Kim Seo Rin, Eirin Alfonso, Zhang Xin, Lerman Amir, Lerman Lilach O
Division of Nephrology and Hypertension, Mayo Clinic, Rochester, USA.
Cardiovascular Diseases, Mayo Clinic, Rochester, USA.
Cell Physiol Biochem. 2019;52(3):617-632. doi: 10.33594/000000044.
BACKGROUND/AIMS: Atherosclerotic renal artery stenosis (ARAS) may cause kidney injury and mitochondrial dysfunction, which is linked to cellular senescence. Elamipretide, a mitochondria-targeted peptide, improves renal function in ARAS, but whether it alleviates senescence is unknown. We hypothesized that elamipretide would reduce senescence stenotic kidney (STK) in ARAS.
Domestic pigs were randomized to control and unilateral ARAS untreated or treated with subcutaneous elamipretide (5d/wk) for 4 weeks starting after 6 weeks of ARAS or sham (n=6 each). After completion of treatment, STK renal blood flow (RBF) and glomerular filtration rate (GFR) were assessed in-vivo using multi-detector computed-tomography. Renal fibrosis and oxidative stress were analyzed in trichrome- and dihydroethidium-stained slides, respectively. Mitochondrial markers involved in the electrontransport chain (COX4, ATP/ADP ratio), biogenesis (PGC1α, PPARα), dynamics (MFN2, DRP1), and mitophagy (parkin, p62) were measured in the kidney using ELISA, western-blot, and immunohistochemistry. Cellular senescence (senescence-associated β-galactosidase and heterochromatin foci, phosphorylated-H2AX, and p16/21/53) and senescence-associated secretory phenotype (SASP; PAI-1, MCP-1, TGFβ, and TNFα) markers were studied by microscopy, quantitative reverse transcription-polymerase chain reaction, and western-blot.
Blood pressure was elevated whereas STK-RBF and GFR were decreased in ARAS pigs, and tissue scarring was increased. ARAS induced STK cellular senescence and accumulated dysfunctional mitochondria, which were associated with cardiolipin loss, upregulated mitochondrial biogenesis, and defective mitophagy. Elamipretide normalized STK-RBF and GFR, alleviated fibrosis and oxidative stress, and restored mitochondrial cardiolipin, biogenesis, and mitophagy in ARAS, but did not change SASP markers, and attenuated only senescenceassociated β-galactosidase activity and p53 gene expression.
Mitochondrial protection improved renal function and fibrosis in the ARAS STK, but only partly mitigated cellular senescence. This finding suggests that mitochondrial dysfunction may not be a major determinant of cellular senescence in the early stage of ARAS.
背景/目的：动脉粥样硬化性肾动脉狭窄（ARAS）可能导致肾损伤和线粒体功能障碍，这与细胞衰老有关。艾拉米肽是一种靶向线粒体的肽，可改善ARAS患者的肾功能，但它是否能减轻衰老尚不清楚。我们假设艾拉米肽可以减少ARAS患者衰老的狭窄肾脏（STK）。
将家猪随机分为对照组和单侧ARAS组，单侧ARAS组在ARAS或假手术6周后开始，未治疗或皮下注射艾拉米肽（每周5天）治疗4周（每组n = 6）。治疗结束后，使用多排螺旋计算机断层扫描在体内评估STK肾血流量（RBF）和肾小球滤过率（GFR）。分别在三色染色和二氢乙锭染色的切片上分析肾纤维化和氧化应激。使用酶联免疫吸附测定、蛋白质免疫印迹法和免疫组织化学法测量肾脏中参与电子传递链（COX4、ATP/ADP比率）、生物合成（PGC1α、PPARα）、动力学（MFN2、DRP1）和线粒体自噬（parkin、p62）的线粒体标志物。通过显微镜检查、定量逆转录-聚合酶链反应和蛋白质免疫印迹法研究细胞衰老（衰老相关β-半乳糖苷酶和异染色质灶、磷酸化-H2AX和p16/21/53）和衰老相关分泌表型（SASP；PAI-1、MCP-1、TGFβ和TNFα）标志物。
ARAS猪的血压升高，而STK-RBF和GFR降低，组织瘢痕增加。ARAS诱导STK细胞衰老并积累功能失调的线粒体，这与心磷脂丢失、线粒体生物合成上调和线粒体自噬缺陷有关。艾拉米肽使ARAS患者的STK-RBF和GFR恢复正常，减轻纤维化和氧化应激，并恢复线粒体心磷脂、生物合成和线粒体自噬，但未改变SASP标志物，仅减弱了衰老相关β-半乳糖苷酶活性和p53基因表达。
线粒体保护改善了ARAS患者STK的肾功能和纤维化，但仅部分减轻了细胞衰老。这一发现表明，线粒体功能障碍可能不是ARAS早期细胞衰老的主要决定因素。
