Nunes Kenia P, Bomfim Gisele F, Toque Haroldo A, Szasz Theodora, Clinton Webb R
Department of Physiology, Augusta University, USA; Department of Biological Sciences, Florida Institute of Technology, USA.
Federal University of Mato Grosso, Brazil.
Life Sci. 2017 Dec 15;191:219-226. doi: 10.1016/j.lfs.2017.10.014. Epub 2017 Oct 12.
Angiotensin II (AngII), a corpus cavernosum (CC) constrictor peptide, modulates Toll like receptor (TLR) expression, a key element of the innate immune system, contributing to impaired vascular function in pathological conditions. However, it is unknown whether TLR4 is involved in AngII-induced erectile dysfunction. In this study, we investigated whether TLR4 plays a role in cavernosal dysfunction caused by AngII upregulation.
Cavernosal smooth muscle cells (CSMC) from C57/BL6 mice were treated with AngII (0.1μM) or bacterial LPS (50ng/ml) for 12-24h and TLR4 expression was assessed. Mice were infused with AngII (90ng/min, 28days) and treated with anti-TLR4 antibody (0.1mg/daily, i.p.) for the last 14days of the treatment. CC tissue was used for functional studies and for Western blotting. Nitric Oxide Synthase (NOS) activity was measured by conversion of [H]-l-arginine to [H]-l-citrulline, systemic TNF-α levels by ELISA, and reactive oxygen species (ROS) by immunofluorescence.
We report upregulation of TLR4 in CSMC following AngII or LPS stimulation. In AngII-infused mice, chronic treatment with anti-TLR4 antibody (28±2.1%) attenuates adrenergic CC contraction, which also ameliorates nitrergic (68.90±0.21 vs. 51.07±0.63, 8Hz, AngII-infused mice treated vs. non-treated). Decreased endothelial NOS expression, reduced NOS activity, and augmented levels of TNF-α, and ROS were found following AngII-infusion. These alterations were prevented, or at least decreased by anti-TLR4 antibody treatment.
Inhibition of TLR4 ameliorates AngII-impaired cavernosal relaxation, decreases TNF-α levels, and restores NO bioavailability, demonstrating that TLR4 partly mediates AngII-induced cavernosal dysfunction.
血管紧张素II（AngII）是一种海绵体（CC）收缩肽，可调节Toll样受体（TLR）的表达，TLR是固有免疫系统的关键要素，在病理状态下会导致血管功能受损。然而，TLR4是否参与AngII诱导的勃起功能障碍尚不清楚。在本研究中，我们调查了TLR4是否在由AngII上调引起的海绵体功能障碍中发挥作用。
用AngII（0.1μM）或细菌脂多糖（LPS，50ng/ml）处理C57/BL6小鼠的海绵体平滑肌细胞（CSMC）12 - 24小时，并评估TLR4的表达。给小鼠输注AngII（90ng/分钟，持续28天），并在治疗的最后14天用抗TLR4抗体（0.1mg/天，腹腔注射）进行治疗。CC组织用于功能研究和蛋白质免疫印迹分析。通过[H]-L-精氨酸向[H]-L-瓜氨酸的转化来测量一氧化氮合酶（NOS）活性，通过酶联免疫吸附测定法测量全身肿瘤坏死因子-α（TNF-α）水平，通过免疫荧光法测量活性氧（ROS）。
我们报道了AngII或LPS刺激后CSMC中TLR4的上调。在输注AngII的小鼠中，用抗TLR4抗体进行慢性治疗（28±2.1%）可减弱肾上腺素能CC收缩，这也改善了硝酸能收缩（8Hz时，输注AngII的治疗组小鼠为68.90±0.21，未治疗组为51.07±0.63）。输注AngII后发现内皮型NOS表达降低、NOS活性降低、TNF-α水平升高以及ROS增加。这些改变通过抗TLR4抗体治疗得以预防或至少减轻。
抑制TLR4可改善AngII受损的海绵体舒张功能，降低TNF-α水平，并恢复一氧化氮的生物利用度，表明TLR4部分介导了AngII诱导的海绵体功能障碍。
