Relative Uptake of Tomato Carotenoids by In Vitro Intestinal and Prostate Cancer Cells

Abstract

Background: Consumption of tomatoes and tomato carotenoids is associated with a reduced risk of prostate cancer. Prostate tissue accumulates tomato carotenoids, including lycopene, β-carotene, and phytoene. Phytoene accumulation is relatively greater in the prostate than that of lycopene, but the metabolic determinants of tissue carotenoid profiles are poorly understood.

Objectives: The purpose of this study was to determine if differences in stability, cellular uptake, and clearance of phytoene compared with lycopene or β-carotene by prostate and intestinal cells may explain differences in observed tissue carotenoid profiles.

Methods: Gene and protein expression for carotenoid metabolism in prostate cell lines were analyzed by qRT-PCR and Western blot, respectively. Uptake, efflux, and clearance of phytoene, lycopene, or β-carotene by prostate cell [LNCaP (Lymph Node Carcinoma of the Prostate cell line), RWPE-1 (a human prostate epithelial cell line), and PC-3 (aprostate cancer cell line)] and absorptive enterocyte (Caco-2) cultures were compared. The effect of scavenger receptor class B member 1 (SCARB1) inhibition on carotenoid uptake by LNCaP, RWPE-1, and Caco-2 cells was tested.

Results: SCARB1 was expressed across prostate cell lines. Lycopene, phytoene, and β-carotene uptakes were similar in LNCaP and PC-3 cells, whereas RWPE-1 cells absorbed a smaller portion of the phytoene dose than lycopene or β-carotene doses. The clearance rates of carotenoids from LNCaP cells did not differ. Intestinal cell uptake of phytoene was greatest, followed by β-carotene and lycopene. SCARBI inhibitor treatment did not significantly reduce the uptake or efflux of carotenoids by LNCaP or Caco-2 cells at the dose concentration provided.

Conclusions: Overall, this study suggests that greater bioavailability at the point of the intestine and greater stability of phytoene are determinants of the relative enrichment of phytoene in prostate tissue.

Keywords: Caco-2; SCARB1; lycopene; phytoene; β-carotene.

FIGURE 1

Recovery of lycopene, phytoene, and β-carotene from human serum over a 24 h incubation (A). (Significant differences were α = 0.050. n = 15, error bars = SD). Distribution of lycopene, phytoene, β-carotene, and α-tocopherol within lipoprotein fractions. Supplemented α-tocopherol levels were calculated by subtracting endogenous α-tocopherol measured in nontocopherol treated serum, n = 3 (B). Relative distribution of endogenous α-tocopherol in lycopene, phytoene, and β-carotene enriched serum lipoprotein fractions, n = 9 (C).

FIGURE 2

Stability of carotenoids over a 24-h preincubation in serum and 8 h in cell-free PC-3 medium. Recovery of carotenoids from serum at 0 h (1-way ANOVA; P > 0.05) (A). The concentration of carotenoids in the cell-free medium at 0 h (1-way ANOVA; P > 0.05) (B). Percent of carotenoid recovered from serum-supplemented media over 8 h, relative to amount present at 0 h (2-way repeated measures ANOVA, P_carotenoid ≥ 0.05; P_time < 0.001) (C). The concentration of carotenoids measured from serum-supplemented media over 8 h (2-way repeated measures ANOVA on log10 transformed data, P_carotenoid = 0.04 (no post hoc diffs.); P_time < 0.001, post hoc diffs shown with lowercase letters) (D). Points represent the mean of n = 3, error bars = SD. Samples analyzed in duplicate. ANOVA, analysis of variance.

FIGURE 3

Gene and protein expression of carotenoid and lipid metabolic genes in prostate cell lines maintained in routine culturing conditions. Gene expression (A, B) is the average of n = 3 samples analyzed in duplicate, note different y-axis scales, bars represent the standard error of n = 3. Gene expression <2.9 × 10^–11 was considered below detection. Protein expression of SCARB1 (n = 3) (C). SCARB1, scavenger receptor class B member 1.

FIGURE 4

Carotenoid uptake by PC-3 (A, D, G), LNCaP (B, E, H), RWPE-1 (C, F, I) cells after 6 h expressed in ng per 1 × 10⁶ cells (A, B, C), and by % of dose per 1 × 10⁶ cells (D, E, F). Carotenoid stability in cell-free media for PC-3 (a), LNCaP (b), and RWPE-1 (c) cell lines after 6 h expressed as % of the dose. Different lowercase letters above bars indicate significant differences (P < 0.05). Error bars are SD. n = 5 trials analyzed in triplicate.

FIGURE 5

Carotenoid clearance time course from LNCaP cells over 48 h (n = 5 trials). Starting media concentrations of carotenoids (A), time courses of the percent of starting amount of lycopene (B), β-carotene (C), and phytoene (D) recovered from LNCaP cells over time. Time course data represent the mean ± SD of 5 trials. The were no significant differences by carotenoid species or time on the percent of baseline cellular carotenoids remaining as analyzed by repeated measures ANOVA.ANOVA, analysis of variance.

FIGURE 6

Carotenoid uptake (A, C) and absorption (B, D) to the basolateral compartment by Caco-2 cells over measured at 4 and 24 h. The effect of time and carotenoid species was tested using 2-way ANOVA. Significant differences in carotenoid uptake by carotenoid species are denoted by bars and different lowercase letters. There was no difference in uptake between time points. There was no significant difference in carotenoid absorption to the basolateral media compartment by carotenoid species or by time point. ANOVA, analysis of variance.