Nitromusk and polycyclic musk compounds as long-term inhibitors of cellular xenobiotic defense systems mediated by multidrug transporters

Abstract

Synthetic musk compounds, widely used as fragrances in consumer products, have been detected in human tissue and, surprisingly, in aquatic organisms such as fish and mollusks. Although their persistence and potential to bioaccumulate are of concern, the toxicity and environmental risks of these chemicals are generally regarded as low. Here, however, we show that nitromusks and polycyclic musks inhibit the activity of multidrug efflux transporters responsible for multixenobiotic resistance (MXR) in gills of the marine mussel Mytilus californianus. The IC(subscript)10(/subscript) (concentration that inhibits 10%) values for the different classes of musks were in the range of 0.09-0.39 microM, and IC(subscript)50(/subscript) values were 0.74-2.56 microM. The immediate consequence of inhibition of efflux transporters is that normally excluded xenobiotics will now be able to enter the cell. Remarkably, the inhibitory effects of a brief 2-hr exposure to musks were only partially reversed after a 24- to 48-hr recovery period in clean seawater. This unexpected consequence of synthetic musks--a long-term loss of efflux transport activity--will result in continued accumulation of normally excluded toxicants even after direct exposure to the musk has ended. These findings also point to the need to determine whether other environmental chemicals have similar long-term effects on these transporters. The results are relevant to human health because they raise the possibility that exposure to common xenobiotics and pharmaceuticals could cause similar long-term inhibition of these transporters and lead to increased exposure to normally excluded toxicants.

Figure 1. Correlation between basal transporter activity [F(basal)] and relative increase in fluorescence in mussel gill tissue with transporter activity inhibited maximally versus basal transporter activity [R_{F(max)/F(basal)}]. y = −0.07x + 3.93.

Figure 2. Dose–effect curves for in vivo inhibition of MXR transporters in mussel gill tissue by nitromusks [MX (A) and MK (B)], polycyclic musks [HHCB (C), ADBI (D), AHTN (E), and ATII (F)], and reference inhibitors [verapamil (G) and quinidine (H)]. See “Materials and Methods” for details. Each curve represents data (obtained by probit regression) from an individual animal.

Figure 3. Inhibitory effects by musks as single compounds and in combinations as measured by retention of rhodamine B (see “Materials and Methods” for details). Bars represent percent increase (mean ± SD) of fluorescence versus respective controls; n = 7 for experiments with solvent, 1 μM MX, 1 μM MK, 1 μM HHCB, and 1 μM ADBI; n = 3 for MX + HHCB, MX + MK, HHCB + ADBI, and MX + MK + HHCB + ADBI. Brackets indicate experiments that are compared.

Figure 4. Long-term inhibition of efflux transporters by nitromusks [MX (A) and MK (B)], polycyclic musks [HHCB (C), ADBI (D), AHTN (E), and ATII (F)], and reference inhibitors [verapamil (G) and quinidine (H)]. Bars represent fluorescence versus respective controls (mean % ± SD); n = 3 (11 for verapamil). : *p < 0.05 by paired t-test.

Figure 5. Correlation of MXR inhibitory efficiency (indicated by IC₅₀ values) and log K_ow of test compounds. Abbreviations: QUI, quinidine; VER, verapamil. For the calculation of the regression curve, (with 95% confidence intervals) only musk compounds were included.