Curcumin and Turmeric Modulate the Tumor-Promoting Effects of Iron In Vitro

Abstract

Free or loosely chelated iron has tumor-promoting properties in vitro. Curcumin, a polyphenol derived from the food spice turmeric (Curcuma longa), is a potent antioxidant that binds iron. The primary aim of this study was to investigate whether curcuminoids prevent tumor-promoting effects of iron in T51B cells, a non-neoplastic rat liver epithelial cell line. Purified curcuminoids (curcumin) or a standardized turmeric extract similarly reduced oxidative stress and cytotoxicity associated with iron overload (IC₅₀ values near 10 μM, P < 0.05). Inhibition of iron-induced tumor promotion (seen upon treatment with 200 μM ferric ammonium citrate ± curcumin/turmeric for 16 wk in culture; subsequently assayed by soft agar colony formation) was nearly complete at 20 μM of total curcuminoids (P < 0.05), a concentration predicted to only partially chelate the added iron. Surprisingly, lower curcumin concentrations (10 μM) increased tumor promotion (P < 0.01). Curcuminoids delivered as a standardized turmeric extract were taken up better by cells, had a longer half-life, and appeared more effective in blocking tumor promotion (P < 0.01), suggesting enhanced curcuminoid delivery to cells in culture. The primary finding that curcuminoids can inhibit tumor promotion caused by iron in T51B cells is tempered by evidence for an underlying increase in neoplastic transformation at lower concentrations.

Figure 1

Structures of curcumin and related curcuminoids.

Figure 2

Curcuminoids prevent iron-induced lipid peroxidation. Confluent T51B cells were treated for 2 h ± 200 μM iron and ±10 μM 8-hydroxyquinoline (8HQ) as indicated. Curcuminoids were added with iron/8HQ as either purified curcuminoids (curc at 2, 10, or 50 μM) or a standardized turmeric extract (turm at 2, 10, or 50 μM). Cell lysates were analyzed by TBARS assay and results expressed in MDA equivalents. Means ± SE from 4 replicate experiments are presented. (***P < 0.001 vs. iron + 8HQ reference (ref.); **P < 0.01; *P < 0.05.)

Figure 3

Curcuminoids reduce acute iron cytotoxicity. Subconfluent, proliferating T51B cells were treated for 2 days ± 200 μM iron and ± 10 μM 8-hydroxyquinoline (8HQ) as indicated. Curcuminoids were added with iron/8HQ as either purified curcuminoids (curc at 5, 10, 20, or 50 μM) or a standardized turmeric extract (turm at 5, 10, 20, or 50 μM). Cell number and viability were determined by MTT assay. Means ± SE from 3 replicate experiments are presented. (***P < 0.001 vs. iron + 8HQ reference (ref.); **P < 0.01; *P < 0.05.)

Figure 4

Curcuminoids reduce the cytotoxicity of chronic iron loading. Proliferating T51B cells were treated for 5 days ± 500 μM iron. Curcuminoids were added with iron as either purified curcuminoids (curc at 2, 5, 10, 20, or 50 μM) or a standardized turmeric extract (turm at 2, 5, 10, 20, or 50 μM). Cell number and viability were determined by MTT assay. Means ± SE from 3 replicates are presented. (**P < 0.01 vs. 500 μM iron only reference (ref.); *P < 0.05.)

Figure 5

Curcuminoids inhibit tumor promotion by iron. Proliferating T51B cells were treated with 0.5 μg/ml MNNG for 24 h (initiated) and cultured for 16 wk with 200 μM iron and ±curcuminoids or turmeric (≥20 μM) as described under “Methods” section. Control cells (no MNNG or iron) were run in parallel. Colony formation in soft agar was measured and results compiled from three independent experiments. The total number of soft agar dishes and treatment conditions were as follows: Control (n = 24 total); MNNG + iron (n = 75 total); purified curcuminoids (curcumin; n = 60 total: 16 at 20 μM, 16 at 30 μM, and 28 at 50 μM); and turmeric (n = 32 total: 16 at 20 μM and 16 at 30 μM). (***P < 0.001 vs. MNNG + iron; **P < 0.01; *P < 0.05.)

Figure 6

Inhibition of tumor promotion by curcuminoids through 20 wk. Proliferating T51B cells were initiated with MNNG and cultured with 200 μM iron and ±20 μM curcuminoids or turmeric with media renewal every 2.6 days on average. Colony formation in soft agar was assessed at 16, 18, and 20 wk. The number of soft agar dishes in this experiment was: Control (n = 8 each at weeks 16–20); MNNG + iron (n = 40 each at weeks 16–20); 20 μM purified curcuminoids (curcumin; n = 8 each at weeks 16–20); 20 μM turmeric (n = 8 each at weeks 16–20).

Figure 7

The in vitro pharmacokinetic properties of purified curcuminoids differ from those of the turmeric extract. Confluent T51B cells were treated at time zero with 200 μM iron and 20 μM curcuminoids, given either as purified curcuminoids (circles) or standardized turmeric extract (squares). Curcuminoid levels in the cells at the indicated times were determined by absorbance at 425 nm as described under “Methods” section. The data represent means and SE from n = 6 independent experiments, each set with a best fit line drawn using a one-phase decay model.