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Chronic inflammation in the benign prostate has been associated with a higher risk of developing prostate cancer. While a range of immune lineages is found in the prostate including T cells, B cells and myeloid cells, the specific subsets of immune cells with each major lineage have not been well described. In this study, we use mass cytometry (CyTOF) to comprehensively and reproducibly profile immune cells in mouse and human prostate. Using 4 myeloid markers (CD11b, CD11c, F4/80, Ly6C) in the mouse, we identified 8 phenotypically-distinct myeloid populations, demonstrating considerable heterogeneity within the immune compartment of the mouse prostate. We then profiled the prostate immune microenvironment from 9 human patients. Unlike the mouse prostate which is myeloid-dominant, the immune compartment in the benign human prostate is consistently T-lymphocyte-dominant. Using the X-shift algorithm to identify individual immune subsets based on marker expression, we found 57 phenotypically-distinct immune cell types in the human prostate. Despite similar proportions of T, B and myeloid lineage cells in the benign human prostate of all patients evaluated, we observed considerable interpatient heterogeneity in the abundance of more specific immune subsets. These findings highlight the importance of studying the immune compartment in the prostate at a granular level and will lead to future studies addressing the functional role of specific immune subsets in prostate epithelial transformation.

BackgroundProstate stem/progenitor cells function in glandular development and maintenance. They may be targets for tumor initiation, so characterization of these cells may have therapeutic implications. Cells from dissociated tissues that form spheres in vitro often represent stem/progenitor cells. A subset of human prostate cells that form prostaspheres were evaluated for self-renewal and tissue regeneration capability in the present study.MethodsProstaspheres were generated from 59 prostatectomy specimens. Lineage marker expression and TMPRSS-ERG status was determined via immunohistochemistry and fluorescence in situ hybridization (FISH). Subpopulations of prostate epithelial cells were isolated by cell sorting and interrogated for sphere-forming activity. Tissue regeneration potential was assessed by combining sphere-forming cells with rat urogenital sinus mesenchyme (rUGSM) subcutaneously in immunocompromised mice.ResultsProstate tissue specimens were heterogeneous, containing both benign and malignant (Gleason 3-5) glands. TMPRSS-ERG fusion was found in approximately 70% of cancers examined. Prostaspheres developed from single cells at a variable rate (0.5-4%) and could be serially passaged. A basal phenotype (CD44+CD49f+CK5+p63+CK8-AR-PSA-) was observed among sphere-forming cells. Subpopulations of prostate cells expressing tumor-associated calcium signal transducer 2 (Trop2), CD44, and CD49f preferentially formed spheres. In vivo implantation of sphere-forming cells and rUGSM regenerated tubular structures containing discreet basal and luminal layers. The TMPRSS-ERG fusion was absent in prostaspheres derived from fusion-positive tumor tissue, suggesting a survival/growth advantage of benign prostate epithelial cells.ConclusionHuman prostate sphere-forming cells self-renew, have tissue regeneration capability, and represent a subpopulation of basal cells.

The capacity of peripheral lymphocytes from rhesus monkeys which had been vasectomized for 7 or 11 years to stimulate and respond to normal rhesus lymphocytes in mixed lymphocyte cultures (MLC) was tested to determine whether vasectomy affects immunologic reactivity. The ability to respond in MLC, a T cell function, was significantly reduced in the 11-year vasectomized animals and in two 7-year vasectomized animals. The ability to stimulate in MLC, a B cell function, was significantly increased in the 11-year vasectomized group. MLC reactivity of normal lymphocytes cultured in plasma from vasectomized animals and lymphocytes from vasectomized animals cultured in normal plasma was not altered, ruling out serum effects in the reduction of MLC responsiveness in these vasectomized animals. Seventy-five per cent of the vasectomized animals with markedly reduced MLC reactivity had the RhL-A determinates 19 and 24, indicating an association between the tendency toward reduced MLC reactivity after vasectomy and histocompatibility type.

Activation of human T cells requires presentation of antigen by Ia (HLA-DR in man) bearing cells of the mononuclear phagocytic series (macrophages, MO), and more recently Langerbans cells, dendritic cells, and vascular endothelial cells. Since T cells must cross endothelial barriers to enter extavasulcar tissues during immune reactions, we investigated the role of endothelial cells in antigen presentation. Endothelial cells were cultured from human umbilical veins and identified by classic morphology and specific markers (factor VIII related antigen, and so on). Antigen-pulsed endothelial cells were used to present antigen to MO-depleted human T cells; activation was assessed by 3 H-thymidine uptake. The HLA-DR compatible endothelial cells were as effective as MO-in reconstituting MO-depleted T-cell responses. The endothelial cell reconstituted responses were antigen specific, HLA-DR restricted, and blocked by monoclonal antibodies to HLA-DR framework structures. Moreover, the T-cell responses were clonal with respect to HLA-DR. A monoclonal antibody completely eliminated MO reconstitution of the MO-depleted response without diminution of endothelial cell reconstitution of the same response. Fibroblasts and smooth muscle cells cultured from the same umbilical veins could not reconstitute the MO-depleted T-cell response. These data indicate that endothelial cells play an important and distinctive role in lymphocyte triggering.

An increasing number of diseases has been associated with specific human leukocyte antigens. A total of 100 white men with adenocarcinoma of the prostate was serotyped for 32 A and B antigens. Compared to a control population of 169 white subjects relative risks of 2 or more were associated with the antigens All and A29. Relative risks of 0.20 or less were associated with the antigens AW23, BW21 and BW22. None of these relative risks was significant statistically. We were unable to demonstrate a significant human leukocyte A or B antigen association with adenocarcinoma of the prostate.