Vitamin D and breast cancer prevention

Yan Jiang, Purdue University

Abstract

Evidence suggests that high vitamin D status (serum 25 hydroxyvitamin D, 25(OH)D) is associated with reduced risk of breast cancer. Since hypoxia responses in breast epithelial cells during carcinogenesis and epithelial cell architecture in mammary gland are so important in breast cancer initiation, current research focused on how 1α, 25-dihyoxyvitamin D (1,25(OH) 2D) regulates hypoxia-inducible factor-1α (HIF-1α), a master transcription factor which mediates hypoxia responses, and apical polarity, a structure in breast epithelial cells that is essential for mammary tissue function. A multistage carcinogenesis model, based on untransformed breast epithelial cell line (MCF10A cells), for breast cancer was used to investigate 1,25(OH)2D regulation of HIF-1α protein. The cells employed were MCF10A cells untransformed breast epithelial cells and Harvey- ras oncogene transfected breast cancer cells (MCF10A-ras cells). In MCF10A cells, treatment with1,25(OH)2D (10 nM) significantly increased both mRNA and protein levels of HIF-1α compared to vehicle. However, in H-ras transfected MCF10A cells, 1,25(OH) 2D treatment induced a transient increase of HIF-1α protein level at 12 hrs, with no change in mRNA level. The vitamin D receptor protein (VDR) levels between these two cell lines were not significantly different, but basal and 1,25(OH)2D stimulated 1α-hyoxylase protein levels, which converts 25(OH)D to 1,25(OH)2D, were significantly higher in H-ras transfected MCF10A cells compared to MCF10A cells. A transcription inhibitor prevented the 1,25(OH)2D induction of HIF-1α protein and mRNA levels in MCF10A cells but failed to alter the induction of HIF-1α protein level in H-ras transfected MCF10A cells. Inhibition of proteasomal degradation prevented the 1,25(OH) 2D-induced HIF-1α protein level in H-ras transfected MCF10A but not in MCF10A cells. Therefore, 1,25(OH)2D regulates HIF-1α protein level via transcriptional regulation in MCF10A cells which is VDR dependent. A putative VDR response element (VDRE) was identified in the HIF-1α gene promoter (-2910 to -2896) by using chromatin immunoprecipitation assay in MCF10A cells. In contrast, 1,25(OH)2D altered HIF-1α protein degradation independent of the VDR in H-ras transfected MCF10A cells. VDR transcriptional activity is reduced in H-ras transfected MCF10A cells compared to MCF10A cells. Extracellular receptor kinase (ERK) signaling is involved in 1,25(OH)2D-induced HIF-1α protein in MCF10A-ras cells. In addition, mRNA level of prolyl hydroxylases (PHD) was up-regulated by 1,25(OH)2D treatment in MCF10A cells but down-regulated in MCF10A-ras cells, indicating that PHD may play a role in 1,25(OH)2D regulation on HIF-1α protein in both MCF10A and MCF10A-ras cells. These results suggest that 1,25(OH)2D up-regulates HIF-1α gene transcription through the VDR in MCF10A cells. Further, in the presence of H-ras oncogene, the VDR transcriptional activity was reduced and 1,25(OH) 2D increases HIF-1α protein stability through ERK signaling. On the other hand, we investigated the influences of hormones that control vitamin D status (1,25(OH)2D and parathyroid hormone (PTH)), on apical polarity in differentiated mammary epithelial cells (S1 cells). Preliminary results indicated the w-6 fatty acid, arachidonic acid (AA) induced the loss of apical polarity in the S1 cells in 3-D culture. In the current study, the downstream product of AA, prostaglandin E2 (PGE2), not AA was observed to decrease apical polarity, however treatment with 1,25(OH)2D did not alter the effect of AA or PGE2 in the S1 cells in 3-D culture. In addition, PTH, which is down-regulated with higher vitamin D status, is not toxic to S1 cell growth. PTH treatment (1 nM and 10 nM) did not affect either apical polarity or basal polarity or the S1 cells proliferation. Thus, hormones regulated by vitamin D status did not prevent the loss of apical polarity induced by PGE2. In conclusion, better vitamin D status may enhance HIF-1α protein level in untransformed breast tissue, but not in cells in the stage of cancer progression. Potential alternative pathways for vitamin D signaling may exist and play a role in vitamin D function in breast cancer development. Additionally, hormones regulated by vitamin D status do not affect breast epithelial cells apical polarity. Overall, our data support the hypothesis that high vitamin D status alters hypoxia responses and potentially angiogenesis with the presence of carcinogenesis-related molecular changes (i.e. H-ras transformation), but may not cause a negative effect on normal breast tissue or epithelial structure. Therefore vitamin D may be a good chemopreventive agent.

Degree

Ph.D.

Subject Area

Nutrition|Oncology

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