Effects of Vitamin D and Calcium Supplementation on Markers of Apoptosis in Normal Colon Mucosa: A Randomized, Double-Blind, Placebo-Controlled Clinical Trial

Abstract

To further clarify and/or develop calcium and vitamin D as chemopreventive agents against colorectal cancer in humans, understand the mechanisms by which these agents reduce risk for the disease, and develop "treatable" biomarkers of risk for colorectal cancer, we conducted a pilot, randomized, double-blind, placebo-controlled, 2 × 2 factorial clinical trial to test the effects of calcium and vitamin D 3 , alone and in combination on markers of apoptosis, in the normal colorectal mucosa. Ninety-two men and women with at least one pathology-confirmed colorectal adenoma were treated with 2.0 g/d calcium or 800 IU/d vitamin D 3 , alone or in combination, versus placebo over 6 months. Overall expression and colorectal crypt distributions of Bcl-2 (an apoptosis inhibitor) and Bax (an apoptosis promoter) in biopsies of normal-appearing rectal mucosa were detected by automated immunohistochemistry and quantified by image analysis. After 6 months of treatment, Bax expression along the full lengths of crypts increased 56% (P = 0.02) in the vitamin D group and 33% in both the calcium (P = 0.31) and calcium plus vitamin D (P = 0.36) groups relative to the placebo group. The vitamin D treatment effect was more pronounced in the upper 40%, or differentiation zone, of crypts (80%; P = 0.01). There were no statistically significant treatment effects on Bcl-2 expression. Overall, these preliminary results suggest that calcium and vitamin D, individually or together, may enhance apoptosis in the normal human colorectal epithelium, and the strongest treatment effects may be vitamin D related and in the upper sections of the colorectal crypts.

Colorectal Mucosal Expression of MSH2 as a Potential Biomarker of Risk for Colorectal Neoplasms

“…1) and cannot be quantified by eye (e.g., by counting cells). Therefore, MSH2 expression density, detected by immunohistochemical staining, was quantified in the stained slides using image analysis densitometry methods (22,23). The procedure was conducted by one trained "scorer" using a light microscope (Olympus BX40), digital camera (Polaroid DMC Digital Light Microscope Camera), digital drawing tablet, and a customdeveloped plug-in to ImagePro Plus (Media Cybernetics) image analysis software.…”

“…1; refs. 22,23). The software program divided the traced hemicrypt into segments corresponding in width to that of an average normal crypt epithelial cell (6.59 μm; ref.…”

“…The software program divided the traced hemicrypt into segments corresponding in width to that of an average normal crypt epithelial cell (6.59 μm; ref. 21) and then calculated overall hemicrypt-and segment-specific labeling optical density and entered these data into a Microsoft Access database along with various dimensional parameters of the hemicrypt (22,23).…”

Whole Slide Imaging for Analytical Anatomic Pathology and Telepathology: Practical Applications Today, Promises, and Perils

“…58 Investigators in our department have also used IA to measure apoptosis in colonic mucosa by using the apoptosis inhibitor Bcl-2 and the apoptosis promoter Bax. 59 We have also used IA in an on-slide ''flow cytometry''-type approach to quantitate inflammatory cells in autoimmune pancreatitis, 39 and it is possible that such methods may become important in quantitating inflammatory cells in a variety of applications (eg, tumor-infiltrating lymphocytes in gastrointestinal tract cancers). These diverse applications demonstrate the many possibilities of WSI IA.…”

Oral cancer risk and vitamin D status, intake and supplementation: A review

“…The 1,25(OH)2D-VDR-RXR complex binds to vitamin D-response elements in DNA which operate to initiate gene transcription. Activation of the VDR by 1,25(OH) 2 D can restore or enhance proapoptotic effects in different cancer cells through transcriptional activation of bax and p-calpain, two effective proapoptotic proteins 35,36 . VDR-vitamin D activation also been demonstrated to increase mRNA expression of transforming growth factor, a potent antiproliferative cytokine in normal and early stage cancer cells; superoxide dismutase, which may reduce oxidative stress-induced DNA damage and loss to DNA repair mechanisms that contribute to carcinogenesis and inflammatory cytokine production; as well as cyclin-dependent kinase (CDK) inhibitor p21, RBL2, RBLP6 and forkhead box O (FOXO) tumour suppressors that function to counteract MAPK-mediated phosphorylation and growth 35,[37][38][39] .…”