Renal Cell Proliferation Research Articles

Let‑7a‑5p may participate in the pathogenesis of diabetic nephropathy through targeting HMGA2.

Diabetic nephropathy (DN) is one of the most common complications of diabetes mellitus (DM), and has been demonstrated as one of the major causes of renal failure. In a previous study, it was noted that microRNA let-7a-5p was downregulated in DN; however, the underlying mechanism requires additional investigation. The aim of the present study was to investigate the roles of let-7a-5p in the pathogenesis of DN and its associated mechanism. The renal tissues of db/db and db/m mice, and renal mesangial cells treated with high concentrations of glucose were obtained; reverse transcription-quantitative polymerase chain reaction, and western blot analysis were applied to detect the expression of let-7a-5p and high-mobility group AT-hook 2 (HMGA2) in vivo and in vitro. In addition, renal mesangial cells cultured under high-glucose conditions (20 and 30 mmol/l) were transfected with either let-7a-5p mimics or let-7a-5p inhibitors. The effects of let-7a-5p on the proliferation and apoptosis of renal mesangial cells were examined using CCK-8 and flow cytometry methods. Additionally, cells were collected and the expression of phosphoinositide 3-kinase (PI3K), phosphorylated protein kinase B (p-AKT) and HMGA2 was analyzed with western blot analysis. Finally, a dual luciferase reporter assay was performed to confirm whether HMGA2 was a direct target of let-7a-5p. Let-7a-5p was significantly downregulated and HMGA2 was markedly upregulated in the tissue samples of DN mice and renal mesangial cells cultured under high-glucose conditions. In addition, transfection of let-7a-5p mimics induced a significant decrease in the proliferation and increase in the apoptosis of renal mesangial cells cultured under high-glucose conditions; transfection of let-7a-5p inhibitors exhibited the opposite effects. Furthermore, transfection of let-7a-5p mimics also led to the inhibition of the PI3K-AKT signaling pathway; transfection of let-7a-5p inhibitors may activate the PI3K-AKT signaling pathway through the increase in PI3K and AKT levels. Finally, a dual luciferase reporter assay confirmed that HMGA2 is a direct target of let-7a-5p. Let-7a-5p was downregulated in DN and may participate in the pathogenesis of DN via regulating HMGA2 expression and the PI3K-AKT signaling pathway.

Effect of over-expressed miR-487A-3p on proliferation and invasion of renal cancer cells and its mechanism

Objective To investigate whether microRNA-487a-3p (miR-487a-3p) can inhibit the proliferation and invasion of renal cancer cells by inhibiting the expression of its target gene TM4SF1 gene. Methods The miR-487a-3p mimics or miR-NC were transfected into renal cell carcinoma cells (ACHN) by LipofectamineTM 2000 and divided into experimental group and control group.The expression of miR-487a-3p after transfection was detected by real-time fluorescence quantitative polymerase chain reaction (qPCR). The cell cycle was evaluated by flow cytometry. MTT assay was used to detect cell proliferation. Transwell invasion assay was used to detect cell invasion. TM4SF1 wild-type (WT) and mutant 3'UTR-luciferase reporter vector were contructed. The dual luciferase reporter gene system was used to detect the effect of miR-487a-3p on the luciferase activity of TM4SF1 wild-type and mutant 3'UTR. The expression of TM4SF1 mRNA and protein in transfected cells were detected by qPCR and Western blot. Results The expression of miR-487a-3p in ACHN cells in the experimental and control groups was (33.12±3.35) and (1.04±0.18), respectively, with significant difference (t=9.553, P<0.001). The proportion of cells in experimental group(42.07±1.62) at G0-G1 phase was significantly higher than that(58.42±2.15) in control group (t=6.087, P=0.001). The proliferation activity of ACHN cell in the experimental group was significantly lower than the control group (P<0.05). The number of invasiveness of ACHN cells in the experimental group(114.70±20.38) was significantly lower than that(269.30±18.12) in the control group (t=5.670, P=0.001). Bioinformatics software and dual luciferase reporter assay showed that TM4SF1 was the target gene of miR-487a-3p (P<0.01). The expression of TM4SF1 mRNA in ACHN cells in experimental group(0.47±0.08) was significantly lower than that(1.02±0.10) in control group, and the difference was statistically significant (t=4.122, P=0.006). The expression of TM4SF1 protein in experimental ACHN cells was significantly lower than that in control group. Conclusions miR-487a-3p can inhibit the proliferation and invasion of ACHN cells in renal cell carcinoma by inhibiting the expression of its target gene TM4SF1, which is expected to become an important target for treatment of renal cell carcinoma in the future. Key words: Kidney Neoplasms; MicroRNAs; Cell Proliferation; Neoplasm Invasiveness

RBCK1 promotes p53 degradation via ubiquitination in renal cell carcinoma

Renal cell carcinoma (RCC) accounts for approximately 3% of adult malignancies, and the incidence of RCC continues to rise worldwide. Although RCC can be treated with surgery at an early stages, the five-year survival rates have been observed to decline dramatically in patients with advanced disease. Most patients with RCC treated with cytotoxic or targeted drugs will develop resistance at some point during therapy. Thus, it is necessary to identify novel therapeutic targets for RCC. Here, we found that RANBP2-type and C3HC4-type zinc finger-containing 1 (RBCK1) expression was upregulated in human RCC samples. Analysis of multiple public databases revealed the correlation between RBCK1 expression and poor prognosis in RCC patients. Subsequently, we performed RBCK1 depletion experiments in RCC cells that severely affected the in vivo and in vitro proliferation of renal cancer cells. The effects of RBCK1 on cell proliferation could be rescued with p53 expression knockdown in two cell lines expressing wild-type p53. Further experiments demonstrated that RBCK1 could facilitate p53 poly-ubiquitination and degradation by direct interaction with p53. Together, our results show that RBCK1 may serve as a promising target for RCC therapy by restoring p53 functions.

Bone Marrow Derived Mesenchymal Stromal Cells Ameliorate Ischemia/Reperfusion Injury-Induced Acute Kidney Injury in Rats via Secreting Tumor Necrosis Factor-Inducible Gene 6 Protein.

Aims To investigate whether bone marrow derived mesenchymal stromal cells (BMSC) have ameliorated ischemia/reperfusion injury-induced acute kidney injury (IRI-AKI) via tumor necrosis factor-inducible gene 6 protein (TSG-6) and how TSG-6 exerted this effect. Methods We used lentiviral vectors of short hairpin RNA (shRNA) targeting TSG-6 gene to silence TSG-6 in BMSC. And TSG-6-silenced BMSC were administrated into IRI-AKI rats. Then we analyzed serum creatinine (Scr) and renal histology of IRI-AKI rats treated with BMSC after different pretreatments. Furthermore, we explored the effect of TSG-6 on renal tubular epithelial cells proliferation in vivo and in vitro assays. Results The Scr levels of IRI-AKI rats treated with BMSC (73.5±7.8 μmol/L) significantly decreased compared to those of IRI-AKI rats treated without BMSC (392.5±24.8 μmol/L, P<0.05) or with DMEM (314.0±19.8 μmol/L, P<0.05). Meanwhile, the renal tissue injury in IRI-AKI rats treated with BMSC improved markedly. However, the Scr levels of IRI-AKI rats treated with TSG-6-silenced BMSC (265.1±21.2 μmol/L) significantly increased compared to those with BMSC (74.0±8.5 μmol/L, P<0.05). The proportion of Ki67-positive cells was reduced in IRI-AKI rats treated with TSG-6-silenced BMSC compared to that in IRI-AKI rats treated with BMSC (29.7±0.8% versus 43.4±3.0%, P<0.05). In vitro, the cell proliferation rate of TSG-6-stimulated NRK-52E cells under hypoxia (89.2±3.9%) increased significantly compared to that of NRK-52E cells alone under hypoxia (82.4±0.8%, P<0.05). Similarly, the proportion of Ki67-positive cells was significantly elevated in TSG-6-stimulated NRK-52E cells under hypoxia. Furthermore, TSG-6 could inhibit infiltration of neutrophils in kidney tissue of IRI-AKI. Conclusions TSG-6 plays a key role in the treatment of IRI-AKI with BMSC, which may be due to its effect on promoting renal tubular epithelial cells proliferation by modulating inflammation.

Low-dose 2-deoxyglucose and metformin synergically inhibit proliferation of human polycystic kidney cells by modulating glucose metabolism

Polycystic kidney disease (PKD) is a common hereditary kidney disease with abnormal proliferation and apoptosis of kidney cystic epithelial cells, eventually leading to chronic renal failure. Currently, there are no effective treatment methods. Similar to tumor cells, cystic epithelial cells have abnormal glycolysis and over-activation of proliferation signaling pathways. In the present study, for the first time, we investigated the effects of low-dose combinational use of 2-deoxyglucose (2-DG) and metformin (MET) on the proliferation and apoptosis in the human cystic kidney epithelial cells. Cystic epithelia cells were divided into control group, 2-DG group, MET group and 2-DG+MET group. Cell Proliferation, apoptosis and glucose metabolism were measured in each group. The results showed that low-dose combinational treatment of 2-DG and MET significantly inhibited the proliferation of renal cystic epithelial cells by suppressing the activities of PKA, mTOR and ERK signaling pathways and upregulating PI3K/Akt pathway. Combination of both drugs increased the apoptosis rates of cystic epithelial cells. Two drugs inhibited glucose metabolic phenotypes, glycolysis and oxidative phosphorylation, and significantly lowered the intracellular ATP level in cystic epithelial cells. 2-DG could also neutralize excessive production of lactate (lactic acidosis) caused by MET and both drugs had complementary effect for cystic epithelial cells. These results reveal that combinational use of low-dose 2-DG and MET can markedly inhibit proliferation via modulating glucose metabolic phenotypes in human polycystic kidney epithelial cells, low-dose combinational use of both drugs can also lower the toxic effects of each drug, and is a novel strategy for future treatment of human polycystic kidney disease.

Chitosan Oligosaccharides Induce Apoptosis in Human Renal Carcinoma via Reactive-Oxygen-Species-Dependent Endoplasmic Reticulum Stress.

In recent years, various studies have confirmed the role of natural products as effective cancer prevention and treatment drugs. The present study demonstrated that chitosan oligosaccharide (COS) from shells of shrimp and crab caused an inhibitory effect on the proliferation of human renal carcinoma in vitro and in vivo. First, the in vivo biodistribution of COS was investigated by the synthesis of cyanine-7-labeled COS (COS-Cy7) following tail vein injection. The kidney was found to be a major target organ. Then, the impacts on renal carcinoma cell proliferation, apoptosis, and reactive oxygen species (ROS) production were observed in vitro, and an orthotopic xenograft tumor model was designed to evaluate the antitumor efficacy of COS in vivo. In renal carcinoma cells, COS induced G2/M phase arrest and apoptosis in a ROS-dependent fashion. COS significantly promoted mRNA expression of nuclear factor erythroid 2-related factor (Nrf2) and Nrf2 target genes, such as heme oxygenase 1, modifier subunit of glutamate cysteine ligase, and solute carrier family 7 member 11. Additionally, COS significantly upregulated the protein expression of glucose-regulated protein 78, protein RNA-like endoplasmic reticulum (ER) kinase, eukaryotic initiation factor 2α, activating transcription factor 4, C/EBP homologous protein, and cytochrome c, which justified the activation of the ER stress signaling pathway. In vivo, COS repressed tumor growth and induced apoptosis and ROS accumulation, consistent with the in vitro results. Taken together, COS repressed human renal carcinoma growth and induced apoptosis both in vitro and in vivo, mainly via ROS-dependent ER stress pathways.

Regulation of TGF-β1-Mediated Wnt/β-catenin Pathways Plays an Important Role in EMT-like Transformation Induced by Cadmium

Backgrounds Some studies have reported that different dosage of cadmium (Cd) had a biphasic effect on cells viability, Cd at a dose of more than 20 μM can induce excessive apoptosis which past studies were mainly concentrated on while there has been few reports on a relatively lower dose of Cd causing abnormal proliferation. Kidney is the most sensitive target organ of Cd, however, the mechanism how Cd at a relatively lower dose affects the proliferation of renal tubular epithelial cells (RTEC) is not yet clear at present. Objectives To explore the toxic effect of Cd at a relatively lower dose on RTEC and study the mechanism how the TGF-β1-mediated Wnt/β-catenin pathways involved in EMT-like transformation induced by Cadmium. Methods The RTEC were isolated by mixed collagenase digestion methods and the effect of Cd at different dose on cell viability was detected by MTT assay. The effect of Cd at Hormesis zone dose on the expression of c-myc, cyclinD1, α-SMA, TGF-β1, Wnt and β-catenin were determined by qRT-PCR. Results When exposed for 24h-72h, Cd at a dose of 2.5 μM had maximum proliferation promoting effect. Cd at Hormesis zone dose could up-regulate the expression of cell cycle and proliferation key regulators c-myc and cyclinD1 in an obvious time-dependent manner. Cd at Hormesis zone dose could significantly promote α-SMA, TGF-β1, Wnt and β-catenin mRNA expression. Conclusions Cd at Hormesis zone dose could induce RTEC significant abnormal proliferation and EMT-like transformation. The activation of TGF-β1-mediated Wnt/β-catenin pathways might play a key role in EMT-like transformation induced by Cd at Hormesis zone dose.

FBXO22 Suppresses Metastasis in Human Renal Cell Carcinoma via Inhibiting MMP-9-Mediated Migration and Invasion and VEGF-Mediated Angiogenesis.

F-box only protein 22 (FBXO22), a substrate receptor of the SKP1-Cullin 1-F-box protein (SCF) E3 ubiquitin ligase that targets key regulators of cellular activities for ubiquitylation and degradation, plays important roles in the progression of human cancer. However, little is known about the role of FBXO22 in renal cell carcinoma (RCC). This study aims to explore the biological function of FBXO22 in RCC progression and its specific regulation mechanism. We performed immunohistochemistry analysis and found that the expression level of FBXO22 was significantly lower in RCC tissues than in normal renal tissues. Reduced FBXO22 expression in RCC tissues is related to tumor size and TNM stage and to worse overall and disease-free survival. Through an in vitro assay, we demonstrated that FBXO22 has no effect on renal cancer cells proliferation, whereas FBXO22 remarkably restricted RCC cell migration and invasion, thereby reversing EMT transition and elevating the activity of tissue inhibitor of matrix metalloproteinase-1, which subsequently inhibited metalloproteinase-9 (MMP-9) expression and activity in vitro. We also found that FBXO22 suppresses tube formation by disrupting the secretion of vascular endothelial growth factor. Meanwhile, in vivo studies verified that FBXO22 suppresses RCC metastasis. These findings suggested that FBXO22 is a novel prognostic indicator and plays an important role in RCC metastasis.

Inhibition of SMYD2 suppresses tumor progression by down-regulating microRNA-125b and attenuates multi-drug resistance in renal cell carcinoma

SMYD2 is a histone methyltransferase that has been reported to be an important epigenetic regulator. This study aims to investigate SMYD2 as a prognostic indicator of clear cell renal cell carcinoma (ccRCC) and explore its role in tumorigenesis and multi-drug resistance.Methods: Tumor specimens, clinicopathologic information, and prognostic outcomes of 186 ccRCC patients from three hospitals in China were collected for SMYD2 immunohistochemistry staining, Kaplan-Meier analysis, and Cox proportional hazards-regression analysis. MicroRNA (miRNA)-microarray profiling identified differentially expressed miRNAs in renal cancer cells subjected to SMYD2 knockdown or treatment with the SMYD2 inhibitor AZ505. The effects of SMYD2 and candidate SMYD2-mediated miRNAs on renal cancer cell proliferation, migration, clonogenicity, and tumorigenicity were determined via cell-function assays and murine xenograft experiments. The half-inhibitory concentrations (IC50) of five antineoplastic drugs (cisplatin, doxorubicin, fluorouracil, docetaxel, and sunitinib) in AZ505-treated and control cells were calculated, and the effects of SMYD2 inhibition on P-glycoprotein (P-gP) expression and multiple-drug resistance were verified.Results: SMYD2 was overexpressed and acted as an oncogene in ccRCC. High SMYD2 expression correlated with a high TNM stage (P = 0.007) and early tumor relapse (P = 0.032). SMYD2 independently predicted a worse overall survival (P = 0.022) and disease-free survival (P = 0.048). AZ505 inhibited the binding of SMYD2 to the miR-125b promoter region (based on chromatin immunoprecipitation assays) and suppressed ccRCC cell migration and invasion by inhibiting the SMYD2/miR-125b/DKK3 pathway. SMYD2 and miR-125b inhibition acted synergistically with anticancer drugs via P-gP suppression in vitro and in vivo.Conclusions: These findings suggested that SMYD2 plays an important role in ccRCC development and could be a potential biomarker for the treatment and prognosis of RCC.

Long-Term Cell Fate Tracking of Individual Renal Cells Using Serial Intravital Microscopy.

Intravital multiphoton microscopy of the kidney is a powerful technique to study alterations in tissue morphology and function simultaneously in the living animal and represents a dynamic and developing research tool in the field. Recent technological advances include serial intravital multiphoton microscopy of the same kidney regions over several weeks and combined with ex vivo histology for cellular biomarker expression of the same cells, which had been subject to serial imaging before. Thus, serial intravital multiphoton microscopy followed by ex vivo histology provides unique tools to perform long-term cell fate tracing of the same renal cells during physiological and pathophysiological conditions, thereby allowing the detection of structural changes of the same renal cells over time. Examples include renal cell migration and proliferation while linking these events to local functional alterations and eventually to the expression of distinct cellular biomarkers. Here, we provide a detailed step-by-step protocol to facilitate serial intravital multiphoton microscopy for long-term in vivo tracking of renal cells and subsequent ex vivo histology for immunohistological staining of the same cells in the fixed tissue.

Activation of AMPK by metformin promotes renal cancer cell proliferation under glucose deprivation through its interaction with PKM2.

Metformin, a common therapeutics for type 2 diabetics, was recently demonstrated to possess antitumor activity in various cancer types. However, its therapy effect in renal cell carcinoma (RCC) still remains controversial. In this study, we found that metformin treatment in RCC cells lead to activation of AMPK, which suppressed the cell proliferation under normal condition, but enhanced cell proliferation under glucose deprivation (GD) condition. Depletion of AMPK by siRNA abolished the proliferation effect of MF under GD condition. Mechanistic investigations revealed that the effect of AMPK on cell proliferation under GD condition is dependent on its nuclear translocation. Moreover, the nuclear AMPK recruits PKM2 and β-Catenin to form a complex, which promotes the transcription of cell proliferation related genes, including CCND1 and c-Myc. Furthermore, depletion of PKM2 or β-Catenin abrogated the proliferative effects of metformin under GD condition. And inhibition of PKM2 also re-sensitized the A498 xenograft in response to metformin treatment. Together, our results suggested that combined of AMPK activation and PKM2 depletion or inhibition can achieve better therapeutic effect for RCC patients.

Role of miR-486-5p in regulating renal cell carcinoma cell proliferation and apoptosis via TGF-β-activated kinase 1.

Renal cell carcinoma (RCC) is a common kidney tumor in adults. The role of miR-486-5p in RCC is unknown. The aim of our study was to identify new targets regulated by miR-486-5p in RCC, to obtain a deeper insight into the network and to better understand the role of these microRNAs and their targets in carcinogenesis of RCC. We performed a series of tests and found consistently lower expression levels of miR-486-5p in kidney cancer cells. Restoration of miR-486-5p expression in RCC cells could lead to the suppression of cell proliferation and the increase of cell apoptosis. Further studies demonstrated that TGF-β-activated kinase 1 was a target gene of miR-486-5p in kidney cancer cells. It was also shown that C-C motif chemokine ligand 2(CCL2) from tumor-associated macrophages downregulated miR-486-5p expression, and miR-486-5p inhibited RCC cell proliferation and apoptosis resistance induced by CCL2. The study demonstrates that there are potential diagnosis and therapy values of miR-486-5p in RCC.

CircPCNXL2 sponges miR-153 to promote the proliferation and invasion of renal cancer cells through upregulating ZEB2

ABSTRACTIncreasing evidence showed that circular RNAs (circRNAs) play critical roles in tumorigenesis. However, the roles and underlying mechanisms of circRNAs in clear cell renal cell carcinoma (ccRCC) remain unclear. In the present study, we identified a novel circRNA circPCNXL2, which was significantly upregulated in ccRCC by circular RNA microarray. Further analysis revealed that circPCNXL2 was significantly increased and correlated with poor overall survival of ccRCC patients. Function assays revealed that circPCNXL2 knockdown reduced RCC cells proliferation, invasion in vitro, and decreased tumor growth in vivo. In mechanism study, we showed that circPCNXL2 could be bind to miR-153 as a miRNA sponge to regulate ZEB2 expression in RCC progression. In addition, our data reported that the effects of circPCNXL2 inhibition on RCC cells proliferation and invasion could be abolished by miR-153 inhibitors. Altogether, we demonstrated that circPCNXL2 could regulate RCC cells proliferation and invasion by miR-153/ZEB2 axis, suggesting circPCNXL2 might serve as a potential therapeutic target for ccRCC treatment.

Low expression of PDK1 inhibits renal cell carcinoma cell proliferation, migration, invasion and epithelial mesenchymal transition through inhibition of the PI3K-PDK1-Akt pathway

As the most commonly occurring form of primary renal tumor, renal cell carcinoma (RCC) is a malignancy accompanied by a high mortality rate. 3-phosphoinositide-dependent protein kinase 1 (PDK1) has been established as a protein target and generated considerable interest in both the pharmaceutical and academia industry. The aim of the current study was to investigate the effect of si-PDK1 on the RCC cell apoptosis, proliferation, migration, invasion and epithelial mesenchymal transition (EMT) in connection with the PI3K-PDK1-Akt pathway. Microarray analysis from the GEO database was adopted to identify differentially expressed genes (DEGs) related to RCC, after which the positive expression of the PDK1 protein in tissue was determined accordingly. The optimal silencing si-RNA was subsequently selected and RCC cell lines 786-O and A498 were selected and transfected with either a si-PDK1 or activator of the PI3K-PDK1-Akt pathway for grouping purposes. The mRNA and protein expressions of PDK1, the PI3K-PDK1-Akt pathway-, EMT- and apoptosis-related genes were then evaluated. The effect of si-PDK1 on cell proliferation, apoptosis, invasion and migration was then analyzed. Through microarray analysis of GSE6344, GSE53757, GSE14762 and GSE781, PDK1 was examined. PDK1 was determined to be highly expressed in RCC tissues. Si-PDK1 exhibited marked reductions in relation to the mRNA and protein expression of PDK1, PI3K, AKT as well as Vimentin while elevated mRNA and protein expressions of E-cadherin were detected, which ultimately suggested that cell migration, proliferation and invasion had been inhibited coupled with enhanced levels of cell apoptosis. While a notable observation was made highlighting that the PI3K-PDK1-Akt pathway antagonized the effect of PDK1 silencing. Taken together, the key observations of this study provide evidence suggesting that high expressions of PDK1 are found in RCC, while highlighting that silencing PDK1 could inhibit RCC cell proliferation, migration, invasion and EMT by repressing the PI3K-PDK1-Akt pathway.

SCUBE1-enhanced bone morphogenetic protein signaling protects against renal ischemia-reperfusion injury

We previously reported that the membrane-bound SCUBE1 (signal peptide-CUB-epithelial growth factor domain-containing protein 1) forms a complex with bone morphogenetic protein 2 (BMP2) ligand and its receptors, thus acting as a BMP co-receptor to augment BMP signal activity. However, whether SCUBE1 can bind to and facilitate signaling activity of BMP7, a renal protective molecule for ischemia-reperfusion (I/R) insult, and contribute to the proliferation and repair of renal tubular cells after I/R remains largely unknown. In this study, we first showed that I/R-induced SCUBE1 was expressed in proximal tubular cells, which coincided with the expression of renoprotective BMP7. Molecular and biochemical analyses revealed that SCUBE1 directly binds to BMP7 and its receptors, functioning as a BMP co-receptor to promote BMP7 signaling. Furthermore, we used a new Scube1 deletion (Δ2) mouse strain to further elucidate the renal pathophysiological function of SCUBE1 after I/R injury. As compared with wild-type littermates, Δ2 mice showed severe renal histopathologic features (extensive loss of brush border, tubular necrosis, and tubular dilation) and increased inflammation (neutrophil infiltrate and induction of monocyte chemoattractant protein-1, tumor necrosis factor-α and interleukin-6) during the resolution of I/R damage. They also showed reduced BMP signaling (phosphorylated Smad1/5/8) along with decreased proliferation and increased apoptosis of renal tubular cells. Importantly, lentivirus-mediated overexpression of SCUBE1 enhanced BMP signaling and conferred a concomitant survival outcome for Δ2 proximal tubular epithelial cells after hypoxia–reoxygenation treatment. The protective BMP7 signaling may be facilitated by stress-inducible SCUBE1 after renal I/R, which suggests potential targeted approaches for acute kidney injury.

Identifying cystogenic paracrine signaling molecules in cyst fluid of patients with polycystic kidney disease.

In autosomal dominant polycystic kidney disease (ADPKD) paracrine signaling molecules in cyst fluid can induce proliferation and cystogenesis of neighboring renal epithelial cells. However, the identity of this cyst-inducing factor is still unknown. The aim of this study was to identify paracrine signaling proteins in cyst fluid using a 3D in vitro cystogenesis assay. We collected cyst fluid from 15 ADPKD patients who underwent kidney or liver resection (55 cysts from 13 nephrectomies, 5 cysts from 2 liver resections). For each sample, the ability to induce proliferation and cyst formation was tested using the cystogenesis assay (RPTEC/TERT1 cells in Matrigel with cyst fluid added for 14 days). Kidney cyst fluid induced proliferation and cyst growth of renal epithelial cells in a dose-dependent fashion. Liver cyst fluid also induced cystogenesis. Using size exclusion chromatography, 56 cyst fluid fractions were obtained of which only the fractions between 30 and 100 kDa showed cystogenic potential. Mass spectrometry analysis of samples that tested positive or negative in the assay identified 43 candidate cystogenic proteins. Gene ontology analysis showed an enrichment for proteins classified as enzymes, immunity proteins, receptors, and signaling proteins. A number of these proteins have previously been implicated in ADPKD, including secreted frizzled-related protein 4, S100A8, osteopontin, and cysteine rich with EGF-like domains 1. In conclusion, both kidney and liver cyst fluids contain paracrine signaling molecules that drive cyst formation. Using size exclusion chromatography and mass spectrometry, we procured a candidate list for future studies. Ultimately, cystogenic paracrine signaling molecules may be targeted to abrogate cystogenesis in ADPKD.

MiR-384 suppressed renal cell carcinoma cell proliferation and migration through targeting RAB23.

microRNAs (miRNAs) are noncoding, short, and endogenous RNAs that play crucial roles in tumor progression at the post-transcriptional level. Here, we studied the role of miR-384 in the pathogenesis of renal cell carcinoma (RCC). We demonstrated that miR-384 expression was downregulated in the RCC specimens compared with nontumor specimens. Moreover, we showed that RAB23 expression was upregulated in the RCC tissues compared with nontumor tissues. Furthermore, we demonstrated that low expression of miR-384 was correlated with high levels of RAB23 in RCC tissues. We also demonstrated that the RAB23 was a direct target gene of miR-384 in RCC cells. In addition, overexpression of miR-384 suppressedRCC cell proliferation, cell cycle, and cell migration. Furthermore, ectopic expression of RAB23 promotedRCC cell proliferation, cell cycle, and cell migration. These data suggested that miR-384 played a tumor suppressor microRNA in the development of RCC partly through inhibiting RAB23 expression.

Long noncoding RNA TUG1 promotes renal cell carcinoma cell proliferation, migration and invasion by downregulating microRNA‑196a.

Long noncoding RNA taurine upregulated gene1 (lncRNA TUG1) and microRNA‑196a (miR‑196a) have been reported to serve important roles in the development of renal cell carcinoma (RCC). However, their potential mechanisms have not been completely elucidated. The aim of the present study was to clarify the biological functions of lncRNA‑TUG1 and miR‑196a, in addition to investigating the interaction between lncRNA‑TUG1 and microRNA‑196a, providing a novel insight into RCC tumorigenesis. The present study comprised two parts. In the first part, lncRNA‑TUG1 was confirmed as an oncogene, via reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) analysis, MTT assay, flow cytometry analysis, and migration and invasion assays. In the second part, the association between lncRNA‑TUG1 and miR‑196a, and the molecular mechanism, was illustrated via RT‑qPCR analysis, MTT assay, dual luciferase reporter assay and western blotting. The results of the present study demonstrated that lncRNA‑TUG1 was able to promote RCC cell proliferation, migration and invasion invitro by suppressing miR‑196a. Additionally, lncRNA‑TUG1 achieved its biological functions by regulating the expression levels of RAC‑α serine/threonine‑protein kinase, mitogen‑activated protein kinase and extracellular signal‑regulated kinase via inhibition of miR‑196a. In conclusion, the present findings proposed a novel potential therapeutic target, the lncRNA‑TUG1‑miR‑196a axis, which may be applicable to the treatment of RCC.

Effects of long non-coding RNA LINC00667 on renal tubular epithelial cell proliferation, apoptosis and renal fibrosis via the miR-19b-3p/LINC00667/CTGF signaling pathway in chronic renal failure

The global prevalence of chronic renal failure (CRF) has significantly elevated with various reports indicating there to be a 10% worldwide rate. The functions of long non-coding RNAs (lncRNAs) and their deeper association with CRF at present remain poorly understood. Hence, the aim of the present study was to investigate the altered expressions of lncRNA LINC00667 in CRF and its associated effects on renal tubular epithelial cell proliferation, apoptosis and renal fibrosis through the microRNA-19b-3p (miR-19b-3p)/LINC00667/connective tissue growth factor (CTGF) signaling pathway. Initially, verification of the targeting relationship between LINC00667, CTGF and miR-19b-3p was performed, after which evidence was obtained indicating that miR-19b-3p could negatively regulate LINC00667 and CTGF. The expressions of CTGF in both the CRF and normal renal tissues were determined by immunohistochemistry means, with LINC00667 and CTGF determined to be highly expressed, while poor expression levels of miR-19b-3p were detected among the CRF tissues. The expressions of LINC00667, miR-19b-3p, fibrosis- and epithelial-mesenchymal transition (EMT)-related genes were also examined. The successfully established CRF rat models were treated with varying mimics, inhibitors, and siRNA. ELISA was applied to determine the renal function-related factors. Besides, the renal cell proliferation, migration and apoptosis were detected. In response to LINC00667 silencing, the renal tubular epithelial cells displayed increased proliferation and migration accompanied by reduced apoptosis based on upregulated miR-19b-3p, along with inhibited renal fibrosis and EMT detected. Taken together, the key findings of our study demonstrated that decreased lncRNA LINC00667 could promote renal tubular epithelial cell proliferation and ameliorate renal fibrosis in CRF via the miR-19b-3p/LINC00667/CTGF signaling pathway.

Downregulation of long noncoding RNA FENDRR predicts poor prognosis in renal cell carcinoma.

Long noncoding RNA FOXF1 adjacent non-coding developmental regulatory RNA (FENDRR) dysregulation associates with multiple types of human cancer. However, the biological functions of FENDRR in renal cell carcinoma are unresolved. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was performed to determine the expression level of FENDRR in renal cell carcinoma tissues. An RNA interference assay and ectopic expression experiments were conducted to evaluate the effects of FENDRR on cell proliferation, migration, invasion and colony formation in vitro. RNA immunoprecipitation was conducted to identify proteins associated with FENDRR. It was observed that FENDRR is frequently downregulated in renal cell carcinoma and overexpression of FENDRR attenuated proliferation, migration, invasion and colony growth of renal carcinoma cells. Conversely, knockdown of FENDRR promotes proliferation and invasiveness of renal carcinoma cells. Downregulation of FENDRR associates with poor prognosis of renal cell carcinoma. Mechanistically, it was identified that FENDRR may bind to Polycomb Repressive Complex 2 and lysin methyltransferase 2A histone modifying complexes. In summary, FENDRR acts as an tumor suppressor in renal cell carcinoma and may serve as a candidate target for gene therapy.