Cancer-associated Fibroblast-derived Extracellular Vesicles Mediate Immune Escape of Bladder Cancer via PD-L1/PD-1 Expression.

OBJECTIVE: Bladder cancer (BCa) is a malignant urological tumor with a high prevalence and poor prognosis. Extracellular vesicles (EVs) are increasingly becoming current hotspots owing to their involvement in cancer progression. This paper probed into the action of cancer-associated fibroblast-derived EVs (CAF-EVs) in the immune escape of BCa. METHODS: CAFs were identified by immunofluorescence. EVs were extracted from CAFs via ultracentrifugation and later characterized. BCa cells (T24 cell line) were co-cultured with CD8+ T cells and then treated with CAF-EVs. The uptake of EVs by T24 cells was examined by confocal laser microscopy. T24 cell apoptosis and invasion were assessed using flow cytometry and invasion assay. CD8+ T cell proliferation was evaluated using CFSE staining. The levels of cytokines (IFN-γ, IL-2, and TNF-α) were measured by ELISA. PD-L1 and PD-1 levels were determined utilizing RT-qPCR and flow cytometry. BCa mouse models were established to identify the effect of CAF-EVs on BCa progression in vivo. RESULTS: CAF-EVs decreased apoptosis and enhanced invasion of T24 cells, reduced proliferation of CD8+ T cells, and diminished levels of IFN-γ, IL-2, and TNF-α secreted by CD8+ T cells. CAF-EVs promoted the immune escape of T24 cells by carrying PD-L1. Downregulation of PDL1 expression in T24 cells or EVs partially counteracted the promotion of CAF-EVs on immune escape by reducing the binding of PD-L1 and PD-1. Additionally, CAF-EVs raised tumor volume and weight, upregulated PD-L1 expression, and weakened CD8+ T cell infiltration in BCa mice. CONCLUSION: CAF-EVs facilitate the immune escape of BCa by upregulating PD-L1/PD-1.

NEDD4L represses prostate cancer cell proliferation via modulating PHF8 through the ubiquitin–proteasome pathway

Purpose Prostate cancer (PC) is a heterogeneous malignancy that greatly threatens man’s health. E3 ubiquitin-protein ligase neural precursor cell expressed developmentally downregulated 4-like (NEDD4L) imparts an regulatory role in various malignancies. This study focused on the modulatory mechanism of NEDD4L in proliferation of prostate cancer cells (PCCs) via regulating histone demethylase plant homeodomain finger protein 8 (PHF8/KDM7B) through the ubiquitin–proteasome system. Methods The expression levels of NEDD4L, PHF8, H3 lysine 9 dimethylation (H3K9me2) and activating transcription factor 2 (ATF2) in PC tissues and cell lines were detected via real-time quantitative polymerase chain reaction and Western blotting. After transfection of pcDNA3.1-NEDD4L, pcDNA3.1-PHF8, and pcDNA3.1-ATF2 into PCCs, cell proliferation was assessed via the cell counting kit-8 and 5-ethynyl-2'-deoxyuridine assays. Interaction between NEDD4L and PHF8 was identified via the protein immunoprecipitation. The ubiquitination level of PHF8 was determined via the ubiquitination detection. The enrichments of H3K9me2 and PHF8 in the ATF2 promotor region were detected via the chromatin-immunoprecipitation assay. Results PHF8 and ATF2 were highly expressed while NEDD4L was poorly expressed in PC tissues and cells. NEDD4L overexpression reduced proliferation of PCCs. NEDD4L induced degradation of PHF8 via ubiquitination. PHF8 limited the enrichment of H3K9me2 in the ATF2 promotor region and enhanced ATF2 transcription. Up regulation of PHF8 or ATF2 abolished the inhibitory role of NEDD4L in proliferation of PCCs. Conclusion NEDD4L facilitated degradation of PHF8 to limit ATF2 transcription, thereby suppressing proliferation of PCCs. (AU)

NEDD4L represses prostate cancer cell proliferation via modulating PHF8 through the ubiquitin-proteasome pathway.

PURPOSE: Prostate cancer (PC) is a heterogeneous malignancy that greatly threatens man's health. E3 ubiquitin-protein ligase neural precursor cell expressed developmentally downregulated 4-like (NEDD4L) imparts an regulatory role in various malignancies. This study focused on the modulatory mechanism of NEDD4L in proliferation of prostate cancer cells (PCCs) via regulating histone demethylase plant homeodomain finger protein 8 (PHF8/KDM7B) through the ubiquitin-proteasome system. METHODS: The expression levels of NEDD4L, PHF8, H3 lysine 9 dimethylation (H3K9me2) and activating transcription factor 2 (ATF2) in PC tissues and cell lines were detected via real-time quantitative polymerase chain reaction and Western blotting. After transfection of pcDNA3.1-NEDD4L, pcDNA3.1-PHF8, and pcDNA3.1-ATF2 into PCCs, cell proliferation was assessed via the cell counting kit-8 and 5-ethynyl-2'-deoxyuridine assays. Interaction between NEDD4L and PHF8 was identified via the protein immunoprecipitation. The ubiquitination level of PHF8 was determined via the ubiquitination detection. The enrichments of H3K9me2 and PHF8 in the ATF2 promotor region were detected via the chromatin-immunoprecipitation assay. RESULTS: PHF8 and ATF2 were highly expressed while NEDD4L was poorly expressed in PC tissues and cells. NEDD4L overexpression reduced proliferation of PCCs. NEDD4Linduced degradation of PHF8 via ubiquitination. PHF8 limited the enrichment of H3K9me2 in the ATF2 promotor region and enhanced ATF2 transcription. Upregulation of PHF8 or ATF2 abolished the inhibitory role of NEDD4L in proliferation of PCCs. CONCLUSION: NEDD4L facilitated degradation of PHF8 to limit ATF2 transcription, thereby suppressing proliferation of PCCs.

Silenced lncRNA SNHG14 restrains the biological behaviors of bladder cancer cells via regulating microRNA-211-3p/ESM1 axis.

BACKGROUND: Bladder cancer (BCa) is a malignant tumor that occurs on the mucosa of the bladder, in which dysregulated long non-coding RNAs (lncRNAs) are involved. This study investigated the effect of lncRNA small nucleolar RNA host gene 1 (SNHG14) on the biological characteristics of BCa cells from microRNA (miR)-211-3p/ESM1 signaling axis. METHODS: BCa tissues and the matched normal tissues were collected to test SNHG14, miR-211-3p and ESM1 levels. SNHG14, miR-211-3p and ESM1 levels in BCa cell lines (T24, 5637, UMUC-3 and EJ) and normal bladder epithelial cells SV-HVC-1 were detected for screening the cell lines for follow-up experiments. T24 and UMUC-3 cells were transfected with different plasmids of SNHG14, miR-211-3p or ESM1 to observe the biological characteristics of BCa cells by MTT, colony formation, Transwell assays and flow cytometry. Tumor xenograft was implemented to inspect tumor growth in vivo. The targeting relationships of SNHG14, miR-211-3p and ESM1 were verified by bioinformatics software, RNA pull down assay and luciferase reporter assay. RESULTS: Enhanced SNHG14, ESM1 and suppressed miR-211-3p were found in BCa tissues and cells. SNHG14 up-regulated ESM1 via competitive binding with miR-211-3p. Decreased SNHG14 or up-regulated miR-211-3p depressed cell cycle entry, colony formation, invasion, migration and proliferation abilities, and facilitated apoptosis of BCa cells. Decreased SNHG14 or up-regulated miR-211-3p reduced the tumor volume and weight of nude mice with BCa, as well as promoted apoptosis and restrained proliferation of tumor cells. miR-211-3p inhibition or ESM1 overexpression reversed the effects of down-regulation of SNHG14 on BCa, and miR-211-3p up-regulation or ESM1 downregulation reversed the effect of SNHG14 overexpression on BCa. SNHG14 targeted miR-211-3p to regulate ESM1 expression. CONCLUSION: Our study highlights that silenced SNHG14 or elevated miR-211-3p represses the tumorigenic ability of BCa cells, which may be linked to ESM1 knockdown.

[Expression of vasodilator-stimulated phosphoprotein relates to prostate cancer metastasis].

OBJECTIVE: To investigate the relationship of the expression of vasodilator-stimulated phosphoprotein (VASP) with the metastasis and prognosis of prostate cancer. METHODS: Prostate cancer PC3 cells were infected with VASP shRNA and control shRNA lentiviruses, respectively. The invasive ability of the PC3 cells was determined by transwell migration assay, the expression of VASP in the prostate cancer tissue from 56 patients was detected by immunohistochemistry, and the survival rate of the patients was analyzed according to the VASP expression levels and follow-up data after radical prostatectomy. RESULTS: VASP shRNA lentivirus significantly inhibited the expression of VASP and decreased the invasive ability of the PC3 cells as compared with the results obtained in the scramble shRNA and blank control groups (P<0.05). The survival analysis of the 56 prostate cancer patients showed that the time of biochemical recurrence was markedly shorter in the VASP positive and strongly positive groups than in the VASP-negative cases (P<0.05), but with no statistically significant difference between the former two groups (P>0.05). CONCLUSIONS: VASP is involved in the regulation of the invasive ability of prostate cancer PC3 cells, and the differences in the VASP expression are related to the prognosis of prostate cancer.

A new method of chronic and recurrent seminal vesiculitis treatment.

PURPOSE: To investigate a new method and its effect on the procedure of dilating the ejaculatory duct and flushing the seminal vesicle with an F9 seminal vesicle scope in patients with chronic and recurrent seminal vesiculitis. PATIENTS AND METHODS: Twenty-six patients with a diagnosis based to signs, laboratory detection, digital rectal examination, and transrectal ultrasonography were involved in present study. The patients underwent a surgical procedure of dilating the ejaculatory duct and flushing the seminal vesicles with an F9 seminal vesicle endoscope. All patients were followed for 3 months to 1 year after treatment. RESULTS: There were significant reductions in symptoms, signs, white blood cell and red blood cell counts on microscopic examination, seminal vesicles size, improvement of inner walls echo in transrectal ultrasonography, and semen culture positive rate. Moreover, all patients showed improvement. CONCLUSIONS: The present study provides a new transurethral seminal tract endoscopic technique with seminal vesicle scope through the normal anatomic tract to treat patients with chronic seminal vesiculitis. It proved to be easily conducted with minimized complications. Further investigations are needed to confirm our results.

Novel methods for the diagnosis and treatment of ejaculatory duct obstruction.

OBJECTIVES: • To investigate a new method of vas deferens radiography for ejaculatory duct obstruction (EDO). • To evaluate the effect of a procedure involving dilation of the ejaculatory duct by F9 seminal vesicoscopy. PATIENTS AND METHODS: • Twenty-two patients with EDO were diagnosed using semen analysis, semen fructose measurement, transrectal ultrasonography (TRUS) and vas deferens radiography. • Of these, 18 patients were successfully treated by dilation of ejaculatory duct using F9 seminal vesicoscopy and four patients, whose treatment was unsuccessful, were treated by transurethral resection of the ejaculatory ducts (TURED). • All patients were followed up for at least 3 months after treatment. RESULTS: • Semen analyses in all 22 patients showed oligoasthenozoospermia or azoospermia, low semen volume (0-1.9 mL), low pH level (5.6-7.0) and absent or low semen fructose. TRUS and radiography showed pure dilated seminal vesicles on both sides in three patients, partial dilated seminal vesicles in one patient, dilation of both the ejaculatory duct and seminal vesicles in ten patients, dilated seminal vesicles and a prostatic cyst in four patients, and dilated ejaculatory duct or cystic lesions without dilated seminal vesicles in the remaining four patients. • At >3-month follow-up after dilation or TURED, the semen characteristics of 18 patients were improved and sperm were present in the semen in 13 cases. Normal semen analyses were found in 7 patients and 6 patients had conceived. • Voiding urethral radiography showed that no patients who had undergone dilation by seminal vesicoscopy had urine reflux into the ejaculatory duct. Only one patient showed urine reflux into the seminal vesicle after TURED. • All patients felt that their symptoms had improved after treatment. CONCLUSIONS: • The approach to vas deferens radiography using vas deferens aspiration has proved to be an effective and safe method for EDO diagnosis. • The procedure involving the dilation of the ejaculatory duct using F9 seminal vesicoscopy is equally effective but has fewer postoperative complications than TURED.

Analysis and difference of voltage-dependent anion channel mRNA in ejaculated spermatozoa from normozoospermic fertile donors and infertile patients with idiopathic asthenozoospermia.

PURPOSE: to analyze the abundance and difference of voltage-dependent anion channel (VDAC) mRNA in ejaculated spermatozoa from normozoospermic fertile donors and infertile patients with idiopathic asthenozoospermia. METHODS: high motile and low motile spermatozoa were separated respectively from ejaculates of 36 donors and 40 patients using a discontinuous Percoll gradient centrifugation. Real-Time PCR was performed to detect mRNA abundance and difference of three VDAC subtypes between two groups with different sperm motility. RESULTS: real-Time PCR demonstrated that three VDAC mRNAs were present in mature spermatozoa. The VDAC2 mRNA level in ejaculated spermatozoa of patients was significantly higher than that of donors. No significant differences of VDAC1 and VDAC3 mRNA levels were found between two groups. CONCLUSION: the high abundance of VDAC2 mRNA seemed to have a positive correlation with low sperm motility. The abnormal expression of VDAC might be related to male infertility with idiopathic asthenozoospermia.