Schneider André, Zhang YaHua, Zhang Mingzhi, Lu Wendell J, Rao Reena, Fan Xuefeng, Redha Reyadh, Davis Linda, Breyer Richard M, Harris Raymond, Guan YouFei, Breyer Matthew D
Division of Nephrology, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee 37232, USA.
Kidney Int. 2004 Apr;65(4):1205-13. doi: 10.1111/j.1523-1755.2004.00493.x.
Prostaglandin E2 (PGE2) plays an important role in many physiologic and pathophysiologic processes in the kidney. Multiple enzymes are involved in PGE2 biosynthesis, including phospholipases, cyclooxygenases (COX), and the PGE2 synthases (PGES). The present studies were aimed at determining the intrarenal localization of mPGES-1 and whether it is coexpressed with COX-1 or COX-2.
Rabbit mPGES-1 and COX-1 cDNAs were cloned using reverse transcription-polymerase chain reaction (RT-PCR) and screening a cDNA library. RNase protection assay and immunoblotting were used to examine mPGES-1 expression levels. In situ hybridization and immunostaining were used to determine the intrarenal localization of mPGES-1 and cyclooxygenases.
Rabbit mPGES-1 shares high sequence similarity to the human homolog. Nuclease protection studies showed that the kidney expresses among the highest level of mPGES-1 of any rabbit tissue. In situ hybridization showed COX-1 and mPGES-1 mRNA was highly expressed in renal medullary collecting ducts (MCD), and to a lesser extent in cortical collecting ducts (CCD). Fainter mPGES-1 expression was also observed in macula densa (MD) and medullary interstitial cells (RMICs), where COX-2 is highly expressed. Double-labeling studies (immunostaining plus in situ hybridization) and immunohistochemistry of mouse tissues confirmed that mPGES-1 predominantly colocalizes with COX-1 in distal convoluted tubule (DCT), CCD, and MCD, and is coexpressed with COX-2 at lower levels in MD and RMICs.
Together, these studies suggest mPGES-1 colocalizes with both COX-1 and COX-2 to mediate the biosynthesis of PGE2 in the kidney.
前列腺素E2（PGE2）在肾脏的许多生理和病理生理过程中发挥重要作用。多种酶参与PGE2的生物合成，包括磷脂酶、环氧化酶（COX）和PGE2合酶（PGES）。本研究旨在确定微粒体前列腺素E合酶-1（mPGES-1）在肾脏内的定位以及它是否与COX-1或COX-2共表达。
使用逆转录聚合酶链反应（RT-PCR）并筛选cDNA文库克隆兔mPGES-1和COX-1 cDNA。采用核糖核酸酶保护试验和免疫印迹法检测mPGES-1表达水平。运用原位杂交和免疫染色法确定mPGES-1和环氧化酶在肾脏内的定位。
兔mPGES-1与人同源物具有高度的序列相似性。核酸酶保护研究表明，肾脏中mPGES-1的表达水平在兔的任何组织中都是最高的。原位杂交显示，COX-1和mPGES-1 mRNA在肾髓质集合管（MCD）中高度表达，在皮质集合管（CCD）中表达程度较低。在致密斑（MD）和肾髓质间质细胞（RMICs，COX-2高度表达的部位）中也观察到较弱的mPGES-1表达。对小鼠组织进行的双标记研究（免疫染色加原位杂交）和免疫组织化学证实，mPGES-1主要与COX-1在远曲小管（DCT）、CCD和MCD中共定位，并在MD和RMICs中与COX-2以较低水平共表达。
总之，这些研究表明mPGES-1与COX-1和COX-2共定位，以介导肾脏中PGE2的生物合成。
