Thyroid and neurotoxicity potentials of synthetic musk compounds (MK, HHCB, AHTN) in early-life stage of zebrafish (Danio rerio)

Abstract

Synthetic musk compounds (SMCs) are widely used in many applications, including consumer chemical products such as perfumes, cosmetics, soap, and fabric softener, to replace perfumes of natural origin. The potential consequences of persistence in the environment and accumulation in the biota are of concern. Their toxicities, however, are generally unknown in the aquatic environment, especially for the thyroid and neurotoxicity. In the present study, three frequently detected SMCs of musk ketone (MK), 1,3,4,6,7,8-hexahydro-4,6,6,7,8,8-hexamethyl-cyclopenta[g]- benzopyran (HHCB), and 6-acetyl-1,1,2,4,4,7-hexamethyltetralin (AHTN), were chosen, and their thyroid disruption and neurotoxicity were assessed using early-life stage zebrafish (Danio rerio). Zebrafish embryos were exposed to MK, HHCB, and AHTN for 5 days. At the end of the exposure, whole body larvae were collected to measure thyroid hormone, transcriptional change, and behavioral change. Daniovision® Ethovision XT was used to determine behavioral changes. Following 120 hours of exposure to synthetic musks, significant changes in the expression of thyroid hormone and neurodevelopment related genes were observed in the zebrafish larvae. After exposure to MK, T4 was significantly decreased, and thyroid hormone-related genes such as crhβ were significantly up-regulated. But ugt1ab was significantly down-regulated, possibly explaining the observed decrease of whole-body T4 concentration. Following MK exposure, behavior changes were decreased. After exposure to HHCB, T4 concentration was also decreased. crhβ gene showed an up-regulating trend, and mbp, gap43, and syn2a were down-regulated. HHCB exposure caused hypoactivity of the larvae was observed. Following AHTN exposure, up-regulations of crhβ, nis, ugt1ab, dio2, and gap43 genes were observed, while thyroid hormones were not influenced. Thyroid disruption effects of ATHN appear to be different from the other SMCs. After light stimulation, zebrafish larvae activity was decreased, and thigmotaxis had no effects. The present observations showed that SMCs such as MK and HHCB, have the potentials to disrupt thyroid hormone or neurodevelopment in the zebrafish at the development stage. This study was conducted at concentrations several orders of magnitude higher than the environmental concentrations. Consequences of long-term exposure at the environmentally relevant concentrations in the aquatic environment warrant further investigations.

합성머스크류는 천연향료의 대체 목적으로 향수, 화장품, 비누, 섬유유연제와 같은 생활화학제품 및 위생용품 등 광범위한 분야에서 사용되고 있다. 합성머스크류는 일반적으로 친지질성 물질이기 때문에 환경 중 잔류성과 체내 축적 가능성이 있어 주의가 필요하다. 하지만, 현재 합성머스크류에 대한 내분비 및 신경독성 관련 지식은 부족하다. 제브라피쉬 배아(수정 후 4시간 미만)를 5일간 musk ketone(MK), 1,3,4,6,7,8-hexahydro-4,6,6,7,8,8-hexamethyl-cyclopenta[g]-benzopyran(HHCB), 6-Acetyl-1,1,2,4,4,7-hexamethyltetralin(AHTN)에 노출시킨 후 갑상선 호르몬을 측정하고, 갑상선 및 신경관련 유전자 수준 변화를 살펴보았고 Daniovision® Ethovision XT를 이용하여 물고기의 행동변화를 관찰하였다. 합성머스크류 노출은 제브라피쉬의 갑상선호르몬, 신경발달 및 행동에 유의한 변화를 초래하였다. MK와 HHCB에 노출된 후 T4농도는 감소하였다. 갑상선 관련 유전자 중 crhβ는 유의하게 증가하였으나 뇌하수체와 갑상선의 조절유전자는 이에 상응하지 않았다. 이 두가지 물질 노출에 의해 관찰된 ugt1ab의 전사 감소가 갑상선호르몬의 저하를 설명하는 것으로 추측된다. 또한, 신경발달 관련 유전자인 mbp, gap43, 또는 c-fos 등의 전사가 유의하게 감소하였다. 행동학적 변화에서도 농도의존적으로 이동거리가 감소하는 패턴을 보였다. AHTN에 노출된 치어도 T4가 감소하는 경향성이 나타났다. 그러나 유전자 발현의 변화를 살펴본 결과 갑상선호르몬 교란의 기전과 신경발달에 초래하는 영향의 방향이 다른 물질과 상이한 것으로 관찰되었다. 제브라피쉬 치어 노출 결과 MK, HHCB는 갑상선 및 신경발달에 영향을 미치나, AHTN은 상대적으로 갑상선에만 영향을 미치는 것을 확인하였다. 합성머스크류에 노출된 생애초기발달단계의 제브라피쉬는 갑상선 호르몬 조절 및 신경발달 독성을 일으킬 수 있음을 확인하였다. 하지만, 고농도에서 단시간 노출로 인한 독성영향을 확인하였기에, 환경 중 농도에서의 장기노출로 인한 독성연구가 필요하다.