Khan Niamat, Lenz Christof, Binder Lutz, Pantakani Dasaradha Venkata Krishna, Asif Abdul R
Institute for Clinical Chemistry/UMG-Laboratories, University Medical Center, Robert-Koch-Str. 40, 37075 Göttingen, Germany.
Department of Biotechnology & Genetic Engineering, Kohat University of Science and Technology, Kohat 26000, Khyber Pakhtunkhwa, Pakistan.
Int J Mol Sci. 2016 Apr 20;17(4):597. doi: 10.3390/ijms17040597.
Mycophenolic acid (MPA) is prescribed to maintain allografts in organ-transplanted patients. However, gastrointestinal (GI) complications, particularly diarrhea, are frequently observed as a side effect following MPA therapy. We recently reported that MPA altered the tight junction (TJ)-mediated barrier function in a Caco-2 cell monolayer model system. This study investigates whether MPA induces epigenetic changes which lead to GI complications, especially diarrhea.
We employed a Chromatin Immunoprecipitation-O-Proteomics (ChIP-O-Proteomics) approach to identify proteins associated with active (H3K4me3) as well as repressive (H3K27me3) chromatin histone modifications in MPA-treated cells, and further characterized the role of midkine, a H3K4me3-associated protein, in the context of epithelial monolayer permeability.
We identified a total of 333 and 306 proteins associated with active and repressive histone modification marks, respectively. Among them, 241 proteins were common both in active and repressive chromatin, 92 proteins were associated exclusively with the active histone modification mark, while 65 proteins remained specific to repressive chromatin. Our results show that 45 proteins which bind to the active and seven proteins which bind to the repressive chromatin region exhibited significantly altered abundance in MPA-treated cells as compared to DMSO control cells. A number of novel proteins whose function is not known in bowel barrier regulation were among the identified proteins, including midkine. Our functional integrity assays on the Caco-2 cell monolayer showed that the inhibition of midkine expression prior to MPA treatment could completely block the MPA-mediated increase in barrier permeability.
The ChIP-O-Proteomics approach delivered a number of novel proteins with potential implications in MPA toxicity. Consequently, it can be proposed that midkine inhibition could be a potent therapeutic approach to prevent the MPA-mediated increase in TJ permeability and leak flux diarrhea in organ transplant patients.
霉酚酸（MPA）被用于维持器官移植患者的同种异体移植物。然而，胃肠道（GI）并发症，尤其是腹泻，是MPA治疗后常见的副作用。我们最近报道，在Caco-2细胞单层模型系统中，MPA改变了紧密连接（TJ）介导的屏障功能。本研究调查MPA是否会诱导导致胃肠道并发症尤其是腹泻的表观遗传变化。
我们采用染色质免疫沉淀 - O蛋白质组学（ChIP - O - 蛋白质组学）方法来鉴定MPA处理细胞中与活性（H3K4me3）以及抑制性（H3K27me3）染色质组蛋白修饰相关的蛋白质，并进一步表征中期因子（一种与H3K4me3相关的蛋白质）在上皮单层通透性方面的作用。
我们分别鉴定出了333种和306种与活性和抑制性组蛋白修饰标记相关的蛋白质。其中，241种蛋白质在活性和抑制性染色质中都有，92种蛋白质仅与活性组蛋白修饰标记相关，而65种蛋白质仅存在于抑制性染色质中。我们的结果表明，与二甲基亚砜（DMSO）对照细胞相比，45种与活性染色质结合的蛋白质和7种与抑制性染色质区域结合的蛋白质在MPA处理的细胞中丰度显著改变。在鉴定出的蛋白质中包括一些在肠道屏障调节中功能未知的新蛋白质，其中有中期因子。我们对Caco - 2细胞单层进行的功能完整性分析表明，在MPA处理前抑制中期因子的表达可以完全阻断MPA介导的屏障通透性增加。
ChIP - O - 蛋白质组学方法鉴定出了许多对MPA毒性可能有影响的新蛋白质。因此，可以提出抑制中期因子可能是预防器官移植患者中MPA介导的TJ通透性增加和渗漏性腹泻的有效治疗方法。
