Hart B A, Gong Q, Eneman J D
Department of Biochemistry, University of Vermont College of Medicine, Burlington 05405, USA.
Toxicology. 1996 Sep 2;112(3):205-18. doi: 10.1016/0300-483x(96)03397-5.
This study examined the expression of metallothionein (MT) isoforms in the lungs of Lewis rats exposed to Cadmium (Cd) aerosols. With the use of isoform-specific oligonucleotide probes and Northern hybridization analysis, we demonstrated that a dramatic, rapid, and coordinate increase occurred in pulmonary MT-1 mRNA and MT-2 mRNA following Cd inhalation exposure. MT mRNAs levels reached a maximum at 2 h post-exposure and remained above control levels at 96 h after exposure. A considerable lag between the time of maximal elevations in MT mRNAs and MT protein accumulation was observed and suggested that regulatory mechanisms in addition to transcriptional control could be involved. MT expression (protein and mRNA) and Cd lung burden were directly related to aerosol Cd concentration. In situ hybridization and immunohistochemistry studies showed good correlation between the localization of MT protein and MT mRNAs. However, staining for MT protein and MT mRNA was not uniformly distributed in the lung. MT was particularly prominent within the alveolar compartment. Even within this area, however, heterogeneity of MT expression was evident. Experiments were subsequently conducted to determine whether prior exposure to Cd modulates the transcriptional activity of MT genes such that there is a greater elevation in gene expression upon reexposure to Cd. Surprisingly, animals pretreated with Cd exhibited a smaller incremental increase in MT mRNA levels in response to subsequent Cd exposure than controls with no prior treatment. Moreover, MT mRNA levels were elevated to a similar extent regardless of whether animals were exposed to Cd aerosols for 1, 2, or 3 weeks (3 h/day; 5 days/week). MT protein and lung Cd burden, on the other hand, exhibited an increasing linear trend as a function of exposure number. In summary, this study has demonstrated that: (1) the lung responds to Cd inhalation exposure by increasing MT mRNA and MT protein levels; (2) MT expression is prominent within alveolar cells but not all cells are MT positive; and (3) Cd-pretreatment does not increase the transcriptional potential of MT genes when the animal is subsequently reexposed to Cd.
本研究检测了暴露于镉（Cd）气溶胶的Lewis大鼠肺中金属硫蛋白（MT）亚型的表达。通过使用亚型特异性寡核苷酸探针和Northern杂交分析，我们证明吸入Cd暴露后，肺中MT-1 mRNA和MT-2 mRNA出现了显著、快速且协同的增加。MT mRNA水平在暴露后2小时达到峰值，并在暴露后96小时仍高于对照水平。观察到MT mRNA最大升高时间与MT蛋白积累之间存在相当长的滞后，这表明除转录控制外，可能还涉及其他调节机制。MT表达（蛋白和mRNA）与肺中Cd负荷直接相关。原位杂交和免疫组织化学研究表明MT蛋白和MT mRNA的定位之间具有良好的相关性。然而，MT蛋白和MT mRNA的染色在肺中并非均匀分布。MT在肺泡隔内特别突出。然而，即使在这个区域内，MT表达的异质性也很明显。随后进行了实验，以确定预先暴露于Cd是否会调节MT基因的转录活性，从而使再次暴露于Cd时基因表达有更大的升高。令人惊讶的是，与未预先处理的对照相比，预先用Cd处理的动物在随后暴露于Cd时，MT mRNA水平的增量增加较小。此外，无论动物暴露于Cd气溶胶1、2或3周（每天3小时；每周5天），MT mRNA水平升高的程度相似。另一方面，MT蛋白和肺Cd负荷随暴露次数呈线性增加趋势。总之，本研究表明：（1）肺通过增加MT mRNA和MT蛋白水平对吸入Cd暴露作出反应；（2）MT表达在肺泡细胞内突出，但并非所有细胞都是MT阳性；（3）当动物随后再次暴露于Cd时，Cd预处理不会增加MT基因的转录潜能。
