TDG suppresses the migration and invasion of human colon cancer cells via the DNMT3A/TIMP2 axis.

Background: Colorectal cancer (CRC) is one of the most common malignant tumors with high rates of recurrence and mortality. Thymine DNA glycosylase (TDG) is a key molecule in the base excision repair pathway. Recently, increasing attention has been paid to the role of TDG in tumor development. However, the specific functions of TDG in CRC remain unclear. Methods: The biological functions of TDG and DNA methyltransferase 3 alpha (DNMT3A) in CRC were evaluated using migration and invasion assays, respectively. A tumor metastasis assay was performed in nude mice to determine the in vivo role of TDG. The interaction between TDG and DNMT3A was determined via co-immunoprecipitation (Co-IP). Chromatin immunoprecipitation analysis (ChIP) was used to predict the DNA-binding site of DNMT3A. We also performed methylation-specific PCR (MSP) to detect changes in TIMP2 methylation. Results: TDG inhibited the migration and invasion of human colon cancer cells both in vitro and in vivo. TDG promoted the ubiquitination and degradation of DNMT3A by binding to it. Its interference with siDNMT3A also inhibits the migration and invasion of human colon cancer cells. Furthermore, the ChIP, MSP, and rescue experiments results confirmed that TDG accelerated the degradation of DNMT3A and significantly regulated the transcription and expression of TIMP2, thereby affecting the migration and invasion of human colon cancer cells. Conclusion: Our findings reveal that TDG inhibits the migration and invasion of human colon cancer cells through the DNMT3A-TIMP2 axis, which may be a potential therapeutic strategy for the development and treatment of CRC.

THAP11 Functions as a Tumor Suppressor in Gastric Cancer through Regulating c-Myc Signaling Pathways.

We aim to investigate the role of THAP11 (thanatos-associated protein11) in gastric cancer and its regulation mechanisms. THAP11 expression was analyzed in 51 pairs of GC tissues and the corresponding paracancerous tissues by qRT-PCR and Western blot. After THAP11 was overexpressed or knocked-down, cell proliferation, cell cycle, and apoptosis were detected in MKN-45 cells. We found that THAP11 was significantly downregulated in GC tissues and GC cell lines. Functionally, THAP11 overexpression markedly inhibited cell growth, induced G1/G0 cell-cycle arrest, and promoted cell apoptosis of MKN-45 cells, while silencing of THAP11 led to increased cell growth, increased DNA synthesis, and inhibited apoptosis. In addition, THAP11 negatively regulated the expression of c-Myc, decreased cyclinD1 protein, and increased p27 and p21 protein levels. We also found cell growth suppression induced by THAP11 was rescued by c-Myc overexpression, further confirming that THAP11 suppresses gastric cancer cell growth via the c-Myc pathway. THAP11 acts as a cell growth suppressor and exerts its role possibly through negatively regulating c-Myc pathway in gastric cancer.

Nei Endonuclease VIII-Like1 (NEIL1) Inhibits Apoptosis of Human Colorectal Cancer Cells.

The study is aimed at investigating the role of Nei endonuclease VIII-like1 (NEIL1) in the pathogenesis of colorectal cancer (CRC). The human CRC (HCT116 and SW480) cells were subjected to the siRNA silencing and recombinant plasmid overexpression of NEIL1. Transfection of siNEIL1 significantly inhibited the cell growth. It also increased the Bax expression levels, while it decreased the Bcl-2 expression levels in human CRC cells, leading the Bax/Bcl-2 balance toward apoptosis. Moreover, the apoptosis was promoted through the caspase-9 signaling pathway. One the other hand, high expression of NEIL1 promoted the cell viability and reduced the apoptosis, inducing the balance of Bax/Bcl-2 in the human colon cancer cells to be antiapoptotic. In addition, the caspase-9 signaling pathway inhibited apoptosis, contrary to the results obtained by downregulating NEIL1 expression. Furthermore, NEIL1 was negatively regulated by miR-7-5p, indicating that miR-7-5p inhibited the NEIL1 expression after transcription. Overexpression of miR-7-5p reversed the effects of NEIL1 on these CRC cells. In conclusion, NEIL1 promotes the proliferation of CRC cells, which is regulated negatively by miR-7-5p. These findings suggest that NEIL1 is a potential therapeutic target for CRC.

miR-203a suppresses cell proliferation by targeting RING-finger protein 6 in colorectal cancer.

Colorectal cancer (CRC) is one of most common cancers worldwide. Although miR-203a is reported as a tumor suppressor involved in cell progression in some cancers, the role of miR-203a in CRC is still controversial and the underling mechanism of miR-203a in CRC remains unclear. Here, we demonstrated that low expression of miR-203a had poorer survival in CRC patients. miR-203a was down-regulated in most human colon cancer cells. Overexpression of miR-203a could inhibit colon cancer cell proliferation and arrest cell cycle in G1 phase. Bioinformatics and dual luciferase reporter assay confirmed that RING-finger protein 6 (RNF6) was a target gene of miR-203a. Silencing RNF6 inhibited cell proliferation and arrest cell cycle in G1 phase. RNF6 overexpression reversed the effects of miR-203a overexpression in colon cancer cells. Taken together, our data indicate that miR-203a inhibits colon cancer cell proliferation by targeting RNF6, offer novel insights into the regulatory network of miR-203a-modulated cell cycle and proliferation, and suggest that miR-203a a potential therapeutic target in CRC treatment.

Norepinephrine transporter promotes the invasion of human colon cancer cells.

Epidemiological studies suggested the use of antidepressants to be associated with decreased risk of colorectal cancer (CRC). However, the underlying mechanism through which this decreased risk occurs remains elusive. The norepinephrine transporter (NET) is a target of antidepressants that maintains noradrenergic transmission homeostasis; however, little is known about its function in human CRC cells. The present study, using public datasets and immunohistochemistry approaches, revealed that NET was highly expressed in human CRC tissues with metastasis and in human colon cancer cells. Furthermore, knockdown of NET inhibited the invasive capability of human colon cancer cells. Additionally, epithelial (E)-cadherin expression was increased and Notch1 signaling was inhibited in NET-depleted colon cancer cells. These findings suggest that NET is highly expressed in human colon cancer, which is associated with the invasion of human colon cancer cells by influencing cell-cell adhesion through the Notch1-E-cadherin pathway. Thus, the present study revealed a novel function for NET and its downstream effectors in colon cancer cells, which will be valuable for future studies in a clinical setting.

MicroRNA-497-5p Induces Cell Cycle Arrest Of Cervical Cancer Cells In S Phase By Targeting CBX4.

PURPOSE: miR-497-5p can inhibit cervical cancer cell proliferation. However, the underlying mechanism remains to be elucidated. METHODS: Bioinformatics was used to analyze the target genes of miR-497-5p. qRT-PCR and Western blot were used to analyze mRNA and protein expression, respectively. Dual-luciferase reporter assay was used to analyze the direct binding between miR-497-5p and 3'-untranslated region of CBX4. Cell viability was measured with MTT assay. Flow cytometry was performed to detect cell cycle distribution. RESULTS: Here, using bioinformatics methods we firstly found that miR-497-5p regulated cervical carcinoma proliferation by targeting polycomb chromobox4 (CBX4). Expression of miR-497-5p in cervical carcinoma tissues was negatively correlated with CBX4. A binding region of miR-497-5p in 3'-untranslated region of CBX4 was predicted. Further experiments confirmed that miR-497-5p directly targeted CBX4. Besides, RNA interference of CBX4 inhibited cervical cancer cell proliferation, arrested cells at S phase and reduced the expression of CDK2 and Cyclin A2 proteins. The use of miR-497-5p inhibitor compromised CBX4 interference RNAs induced cycle arrest of cervical cancer cells. Cells co-transfected with miR-497-5p inhibitors and CBX4 interference RNAs had a higher proliferation rate than CBX4 inference RNA-transfected cells. CONCLUSION: All together, the present study demonstrates that miR-497-5p inhibits cervical cancer cells proliferation by directly targeting CBX4.

The effect of MTHFD2 on the proliferation and migration of colorectal cancer cell lines.

PURPOSE: Methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) is a tetramethylfolate dehydrogenase enzyme involved in folate metabolism. The aim of this study is to determine the effect of MTHFD2 on the proliferation and metastasis of colorectal cancer (CRC). PATIENTS AND METHODS: MTHFD2 was silenced or overexpressed in CRC cells. qRT-PCR and Western blotting were used to analyze mRNA and protein expression, respectively. The MTT assay and colony forming assay were used to detect cell proliferation and colony formation ability. The cycle and apoptosis changes were detected by flow cytometry. Transwell experiments were used to analyze the migration ability of CRC cells. RESULTS: The expression of MTHFD2 in 31 kinds of cancers was analyzed by bioinformatics, and MTHFD2 was found highly expressed in various cancer cells including CRC cells. Silencing the expression of MTHFD2 resulted in inhibition of the proliferation of CRC cells, weakening of the migration ability, blocking of the cell cycle in G0/G1-S phase, and promotion of the apoptosis of CRC cells. On the contrary, overexpression of MTHFD2 in CRC cells resulted in enhancement of the proliferation and migration ability, promotion of cell cycle progression and inhibition of cell apoptosis. CONCLUSION: MTHFD2 is positively related with colorectal cancer and the MTHFD2 gene is a tumor promoting gene in CRC cells.

Effect of mGluR7 on proliferation of human embryonic neural stem cells.

This study is to investigate the effect of metabotropic glutamate receptor 7 (mGluR7) on the proliferation of human embryonic neural stem cells (NSCs) and its molecular mechanism.Human embryonic NSCs were isolated. The pCMV2-GV146-GFP-mGluR7 plasmid was transfected to over-express mGluR7 while mGluR7 siRNA was transfected to knockdown mGluR7. MTT assay was used to analyze cell proliferation. Flow cytometry was used to detect cell cycle and apoptosis. Protein and mRNA levels were analyzed by Western blot and RT-qPCR, respectively.The viability of human NSCs and the diameter of neurospheres after 24 hours, 48 hours, and 72 hours of transfection significantly increased by mGluR7 overexpression whereas significantly decreased by mGluR7 knockdown. Ki-67 expression was up-regulated by mGluR7 overexpression whereas down-regulated by mGluR7 siRNA, indicating a promotive effect of mGluR7 on NSC proliferation. After mGluR7 overexpression, G1/G0 phase cell ratio dropped significantly compared with control group, while the S phase cell ratio increased. mGluR7 silencing arrested human NSCs at G1/G0 phase. After 48 hours of transfection, there was a decrease of apoptosis by mGluR7 overexpression, while mGluR7 silencing induced apoptosis of human NSCs. Additionally, overexpression of mGluR7 up-regulated the expression of p-serine/threonine kinase (AKT), cyclin D1, and cyclin-dependent kinase 2 (CDK2). The mGluR7 knockdown had opposite effects. Similarly, mGluR7 down-regulated the expression of Caspase-3/9, while the mGluR7 knockdown promoted this.mGluR7 can promote the proliferation of human embryonic cortical NSCs in vitro. This effect may be mediated by promoting cell cycle progression, inhibiting cell apoptosis, activating the AKT signaling pathway, and inhibiting the Caspase-3/9 signaling pathway.

Effect and mechanism of mGluR6 on the biological function of rat embryonic neural stem cells.

Here, we investigated the effects and molecular mechanisms of metabotropic glutamate receptor 6 (mGluR6) on rat embryonic neural stem cells (NSCs). Overexpression of mGluR6 significantly promoted the proliferation of NSCs and increased the diameter of neutrospheres after treatment for 24 h, 48 h and 72 h. Overexpression of mGluR6 promoted G1 to S phase transition, with significantly decreased cell ratio in G1/G0 phase but significantly increased cell ratio in S phase. Additionally, mGluR6 overexpression for 48 h decreased the early and late apoptosis significantly. Moreover, overexpression of mGluR6 significantly increased the expression of p-ERK1/2, Cyclin D1 and CDK2, while the expression of p-p38 was significantly decreased. On the contrary, these effects of mGluR6 overexpression were reversed by mGluR6 knockdown. In conclusion, mGluR6 promotes the proliferation of NSCs by activation of ERK1/2-Cyclin D1/CDK2 signaling pathway and inhibits the apoptosis of NSCs by blockage of the p38 MAPK signaling pathway.

Proteomic Screening for Serum Biomarkers for Cervical Cancer and Their Clinical Significance.

BACKGROUND The present study aimed to determine serum markers for cervical cancer (CC) and to provide valuable references for clinical diagnosis and treatment. MATERIAL AND METHODS Serum samples were collected from age-matched healthy control women, and from female CC patients before and after surgery. Serum biomarkers were selected by comparing serum peptides profiles among the 3 groups by magnetic bead-based weak cation - exchange chromatography fractionation combined with matrix-assisted laser desorption/ionization-time of flight mass spectrometry. Probable serum biomarkers for cervical cancer were then further identified by liquid chromatography-electrospray ionization-tandem mass spectrometry system and the identified proteins were verified by enzyme-linked immunosorbent assay (ELISA). RESULTS Three peptide biomarkers were identified for distinguishing CC patients from normal individuals, and distinguishing preoperative CC patients from postoperative CC patients. Of these 3 identified protein peptide regions, 2 peptide regions - TKT (Peak 2, 2435.63 m/z, 499-524) and FGA (Peak 4, 2761.79 m/z, 603-629) - were identified as upregulated markers, and peptide region of APOA1 (Peak 9, 2575.3 m/z, 245-260) was identified as a downregulated biomarker in preoperative CC patients compared with healthy women. CONCLUSIONS The present study provides a new method for identifying potential serum biomarkers for CC patients.

miR-30a-3p Targets MAD2L1 and Regulates Proliferation of Gastric Cancer Cells.

PURPOSE: This study was done to investigate the inhibition effects of miR-30a-3p on mitotic arrest deficient 2 like 1 (MAD2L1) expression and the proliferation of gastric cancer cells. PATIENTS AND METHODS: Cluster analysis and the TCGA database were used to screen the key genes highly expressed in gastric cancer. Based on the LinkedOmics website, the correlation between the miR-30a-3p and the cell cycle-related target gene MAD2L1 in gastric cancer was analyzed. The mRNA and protein expression levels were detected with the quantitative real-time PCR and Western blot analysis. The cell proliferation and cell cycle were also detected and analyzed. RESULTS: Bioinformatics analysis showed that MAD2L1 was highly expressed in tumor tissues compared with normal tissues. Compared with normal tissues, the miR-30a-3p was significantly decreased in the gastric cancer tissues. Moreover, MAD2L1 was significantly negatively correlated with the miR-30a-3p expression. Furthermore, over-expression of miR-30a-3p decreased the expression of MAD2L1 at the protein level, which inhibited the proliferation of AGS and BGC-823 gastric cancer cells. In addition, the cell cycles of AGS and BGC-823 cells were arrested at the G0/G1 phase. CONCLUSION: MAD2L1 is a pro-oncogene which is up-regulated in gastric cancer. The miR-30a-3p can down-regulate the MAD2L1 expression, inhibiting the proliferation of gastric cancer cells and affect the cell cycle.