Uptake and Immunomodulatory Properties of Betanin, Vulgaxanthin I and Indicaxanthin towards Caco-2 Intestinal Cells

Abstract

The present study aimed to compare the absorption and transport patterns of three main betalains, betanin, vulgaxanthin I and indicaxanthin, into intestinal epithelial cells and to assess their distinct molecular effects on inflammatory and redox-related cell signalling in association with their radial scavenging potencies. All three betalains showed anti-inflammatory effects (5–80 μM), reflected by attenuated transcription of pro-inflammatory mediators such as cyclooxygenase-2 and inducible NO-synthase. Concomitant increases in antioxidant enzymes such as heme oxygenase-1 were only observed for betanin. Moreover, betanin uniquely demonstrated a potent dose-dependent radical scavenging activity in EPR and cell-based assays. Results also indicated overall low permeability for the three betalains with Papp of 4.2–8.9 × 10−7 cm s−1. Higher absorption intensities of vulgaxanthin and indicaxanthin may be attributed to smaller molecular sizes and greater lipophilicity. In conclusion, betanin, vulgaxanthin I and indicaxanthin have differentially contributed to lowering inflammatory markers and mitigating oxidative stress, implying the potential to ameliorate inflammatory intestinal disease. Compared with two betaxanthins, the greater efficacy of betanin in scavenging radical and promoting antioxidant response might, to some extent, compensate for its poorer absorption efficiency, as demonstrated by the Caco-2 cell model.

Redox Properties, Bioactivity and Health Effects of Indicaxanthin, a Bioavailable Phytochemical from Opuntia ficus indica, L.: A Critical Review of Accumulated Evidence and Perspectives

“…In other studies [ 59 ], 5–80 μM Indicaxanthin down-regulated a number of redox-dependent, pro-inflammatory factors including NOX-1, IL-6, IL-8, COX2, and NOS in Caco-2 cells exposed to a cocktail of cytokines (TNF-α, IL-1β, IFN-γ). The phytochemical, instead, appeared poorly effective in decreasing ROS in the same cells treated with 200 μM H 2 O 2 , while EPR measurements carried out in parallel provided evidence that Indicaxanthin did not possess direct radical scavenging capacity against superoxide [ 59 ], in accordance with other observations [ 32 ].…”

“…Molecular mass and physico-chemical characteristics (electric charge, lipophilicity, Section 3.2 ) appeared to be essential to the transport, by paracellular diffusion, with a high permeability coefficient (P = 4.4 × 10 −6 cm s −1 ), at pH 6.0, in a concentration-dependent mode, without involvement of H + -dependent carriers, nor efflux transporters [ 60 ]. Other studies report a lower coefficient (P = 8.9 × 10 −7 cm s −1 ) at pH 6.5 [ 59 ]. Overall, the molecule’s high resistance to the digestive steps, and easy of diffusion at the intestinal epithelium account for the high bioavailability of Indicaxanthin from fruits of cactus pear [ 31 ].…”

“…Thanks to its amphiphilicity, the molecule can cross cell membranes and enters cells. Indicaxanthin was recovered in the soluble fraction of red blood cells, after spiking of human plasma [ 29 ]; likewise, uptake was observed after incubation with intestinal-like differentiated Caco-2 cells [ 32 , 59 ]. In addition, Indicaxanthin entered the cells of mouse ileum organ preparations [ 34 , 45 ], of C. elegans [ 47 , 48 ] and, remarkably, was distributed in several specific areas of rat brain after oral administration of amounts consistent with diet [ 37 ].…”

“…The models presented in the studies above [ 32 , 33 , 59 ] reveal interesting issues concerning the potential of Indicaxanthin to prevent oxidative damage in healthy cells: Indicaxanthin (i) does not act as a scavenger of reactive oxygen species; (ii) prevents agonist-induced ROS generation by inhibiting enzymes of the NOX family. The membrane microenvironment has been shown essential in modulating the activity of NOX enzymes [ 104 , 115 ].…”

“…Ultimately, a cell-independent procedure showing that Indicaxanthin inhibited the total nuclear DNMT, and an in silico computational modelling analysis revealing a stable binding of the phytochemical at the DNMT1 catalytic site [ 46 ], suggest that Indicaxanthin could block the DNA methylation process operating at a gene-specific level [ 177 ]. Intracellular uptake and accumulation of Indicaxanthin in Caco-2 cells [ 32 , 59 ], as a necessary precondition for this activity have been observed. Redox regulation and signaling may be upstream of epigenetic modulation [ 172 ]; direct inhibition of methylating enzymes could explain the activity of Indicaxanthin in proliferating Caco-2 cells where the phytochemical did not appear to alter the redox tone [ 42 ].…”