Abstract
Astrocytes provide a vital protective function in the brain. These cells are also vulnerable to oxidative stress, thus their loss of function could contribute to neurodegeneration. The goal of this study is to develop a cell culture model to study oxidative stress in astrocytes. Enriched astrocytic cultures were generated from neonatal mice. tertiary-butyl hydroperoxide (t- bOOH) was used as an exogenous peroxide and lactate dehydrogenase (LDH) release as a measure of loss of viability. Exposure to t-bOOH resulted in a linear increase in astrocytic death reaching 91.2% after 4 h exposure. That cell death was due to oxidative injury, was shown by the ability of the antioxidant N,N'-diphenyl-1,4-phenylenediamine (DPPD) to protect the t-bOOH treated cells. The involvement of iron in cell toxicity was demonstrated by the ability of the iron specific chelator desferal (DF) to completely prevent t-bOOH induced LDH release. Cells treated with a lipid soluble iron compound 3,5,5-trimethyl (hexanoyl) ferrocene (TMH-Ferrocene), were more vulnerable to t-bOOH whereas neither ferrous ammonium sulfate (FAS) nor ferric ammonium citrate (FAC) had an effect. The increased sensitivity of the cells exposed to TMHF was reversible with the iron chelator desferal. Addition of recombinant human heavy chain ferritin or human apo-transferrin (Tf) did not alter LDH release. Electron microscopic analysis indicated astrocytes exposed to t-bOOH exhibited mitochondrial swelling prior to cell death (lactate dehydrogenase release). Additional increases in mitochondrial swelling were seen when the astrocytes were exposed to the lipophilic iron compound TMH- ferrocene and t-bOOH. These studies show that astrocytes are exquisitely sensitive to oxidative stress and that their vulnerability is related to and enhanced by iron. Decreased mitochondrial function in response to oxidative stress may precede cell death.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 125-132 |
| Number of pages | 8 |
| Journal | Brain research |
| Volume | 788 |
| Issue number | 1-2 |
| DOIs | |
| State | Published - Mar 30 1998 |
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All Science Journal Classification (ASJC) codes
- Neuroscience(all)
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An in vitro model for analysis of oxidative death in primary mouse astrocytes. / Robb, S. J.; Connor, James.
In: Brain research, Vol. 788, No. 1-2, 30.03.1998, p. 125-132.Research output: Contribution to journal › Article
TY - JOUR
T1 - An in vitro model for analysis of oxidative death in primary mouse astrocytes
AU - Robb, S. J.
AU - Connor, James
PY - 1998/3/30
Y1 - 1998/3/30
N2 - Astrocytes provide a vital protective function in the brain. These cells are also vulnerable to oxidative stress, thus their loss of function could contribute to neurodegeneration. The goal of this study is to develop a cell culture model to study oxidative stress in astrocytes. Enriched astrocytic cultures were generated from neonatal mice. tertiary-butyl hydroperoxide (t- bOOH) was used as an exogenous peroxide and lactate dehydrogenase (LDH) release as a measure of loss of viability. Exposure to t-bOOH resulted in a linear increase in astrocytic death reaching 91.2% after 4 h exposure. That cell death was due to oxidative injury, was shown by the ability of the antioxidant N,N'-diphenyl-1,4-phenylenediamine (DPPD) to protect the t-bOOH treated cells. The involvement of iron in cell toxicity was demonstrated by the ability of the iron specific chelator desferal (DF) to completely prevent t-bOOH induced LDH release. Cells treated with a lipid soluble iron compound 3,5,5-trimethyl (hexanoyl) ferrocene (TMH-Ferrocene), were more vulnerable to t-bOOH whereas neither ferrous ammonium sulfate (FAS) nor ferric ammonium citrate (FAC) had an effect. The increased sensitivity of the cells exposed to TMHF was reversible with the iron chelator desferal. Addition of recombinant human heavy chain ferritin or human apo-transferrin (Tf) did not alter LDH release. Electron microscopic analysis indicated astrocytes exposed to t-bOOH exhibited mitochondrial swelling prior to cell death (lactate dehydrogenase release). Additional increases in mitochondrial swelling were seen when the astrocytes were exposed to the lipophilic iron compound TMH- ferrocene and t-bOOH. These studies show that astrocytes are exquisitely sensitive to oxidative stress and that their vulnerability is related to and enhanced by iron. Decreased mitochondrial function in response to oxidative stress may precede cell death.
AB - Astrocytes provide a vital protective function in the brain. These cells are also vulnerable to oxidative stress, thus their loss of function could contribute to neurodegeneration. The goal of this study is to develop a cell culture model to study oxidative stress in astrocytes. Enriched astrocytic cultures were generated from neonatal mice. tertiary-butyl hydroperoxide (t- bOOH) was used as an exogenous peroxide and lactate dehydrogenase (LDH) release as a measure of loss of viability. Exposure to t-bOOH resulted in a linear increase in astrocytic death reaching 91.2% after 4 h exposure. That cell death was due to oxidative injury, was shown by the ability of the antioxidant N,N'-diphenyl-1,4-phenylenediamine (DPPD) to protect the t-bOOH treated cells. The involvement of iron in cell toxicity was demonstrated by the ability of the iron specific chelator desferal (DF) to completely prevent t-bOOH induced LDH release. Cells treated with a lipid soluble iron compound 3,5,5-trimethyl (hexanoyl) ferrocene (TMH-Ferrocene), were more vulnerable to t-bOOH whereas neither ferrous ammonium sulfate (FAS) nor ferric ammonium citrate (FAC) had an effect. The increased sensitivity of the cells exposed to TMHF was reversible with the iron chelator desferal. Addition of recombinant human heavy chain ferritin or human apo-transferrin (Tf) did not alter LDH release. Electron microscopic analysis indicated astrocytes exposed to t-bOOH exhibited mitochondrial swelling prior to cell death (lactate dehydrogenase release). Additional increases in mitochondrial swelling were seen when the astrocytes were exposed to the lipophilic iron compound TMH- ferrocene and t-bOOH. These studies show that astrocytes are exquisitely sensitive to oxidative stress and that their vulnerability is related to and enhanced by iron. Decreased mitochondrial function in response to oxidative stress may precede cell death.
UR - http://www.scopus.com/inward/record.url?scp=0032579829&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0032579829&partnerID=8YFLogxK
U2 - 10.1016/S0006-8993(97)01543-6
DO - 10.1016/S0006-8993(97)01543-6
M3 - Article
C2 - 9554979
AN - SCOPUS:0032579829
VL - 788
SP - 125
EP - 132
JO - Brain Research
JF - Brain Research
SN - 0006-8993
IS - 1-2
ER -