Gangwar Ruchika, Meena Avtar S, Shukla Pradeep K, Nagaraja Archana S, Dorniak Piotr L, Pallikuth Sandeep, Waters Christopher M, Sood Anil, Rao RadhaKrishna
Department of Physiology, University of Tennessee Health Science Center, Memphis, TN, U.S.A.
Department of Gynecologic Oncology and Center for RNA Interference and Non-Coding RNA, M.D. Anderson Cancer Center, Houston, TX, U.S.A.
Biochem J. 2017 Feb 20;474(5):731-749. doi: 10.1042/BCJ20160679.
The role of reactive oxygen species (ROS) in osmotic stress, dextran sulfate sodium (DSS) and cyclic stretch-induced tight junction (TJ) disruption was investigated in Caco-2 cell monolayers and restraint stress-induced barrier dysfunction in mouse colon Live cell imaging showed that osmotic stress, cyclic stretch and DSS triggered rapid production of ROS in Caco-2 cell monolayers, which was blocked by depletion of intracellular Ca by 1,2-bis-(-aminophenoxy)ethane-,,','-tetraacetic acid. Knockdown of Ca1.3 or TRPV6 channels blocked osmotic stress and DSS-induced ROS production and attenuated TJ disruption and barrier dysfunction. -Acetyl l-cysteine (NAC) and l--Nitroarginine methyl ester (l-NAME) blocked stress-induced TJ disruption and barrier dysfunction. NAC and l-NAME also blocked stress-induced activation of c-Jun -terminal kinase (JNK) and c-Src. ROS was colocalized with the mitochondrial marker in stressed cells. Cyclosporin A blocked osmotic stress and DSS-induced ROS production, barrier dysfunction, TJ disruption and JNK activation. Mitochondria-targeted Mito-TEMPO blocked osmotic stress and DSS-induced barrier dysfunction and TJ disruption. Chronic restraint stress in mice resulted in the elevation of intracellular Ca, activation of JNK and c-Src, and disruption of TJ in the colonic epithelium. Furthermore, corticosterone administration induced JNK and c-Src activation, TJ disruption and protein thiol oxidation in colonic mucosa. The present study demonstrates that oxidative stress is a common signal in the mechanism of TJ disruption in the intestinal epithelium by different types of cellular stress and bio behavioral stress .
在Caco-2细胞单层中研究了活性氧（ROS）在渗透应激、硫酸葡聚糖钠（DSS）和周期性拉伸诱导的紧密连接（TJ）破坏中的作用，以及在小鼠结肠中束缚应激诱导的屏障功能障碍。活细胞成像显示，渗透应激、周期性拉伸和DSS在Caco-2细胞单层中触发了ROS的快速产生，这被1,2-双（-氨基苯氧基）乙烷-N,N,N',N'-四乙酸耗尽细胞内钙所阻断。敲低Ca1.3或TRPV6通道可阻断渗透应激和DSS诱导的ROS产生，并减轻TJ破坏和屏障功能障碍。N-乙酰半胱氨酸（NAC）和L-硝基精氨酸甲酯（L-NAME）可阻断应激诱导的TJ破坏和屏障功能障碍。NAC和L-NAME还可阻断应激诱导的c-Jun氨基末端激酶（JNK）和c-Src的激活。ROS与应激细胞中的线粒体标志物共定位。环孢素A可阻断渗透应激和DSS诱导的ROS产生、屏障功能障碍、TJ破坏和JNK激活。靶向线粒体的Mito-TEMPO可阻断渗透应激和DSS诱导的屏障功能障碍和TJ破坏。小鼠的慢性束缚应激导致结肠上皮细胞内钙升高、JNK和c-Src激活以及TJ破坏。此外，给予皮质酮可诱导结肠黏膜中JNK和c-Src激活、TJ破坏和蛋白质硫醇氧化。本研究表明，氧化应激是不同类型的细胞应激和生物行为应激导致肠上皮TJ破坏机制中的共同信号。
