Predictors for spontaneous pregnancy after microsurgical subinguinal varicocelectomy: a prospective cohort study.

PURPOSE: The purpose of this study is to determine the possible preoperative predictors of spontaneous pregnancy (SPR) for infertile males with varicocele after microsurgical subinguinal varicocelectomy (MVL) performed in two medical centers in a prospective cohort study. METHODS: A total of 120 males with varicocele that underwent MVL between June 2013 and June 2014 in two medical centers were documented. Related data, including male and female partner age, male body mass index (BMI), female BMI, preoperative semen parameters, hormone levels, testicular volume, grade and side of varicocele, were collected and analyzed. The follow-up assessment was also conducted within a 2-year period after the surgery. The outcome measure was SPR within the 2-year follow-up reported. The possible determinants of SPR were also analyzed and indentified using Cox regression analysis. RESULTS: Of the 110 patients that accomplished the 2-year follow-up, 42 patients reported pregnancy outcome. Using Cox regression analysis, total motile sperm count [TMC; RR (95% CI) = 1.362 (1.120-1.560), p = 0.003] and follicle-stimulating hormone [FSH; RR (95% CI) = 0.726 (0.541-0.980), p = 0.020] levels posed significant determinants for SPR. CONCLUSION: Our study indicated that males who underwent MVL with higher TMC and lower FSH preoperatively have higher possibility of pregnancy postoperatively.

ROS activates JNK-mediated autophagy to counteract apoptosis in mouse mesenchymal stem cells in vitro.

AIM: Transplantation of mesenchymal stem cells (MSCs) for the treatment of diabetic erectile dysfunction (ED) is hampered by apoptosis of the transplanted cells. In diabetic ED, there is increased oxidative stress and decreased NO in the corpora cavernosa, and reactive oxygen species (ROS) induce apoptosis of the transplanted cells. In this study we examined whether and how autophagy was involved in ROS-induced apoptosis of MSCs. METHODS: Mouse C3H10 MSCs were treated with H2O2 to simulate the high oxidative condition in diabetic ED. Cell viability was measured using MTT assay. Apoptosis was analyzed by flow cytometry. Apoptosis- and autophagy-related proteins were detected with Western blot assays. Intracellular autophagosome accumulation was studied using transmission electron microscopy. RESULTS: Treatment of MSCs with H2O2 (50-400 µmol/L) inhibited the cell viability in concentration- and time-dependent manners. Furthermore, H2O2 (300 µmol/L) induced apoptosis, as well as activated autophagy in MSCs. Pretreatment with lysosome inhibitor chloroquine (10 µmol/L) or PI3K inhibitor 3-methyladenine (5 mmol/L) significantly enhanced H2O2-induced cell death. Pretreatment with JNK inhibitor SP600125 (10 µmol/L) abrogated H2O2-induced accumulation of LC3-II, and attenuated H2O2-induced reduction of Bcl-2 levels in MSCs. CONCLUSION: ROS induce autophagy to counteract apoptosis in MSCs by activation of JNK. Thus, augmentation of autophagy may reduce apoptosis, prolonging MSC survival and improving MSC-based therapeutic efficacy for diabetic ED.

Over-expressing transporters associated with antigen processing increases antitumor immunity response in prostate cancer.

As we know, prostate cancer down-regulates expression of HLA-1 Antigen Processing Machinery (APM) and has defects in the antigen presentation pathway. In vitro, the prostate cancer cell (PC-3 cells) infected with Lentivirus TAP1 can efficiently over-express TAP1 and Tapasin, and HLA-1 was also up-regulated on the surface of the infected cells. The lentivirus TAP1 infection increased the apoptosis rate of PC-3 cells. In addition, with the co-cluture PC-3 cells and lymphocytes, TAP1 augmented the expression of CD3⁺CD8⁺CD38⁺ T cell. Importantly, administration of Lentivirus TAP1 to prostate cancer cells in a xenograft mouse model can prolong survival and increase the CD4⁺ T cells, and CD8⁺ T cells as well as decrease Foxp3⁺ T cells in the tumor microenvironment. In summary, a recombinant lentivirus expressing TAP1 can effectively increase prostate cancer tumor-specific immune response.

Tolerogenic dendritic cells generated by RelB silencing using shRNA prevent acute rejection.

It is well known that adoptive transfer of donor-derived tolerogenic dendritic cells (DCs) helps to induce immune tolerance. RelB, one of NF-κB subunits, is a critical element involved in DC maturation. In the present study, our results showed tolerogenic DCs could be acquired via silencing RelB using small interfering RNA. Compared with imDCs, the tolerogenic DCs had more potent ability to inhibit mixed lymphocyte reaction (MLR) and down-regulate Th1 cytokines and prompt the production of Th2 cytokines. They both mediated immune tolerance via the increased of T cell apoptosis and generation of regulatory T cells. Administration of donor-derived tolerogenic DCs significantly prevented the allograft rejection and prolonged the survival time in a murine heart transplantation model. Our results demonstrate donor-derived, RelB-shRNA induced tolerogenic DCs can significantly induce immune tolerance in vitro and in vivo.

Lentiviral-mediated shRNA against RelB induces the generation of tolerogenic dendritic cells.

OBJECTIVE: Lentiviral-mediated shRNA against RelB was used to produce tolerogenic dendritic cells from murine bone marrow derived dendritic cells (BMDCs). METHOD: RelB expression in the BMDCs was silenced by lentivirus carrying RelB shRNA. The apoptosis rate and surface markers of DCs were assessed by flow cytometry. IL-12,IL-10,TGF-ß1 secreted by DCs and DNA binding capacity of NF-κB subunits in the nucleus were measured by ELISA, independently. MLR was used to analyze the capacity of DCs to inhibit immune response. RESULTS: RelB expression was significantly inhibited in DCs following lentiviral mediated delivery of RelB specific shRNA. The RelB shRNA-DC produced lower IL-12 and higher IL-10 than mature dendritic cells (mDCs) and silencing control DCs. There was no difference in the apoptosis rate between shRNA RelB-DCs and mDCs. The expression levels of co-stimulatory molecules (CD80, CD86 and CD83) and MHC-II class molecule were lower in the RelB shRNA-DCs than in the mDCs and silencing control DCs. In addition, RelB shRNA also inhibited the RelB DNA binding capacity but had no effect on other NF-κB subunits. The shRNA RelB-DCs can significantly inhibit mixed lymphocyte reaction (MLR) and down-regulate Th1 cytokines and prompt the production of Th2 cytokines. CONCLUSION: Our results indicate RelB shRNA transfection of DCs can induce the immature status, and produce tolerogenic DCs.

[Prostate cancer cell vaccine transfected with 4-1BBL induces anti-tumor immunity in vitro].

OBJECTIVE: To explore the anti-tumor immunity in vitro induced by prostate cancer cell vaccine transfected with recombinant adenovirus encoding 4-1BBL in mice. METHODS: The replication-deficient adenovirus AdEasy-1 system was used to construct recombinant adenovirus Ad-m4-1BBL and Ad-eGFP. The prostate cancer cell RM-1 of mice was transfected with Ad-m4-1BBL and Ad-eGFP, and treated with mitomycin (MMC) to produce TCV, TCV-Ad-eGFP and TCV-Ad-m4-1BBL, followed by co-culture with syngeneic murine spleen cells. Then the cytotoxic activity of the lymphocytes against RM-1 cells was analyzed with CCK-8 solution, and IL-2 and INF-gamma were detected by ELISA. RESULTS: The 4-1BBL protein was highly expressed in the TCV-Ad-m4-1BBL of the 4-1BBL-transfected mice. TCV-Ad-m4-1BBL significantly increased the expressions of IL-2 ([180.24 +/- 2.22] pg/ml) and INF-gamma ([1512.46 +/- 23.64] pg/ml) as compared with TCV and TCV-Ad-eGFP (P < 0.05), and induced higher RM-1 cell specific cytotoxicity ([34.24 +/- 2.64]%) than the latter two ([9.82 +/- 1.48]%) and ([14.65 +/- 3. 21]%), (P < 0.05). But none of them exhibited significant cytotoxicity against hepatocellular carcinoma Hepal-6. CONCLUSION: The m4-1BBL-expressing prostate cancer cell vaccine can effectively induce anti-tumor immune responses.