The enzymatic activity of human aldehyde dehydrogenases 1A2 and 2 (ALDH1A2 and ALDH2) is detected by Aldefluor, inhibited by diethylaminobenzaldehyde and has significant effects on cell proliferation and drug resistance.
The enzymatic activity of human aldehyde dehydrogenases 1A2 and 2 (ALDH1A2 and ALDH2) is detected by Aldefluor, inhibited by diethylaminobenzaldehyde and has significant effects on cell proliferation and drug resistance.
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DOI：
10.1016/j.cbi.2011.10.007
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发表时间：
2012-01-05
影响因子：
5.1
通讯作者：
Chang, Lung-Ji
中科院分区：
文献类型：
--
作者：
Moreb, Jan S.;Ucar, Deniz;Han, Shuhong;Amory, John K.;Goldstein, Alex S.;Ostmark, Blanca;Chang, Lung-Ji
自从基于Aldefluor流式细胞术的测定方法出现以来，人们对使用醛脱氢酶（ALDH）活性作为干细胞的一种标志物产生了新的兴趣。二乙基氨基苯甲醛（DEAB）被认为是ALDH1A1异构体的特异性抑制剂。在本研究中，我们探讨了人ALDH同工酶ALDH1A2和ALDH2对耐药和增殖的影响，以及DEAB作为抑制剂的特异性。我们还使用TaqMan低密度阵列（TLDA）筛选了19种ALDH同工酶在K562细胞中的表达。我们使用含有ALDH2或ALDH1A2全cDNA长度的慢病毒载体在K562白血病和H1299肺癌细胞系中过表达这些酶。通过活性测定、Western blot、RT-PCR和Aldefluor法检测成功表达。携带ALDH1A2或ALDH2的两种细胞系均表现出更高的细胞增殖率和克隆效率，并且对4-氢过氧环磷酰胺和阿霉素的耐药性增强。为了研究已知的ALDH活性抑制剂DEAB和diulfiram的特异性，我们用每种抑制剂孵育每个细胞系，并测量剩余的ALDH酶活性。两种抑制剂均使两种同工酶的ALDH活性降低65-90%。此外，我们的TLDA结果显示ALDH1， ALDH7， ALDH3和ALDH8在K562细胞中表达。我们得出结论，DEAB不是ALDH1A1的特异性抑制剂，Aldefluor试验对ALDH1A1的活性也不是特异性的。此外，其他ALDH同工酶似乎在各种恶性细胞的生物学和耐药过程中发挥重要作用。
There has been a new interest in using aldehyde dehydrogenase (ALDH) activity as one marker for stem cells since the Aldefluor flow cytometry-based assay has become available. Diethylaminobenzaldehyde (DEAB), used in the Aldeflour assay, has been considered a specific inhibitor for ALDH1A1 isoform. In this study, we explore the effects of human ALDH isoenzymes, ALDH1A2 and ALDH2, on drug resistance and proliferation, and the specificity of DEAB as an inhibitor. We also screened for the expression of 19 ALDH isoenzymes in K562 cells using TaqMan Low Density Array (TLDA). We used lentiviral vectors containing the full cDNA length of either ALDH2 or ALDH1A2 to over express the enzymes in K562 leukemia and H1299 lung cancer cell lines. Successful expression was measured by activity assay, Western blot, RT-PCR, and Aldefluor assay. Both cell lines, with either ALDH1A2 or ALDH2, exhibited higher cell proliferation rates, higher clonal efficiency, and increased drug resistance to 4-hydroperoxycyclophosphamide and doxorubicin. In order to study the specificity of known ALDH activity inhibitors, DEAB and disulfiram, we incubated each cell line with either inhibitor and measured the remaining ALDH enzymatic activity. Both inhibitors reduced ALDH activity of both isoenzymes by 65–90%. Furthermore, our TLDA results revealed that ALDH1, ALDH7, ALDH3 and ALDH8 are expressed in K562 cells. We conclude that DEAB is not a specific inhibitor for ALDH1A1 and that Aldefluor assay is not specific for ALDH1A1 activity. In addition, other ALDH isoenzymes seem to play a major role in the biology and drug resistance of various malignant cells.
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影响因子：
3
作者：
Moreb, Jan S.;Muhoczy, Dagmara;Zucali, James R.
通讯作者：
Zucali, James R.
影响因子：
2.6
作者：
Black WJ;Stagos D;Marchitti SA;Nebert DW;Tipton KF;Bairoch A;Vasiliou V
通讯作者：
Vasiliou V
影响因子：
20.3
作者：
JONES, RJ;BARBER, JP;HILTON, J
通讯作者：
HILTON, J
影响因子：
3
作者：
Barth, Kelly S.;Malcolm, Robert J.
通讯作者：
Malcolm, Robert J.
影响因子：
2.6
作者：
Niederreither, K;McCaffery, P;Dolle, P
通讯作者：
Dolle, P