Foreign metallothionein-I expression by transient transfection in MT-I and MT-II null astrocytes confers increased protection against acute methylmercury cytotoxicity

Chang Ping Yao, Jeffrey W. Allen, Lysette A. Mutkus, Shao Bin Xu, Kim H. Tan, Michael Aschner

Research output: Contribution to journalArticlepeer-review

31 Scopus citations

Abstract

The mechanisms associated with metallothionein (MT) gene regulation are complex and poorly understood. Only a modest increase in brain MT expression levels is attained by exposure to metals, MT gene transfection, and MT gene knock-in techniques. Accordingly, in the present study, MT null astrocytes isolated from transgenic mice deficient in MT-I and MT-II genes were introduced as a zero background model of MT expression. MT protein levels were determined by western blot analysis. MT proteins in MT-I and MT-II null astrocytes were undetectable. Transient MT-I gene transfection increased the levels of foreign MT expression in MT-I and MT-II null astrocytes by 2.3-fold above basal levels in wild-type astrocytes. Intracellular Na251CrO4 efflux and D-[2,3-3H]aspartate uptake were studied as indices of acute methylmercury (MeHg) (5 μM) cytotoxicity. In MT-I and MT-II knockout astrocytes MeHg led to significant (p < 0.01) increase in Na251CrO4 efflux and a significant (p < 0.05) decrease in the initial rate (1 min) of D-[2,3-3H]aspartate uptake compared to MT-I and MT-II knockout controls. Transfection of the MT-I gene in MT-I and MT-II null mice significantly (p < 0.01) decreased the effect of MeHg on Na251CrO4 efflux in MT null, as well as wild-type astrocytes. MT-I gene transfection in MT-I and MT-II null astrocytes reversed the inhibitory effect of MeHg on D-[2,3-3H]aspartate uptake, such that initial rates of uptake in MT-I transfected cells in the presence and absence of MeHg (5 μM) were indistinguishable. These results demonstrate that: (1) astrocytes lacking MTs are more sensitive to MeHg than those with basal MT protein levels, (2) the MT-I gene can be overexpressed in MT-I and MT-II null astrocytes by transient MT-I gene transfection, and (3) that foreign MT expression endows astrocytes with increased resistance to MeHg. (C) 2000 Elsevier Science B.V.

Original languageEnglish (US)
Pages (from-to)32-38
Number of pages7
JournalBrain Research
Volume855
Issue number1
DOIs
StatePublished - Feb 7 2000
Externally publishedYes

Keywords

  • Knock-in
  • Knockout
  • MT-I and MT-II null astrocytes
  • Metallothionein
  • Methylmercury
  • Transgenic mouse

ASJC Scopus subject areas

  • Neuroscience(all)
  • Molecular Biology
  • Clinical Neurology
  • Developmental Biology

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