B-cell Translocation Gene 2 Gene Research Articles

Prognostic effect of DNA methylation of BTG2 gene in Chinese hepatocellular carcinoma

BackgroundThis study aims to develop a prognostic model for overall survival based on potential methylation sites within B-cell translocation gene 2 (BTG2) in Chinese patients with hepatocellular carcinoma (HCC). MethodsThis is a retrospective study. The beta values of nine CpG sites and RSEM normalized count values of BTG2 gene were extracted from the Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) (TCGA-LIHC) dataset, with the beta value representing the methylation level by indicating the ratio of the intensity of the methylated bead type to the combined locus intensity. Pyrosequencing was performed to determine the range of methylation values surrounding cg01798157 site in BTG2 gene. A weighted linear model was developed to predict the overall survival (OS). ResultsThe beta value of cg01798157 was significantly negatively associated with the mRNA expression of BTG2 in the TCGA-LIHC dataset (Spearman's rho = −0.5306, P = 2.27 × 10−27). The methylation level of cg01798157 was significantly associated with OS in the cohort of 51 Chinese HCC patients (Hazard ratio = 0.597, 95% CI: 0.434–0.820, P = 0.001). Multivariate Cox regression analysis identified methylation level of cg01798157, cirrhosis, and microvascular invasion as independent prognostic factors. The prognostic efficiency of death risk score was superior to that of cirrhosis or microvascular invasion alone. ConclusionsThe methylation level of cg01798157 in BTG2 may be an epigenetic biomarker in Chinese patients with resectable HCC.

Platelet microparticles-derived miR-25-3p promotes the hepatocyte proliferation and cell autophagy via reducing B-cell translocation gene 2.

Platelets are critical regulators of liver regeneration, but the mechanisms are still not fully understood. Platelets have been shown to contain a wide variety of microRNAs (miRNAs) and play an important role in many diseases. However, the mechanism that how the platelet microparticles (PMPs)-derived miRNA regulate the hepatocyte proliferation is not very clear. In this study, we have successfully isolated and identified PMPs. We also found that PMPs, which could be well integrated into the HHL-5 cells, could upregulate the level of miR-25-3p in HHL-5 cells. Meanwhile, we found that PMPs-derived miR-25-3p promoted HHL-5 cells proliferation by accelerating cells into the S phase, and enhanced the autophagy by increasing the LC3II expression and reducing the P62 expression. Then, we proved that the miR-25-3p could target the B-cell translocation gene 2 (BTG2) and downregulate the expression levels of the BTG2 gene in HHL-5 cells. In addition, the overexpression of BTG2 significantly inhibited the proliferation and autophagy abilities of HHL-5 cells, while cotransfected miR-25-3p mimics or PMPs could partially rescue HHL-5 cells proliferation and autophagy. Furthermore, we proved that PMPs accelerated hepatocyte proliferation by regulating autophagy pathways. Therefore, PMPs-derived miR-25-3p promoted HHL-5 cell proliferation and autophagy by targeting BTG2, which may be a new therapeutic method for liver regeneration.

NUSAP1 knockdown inhibits cell growth and metastasis of non-small-cell lung cancer through regulating BTG2/PI3K/Akt signaling.

Non-small-cell lung cancer (NSCLC) is the most common malignancy along with high mortality rate worldwide. Recently, nucleolar and spindle-associated protein 1 (NUSAP1) has been reported to be involved in the malignant progression of several cancers. However, in NSCLC, the biological function of NUSAP1 and its molecular mechanism have not been reported. Here, our findings indicated that the NUSAP1 messenger RNA expression level was remarkably upregulated in NSCLC tissues compared with that of adjacent normal tissues. We also found that NUSAP1 gene expression was notably upregulated in NSCLC cell lines (A549, 95-D, H358, and H1299) compared with that of normal human bronchial epithelial cell line (16HBE). Subsequently, the biological function of NUSAP1 was investigated in A549 and H358 cells transfected with NUSAP1 small interfering RNA (siRNA), respectively. Results showed that NUSAP1 knockdown inhibited NSCLC cell proliferation, and promoted cell apoptosis. Furthermore, the number of cell migration and invasion was significantly suppressed by NUSAP1 knockdown. In addition, our results indicated that NUSAP1 knockdown increased the gene expression of B-cell translocation gene 2 (BTG2), but decreased the expression levels of phosphoinositide 3-kinase (PI3K) and phosphorylated serine/threonine kinase (p-AKT). BTG2 siRNA partly abrogates the effect of NUSAP1 knockdown on BTG2 gene expression. Fumonisin B1 (FB1), a AKT activator, reversed the effect of NUSAP1 knockdown on the biological function in NSCLC. Taken together, NUSAP1 knockdown promotes NSCLC cell apoptosis, and inhibits cell proliferation, cell migration, and invasion, which is associated with regulating BTG2/PI3K/Akt signal pathway. Our findings suggest that NUSAP1 is a promising molecular target for NSCLC treatment.

BTG2 is a tumor suppressor gene upregulated by p53 and PTEN in human bladder carcinoma cells.

Although widely deemed as a tumor suppressor gene, the role of B‐cell translocation gene 2 (BTG2) in bladder cancer is still inconclusive. We investigated the role and regulatory mechanism of BTG2 in bladder cancer. BTG2 expression in human bladder tissues was determined by RT‐qPCR and immunoblotting assays. Expressions of BTG2 and PTEN in bladder carcinoma cells were determined by immunoblotting, RT‐qPCR, or reporter assays. The 3H‐thymidine incorporation assay, flow cytometry, and the xenograft animal model were used to determine the cell growth. BTG2 expression was lower in human bladder cancer tissues than normal bladder tissues. Highly differentiated bladder cancer cells, RT4, expressed higher BTG2 than the less‐differentiated bladder cancer cells, HT1376 and T24. Overexpression of BTG2 in T24 cells inhibited cell growth in vitro and in vivo. Camptothecin and doxorubicin treatments in RT‐4 cells or transient overexpression of p53 into p53‐mutant HT1376 cells induced p53 and BTG2 expression. Further reporter assays with site‐mutation of p53 response element from GGGAAAGTCC to GGAGTCC within BTG2 promoter area showed that p53‐induced BTG2 gene expression was dependent on the p53 response element. Ectopic PTEN overexpression in T24 cells blocked the Akt signal pathway which attenuated cell growth via upregualtion of BTG2 gene expression, while reverse effect was found in PTEN‐knockdown RT‐4 cells. PTEN activity inhibitor (VO‐OHpic) treatment decreased BTG2 expression in RT‐4 and PTEN‐overexpressed T24 cells. Our results suggested that BTG2 functioned as a bladder cancer tumor suppressor gene, and was induced by p53 and PTEN. Modulation of BTG2 expression seems a promising way to treat human bladder cancer.

BTG2 Is Down-Regulated and Inhibits Cancer Stem Cell-Like Features of Side Population Cells in Hepatocellular Carcinoma.

Our previous study found that B cell translocation gene 2 (BTG2) was hyper-methylated and down-regulated in side population (SP) cells of hepatocellular carcinoma (HCC) cell line. However, its clinical significances and biological impacts on HCC SP cells remained unclear. To investigate the prognostic value of BTG2 gene in HCC and its influences on cancer stem cells (CSCs)-like traits of HCC cell line SP cells. BTG2 expression in human HCC and adjacent non-cancerous tissues was detected by immunohistochemical staining and quantitative real-time PCR, and also obtained from GEO and TCGA data. Its prognostic values were assessed. Its biological influences on HCC cell line SP cells were evaluated using cell viability, cell cycle, plate clone-forming assay, and chemoresistance in vitro and tumorigenicity in vivo. BTG2 expression was significantly suppressed in human HCC compared to adjacent non-cancerous tissues. BTG2 expression was correlated with TNM stage, tumor size and vascular invasion. Lower expression of BTG2 was associated with poorer overall survival and disease-free survival. In vitro, overexpression of BTG2 substantially suppressed cell proliferation and accumulation of HCC cell line SP cells in G0/G1 phase. Colony formation ability was markedly suppressed by BTG2 overexpression. Moreover, sensitivity of HCC cell line SP cells to 5-fluorouracil was substantially increased by overexpression of BTG2. Furthermore, tumorigenicity of HCC cell line SP cells transfected with BTG2 plasmids was significantly reduced in vivo. BTG2 gene could regulate the CSC-like traits of HCC cell line SP cells, and it represented as a molecular prognostic marker for HCC.

Prostate-derived ets factor represses tumorigenesis and modulates epithelial-to-mesenchymal transition in bladder carcinoma cells

Prostate-derived Ets (E-twenty six) factor (PDEF), an epithelium-specific member of the Ets family of transcription factors, has been shown to play a role in suppressing the development of many epithelium-derived cancers such as prostate and breast cancer. It is not clear, however, whether PDEF is involved in the development or progression of bladder cancer. In a comparison between normal urothelium and bladder tumor tissue, we identified significant decreases of PDEF in the tumor tissue. Further, the immunohistochemistry assays indicated a significantly higher immunostaining of PDEF in low-grade bladder tumors. Additionally, the highly differentiated transitional-cell bladder carcinoma RT-4 cells expressed significantly more PDEF levels than the bladder carcinoma HT1376 and the T24 cells. Ectopic overexpression of PDEF attenuated proliferation, invasion, and tumorigenesis of bladder carcinoma cells in vitro and in vivo. PDEF enhanced the expression levels of mammary serine protease inhibitor (MASPIN), N-myc downstream regulated gene 1 (NDRG1), KAI1, and B-cell translocation gene 2 (BTG2). PDEF modulated epithelial-mesenchymal-transition (EMT) by upregulating E-cadherin expression and downregulating the expression of N-cadherin, SNAIL, SLUG, and vimentin, leading to lower migration and invasion abilities of bladder carcinoma cells. Filamentous actin (F-actin) polarization and remodeling were observed in PDEF-knockdown RT-4 cells. Our results suggest that PDEF gene expression is associated with the extent of bladder neoplasia and PDEF modulated the expressions of EMT-related genes. The induction of BTG2, NDRG1, MASPIN, and KAI1 gene expressions by PDEF may explain the inhibitory functions of PDEF on the proliferation, invasion, and tumorigenesis in bladder carcinoma cells.

MicroRNA‑21 regulates the expression of BTG2 in HepG2 liver cancer cells.

B-cell translocation gene 2 (BTG2) is a tumor suppressor gene, which belongs to the anti-proliferation gene family. Our previous study demonstrated that microRNA (miR)-21 and the expression of BTG2 were negatively correlated during hepatocarcinogenesis. The aim of the present study was to investigate the effects of miR-21 on the growth and progression of liver cancer cells, and to determine the underlying mechanism. A luciferase reporter assay was used to demonstrate that the BTG2 gene was a direct target of miR-21. In addition, the effects of miR-21 on cell growth and gene expression in HepG2 human hepatocellular carcinoma (HCC) cells were analyzed using reverse transcription-quantitative polymerase chain reaction, western blotting, an MTT assay, flow cytometry, a Transwell invasion assay and a wound healing assay. The expression levels of miR-21 in the HepG2 cells were significantly higher, compared with those in L02 normal liver cells. The expression levels of BTG2 in liver cancer cell lines (HepG2 and Huh7) were significantly lower, compared with that in the L02 cells. These results suggested that BTG2 was the direct target gene of miR-21. The protein expression levels of BTG2 were inhibited by high expression levels of miR-21, and increased by inhibition of the expression of miR-21 in the HepG2 cells. Inhibition of miR-21 reduced cell proliferation and invasion, and increased the rate of apoptosis in the HepG2 cells. These results indicated that miR-21 regulates cell proliferation, invasion, migration and apoptosis in HepG2 cells, which may be associated with its effects on the expression of BTG2. The results of the present study may provide a basis for targeting the miR-21/BTG2 interaction for the treatment of HCC.

[Enhancement of B-cell translocation gene-2 inhibits proliferation and metastasis of colon cancer cells].

To inhibit the proliferation and metastasis of colon cancer cells by increasing the expression level of B-cell translocation gene-2 (BTG2). Western blot assay was used to detect the expression level of BTG2 protein in the normal intestinal epithelial HIEC cells and three colon cancer cell lines SW620, HT-29 and LS174T. The expression of BTG2 protein in normal colonic epithelial tissue, colon adenoma and colon cancer tissue was detected by immunohistochemistry. The plasmid with BTG2 gene full-length sequence was transfected into colon cancer SW620 cells, and the expression of BTG2 protein was detected by Western blot. The cell growth curve was drawn by MTT test. The Ki-67-positive rate was calculated using immunofluorescence staining. The cell migration of colon cancer cells was detected by scratch test and Transwell double chamber culture system, and the pseudopodia growth of tumor cells was detected by Matrigel 3D culture system. Western blot results showed that BTG2 relative expression levels were 0.83 ± 0.12, 0.18 ± 0.04, 0.20 ± 0.05 and 0.36 ± 0.07 in normal human intestinal epithelial cells HIEC, and human colon cancer cell line SW620, HT-29 and LS174T, respectively. The results of immunohistochemistry showed that the positive expression of BTG2 protein in normal colorectal tissue, colorectal adenoma and colorectal carcinoma tissues were 82.5% (33/40), 77.5%(31/40) and 17.5% (7/40), respectively, with a significant difference between two groups (P < 0.05). Immunofluorescence results showed that the positive rate of Ki-67 in the control group, empty vector group and BTG2 transfection group was (76.2 ± 8.0)%, (81.4 ± 9.7)% and (50.1 ± 7.1)%, respectively, showing a significant difference between two groups (P < 0.05). The scratch test results showed that in the control group, empty vector group and BTG2 transfection group, the distance of SW620 cells between two sides was (79.27 ± 11.24) µm, (80.65 ± 12.17) µm and (124.77 ± 19.63) µm, respectively, with a significant difference between two groups (P < 0.05). Transwell results showed that in the control group, empty plasmid group and BTG2 transfection group, the SW620 cell migration rate was (78.5 ± 13.1)%, (73.2 ± 12.9)% and (47.4 ± 9.1)%, respectively, showing a significant difference between two groups (P < 0.05). The number of neurospheres of BTG2 transfection group was decreased SW620, which had poor ductility. BTG2 gene is involved in colon cancer cell proliferation and metastasis, and effectively restores the function of BTG2 protein. Therefore, it may be expected to become a new option in gene therapy for colon cancer.

B-cell translocation gene 2 promotes hepatic hepcidin production via induction of Yin Yang 1

Hepcidin is a peptide hormone secreted in the liver and plays a key role in maintaining iron homeostasis. Here, we demonstrate that B-cell translocation gene 2 (BTG2) is a key player in hepatic hepcidin regulation via induction of Yin Yang 1 (YY1). Hepatic hepcidin gene expression significantly enhanced by fasting states and glucagon exposure led to induction of gluconeogenic gene expression, and elevated serum hepcidin production in mice. Notably, overexpression of BTG2 using adenoviral system (Ad-BTG2) significantly elevated serum hepcidin levels via a significant induction of YY1 gene transcription. Immunoprecipitation studies demonstrated that BTG2 physically interacted with YY1 and recruited on the hepcidin gene promoter. Finally, ablation of hepatic BTG2 gene by gene silencing markedly attenuated the elevation of serum hepcidin production along with YY1 and hepcidin mRNA expression in fasting state. Likewise, forskolin (FSK)-stimulated hepcidin promoter activity was dramatically disrupted by endogenous BTG2 knockdown. Overall, our current study provides a novel molecular mechanism of BTG2-mediated induction of hepcidin gene expression, thereby contributing to a better understanding of the hepatic hepcidin production involved in iron homeostasis.

Upregulation of B-cell translocation gene 2 by epigallocatechin-3-gallate via p38 and ERK signaling blocks cell proliferation in human oral squamous cell carcinoma cells

Oral squamous cell carcinoma (OSCC) is a well-known malignancy that accounts for the majority of oral cancers. B-cell translocation gene 2 (BTG2) is an important regulator of cell cycle dynamics in cancer cells. However, the role of BTG2 in OSCC cells and the influences of epigallocatechin-3-gallate (EGCG) on BTG2 gene expressions have not been well evaluated. The objectives of this study were to examine the effect of EGCG-induced BTG2 expression and the potential signal pathways involved. The 3H-thymidine incorporation and Western-blot assays revealed cell proliferation was attenuated by EGCG via upregulation of BTG2 expression causing cell cycle G1 phase arrest in OSCC cells. BTG2 overexpression decreased tumor cell growth, while BTG2 knockdown illuminated the opposite effect in xenograft animal studies. Overexpressed BTG2 arrested the cell cycle at the G1 phase and downregulated protein expressions of cyclin A, cyclin D, and cyclin E. Western-blot assays indicated that EGCG induced phosphorylation of p38, JNK, and ERK. However, pretreatments with selective mitogen-activated protein kinase (MAPK) inhibitors, SB203580 (p38 inhibitor) and PD0325901 (ERK1/2 inhibitor), significantly suppressed the activation of EGCG on BTG2 expression. Our results indicate that EGCG attenuates cell proliferation of OSCC cells by upregulating BTG2 expression via p38 and ERK pathways.

BTG2: a rising star of tumor suppressors (review).

B-cell translocation gene 2 (BTG2), the first gene identified in the BTG/TOB gene family, is involved in many biological activities in cancer cells acting as a tumor suppressor. The BTG2 expression is downregulated in many human cancers. It is an instantaneous early response gene and plays important roles in cell differentiation, proliferation, DNA damage repair, and apoptosis in cancer cells. Moreover, BTG2 is regulated by many factors involving different signal pathways. However, the regulatory mechanism of BTG2 is largely unknown. Recently, the relationship between microRNAs and BTG2 has attracted much attention. MicroRNA-21 (miR-21) has been found to regulate BTG2 gene during carcinogenesis. In this review, we summarize the latest findings in the investigations of biological functions of BTG2 and regulation of its expression, with an emphasis on miR-21 in regulation of BTG2 gene in various cancers. B-cell translocation gene 2 (BTG2), also known as PC3 or TIS21, belongs to the antiproliferative (APRO) gene family. Several studies have demonstrated that BTG2 is involved in a large number of physiological and pathological processes, such as cell differentiation, proliferation, apoptosis, and other cellular functions, acting as a tumor suppressor. In this review, we summarize the latest findings in BTG2 studies, highlighting the mechanisms for the regulatory effects of microRNAs (miRNAs) on BTG2 gene expression in the most common human cancers.

Inhibition of bladder cancer invasion by Sp1-mediated BTG2 expression via inhibition of DNA methyltransferase 1.

Significantly lower endogenous expression of B-cell translocation gene 2 (BTG2) was observed in human muscle-invasive bladder cancers (MIBC) than matched normal tissues and non-muscle invasive bladder cancers (NMIBC). BTG2 expression was inversely correlated with increased expression of the DNA methyltransferases DNMT1 and DNMT3a in MIBC, but not NMIBC, suggesting a potential role for BTG2 expression in muscle invasion of bladder cancer. Over 90% of tumor tissues revealed strong methylation at CpG islands of the BTG2 gene, compared with no methylation in the normal tissues, implying epigenetic regulation of BTG2 expression in bladder carcinogenesis. By using EJ bladder cancer cells and the demethylating agent decitabine, transcription of BTG2 was shown to be up-regulated by inhibiting DNMT1 expression via modification at CpG islands. DNMT1 binding to the BTG2 gene further regulated BTG2 expression by chromatin remodeling, such as H3K9 dimethylation and H3K4 trimethylation, and Sp1 activation. Induced BTG2 expression significantly reduced EJ cell tumorigenesis and invasiveness together with induction of G2 /M arrest. These results demonstrate an important role for the BTG2(/TIS21/PC3) gene in the progression of bladder cancers, and suggest that BTG2(/TIS21/PC3) is a promising epigenetic target for prevention of muscle invasion in human bladder cancers.

Cisplatin modulates B-cell translocation gene 2 to attenuate cell proliferation of prostate carcinoma cells in both p53-dependent and p53-independent pathways.

Cisplatin is a widely used anti-cancer drug. The B-cell translocation gene 2 (BTG2) is involved in the cell cycle transition regulation. We evaluated the cisplatin effects on prostate cancer cell proliferation and the expressions of BTG2, p53, androgen receptor (AR) and prostate specific antigen (PSA) in prostate carcinoma, p53 wild-type LNCaP or p53-null PC-3, cells. Cisplatin treatments attenuated cell prostate cancer cell growth through inducing Go/G1 cell cycle arrest in lower concentration and apoptosis at higher dosage. Cisplatin treatments enhanced p53 and BTG2 expression, repressed AR and PSA expression, and blocked the activation of androgen on the PSA secretion in LNCaP cells. BTG2 knockdown in LNCaP cells attenuated cisplatin-mediated growth inhibition. Cisplatin enhanced BTG2 gene expression dependent on the DNA fragment located within -173 to -82 upstream of BTG2 translation initiation site in prostate cancer cells. Mutation of the p53 response element from GGGCAGAGCCC to GGGCACC or mutation of the NFκB response element from GGAAAGTCC to GGAAAGGAA by site-directed mutagenesis abolished the stimulation of cisplatin on the BTG2 promoter activity in LNCaP or PC-3 cells, respectively. Our results indicated that cisplatin attenuates prostate cancer cell proliferation partly mediated by upregulation of BTG2 through the p53-dependent pathway or p53-independent NFκB pathway.

B-Cell Translocation Gene 2 Regulates Hepatic Glucose Homeostasis via Induction of Orphan Nuclear Receptor Nur77 in Diabetic Mouse Model

B-cell translocation gene 2 (BTG2) is a member of an emerging gene family that is involved in cellular functions. In this study, we demonstrate that BTG2 regulates glucose homeostasis via upregulation of Nur77 in diabetic mice. Hepatic BTG2 gene expression was elevated by fasting and forskolin. Overexpression of Btg2 increased the expression of hepatic gluconeogenic genes and blood glucose output and subsequently impaired glucose and insulin tolerance. Upregulation of the transcriptional activity of Nur77, gluconeogenic genes, and glucose production by forskolin was observed by Btg2 transduction, but not in Btg2 knockdown. BTG2-stimulated glucose production and glucose-6-phosphatase promoter activity were attenuated by dominant-negative Nur77. Coimmunoprecipitation and chromatin immunoprecipitation assays showed that BTG2 induced Nur77 occupancy on the glucose-6-phosphatase promoter via a physical interaction. Btg2 gene expression was increased in streptozotocin-treated and db/db mice. Finally, impairment of glucose homeostasis, such as the increase of blood glucose, glucose intolerance, and insulin intolerance, was elevated in diabetic mice, whereas this phenomenon was abolished in knockdown of Btg2. Together, these data suggest that BTG2 participates in the regulation of hepatic glucose homeostasis, which means that BTG2 might serve as a potential therapeutic target for combating metabolic dysfunction.

Topoisomerase Inhibitors Modulate Gene Expression of B-Cell Translocation Gene 2 and Prostate Specific Antigen in Prostate Carcinoma Cells

Camptothecin (CPT) and doxorubicin (DOX) have been demonstrated to have potent anti-tumor activity. The B-cell translocation gene 2 (BTG2) is involved in the regulation of cell cycle progression. We evaluated the molecular mechanisms of CPT and DOX on cell proliferation and the expressions of BTG2 and prostate specific antigen (PSA) in prostate carcinoma cells. Our results indicated that CPT or DOX treatments induced Go/G1 cell cycle arrest in LNCaP cells and apoptosis at higher dosage. Immunoblot and transient gene expression assay indicated that CPT or DOX treatments induced p53 and BTG2 gene expression, with the later effect dependent on the p53 response element within BTG2 promoter area since mutation of the p53 response element from GGGAAAGTCC to GGAGTCC or from GGCAGAGCCC to GGCACC by site-directed mutagenesis abolished the stimulation of CPT or DOX on the BTG2 promoter activity, which is also supported by our results that cotreatments of pifithrin-α, an inhibitor of p53 dependent transcriptional activation, blocked the induction of CPT or DOX on BTG2 gene expression. CPT or DOX also downregulated the protein expressions of androgen receptor (AR) and PSA. Transient gene expression assays suggested that CPT or DOX’s attenuation of PSA promoter activity is dependent on both the androgen and p53 response elements within of the PSA promoter. Our results indicated that CPT and DOX attenuate cell proliferation via upregulation of BTG2 gene expression through the p53-dependent pathway. The CPT and DOX block the PSA gene expression by upregulation of p53 activity and downregulation of androgen receptor activity.

Expression and significance of miRNA-21 and BTG2 in lung cancer

This study investigates the expression of micro-ribonucleic acid-21 (miRNA-21) and B cell translocation gene 2 (BTG2) in lung cancer cells. We examined the impact of miRNA-21 on biological characteristics of lung cancer cells, such as growth, proliferation, apoptosis, and invasion. The expression of miRNA-21 and BTG2 protein in lung cancer cell lines (A549, HCC827, NCI-H292, and 95-D) was examined using quantitative reverse transcription-polymerase chain reaction and Western blot analysis, respectively. Subsequently, the regulatory role of miRNA-21 on BTG2 was explored by inhibiting miRNA-21 expression in 95-D cells using miRNA-21-antisense oligonucleotides (miRNA-21 ASO). The impact of miRNA-21 on the biological characteristics of 95-D cells was further studied using methylthiazol tetrazolium assays, flow cytometry, and Transwell invasion chamber assays. The impact of miRNA-21 on the expression of cyclin D1, caspase-3, and matrix metalloprotease-9 (MMP9) was also studied. miRNA-21 expression was significantly higher in lung cancer cell lines (A549, HCC827, NCI-H282, and 95-D) than that in normal human bronchial epithelial cells (HBE; p < 0.05). The pattern of BTG2 protein expression was exactly the opposite of miRNA-21 expression in lung cancer cells. BTG2 was highly expressed in HBE cells and was expressed at very low levels in lung cancer cell lines (A549, HCC827, NCI-H292, and 95-D). High miRNA-21 expression may inhibit BTG2 protein expression, whereas the inhibition of miRNA-21 expression may promote BTG2 protein expression in 95-D cells. Cell viability and invasion of 95-D cells were significantly lower in the miRNA-21 ASO-transfected group than that in the control ASO-transfected group and untransfected group (p < 0.05). The number of apoptotic cells was significantly higher in the miRNA-21 ASO-transfected group than that in the control ASO-transfected and untransfected groups (p < 0.05). The expression level of cyclin D1 and MMP9 in 95-D cells was significantly lower in the miRNA-21 ASO-transfected group than in the control ASO-transfected and untransfected groups (p < 0.05). Meanwhile, caspase-3 expression was significantly higher in the miRNA-21 ASO-transfected group than that in the control ASO-transfected and untransfected groups (p < 0.05). miRNA-21 overexpression may inhibit the BTG2 gene in lung cancer cells. miRNA-21 may promote cell proliferation and invasion and inhibit cell apoptosis in 95-D cells.

B-cell translocation gene 2 positively regulates GLP-1-stimulated insulin secretion via induction of PDX-1 in pancreatic β-cells

Glucagon-like peptide-1 (GLP-1) is a potent glucoincretin hormone and an important agent for the treatment of type 2 diabetes. Here we demonstrate that B-cell translocation gene 2 (BTG2) is a crucial regulator in GLP-1-induced insulin gene expression and insulin secretion via upregulation of pancreatic duodenal homeobox-1 (PDX-1) in pancreatic β-cells. GLP-1 treatment significantly increased BTG2, PDX-1 and insulin gene expression in pancreatic β-cells. Notably, adenovirus-mediated overexpression of BTG2 significantly elevated insulin secretion, as well as insulin and PDX-1 gene expression. Physical interaction studies showed that BTG2 is associated with increased PDX-1 occupancy on the insulin gene promoter via a direct interaction with PDX-1. Exendin-4 (Ex-4), a GLP-1 agonist, and GLP-1 in pancreatic β-cells increased insulin secretion through the BTG2–PDX-1–insulin pathway, which was blocked by endogenous BTG2 knockdown using a BTG2 small interfering RNA knockdown system. Finally, we revealed that Ex-4 and GLP-1 significantly elevated insulin secretion via upregulation of the BTG2–PDX-1 axis in pancreatic islets, and this phenomenon was abolished by endogenous BTG2 knockdown. Collectively, our current study provides a novel molecular mechanism by which GLP-1 positively regulates insulin gene expression via BTG2, suggesting that BTG2 has a key function in insulin secretion in pancreatic β-cells.

Upregulation of prostate‐derived Ets factor by luteolin causes inhibition of cell proliferation and cell invasion in prostate carcinoma cells

Luteolin is a polyphenolic flavone and has antitumor activity for many cancers. The prostate-derived Ets factor (PDEF), a novel epithelium-specific Ets transcription factor, acts as an androgen-independent transcriptional activator of the prostate-specific antigen (PSA) promoter. We determined the antitumor function of luteolin via upregulation of PDEF gene expression in human prostate carcinoma LNCaP cells. Results from flow cytometry and (3) H-thymidine incorporation assays revealed that luteolin treatments attenuated cell proliferation and arrested the cell cycle at the G1/S phase. High concentration of luteolin (30 μM) induced cell apoptosis. Immunoblot assays and enzyme linked immunosorbent assay revealed that luteolin treatment upregulated PDEF but downregulated androgen receptor (AR) gene expression, which decreased PSA gene expression in LNCaP cells. Results of immunoblot and transient gene expression assays revealed that luteolin treatments at proapoptosis dosage, enhanced gene expression of PDEF, B-cell translocation gene 2 (BTG2), N-myc downstream regulated gene 1 (NDRG1) and Maspin. Transient gene expression assays indicated that cotransfection of the PDEF expression vector enhanced the promoter activities of the BTG2, NDRG1 and Maspin genes. Stable overexpression of PDEF significantly induced BTG2, NDRG1 and Maspin gene expression, which markedly attenuated in vitro cell proliferation and invasion of LNCaP cells. The modulatory effect of luteolin on BTG2, NDRG1 and Maspin gene expression were attenuated when PDEF was knocked-down. These results suggest that luteolin blocks PSA gene expression by downregulation of AR expression. The enhancement of PDEF expression, which induced BTG2, NDRG1 and Maspin gene expression, could account for the function of luteolin for antiproliferation and anti-invasion in LNCaP cells.

Molecular characterization, expression pattern and association analysis of the porcine BTG2 gene

B-cell translocation gene 2 (BTG2), a member of the B-cell translocation gene family with anti-proliferative properties, have been characterized to be involved in cell growth, differentiation and survival. In this study, we cloned the full length sequences of cDNA and genomic DNA of BTG2 gene from the porcine skeletal muscle. Spatial expression analysis showed that the porcine BTG2 gene is expressed predominantly in muscle. Temporal expression analysis in longissimus dorsi muscle demonstrated that the expression of BTG2 gene has the highest expression at 60 days old in Large White while with a peak expression at 120 days old in Meishan. Temporal analysis also revealed that the expression of BTG2 gene is generally higher in Large White than in Meishan at all the developmental stages tested (65 days of conception and 3, 35, 60, 120, and 180 days of postnatal). A single nucleotide polymorphism (G417C) in the intron of BTG2 gene was then detected by PCR-RFLP in Large White × Meishan F2 resource population and association analysis suggested that this polymorphic site had significant association (P < 0.05) with the buttock fat thickness, fat percentage, lean muscle percentage, ratio of lean to fat and carcass length.

Impact of BTG2 expression on proliferation and invasion of gastric cancer cells in vitro

BTG2 (B cell translocation gene 2) is downregulated in several human tumors and has been known as a tumor suppressor in carcinogenesis of thymus, prostate, kidney, and liver. However, little is known about the role BTG2 plays in gastric adenocarcinoma. In the present study, we intended to investigate the influence of BTG2 on the growth, proliferation, apoptosis, invasion and cell cycle of the gastric cancer cell lines SGC7901 and MKN45. BTG2 cDNA was insected into a constitutive vector pcDNA3.1 followed by transfection in gastric cancer cell line MKN45 and SGC7901 by using liposome. Then stable transfectants were selected and appraised. The apoptosis and cell cycles of these transfectants were analyzed by using flow cytometric assay. The growth and proliferation were analyzed by cell growth curves and colony-forming assay, respectively. The invasion of these clones was analyzed by using cell migration assay. MKN-BTG2 (MKN45 with stable transfection of BTG2 gene) and SGC-BTG2 (SGC7901 with stable transfection of BTG2 gene) grew slower than their control groups, respectively. The cell counts of MKN-BTG2 in the fourth, fifth, sixth and seventh days were significantly fewer than those of control groups (P < 0.05). Those of SGC-BTG2 in the fourth fifth, sixth and seventh days were significantly fewer than those of control groups too (P < 0.05). Cell cycle analysis showed that proportions of MKN-BTG2 and SGC-BTG2 cells in G0-G1 and S were different significantly with those of their control groups, respectively (P < 0.05). The apoptosis rate of MKN-BTG2 was significantly higher than those of control groups (P < 0.05). Results of colony-forming assay showed that the colon formation rates of MKN-BTG2 and SGC-BTG2 were lower than those of their control groups (P < 0.05). The results of cell migration assay showed that the cell migration rates of MKN-BTG2 and SGC-BTG2 were not significantly different with those of their control groups (P > 0.05). BTG2 can restrain the growth and proliferation of gastric cancer cells powerfully. It can reduce some malignant phenotype of these tumor cells. But it could not impact the ability of invasion of gastric cancer cells, so could not restrain the metastasis of gastric cancer. In gastric cancer, BTG2 could be thought as a tumor-inhibiting gene in some distance, so the gene could be a potential target of gene therapy.