Adenosine A_2A Receptor Is An Important Target For Regulating Renal Interstitial Fibrosis In Obstructive Nephropathy

BackgroundRegardless of the etiology, almost all forms of end stage renal disease share thecommon pathological feature of progressive renal interstitial fibrosis （RIF） and tubularatrophy. RIF is commonly triggered by inflammatory processes and succedentepithelial-mesenchymal transition （EMT）. An early initiated anti-inflammatory strategy istherefore of the importance to prevent the progression of RIF, however, no therapeuticapproach is currently available to achieve this goal. Thus, exploring new therapeutic targetis in urgent need.Renal inflammation after sustained injuries, e.g. IgA nephropathy and lupus nephritis,serves as a primer that sets up the fibrogenic stage and triggers tissue fibrogenesis. In thispathological progress inflammatory cells, such as macrophage and lymphocyte, play crucialroles. RIF is characterized by the myofibroblast activation and the accumulation of matrixproteins including collagen types I （Col I） and type III （Col III）. While produced via theEMT event in the pathologic progress of RIF myofibroblast with identified expression ofα-SMA is the major source of the increased production of matrix protein in RIF. A unilateralureteral obstruction （UUO） model has been refined to elucidate the pathogenesis andmechanisms responsible for RIF. It has been shown that the infiltration of macrophages andT cells and lymphocyte dysfunction are two major mechanisms contributing toUUO-induced RIF model. In this model, at cellular level, tubular dilatation leads tubularepithelia lose their epithelial characteristics and acquire mesenchymal traits such as α-SMAexpression and actin reorganization. Therefore, tubular epithelial cells become the majorsource of renal myofibroblast during EMT process. At molecular level, TGF-β1plays a keyrole in EMT via activation of its downstream Rho/ROCK signaling pathway. Recently, adenosine A_2A receptor (A_2AR) emerges as a novel inflammation regulatoraffecting on inflammation process and tissue repair. Pharmacology studies showed thatA_2AR agonist, CGS21680and ATL193, can effectively suppress inflammation. Activationof A_2AR leads to attenuation of glomerulonephritis and renal injury. Further, recent studyidentified that A_2AR activation inhibits Rho/ROCK1in hepatic stellate cells. All abovestrongly suggest that A_2AR manipulation plays an important regulatory role oninflammation on EMT event. Therefore, we hypothesize that activation of A_2AR maysuppress cellular infiltration, EMT event and profibrogenic factors, thereby to preventconsequent pathology of RIF. Conversely, inactivation of A_2AR may lead exacerbation ofRIF.Using experimental UUO-induced RIF mouse model, we evaluated the modulatoryeffect of A_2AR-based manipulation on several aspects of RIF progression, includinginterstitial lymphocyte infiltration, cellular biomarkers of EMT, expression of profibrogenicfactor TGF-β1and its downstream Rho/ROCK1pathway, as well as the consequentextracellular matrix accumulation.MethodsTo test this hypothesis we applied unilateral ureteral obstruction （UUO） model of RIFon A_2AR knockout mice and their littermates, and combined the intervention of selectiveA_2AR agonist, CGS21680. At the day3,7and14post-RIF model establishment, weevaluated the effects of A_2AR manipulation on molecular pathological progress of RIF,including imunohistochemistry of collagen types I, III, α-SMA and CD4+T lymphocyte,CD4+CD25+FoxP3+regulatory T cells （Treg）, Western blot of E-cadherin and α-smoothmuscle actin （α-SMA）, and quantitative PCR for ROCK1and TGF-β1mRNAmeasurement.ResultsOur data showed that activation of A_2AR significantly suppressed the deposition ofcollagen types I, III, the infiltration of CD4+T lymphocytes as well as the expression ofTGF-β1and ROCK1that inhibited and postponed the process of EMT. Conversely, geneticinactivation of A_2AR exacerbated aforementioned pathophysiological processes of RIF. ConclusionTogether, activation of A2AR effectively alleviated EMT and RIF in mice, suggestingA2AR as a potential therapeutic target for the treatment of RIF.