While it is known that spermatogonial stem cells (SSCs) initiate the production of male germ cells, the mechanisms of SSC self-renewal, proliferation, and differentiation remain poorly understood. We have previously identified Strawberry Notch 1 (SBNO1), a vertebrate strawberry notch family protein, in the proteome profile for mouse SSC maturation and differentiation, revealing SBNO1 is associated with neonatal testicular development. To explore further the location and function of SBNO1 in the testes, we performed Sbno1 gene knockdown in mice to study the effects of SBNO1 on neonatal testicular and SSC development. Our results revealed that SBNO1 is required for neonatal testicular and SSC development in mice. Particularly, in vitro Sbno1 gene knockdown with morpholino oligonucleotides caused a reduction of SSCs and inactivation of the noncanonical Wnt pathway, through Jun N-terminal kinases. Our study suggests SBNO1 maintains SSCs by promoting the noncanonical Wnt pathway.
Adult Germline Stem Cells/metabolism* ; Animals ; Cell Proliferation/physiology* ; Gene Knockdown Techniques ; Male ; Mice ; Proteome ; Repressor Proteins/metabolism* ; Testis/metabolism* ; Wnt Signaling Pathway/physiology*

Hepatocellular carcinoma (HCC) is one of the most common types of liver cancer and is the second leading cause of cancer mortality with an estimated 745 500 deaths annually (Jemal et al., 2011). Although new therapeutic modalities including novel chemotherapeutic interventions and targeted therapy have been applied, the prognosis of HCC patients remains unsatisfactory due to the high incidence of intrahepatic and distal metastases (Siegel et al., 2018).
Apoptosis Regulatory Proteins/physiology* ; Biomarkers ; Carcinoma, Hepatocellular/pathology* ; Female ; Genome ; Humans ; Hypoxia ; Liver Neoplasms/pathology* ; Male ; MicroRNAs/analysis* ; Neoplasm Staging ; Prognosis ; Repressor Proteins/physiology*

Proline-, glutamic acid-, and leucine-rich protein 1 (PELP1) is a scaffolding protein involved in both genomic and nongenomic estrogen signal transduction pathways. To date, the role of PELP1 protein has yet to be characterized in human sperm and has not been associated with sperm parameters. To confirm the presence of PELP1 in human sperm, fresh semen samples were obtained from 178 donors. The study was designed to establish both mRNA and protein presence, and protein cellular localization. Additionally, the number of PELP1-positive spermatozoa was analyzed in men with normal and abnormal semen parameters. Sperm parameters were assessed according to the World Health Organization (WHO) 2010 standards. The presence of PELP1 in spermatozoa was investigated using four precise, independent techniques. The qualitative presence of transcripts and protein was assessed using reverse transcription-polymerase chain reaction (RT-PCR) and western blot protocols, respectively. The cellular localization of PELP1 was investigated by immunocytochemistry. Quantitative analysis of PELP1-positive cells was done by flow cytometry. PELP1 mRNA and protein was confirmed in spermatozoa. Immunocytochemical analysis identified the presence of PELP1 in the midpieces of human sperm irrespective of sperm parameters. Becton Dickinson fluorescence-activated cell sorting (FACSCalibur™) analysis revealed a significantly lower number of PELP1-positive cells in males with normal semen parameters versus abnormal samples (42.78% ± 11.77% vs 61.05% ± 21.70%, respectively; P = 0.014). The assessment of PELP1 may be a time-saving method used to obtain information about sperm quality. The results of our study suggest that PEPL1 may be utilized as an indicator of sperm quality; thereby, PELP1 may be an additional biomarker useful in the evaluation of male infertility.
Adolescent ; Adult ; Biomarkers/metabolism* ; Co-Repressor Proteins/metabolism* ; Humans ; Infertility, Male/metabolism* ; Male ; Middle Aged ; Semen Analysis ; Sperm Motility/physiology* ; Spermatozoa/metabolism* ; Transcription Factors/metabolism* ; Young Adult

We sought to investigate the underlying mechanism of action of the long noncoding RNA (lncRNA) LOC283070 in the development of androgen independence in prostate cancer. The interactions between LOC283070 and target proteins were investigated by RNA pull-down and RNA-binding protein immunoprecipitation (RIP) assays. Subcellular fractionation and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) were used to detect the subcellular localization of LOC283070. Western blotting was performed to detect the expression of prohibitin 2 (PHB2). Luciferase activity assays were performed to evaluate the effects of LOC283070 and PHB2 on the androgen receptor (AR) signaling pathway. A methyl thiazolyl tetrazolium (MTT) assay and a growth curve assay were used to test cell viability. Flow cytometry was performed to analyze cell cycles. A transwell assay was employed to test cell migration. We identified PHB2 as an interaction partner of LOC283070 in the pull-down and RIP experiments. Furthermore, we confirmed that the enrichment of LOC283070 with PHB2 in androgen-independent LNCaP (LNCaP-AI) cells was much greater than that in LNCaP cells. Moreover, the expression of PHB2 was not significantly different between the two cell lines, and the expression of LOC283070 in the nuclei of the LNCaP-AI cells was significantly greater than that in the LNCaP cells. In vitro data revealed that PHB2 overexpression significantly inhibited AR activity and cell proliferation and migration and induced accumulation of prostate cancer cells in G0/G1 phase. Moreover, the overexpression of LOC283070 fully abrogated the effects of PHB2 overexpression. In conclusion, we found that LOC283070 can bind to PHB2 located in the nucleus and inhibit its effect, and this is one of the mechanisms by which LOC283070 is involved in the transition of LNCaP cells into androgen-independent cells.
Androgens/metabolism* ; Cell Line, Tumor ; Cell Proliferation/physiology* ; Gene Expression Regulation, Neoplastic ; Humans ; Male ; Prohibitins ; Prostatic Neoplasms/metabolism* ; RNA, Long Noncoding/metabolism* ; Receptors, Androgen/metabolism* ; Repressor Proteins/metabolism* ; Signal Transduction/physiology*

PURPOSE: In the gastric mucosa of Helicobacter pylori (H. pylori)-infected patients with gastritis or adenocarcinoma, proliferation of gastric epithelial cells is increased. Hyperproliferation is related to induction of oncogenes, such as β-catenin and c-myc. Even though transcription factors NF-κB and AP-1 are activated in H. pylori-infected cells, whether NF-κB or AP-1 regulates the expression of β-catenein or c-myc in H. pylori-infected cells has not been clarified. The present study was undertaken to investigate whether H. pylori-induced activation of NF-κB and AP-1 mediates the expression of oncogenes and hyperproliferation of gastric epithelial cells. MATERIALS AND METHODS: Gastric epithelial AGS cells were transiently transfected with mutant genes for IκBα (MAD3) and c-Jun (TAM67) or treated with a specific NF-κB inhibitor caffeic acid phenethyl ester (CAPE) or a selective AP-1 inhibitor SR-11302 to suppress activation of NF-κB or AP-1, respecively. As reference cells, the control vector pcDNA was transfected to the cells. Wild-type cells or transfected cells were cultured with or without H. pylori. RESULTS: H. pylori induced activation of NF-κB and AP-1, cell proliferation, and expression of oncogenes (β-catenein, c-myc) in AGS cells, which was inhibited by transfection of MAD3 and TAM67. Wild-type cells and the cells transfected with pcDNA showed similar activities of NF-κB and AP-1, proliferation, and oncogene expression regardless of treatment with H. pylori. Both CAPE and SR-11302 inhibited cell proliferation and expression of oncogenes in H. pylori-infected cells. CONCLUSION: H. pylori-induced activation of NF-κB and AP-1 regulates transcription of oncogenes and mediates hyperproliferation in gastric epithelial cells.
Blotting, Western ; Caffeic Acids ; Cell Line, Tumor ; Cell Proliferation ; DNA, Bacterial/analysis/genetics ; DNA-Binding Proteins/*metabolism ; Epithelial Cells/*metabolism ; Gastric Mucosa/*metabolism/pathology ; Gastritis/pathology ; Gene Expression Regulation, Bacterial ; Helicobacter Infections/metabolism/pathology/physiopathology ; Helicobacter pylori/pathogenicity/physiology ; Humans ; NF-kappa B/antagonists & inhibitors/*biosynthesis/metabolism ; Peptide Fragments ; Phenylethyl Alcohol/analogs & derivatives ; Proto-Oncogene Proteins c-jun ; Repressor Proteins ; Transcription Factor AP-1/*biosynthesis ; Transcription Factors/*metabolism ; beta Catenin/*metabolism

BACKGROUNDBrain acid soluble protein 1 (BASP1) is identified as a novel potential tumor suppressor in several cancers. However, its role in thyroid cancer has not been investigated yet. In the present study, the antitumor activities of BASP1 against the growth and migration of thyroid cancer cells were evaluated.

METHODSBASP1 expression in thyroid cancer tissues and normal tissues were examined by immunohistochemical staining and the association between its expression and prognosis was analyzed. pcDNA-BASP1 carrying full length of BASP1 cDNA was constructed to restore the expression of BASP1 in thyroid cancer cell lines (BHT-101 and KMH-2). The cell proliferation in vitro and in vivo was evaluated by WST-1 assay and xenograft tumor models, respectively. Cell cycle distribution after transfection was analyzed using flow cytometry. Cell apoptosis after transfection was examined by annexin V/propidium iodide assay. The migration was examined using transwell assay.

RESULTSBASP1 expression was abundant in normal tissues while it is significantly decreased in cancer tissues (P = 0.000). pcDNA-BASP1 restored the expression of BASP1 and significantly inhibited the growth of BHT-101 and KMH-2 cells as well as xenograft tumors in nude mice (P = 0.000). pcDNA-BASP1 induced G1 arrest and apoptosis in BHT-101 and KMH-2 cells. In addition, pcDNA-BASP1 significantly inhibited the cell migration.

CONCLUSIONSDownregulation of BASP1 expression may play a role in the tumorigenesis of thyroid cancer. Restoration of BASP1 expression exerted extensive antitumor activities against growth and migration of thyroid cancer cells, which suggested that BASP1 gene might act as a potential therapeutic agent for the treatment of thyroid cancer.

Aged ; Animals ; Apoptosis ; genetics ; physiology ; Calmodulin-Binding Proteins ; genetics ; metabolism ; Cell Cycle ; genetics ; physiology ; Cell Line, Tumor ; Cell Movement ; genetics ; physiology ; Cell Proliferation ; genetics ; physiology ; Cytoskeletal Proteins ; genetics ; metabolism ; Female ; Gene Expression Regulation, Neoplastic ; genetics ; physiology ; Humans ; Male ; Membrane Proteins ; genetics ; metabolism ; Mice ; Mice, Nude ; Middle Aged ; Nerve Tissue Proteins ; genetics ; metabolism ; Repressor Proteins ; genetics ; metabolism ; Thyroid Neoplasms ; metabolism ; pathology ; Xenograft Model Antitumor Assays

OBJECTIVETo investigate the expression and possible roles of Wnt inhibitory factor-1 (Wif-1) and β-catenin in the Wnt pathway in childhood acute lymphoblastic leukemia (ALL).

METHODSThe clinical data of 35 children who had newly-diagnosed ALL and achieved complete remission on day 33 of remission induction therapy were retrospectively reviewed. The children before treatment were considered as the incipient group, and those who achieved complete remission on day 33 were considered as the remission group. Fifteen children with non-malignant hematologic diseases were enrolled as the control group. RT-PCR was used to measure the mRNA expression of Wif-1 and β-catenin. ELISA was used to measure the protein expression of Wif-1.

RESULTSCompared with the control and remission groups, the incipient group had significantly lower mRNA and protein expression of Wif-1 and significantly higher mRNA expression of β-catenin (P<0.05). In the incipient and remission groups, high-risk children showed significantly higher mRNA expression of β-catenin and significantly lower mRNA and protein expression of Wif-1 than the medium- and low-risk children (P<0.05). In the incipient and remission group, the children with T-cell acute lymphoblastic leukemia showed significantly higher mRNA expression of β-catenin and significantly lower mRNA and protein expression of Wif-1 compared with those with B-lineage acute lymphoblastic leukemia (P<0.05). In each group, there was a negative correlation between the mRNA expression of Wif-1 and β-catenin (P<0.05).

CONCLUSIONSReduced expression of Wif-1 and increased expression of β-catenin may be involved in the pathogenesis of childhood ALL, and the degree of reduction in Wif-1 and/or increase in β-catenin may be related to prognosis.

Adaptor Proteins, Signal Transducing ; genetics ; physiology ; Adolescent ; Child ; Child, Preschool ; Female ; Humans ; Infant ; Male ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; etiology ; physiopathology ; RNA, Messenger ; analysis ; Repressor Proteins ; genetics ; physiology ; Wnt Signaling Pathway ; physiology ; beta Catenin ; genetics ; physiology

To investigate the expression of metastasis tumor-associated protein 2 (MTA2) in cervical squamous carcinoma and its relationship with prognosis.
 Methods: Immunohistochemistry and real-time PCR were performed to determine the expression and distribution of MTA2 mRNA and protein in normal cervical tissue, cervical intraepithelial neoplasia (CIN) and cervical squamous carcinomas tissues, then its relationship with clinical pathological factors and prognosis was analyzed.
 Results: The positive rate of MTA2 protein in normal cervical tissue, CIN and cervical squamous cell carcinomas tissues were 0, 30.0%, 73.4%, respectively. The positive rate was associated with international federation of gynecology and obstetrics (FIGO) stage and lymph node metastasis, whereas there was no correlation with the age of patients or the degree of tumor differentiation. The expression of MTA2 mRNA in normal cervical tissue, CIN and cervical squamous carcinomas tissues was 0.437±0.028, 0.737±0.102 and 1.172±0.068, respectively. The positive rate was associated with FIGO stage and lymph node metastasis, whereas there was no correlation with the age of patients or the degree of tumor differentiation. The result of survival analysis showed poor overall survival time in the patients with high expression of MTA2. Multivariate COX proportional hazards model showed that the positive expression of MTA2 protein, FIGO stage and the metastasis of lymph node were independent prognostic factors for unfavorable clinical outcome of cervical cancer.
 Conclusion: The positive expression of MTA2 was closely related to the development, invasion and metastasis of cervical squamous cell carcinomas. The positive expression of MTA2 protein, FIGO stage and the metastasis of lymph node were independent prognostic factors for unfavorable clinical outcome of cervical cancer. The expression of MTA2 could be used as a potential molecular marker in evaluating the prognosis of cervical squamous cell carcinomas.
Biomarkers, Tumor ; genetics ; Carcinoma, Squamous Cell ; genetics ; mortality ; Cervical Intraepithelial Neoplasia ; genetics ; Female ; Gene Expression ; physiology ; Histone Deacetylases ; physiology ; Humans ; Immunohistochemistry ; Lymph Nodes ; Lymphatic Metastasis ; genetics ; Neoplasm Staging ; Prognosis ; Proportional Hazards Models ; Real-Time Polymerase Chain Reaction ; Repressor Proteins ; physiology ; Survival Analysis ; Uterine Cervical Neoplasms ; genetics ; mortality

BACKGROUNDAs a novel molecular markerof non-small cell lung cancer (NSCLC), PRDI-BF1 and RIZ homology domain containing protein 14 (PRDM14) is over-expressed in NSCLC tumor tissues. Extracellular matrix degradation mediated by the balance between matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases (TIMPs) is one of the most important mechanism in lung cancer metastasis. This study aimed to determine if PRDM14 promoted the migration of NSCLC cells through extracellular matrix degradation mediated by change of MMP/TIMP expression.

METHODSThe expression of PRDM14 was down-regulated in human cell line A 549 after transfection with lentiviral vector-mediated short-hairpin ribonucleic acids (shRNAs) which targeted the PRDM14 promoter. Cellular migration of shRNA-infected cells was detected by a scratch wound healing assay and transwell cell migration assay. Expression levels of MMP1, MMP2, TIMP1, and TIMP2 were measured by quantitative real-time polymerase chain reaction (RT-PCR).

RESULTSMigration of PRDM14-shRNA-infected cells was significantly inhibited relative to control cells as measured by the scratch wound healing (P < 0.05) and transwell cell migration assays (P < 0.01). The expression of MMP1 in A549 cells infected by PRDM14-shRNA was down-regulated significantly (P < 0.01), whereas the expression of TIMP1 and TIMP2 was up-regulated significantly (P < 0.01).

CONCLUSIONSPRDM14 accelerates A549 cells migration in vitro through extracellular matrix degradation. PRDM14 is considered as a potential therapeutic target in metastatic NSCLC.

Carcinoma, Non-Small-Cell Lung ; metabolism ; Cell Line, Tumor ; Cell Movement ; genetics ; physiology ; Extracellular Matrix ; metabolism ; Humans ; Matrix Metalloproteinase 1 ; metabolism ; Matrix Metalloproteinase 2 ; metabolism ; Neoplasm Metastasis ; genetics ; Repressor Proteins ; metabolism ; Tissue Inhibitor of Metalloproteinase-1 ; metabolism ; Tissue Inhibitor of Metalloproteinase-2 ; metabolism

Human papillomaviruses (HPVs) including high-risk (HR) and low-risk (LR) subtypes have distinguishable variation on both genotypes and phenotypes. The co-infection of multiple HR-HPVs, headed by HPV16, is common in cervical cancer in female. Recently accumulating reports have focused on the interaction between virus and host, particularly the role of human microRNAs (miRNAs) in anti-viral defense by targeting viral genome. Here, we found a well-conserved target site of miRNAs in the genomes of most HR-HPVs, not LR-HPVs, by scanning all potential target sites of human miRNAs on 24 HPVs of unambiguous subtypes of risk. The site is targeted by two less common human miRNAs, miR-875 and miR-3144, and is located in E6 oncogene open reading frame (ORF) and overlap with the first alternative splice exon of viral early transcripts. In validation tests, miR-875 and miR-3144 were identified to suppress the target reporter activity markedly and inhibit the expression of both synthetically exogenous E6 and endogenous E6 oncogene. High level of two miRNAs can inhibit cell growth and promote apoptosis in HPV16-positive cervical cancer cells. This study provides a promising common target of miRNAs for most HR-HPVs and highlights the effects of two low expressed human miRNAs on tumour suppression.
Apoptosis ; genetics ; Base Sequence ; Binding Sites ; genetics ; Cell Line, Tumor ; Cell Proliferation ; genetics ; Female ; Gene Expression ; Host-Pathogen Interactions ; genetics ; Human papillomavirus 16 ; genetics ; physiology ; Humans ; MicroRNAs ; genetics ; Microscopy, Fluorescence ; Molecular Sequence Data ; Oncogene Proteins, Viral ; genetics ; Repressor Proteins ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Homology, Nucleic Acid ; Uterine Cervical Neoplasms ; genetics ; pathology ; virology