Low intraprostatic DHT promotes the infiltration of CD8+ T cells in BPH tissues via modulation of CCL5 secretion

Abstract

Clinical studies suggested thatandrogen might be associated with infiltrating T cells in prostate of benign prostatic hyperplasia (BPH) patients, but detail of T-cell subset and mechanism still remained unclear. The present study tested the hypothesis that intraprostatic 5 α -dihydrotestosterone (DHT) exerts effects on T cells recruitment by BPH epithelial cells. Prostate tissues from 64 cases of BPH patients after transurethral resection of prostate (TURP) were divided into 2 groups: (1) no medication history; (2) administration of 5 α -reductase type II inhibitor-finasteride 5 mg daily for at least 6 months before surgery. Group 2 presented significantly higher CD8+ T cells infiltration than group 1, but no changes in CD4+ T cells (immunohistochemistry and flow cytometry). In vitro study more CD8+ T cell migrated to the prostate tissue lysates from group 2 and BPH-1 cells in low DHT condition. Transcription of chemokine (C-C motif) Ligand 5 (CCL5) mRNA in BPH-1 cells and chemokine (C-C motif) receptor 5 (CCR5) mRNA in CD8+ T cells were upregulated in low DHT condition (q-PCR). CCL5 expression was also identified to be higher in group 2 prostate tissues by IHC. This study suggested that intraprostatic DHT may participate in regulating inflammatory response which was induced by human prostatic epithelial cell, via modulating CCL5 secretion.

Figure 1

The T-cell population infiltrating prostate tissue with/without finasteride treatment. (a) CD8 was stained from no medication group and finasteride group; scale bar: 100 μm and 20 μm. Negative controls were showed in the bottom left corner, respectively. Data presented as the percentage of CD8+ T-cell number in all of nuclear cell number (mean ± SEM); *P = 0.013 (t-test). (b) Immunohistochemistry staining for CD4+ T infiltration. Magnification and negative control are the same as mentioned before. (c) Flow cytometry assay. Data presented as the rate of CD8+ or CD4+ T cells in total T cell (mean ± SEM). *P = 0.003 in the right column, (t-test).

Figure 2

CD8+ T cells migration in vitro. (a) CD8+ T cell migrated to lower chamber after 6 hrs. Scale bar: 50 μm. (b) Data showed as the percentage of migrated cell number in total cell number (mean ± SEM). *P < 0.000. (t-test). (c) The migration of molt-3 cells to BPH-1 cells with/without charcoal medium treatment in the lower chamber. Data presented as the average cell numbers (mean ± SEM). *P = 0.026. (ANOVA and Newman-Keuls test).

Figure 3

Induction of chemokines in BPH-1 cells stimulated by changes of DHT level. (a) Q-PCR screening of a panel of cytokine factors that could be responsible for BPH-1 cell promoted T-cell migration. Compared to the BPH-1 cells cultured with normal medium, mRNA transcription of CCL5 was upregulated in BPH-1 cells with charcoal medium treatment; *P = 0.014 (t-test). (b) Transcription of chemokine related receptors mRNA was detected by q-PCR. *P = 0.018 (ANOVA and Newman-Keuls test). (c) The interruption assay by adding CCL5 neutralizing antibody in above migration system. Data presented as the average cell numbers in lower chamber (mean ± SEM). *P = 0.031 (ANOVA and Newman-Keuls test).

Figure 4

CCL5 immunolocalization in prostate tissue samples by IHC. (a) CCL5 was stained from no medication group and finasteride group. Magnification and negative control are the same as mentioned before. (b) The average immunoreactive score of CCL5 in different group was quantified (mean ± SEM). *P = 0.002 (Mann-Whitney test).

Figure 5

Mechanism and regulatory pathway of low intraprostatic DHT-promoted CD8+ T cell infiltration in BPH prostate tissue. Low intraprostatic DHT level could promote BPH epithelial cells to recruit more CD8+ T cells via upregulation of CCL5 mRNA transcription.