An efficient autometallography approach to localize lead at ultra-structural levels of cultured cells  

作　　者：Han Song Gang Zheng Xue-Feng Shen Zai-Hua Zhao Yang Liu Yang Liu Ying-Ying Liu Jun-Jun Kang Jing-Yuan Chen Wen-Jing Luo 

机构地区：[1]Department of Health Service,PLA General Hospital,Beijing 100853,China [2]Department of Occupational and Environmental Health and the Ministry-of-Education’s Key Laboratory of Hazard Assessment and Control in Special Operational Environment,School of Public Health,Fourth Military Medical University,Xi’an 710032,China [3]Department of Neurology,Xijing Hospital,Fourth Military Medical University,Xi’an 710032,China [4]Institute of Neurosciences,Fourth Military Medical University,Xi’an 710032,China

出　　处：《Biophysics Reports》2020年第5期211-221,共11页生物物理学报（英文版）

基　　金：the National Basic Research Program of China(973 Program)(2012CB525002);the National Natural Science Foundation of China(81230063 and 81172700);in part by the Program for Changjiang Scholars and Innovative Research Team in University(PCSIRT);NIH Grants ES-R0110563 and ES-R0107331.

摘　　要：Understanding the precise intracellular localization of lead(Pb)is a key in deciphering processes in Pbinduced toxicology.However,it is a great challenge to trace Pb in vitro,especially in cultured cells.We aimed to find an innovative and efficient approach to investigate distribution of Pb in cells and to validate it through determining the subcellular Pb content.We identified its ultra-structural distribution with autometallography under electron microscopy in a choroidal epithelial Z310 cell line.Electron microscopy in combination with energy-dispersive X-ray spectroscope(EDS)was employed to provide further evidence of Pb location.In addition,Pb content was determined in the cytosol,membrane/organelle,nucleus and cytoskeleton fractions with atomic absorption spectroscopy.Pb was found predominantly inside the nuclear membranes and some was distributed in the cytoplasm under lowconcentration exposure.Nuclear existence of Pb was verified by EDS under electron microscopy.Once standardized for protein content,Pb percentage in the nucleus fraction reached the highest level(76%).Our results indicate that Pb is accumulated mainly in the nucleus of choroid plexus.This method is sensitive and precise in providing optimal means to study the ultra-structural localization of Pb for in vitro models.In addition,it offers the possibility of localization of other metals in cultured cells.Some procedures may also be adopted to detect target proteins via immunoreactions.

关 键 词：LEAD Autometallography Subcellular fraction Energy-dispersive X-ray spectroscope Electron microscopy 

分 类 号：R99[医药卫生—毒理学]