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1 alpha, 25-dihydroxylvitamin D3 promotes Bacillus Calmette-Guérin immunotherapy of bladder cancer.

Hsu JW, Yin PN, Wood R, Messing J, Messing E, Lee YF - Oncotarget (2013)

Bottom Line: However, the mechanisms of BCG action have not been completely understood, thereby limiting the improvement of BCG therapy.Vitamin D deficiency has been associated with a high risk of TB infection, and the beneficial effect of UV exposure in TB patients was proven to be mediated via activation of vitamin D signals of innate immune cells.This THP-1 cell migration promoted by 1,25-VD can be blocked by IL-8 neutralized antibody.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, University of Rochester, Rochester, New York, USA.

ABSTRACT
Bacillus Calmette-Guérin (BCG), a vaccine against tuberculosis(TB), has been used and proven to be one of the most effective treatments for non-muscle invasive bladder cancer (BCa). However, the mechanisms of BCG action have not been completely understood, thereby limiting the improvement of BCG therapy. Vitamin D deficiency has been associated with a high risk of TB infection, and the beneficial effect of UV exposure in TB patients was proven to be mediated via activation of vitamin D signals of innate immune cells. Thus, vitamin D signals might be involved in mediating BCG immunotherapy. To test this hypothesis, we examined the impact of 1 alpha, 25-dihydroxyvitamin D3 (1,25-VD) on BCG-induced response in BCa cells and macrophage cells. Our data revealed that 1,25-VD promotes BCG-induced interleukin 8 (IL-8) secretion by BCa cells, consequently inducing the migration of macrophage, THP-1. This THP-1 cell migration promoted by 1,25-VD can be blocked by IL-8 neutralized antibody. Furthermore, 1,25-VD increased BCG-induced expression of macrophage markers in THP-1 cell, and enhanced the BCG-induced THP-1 cytotoxicity against low-grade BCa cells. Importantly, a pre-clinical trial using the N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN)-induced BCa mouse model revealed that intravesical co-treatment of 1,25-VD with BCG can prolong mice survival. These data demonstrate a novel mechanism by which 1,25-VD promotes BCG-mediated anti-BCa pathways and provides a platform for improving BCG efficacy with combination of 1,25-VD.

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1,25-VD promotes THP-1 cell migration through IL-8(A) 1,25-VD promotes BCG induced THP-1 cell migration by a transwell migration assay. TCC-SUP cells were treated with vehicle (ethanol), 100 nM 1,25-VD, 2 × 105 CFU BCG, or in combination of 1,25-VD and BCG. Culture media were replaced with fresh media after 2 hours' treatment of drugs, then cells were cultured for another 22 hours. The conditioned media were collected and loaded in the lower chamber, and Thp-1 cells after serum-free starvation were added to transwell insert with 5 μm pore. After 24 hours the infiltrating THP-1 cells in the lower chamber were collected and counted. (B) Blockage of IL-8 by neutralized antibody reverses 1,25-VD-induced THP-1 migration. A transwell migration assay was performed as described in A, except adding IL-8 neutralizing antibody (20 μg/ml) with BCG and BCG+1,25-VD treated TCC-SUP cells. For statistical analysis, differences in mean values between two groups were analyzed by two-tailed Student's test. Results are the mean ± SEM of three experiments.
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Figure 3: 1,25-VD promotes THP-1 cell migration through IL-8(A) 1,25-VD promotes BCG induced THP-1 cell migration by a transwell migration assay. TCC-SUP cells were treated with vehicle (ethanol), 100 nM 1,25-VD, 2 × 105 CFU BCG, or in combination of 1,25-VD and BCG. Culture media were replaced with fresh media after 2 hours' treatment of drugs, then cells were cultured for another 22 hours. The conditioned media were collected and loaded in the lower chamber, and Thp-1 cells after serum-free starvation were added to transwell insert with 5 μm pore. After 24 hours the infiltrating THP-1 cells in the lower chamber were collected and counted. (B) Blockage of IL-8 by neutralized antibody reverses 1,25-VD-induced THP-1 migration. A transwell migration assay was performed as described in A, except adding IL-8 neutralizing antibody (20 μg/ml) with BCG and BCG+1,25-VD treated TCC-SUP cells. For statistical analysis, differences in mean values between two groups were analyzed by two-tailed Student's test. Results are the mean ± SEM of three experiments.

Mentions: The pro-inflammatory cytokine secretion by BCa cells following BCG instillation would consequently induce innate immune cells infiltration into the bladder wall, and this first wave immune response is required for a successful BCG therapy [30]. Therefore, the 1,25-VD induction of IL-8 secretion by BCa cells could then promote BCG-induced innate immune cell migration. To test this, we performed a transwell migration assay whereby we treated TCC-SUP cells with vehicle control, 1,25-VD, BCG, and BCG+1,25-VD for two hours, then replaced with culture media for 22 hours incubation. Afterwards, the conditioned media that served as chemo-attractant sources were collected for migration of monocyte/macrophage THP-1 cells. We found that BCG treatment induced more THP-1 cell migration than vehicle control. Importantly, BCG+1,25-VD treatment induced statistically more THP-1 migration than BCG alone (Fig. 3 upper panel).


1 alpha, 25-dihydroxylvitamin D3 promotes Bacillus Calmette-Guérin immunotherapy of bladder cancer.

Hsu JW, Yin PN, Wood R, Messing J, Messing E, Lee YF - Oncotarget (2013)

1,25-VD promotes THP-1 cell migration through IL-8(A) 1,25-VD promotes BCG induced THP-1 cell migration by a transwell migration assay. TCC-SUP cells were treated with vehicle (ethanol), 100 nM 1,25-VD, 2 × 105 CFU BCG, or in combination of 1,25-VD and BCG. Culture media were replaced with fresh media after 2 hours' treatment of drugs, then cells were cultured for another 22 hours. The conditioned media were collected and loaded in the lower chamber, and Thp-1 cells after serum-free starvation were added to transwell insert with 5 μm pore. After 24 hours the infiltrating THP-1 cells in the lower chamber were collected and counted. (B) Blockage of IL-8 by neutralized antibody reverses 1,25-VD-induced THP-1 migration. A transwell migration assay was performed as described in A, except adding IL-8 neutralizing antibody (20 μg/ml) with BCG and BCG+1,25-VD treated TCC-SUP cells. For statistical analysis, differences in mean values between two groups were analyzed by two-tailed Student's test. Results are the mean ± SEM of three experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3926835&req=5

Figure 3: 1,25-VD promotes THP-1 cell migration through IL-8(A) 1,25-VD promotes BCG induced THP-1 cell migration by a transwell migration assay. TCC-SUP cells were treated with vehicle (ethanol), 100 nM 1,25-VD, 2 × 105 CFU BCG, or in combination of 1,25-VD and BCG. Culture media were replaced with fresh media after 2 hours' treatment of drugs, then cells were cultured for another 22 hours. The conditioned media were collected and loaded in the lower chamber, and Thp-1 cells after serum-free starvation were added to transwell insert with 5 μm pore. After 24 hours the infiltrating THP-1 cells in the lower chamber were collected and counted. (B) Blockage of IL-8 by neutralized antibody reverses 1,25-VD-induced THP-1 migration. A transwell migration assay was performed as described in A, except adding IL-8 neutralizing antibody (20 μg/ml) with BCG and BCG+1,25-VD treated TCC-SUP cells. For statistical analysis, differences in mean values between two groups were analyzed by two-tailed Student's test. Results are the mean ± SEM of three experiments.
Mentions: The pro-inflammatory cytokine secretion by BCa cells following BCG instillation would consequently induce innate immune cells infiltration into the bladder wall, and this first wave immune response is required for a successful BCG therapy [30]. Therefore, the 1,25-VD induction of IL-8 secretion by BCa cells could then promote BCG-induced innate immune cell migration. To test this, we performed a transwell migration assay whereby we treated TCC-SUP cells with vehicle control, 1,25-VD, BCG, and BCG+1,25-VD for two hours, then replaced with culture media for 22 hours incubation. Afterwards, the conditioned media that served as chemo-attractant sources were collected for migration of monocyte/macrophage THP-1 cells. We found that BCG treatment induced more THP-1 cell migration than vehicle control. Importantly, BCG+1,25-VD treatment induced statistically more THP-1 migration than BCG alone (Fig. 3 upper panel).

Bottom Line: However, the mechanisms of BCG action have not been completely understood, thereby limiting the improvement of BCG therapy.Vitamin D deficiency has been associated with a high risk of TB infection, and the beneficial effect of UV exposure in TB patients was proven to be mediated via activation of vitamin D signals of innate immune cells.This THP-1 cell migration promoted by 1,25-VD can be blocked by IL-8 neutralized antibody.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, University of Rochester, Rochester, New York, USA.

ABSTRACT
Bacillus Calmette-Guérin (BCG), a vaccine against tuberculosis(TB), has been used and proven to be one of the most effective treatments for non-muscle invasive bladder cancer (BCa). However, the mechanisms of BCG action have not been completely understood, thereby limiting the improvement of BCG therapy. Vitamin D deficiency has been associated with a high risk of TB infection, and the beneficial effect of UV exposure in TB patients was proven to be mediated via activation of vitamin D signals of innate immune cells. Thus, vitamin D signals might be involved in mediating BCG immunotherapy. To test this hypothesis, we examined the impact of 1 alpha, 25-dihydroxyvitamin D3 (1,25-VD) on BCG-induced response in BCa cells and macrophage cells. Our data revealed that 1,25-VD promotes BCG-induced interleukin 8 (IL-8) secretion by BCa cells, consequently inducing the migration of macrophage, THP-1. This THP-1 cell migration promoted by 1,25-VD can be blocked by IL-8 neutralized antibody. Furthermore, 1,25-VD increased BCG-induced expression of macrophage markers in THP-1 cell, and enhanced the BCG-induced THP-1 cytotoxicity against low-grade BCa cells. Importantly, a pre-clinical trial using the N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN)-induced BCa mouse model revealed that intravesical co-treatment of 1,25-VD with BCG can prolong mice survival. These data demonstrate a novel mechanism by which 1,25-VD promotes BCG-mediated anti-BCa pathways and provides a platform for improving BCG efficacy with combination of 1,25-VD.

Show MeSH
Related in: MedlinePlus