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Decreased Cyclic Guanosine Monophosphate–Protein Kinase G Signaling Impairs Angiogenesis in a Lamb Model of Persistent Pulmonary Hypertension of the Newborn
American Journal of Respiratory Cell and Molecular Biology ( IF 5.9 ) Pub Date : 2021-11-01 , DOI: 10.1165/rcmb.2020-0434oc
Megha Sharma 1 , Ujala Rana 2 , Chintamani Joshi 2 , Teresa Michalkiewicz 2 , Adeleye Afolayan 2 , Abdul Parchur 3 , Amit Joshi 3 , Ru-Jeng Teng 2 , Girija G Konduri 2
Affiliation  

Impaired angiogenesis function in pulmonary artery endothelial cells (PAEC) contributes to persistent pulmonary hypertension of the newborn (PPHN). Decreased nitric oxide (NO) amounts in PPHN lead to impaired mitochondrial biogenesis and angiogenesis in the lung; the mechanisms remain unclear. We hypothesized that decreased cyclic guanosine monophosphate (cGMP)–PKG (protein kinase G) signaling downstream of NO leads to decreased mitochondrial biogenesis and angiogenesis in PPHN. PPHN was induced by ductus arteriosus constriction from 128–136 days’ gestation in fetal lambs. Control animals were gestation-matched lambs that did not undergo ductal constriction. PAEC isolated from PPHN lambs were treated with the sGC (soluble guanylate cyclase) activator cinaciguat, the PKG activator 8-bromo-cGMP, or the PDE-V (PDE type V) inhibitor sildenafil. Lysates were immunoblotted for mitochondrial transcription factors and electron transport chain C-I (complex I), C-II, C-III, C-IV, and C-V proteins. The in vitro angiogenesis of PAEC was evaluated by using tube-formation and scratch-recovery assays. cGMP concentrations were measured by using an enzyme immunoassay. Fetal lambs with ductal constriction were given sildenafil or control saline through continuous infusion in utero, and the lung histology, capillary counts, vessel density, and right ventricular pressure were assessed at birth. PPHN PAEC showed decreased mitochondrial transcription factor levels, electron transport chain protein levels, and in vitro tube formation and cell migration; these were restored by cinaciguat, 8-bromo-cGMP, and sildenafil. Cinaciguat and sildenafil increased cGMP concentrations in PPHN PAEC. Radial alveolar and capillary counts and vessel density were lower in PPHN lungs, and the right ventricular pressure and Fulton Index were higher in PPHN lungs; these were improved by in utero sildenafil infusion. cGMP–PKG signaling is a potential therapeutic target to restore decreased mitochondrial biogenesis and angiogenesis in PPHN.



中文翻译:

减少的环磷酸鸟苷-蛋白激酶 G 信号转导损害新生儿持续性肺动脉高压羔羊模型中的血管生成

肺动脉内皮细胞 (PAEC) 血管生成功能受损导致新生儿持续性肺动脉高压 (PPHN)。PPHN 中一氧化氮 (NO) 量的减少导致肺中线粒体生物发生和血管生成受损;机制仍不清楚。我们假设 NO 下游的环磷酸鸟苷 (cGMP)-PKG (蛋白激酶 G) 信号传导减少导致 PPHN 中线粒体生物发生和血管生成减少。PPHN 是由胎儿羔羊妊娠 128-136 天的动脉导管收缩引起的。对照动物是未经历导管收缩的妊娠匹配羔羊。从 PPHN 羔羊中分离出来的 PAEC 用 sGC(可溶性鸟苷酸环化酶)激活剂 cinaciguat、PKG 激活剂 8-溴-cGMP 或 PDE-V(PDE V 型)抑制剂西地那非处理。针对线粒体转录因子和电子传递链 CI(复合物 I)、C-II、C-III、C-IV 和 CV 蛋白对裂解物进行免疫印迹。这通过使用管形成和划痕恢复测定法评估 PAEC的体外血管生成。通过使用酶免疫测定法测量cGMP浓度。有导管收缩的胎羔在子宫内连续输注西地那非或对照生理盐水,出生时评估肺组织学、毛细血管计数、血管密度和右心室压力。PPHN PAEC 显示线粒体转录因子水平、电子传递链蛋白水平降低,并且在体外管形成和细胞迁移;这些通过 cinaciguat、8-溴-cGMP 和西地那非恢复。Cinaciguat 和西地那非增加了 PPHN PAEC 中的 cGMP 浓度。PPHN 肺的径向肺泡和毛细血管计数和血管密度较低,而 PPHN 肺的右心室压力和富尔顿指数较高;这些通过子宫内西地那非输注得到改善。cGMP-PKG 信号传导是恢复 PPHN 中减少的线粒体生物发生和血管生成的潜在治疗靶点。

更新日期:2021-11-01
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