating Smad1/5 phosphorylation.

Growth differentiation factor 15 (GDF15) belongs to the Transforming growth factor ß(TGF-ß) superfamily. The decrease of GDF15 in the serum of pregnant women was associated with miscarriage. Both IHC and ELISA assays showed that GDF15 in trophoblast tissue and serum of pregnant women who miscarried was significantly lower than in those who had a live birth. GDF15 deficiency was associated with embryo resorption in GDF15 knockout mice through CRIPSR editing. In addition, the migration and invasion ability of HTR-8/SVneo and JEG-3 cells were promoted by GDF15. Mechanistically, GDF15 increased Smad1/5 phosphorylation, resulting in upregulating SNAI1/2, VIMENTIN and downregulating E-CADHERIN. A dual-luciferase reporter assay confirmed that Smad-binding elements (SBE) and/or GC-rich motifs were activated and target genes such as SNAI1/2, SERPINE1, and TIMP3 were transcriptionally regulated by GDF15/Smad5 signaling. Therefore, our data revealed a crucial role of GDF15 on invasion of trophoblast by upregulating the activity of TGF-ß/Smad1/5 pathway.

ESRRB Inhibits the TGFß Signaling Pathway to Drive Cell Proliferation in Cervical Cancer.

Estrogen-related receptor ß (ESRRB) is a member of the orphan nuclear receptor family and mediates stem cell self-renewal and early embryonic development. Previous studies have also reported that ESRRB plays a role in the development and progression of breast cancer and prostate cancer. In this study, we observed that ESRRB was highly expressed in cervical cancer and was associated with disease progression. Knocking out ESRRB using CRISPR/Cas9 gene editing in cervical cancer cells induced cell-cycle arrest at the transition from the G0-G1 phase to the S phase, resulting in inhibition of cell proliferation in vitro and reduced tumor growth in vivo. Conversely, ectopic expression of ESRRB significantly promoted the proliferation of cervical cancer cells. ESRRB activated transcription of SMAD7, a TGFß pathway inhibitor, which blocked phosphorylation and nuclear translocation of SMAD2/3 to the nucleus, thereby downregulating CDKN1A and upregulating CCNA2 and MYC. In turn, MYC transactivated ESRRB and upregulated SMAD7, thus forming a positive feedback loop with ESRRB. Together, these findings identify the tumor-promoting function of ESRRB in cervical cancer and reveal a mechanism by which ESRRB stimulates cell proliferation to promote cancer progression. SIGNIFICANCE: The ESRRB/SMAD7/MYC-positive feedback loop inhibits TGFß signaling to activate cell-cycle progression and promote proliferation in cervical cancer, thereby driving tumor growth.

NEK2 inactivates the Hippo pathway to advance the proliferation of cervical cancer cells by cooperating with STRIPAK complexes.

The never in mitosis gene A (NIMA)-related kinase 2 (NEK2) protein has been reported to be an oncoprotein that plays different oncogenic roles in multiple cancers. Here, we confirmed that NEK2 highly expressed in cervical cancer cells rather than in normal epithelial basal layer cells in cervical tissues and correlated with worse outcomes. We also demonstrated that NEK2 promoted the in vivo growth of subcutaneous xenograft tumors stemming from cervical cancer cells and the in vitro cell proliferation by decreasing Ser127-phosphorylation of the YAP protein retained in the cytoplasm while increasing the levels of active nucleus-associated YAP protein, which was followed by increases in the targeted proteins CTGF, CYR61 and GLI2. Furthermore, the Hippo signaling pathway was inactivated in manipulated NEK2-overexpressing cervical cancer cells by regulating the levels of MST1/2 dephosphorylation. Additionally, mass spectrometric sequencing and bilateral coimmunoprecipitation were employed suggested that NEK2 acted at an early upstream step to promote dephosphorylation of MST2 and inactivate the Hippo signaling cascade by cooperating with STRIPAK complexes. We conjecture that NEK2 may be a future target for cervical cancer therapy.

Systematic review of subsequent pregnancy outcomes in couples with parental abnormal chromosomal karyotypes and recurrent pregnancy loss.

OBJECTIVE: To evaluate the current evidence of pregnancy outcomes among couples with recurrent pregnancy loss (RPL) with abnormal karyotypes vs. those with normal karyotypes and among couples with RPL and abnormal karyotypes after receiving expectant management vs. preimplantation genetic diagnosis (PGD). DESIGN: Systematic review and meta-analysis. SETTING: Academic medical centers. PATIENT(S): Pregnancy outcomes in 6,301 couples with RPL who conceived without medical intervention in 11 studies were analyzed. However, only 2 studies addressed the outcomes of couples with RPL and abnormal karyotypes after expectant management (75 cases) vs. PGD (50 cases). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): The pregnancy outcomes in couples with RPL with abnormal and normal karyotypes across included studies were evaluated. RESULT(S): Compared with those with a normal karyotype, a significantly lower first pregnancy live birth rate (LBR) was found in couples with RPL with abnormal karyotypes (58.5% vs. 71.9%; odds ratio [OR], 0.55; 95% confidence interval [CI], 0.46-0.65; I2 =27%). A markedly decreased first pregnancy LBR was found in couples with a translocation (52.9% vs. 72.4%; OR, 0.44; 95% CI, 0.31-0.61; I2 =33%) but not in couples with an inversion. However, the differences in accumulated LBR (81.4% vs. 74.8%; OR, 0.96; 95% CI, 0.90-1.03; I2 = 0) were nonsignificant, whereas the miscarriage rate was distinctly higher in couples with RPL and abnormal karyotypes (53.0% vs. 34.7%; OR, 2.21; 95% CI, 1.69-2.89; I2 = 0). Compared with those who chose expectant management, differences in accumulated LBR were nonsignificant (60% vs. 68%; OR, 0.55; 95% CI, 0.11-2.62; I2 =71%), whereas the miscarriage rate (24% vs. 65.3%; OR, 0.15; 95% CI, 0.04-0.51; I2 = 45) was markedly low in couples with RPL and abnormal karyotypes who chose PGD. CONCLUSION(S): Couples with RPL and abnormal karyotypes had a higher miscarriage rate than couples with normal karyotypes but achieved a noninferior accumulated LBR through multiple conception attempts. In couples with RPL and abnormal karyotypes, PGD treatment did not increase the accumulated LBR but markedly reduced miscarriage rate compared with expectant management.

Cytotoxic effect of disulfiram/copper on human cervical cancer cell lines and LGR5-positive cancer stem-like cells.

BACKGROUND: Tumor resistance is a global challenge for tumor treatment. Cancer stem cells (CSCs) are the main population of tumor cells for drug resistance. We have reported that high aldehyde dehydrogenase (ALDH) activity represents a functional marker for cervical CSCs. Here, we aimed at disulfiram (DSF), an ALDH inhibitor, that has the potential to be used for cervical cancer treatment. METHODS: MTT assay, western blot, vector construction and transfection, cell sorting and in vivo anti-tumor assays were performed using cervical cancer cell lines SiHa and HeLa. Cell cycle distribution and cell apoptosis were carried out by flow cytometry. The cytotoxicity of DSF was detected by MTT assay and cervical cancer xenograft models. RESULTS: DSF was cytotoxic to cervical cancer cell lines in a copper (Cu)-dependent manner. Disulfiram/copper (DSF/Cu) complex induced deregulation of S-phase and inhibited the expression of stemness markers in cervical cancer cells. Furthermore, DSF/Cu could also reduce the cancer stem cell-like LGR5+ cells which lead to cisplatin resistance in cervical cancer cells. DSF/Cu complex had the greater antitumor efficacy on cervical cancer than cisplatin in vitro and in vivo. CONCLUSION: Our findings indicate that the cytotoxicity of DSF/Cu complex may be superior to cisplatin because of targeting LGR5-positive cervical cancer stem-like cells in cervical cancer. Thus, the DSF/Cu complex may represent a potential therapeutic strategy for cervical cancer patients.

Recurrent pregnancy loss: fewer chromosomal abnormalities in products of conception? a meta-analysis.

OBJECTIVE: To compare the prevalence of chromosomal abnormalities detected in products of conception (POCs) between recurrent pregnancy loss and sporadic pregnancy loss. METHODS: A systematic search was performed in the PubMed and Embase databases from inception to December 31, 2020. Relevant studies analysing the association between the number of pregnancy losses and the incidence of chromosomal abnormalities were included. Independent data extraction was conducted and study quality was assessed. Meta-analyses were carried out to calculate odds ratios by using fixed- or random-effects models according to statistical homogeneity. RESULTS: A total of 8320 POCs in 19 studies were identified for the meta-analyses. The incidence of chromosomal abnormalities in sporadic pregnancy loss was significantly higher than that in recurrent pregnancy loss. In subgroup analyses, the following studies reported a high incidence of abnormal outcomes of sporadic pregnancy loss: studies with ≥ 300 samples, studies published before 2014, studies conducted in European and American countries, and studies with analyses using conventional karyotype techniques. Moreover, the incidence of chromosomal abnormalities in women with two pregnancy losses was significantly higher than that in women with three or more pregnancy losses. However, there was no difference in the distribution of abnormal types between sporadic and recurrent pregnancy loss or between two and three or more pregnancy losses. CONCLUSIONS: The prevalence of chromosomal abnormalities detected in POCs was lower in recurrent pregnancy loss than in sporadic pregnancy loss, and decreased with an increasing number of pregnancy losses.

Analysis of parental abnormal chromosomal karyotype and subsequent live births in Chinese couples with recurrent pregnancy loss.

The frequency and distribution of chromosomal abnormalities and the impact of parental chromosomal aberration on the pregnancy outcomes of couples with recurrent pregnancy loss remains controversial. 3235 RPL couples who experienced two or more miscarriages before 20 weeks were diagnosed in our tertiary referral hospital during 2008-2018 and included in the single-center retrospective cohort study covering a 10-year period. Chromosome aberration was detected in 121 (3.74%) among 3235 RPL couples which included 75 female and 46 male cases at an individual level. 101 cases were structural aberrations including balanced translocations in 46(38.0%) cases, Robertsonian translocations in 13(10.7%) cases, inversions in 42(34.7%) cases and 20(16.5%) cases were numerical aberrations. 121 carriers and 428 non-carriers were followed up for two years, 55 carriers and 229 non-carriers were subsequent pregnant after diagnosis by natural conception or intrauterine insemination. The frequency of carriers to have a health newborn was not significantly different with non-carriers (72.7% vs. 71.2%, adjusted P = 0.968). This study described the majority of carriers were balanced translocations and chromosome aberrations had a limited influence on live birth rate from the present data. The results of the study also remind us that natural conception may be also a good alternative rather than PGD (Pre-implantation Genetic Diagnosis) which is common in many other reproductive centers for such patients.

LGR6 activates the Wnt/ß-catenin signaling pathway and forms a ß-catenin/TCF7L2/LGR6 feedback loop in LGR6high cervical cancer stem cells.

The leucine-rich repeat-containing G-protein-coupled receptor 6 (LGR6) is considered to be a stem cell marker in many normal tissues and promotes tissue development, regeneration, and repair. LGR6 is also related to the initiation and progression of some malignant tumors. However, the role of LGR6 in cervical cancer has not been reported. Here, immunohistochemistry and western blotting showed that LGR6 was significantly upregulated in cervical cancer, compared with the normal cervix. By analyzing The Cancer Genome Atlas database, LGR6 was found to be correlated with a poor prognosis of cervical cancer. Then, a small population of LGR6high cells isolated by using the fluorescence-activated cell sorting exhibited enhanced properties of cancer stem cells including self-renewal, differentiation, and tumorigenicity. Moreover, RNA sequencing revealed that LGR6 was correlated with the Wnt signaling pathway and TOP/FOP, reverse transcription-PCR, and western blotting further proved that LGR6 could activate the Wnt/ß-catenin signaling pathway. Interestingly, LGR6 upregulated the expression of TCF7L2 by activating the Wnt/ß-catenin pathway. Then, TCF7L2 combining with ß-catenin in the nucleus enhanced LGR6 transcription by binding the promoter of LGR6, which further activated the Wnt signaling to form a positive feedback loop. Thus, our study demonstrated that LGR6 activated a novel ß-catenin/TCF7L2/LGR6-positive feedback loop in LGR6high cervical cancer stem cells (CSCs), which provided a new therapeutic strategy for targeting cervical CSCs to improve the prognosis of cervical cancer patients.

PRDM4 inhibits cell proliferation and tumorigenesis by inactivating the PI3K/AKT signaling pathway through targeting of PTEN in cervical carcinoma.

PR domain zinc finger protein 4 (PRDM4) is a transcription factor that plays key roles in stem cell self-renewal and tumorigenesis. However, its biological role and exact mechanism in cervical cancer remain unknown. Here, both immunohistochemistry (IHC) and Western blot assays demonstrated that the expression of PRDM4 in cervical cancer tissues was much lower than that in the normal cervix. A xenograft assay showed that PRDM4 overexpression in the cervical cancer cell lines SiHa and HeLa dramatically inhibited cell proliferation and tumorigenic potential in vivo. Conversely, the silencing of PRDM4 promoted cervical cancer cell proliferation and tumorigenic potential. Mechanistically, PRDM4 induced cell cycle arrest at the transition from G0/G1 phase to S phase by upregulating p27 and p21 expression and downregulating Cyclin D1 and CDK4 expression. Furthermore, the PI3K/AKT signaling pathway was inactivated in PRDM4-overexpressing cells, which decreased the levels of p-AKT and upregulated the expression of PTEN, an inhibitor of the PI3K/AKT signaling pathway, at both the transcriptional and translational levels. Dual-luciferase reporter assays and qChIP assays confirmed that PRDM4 transactivated the expression of PTEN by binding to two specific regions in the PTEN promoter. Furthermore, PTEN silencing or a PTEN inhibitor rescued the cell defects induced by PRDM4 overexpression. Therefore, our data suggest that PRDM4 inhibits cell proliferation and tumorigenesis by downregulating the activity of the PI3K/AKT signaling pathway by directly transactivating PTEN expression in cervical cancer.

HOXB4 inhibits the proliferation and tumorigenesis of cervical cancer cells by downregulating the activity of Wnt/ß-catenin signaling pathway.

Homeobox B4 (HOXB4), which belongs to the homeobox (HOX) family, possesses transcription factor activity and has a crucial role in stem cell self-renewal and tumorigenesis. However, its biological function and exact mechanism in cervical cancer remain unknown. Here, we found that HOXB4 was markedly downregulated in cervical cancer. We demonstrated that HOXB4 obviously suppressed cervical cancer cell proliferation and tumorigenic potential in nude mice. Additionally, HOXB4-induced cell cycle arrest at the transition from the G0/G1 phase to the S phase. Conversely, loss of HOXB4 promoted cervical cancer cell growth both in vitro and in vivo. Bioinformatics analyses and mechanistic studies revealed that HOXB4 inhibited the activity of the Wnt/ß-catenin signaling pathway by direct transcriptional repression of ß-catenin. Furthermore, ß-catenin re-expression rescued HOXB4-induced cervical cancer cell defects. Taken together, these findings suggested that HOXB4 directly transcriptional repressed ß-catenin and subsequently inactivated the Wnt/ß-catenin signaling pathway, leading to significant inhibition of cervical cancer cell growth and tumor formation.

Hexokinase 2 Promotes Cell Growth and Tumor Formation Through the Raf/MEK/ERK Signaling Pathway in Cervical Cancer.

Hexokinase 2 (HK2) is a member of the hexokinases (HK) that has been reported to be a key regulator during glucose metabolism linked to malignant growth in many types of cancers. In this study, stimulation of HK2 expression was observed in squamous cervical cancer (SCC) tissues, and HK2 expression promoted the proliferation of cervical cancer cells in vitro and tumor formation in vivo by accelerating cell cycle progression, upregulating cyclin A1, and downregulating p27 expression. Moreover, transcriptome sequencing analysis revealed that MAPK3 (ERK1) was upregulated in HK2-overexpressing HeLa cells. Further experiments found that the protein levels of p-Raf, p-MEK1/2, ERK1/2, and p-ERK1/2 were increased in HK2 over-expressing SiHa and HeLa cells. When ERK1/2 and p-ERK1/2 expression was blocked by an inhibitor (FR180204), reduced cyclin A1 expression was observed in HK2 over-expressing cells, with induced p27 expression and inhibited cell growth. Therefore, our data demonstrated that HK2 promoted the proliferation of cervical cancer cells by upregulating cyclin A1 and down-regulating p27 expression through the Raf/MEK/ERK signaling pathway.

NF-YA promotes the cell proliferation and tumorigenic properties by transcriptional activation of SOX2 in cervical cancer.

NF-YA is considered as a crucial regulator for the maintenance of cancer stem cell (CSC) and involved in various types of malignant tumours. However, the exact function and molecular mechanisms of NF-YA in the progression of cervical cancer remains poorly understood. Here, the expression of NF-YA detected by immunohistochemistry was gradually increased from normal cervical tissues, to the high-grade squamous intraepithelial lesions, and then to cervical cancer tissues. NF-YA promoted the cell proliferation and tumorigenic properties of cervical cancer cells as well as tumorsphere formation and chemoresistance in vitro. The luciferase reporter assay combined with mutagenesis analyses and Western blotting showed that NF-YA trans-activated the expression of SOX2 in cervical cancer. Furthermore, quantitative chromatin immunoprecipitation (qChIP) and electrophoretic mobility shift assay (EMSA) confirmed that NF-YA protein directly bound to the CCAAT box region located upstream of the SOX2 promoter. Together, our data demonstrated that NF-YA was highly expressed in cervical cancer and promoted the cell proliferation, tumorigenicity and CSC characteristic by trans-activating the expression of SOX2.

HOXA5 inhibits the proliferation and neoplasia of cervical cancer cells via downregulating the activity of the Wnt/ß-catenin pathway and transactivating TP53.

HOXA5 is considered a regulator involved in embryonic development and cellular differentiation and a tumor suppressor. Nevertheless, its biological role in cervical carcinoma is still unclear. In the present study, immunohistochemistry showed that HOXA5 expression gradually decreased as the degree of cervical lesions deepened. Ectopic expression of HOXA5 restrained cell proliferation, decreased cell viability, and inhibited tumor formation in vitro and in vivo. Furthermore, the expression of HOXA5 could arrest cell cycle from G0/G1 to S phase. RNA-seq revealed that p21 and cyclinD1 were involved in this process. Moreover, the gene set enrichment analysis and the TOP/FOP reporter assay both suggested that HOXA5 could restrain the activity of the Wnt/ß-catenin pathway. Further study using dual-luciferase reporter assay and quantitative chromatin immunoprecipitation assay demonstrated that HOXA5 could directly bind to the TAAT motif within the promoter of TP53 by its HD domain and transactivate TP53, which can upregulate p21. Altogether, our data suggest that HOXA5 inhibits the proliferation and neoplasia via repression activity of the Wnt/ß-catenin pathway and transactivating TP53 in cervical cancer.

NF-YA transcriptionally activates the expression of SOX2 in cervical cancer stem cells.

Roles for SOX2 have been extensively studied in several types of cancer, including colorectal cancer, glioblastoma and breast cancer, with particular emphasis placed on the roles of SOX2 in cancer stem cell. Our previous study identified SOX2 as a marker in cervical cancer stem cells driven by a full promoter element of SOX2 EGFP reporter. Here, dual-luciferase reporter and mutagenesis analyses were employed, identifying key cis-elements in the SOX2 promoter, including binding sites for SOX2, OCT4 and NF-YA factors in SOX2 promoter. Mutagenesis analysis provided additional evidence to show that one high affinity-binding domain CCAAT box was precisely recognized and bound by the transcription factor NF-YA. Furthermore, overexpression of NF-YA in primitive cervical cancer cells SiHa and C33A significantly activated the transcription and the protein expression of SOX2. Collectively, our data identified NF-YA box CCAAT as a key cis-element in the SOX2 promoter, suggesting that NF-YA is a potent cellular regulator in the maintenance of SOX2-positive cervical cancer stem cell by specific transcriptional activation of SOX2.

REX1 promotes EMT-induced cell metastasis by activating the JAK2/STAT3-signaling pathway by targeting SOCS1 in cervical cancer.

ZFP42 zinc finger protein (REX1), a pluripotency marker in mouse pluripotent stem cells, has been identified as a tumor suppressor in several human cancers. However, the function of REX1 in cervical cancer remains unknown. Both IHC and western blot assays demonstrated that the expression of REX1 protein in cervical cancer tissue was much higher than that in normal cervical tissue. A xenograft assay showed that REX1 overexpression in SiHa and HeLa cells facilitated distant metastasis but did not significantly affect tumor formation in vivo. In addition, in vitro cell migration and invasion capabilities were also promoted by REX1. Mechanistically, REX1 overexpression induced epithelial-to-mesenchymal transition (EMT) by upregulating VIMENTIN and downregulating E-CADHERIN. Furthermore, the JAK2/STAT3-signaling pathway was activated in REX1-overexpressing cells, which also exhibited increased levels of p-STAT3 and p-JAK2, as well as downregulated expression of SOCS1, which is an inhibitor of the JAK2/STAT3-signaling pathway, at both the transcriptional and translational levels. A dual-luciferase reporter assay and qChIP assays confirmed that REX1 trans-suppressed the expression of SOCS1 by binding to two specific regions of the SOCS1 promoter. Therefore, all our data suggest that REX1 overexpression could play a crucial role in the metastasis and invasion of cervical cancer by upregulating the activity of the JAK2/STAT3 pathway by trans-suppressing SOCS1 expression.

SALL4 promotes the tumorigenicity of cervical cancer cells through activation of the Wnt/ß-catenin pathway via CTNNB1.

SALL4 is overexpressed in many cancers and is found to be involved in tumorigenesis and tumor progression. However, the function of SALL4 in cervical cancer remains unknown. Here, we showed that the expression of SALL4 was gradually increased from normal cervical tissue to high-grade squamous intraepithelial lesions and then to squamous cervical carcinoma. SALL4 was upregulated or downregulated in cervical cancer cells by stably transfecting a SALL4-expressing plasmid or a shRNA plasmid targeting SALL4, respectively. In vitro, cell growth curves and MTT (3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide) assays showed that SALL4 promoted the cell proliferation of cervical cancer cells. In vivo, xenograft experiments verified that SALL4 enhanced the tumor formation of cervical cancer cells in female BALB/c Nude mice. Cell cycle analysis by fluorescence-activated cell sorting found that SALL4 accelerates cell cycle transition from the G0 /G1 phase to the S phase. TOP/FOP-Flash reporter assay revealed that SALL4 significantly upregulates the activity of Wnt/ß-catenin pathway. Western blotting showed that the expression levels of ß-catenin and important downstream genes, including c-Myc and cyclin D1, were increased by SALL4 in cervical cancer cells. Furthermore, dual-luciferase reporter and chromatin immunoprecipitation assays confirmed that SALL4 transcriptionally activated CTNNB1 by physically interacting with its promoters. Taken together, The results of this study demonstrated that SALL4 may promote cell proliferation and tumor formation of cervical cancer cells by upregulating the activity of the Wnt/ß-catenin signaling pathway by directly binding to the CTNNB1 promoter and trans-activating CTNNB1.

SOX17 restrains proliferation and tumor formation by down-regulating activity of the Wnt/ß-catenin signaling pathway via trans-suppressing ß-catenin in cervical cancer.

The SRY-box containing gene 17 (SOX17) is considered as a regulator in stemness maintenance and a suppressor in some malignant tumors. However, the biological function and molecular mechanism of SOX17 in the process of initiation and progression of cervical cancer remain obscure. In this study, immunohistochemistry showed that the expression of SOX17 was high in the normal cervix, moderate in the high-grade squamous intraepithelial lesion, and low in the cervical cancer. SOX17 inhibited the proliferation and viability of cervical cancer cells in vitro as well as tumor formation in vivo. Additionally, SOX17 induced the cell cycle arrest at the transition from the G0/G1 phase to the S phase. The TOP/ FOP-Flash reporter assay and Western blotting showed SOX17 inhibited the activity of the Wnt/ß-catenin signaling pathway in cervical cancer. Further, firefly luciferase reporter assay and quantitative chromatin immunoprecipitation (qChIP) assays confirmed that SOX17 trans-suppressed the expression of ß-catenin by directly binding to the specific region of the ß-catenin promoter. Together, our data demonstrated that SOX17 restrained the proliferation and tumor formation by down-regulating the activity of the Wnt/ß-catenin signaling pathway via trans-suppression of ß-catenin in cervical cancer.