ITPKA1 Promotes Growth, Migration and Invasion of Renal Cell Carcinoma via Activation of mTOR Signaling Pathway.

BACKGROUND: Renal cell cancer (RCC) is one of the most lethal malignancies of the kidney in adults. mTOR (mammalian target of rapamycin) signaling pathway plays a pivotal role in RCC tumorigenesis and progression and inhibitors targeting the mTOR pathway have been widely used in advanced RCC treatment. Therefore, it is of great significance to explore the potential regulators of the mTOR pathway as RCC therapeutic targets. MATERIALS AND METHODS: Bioinformatics analysis was used to screen out the most significant differentially expressed genes in the RCC dataset of The Cancer Genome Atlas (TCGA). Real-time PCR and Western-blot analysis were utilized to examine the expression of inositol-1,4,5-trisphosphate-3-kinase-A (ITPKA) in four RCC cell lines and one human embryonic kidney cell line. Cell counting Kit-8 and colony formation assay were performed to estimate the effect of ITPKA on the proliferation ability of RCC cells. Wound healing and Transwell assays were used to test the effect of ITPKA on RCC cell migration and invasion. Xenograft formation assay was performed in nude mice to investigate the effect of ITPKA in vivo. mTORC1 pathway inhibitor was added to explore the mechanisms by which ITPKA regulates RCC cell growth and progression. RESULTS: Based on bioinformatics analysis, ITPKA is screened out as one of the most significant differentially expressed genes in RCC. ITPKA is upregulated and positively correlated with RCC malignancy and poorer prognosis. ITPKA promotes RCC growth, migration and invasion in cultured cells, and accelerates tumor growth in nude mice. Mechanistically, ITPKA stimulates the mTORC1 signaling pathway which is a requirement for ITPKA modulation of RCC cell proliferation, migration and invasion. CONCLUSION: Our data demonstrate a critical regulatory role of the ITPKA in RCC and suggest that ITPKA/mTORC1 axis may be a promising target for diagnosis and treatment of RCC.

miR-325-3p Overexpression Inhibits Proliferation and Metastasis of Bladder Cancer Cells by Regulating MT3.

BACKGROUND miRNAs have been widely used in cancer treatment. Our study was designed to explore the effects of miR-325-3p in bladder cancer cells. MATERIAL AND METHODS Levels ofd miR-325-3p and MT3 in bladder cancer tissues and cells were assessed by quantitative real-time polymerase chain reaction (qRT-PCR). miR-325-3p mimics were transfected into bladder cancer T24 cells, and cell migration and invasion rates and cell proliferation were assessed by transwell assay and Cell Counting Kit-8 (CCK-8). The target mRNA for miR-325-3p was predicted by Targetscan7.2 and confirmed by dual-luciferase reporter assay. More experiments were performed to confirm the effects of miR-325-3p and MT3 in T24 cells. Additionally, the levels of TIMP-2, MMP9, and E-cadherin were assessed by Western blotting to identify the effects of miR-325-3p and MT3 on epithelial-mesenchymal transition (EMT). RESULTS miR-325-3p expression was reduced and MT3 was increased in bladder cancer tissues and bladder cancer cells. miR-325-3p mimics suppressed cell proliferation ability and invasion and migration rates of T24 cells. Moreover, miR-325-3p was confirmed to target MT3. Further experiments showed that the effects of increased cell proliferation, invasion, migration, and EMT promoted by MT3 overexpression were abolished by miR-325-3p mimics, proving that miR-325-3p is a tumor suppressor through targeting MT3 in bladder cancer cells. CONCLUSIONS Downregulation of miR-325-3p in bladder cancer regulates cell proliferation, migration, invasion, and EMT by targeting MT3. Furthermore, miR-325-3p is a potential therapeutic target in treating bladder cancer.

FOXJ1 promotes bladder cancer cell growth and regulates Warburg effect.

Forkhead Box J1 (FOXJ1) which belongs to Fox gene family, plays complex and crucial roles in processes of development, organogenesis, regulation of the immune system, as well as progression of several malignancies. However, how FOXJ1 functions in bladder cancer remains unclear. Here, we report that FOXJ1 is upregulated in most bladder cancer patients, and predicts poor clinical outcomes. FOXJ1 facilitates bladder cancer cell proliferation and colony formation. FOXJ1 knockdown suppresses bladder tumor growth in nude mice. Mechanistically, FOXJ1 enhances glycolysis by increasing glucose uptake, lactate production and extracellular acidification rate (ECAR), and decreasing ATP generation and oxygen consumption rate (OCR) in bladder cancer cells. Our findings provide clues regarding the role of FOXJ1 as a tumor inducer in bladder cancer and an enhancer in glycolysis. Targeting FOXJ1 could be a potential therapeutic strategy in bladder cancer.

VHL-TGFBI signaling is involved in the synergy between 5-aza-2'-deoxycytidine and paclitaxel against human renal cell carcinoma.

PURPOSE: To analyse the role of von Hippel-Lindau (VHL) and transforming growth factor ß-induced (TGFBI) in synergistic mechanisms of 5-aza-2'-deoxycytidine (DAC) and paclitaxel (PTX) against renal cell carcinoma (RCC). METHODS: To elucidate the role in the synergy between DAC and PTX against RCC cells, TGFBI expression was regulated using siRNA technology and an expression vector containing the full-length cDNA for TGFBI was also transfected into RCC cells. The proliferation of RCC cells was evaluated using the WST-1 assay and TGFBI expression was detected by real-time PCR (RT-PCR), and Western blot. RESULTS: The results indicated that the expression of TGFBI was significantly decreased by DAC or PTX alone in vitro and in vivo. Moreover, the combination of DAC and PTX caused a synergistic decrease in the expression of TGFBI in RCC cells. We also investigated the effect of VHL-TGFBI signaling on the synergy between DAC and PTX, although the synergy between the two medications was not abolished by interfering with VHL activity or TGFBI expression. RCC cells without VHL activity and RCC cells expressing high levels of TGFBI displayed an increased synergistic effect compared to control cells. CONCLUSION: Our study suggests that VHL-TGFBI signaling is involved in the synergy between DAC and PTX against RCC cells. In addition, the synergy between DAC and PTX is more effective in VHL inactive RCC cells.

VHL-TGFBI signaling is involved in the synergy between 5-aza-2'-deoxycytidine and paclitaxel against human renal cell carcinoma.

PURPOSE: To analyse the role of von Hippel-Lindau (VHL) and transforming growth factor ß-induced (TGFBI) in synergistic mechanisms of 5-aza-2'-deoxycytidine (DAC) and paclitaxel (PTX) against renal cell carcinoma (RCC). METHODS: To elucidate the role in the synergy between DAC and PTX against RCC cells, TGFBI expression was regulated using siRNA technology and an expression vector containing the full-length cDNA for TGFBI was also transfected into RCC cells. The proliferation of RCC cells was evaluated using the WST-1 assay and TGFBI expression was detected by real-time PCR (RT-PCR), and Western blot. RESULTS: The results indicated that the expression of TGFBI was significantly decreased by DAC or PTX alone in vitro and in vivo. Moreover, the combination of DAC and PTX caused a synergistic decrease in the expression of TGFBI in RCC cells. We also investigated the effect of VHL-TGFBI signaling on the synergy between DAC and PTX, although the synergy between the two medications was not abolished by interfering with VHL activity or TGFBI expression. RCC cells without VHL activity and RCC cells expressing high levels of TGFBI displayed an increased synergistic effect compared to control cells. CONCLUSIONS: Our study suggests that VHL-TGFBI signaling is involved in the synergy between DAC and PTX against RCC cells. In addition, the synergy between DAC and PTX is more effective in VHL inactive RCC cells.

Effect of icarisid II on diabetic rats with erectile dysfunction and its potential mechanism via assessment of AGEs, autophagy, mTOR and the NO-cGMP pathway.

Erectile dysfunction (ED) is a major complication of diabetes mellitus. Icariin has been shown to enhance erectile function through its bioactive form, icarisid II. This study investigates the effects of icarisid II on diabetic rats with ED and its potential mechanism via the assessment of advanced glycosylation end products (AGEs), autophagy, mTOR and the NO-cGMP pathway. Icarisid II was extracted from icariin by an enzymatic method. In the control and diabetic ED groups, rats were administered normal saline; in the icarisid II group, rats were administered icarisid II intragastrically. Erectile function was evaluated by measuring intracavernosal pressure/mean arterial pressure (ICP/MAP). AGE concentrations, nitric oxide synthase (NOS) activity and cGMP concentration were assessed by enzyme immunoassay. Cell proliferation was analysed using methyl thiazolyl tetrazolium assay and flow cytometry. Autophagosomes were observed by transmission electron microscopy, monodansylcadaverine staining and GFP-LC3 localisation. The expression of NOS isoforms and key proteins in autophagy were examined by western blot. Our results have shown that Icarisid II increased ICP/MAP values, the smooth muscle cell (SMC) growth curve, S phase and SMC/collagen fibril (SMC/CF) proportions and decreased Beclin 1 (P<0.05). Icarisid II significantly increased the proliferative index and p-p70S6K(Thr389) levels and decreased the numbers of autophagosomes and the levels of LC3-II (P<0.01). Icarisid II decreased AGE concentrations and increased cGMP concentration, NOS activity (P<0.05) and cNOS levels (P<0.01) in the diabetic ED group. Therefore, Icarisid II constitutes a promising compound for diabetic ED and might be involved in the upregulation of SMC proliferation and the NO-cGMP pathway and the downregulation of AGEs, autophagy and the mTOR pathway.