Integrating GC-MS and comparative transcriptome analysis reveals that TsERF66 promotes the biosynthesis of caryophyllene in Toona sinensis tender leaves.

Introduction: The strong aromatic characteristics of the tender leaves of Toona sinensis determine their quality and economic value. Methods and results: Here, GC-MS analysis revealed that caryophyllene is a key volatile compound in the tender leaves of two different T. sinensis varieties, however, the transcriptional mechanisms controlling its gene expression are unknown. Comparative transcriptome analysis revealed significant enrichment of terpenoid synthesis pathway genes, suggesting that the regulation of terpenoid synthesis-related gene expression is an important factor leading to differences in aroma between the two varieties. Further analysis of expression levels and genetic evolution revealed that TsTPS18 is a caryophyllene synthase, which was confirmed by transient overexpression in T. sinensis and Nicotiana benthamiana leaves. Furthermore, we screened an AP2/ERF transcriptional factor ERF-IX member, TsERF66, for the potential regulation of caryophyllene synthesis. The TsERF66 had a similar expression trend to that of TsTPS18 and was highly expressed in high-aroma varieties and tender leaves. Exogenous spraying of MeJA also induced the expression of TsERF66 and TsTPS18 and promoted the biosynthesis of caryophyllene. Transient overexpression of TsERF66 in T. sinensis significantly promoted TsTPS18 expression and caryophyllene biosynthesis. Discussion: Our results showed that TsERF66 promoted the expression of TsTPS18 and the biosynthesis of caryophyllene in T. sinensis leaves, providing a strategy for improving the aroma of tender leaves.

Synthesis, DNA binding of bis-naphthyl ferrocene derivatives and the application as new electroactive indicators for DNA biosensor.

A series of bis-naphthyl ferrocene derivatives were synthesized and characterized. Based on the results obtained from UV-visible absorption titration and ethidium bromide (EB) displacement experiments, it was observed that the synthesized compounds exhibited a strong binding ability to dsDNA. In comparison to the viscosity curve of EB, the tested compounds demonstrated a bisintercalation binding mode when interacting with CT-DNA. Differential pulse voltammetry (DPV) was employed to assess the binding specificity of these indicators towards ssDNA and dsDNA. All tested indicators displayed more pronounced signal differences before and after hybridization between probe nucleic acids and target nucleic acids compared to Methylene Blue (MB). Among the evaluated compounds, compound 3j containing an ether chain showed superior performance as an indicator, making it suitable for constructing DNA-based biosensors. Under optimized conditions including probe ssDNA concentration and indicator concentration, this biosensor exhibited good sensitivity, reproducibility, stability, and selectivity. The limit of detection was calculated as 4.53 × 10-11 mol/L. Furthermore, when utilizing 3j as the indicator in serum samples, the biosensor achieved satisfactory recovery rates for detecting the BRCA1 gene.

PLCε promotes the Warburg effect and tumorigenesis through AKT/GSK3ß/Cdc25a in bladder cancer.

Phospholipase C epsilon (PLCε) is a oncogene in various malignancies and regulates diverse cellular functions. But understanding of the relation between PLCε and glycolytic pathways has not been clearly identified. In the present study, we explored the effect of PLCε on the Warburg effect and tumorigenesis in bladder cancer (BCa). In our study, we showed that PLCε expression was elevated in BCa samples compared with matched adjacent nonmalignant bladder tissues. PLCε depletion using Lentivirus-shPLCε (LV-shPLCε) dramatically decreased cell growth, glucose consumption and lactate production, arresting T24 and BIU cells in the S phase of the cell cycle. We also observed that PLCε was correlated with the activation of protein kinase B (AKT) and cell division cycle 25 homolog A (Cdc25a) overexpression. In addition, we demonstrated that AKT/glycogen synthase kinase 3 beta (GSK3ß)/Cdc25a signaling pathways are involved in the PLCε-mediated Warburg effect in BCa. Moreover, we showed that PLCε had an effect on tumorigenesis in in vivo experiments. In summary, our findings demonstrate that AKT/GSK3ß/Cdc25a is critical for the effect PLCε on Warburg effect and tumorigenesis.

Discovery of protein biomarkers for venous thromboembolism in non-small cell lung cancer patients through data-independent acquisition mass spectrometry.

Objective: Non-small cell lung cancer (NSCLC) patients present a high incidence of venous thromboembolism (VTE) with poor prognosis. It is crucial to identify and diagnose VTE early. The study aimed to identify potential protein biomarkers and mechanism of VTE in NSCLC patients via proteomics research. Methods: Proteomic analysis of the human plasma was performed through data-independent acquisition mass spectrometry for 20 NSCLC patients with VTE, and 15 NSCLC patients without VTE. Significantly differentially expressed proteins were analyzed by multiple bioinformatics method for further biomarker analysis. Results: A total of 280 differentially expressed proteins were identified in VTE and non-VTE patients, where 42 were upregulated and 238 were downregulated. These proteins were involved in acute-phase response, cytokine production, neutrophil migration and other biological processes related to VTE and inflammation. Five proteins including SAA1, S100A8, LBP, HP and LDHB had significant change between VTE and non-VTE patients, with the area under the curve (AUC) were 0.8067, 0.8308, 0.7767, 0.8021, 0.8533, respectively. Conclusions: SAA1, S100A8, LBP, HP and LDHB may serve as potential plasma biomarkers for diagnosis VTE in NSCLC patients.

STAT3 phosphorylation is required for the HepaCAM-mediated inhibition of castration-resistant prostate cancer cell viability and metastasis.

BACKGROUND: Castration-resistant prostate cancer (CRPC) is an advanced disease that is difficult to treat, the mechanism of it is unclear. This study illustrated the function of hepatocyte cell adhesion molecule (HepaCAM) on CRPC cell viability and metastasis. METHODS: The expression of HepaCAM and p-STAT3 in CRPC tissues were determined by immunohistochemistry and western blot analysis. Cell Counting Kit-8 and colony formation assays were deployed to analyze the growth ability of CRPC cells following the adenovirus-mediated re-expression of HepaCAM. CRPC cell migration and invasion capacity were investigated by wound healing and Matrigel-coated transwell assays, respectively. The messenger RNA or protein levels of p-STAT3, CyclinD1, cMyc, MMP2, MMP9, and VEGF were determined by reverse transcription (RT) followed by quantitative real-time polymerase chain reaction (RT-qPCR), and western blot analysis after either HepaCAM re-expression alone or in combination with IL-22 treatment. A CRPC orthotopic xenograft mouse model was applied to investigate the functional effect of HepaCAM on the metastasis of CRPC cells to the lungs. RESULTS: The expression levels of HepaCAM were decreased while those of p-STAT3 were elevated in CRPC cells compare with surrounding benign tissues (p < .001). The overexpression of HepaCAM in CRPC cells notably reduced proliferation, migration, and invasion by inhibiting the expression of p-STAT3, CyclinD1, cMyc, MMP2, MMP9, and VEGF (p < .05). In addition, the expression of HepaCAM significantly inhibited the IL-22/p-STAT3 axis and the metastasis of CRPC cells to the lungs. CONCLUSIONS: Our data suggested that HepaCAM suppressed the viability of CRPC cells via the IL-22/p-STAT3 axis and inhibited the metastasis of CRPC cells from the prostate to the lungs (p < .05).

Synthesis, electrochemistry, DNA binding and in vitro cytotoxic activity of tripodal ferrocenyl bis-naphthalimide derivatives.

A series of tripodal ferrocenyl bis-naphthalimide derivatives were synthesized and characterized. All of the bis-naphthalimide derivatives exhibited good DNA binding ability which was confirmed by ethidium bromide (EB) displacement experiment and ultraviolet (UV)-visible absorption titration. And the binding mode of these compounds was proved to be a hybrid binding mode by experiments. The cytotoxicity of synthesized compounds against 4 different human cancer cell lines (EC109, BGC823, SGC7901 and HEPG2) was evaluated by thiazolyl blue tetrazolium bromide (MTT) assay. All of the bis-naphthalimide derivatives exhibited good anticancer activity than the positive control drug (amonafide), which was due to the promotion of reactive oxygen species (ROS) level in test cancer cells by the reversible one-electron redox process of ferrocenyl bis-naphthalimide derivatives. Although there was no obvious relationship between the binding constants and the chain length, the structure cytotoxicity relationship revealed that the linker of n = 3, m = 1 was the best choice for the tested tripodol bis-naphthalimide derivatives. SYNOPSIS: A series of tripodal ferrocenyl bis-naphthalimide derivatives were synthesized to study the DNA binding ability and the cytotoxicity induced by reactive oxygen species. All of the compounds exhibited good DNA binding ability. And the structure cytotoxicity relationship revealed that the structure of 5h was the best choice.

CircHIPK3 Facilitates the G2/M Transition in Prostate Cancer Cells by Sponging miR-338-3p.

BACKGROUND: Circular RNAs (circRNAs) play a crucial role in gene expression regulation. CircHIPK3 is a circRNA derived from Exon 2 of HIPK3 gene and its role in prostate cancer (PCa) is still unclear. METHODS: CCK8 assays, flow cytometry and colony formation assays were performed to assess the effects of circHIPK3 in PCa cells. Bioinformatics analysis, RNA pull-down assay, RNA immunoprecipitation assay (RIP), and luciferase activity assay were performed to dissect the mechanism underlying circHIPK3-mediated G2/M transition in PCa cells. RESULTS: CircHIPK3 expression was upregulated in PCa cells and prostate cancer tissues. Overexpression of circHIPK3 or circHIPK3 silencing altered PCa viability, proliferation and apoptosis in vitro. CircHIPK3 could sponge miR-338-3p and inhibit its activity, resulting in increased expression of Cdc25B and Cdc2 in vitro. CONCLUSION: CircHIPK3 promotes G2/M transition and induces PCa cell proliferation by sponging miR-338-3p and increasing the expression of Cdc25B and Cdc2. CircHIPK3 may play an oncogenic role in PCa.

PLCε regulates metabolism and metastasis signaling via HIF-1α/MEK/ERK pathway in prostate cancer.

Phospholipase C-ε (PLCε) is frequently overexpressed in tumors and plays an important role in the regulation of tumorigenesis. Although great progress has been made in understanding biological roles of PLCε, the relevant molecular mechanisms underlying its pro-tumor activity remain largely unclear. Here, we demonstrated that PLCε knockdown reduced cell metastasis, glucose consumption and lactate production in a manner that depended on hypoxia inducible factor 1α (HIF-1α) expression in prostate cancer cells. Interestingly, our findings showed that the expression levels of PLCε were positively associated with those of HIF-1α in clinical prostate carcinoma samples. Knockdown of PLCε impaired HIF-1α levels and transcriptional activity by regulating the extracellular-signal-regulated kinase pathway, and blocking HIF-1α nuclear translocation. Furthermore, PLCε could interact with the von Hippel-Lindau E3 ligase complex to modulate the stability of HIF-1α. Collectively, our findings demonstrate that PLCε could be a crucial positive regulator of HIF-1α, which would promote PLCε-enhanced tumorigenesis.

PLCε regulates prostate cancer mitochondrial oxidative metabolism and migration via upregulation of Twist1.

BACKGROUND: Metabolic rewiring is a common feature of many cancer types, including prostate cancer (PCa). Alterations in master genes lead to mitochondrial metabolic rewiring and provide an appealing target to inhibit cancer progression and improve survival. Phospholipase C (PLC)ε is a regulator of tumor generation and progression. However, its role in mitochondrial metabolism remains unclear. METHODS: The GEO, The Cancer Genome Atlas, and the GTEx databases were used to determine Twist1 mRNA levels in tumors and their non-tumor counterparts. Fifty-five PCa and 48 benign prostatic hypertrophy tissue samples were tested for the presence of PLCε and Twist1 immunohistochemically. An association between PLCε and Twist1 was determined by Pearson's correlation analysis. PLCε was knocked down with a lentiviral short hairpin RNA. Mitochondrial activity was assessed by measuring the oxygen consumption rate. Western blotting analyses were used to measure levels of PPARß, Twist1, phosphorylated (p)-Twist1, p-MEK, p-ERK, p-P38, and p-c-Jun N-terminal kinase (JNK). Cells were treated with inhibitors of MEK, JNK, and P38 MAPK, and an agonist and inhibitor of peroxisome proliferator activated receptor (PPAR) ß, to evaluate which signaling pathways were involved in PLCε-mediated Twist1 expression. The stability of Twist1 was determined after blocking protein synthesis with cycloheximide. Reporter assays utilized E-cadherin or N-cadherin luciferase reporters under depletion of PLCε or Twist1. Transwell assays assessed cell migration. Finally, a nude mouse tumor xenograft assay was conducted to verify the role of PLCε in tumor formation. RESULTS: Our findings revealed that the expression of PLCε was positively associated with Twist1 in clinical PCa samples. PLCε knockdown promoted mitochondrial oxidative metabolism in PCa cells. Mechanistically, PLCε increased phosphorylation of Twist1 and stabilized the Twist1 protein through MAPK signaling. The transcriptional activity of Twist1, and the Twist1-mediated epithelial-to-mesenchymal transition, cell migration, and transcription regulation, were suppressed by PLCε knockdown and by blocking PPARß nuclear translocation. The tumor xenograft assay demonstrated that PLCε depletion diminished PCa cell tumorigenesis. CONCLUSIONS: These findings reveal an undiscovered physiological role for PLCε in the suppression of mitochondrial oxidative metabolism that has significant implications for understanding PCa occurrence and migration.

Simvastatin delays castration­resistant prostate cancer metastasis and androgen receptor antagonist resistance by regulating the expression of caveolin­1.

The failure of androgen deprivation therapy in prostate cancer treatment mainly results from drug resistance to androgen receptor antagonists. Although an aberrant caveolin­1 (Cav­1) expression has been reported in multiple tumor cell lines, it is unknown whether it is responsible for the progression of castration­resistant prostate cancer (CRPC). Thus, the aim of the present study was to determine whether Cav­1 can be used as a key molecule for the prevention and treatment of CRPC, and to explore its mechanism of action in CRPC. For this purpose, tissue and serum samples from patients with primary prostate cancer and CRPC were analyzed using immunohistochemistry and enzyme­linked immunosorbent assay, which revealed that Cav­1 was overexpressed in CRPC. Furthermore, Kaplan­Meier survival analysis and univariate Cox proportional hazards regression analysis demonstrated that Cav­1 expression in tumors was an independent risk factor for the occurrence of CRPC and was associated with a shorter recurrence­free survival time in patients with CRPC. Receiver operating characteristic curves suggested that serum Cav­1 could be used as a diagnostic biomarker for CRPC (area under the curve, 0.876) using a cut­off value of 0.68 ng/ml (with a sensitivity of 82.1% and specificity of 80%). In addition, it was determined that Cav­1 induced the invasion and migration of CRPC cells by the activation of the H­Ras/phosphoinositide­specific phospholipase Cε signaling cascade in the cell membrane caveolae. Importantly, simvastatin was able to augment the anticancer effects of androgen receptor antagonists by downregulating the expression of Cav­1. Collectively, the findings of this study provide evidence that Cav­1 is a promising predictive biomarker for CRPC and that lowering cholesterol levels with simvastatin or interfering with the expression of Cav­1 may prove to be a useful strategy with which to prevent and/or treat CRPC.

PLCε knockdown overcomes drug resistance to androgen receptor antagonist in castration-resistant prostate cancer by suppressing the wnt3a/ß-catenin pathway.

Most prostate cancers (Pcas) develop into castration-resistant prostate cancer (CRPC) after receiving androgen deprivation therapy (ADT). The expression levels of PLCε and wnt3a are increased in Pca and regulate androgen receptor (AR) activity. However, the biological function and mechanisms of PLCε and wnt3a in CRPC remain unknown. In this study, we found that the expression levels of PLCε, wnt3a, and AR were significantly increased in CRPC tissues as well as bicalutamide-resistant-LNCaP and enzalutamide-resistant-LNCaP cells. In addition, PLCε knockdown partly restored the sensitivity of drug-resistant cells to bicalutamide and enzalutamide by inhibiting the activity of the wnt3a/ß-catenin/AR signaling axis. Interestingly, the resistance of LNCaP cells docetaxel is related to PLCε but not the wnt3a/ß-catenin pathway. We also found that the combination of PLCε knockdown and enzalutamide treatment synergistically suppressed cell proliferation, tumor growth, and bone metastasis using in vitro and in vivo experiments. Our study revealed that PLCε is involved in the progression of drug-resistance in CRPC and could be a new target for the treatment of CRPC.

HepaCAM Regulates Warburg Effect of Renal Cell Carcinoma via HIF-1α/NF-κB Signaling Pathway.

OBJECTIVE: To investigate how hepatocyte cell adhesion molecule (hepaCAM) regulates cancer energy metabolism through hypoxia-inducible factor (HIF-1α) in renal cell carcinoma (RCC). MATERIALS AND METHODS: The expression of hepaCAM and HIF-1α in RCC tissue samples was examined by immunohistochemistry. Glucose consumption and lactate production assays were used to detect metabolic activity in RCC cell lines. P65 and IκB kinase (IKKß) mRNA and protein expression were detected using quantitative real-time polymerase chain reaction and western blotting, respectively. Nuclear translocation of P65 was observed by immunofluorescence staining after re-expressing hepaCAM. The luciferase reporter assay was applied to validate the transcriptional activity of HIF-1α. RESULTS: HIF-1α expression was elevated and hepaCAM suppressed in RCC compared with adjacent normal tissues. Furthermore, hepaCAM re-expression significantly decreased glycolytic metabolism in RCC cell lines, and reduced HIF-1α, IKKß, and P65 expression. The expression of HIF-1α, GLUT1, LDHA, and PKM2 were further reduced with combined hepaCAM overexpression and treatment with the NF-κB inhibitor BAY11-7082, compared to hepaCAM overexpression alone. Additionally, hepaCAM decreased the transcriptional activity of HIF-1α and blocked P65 nuclear translocation by the NF-κB pathway. CONCLUSION: Our data suggest that hepaCAM suppresses the Warburg effect via the HIF-1α/NF-κB pathway in RCC, which is a facilitating factor in hepaCAM-reduced tumorigenesis.

Synthesis, DNA Binding, and Anticancer Properties of Bis-Naphthalimide Derivatives with Lysine-Modified Polyamine Linkers.

A series of bis-naphthalimide derivatives with different diamine linkers were designed and synthesized. All of the synthesized bis-naphthalimide derivatives were characterized by NMR and HRMS spectra. The binding ability between the compounds and CT DNA was evaluated by using UV-Vis titration experiments. The bis-naphthalimide compound with an ethylenediamine linker showed the largest binding constant with CT DNA. Hence, it was used as the model compound to study the DNA binding selectivity by UV-Vis titration aiming at different DNA duplexes. As a result, this compound showed binding preference to AT-rich duplexes. The DNA binding modes of the compounds were also measured by viscosity titration. The cytotoxicity of the compounds was evaluated by MTT assay. Compounds with 1,6-diaminohexane or 1,4-phenylenedimethanamine linkers showed higher cytotoxicity compared with other bis-naphthalimide derivatives.

Knockdown of Phospholipase Cε (PLCε) Inhibits Cell Proliferation via Phosphatase and Tensin Homolog Deleted on Chromosome 10 (PTEN)/AKT Signaling Pathway in Human Prostate Cancer.

BACKGROUND Phospholipase Cε (PLCε), a member of the plc family, has been extensively studied to reveal its role in the regulation of different cell functions, but understanding of the underlying mechanisms remains limited. In the present study, we explored the effects of PLCε on PTEN (phosphatase and tensin homolog deleted on chromosome 10) in cell proliferation in prostate cancer cells. MATERIAL AND METHODS We assessed PLCε and PTEN expression in human benign prostate tissues compared to prostate cancer tissues by immunohistochemistry. Lentivirus-shPLCε (LV-shPLCε) was designed to silence PLCε expression in DU145 and PC3 cell lines, and the effectiveness was tested by qRT-PCR and Western blotting. MTT assay and colony formation assay were conducted to observe cell proliferation. Western blotting and immunofluorescence assays were used to detect changed PTEN expression in DU145. RESULTS We observed that PLCε expression was reduced in human benign prostate tissues compared to prostate cancer tissues, while PTEN expression showed the opposite trend. Silencing of the PLCε gene significantly inhibited cell proliferation in DU145 and PC3 cell lines. DU145 is a PTEN-expressing cell, while PC3 is PTEN-deficient. After infection by LV-shPLCε, we noticed that PTEN expression was up-regulated in DU145 cells but not in PC3 cells. Furthermore, we found that PLCε gene knockdown decreased P-AKT protein levels, but AKT protein levels were not affected. Immunofluorescence assays showed that PTEN expression had an intracellular distribution change in the DU145 cell line, and Western blot analysis showed that PTEN was obviously up-regulated in cell nucleus and cytoplasm. CONCLUSIONS PLCε is an oncogene, and knockdown of expression of PLCe inhibits PCa cells proliferation via the PTEN/AKT signaling pathway.

Co-Cluster-Based Metal-Organic Frameworks as Selective Catalysts for Benzene Tandem Acylation-Nazarov Cyclization to Benzocyclopentanone.

One-step selective benzene acylation-Nazarov cyclization is an attractive, yet challenging method for the synthesis of the benzocyclopentanone skeleton, which is key intermediate of many natural products. Herein, two metal-cluster-based metal-organic frameworks (MCOFs) {(H3 O)2 [Co4 (pbcd)2 (µ3 -OH)2 ]⋅CH3 CH2 OH⋅4 H2 O}n (1; pbcd=9,9'-(propan-1,3-diyl)bis(9H-carbazole-3,6-dicarboxylic acid) and {[Co5.25 (mcd)2 (HCO2 )(µ2 -O)0.5 (µ3 -OH)0.5 (H2 O)4 ]⋅6 H2 O⋅5 DMF}n (2; mcd=9,9'-methylenebis(9H-carbazole-3,6-dicarboxylic acid) were developed to catalyze a tandem Nazarov cyclization reaction of 1,3-dimethoxybenzene with α,ß-unsaturated carboxylic acids for the synthesis of cyclopentenone[b]benzenes. MCOFs 1 and 2, which were constructed from tetranuclear CoII cluster [Co4 (µ3 -OH)2 ] and hexanuclear CoII cluster [Co6 (HCO2 )(µ2 -O)(µ3 -OH)], respectively, exhibit high catalytic activity arising not only from their suitable channel size and accessible catalytic sites, but also because of their high density of Lewis acidic sites within the frameworks and the synergic activity among CoII ions. In contrast, {[Co2 (pbcd)(bpe)]⋅2 H2 O⋅2 DMF}n (3; bpe=1,2-bis(pyridin-4-yl)ethane) containing binuclear CoII and having large pore windows is a highly selective catalyst for obtaining exclusively the acylation products. Easy product separation, simple reaction procedures, and catalyst recycling make the catalyst system attractive. This work highlights the synergistic effect among ions of MCOFs in interacting with substrates in a sequential or cooperative manner to achieve tandem catalysis.

Expression of hepaCAM inhibits bladder cancer cell proliferation via a Wnt/ß-catenin-dependent pathway in vitro and in vivo.

We previously established that hepatocyte cell adhesion molecule (hepaCAM), a typical structure of immunoglobulin (Ig)-like adhesion molecules, inhibited the proliferation and the progression of cultured human bladder cancer cells. As increasing evidence reveals that aberrant activation of canonical Wnt pathway is involved in the pathogenesis of bladder cancer, and ß-catenin serves as a pivotal molecule of Wnt pathway. Then, we explored whether the anti-proliferation effect of hepaCAM was associated with Wnt/ß-catenin pathway in human bladder cancer cells. The negative correlation between hepaCAM and ß-catenin in transitional cell carcinoma of bladder (TCCB) was found. Follow by, studied the effect of hepaCAM on the key elements of Wnt pathway. Here, Our researches showed that hepaCAM played a central role in modulating the Wnt/ß-catenin signaling pathway by interfering nuclear protein levels of ß-catenin, leading to down-regulate transcriptional activity of LEF/TCF and its target genes c-Myc and cyclinD1. Mechanistically, we demonstrated that hepaCAM-activated GSK3ß led to elevate the phosphorylation of ß-catenin, contributing to the aberrant translocation of ß-catenin. In addition, Anti-proliferation and associated molecular mechanisms of hepaCAM were demonstrated by using vivo experiment. In conclusion, our reports uncover that expression of hepaCAM suppresses the proliferation of bladder cancer cells through a Wnt/ß-catenin-dependent signaling pathway in vitro and in vivo.

Down-regulation of miR-29c in human bladder cancer and the inhibition of proliferation in T24 cell via PI3K-AKT pathway.

The purpose of this study was to explore new tumor suppressor microRNA in bladder cancer and to conduct functional analysis of its suppressive role. To investigate the expression of miR-29c, qRT-PCR was used in 30 pairs of bladder cancer tissues and normal tissues (adjacent bladder tissue samples). The expression of miR-29c was down regulated in bladder cancer tissues compared with normal tissues. Also, the low-level expression of miR-29c was associated with tumor stage (P = 0.002), and ectopic over-expression of miR-29c in T24 cells can significantly inhibit cell proliferation, decrease motility, suppress the G1/S cell cycle transition and induce apoptosis. Furthermore, it could cause a decrease in AKT and GSK-3ß phosphorylation. While LY294002 reduced the protein level of pAKT, the over-expression of miR-29c can further decrease its level in T24 cells pretreated with LY294002. Our study also indicated that the proliferation inhibition of T24 may take place via AKT-GSK3ß pathway. Thus, miR-29c could be an active player in disease state of bladder cancer and it may be a promising tumor suppressor in bladder cancer.

Overexpression of HepaCAM inhibits cell viability and motility through suppressing nucleus translocation of androgen receptor and ERK signaling in prostate cancer.

BACKGROUND: HepaCAM is suppressed in a variety of human cancers, and involved in cell adhesion, growth, migration, invasion, and survival. However, the expression and function of HepaCAM in prostate cancer are still unknown. METHODS: HepaCAM expression has been detected by RT-PCR, Western blotting and immunohistochemistry staining in prostate cell lines RWPE-1, LNCap, DU145, PC3, and in 75 human prostate tissue specimens, respectively. Meanwhile, the cell proliferation ability was detected by WST-8 assay. The role of HepaCAM in prostate cancer cell migration and invasion was examined by wound healing and transwell assay. And flow cytometry was used to observe the apoptosis of prostate cancer cells. Then we detected changes of Androgen Receptor translocation and ERK signaling using immunofluorescence staining and western blot after overexpression of HepaCAM. RESULTS: The HepaCAM expression was significantly down-regulated in prostate cancer tissues and undetected in prostate cancer cells. However, the low HepaCAM expression was not statistically associated with clinicopathological characteristics of prostate cancer. Overexpression of HepaCAM in prostate cancer cells decreased the cell proliferation, migration and invasion, and induced the cell apoptosis. Meanwhile, HepaCAM prevented the androgen receptor translocation from the cytoplasm to the nucleus and down-regulated the MAPK/ERK signaling. CONCLUSION: Our results suggested that HepaCAM acted as a tumor suppressor in prostate cancer. HepaCAM inhibited cell viability and motility which might be through suppressing the nuclear translocation of Androgen Receptor and down-regulating the ERK signaling. Therefore, it was indicated that HepaCAM may be a potential therapeutic target for prostate cancer.

Exosome-derived microRNA-29c induces apoptosis of BIU-87 cells by down regulating BCL-2 and MCL-1.

BACKGROUND: Aberrant expression of the microRNA-29 family is associated with tumorigenesis and cancer progression. As transport carriers, tumor-derived exosomes are released into the extracellular space and regulate multiple functions of target cells. Thus, we assessed the possibility that exosomes could transport microRNA- 29c as a carrier and correlations between microRNA-29c and apoptosis of bladder cancer cells. MATERIALS AND METHODS: A total of 28 cancer and adjacent tissues were examined by immunohistochemistry to detect BCL-2 and MCL-1 expression. Disease was Ta-T1 in 12 patients, T2-T4 in 16, grade 1 in 8, 2 in 8 and 3 in 12. The expression of microRNA-29c in cancer tissues was detected by quantitative reverse transcriptase PCR (QRT- PCR). An adenovirus containing microRNA-29c was used to infect the BIU-87 human bladder cancer cell line. MicroRNA-29c in exosomes was measured by QRT-PCR. After BIU-87 cells were induced by exosomes-derived microRNA-29c, QRT-PCR was used to detect the level of microRNA-29c. Apoptosis was examined by flow cytometry and BCL-2 and MCL-1 mRNA expressions were assessed by reverse transcription-polymerase chain reaction. Western blotting was used to determine the protein expression of BCL-2 and MCL-1. RESULTS: The expressions of BCL-2 and MCL-1 protein were remarkably increased in bladder carcinoma (p<0.05), but was found mainly in the basal and suprabasal layers in adjacent tissues. The expression of microRNA-29c in cancer tissues was negatively correlated with the BCL-2 and MCL-1. The expression level of microRNA-29c in exosomes and BIU-87 cells from the experiment group was higher than that in control groups (p<0.05). Exosome-derived microRNA-29c induced apoptosis (p<0.01). Although only BCL-2 was reduced at the mRNA level, both BCL-2 and MCL-1 were reduced at the protein level. CONCLUSIONS: Human bladder cancer cells infected by microRNA- 29c adenovirus can transport microRNA-29c via exosomes. Moreover, exosome-derived microRNA29c induces apoptosis in bladder cancer cells by down-regulating BCL-2 and MCL-1.

Nuclear factor-κB signaling pathway is involved in phospholipase Cε-regulated proliferation in human renal cell carcinoma cells.

Phospholipase Cε (PLCε), a downstream effector of small GTPase superfamily, has been identified to play a crucial role in tumorigenesis. Previously, our studies have showed that PLCε promotes proliferation of renal cell carcinoma (RCC) cells. However, the molecular mechanisms by which PLCε enhances the survival phenotype of RCC cells are still not fully instructed. In the present study, we first demonstrated that PLCε was highly expressed and had a close correlation with Ki67 protein expression in RCC tissue samples. Further, we found that downregulation of PLCε expression repressed growth and induced apoptosis in RCC cells. In addition, we reported a mechanism by which knockdown of PLCε gene potently suppressed the nuclear factor kappa (NF-κB) signaling pathway through action on inhibitor of κB kinase. Moreover, silencing PLCε gene decreased vascular endothelial growth factor (VEGF) expression, which was a downstream growth factor of NF-κB signaling pathway. Finally, downregulation of VEGF was severely enhanced by treatment cells with NF-κB specific inhibitor BAY11-7028 in PLCε knockdown cells. Taken together, these findings suggest that PLCε promotes RCC cell growth via NF-κB-mediated upregulation of VEGF.