[Correlation of oxidative stress with sperm DNA integrity and semen parameters in infertile men with varicocele].

OBJECTIVE: To investigate the relationship of oxidative stress with DNA integrity and semen parameters in infertile men with varicocele (VC). METHODS: This prospective study included 98 infertile males with VC. According to the levels of reactive oxygen species (ROS) in the semen, we divided the patients into a high ROS group (n=44) and a low ROS group (n=54), determined the sperm DNA fragmentation index (DFI), motility and morphology, and analyzed their correlation with ROS in the two groups of patients. RESULTS: Compared with the patients of the low ROS group, those of the high ROS group showed a significantly higher DFI (27.38±8.10 vs 34.49±6.05, P=0.039) and a higher concentration of seminal leukocytes (ï¼»0.65±0.15ï¼½×106/ml vs ï¼»0.86±0.41ï¼½×106/ml, P=0.022), but lower sperm motility (ï¼»36.16±22.83ï¼½% vs ï¼»18.22±25.21ï¼½%, P=0.017), percentage of progressively motile sperm (ï¼»23.34±11.53ï¼½% vs ï¼»16.34±9.22ï¼½%, P=0.041), sperm curvilinear velocity (ï¼»27.03±6.21ï¼½ vs ï¼»20.62±4.38ï¼½ µm/s, P=0.013), and sperm linearity (ï¼»29.75±8.24ï¼½% vs ï¼»18.30±7.93ï¼½%, P=0.024). Spearman correlation analysis indicated that the ROS level was correlated positively with the concentration of seminal leukocytes (r=0.41, P<0.01) and DFI (r=0.21, P=0.006), but negatively with sperm curvilinear velocity (r=－0.24, P=0.017), linearity (r=－0.24, P=0.021), motility (r=－0.31, P=0.002), and the percentage of progressively motile sperm (r=－0.41, P=0.012). Additionally, the sperm DFI manifested a significant negative correlation with sperm motility (r=－0.29, P<0.01) and the percentage of progressively motile sperm (r=－0.34, P<0.01). CONCLUSIONS: The level of seminal ROS is positively correlated with the sperm DFI in infertile men with varicocele, and both the ROS level and DNA integrity are associated with semen parameters.

MicroRNA-224 inhibits progression of human prostate cancer by downregulating TRIB1.

Our previous microarray data showed that microRNA-224 (miR-224) was downregulated in human prostate cancer (PCa) tissues compared with adjacent benign tissues. However, the underlying mechanisms by which miR-224 is involved in PCa remain unclear. In this study, we identified TRIB1 as a target gene of miR-224. Forced expression of miR-224 suppressed PCa cell proliferation, invasion and migration, and promoted cell apoptosis by downregulating TRIB1. Moreover, the expression level of miR-224 in PCa tissues was negatively correlated with that of TRIB1. miR-224 downregulation was frequently found in PCa tissues with metastasis, higher PSA level and clinical stage, whereas TRIB1 upregulation was significantly associated with metastasis. Both miR-224 downregulation and TRIB1 upregulation were significantly associated with poor biochemical recurrence-free survival of patients with PCa. In conclusion, these findings reveal that the aberrant expression of miR-224 and TRIB1 may promote PCa progression and have potentials to serve as novel biomarkers for PCa prognosis.

MicroRNA-30c serves as an independent biochemical recurrence predictor and potential tumor suppressor for prostate cancer.

MicroRNA-30c (miR-30c) acts as a tumor suppressor or a tumor promoter in various human malignancies. However, the involvement of miR-30c in prostate cancer (PCa) is still unclear. The aim of this study was to investigate the molecular function and the clinical significance of miR-30c in PCa. Expression levels of miR-30c in PCa tissues and cells were detected by quantitative real-time-PCR (qRT-PCR). Additionally, the associations of miR-30c expression with clinicopathological features and prognosis in PCa patients were analyzed. The potential role of miR-30c in tumorigenesis of PCa cells was further evaluated by in vitro cell assays. MiR-30c was significantly down-regulated in PCa tissues and cells compared with the corresponding controls (P<0.05). In addition, the downregulation of miR-30c in PCa tissues was significantly associated with higher Gleason score (P=0.009), advanced pathological stage (P=0.016) and biochemical recurrence (P=0.034). Moreover, Kaplan-Meier survival analysis showed that the reduced expression of miR-30c was correlated with shorter biochemical recurrence-free survival (P=0.023). The multivariate analysis also identified miR-30c as an independent prognostic predictor for biochemical recurrence-free survival in patients with PCa. Furthermore, the enforced expression of miR-30c suppressed proliferation, migration and invasion of PCa cells in vitro. Our data indicated the involvement of miR-30c in PCa progression and suggested its potential role as an independent predictor of biochemical recurrence in PCa. On cellular level, miR-30c may function as a tumor suppressor for PCa cells by inhibiting tumor cell proliferation, migration and invasion.

Global analysis of the differentially expressed miRNAs of prostate cancer in Chinese patients.

BACKGROUND: Our recent study showed the global physiological function of the differentially expressed genes of prostate cancer in Chinese patients was different from that of other non-Chinese populations. microRNA are estimated to regulate the expression of greater than 60% of all protein-coding genes. To further investigate the global association between the transcript abundance of miRNAs and their target mRNAs in Chinese patients, we used microRNA microarray approach combined with bioinformatics and clinical-pathological assay to investigate the miRNA profile and evaluate the potential of miRNAs as diagnostic and prognostic markers in Chinese patients. RESULTS: A total of 28 miRNAs (fold change ≥ 1.5; P ≤ 0.05) were differentially expressed between tumor tissue and adjacent benign tissue of 4 prostate cancer patients.10 top Differentially expressed miRNAs were validated by qRT-PCR using all 20 tissue pairs. Compared to the miRNA profile of non-Chinese populations, the current study showed that miR-23b, miR-220, miR-221, miR-222, and miR-205 maybe common critical therapeutic targets in different populations. The integrated analysis for mRNA microarray and miRNA microarray showed the effects of specifically inhibiting and/or enhancing the function of miRNAs on the gene transcription level. The current studies also identified 15 specific expressed miRNAs in Chinese patients. The clinical feature statistics revealed that miR-374b and miR-19a have significant correlations with clinical-pathological features in Chinese patients. CONCLUSIONS: Our findings showed Chinese prostate cancer patients have a common and specific miRNA expression profile compared with non-Chinese populations. The miR-374b is down-regulated in prostate cancer tissue, and it can be identified as an independent predictor of biochemical recurrence-free survival.

Down-regulation of the ErbB3 binding protein 1 in human bladder cancer promotes tumor progression and cell proliferation.

The ErbB3 binding protein 1 (Ebp1) represents a downstream effector of the ErbB signaling network and has been demonstrated to be a potent tumor suppressor in various human malignancies, however, its involvement in human bladder cancer is still unclear.To investigate the clinical significance and potential role of ErbB3 binding protein 1 (Ebp1) in bladder cancer. Ebp1 expression at protein and gene levels in 52 surgically removed bladder cancer specimens as well as 21 adjacent normal bladder specimens were respectively detected by immunohistochemistry and qRT-PCR. The association of Ebp1 protein expression with the clinicopathological features of bladder cancer was also statistically analyzed. Its roles in bladder cancer cell line were further evaluated. The expression level of Ebp1 protein and gene in bladder cancer tissues was significantly lower than that in adjacent normal bladder tissues (P < 0.01). When categorized into low vs. high expression, the down-regulation of Ebp1 protein was associated with the advanced pathologic stage (P = 0.036) and the high histologic grade (P = 0.001) of patients with bladder cancer. Moreover, following the transfection of Ebp1 in bladder cancer cells, not only cell proliferation and cell invasion decreased significantly, but also the cell cycle was blocked at G0/G1 stage. Our data suggest for the first time that the down-regulation of Ebp1 closely correlates with advanced clinicopathological characteristics of human bladder cancer. Furthermore, Ebp1 plays an important role in the bladder cancer cells' proliferation by regulating the cancer cell cycle from G0/G1 to S.

SOXs in human prostate cancer: implication as progression and prognosis factors.

BACKGROUND: SOX genes play an important role in a number of developmental processes. Potential roles of SOXs have been demonstrated in various neoplastic tissues as tumor suppressors or promoters depending on tumor status and types. The aim of this study was to investigate the involvement of SOXs in the progression and prognosis of human prostate cancer (PCa). METHODS: The gene expression changes of SOXs in human PCa tissues compared with non-cancerous prostate tissues was detected using gene expression microarray, and confirmed by real-time quantitative reverse transcriptase-polymerase chain reaction (QRT-PCR) analysis and immunohositochemistry. The roles of these genes in castration resistance were investigated in LNCaP xenograft model of PCa. RESULTS: The microarray analysis identified three genes (SOX7, SOX9 and SOX10) of SOX family that were significantly dis-regulated in common among four PCa specimens. Consistent with the results of the microarray, differential mRNA and protein levels of three selected genes were found in PCa tissues by QRT-PCR analysis and immunohistochemistry. Additionally, we found that the immunohistochemical staining scores of SOX7 in PCa tissues with higher serum PSA level (P = 0.02) and metastasis (P = 0.03) were significantly lower than those with lower serum PSA level and without metastasis; the increased SOX9 protein expression was frequently found in PCa tissues with higher Gleason score (P = 0.02) and higher clinical stage (P < 0.0001); the down-regulation of SOX10 tend to be found in PCa tissues with higher serum PSA levels (P = 0.03) and advanced pathological stage (P = 0.01). Moreover, both univariate and multivariate analyses showed that the down-regulation of SOX7 and the up-regulation of SOX9 were independent predictors of shorter biochemical recurrence-free survival. Furthermore, we discovered that SOX7 was significantly down-regulated and SOX9 was significantly up-regulated during the progression to castration resistance. CONCLUSIONS: Our data offer the convince evidence that the dis-regulation of SOX7, SOX9 and SOX10 may be associated with the aggressive progression of PCa. SOX7 and SOX9 may be potential markers for prognosis in PCa patients. Interestingly, the down-regulation of SOX7 and the up-regulation of SOX9 may be important mechanisms for castration-resistant progression of PCa.

[Corrigendum] miR­505 suppresses prostate cancer progression by targeting NRCAM.

Subsequently to the publication of this article, one of the corresponding authors, Dr Wei­De Zhong, has realized that the information presented in the box for correspondence for him was incorrect. Although Dr Zhong is correctly shown as having three affiliation addresses in the paper, the address affiliation listed first on the paper should have been presented as the address for correspondence, not the second one. Therefore, the authors' affiliation information should have appeared as follows (changes are highlighted in bold): Correspondence to: Dr Wei­ De Zhong, Guangdong Provincial Institute of Nephrology, Southern Medical University, Guangzhou, Guangdong 510515, P.R. China. Dr Zhong deeply regrets his oversight in this regard, and apologizes for any inconvenience caused. [the original article was published in Oncology Reports 42: 991-1004, 2019; DOI: 10.3892/or.2019.7231].

miR­505 suppresses prostate cancer progression by targeting NRCAM.

Previous researchers have demonstrated that microRNA­505 (miR­505) is negatively correlated with progression in various malignancies. However, the detailed function and molecular mechanisms of miR­505 have yet to be completely elucidated in prostate cancer (PCa). The present study initially identified the potential role of miR­505 in PCa using in vitro experiments, and demonstrated that restoration of miR­505 inhibited proliferation, invasion and migration, yet induced cell cycle arrest and promoted apoptosis in PCa cells. The present study also demonstrated that the expression of neuron­glial­related cell adhesion molecule (NRCAM) was markedly upregulated in PCa cells when compared with benign prostate epithelium. A luciferase reporter assay demonstrated that miR­505 directly targeted NRCAM in PCa cells. In addition, NRCAM stimulation antagonized the inhibitory effects of miR­505 on the proliferation, migration, and invasion of PCa cells. Furthermore, lower levels of miR­505 and higher levels of NRCAM may serve as a predictor of worse biochemical recurrence­free survival or disease­free survival in patients with PCa. In conclusion, the present study revealed the inhibitory effects of miR­505 on PCa tumorigenesis, which potentially occur by targeting NRCAM. The combined analysis of NRCAM and miR­505 may predict disease progression in patients with PCa following radical prostatectomy.

miR­30c suppresses prostate cancer survival by targeting the ASF/SF2 splicing factor oncoprotein.

Our previous study revealed that microRNA (miR) ­30c represents a potential tumor suppressor gene, the expression of which is associated with decreased oncogenic potential in prostate cancer (PCa) cell lines. However, the functional role and underlying mechanisms of miR­30c in PCa remain to be fully elucidated. Reverse transcription­quantitative polymerase chain reaction and immunohistochemical analysis were used to detect the expression levels of alternative splicing factor/splicing factor 2 (ASF/SF2) in PCa tissues. A luciferase reporter assay was used to investigate whether ASF/SF2 may be a direct target gene of miR­30c. In addition, the effects of miR­30c on the proliferation and apoptosis of PCa cell lines were examined, following transfection with miR­30c mimics. Furthermore, correlation analysis was performed to investigate the relationship between the expression of miR­30c and ASF/SF2 and various clinicopathological parameters of patients with PCa. The present results demonstrated that PCa tissues exhibited higher levels of alternative splicing factor/splicing factor 2 (ASF/SF2), compared with normal tissues. In addition, miR­30c was revealed to targete the 3'­untranslated region of the ASF/SF2 gene, causing a decrease in the mRNA and protein levels of ASF/SF2. Furthermore, miR­30c was reported to decrease cell proliferation, increase the percentage of cells in the G1 cell cycle phase, and promote apoptosis through the inhibition of ASF/SF2. Following correlation analysis using patient samples, the expression of ASF/SF2 was revealed to be tightly correlated with the pathological stage of PCa and biochemical recurrence (BCR). In addition, patients with PCa exhibiting low expression levels of miR­30c and high expression of ASF/SF2 had significantly lower rates of BCR­free survival. In conclusion, the present study suggested that the tumor suppressor miR­30c may be involved in PCa tumorigenesis, possibly via targeting ASF/SF2. The combined analysis of the expression of ASF/SF2 and miR­30c may be a valuable tool for early prediction of BCR in patients with PCa following radical prostatectomy.

BCL9, a coactivator for Wnt/ß-catenin transcription, is targeted by miR-30c and is associated with prostate cancer progression.

B-cell lymphoma 9 (BCL9), a component of aberrantly activated Wnt signaling, is an important contributing factor to tumor progression. Our previous data indicated that downregulation of the tumor suppressor microRNA-30c (miR-30c) was a frequent pathogenetic event in prostate cancer (PCa). However, a functional link between miR-30c and BCL9/Wnt signaling, and their clinical and pathological significance in PCa, have not been well established. The present study demonstrated that miR-30c serves as a key negative regulator targeting BCL9 transcription in PCa cells. Ectopic expression of miR-30c was associated with reduced expression of Wnt pathway downstream targets, including c-Myc, cluster of differentiation 44 and sex determining region Y-box 9 in DU145 human PCa cells. Examination of clinical prostate specimens revealed higher levels of BCL9 expression in PCa compared with that in benign prostate tissues. After substantiating this finding by patient sample analysis, BCL9 expression or activity was observed to be closely correlated with PCa biochemical recurrence (BCR) and disease progression, whereas it was inversely associated with miR-30c. Furthermore, overexpression of BCL9 in PCa acted cooperatively with miR-30c low expression to predict earlier BCR in PCa. These findings indicate that inhibition of BCL9/Wnt signaling by miR-30c is important in the progression of PCa. Furthermore, the combined analysis of miR-30c and BCL9 may be valuable tool for prediction of BCR in PCa patients following radical prostatectomy.

Overexpression of PDZ-binding kinase confers malignant phenotype in prostate cancer via the regulation of E2F1.

Roles and mechanisms of cell cycle-specific transcription factor E2F1 on prostate cancer (PCa) have not been fully elucidated. To address this problem, we here identified PDZ-binding kinase (PBK) as a direct target for E2F1 through bioinformatics binding site prediction, combined with chromatin immunoprecipitation-PCR (ChIP-PCR), quantitative (Q)-PCR and Western blot analysis. Then, we observed that the knockdown of both E2F1 and PBK could suppress cell proliferation, invasion and migration of PCa cell lines in vitro. Based on Taylor dataset, we found that PBK upregulation occurred more frequently in PCa patients with the older age of patients (P=0.044), the higher Gleason score (P<0.001), the advanced clinical pathological stage (P=0.019), the presence of metastasis (P=0.008), the overall survival (P<0.001) and PSA failure (P=0.004). More interestingly, the survival analysis identified PBK as an independent factor for predicting the biochemical recurrence-free survival of PCa patients (P=0.041). Taken together, these findings offer the convincing evidence for the first time that the overexpression of PBK may lead to high malignant phenotype in PCa cells via the regulation of E2F1. PBK may function as a biomarker that can differentiate patients with biochemical recurrent and non-biochemical recurrent disease following radical prostatectomy, highlighting its potential as a therapeutic target.

Decreased expression of myosin light chain MYL9 in stroma predicts malignant progression and poor biochemical recurrence-free survival in prostate cancer.

The aim of this study was to investigate the associations of myosin light chain (MYL9) downregulation with tumor progression and prognosis in patients with prostate cancer (PCa). MYL9 protein expression in human PCa and non-cancerous prostate tissues was detected by Western blot and immunohistochemistry analyses, which was validated by microarray-based Taylor data at mRNA level. Then, the associations of MYL9 expression with clinicopathological features and clinical outcome of PCa patients were statistically analyzed. Both Western blot and immunohistochemistry analyses found that MYL9 expression was significantly decreased (both P < 0.001) in PCa tissues compared with those in non-cancerous prostate tissues. In addition, MYL9 was mainly expressed in the cytoplasm of stromal cells of prostate tissues, and the decreased expression of MYL9 in PCa tissues was significantly correlated with the older age of patients (P = 0.011), the higher Gleason score (P < 0.001), the advanced pathological stage (P = 0.002), the presence of metastasis (P < 0.001) and PSA failure (P = 0.001). Furthermore, both univariate and multivariate analyses showed that the downregulation of MYL9 was an independent predictor of shorter overall survival (P = 0.026 and P = 0.009, respectively) and biochemical recurrence-free survival (P = 0.001 and P = 0.002, respectively). Our data strongly confirmed for the first time that the decreased expression of MYL9 may play an important role in tumor progression of PCa. More importantly, the downregulation of MYL9 may efficiently predict both overall and biochemical recurrence-free survivals in PCa patients.

Analysis of genetic aberrations on chromosomal region 8q21-24 identifies E2F5 as an oncogene with copy number gain in prostate cancer.

The copy number gain of genes in chromosomal region 8q21-24 has been demonstrated to be associated with genesis and progression of prostate cancer (PCa). The aim of this study was to identify novel and effective molecular markers in this chromosomal region for PCa. The differentially expressed genes in PCa specimens were screened by gene microarray analysis, which was validated by RT-QPCR analysis. Then, the DNA qPCR analysis was carried out to detect the copy number changes of these differentially expressed genes. Moreover, the clinical significance of candidate markers (MYC and E2F5) in PCa were further determined. E2F5 and MYC were identified as candidate markers in PCa tissues and PCa cell lines. The DNA qPCR revealed the significant copy number gains of E2F5 and MYC in PCa tissues but not in PCa cell lines. In addition, Western blot analysis and immunohistochemical staining both found the significant higher expression of E2F5 and MYC proteins in PCa tissues than those in adjacent benign specimens (all P < 0.01). Moreover, the overexpression of E2F5 protein was significantly associated with a high Gleason score (P < 0.01), an advanced clinical stage (P = 0.01), a positive metastasis (P < 0.01) and PSA Failure (P < 0.01). The overexpression of MYC was more frequently found in PCa tissues with positive metastasis (P = 0.02) and PSA failure (P = 0.02). Interestingly, there was a close correlation in the expression level of MYC in PCa tissues with that of E2F5 (r ( s ) = 0.5, P ≤ 0.001). Our data offers the convincing evidence that the copy number gains of E2F5 and MYC may play an important role in genesis and progression of PCa. Especially, E2F5 may be a novel potential candidate marker for malignant PCa.

MicroRNA-335 acts as a candidate tumor suppressor in prostate cancer.

MicroRNA-335 (miR-335) acts as a tumor suppressor or a tumor promoter in different human malignancies. However, the involvement of miR-335 in prostate cancer (PCa) is still unclear. The purpose of this study was to investigate the functional and clinical significance of miR-335 in PCa. miR-335 expression in 3 PCa cell lines (LNCaP/DU145/PC3) and in 20 clinical PCa tissues were detected by real-time quantitative reverse transcriptase-PCR compared with corresponding controls. The function of miR-335 was investigated for cell proliferation, invasion and migration in PCa cells transfected with agents containing EGFP-miR-335 expression vector. Additionally, miR-335 expression in 104 clinical PCa tissues was detected by in situ hybridization. Its assocaitions with clinicopathological features and prognosis in patients with PCa were also determined. miR-335 was significantly down-regulated in PCa cell lines than in the normal prostate cell line (P < 0.01). With the similar results in vitro, the reduced expression of miR-335 was also found in human PCa tissues comparing with paired adjacent benign prostate tissues (P < 0.05). Moreover, the increased expression of miR-335 suppressed cell proliferation, invasion and migration of PCa cell lines in vitro. Turning to its clinical significance, the low expression of miR-335 was significantly associated with high Gleason Score (P = 0.04), advanced clinical stage (P = 0.04), and positive metastasis (P = 0.02), but not with prognosis in PCa patients. Our data demonstrated for the first time the inhibitory effect of miR-335 on cell proliferation and invasion for PCa cells. The loss of this microRNA might be associated with clinical progression of PCa patients.

MicroRNA-23b downregulates peroxiredoxin III in human prostate cancer.

To investigate the mechanism by which peroxiredoxin III (PRDX3) is altered in human prostate cancer (PCa), we used microRNA (miRNA) target prediction program and miRNA microarray to predict and identify miR-23b as a candidate miRNA that targets PRDX3. We showed that miR-23b suppresses PRDX3 protein expression in human DU145 cells under normal and hypoxic conditions. Additionally, the clinical significance of miR-23b and PRDX3 expression in PCa patients was also confirmed. In conclusion, our data suggest that the effects of PRDX3 in PCa progression may be caused by the regulation function of miR-23b, and consequently, miR-23b may be involved in the response of PCa cells to hypoxia stress.