OBJECTIVE: To explore the role of miR-744-5p/CCND1 axis in clear-cell renal cell carcinoma (ccRCC). METHODS: We examined the expression levels of miR-744-5p in 65 pairs of ccRCC and adjacent tissue specimens and in 5 ccRCC cell lines and human renal tubular epithelial (HK2) cells using qRT-PCR. The ccRCC cell lines 786-O and OSRC2 were transfected with miR-744-5p mimic, CCND1 mimic, or their negative control mimics, and the changes in cell proliferation, migration, and invasion were evaluated with CCK-8, wound healing, and Transwell assays. The downstream target molecules of miR-744-5p were predicted by bioinformatics analysis, and the expression level of CCND1 in ccRCC cells was verified by qRT-PCR and Western blotting. The relationship between miR-744-5p and CCND1 was further validated by dual luciferase reporter assay, and the role of the miR-744-5p/CCND1 axis in ccRCC was explored by rescue experiments. RESULTS: MiR-744-5p was significantly downregulated in ccRCC tissues and cell lines (all P < 0.05), and its overexpression inhibited the proliferation, migration, and invasion of ccRCC cells (all P < 0.05). Bioinformatics analysis and dual luciferase reporter assay showed that CCND1 was a downstream target of miR-744-5p. The results of rescue experiments showed that upregulation of CCND1 could partially reverse the inhibitory effect of miR-744-5p overexpression on ccRCC cell proliferation, migration, and invasion (all P < 0.05). CONCLUSION MiR-744-5p inhibits the malignant phenotype of ccRCC cells by targeting CCND1, and the miR-744-5p/CCND1 axis may be a novel target for diagnosis and treatment of ccRCC.
Carcinoma, Renal Cell/metabolism* ; Cell Line, Tumor ; Cell Movement/genetics* ; Cell Proliferation/genetics* ; Cyclin D1/genetics* ; Humans ; Kidney Neoplasms/metabolism* ; MicroRNAs/metabolism*

Adenocarcinoma, Clear Cell ; metabolism ; pathology ; surgery ; Adult ; Carcinoma ; metabolism ; pathology ; Carcinoma, Mucoepidermoid ; metabolism ; pathology ; Carcinoma, Renal Cell ; metabolism ; pathology ; secondary ; Diagnosis, Differential ; Humans ; Keratins ; metabolism ; Male ; Nasal Cavity ; Nose Neoplasms ; metabolism ; pathology ; surgery ; S100 Proteins ; metabolism

PURPOSE: Both insulin and insulin-like growth factor (IGF)-1 signaling are key regulators of energy metabolism, cellular growth, proliferation, and survival. The IGF-1 receptor (IGF-1R) is overexpressed in most types of human cancers including renal cell carcinoma (RCC) with poor prognosis. Insulin receptor (IR) shares downstream effectors with IGF-1R; however, the expression and function of IR in the tumorigenesis of renal cancer remains elusive. Therefore, we examined the expression of IR and its prognostic significance in clear cell RCC (CCRCC). MATERIALS AND METHODS: Immunohistochemical staining for IR was performed on 126 formalin-fixed paraffin-embedded CCRCC tissue samples. Eight of these cases were utilized for western blot analysis. The results were compared with various clinico-pathologic parameters of CCRCC and patient survival. RESULTS: IR was expressed in the nuclei of CCRCC tumor cells in 109 cases (87.9%). Higher IR expression was significantly correlated with the presence of cystic change, lower Fuhrman nuclear grade, lower pathologic T stage, and lower TNM stage, although it wasn't significantly related to diabetes status and patient survival. Western blot analyses supported the results of the immunohistochemistry studies. CONCLUSION: IR expression in CCRCC may be associated with favorable prognostic factors.
Aged ; Blotting, Western ; Carcinoma, Renal Cell/*metabolism/*pathology ; Female ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Prognosis ; Receptor, Insulin/*metabolism

PURPOSE: Both insulin and insulin-like growth factor (IGF)-1 signaling are key regulators of energy metabolism, cellular growth, proliferation, and survival. The IGF-1 receptor (IGF-1R) is overexpressed in most types of human cancers including renal cell carcinoma (RCC) with poor prognosis. Insulin receptor (IR) shares downstream effectors with IGF-1R; however, the expression and function of IR in the tumorigenesis of renal cancer remains elusive. Therefore, we examined the expression of IR and its prognostic significance in clear cell RCC (CCRCC). MATERIALS AND METHODS: Immunohistochemical staining for IR was performed on 126 formalin-fixed paraffin-embedded CCRCC tissue samples. Eight of these cases were utilized for western blot analysis. The results were compared with various clinico-pathologic parameters of CCRCC and patient survival. RESULTS: IR was expressed in the nuclei of CCRCC tumor cells in 109 cases (87.9%). Higher IR expression was significantly correlated with the presence of cystic change, lower Fuhrman nuclear grade, lower pathologic T stage, and lower TNM stage, although it wasn't significantly related to diabetes status and patient survival. Western blot analyses supported the results of the immunohistochemistry studies. CONCLUSION: IR expression in CCRCC may be associated with favorable prognostic factors.
Aged ; Blotting, Western ; Carcinoma, Renal Cell/*metabolism/*pathology ; Female ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Prognosis ; Receptor, Insulin/*metabolism

The aim of this study was to investigate the relationship of cyclooxygenase (COX) -2 and p53 expression with prognosis in patients with conventional renal cell carcinoma (RCC). Formalin-fixed, paraffin-embedded tissue sections of conventional RCC from 92 patients, who had undergone radical nephrectomy, were examined for COX-2 and p53 expression by immunohistochemistry and compared with clinicopathological variables. The COX-2 expression significantly correlated only with tumor size (p=0.049), whereas the p53 expression profoundly correlated with the TNM stage (p=0.024), M stage (p=0.001), and metastasis (synchronous or metachronous; p= 0.004). The COX-2 overexpression did not significantly associate with p53 positivity (p=0.821). The survival rate of patients correlated with the p53 expression (p < 0.0001) but not with the COX-2 expression (p=0.7506). Multivariate analyses indicated that tumor size, M stage, and p53 expression were independent prognostic factors for cancer-specific survival. The COX-2 expression was not an independent factor. These results show that the increased expression of p53 was associated with metastasis and a worse prognosis in conventional RCC, which suggests that p53 might have played an important role in the progression of conventional RCC. The increased expression of COX-2 was associated only with tumor size, but may not be an important prognostic factor in conventional RCC. No association was observed between COX-2 overexpression and p53 positivity in conventional RCC.
Carcinoma, Renal Cell/*metabolism/mortality/pathology ; Humans ; Kidney Neoplasms/*metabolism/mortality/pathology ; Prognosis ; Prostaglandin-Endoperoxide Synthase/*metabolism ; Protein p53/*metabolism ; Tumor Markers, Biological/*metabolism

Adenocarcinoma, Follicular ; Adult ; Carcinoid Tumor ; metabolism ; pathology ; Carcinoma, Renal Cell ; metabolism ; pathology ; surgery ; Diagnosis, Differential ; Female ; Humans ; Keratin-7 ; metabolism ; Kidney Neoplasms ; metabolism ; pathology ; surgery ; Mucin-1 ; metabolism

OBJECTIVE: To explore the potential mechanism of resistance to axitinib in clear cell renal cell carcinoma (ccRCC), with a view to expanding the understanding of axitinib resistance, facilitating the design of more specific treatment options, and improving the treatment effectiveness and survival prognosis of patients. METHODS: By exploring the half maximum inhibitory concentration (IC₅₀) of axitinib on ccRCC cell lines 786-O and Caki-1, cell lines resistant to axitinib were constructed by repeatedly stimulated with axitinib at this concentration for 30 cycles in vitro. Cell lines that were not treated by axitinib were sensitive cell lines. The phenotypic differences of cell proliferation and apoptosis levels between drug resistant and sensitive lines were tested. Genes that might be involved in the drug resistance process were screened from the differentially expressed genes that were co-upregulated in the two drug resistant lines by transcriptome sequencing. The expression level of the target gene in the drug resistant lines was verified by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot (WB). The expression differences of the target gene in ccRCC tumor tissues and adjacent tissues were analyzed in the Gene Expression Profiling Interactive Analysis (GEPIA) public database, and the impact of the target gene on the prognosis of ccRCC patients was analyzed in the Kaplan-Meier Plotter (K-M Plotter) database. After knocking down the target gene in the drug resistant lines using RNA interference by lentivirus vector, the phenotypic differences of the cell lines were tested again. WB was used to detect the levels of apoptosis-related proteins in the different treated cell lines to find molecular pathways that might lead to drug resistance. RESULTS: Cell lines 786-O-R and Caki-1-R resistant to axitinib were successfully constructed in vitro, and their IC₅₀ were significantly higher than those of the sensitive cell lines (10.99 μmol/L, P < 0.01; 11.96 μmol/L, P < 0.01, respectively). Cell counting kit-8 (CCK-8) assay, colony formation, and 5-ethynyl-2 '-deoxyuridine (EdU) assay showed that compared with the sensitive lines, the proliferative ability of the resistant lines decreased, but apoptosis staining showed a significant decrease in the level of cell apoptosis of the resistant lines (P < 0.01). Although resistant to axitinib, the resistant lines had no obvious new replicated cells in the environment of 20 μmol/L axitinib. Nuclear protein 1 (NUPR1) gene was screened by transcriptome sequencing, and its RNA (P < 0.0001) and protein expression levels significantly increased in the resistant lines. Database analysis showed that NUPR1 was significantly overexpressed in ccRCC tumor tissue (P < 0.05); the ccRCC patients with higher expression ofNUPR1had a worse survival prognosis (P < 0.001). Apoptosis staining results showed that knockdown ofNUPR1inhibited the anti-apoptotic ability of the resistant lines to axitinib (786-O, P < 0.01; Caki-1, P < 0.05). WB results showed that knocking downNUPR1decreased the protein level of B-cell lymphoma-2 (BCL2), increased the protein level of BCL2-associated X protein (BAX), decreased the protein level of pro-caspase3, and increased the level of cleaved-caspase3 in the resistant lines after being treated with axitinib. CONCLUSION ccRCC cell lines reduce apoptosis through theNUPR1 -BAX/ BCL2 -caspase3 pathway, which is involved in the process of resistance to axitinib.
Humans ; Carcinoma, Renal Cell/metabolism* ; Axitinib/pharmacology* ; Kidney Neoplasms/metabolism* ; bcl-2-Associated X Protein ; Nuclear Proteins ; Cell Line, Tumor ; Apoptosis ; Cell Proliferation

OBJECTIVETo investigate the optimal margin in nephron-sparing surgery (NSS) for renal cell carcinoma (RCC) 4 cm or less in diameter.

METHODSEighty-two kidneys with RCC 4 cm or less in diameter resected by radical nephrectomy were prospectively studied. The kidney samples were sectioned at 3 mm interval and examined for multicentricity. On each layer of tissue sectioned, parenchyma margin of 15 mm beyond pseudocapsule was continuously sectioned and examined for completeness of pseudocapsule and extra-pseudocapsule cancer lesion. The farthest distance between extra-pseudocapsule lesion and primary tumor was measured. PCNA expression was detected in 41 patients by using standard SP immunohistochemistry technique.

RESULTSThe diameter of 82 primary tumors was 3.4 +/- 0.8 cm (range 1.5 - 4.0 cm). Of these, 31.7% (26/82) were found without intact pseudocapsule and 17.1% (14/82) with positive cancer lesions beyond pseudocapsule. The average distance between extra-pseudocapsule cancer lesion and primary tumor was 0.5 +/- 1.3 mm (range 0 - 5.0 mm), with a confidential interval (CI) of 95% (0.11, 0.94). Statistically, the one side percentile P(95) was 4.9 mm, P(97.5) was 5.0 mm and P(100) was 5.0 mm. The mean PCNA index in the 41 patients with RCC was (29.5 +/- 17.6)%, which was (49.6 +/- 21.5)% in the group with extra-pseudocapsule cancer lesions and (24.6 +/- 12.7)% in the group without (t = 3.162, P = 0.013). The ratio of strong expression was 5/8 in the group with extra-pseudocapsule cancer lesions, and 18.2% (6/33) in the group without the lesions (chi(2) = 6.442, P = 0.011). Logistic regression analysis showed that completeness of pseudocapsule and PCNA index were significant predictors of extra-pseudocapsule cancer lesions (P = 0.019).

CONCLUSIONSThese data suggest that when NSS is performed in RCC 4 cm or less in diameter, a margin of more than 5 mm of adjacent parenchyma should be excised with the tumor. Enucleation alone was associated with a significant risk of incomplete excision, and therefore liable for local recurrence. Tumors with incomplete pseudocapsule and(or) high PCNA indices are more likely to have extra-pseudocapsule cancer lesions, so intensive follow-up is necessary after NSS.

Adult ; Carcinoma, Renal Cell ; metabolism ; pathology ; surgery ; Female ; Humans ; Kidney Neoplasms ; metabolism ; pathology ; surgery ; Male ; Middle Aged ; Nephrectomy ; methods ; Proliferating Cell Nuclear Antigen ; metabolism ; Retrospective Studies

OBJECTIVETo detect the expression of apoptosis gene PDCD5 in tissues of normal human kidney, renal clear cell carcinoma, normal bladder and bladder carcinoma, and explore the role of PDCD5 gene in renal clear cell carcinoma and bladder carcinoma.

METHODSIndirect immunohistochemistry was employed to detect PDCD5 expression in 63 kidney specimens and 42 bladder specimens. Positive expression rates and intensity of PDCD5 protein expression in the kidney tissue were investigated microscopically and by computerized image analysis. Positive expression rate in the bladder tissue was investigated by microscopic observation.

RESULTSThe results of immunohistochemical staining showed PDCD5 protein overexpression in the renal tubule of normal human kidney tissues and downregulation with the stage increase of renal clear cell carcinoma. PDCD5 protein expression showed statistical significance in tissues of normal kidney and renal clear cell carcinoma in all stages. No obvious PDCD5 expression was detected in the tissues of normal human bladder and bladder carcinoma.

CONCLUSIONPDCD5 is an important apoptosis-regulating factor in the occurrence of renal clear cell carcinoma, and its expression is extremely low in tissues of normal human bladder and bladder carcinoma.

Adult ; Aged ; Apoptosis Regulatory Proteins ; biosynthesis ; Carcinoma, Renal Cell ; metabolism ; Carcinoma, Transitional Cell ; metabolism ; Female ; Humans ; Immunohistochemistry ; Kidney ; metabolism ; Kidney Neoplasms ; metabolism ; Male ; Middle Aged ; Neoplasm Proteins ; biosynthesis ; Urinary Bladder ; metabolism ; Urinary Bladder Neoplasms ; metabolism

Angiogenesis is a series of processes that include endothelial proliferation, migration and tube formation. Vascular endothelial growth factor (VEGF) is regarded as a potent mediator of angiogenesis, vascular permeability and tumor cell growth in renal cell carcinoma. This study was designed to evaluate the expression of VEGF and the microvessel count (MVC) and to determine their prediction efficacies for prognosis in renal cell carcinoma. The relationship between the expression of VEGF and MVC were evaluated immunohistochemically in 50 patients with renal cell carcinoma who received a radical nephrectomy at Wonju Christian Hospital between 1989 and 1997. Microvessels were identified by immunostaining endothelial cells for CD-31 antigen. The mean follow-up was 96 months (3 - 133 months). Overall 5-year survival rate was 71.5%. VEGF was expressed in the tumor cell cytoplasm. Of the 50 tumors, 23 (46%) were weak to strongly positive for VEGF but 27 (54%) were unreactive. The respective 5-year survival rates for patients with positive and negative expressions of VEGF were 70% and 73% (p > 0.05). The overall mean MVC was 13.4 in a 400x field. Mean MVCs were significantly higher in VEGF-positive tumors (17.6 +/- 12.1) than in VEGF-negative tumors (9.9 +/- 5.4), and the MVCs of the high vascular density group and the low ascular density groups were significantly different. The 5-year survival rates of patients with high vascular density and low vascular density were 59% and 86%. The median survival period for patients with MVCs higher than or equal to 10 vessels/field was 85 months, whereas for those with MVCs lower than 10 vessels/field the median survival time was 102 months. These results suggest that MVC may be a better prognostic factor in renal cell carcinoma than the expression of VEGF.
Adult ; Aged ; Carcinoma, Renal Cell/*blood supply/*metabolism ; Endothelial Growth Factors/*metabolism ; Female ; Human ; Kidney Neoplasms/*blood supply/*metabolism ; Lymphokines/*metabolism ; Male ; Middle Age ; Neovascularization, Pathologic/*pathology ; Prognosis