Tigated if a progressive increase in the levels of Bmi1 in prostatic tissues could be detected during progressive age of CaP. For this purpose we used prostatic tissue samples collected at different ages of TRAMP transgenic mice. As shown in Fig 1A; Bmi1 protein was observed to be detectable in all ages of TRAMP mice. In general, the staining was stronger in prostatic epithelial cells from older mice than in prostatic epithelial cells from younger mice. Smooth muscle cells have much stronger staining than fibroblast cells. The staining pattern of Bmi1 protein was compared in age 17 weeks to 45 weeks old prostatic specimens (Fig. 1A). These data showed increased expression levels of Bmi1 protein in prostate of older aged mice (Fig. 1A). There was an intense staining at apical region of epithelial cells. Stromal regions were observed to have a positive staining (Fig. 1A).BMI1 expression in normal and neoplastic prostatic cells representing CaP KDM5A-IN-1 price disease in Africa- American menAge-adjusted data from SEER study showed that AfricanAmerican men have a 60 higher incidence and 125 higher mortality rates from CaP than Caucasian men [1,13]. Race and family history are the two most widely accepted risk factors for this disease [1]. Understanding the underlying biological mechanismsBMI1:Potential Serum-Biomarker for Prostate CancerFigure 1. BMI1 protein levels in prostatic tumor tissues of humans and TRAMP transgenic mice. (A) Photomicrographs represent immunostaining of BMI1 in prostatic tissues of transgenic TRAMP mice. Arrows indicate staining for BMI1. Magnification 640. (Bi) Photomicrographs show BMI1-positive neoplastic and non-neoplastic regions of prostatic specimens of CaP patients as assessed by immunostaining. Arrows indicate staining for BMI1. Magnification 640. (Bii) Box plots for BMI1 protein based 18325633 on score pertain to immunostaining pattern in normal and CaP specimens in stromal region.*, P,0.05; black bar in box, median values. doi:10.1371/journal.pone.0052993.gresponsible for CaP progression will eventually lead to the development of more effective therapeutic strategies. We determined the levels of BMI1 expression in a cell-based in vitro model representing different phenotypes of CaP disease in AfricanAmerican men. These include RC77N/E (representing normal prostatic epithelial cells in African-American men), RC77T/E (representing androgen-dependent tumorigenic prostatic epithelial cells), E006 (representing androgen-dependent non-tumorigenic prostatic epithelial cells) and PCa-2b (representing CRPC phenotype; however retain androgen responsiveness) [13?4]. As shown in Figure 2(Bi i), all CaP cell lines RC77T/E, PCa2b and E006 Arg8-vasopressin biological activity exhibited a higher expression of BMI1 protein than in normal cells RC77N/E. These data (Fig. 2A ) suggest a possibility that expression of intracellular BMI1 protein may be correlated with the secretory BMI1 levels in human tissues and may play a role in aggressiveness of human CaP.BMI1 is a secretory protein: Detection in serum-free media from CaP cell culturesThe presence of BMI1 in the apical region of prostate epithelial cells and stromal region prompted us to hypothesize that it could be a secretory protein in nature. To test our hypothesis we asked if BMI1 is secreted by tumor cells under culture conditions. In order to detect BMI1 protein in culture media of CaP cells, we employed Slot-blot technique. Cells at a confluency level of 80 were allowed to grow in fresh serum-free media for 24 h. As eviden.Tigated if a progressive increase in the levels of Bmi1 in prostatic tissues could be detected during progressive age of CaP. For this purpose we used prostatic tissue samples collected at different ages of TRAMP transgenic mice. As shown in Fig 1A; Bmi1 protein was observed to be detectable in all ages of TRAMP mice. In general, the staining was stronger in prostatic epithelial cells from older mice than in prostatic epithelial cells from younger mice. Smooth muscle cells have much stronger staining than fibroblast cells. The staining pattern of Bmi1 protein was compared in age 17 weeks to 45 weeks old prostatic specimens (Fig. 1A). These data showed increased expression levels of Bmi1 protein in prostate of older aged mice (Fig. 1A). There was an intense staining at apical region of epithelial cells. Stromal regions were observed to have a positive staining (Fig. 1A).BMI1 expression in normal and neoplastic prostatic cells representing CaP disease in Africa- American menAge-adjusted data from SEER study showed that AfricanAmerican men have a 60 higher incidence and 125 higher mortality rates from CaP than Caucasian men [1,13]. Race and family history are the two most widely accepted risk factors for this disease [1]. Understanding the underlying biological mechanismsBMI1:Potential Serum-Biomarker for Prostate CancerFigure 1. BMI1 protein levels in prostatic tumor tissues of humans and TRAMP transgenic mice. (A) Photomicrographs represent immunostaining of BMI1 in prostatic tissues of transgenic TRAMP mice. Arrows indicate staining for BMI1. Magnification 640. (Bi) Photomicrographs show BMI1-positive neoplastic and non-neoplastic regions of prostatic specimens of CaP patients as assessed by immunostaining. Arrows indicate staining for BMI1. Magnification 640. (Bii) Box plots for BMI1 protein based 18325633 on score pertain to immunostaining pattern in normal and CaP specimens in stromal region.*, P,0.05; black bar in box, median values. doi:10.1371/journal.pone.0052993.gresponsible for CaP progression will eventually lead to the development of more effective therapeutic strategies. We determined the levels of BMI1 expression in a cell-based in vitro model representing different phenotypes of CaP disease in AfricanAmerican men. These include RC77N/E (representing normal prostatic epithelial cells in African-American men), RC77T/E (representing androgen-dependent tumorigenic prostatic epithelial cells), E006 (representing androgen-dependent non-tumorigenic prostatic epithelial cells) and PCa-2b (representing CRPC phenotype; however retain androgen responsiveness) [13?4]. As shown in Figure 2(Bi i), all CaP cell lines RC77T/E, PCa2b and E006 exhibited a higher expression of BMI1 protein than in normal cells RC77N/E. These data (Fig. 2A ) suggest a possibility that expression of intracellular BMI1 protein may be correlated with the secretory BMI1 levels in human tissues and may play a role in aggressiveness of human CaP.BMI1 is a secretory protein: Detection in serum-free media from CaP cell culturesThe presence of BMI1 in the apical region of prostate epithelial cells and stromal region prompted us to hypothesize that it could be a secretory protein in nature. To test our hypothesis we asked if BMI1 is secreted by tumor cells under culture conditions. In order to detect BMI1 protein in culture media of CaP cells, we employed Slot-blot technique. Cells at a confluency level of 80 were allowed to grow in fresh serum-free media for 24 h. As eviden.
