Cometti Barbara, Dubey Raghvendra K, Imthurn Bruno, Jackson Edwin K, Rosselli Marinella
Departments of Medicine and Pharmacology, University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania 15261, USA.
Biol Reprod. 2003 Sep;69(3):868-75. doi: 10.1095/biolreprod.103.017962. Epub 2003 Apr 30.
The extracellular cAMP-adenosine pathway refers to the local production of adenosine mediated by cAMP egress into the extracellular space, conversion of cAMP to AMP by ectophosphodiesterase (PDE), and the metabolism of AMP to adenosine by ecto-5'-nucleotidase. The goal of this study was to assess whether the cAMP-adenosine pathway is expressed in oviduct cells. Studies were conducted in cultured bovine oviduct cells (mixed cultures of fibroblasts and epithelial cells, 1:1 ratio). Confluent monolayers of oviduct cells were exposed to cAMP (0.01-100 micromol/L) in the presence and absence of 3-isobutyl-1-methylxanthine (IBMX, 1 mmol/L, an inhibitor of both extracellular and intracellular PDE activity), 1,3-dipropyl-8-p-sulfophenylxanthine (DPSPX, 100 micromol/L, a xanthine that can inhibit extracellular or ecto-PDE activity at high concentrations), or alpha,beta-methylene-adenosine-5'-diphosphate (AMPCP, 100 micromol/L, an ecto-5'-nucleotidase inhibitor) for 0-60 min. The medium was then sampled and assayed for AMP, adenosine, and inosine. Addition of exogenous cAMP to oviduct cells increased extracellular levels of AMP, adenosine, and inosine in a concentration- and time-dependent manner. This effect was attenuated by blockade of total (extracellular and intracellular) PDE activity (IBMX), ecto-PDE activity (DPSPX), or ecto-5'-nucleotidase (AMPCP). The functional relevance of the cAMP-adenosine pathway is supported by the findings that treatment with adenylyl cyclase stimulants (forskolin plus isoproterenol) resulted in the egress of cAMP (97% extracellular) into the extracellular space and its conversion into adenosine. The extracellular cAMP-adenosine pathway exists in oviduct cells and may play an important role in regulating the biology and physiology of the oviduct. This pathway also may play a critical role in regulating sperm function, fertilization, and early embryo development.
细胞外cAMP - 腺苷途径是指由cAMP释放到细胞外空间介导的腺苷局部产生过程，即胞外磷酸二酯酶（PDE）将cAMP转化为AMP，以及胞外5'-核苷酸酶将AMP代谢为腺苷。本研究的目的是评估cAMP - 腺苷途径是否在输卵管细胞中表达。研究在培养的牛输卵管细胞（成纤维细胞和上皮细胞的混合培养物，比例为1:1）中进行。将汇合的输卵管细胞单层在存在和不存在3 - 异丁基 - 1 - 甲基黄嘌呤（IBMX，1 mmol/L，一种细胞外和细胞内PDE活性抑制剂）、1,3 - 二丙基 - 8 - 对 - 磺基苯基黄嘌呤（DPSPX，100 μmol/L，一种在高浓度下可抑制细胞外或胞外PDE活性的黄嘌呤）或α,β - 亚甲基 - 腺苷 - 5'-二磷酸（AMPCP，100 μmol/L，一种胞外5'-核苷酸酶抑制剂）的情况下，暴露于cAMP（0.01 - 100 μmol/L）中0 - 60分钟。然后对培养基进行采样，并测定其中的AMP、腺苷和肌苷。向输卵管细胞中添加外源性cAMP会以浓度和时间依赖性方式增加细胞外AMP、腺苷和肌苷的水平。这种效应会因阻断总（细胞外和细胞内）PDE活性（IBMX）、胞外PDE活性（DPSPX）或胞外5'-核苷酸酶（AMPCP）而减弱。腺苷酸环化酶刺激剂（福斯高林加异丙肾上腺素）处理导致cAMP（97%位于细胞外）释放到细胞外空间并转化为腺苷，这一发现支持了cAMP - 腺苷途径的功能相关性。细胞外cAMP - 腺苷途径存在于输卵管细胞中，可能在调节输卵管的生物学和生理学方面发挥重要作用。该途径在调节精子功能、受精和早期胚胎发育方面也可能起关键作用。
