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.

Characterization of acute renal allograft rejection by proteomic analysis of renal tissue in rat.

Rapid and reliable biomarkers of renal allograft rejection have not been available. This study aimed to investigate biomarkers in renal allograft tissue using proteomic analysis. Orthotopic kidney transplantations were performed using Fisher (F344) or Lewis rats as donors and Lewis rats as recipients. Syngenic control group (Group I) constituted F344-to-F344 orthotopic kidney allo-transplantations (n = 8); and allogenic group (Group II) consisted of F344-to-Lewis orthotopic kidney allo-transplantations (n = 8). Renal tissues were harvested 7 days after transplantation. Samples were analyzed using 2-D electrophoresis and matrix assisted laser desorption ionization-time of flight mass spectrometry. 6 differentially expressed proteins were identified between allogenic group and syngenic control group. A rat model of acute renal allograft rejection was successfully set up. Differentially expressed proteins in renal allograft tissue of rat were detected using proteomic analysis and might serve as novel diagnostic and therapeutic targets in human. Quantitative proteomics, using MALDL-TOF-MS methodology has the potential to provide a profiling and a deeper understanding of acute renal rejection.

HepaCAM induces G1 phase arrest and promotes c-Myc degradation in human renal cell carcinoma.

Hepatocyte cell adhesion molecule (hepaCAM) encodes a generally inactive phosphorylated glycoprotein which mediates cancer cell proliferation, migration, and differentiation. We have reported that hepaCAM is down-regulated in renal cell carcinoma (RCC) and takes responsibility of cell growth inhibition. However, the precise mechanisms of hepaCAM inhibits cell growth is still unknown. In this study, we demonstrated that re-expression of hepaCAM can cause an accumulation in G0/G1 phase in 786-0 cells. This reaction was accompanied by a substantial reduction of c-Myc expression through using an ectopic hepaCAM expression system. Furthermore, we found a comparable decrease in proliferation and G0/G1 accumulation of 786-0 and RC-2 cells after treatment with a small molecule c-Myc inhibitor, 10058-F4. This indicated that the down regulation of c-Myc was an essential process in controlling growth inhibitory actions of hepaCAM. Nevertheless, re-expression of hepaCAM results in apparent reduction of c-Myc protein with no corresponding reduction of c-Myc mRNA. This suggests that this reaction might take place at a post-transcriptional level rather than transcriptional one. Consistent with these findings, hepaCAM decreased c-Myc stability by increasing the proportion of c-Myc phosphorylation on T58 which can be abrogated by a proteasomal inhibitor (MG132). Thus, our research implies that the decrease in c-Myc protein expression, resulting from ectopic expression of hepaCAM, may contribute to the inhibition of proliferation in these cells.

Normal values for renal parenchymal volume and kidney length as measured by non-enhanced multidetector spiral computed tomography.

BACKGROUND: Renal parenchymal volume (RPV) is considered an important index for clinical decisions. However, normal values have not been established, which hinders the clinical application of RPV. PURPOSE: To test the accuracy and reproducibility of RPV and to investigate the normal values of RPV and kidney length as measured by non-enhanced multidetector computed tomography (CT). MATERIAL AND METHODS: An animal model study was performed to test the accuracy and repeatability of RPV measured by CT. RPV of porcine kidneys was measured by water displacement (actual values) and non-enhanced multidetector CT. Individual RPV and kidney length were measured by non-enhanced CT in patients with no clinical history of renal disease (n = 722). Patient height, age, weight, body surface area (BSA), and body mass index (BMI) were assessed before each patient's CT examination was performed. RESULTS: RPV obtained by the CT method was within 2% of the RPV determined by the water displacement method. The normal values of RPV (M ± 1.96 standard deviation [SD]) were 145.72 ± 54.37 mL for men and 132.46 ± 41.94 mL for women. The normal values of kidney length (M ± 1.96 SD) were 10.27 ± 1.98 cm for men and 9.93 ± 1.58 cm for women. RPV did not significantly correlate with BSA or weight in women, but correlated significantly with height and age in both men and women. Of the assessed factors age, weight, height, BSA, and BMI, age, and height were the independent factors that best reflected RPV, in both men and women. CONCLUSION: The present animal study results showed that non-enhanced multidetector CT findings accurately reflect the RPV. The subsequent study performed in humans showed that the RPV of the presently sampled Chinese population was lower than the commonly quoted reference values obtained from Western populations.

[Glycosyl-phosphatidylinositol-anchored interleukin-2 expressed on tumor-derived exosomes induces anti-tumor immune response].

OBJECTIVE: To prepare IL-2-anchored and tumor-derived exosomes vaccine, and investigate the antitumor efficiency of the special cytotoxic T-lymphocytes induced by Ex/GPI-IL-2. METHODS: To construct pEGFP-N1-IL2gpi plasmid coding a fusion gene of a DNA oligo encoding GPI-anchor signal sequence attaching to human IL-2 cDNA. Then T24 cell lines stably expressing GPI-IL-2 proteins (T24/GPI-IL-2) were established. Ex/GPI-IL-2 were isolated and purified by ultrafiltration and sucrose gradient centrifugation, and the morphology and molecule markers were analyzed. The mixed lymphocyte reaction study and cytotoxic study were performed to determine the proliferative effect of T lymphocytes and the cytotoxicity induced by Ex/GPI-IL-2. RESULTS: The pEGFP-N1-IL2gpi plasmid was successfully constructed, and cell lines stably expressing GPI-IL-2 fusion proteins were established. Ex/GPI-IL-2 were small vesicular and saucer-shaped in diameter of 30-90 nm, containing heat shock protein 70, intercellular adhesion molecule-1, MAGE-1 and GPI-IL-2. Ex/GPI-IL-2-pulsed could dendritic cells induce proliferation of T cells and cytotoxic immune response more efficiently (P<0.05). CONCLUSIONS: GPI-IL-2 gene-modified tumor cells can make the exosomes containing GPI-IL-2 with an increased anti-tumor effect. Our study provides a feasible approach for exosome-based tumor immunotherapy of bladder transitional cell tumors.

[Preparation of renal cancer vaccine of IL-12-anchored exosomes and its antitumor effect in vitro].

OBJECTIVE: To prepare a vaccine of IL-12-anchored exosomes derived from renal cancer cells and to evaluate its antitumor effect in vitro. METHODS: A mammalian co-expression plasmid of glycolipid-anchor-IL-12 (GPI-IL-12) was constructed by subcloning IL-12A chain gene (P35 subunit) and a fusion gene containing GPI-anchor signal sequence and IL-12B chain gene (P40 subunit) in pBudCE4.1. Confocal laser scanning microscopy and flow cytometry were used to analyze the expression of the fusion proteins. Transmission electron microscopy and Western blot were used to identify the morphology and characteristic molecules of exosomes separated by ultrafiltration and sucrose gradient centrifugation. The function of IL-12-anchored exosomes was determined by IFN-gamma release assay. RESULTS: Mammalian co-expression plasmids were successfully constructed. Confocal laser scanning microscopy and flow cytometric analysis of the RC-2-GPI-IL-12 transfectants showed the expression of IL-12 on the cell surface. Exosomes were purified by ultrafiltration and sucrose gradient centrifugation, which were 30-80 nm in diameter, typically saucer-shaped, and expressing HSP70, ICAM-1, G250 and GPI-IL-12. (80.0 +/- 9.6) pg/ml of IL-12 was detected in 10 microg/ml exosomes and it significantly induced the release of IFN-gamma. Stimulation with EXO-IL-12 could efficiently induce antigen-specific cytotoxic T lymphocytes (CTL), resulting in more significant cytotoxic effects in vitro. CONCLUSION: A vaccine of exosomes-GPI-IL-12 can be obtained from the culture supernatant of renal cancer cells modified to express anchored IL-12. This vaccine expressing IL-12 and tumor associated antigen G250 may become a new strategy for the treatment of renal cancer.

PLCε knockdown prevents serine/glycine metabolism and proliferation of prostate cancer by suppressing YAP.

The metabolic reprogramming is an important basis for the development of many tumors, including prostate cancer (PCa). Metabolic changes in many amino acids consist of serine and glycine affect the biological behavior of them. Phospholipase C epsilon (PLCε) plays an important role as an oncogene. However, its role in regulating amino acid metabolism remains unclear. In this study, results found significantly positive correlation between PLCε and Yes-associated protein (YAP) in PCa tissues. LC-MS/MS and GC-MS results further displayed abnormally elevated levels of serine, glycine and its some downstream metabolites in the blood of PCa patients. Secondly, PLCε knockdown can inhibit serine/glycine producing and proliferation of PCa both in vivo and in vitro. Mechanistically, PLCε may affect the serine/glycine metabolism by regulating dephosphorylation and nuclear translocation of YAP. More interestingly, verteporfin (VP, a specific inhibitor of YAP) could effectively enhance the PLCε-depletion induced inhibition of serine/glycine secretion and growth. Overall, this research revealed the possibility of anomalous serine/glycine levels in the blood for the diagnosis of PCa, identified the important role of the PLCε/YAP axis in regulating serine/glycine metabolism, cell proliferation and tumor growth, and suggested the combination of VP with PLCε-depletion may provide a new idea for the treatment of PCa.

[Exosomes derived form bladder transitional cell carcinoma cells induce CTL cytotoxicity in vitro].

OBJECTIVE: To isolate and purify exosomes derived from human bladder transitional cell carcinoma T24 cells, analyze the morphology and protein composition, and investigate the antitumor effect of specific cytotoxic T lymphocytes induced by exosomes. METHODS: Exosomes were isolated and purified by ultrafiltration and sucrose gradient centrifugation, and characterized by electron microscopy and Western blot. Dendritic cells were amplified and purified from peripheral blood and pulsed with exosomes. Then they were co-cultured with T cells, and divided into 3 groups: exosome-pulsed DC group, unplused DC group and control group. Alamar-Blue assay was used to evaluate the specific cytolytic activity. RESULTS: The exosomes were in size about 30 approximately 90 nm saucer-shaped membranous vesicles. HSP70, ICAM-1 and CK20 were detected by Western blot. The CTL induced by DC pulsed with exosomes had significant cytolytic activity (P < 0.01). CONCLUSION: The exosomes derived from T24 cells are loaded with immunoprotein HSP70 and ICAM-1, and DC pulsed with exosomes can promote the anti-tumor effect of CTLs in vitro.

[Effects of silencing transforming growth factor-beta1 by RNAi upon transdifferentiation of renal allograft tubular epithelial cells in rats].

OBJECTIVE: To investigate the effects of shRNA-transforming growth factor (TGF)-beta1 plasmid upon epithelial-myofibroblast transdifferentiation of renal allograft in rats. METHODS: Divided the Wistar rats into 4 groups: Group J (sham-operated group), T (plasmid group), H (vacant plasmid group) and Y (simply transplantation group). The SD to Wistar rat transplant kidney-sclerosis accelerated model was constructed and transfected with the plasmid based on hydromechanics. Transplanted kidneys were collected at Months 1, 2 and 3 post-transplantation. The gene transcriptional levels of TGF-beta1 and E-cadherin were detected by RT-PCR and the protein variation of E-cadherin was examined by Western blotting. The pathological changes and infiltrated inflammatory cells were assessed by HE staining and the immunohistochemical staining of E-cadherin and alpha-SMA used to label epithelial cells and fibroblast in order to exhibit cell transdifferentiation. RESULTS: Compared with Group H and Y, the mRNA transcription of TGF-beta1 was obviously inhibited in the Group T: at Month 3, the TGF-beta1 mRNA of Group T is 0.73 +/- 0.08, significantly lower than Group H and Y (0.92 +/- 0.07 and 0.95 +/- 0.04, both P < 0.01); the expression of E-cadherin was maintained at a high level: at the Month 3, the E-cadherin mRNA of Group T is 0.39 +/- 0.11, significantly higher than Group H and Y (0.15 +/- 0.07, and 0.17 +/- 0.06, both P < 0.01); the E-cadherin protein of Group T is 0.38 +/- 0.08, significantly higher than group H and Y (0.15 +/- 0.07 and 0.15 +/- 0.07, both P < 0.01); epithelial cells were much more and fibroblast was much less than that of Group H and Y; there were also less infiltrated chronic inflammatory cells and extracellular matrix deposition in the Group T. CONCLUSION: The shRNA-TGF-beta1 plasmid can inhibit the epithelial-myofibroblast transdifferentiation of renal allograft in rats. The mechanisms may be associated with its effects of down-regulating TGF-beta1 and up-regulating E-cadherin.

[Proliferation apoptotic influence of crocin on human bladder cancer T24 cell line].

OBJECTIVE: To investigate the proliferation, apoptosis and mechanisms on T24 cell of transitional cell carcinoma of bladder (TCCB) by crocin. METHOD: MTT assay was used to evaluate the proliferation of T24 cells. The changes of cell cycle and cell apoptotic percentage were measured by flow cytometry. T24 cells were inoculated into BALB/c nude mice to establish model of carcinoma of bladder. The mice were randomly divided into control group and experimental group. After treatment with 50 mmol x L(-1) crocin, the inhibited growth of tumor was observed. Electronic microscope was used to observe the morphological changes. The expressions of Bcl-2, Bax, Survivin and Cyclin D1 were detected by immunohistochemistry. RESULT: The growth of T24 cells was remarkably inhibited after treatment of crocin. Flow cytometric profiles revealed that crocin led to the increase of the cells in G0/G1 phase, the percentage of cell apoptosis was also increased. Crocin could inhibit the growth of BALB/c xenograft tumor. The morphology changes of cell apoptosis were observed. Bcl-2, Cyclin D1 and survivin expressions determined by immunohistochemical staining were down-regulated after treatment with Bax expression up-regulated. CONCLUSION: Crocin exerts both in vitro and in vivo anti-cancer effect on TCCB T24 cell line. The mechanisms may change tumour cell cycle and induce tumour cell apoptosis by down-regulating the expression of Bcl-2, Survivin, Cyclin D1 and up-regulating the expression of Bax.

miR-93 Promotes the Growth and Invasion of Prostate Cancer by Upregulating Its Target Genes TGFBR2, ITGB8, and LATS2.

This study aimed to evaluate the effects of miR-93 on the growth and invasiveness of prostate cancer (PC) cells (PCCs). Real-time PCR was carried out to detect the expression of miR-93 in the PC tissues and cell lines. The adjacent normal tissues served as controls. For in vitro experiments, methyl thiazolyl tetrazolium, clone formation, tumor cell invasion assays, and western blot analysis (WBA) were performed to confirm the variations in the proliferation and invasiveness of PCCs, prior and subsequent to transfection with an miR-93 antisense oligonucleotide (ASO), which blocks miR-93 binding to its target. Furthermore, the effect of miR-93 on the proliferation of PCCs was examined. Finally, the expression levels of the target genes of miR-93 were determined by WBA. miR-93 expression was higher in PC tissues than in the adjacent normal tissues, and a reduction in the miR-93 level remarkably inhibited the proliferation and invasiveness of PCCs. Moreover, miR-93 enhanced the expression of its target genes TGFΒR2, ITGB8, and LATS2. The results of this study suggest that miR-93 may promote the proliferation and invasion of PCCs by upregulating its target genes TGFBR2, ITGB8, and LATS2.

[Construction of deltaNp63 specific small hairpin RNA expressing plasmid and its role in bladder cancer--a preliminary study].

OBJECTIVE: To construct deltaNp63 specific small hairpin RNA (shRNA) expressing plasmid,to examine its inhibitory effect to the expression of deltaNp63 protein and mRNA in transitional cell carcinoma of the bladder (TCCB) , its effect on TCCB cells cycle and proliferation. METHODS: DeltaNp63 specific oligonucleotides were designed and synthesized. These oligonucleotides were annealed to form double strand DNA fragments and this fragment was cloned into Pgenesil-1 plasmid. The recombinant deltaNp63-shRNA expression construct was confirmed by using Pst I + Sal I double digestion and by sequencing. Fluorescence staining was used to confirm the success of transfection in TCCB cells under the fluorescence microscope. The inhibitory effect of deltaNp63-shRNA construct was examined with semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemical staining assay. The cell cycle of TCCB cells was assayed by flow cytometry (FCM). The cellular proliferation of TCCB cells was assayed by tetrazolium bromide (MTT) colorimetry. RESULTS: The deltaNp63-shRNA expression plasmid was successfully constructed and transfected into TCCB cells. It can effectively reduce the expression of deltaNp63 protein and mRNA. The reduction rate of deltaNp63 mRNA was 63.0%, and the G0/G1 ratio was increased and S phase was decreased in transfected TCCB cells. The cellular proliferation was also lower in transfected 5637 cells in comparrison with that of non-transfected TCCB cells. CONCLUSION: A deltaNp63-shRNA expression plasmid, constructed from Pgenesil-1 plasmid, can successfully be transfected into TCCB cells and can effectively inhibit the expression of deltaNp63 protein and mRNA. It also can take part in regulation of the cell cycling and inhibit the cellular proliferation of TCCB cells.

[Effect of saffor (Carthamus tinctorius) injection on renal ischemia/reperfusion injury in rats].

OBJECTIVE: To study the effect and mechanism of saffor injection on renal ischemia/reperfusion (I/R) injury in rats. METHOD: Seventy-five SD rats were randomly divided into five groups (n = 15, in each), normal control groups, I/R control groups, low-dose treatment groups, middle-dose treatment groups and high-dose treatment groups. After rat's I/R injury model was established, renal function was assessed by measuring serum creatinine, blood urea nitrogen, urine osmotic pressure and urine osmotic pressure/blood osmotic pressure, the apoptosis rate in I/R renal tissure was measured by TUNEL method and caspase-3 concentration was measured by immunohistochemistry. RESULT: Reperfusion of the ischemic kidney induced marked renal dysfunction. Saffor injection significantly inhibited the reperfusion-associated increase in apoptosis rate and caspase-3 protein absorbance value. Moreover, the renal dysfunction at all treatment groups was markedly ameliorated by Saffor injection. (P < 0.01). CONCLUSION: The results show that saffor injection significantly reduces the renal dysfunction and injury caused by I/R of the kidney, And the protective effect of Saffor injection may be related to the inhibition of cell apoptosis and caspase-3 gene expression following renal I/R.

Hypoxia promotes 786-O cells invasiveness and resistance to sorafenib via HIF-2α/COX-2.

Accumulating evidences indicated that hypoxia-induced factors and COX-2 play a important role in tumorigenesis in various human cancer. Yet, the relationship between HIFs and COX-2 in human renal cancer remains unclear. The present study was to examine the role of HIFs and COX-2 in the invasiveness and the resistance to target agent in renal cancer cell line (786-O). In 786-O cells, hypoxia induced the increase in the protein expression of HIF1 and HIF2. We also demonstrate that hypoxia up-regulated the protein expression of COX-2 and Snail, but down-regulation of E-cadherin expression in 786-O cells promoted the invasiveness of 786-O cells and enhanced the resistance of 786-O cells to sorafenib. siRNA target to HIF1α, HIF2α and NS398, a selective inhibitor of COX-2, were used in this study. Only siRNA-HIF2α significantly suppressed the protein expression of HIF2 and COX-2, then decreased the invasive ability and resistance of 786-O cells to sorafenib under hypoxia. NS398 attenuated the increase in invasive cells number and the IC50 value of sorafenib induced by hypoxia. In conclusion, our results demonstrated that hypoxia promoted the invasiveness and resistance of 786-O cells to sorafenib via HIF2 and COX-2 and induced the activation of Snail/E-cadherin, suggesting that a signalling mechanism involving HIF2/COX2 modulates invasiveness and resistance to sorafenib in 786-O cells under hypoxia.

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.

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.

A new PKCα/ß/TBX3/E-cadherin pathway is involved in PLCε-regulated invasion and migration in human bladder cancer cells.

Although PLCε has been verified to enhance bladder cancer cell invasion, the signaling pathways responsible for this remain elusive. Protein kinase C (PKCα/ß), which is involved in cancer development and progression, has been demonstrated to be activated by PLCε. However, the roles of PKCα/ß in PLCε-mediated bladder carcinoma cell invasion and migration have not been clearly identified. In this study, to determine what role PKCα/ß plays in PLCε-mediated bladder cancer cell invasion and migration, we silenced PLCε gene by adenovirus-shPLCε in T24 and BIU-87 cells and then revealed that it significantly inhibited cell migration and invasion. Further research indicated that cell bio-function of PLCε-regulated was related with PKCα/ß activity. These in vitro findings were supported by data from bladder carcinoma patient samples. In 35 case bladder cancer tumor samples, PLCε-overexpressing tumors showed significantly higher positive rates of PKCα/ß membrane immunohistochemistry staining than PLCε-low-expressing tumors. Mechanistically, study further showed that PLCε knockdown gene induced E-cadherin expression and decreased TBX3 expression, both of which were dependent on PKCα/ß activity. In addition, we demonstrated that treatment cells with TBX3-specific shorting hairpin RNA (shRNA) up-regulated E-cadherin expression and inhibited cell invasion/migration. Moreover, in in vivo experiment, immunohistochemistry analysis of Ad-shPLCε-infected tumor tissue showed low expression levels of phospho-PKCα/ß and TBX3 and high expression levels of E-cadherin compared with those of the control group. In summary, our findings uncover that PKCα/ß is critical for PLCε-mediated cancer cell invasion and migration and provide valuable insights for current and future Ad-shPLCε and PKCα/ß clinical trials.

Review of selenium and prostate cancer prevention.

Prostate cancer is the most common malignancy in men in the United States. Surgery or radiation are sometimes unsatisfactory treatments because of the complications such as incontinence or erectile dysfunction. Selenium was found to be effective to prevent prostate cancer in the Nutritional Prevention of Cancer Trial (NPC), which motivated two other clinical trials: the Selenium and Vitamin E Cancer Prevention Trial (SELECT) and a Phase III trial of selenium to prevent prostate cancer in men with high-grade prostatic intraepithelial neoplasia. However, these two trials failed to confirm the results of the NPC trial and indicated that the selenium may not be preventive of prostate cancer. In this article we review the three clinical trials and discuss some different points which might be potential factors underlying variation in results obtained.

Bladder cancer cell-derived exosomes inhibit tumor cell apoptosis and induce cell proliferation in vitro.

Exosomes are small membrane vesicles released by a variety of mammalian cells into the extracellular space and are involved in cell­to­cell signaling. This study aimed to investigate the effects of bladder cancer cell­derived exosomes on the regulation of tumor cell viability and apoptosis, as well as the underlying molecular events. Exosomes were purified from the supernatants of human bladder cancer T24 cell cultures. Transmission electron microscopy was used to confirm their morphology and western blot analyses determined the protein content of cells. Subsequently, bladder cancer cell lines were treated with different concentrations of exosomes. Tumor cell viability was shown to be reduced, as detected by the Cell Counting Kit­8 assay. Annexin V/flow cytometric assays showed that exosomes inhibited apoptosis of bladder cancer cell lines in a dose- and time­dependent manner. Exosomes were demonstrated to upregulate the expression of Bcl­2 and Cyclin D1 proteins, but reduce the levels of Bax and caspase­3 proteins in these cells. Moreover, exosomes dose­dependently increased the expression of phosphorylated Akt and extracellular signal­regulated protein kinase (ERK). In conclusion, this study demonstrated that bladder cancer cell­derived exosomes inhibited tumor cell apoptosis, which was associated with the activation of Akt and ERK pathway genes, suggesting that tumor­derived exosomes are involved in bladder cancer progression. Inhibition of exosome formation and release may therefore be a novel strategy in future treatment of bladder cancer.