Docosahexaenoic Acid and Butyrate Synergistically Induce Colonocyte Apoptosis by Enhancing Mitochondrial Ca2+ Accumulation

Abstract

We have previously shown that butyrate, a short-chain fatty acid fiber fermentation product, induces colonocyte apoptosis via a nonmitochondrial, Fas-mediated, extrinsic pathway. Interestingly, fermentable fiber when combined with fish oil containing docosahexaenoic acid (DHA, 22:6n-3) exhibits an enhanced ability to induce apoptosis and protect against colon tumorigenesis. To determine the molecular mechanism of action, the effect of DHA and butyrate cotreatment on intracellular Ca 2+ homeostasis was examined. Mouse colonocytes were treated with 50 Mmol/L DHA or linoleic acid (LA) for 72 h F butyrate (0-10 mmol/L) for the final 24 h. Cytosolic and mitochondrial Ca 2+ levels were measured using Fluo-4 and Rhod-2. DHA did not alter basal Ca 2+ or the intracellular inositol trisphosphate (IP 3 ) pool after 6 h butyrate cotreatment. In contrast, at 12 and 24 h, DHA-and butyrate-treated cultures exhibited a 25% and 38% decrease in cytosolic Ca 2+ compared with LA and butyrate. Chelation of extracellular Ca 2+ abolished the effect of thapsigargin on the IP 3 -releasable Ca 2+ pool. DHA and butyrate cotreatment compared with untreated cells increased the mitochondrial-to-cytosolic Ca 2+ ratio at 6, 12, and 24 h by 73%, 18%, and 37%, respectively. The accumulation of mitochondrial Ca 2+ preceded the onset of apoptosis. RU-360, a mitochondrial-uniporter inhibitor, abrogated mitochondrial Ca 2+ accumulation and also partially blocked apoptosis in DHA and butyrate cotreated cells. Collectively, these data show that the combination of DHA and butyrate, compared with butyrate alone, further enhances apoptosis by additionally recruiting a Ca 2+

Synergy between docosahexaenoic acid and butyrate elicits p53-independent apoptosis via mitochondrial Ca2+accumulation in colonocytes

“…We used isogenic p53 wild-type and deficient human colon tumor (HCT-116) cell lines to determine whether chemoprotective nutrients modulate intracellular Ca 2ϩ compartmentalization to induce colonocyte apoptosis. The results confirm and extend our previous observations (36,42) and demonstrate that DHA and butyrate synergistically enhance mitochondrial Ca 2ϩ accumulation, which serves as a trigger for apoptosis in a p53-independent manner. Consistent with previous observations (12,29,30,54), our data demonstrate that pleiotropic bioactive components generated by fermentable fiber (butyrate) and fish oil (DHA) act synergistically to enhance apoptosis.…”

“…After we recently demonstrated using young adult mouse colonocytes that DHA and butyrate combination synergistically alters intracellular Ca 2ϩ compartmentalization by enhancing mitochondrial Ca 2ϩ accumulation through a storeoperated channel-mediated mechanism (36), in this study our primary objective was to corroborate these novel findings utilizing human colonocyte cell lines and an in vivo rat model system. Since mitochondria and the p53 tumor suppressor gene play a critical role in damage-induced apoptosis (52, 57), we postulated that the combination of DHA and butyrate significantly enhances mitochondrial Ca 2ϩ accumulation and apoptosis in a p53-dependent fashion.…”

“…Images were collected via a ϫ20 objective (0.75 numerical aperture) and a ROPER CoolSnap HQ camera. Fluo-4 calibration was performed using the Calcium Calibration buffer kit 2 (Molecular Probes) as previously described (36). In select experiments, MitoTracker Green FM was used to confirm the subcellular localization of rhod-2 fluorescence (10).…”

Aryl Hydrocarbon Receptor Activity of Tryptophan Metabolites in Young Adult Mouse Colonocytes

“…The YAMC cell line was initially generated from the Immorto mouse (Whitehead et al, 1993) and has been previously used in our studies (Kolar et al, 2007;Weige et al, 2009;Turk et al, 2011). Cells were maintained in RPMI 1640 medium with 5% fetal bovine serum, 5 units/ml mouse interferon-g (IF005) (EMD Millipore, Billerica, MA), 1% ITS "2" minus (insulin, transferrin, selenium) (41-400-045; Life Technologies, Grand Island, NY) at 33°C (permissive conditions).…”

Changes in Korean Adult Females’ Intestinal Microbiota Resulting from Kimchi Intake

“…It also suppresses histone deacetylation and cancer cell proliferation by regulating the expression of genes, such as p21. In addition, it induces apoptosis with Ca 2+ accumulation in mitochondria [26][27][28]. The ecosystem of intestinal microorganisms also shows correlations with obesity [13,17].…”