Blommaert F A, Michael C, van Dijk-Knijnenburg H C, Schornagel J H, den Engelse L, Fichtinger-Schepman A M
Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
Cancer Chemother Pharmacol. 1996;38(3):273-80. doi: 10.1007/s002800050482.
The formation and persistence of platinum-DNA adducts were studied with immuno(cyto)chemical methods in male and female Sprague-Dawley rats treated with a single i.p. dose of carboplatin. Linear dose-effect curves were observed for kidney and liver with an immunocytochemical assay using NKI-A59 antiserum that recognizes intrastrand cross-links. With this method, no staining of the nuclei due to platinum-DNA damage could be observed in the spleen, testis, uterus, or ovary after administration of up to 80 mg/kg carboplatin. A homogeneous staining of the nuclei in the liver was observed. The nuclear staining in the kidney was somewhat more intense but less homogeneous, with small groups of intensely stained nuclei occasionally being seen in the outer cortex. An approximately 15 to 20-times lower dose of cisplatin than of carboplatin was needed to reach equal staining levels in the liver and kidney. Plateau staining levels in both tissues were reached at between approximately 8 and 48 h after administration of the carboplatin. This was followed by a significant reduction in the kidney samples, whereas the staining levels in the liver section seemed to be more persistent. No major difference was observed between male and female rats in the formation and removal of DNA damage in these tissues. The levels of the various DNA adducts were measured with a competitive ELISA in liver, kidney, spleen, testis, and combined ovary/uterus samples collected at 8 and 48 h after carboplatin administration. At both 8 and 48 h, the highest platination levels were observed in the kidney, followed--in decreasing order--by the liver, combined uterus and ovary samples, spleen, and testis. At 8 h after administration of carboplatin, the relative occurrence of the bifunctional adducts Pt-GG (34%), Pt-AG (27%), and G-Pt-G (32%), was similar in all tissues. The same held for the monoadducts that amounted to about 7% of the total DNA platination. These data indicate that in the first few hours after carboplatin treatment, no preference for the formation of Pt-GG adducts was observed, which confirms our earlier observations obtained with cultured cells. When the total DNA-platination levels (calculated from the sum of the adducts) seen at 8 and 48 h after treatment were compared, a substantial decrease in DNA platination was observed in the kidney (37%), liver (30%) and ovary/uterus (39%), whereas the repair levels in the testis (9%) and, probably, the spleen (18%) were substantially lower. In all tissues studied, only the relative occurrence of the Pt-GG adducts increased between 8 and 48 h, and as a result, at 48 h, after carboplatin administration the Pt-GG adduct was the major adduct persisting in the DNA samples.
采用免疫（细胞）化学方法，研究了经腹腔注射单次剂量卡铂处理的雄性和雌性Sprague-Dawley大鼠体内铂-DNA加合物的形成和持久性。使用识别链内交联的NKI-A59抗血清进行免疫细胞化学分析，观察到肾脏和肝脏呈线性剂量效应曲线。采用该方法，给予高达80mg/kg卡铂后，在脾脏、睾丸、子宫或卵巢中未观察到因铂-DNA损伤导致的细胞核染色。观察到肝脏细胞核呈均匀染色。肾脏中的细胞核染色稍强但不均匀，在外皮质偶尔可见小群强染色的细胞核。在肝脏和肾脏中达到同等染色水平所需的顺铂剂量比卡铂低约15至20倍。给予卡铂后约8至48小时，两个组织均达到平台期染色水平。随后肾脏样本中的染色水平显著降低，而肝脏切片中的染色水平似乎更持久。在这些组织中，雄性和雌性大鼠在DNA损伤的形成和清除方面未观察到重大差异。在给予卡铂后8小时和48小时收集的肝脏、肾脏、脾脏、睾丸以及卵巢/子宫联合样本中，采用竞争性ELISA法测量了各种DNA加合物的水平。在8小时和48小时时，肾脏中的铂化水平最高，其次（按降序排列）为肝脏、子宫和卵巢联合样本、脾脏和睾丸。给予卡铂后8小时，所有组织中双功能加合物Pt-GG（34%）、Pt-AG（27%）和G-Pt-G（32%）的相对发生率相似。单加合物的情况也是如此，其占总DNA铂化的约7%。这些数据表明，在卡铂治疗后的最初几个小时内，未观察到对Pt-GG加合物形成的偏好，这证实了我们早期在培养细胞中获得的观察结果。比较治疗后8小时和48小时观察到的总DNA铂化水平（根据加合物总和计算），发现肾脏（37%）、肝脏（30%）和卵巢/子宫（39%）中的DNA铂化显著降低，而睾丸（9%）以及可能的脾脏（18%）中的修复水平则低得多。在所有研究的组织中，仅Pt-GG加合物的相对发生率在8小时至48小时之间增加，因此，给予卡铂后48小时，Pt-GG加合物是DNA样本中持续存在的主要加合物。
