Tulane Hypertension and Renal Center of Excellence, Tulane University School of Medicine, New Orleans, LA 70112-2699, USA.
Department of Physiology, Tulane University School of Medicine, New Orleans, LA 70112-2699, USA.
Cells. 2023 May 28;12(11):1492. doi: 10.3390/cells12111492.
The current prevailing paradigm in the renin-angiotensin system dictates that most, if not all, biological, physiological, and pathological responses to its most potent peptide, angiotensin II (Ang II), are mediated by extracellular Ang II activating its cell surface receptors. Whether intracellular (or intracrine) Ang II and its receptors are involved remains incompletely understood. The present study tested the hypothesis that extracellular Ang II is taken up by the proximal tubules of the kidney by an AT (AT) receptor-dependent mechanism and that overexpression of an intracellular Ang II fusion protein (ECFP/Ang II) in mouse proximal tubule cells (mPTC) stimulates the expression of Na/H exchanger 3 (NHE3), Na/HCO cotransporter, and sodium and glucose cotransporter 2 (Sglt2) by AT/MAPK/ERK1/2/NF-kB signaling pathways. mPCT cells derived from male wild-type and type 1a Ang II receptor-deficient mice () were transfected with an intracellular enhanced cyan fluorescent protein-tagged Ang II fusion protein, ECFP/Ang II, and treated without or with AT receptor blocker losartan, AT receptor blocker PD123319, MEK1/MEK2 inhibitor U0126, NF-кB inhibitor RO 106-9920, or p38 MAP kinase inhibitor SB202196, respectively. In wild-type mPCT cells, the expression of ECFP/Ang II significantly increased NHE3, Na/HCO, and Sglt2 expression ( < 0.01). These responses were accompanied by >3-fold increases in the expression of phospho-ERK1/2 and the p65 subunit of NF-кB ( < 0.01). Losartan, U0126, or RO 106-9920 all significantly attenuated ECFP/Ang II-induced NHE3 and Na/HCO expression ( < 0.01). Deletion of AT (AT) receptors in mPCT cells attenuated ECFP/Ang II-induced NHE3 and Na/HCO expression ( < 0.01). Interestingly, the AT receptor blocker PD123319 also attenuated ECFP/Ang II-induced NHE3 and Na/HCO expression ( < 0.01). These results suggest that, similar to extracellular Ang II, intracellular Ang II may also play an important role in Ang II receptor-mediated proximal tubule NHE3, Na/HCO, and Sglt2 expression by activation of AT/MAPK/ERK1/2/NF-kB signaling pathways.
目前，肾素-血管紧张素系统的主流观点认为，其最有效肽段血管紧张素 II（Ang II）的大多数（如果不是全部）生物学、生理学和病理学反应都是通过细胞外 Ang II 激活其细胞表面受体来介导的。而细胞内（或胞内）Ang II 及其受体是否参与其中仍不完全清楚。本研究通过实验验证了以下假说：即细胞外 Ang II 通过 AT（AT）受体依赖性机制被肾脏近端小管摄取，并且在小鼠近端小管细胞（mPTC）中过表达细胞内 Ang II 融合蛋白（ECFP/Ang II）可通过 AT/MAPK/ERK1/2/NF-κB 信号通路刺激 Na/H 交换器 3（NHE3）、Na/HCO 共转运体和钠葡萄糖共转运体 2（Sglt2）的表达。从雄性野生型和 1a 型血管紧张素 II 受体缺陷型小鼠（）中分离出 mPCT 细胞，并用细胞内增强型青色荧光蛋白标记的 Ang II 融合蛋白 ECFP/Ang II 转染，并分别用 AT 受体阻滞剂洛沙坦、AT 受体阻滞剂 PD123319、MEK1/MEK2 抑制剂 U0126、NF-κB 抑制剂 RO 106-9920 或 p38 MAP 激酶抑制剂 SB202196 处理，而不进行处理。在野生型 mPCT 细胞中，ECFP/Ang II 的表达显著增加了 NHE3、Na/HCO 和 Sglt2 的表达（<0.01）。这些反应伴随着 ERK1/2 磷酸化和 NF-κB p65 亚单位的表达增加了 3 倍以上（<0.01）。洛沙坦、U0126 或 RO 106-9920 均显著减弱了 ECFP/Ang II 诱导的 NHE3 和 Na/HCO 的表达（<0.01）。在 mPCT 细胞中敲除 AT（AT）受体也减弱了 ECFP/Ang II 诱导的 NHE3 和 Na/HCO 的表达（<0.01）。有趣的是，AT 受体阻滞剂 PD123319 也减弱了 ECFP/Ang II 诱导的 NHE3 和 Na/HCO 的表达（<0.01）。这些结果表明，与细胞外 Ang II 类似，细胞内 Ang II 可能通过激活 AT/MAPK/ERK1/2/NF-κB 信号通路，在 Ang II 受体介导的近端小管 NHE3、Na/HCO 和 Sglt2 表达中也发挥重要作用。
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