药学学报  2022, Vol. 57 Issue (3): 557-567     DOI: 10.16438/j.0513-4870.2021-1126   PDF    
基于肺血管细胞增殖抑制的肺动脉高压治疗药物
张小楠1,2,3, 李占强1,2, 芦殿香1,2     
1. 青海大学高原医学研究中心, 青海 西宁 810001;
2. 高原医学教育部重点实验室, 青海省高原医学应用基础重点实验室 (青海-犹他高原医学联合重点实验室), 青海 西宁 810001;
3. 青海省人民医院, 青海 西宁 810007
摘要: 肺动脉高压(pulmonary hypertension, PH) 是一种进展迅速的肺血管疾病, 预后不良, 最终导致右心衰竭和死亡。肺小动脉的重塑是PH的重要病理特征。位于肺动脉中层的肺动脉平滑肌细胞(pulmonary arterial smooth muscle cells, PASMCs) 表现出类似于肿瘤细胞的异常增殖和抗凋亡特征, 是肺血管重塑的主要启动因素。本文主要讨论在PASMCs增殖中起关键作用的信号通路以及对细胞增殖通路为靶点的抑制剂的最新研究进展做一综述, 以期为PH的靶向治疗提供新的视角。
关键词: 肺动脉高压    肺动脉平滑肌细胞    肺血管重塑    细胞周期    增殖信号通路    
Treatment of pulmonary hypertension based on inhibition of pulmonary vascular cell proliferation
ZHANG Xiao-nan1,2,3, LI Zhan-qiang1,2, LU Dian-xiang1,2     
1. Research Center for High Altitude Medicine, Qinghai University, Xining 810001, China;
2. Key Laboratory of High Altitude Medicine, Ministry of Education; Key Laboratory of Application and Foundation for High Altitude Medicine Research in Qinghai Province (Qinghai-Utah Joint Research Key Lab for High Altitude Medicine), Xining 810001, China;
3. Qinghai Provincial People's Hospital, Xining 810007, China
Abstract: Pulmonary hypertension is a rapidly progressing disease of the lung vasculature with poor prognosis, ultimately leading to right heart failure and death. The remodeling of small pulmonary arteries represents an important pathological characteristic of pulmonary hypertension. Pulmonary arterial smooth muscle cells (PASMCs) located in the middle layer of pulmonary artery exhibit hyperproliferation and resistance to apoptosis, which is the main initiator of pulmonary vascular remodeling and similar to that seen in tumor cells. In this review we focus on the signaling pathways that play a key role in PASMCs proliferation and the latest research progress on inhibitors targeting cell proliferation pathways to provide a new perspective for the treatment of PH.
Key words: pulmonary hypertension    pulmonary arterial smooth muscle cell    pulmonary vascular remodeling    cell cycle    proliferative signaling pathway    

在生理条件下, 成人肺循环通常是低阻力、低压力、高流量系统, 平均肺动脉压(mean pulmonary arterial pressure, mPAP) 小于20 mmHg。肺动脉高压(pulmonary hypertension, PH) 是指在海平面、静息状态下, 经右心导管检查(right heart catheterization, RHC) 测定时mPAP ≥ 25 mmHg (1 mmHg = 0.133 kPa)。正常成年人静息状态下mPAP为14.0 ± 3.3 mmHg, 其上限不超过20 mmHg[1]。WHO根据潜在的病因, 病理学和血流动力学特征将PH分为5类: ①动脉性PH [pulmonary arterial hypertension, PAH, 世界卫生组织(WHO) 第1组]; ②继发于左心疾病所致PH (WHO第2组); ③肺部疾病和(或) 低氧所致PH (WHO第3组); ④慢性血栓栓塞性PH (chronic thromboembolic pulmonary hypertension, CTEPH, WHO第4组); ⑤未名和(或) 多因素所致PH (WHO第5组)[2]

PH是由多种异源性疾病(病因) 和不同发病机制所致肺血管结构和功能改变, 引起肺血管阻力和肺动脉压力升高的临床和病理生理综合征, 通常会发展为右心衰竭甚至死亡。肺血管重塑(pulmonary vascular remodeling, PVR) 的病理特征包括中膜肥大/增生, 内膜和外膜纤维化, 血管腔呈现同心性和丛状病变、原位血栓形成及血管周围炎症细胞(B和T淋巴细胞、肥大细胞、树突状细胞和巨噬细胞等) 的浸润[3]。血管重塑主要累及远端肌型肺动脉和毛细血管前小动脉, 但也不同程度地累及毛细血管后静脉和支气管动脉, 但是后者的机制尚不明确[4]

肺动脉内皮细胞(pulmonary arterial endothelial cells, PAECs)、肺动脉平滑肌细胞(pulmonary artery smooth muscle cells, PASMCs) 和成纤维细胞(pulmonary arterial fibroblasts, PAFs) 过度增殖和抗凋亡导致肺血管阻塞、肺动脉压力升高和右心室负荷过重[5]。其中, 位于肺动脉中层的PASMCs是肺动脉壁的主要组成部分, PH患者的PASMCs表现出类似肿瘤细胞的表型, 表现为不受控制的复制潜力、逃避生长抑制因子、抗凋亡、DNA不稳定性、特定信号转导过程的激活以及细胞代谢的改变和炎症[6]。同样, 慢性炎症、病理性的血管生成和免疫逃避也是PH和癌症共同的发病机制[7]。从某些方面, PH甚至可以被认为是一种假性恶性疾病[7]。因此, 对于PH的治疗不仅需要考虑肺血管收缩的问题, 还必须通过抑制增殖和激活抗增殖机制来解决PVR的问题。本文重点讨论在肺血管细胞增殖方面起关键作用的信号通路以及可用于改善PH预后的药物(表 1)[8-18]在PH治疗中的临床应用前景, 以期为PH的发病机制及最新的治疗措施的研究提供一些思路。

Table 1 The list of pharmaceutical drugs for pulmonary hypertension (PH). po: Per os; qd: Quaque die; bid: Bis in die; ig: Intragastrical; sc: Subcutaneous; ip: Intrapertoneal; iv: Intravenous; ivgtt: Injection venosa gutta; PDGFR: Platelet-derived growth factor receptor; PASMCs: Pulmonary artery smooth muscle cells; PAH: Pulmonary arterial hypertension; PAECs: Pulmonary artery endothelial cells; EGFR: Epidermal growth factor receptor; MCT-PH: Monocrotaline-induced PH; HPH: Hypoxia-induced pulmonary; FGFR: Fibroblast growth factor receptor; Raf: Rapidly accelerated fibrosarcoma; PI3K: Phosphatidylinositol 3-kinase; Akt: Protein kinase B; PAAFs: Pulmonary artery adventitial fibroblasts; mTOR: Mechanistic target of rapamycin; mTORC1: mTOR complex 1; mTORC2: mTOR complex 2; DAPT: N-[N-(3, 5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine); FOXO: Forkhead-box class O; SuHx: Sugen/hypoxia; PPAR-γ: Peroxisome proliferator-activated receptor gamma; HIF1α: Hypoxia inducible factor 1α; HIF2α: Hypoxia inducible factor 2α
1 肺血管细胞增殖与细胞周期调控失调 1.1 PH与肺血管细胞增殖

PH肺血管细胞增殖/凋亡失衡机制与肿瘤细胞有很多相似之处, 因此结合这两个疾病之间交叉的信号通路, 使用抗癌药物治疗PH可能为PH的治疗带来新的机会。PH肺血管细胞的异常增殖反应发生在血管壁的3层, 涉及3种类型的血管细胞: PAECs、PASMCs和PAFs[19]。研究发现, 在PH患者及PH动物模型来源的原代细胞系, PAECs、PASMCs和PAFs均存在异常增殖、抗凋亡及类癌样表型[20, 21]。虽然PH发生的确切触发因素尚不清楚, 但有几个已知的危险因素和疾病发生的驱动因素, 包括生长因子及炎症因子刺激、基因突变、暴露在缺氧环境或药物/毒素中毒等[22], 促使细胞表面受体激活, 作用于下游促增殖、抗凋亡的内源性信号转导通路, 诱导细胞进入复制周期, 进而出现异常增殖、抗凋亡的细胞表型[23] (图 1)。因此针对这类在肿瘤和PH发生中调节失调的信号通路为靶点, 通过抑制细胞周期由G1期向S期的过渡, 靶向作用于增殖/凋亡失衡的细胞群来逆转PVR, 希望达到治疗PH的目的。

Figure 1 The proliferative signaling pathway of pulmonary vascular remodeling. Ras: Rat sarcoma; MEK: Mitogen extracellular kinase; Hes: Hairy/enhancer of split
1.2 PH与细胞周期调控

细胞增殖与静止之间的平衡受许多介质精细的调节, 细胞周期蛋白依赖性激酶(cyclin-dependent kinases, CDKs) 家族的几个成员协同驱动细胞周期的复杂事件。细胞周期从G1期到S期的过渡取决于CDK1、CDK2、CDK4和CDK6的活性[24]。在细胞周期G1期末, 有一个决定细胞是否进行有丝分裂的关键时期, 称为限制点(restriction point, R point), R点的分子基础被确认为是视网膜母细胞瘤(retinoblastoma, Rb) 蛋白的过度磷酸化, Rb蛋白是一种抗肿瘤和抗增殖的蛋白[25]。D型细胞周期蛋白(D-type cyclins, cyclin D) 与CDK4/6结合, 形成CDK4/6-cyclin D复合物, 然后进入细胞核被CDK激活激酶(CDK-activating kinase, CAK) 激活。CDK4/6-cyclin D复合物在G1期早期部分磷酸化Rb蛋白(低磷酸化)[26]。低磷酸化状态的Rb蛋白部分降低了E2F转录因子家族的活性, 从而促进了E2F靶基因的表达, 包括E-型周期蛋白(cyclins E1和E2) 的表达, 在G1后期, 随着cyclin E的表达升高, 结合并激活CDK2, 形成CDK2-cyclin E复合物, 过度磷酸化Rb使Rb失活, 从而使E2F完全激活并进入S期[25, 27]

CDK4/6和CDK2活性受到内源性CDK抑制剂的调节, 这些抑制剂可以有效地抑制CDKs的活性, 导致G1期停滞, 从而限制细胞增殖。内源性CDKs抑制剂主要有两大类, 第一类是INK4家族, 由p16INK4A、p15INK4B、p18INK4C和p19INK4D组成。这些蛋白与CDK4和CDK6结合, 形成缺乏激酶活性的复合物。第二类是CIP/KIP家族, 包括p27KIP1、p21CIP1和p57KIP2。这些抑制剂可以有效地抑制几种CDK (包括CDK4/6、CDK2和CDK1) 的活性[26]。在癌细胞中, CDK-cyclin复合物的活性经常受诸多因素的调控, 其中包括内源性CDK抑制剂水平降低, 导致细胞周期蛋白CDK复合物活性持续升高, 从而使细胞生长失控[28, 29]。PH肺血管增殖/凋亡失衡机制与肿瘤的发病机制有相似之处。在PH和癌症的发病过程中有几个类似的信号通路, 包括Ras/Raf/MEK、PI3K/Akt/mTOR、Notch-Hes和Wnts信号通路等。这些通路在PH的发病过程中通常处于失调状态, 并且作为G1/S细胞周期检查点的正向调节因子, 通过多种机制调控细胞的生长、增殖、凋亡(图 2)。

Figure 2 The major proliferative signaling pathways and drug targets in pulmonary hypertension and cancer. RTKs: Receptor tyrosine kinases; ERK: Extracellular signal-regulated kinase; GSK3β: Glycogen synthase kinase 3β; GSI: γ-Secretase inhibitor
2 与PH肺血管细胞增殖相关的生长因子受体

酪氨酸激酶受体(receptor tyrosine kinases, RTKs) 是生长因子、细胞因子和激素的高亲和性细胞表面受体, 是位于细胞膜的跨膜受体蛋白。与PH相关的RTKs包括血小板源性生长因子(platelet-derived growth factor, PDGF)、表皮生长因子(epidermal growth factor, EGF)、成纤维细胞生长因子(fibroblast growth factor, FGF)、血管内皮生长因子(vascular endothelial growth factor, VEGF) 和神经生长因子(nerve growth factor, NGF) 受体家族。两条主要的RTK下游信号通路是MEK/ERK信号通路和PI3K/Akt信号通路。

有关RTKs及其配体PDGF、EGF、FCG、NCG等对PASMCs和PAECs过度增殖和迁移的作用已在人和PH动物实验模型中得到证实[30]。几种RTK抑制剂(tyrosine kinase inhibitors, TKIs) 被用于治疗不同种类的癌症, 包括伊马替尼(imatinib)、达沙替尼(dasatinib) 和尼罗替尼(nilotinib), 多靶点多激酶抑制剂索拉非尼(sorafenib) 和舒尼替尼(sunitinib) (抑制PDGF和VEGF受体)。Imatinib是第一个用于PH临床研究的RTK, 也是第一个用于PH直接靶向干预血管重塑的药物。研究也发现imatinib可以抑制啮齿类动物和人的PASMCs的增殖和迁移, 促进PASMCs的凋亡[31, 32]; imatinib逆转了两种PH动物模型的PVR, 阻止PDGFR-b的磷酸化以及下游信号的转导[33]。然而, 在一项Ⅲ期临床试验中, 尽管imatinib降低了肺血管阻力, 但由于风险/收益不成正比, imatinib未被批准用于临床治疗PH, 因为治疗组PH患者的疾病进展没有被缩短; 此外, 在8名同时接受imatinib和维生素K拮抗剂抗凝治疗的患者中出现了硬膜下血肿[34]。但是研究者对使用imatinib做进一步试验的兴趣仍然很高。Imatinib在PH治疗中所显示的疗效为寻找更具优势的TKIs提供了研究思路, 不幸的是, 目前所研究的TKIs没有一个达到预期研究的目标。Nilotinib是另一种比imatinib更有效的选择性抑制BCR (breakpoint cluster region)-ABL的TKI, 其已被证实可以预防啮齿类动物系统性硬化症PH的血管增生[35]。然而, nilotinibd的Ⅱ期临床试验因严重的不良反应而终止(NCT01179737)。也有报道显示, TKI家族中的其他一些药物可以诱发PH, 如用dasatinib治疗白血病可引起PH[36, 37]。另一种蛋白激酶抑制剂Sugen5416在低氧暴露的情况下也能诱导啮齿动物的PH[38]。尽管一些TKIs为治疗PH带来了新希望, 但还需要更多的研究来评估其临床应用前景。

3 与PH肺血管细胞增殖相关的信号通路 3.1 Ras/Raf/MEK/ERK信号通路

Ras/Raf/MEK/ERK是癌症发生的一个重要信号通路, 其在癌症中的作用已被详细描述[39], 致癌的Ras、Raf突变已在人类多种癌症中被发现[40]。在PAEC中, 骨形态发生蛋白受体2 (bone morphogenetic protein type Ⅱ receptor, BMPR2) 的沉默可以导致Ras/Raf/MEK/MAPK/AP1信号系统激活, 进而引起PAEC增殖[41]。此外, 有学者发现在晚期PH患者的肺血管系统中MAPK表达异常[42]。最近的研究表明, 功能获得性Raf突变与两名婴儿的致命性PAH发病相关, 这可能是Raf-1激酶和Ras/MAPK通路的结构性激活所致[43]。同时, 在探索Ras在PH中的重要性时, 发现miR-455-3P-1通过下调Ras/ERK信号通路中关键因子的表达, 抑制PASMCs增殖, 从而降低肺动脉高压[44]。因此, Ras/Raf/MEK/MAPK信号通路可能是治疗PH病理性血管重塑的信号途径及治疗靶点。索拉非尼(sorafenib, 最初被确定为Raf-1抑制剂, 随后被确定为PDGFR、VEGFR、Kit和Flt-3的抑制剂) 已被证明在PH和右心室肥厚的实验动物模型中发挥了有效的作用[45, 46]。2010年, 关于sorafenib治疗PAH患者的安全性、耐受性和疗效评估的一份报告显示, 接受sorafenib治疗(每次200 mg, 2次/日, 口服) 的12名PAH患者对于该药有较好的耐受性[47]。2017年的一项研究证明, sorafenib对重度肺动脉高压和/或右心衰竭患者有很好的治疗效果, 接受治疗的9名患者中, 仅有一名患者出现全身皮肤不良反应而终止试验, 56%的患者虽然出现了轻微的皮肤不良反应, 但也能耐受sorafenib的治疗, 并明显改善了血流动力学指标[48]。因此, sorafenib是治疗难治性PH患者的一种替代策略, 但其临床安全性有待于进一步评估。

3.2 PI3K-Akt-mTOR信号通路

PI3K-Akt-mTOR通路失调是人类肿瘤中最常见的信号通路之一, 目前正在积极开展其作为抗癌治疗靶点的研究[49]。最近的研究表明, PI3K-Akt-mTOR通路可被多种细胞因子激活, 如缺氧诱导因子-1α (hypoxia inducible factor-1α, HIF1α)、PDGF、磷酸酶和紧张素同源物(phosphatase and tensin homolog, PTEN) 等。在低氧诱导的PH模型中, PDGF激活PI3Ks/Akt通路, 诱导肺动脉中膜PASMCs转录因子cAMP反应元件结合蛋白(cAMP response element binding protein, CREB) 的丢失, 促进PASMCs从静止的、收缩表型向增殖、迁移、去分化的合成表型转换, 从而导致PVR[12]。体外研究表明, PI3Ks在生长因子诱导的PASMCs增殖和迁移中起关键作用[50]。在包括特发性肺动脉高压(idiopathic pulmonary artery hypertension, IPAH) 和CTEPH患者以及多种PH动物模型, 包括低氧性PH (hypoxia pulmonary hypertension, HPH)、缺氧/Sugen诱导PH (hypoxia/Sugen induced pulmonary hypertension, SuHx) 和野百合碱诱导PH (monocrotaline-induced PH, MCT-PH) 的动物模型中均发现Akt/mTOR促进PASMC增殖[51-55]

已有研究表明PI3K-Akt-mTOR信号通路在肺血管重塑中起重要作用, 因此使用PI3K、Akt、mTOR的特异性抑制剂被认为是开发新的、特异性治疗PAH的一种很好的治疗策略。PH动物模型的药理学实验发现, 靶向PI3K或Akt的药理抑制剂薯蓣皂甙(dioscin) 可以减轻HPH大鼠的PVR的进展[56]。mTOR抑制剂雷帕霉素(rapamycin, RAP) 在临床上常用于器官移植的抗排斥反应和自身免疫性疾病的治疗[57]。有关使用RAP干预PH的临床前研究有较大争议, 多篇文献报道了不同的实验结果。在联合肺切除和MCT-PH的大鼠模型中(全肺切除后7天, MCT注射前2天) 给予RAP (2.5 mg·kg-1, 1次/日) 可减少PVR和右心室肥厚指数(right ventricular hypertrophy, RVH), 但在逆转模型(全肺切除15天) 给予上述剂量的RAP不能减轻PVR和RVH[58]。在HPH小鼠中, RAP (从缺氧3周后开始, 3 mg·kg-1, 1次/日) 减轻了肺血管的重塑和随后的RVH[59]。预防性给予MCT-PH大鼠大剂量RAP (5 mg·kg-1·d-1, MCT注射当日), 可减轻PVR、肺动脉压力和RVH[51]。mTORC2抑制剂PP242, 能够诱导PASMCs凋亡, 逆转HPH大鼠的肺血管重塑。目前, mTORC2抑制剂正在进行某些癌症的临床研究, 还没有针对PH患者的临床研究[60]。现正在进行NAB-rapamycin (ABI-009, 一种白蛋白结合型rapamycin) 防治PH的Ⅰ期临床试验(Clinical Trials. Gov identifier: NCT02587325)。预计在不久的将来, 这项临床研究结果将有助于开发新的治疗PH策略。

3.3 Notch-Hes信号通路

Notch蛋白是一种细胞膜受体, 在细胞间的信息传递中起重要作用[61]。目前已在哺乳动物中发现了Notch1~4四种受体和Jagged1、Jagged2、Delta1、Delta3和Delta4五种配体[62]。Notch信号通路通过诱导靶基因转录影响细胞分化、成熟、增殖和凋亡, 并在血管平滑肌细胞(vein smooth muscle cell, VSMC) 表型转换(收缩到增殖) 的调节中起关键作用[63]。研究发现, Notch信号的靶基因包括编码发状分裂相关增强子(hairy enhancer of split, Hes) 基因家族、Hes相关蛋白(Hes related proteins, Hey) 家族、c-myc基因和cyclin D等[64]。其中Notch-Hes信号通路对调控VSMC和血管内皮细胞的增殖、分化、凋亡具有重要作用, 参与动脉粥样硬化、PH、心肌梗死等疾病的发生[62]。Hes蛋白是一类包含碱性的螺旋-环-螺旋状结构域(basic helix-loop-helix, bHLH) 的转录因子。人类Hes蛋白家族由Hes1~7七种蛋白组成。目前认为, Hes1、Hes5、Hes7参与Notch信号通路, 可作为Notch效应分子发挥作用[65]。由于Notch3主要存在于VSMC中, 并参与调控VSMCs表型转换, 因此它被认为是包括PH在内的干预血管病理性重塑的候选靶点[66]。腺病毒NICD3 (Notch intracellular domain 3) 在PASMCS中过表达可明显促进细胞增殖, 同时上调Hes1转录因子的表达, 下调p27KIP1细胞周期蛋白的表达, 而p27KIP1的下降可以被Hes1 siRNA抑制[67]。Hes5也参与了Notch3诱导的肺动脉高压中PASMCs的增殖。与Notch3一样, Hes5在大鼠和人的肺小动脉的PASMCs中特异性表达, 利用siRNA敲低Hes5基因的表达, PH患者的PASMCs增殖率比正常供体的PASMCs减低, 说明Notch3-Hes通路在PH的PASMCs增殖中的作用更为显著[68]。在MCT诱导的PH大鼠PASMCs中, NICD3表达的增加与Hes1上调和p27KIP1表达下调有关, 并且发现Notch3-Hes5 mRNA表达的增加与mPAP呈正相关[69]。抑制Notch3-Hes信号通路可能有望成为干预和治疗PH的潜在靶点。

γ-分泌酶抑制剂(γ-secretase inhibitor, GSI) DAPT (N-[N-(3, 5-difluorophenacetyl)-L-alanyl]-S-phenylglycine) 是一种治疗阿尔茨海默病的药物, 先前的研究显示, DAPT可在体内抑制Notch裂解, 减少各种实验性PH的病理过程。在HPH小鼠模型中, 小鼠右心室收缩压升高、右心室肥厚, 伴随着Notch通路被激活, 发生PVR, 给予小鼠腹腔注射Notch信号抑制剂DAPT, 能够抑制PASMCs的过度增殖, 降低肺动脉压力及减轻PVR[70]。在MCT-PH大鼠模型上也证实了DAPT的作用, 给予DAPT治疗组肺动脉肌化程度较对照组减轻, 减缓了PH的发生[14]。丙基硫氧嘧啶(propylthiouracil, PTU) 是另一种GSI, 也已经在MCT-PH模型中进行了体内研究, 得到了与给予DAPT干预类似的结果。每日给予PTU干预可显著减少MCT-PH大鼠肺动脉中Notch3的表达, 肺动脉肌化程度较对照组减轻[71]。GSI在PH模型中的作用表明这些药物有可能成为用于治疗人类PH的新方法, 目前这类药物已经用于治疗某些Notch信号失调的癌症的临床前研究[72]

3.4 Wnt信号通路

由Wnt基因调控的信号转导途径称为Wnt信号通路, 在调节细胞增殖、分化、迁移、黏附和干细胞自我更新方面发挥重要作用[73]。已有研究报道, 根据Wnt信号通路的激活是否依赖β-catenin分为经典Wnt信号通路[Wnt/β-catenin (Wnt/bC) 通路] 和非经典Wnt信号通路[包括Wnt/planar cell polarity (Wnt/PCP) 通路和Wnt/Ca2+通路)][74]。Wnt/Ca2+途径参与磷脂酶C (phospholipase C, PLC) 和蛋白激酶C (protein kinase C, PKC) 的激活, 并参与细胞的增殖过程[75]。在没有细胞外Wnt配体的情况下, β-catenin被磷酸化并抑制下游基因的表达。当Wnt与细胞表面受体结合时, β-catenin被移位到细胞核, 并调控与细胞增殖和存活相关的下游靶基因的表达[76, 77]

已有较多证据表明Wnt信号通路参与PASMCs的增殖[78]。Quasnichka等[79]认为, 生长因子通过刺激β-catenin/T细胞因子(T cell factor, TCF) 信号激活Wnt通路, 调控cyclin D1和p21CIP1基因的表达, 进而调节VSMCs增殖。在MCT诱导的PH大鼠模型中, 糖原合成酶激酶3β (glycogen synthase kinase 3β, GSK3β) 过表达显著增加ERK磷酸化, 促进PASMCs增殖, 这一途径可以被Wnt通路调节[80]。Yu等[81]发现Wnt 5a下调β-catenin及其靶基因cyclin D1的表达, 抵抗缺氧诱导的人PASMCs的增殖。最近的一项研究发现, 激活Wnt/PCP通路是建立人肺内皮细胞-外膜细胞相互作用所必需的。Wnt/PCP信号的丢失可能会降低PH中新生血管的活性, 从而导致PH的发生[82]。鉴于Wnt通路在肺动脉高压肺血管生成和PVR中的关键作用, 预测调控Wnt信号通路的上游和下游靶基因, 可能是一种潜在的治疗PH的策略。

4 与PH肺血管细胞增殖相关的转录因子 4.1 叉头转录因子(forkhead box, FOX)

FOX蛋白家族由19个转录因子组成, 为FOXA、FOXC、FOXO、FOXP和FOXM 5个亚群[83]。FOXO在肿瘤细胞中参与许多重要的生物学活动, 包括细胞凋亡、氧化应激、DNA修饰和细胞代谢等, 作用于几种致癌信号通路的下游, 如PI3K-Akt、ERK等, 基于FOXO的抗凋亡和促增殖功能, 其被认为是肿瘤抑制因子[84, 85]。在PH患者以及MCT-PH和Hyp/SU-PH大鼠的PASMCs中, 总FOXO1蛋白表达水平降低, 磷酸化FOXO1水平升高, 总FOXO1/磷酸化FOXO1比值的变化与PASMCs增殖、PH的发生有关。在动物模型中, FOXO1的激活可以逆转PH。因此, 靶向FOXO1通路可能是治疗PH的一个可行的策略[15]。FOXM1通过调节细胞周期的G1/S和G2/M转变保持有丝分裂纺锤体的完整性, 从而发挥促进细胞增殖、血管生成、上皮-间充质转化和DNA损伤修复的作用[86]。最近的研究表明, FOXM1在PH患者和动物模型的PASMC中的表达水平升高, 并通过促进PASMCs的增殖和去分化以及抑制PASMCs的凋亡而促进PVR。抑制PASMCs特异性FOXM1可以逆转血管重塑并抑制PH的发展[87-90]。这些发现提示, FOXM1在PH的发生发展过程中也起着重要作用, 抑制FOXM1可能是治疗PH的一种有前途的治疗策略[91]

紫杉醇是一种微血管稳定剂, 目前被用作多种癌症的化疗药物, 具有增加FOXO1表达和抑制FOXO1磷酸化的作用。研究发现, 无论是通过静脉注射或吸入紫杉醇重建FOXO1活性, 还是通过基因治疗重建FOXO1的转录活性, 都可以通过调节多种细胞周期信号通路相关因子(cyclin D1、p27KIP1、B细胞淋巴瘤/白血病-2基因和生长阻滞与DNA损伤基因) 的表达和促进BMPR2信号转导, 在体外恢复静止期PASMC的表型, 并可逆转体内肺血管重构和右心肥厚。此外, 紫杉醇可能通过下调FOXM1的表达对PH大鼠起治疗作用[15]

4.2 PPAR-γ

过氧化物酶体增殖物激活受体γ (peroxisome proliferator-activated receptor gamma, PPAR-γ) 属于一组核受体蛋白, 具有转录因子的功能, 当被激活时, PPAR-γ可提高胰岛素敏感性, 并具有抗炎特性。临床前研究和人体研究都证明PPAR-γ参与了PH的发生[92, 93]。罗格列酮和吡格列酮(用于治疗2型糖尿病的PPAR-γ激动剂) 通过激活PPAR-γ信号通路抑制实验动物PH的发生。然而, 尽管临床前实验结果很令人满意, 但由于吡格列酮被质疑可能会导致膀胱癌, 故提前终止了吡格列酮的临床试验(ClinicalTrials.gov identifier: NCT00825266)。由于可能存在心血管不良反应, 罗格列酮也已在某些国家被退出市场[94]。并且两种药物的临床前剂量明显高于患者使用的最大剂量。目前, 靶向PPAR-γ的新疗法的探索仍在继续。在最近发表的一项研究中, 西地那非和罗格列酮联合吸入疗法被应用于PH大鼠的治疗, 发现两者合用可以改善PH大鼠的预后[95]。目前临床上使用不同药物的简单加和, 并无两种药物的联合递送方案, 因此联合递送药物的临床前研究成为炙手可热的研究方向之一[96]

4.3 HIF

HIF是异源二聚体转录因子, 包括组成性表达的β亚基和对氧敏感的α亚基。目前已鉴定出3个HIFα亚基, 包括HIF1α、HIF2α和HIF3ααβ亚基在氨基酸和结构上都具有很高的同源性, 在短期缺氧暴露下, HIF1α、HIF2α和HIF3α具有类似的依赖于氧气的调节方式[97, 98]。所有HIF的α亚基都受到相同的依赖于脯氨酰羟化酶(prolyl hydroxylase domain, PHD) 降解机制的影响[99]。现已鉴定出数千个人类缺氧诱导因子的靶基因, 这些基因共同参与调控细胞增殖、血管新生、能量代谢、血管运动等反应过程[100, 101]

目前, 有关HIF1α和HIF2α亚型在PH发生中的作用已有大量研究。第一个HIF1α和HIF2α参与PH发生的直接证据来自于HIF1α或HIF2α基因敲除小鼠[102, 103], 研究发现, 纯合子HIF1α或HIF1β基因敲除小鼠因心脏和血管问题无法存活。HIF2α完全缺失小鼠50%出现胚胎死亡[102]。然而, 部分敲除HIF1α或HIF2α基因的小鼠仍能正常存活, 当暴露在慢性缺氧(10%) 的环境中时, 与对照组相比, 这些小鼠的肺动脉压、右心室肥厚指数及PVR减轻, 提示HIF系统在PH的发病过程中起重要作用[102, 104]。研究者用PASMCs特异性HIF1α敲除小鼠研究了HIF1α在调节肺血管张力和PVR中的作用, 发现HIF1α的选择性缺失改善了慢性低氧诱导的小鼠肺血管的重塑, 减轻了小鼠的肺动脉高压[105]。此外, 体外研究结果发现, 低氧暴露或HIF1α过表达加速PASMCs的增殖, 低氧条件下收缩型PASMCs表型被合成型表型取代[106, 107]。有学者研究了PH小鼠肺血管组织特异性的HIFα表达谱, 发现HIF2α在PAECs中高表达[105, 108]。PAECs中特异性HIF2α基因敲除可以防止与PH相关的PVR的发生和发展, 在同一模型中HIF1α的缺失对小鼠几乎没有影响, 作者认为PAECs中的HIF2α对该疾病发生具有重要作用[109]。另有几个研究小组报道, 在小鼠中, PAECs中PHD2的缺失导致HIF2α的异常稳定, 并导致闭塞性血管病变和严重的肺动脉高压[110, 111]。HIF信号通路的激活参与了PH的发病过程, HIF1α和HIF2α在PH的PVR中起重要作用。了解HIF信号通路的分子机制有助于开发用于PH治疗的新药物。

由于HIF1α和HIF2α基因序列有高度同源性, 氧依赖调控机制有相似性, 已有几个化合物被证明可以同时抑制HIF1α和HIF2α的转录活性。Zhang等[112]已经报道了心脏苷类, 如地高辛对HIF1α和HIF2α蛋白合成均有抑制作用, 但对HIF2α的抑制作用低于HIF1α。吖啶黄素是一种具有抗菌活性的染料, 可与HIF1α和HIF2α的PAS-B结构域结合, 并抑制HIF的二聚化和转录活性。动物体内实验结果表明, 地高辛治疗可以预防和逆转HPH小鼠PH的发展, 吖啶黄素可以减轻HPH小鼠的右心室收缩压, 改善右心室肥厚[16]。但是由于HIF的亚型特异性及上述药物的不良反应, 其研究仍处于动物实验阶段。PT2567是Peloton Therapeutics公司研发的一种有效的、选择性的口服HIF2α抑制剂, 用于非肿瘤适应症, 最开始针对PH, 目前仍在探索其在炎症性肠病中的开发潜力, 处于临床前研究阶段[113]。国内Hu等[17]利用HIF2α抑制剂PT2567治疗HPH大鼠, 发现PT2567通过抑制炎症因子(C-X-C基序趋化因子受体4、细胞间黏附分子-1和基质细胞衍生因子-1) 及肺血管细胞增殖相关信号通路基因(分化抑制因子1、分化抑制因子13和转录生长因子α) 的表达, 显著抑制了HPH大鼠肺血管细胞增殖, 降低了mPAP, 减轻了PVR。化合物76 (C76) 是一种选择性HIF2α抑制剂, 研究者将C76用于人肺微血管内皮细胞和Egln1Tie2小鼠, 以及Sugen5416/Hyp和MCT-PH大鼠, 发现C76降低了Egln1Tie2Cre小鼠和Sugen5416/HyP大鼠的右心室收缩压, 改善了右心室肥厚, 在MCT大鼠中, C76可降低右心室收缩压、改善肺血管重塑和右心室肥大, 并能够降低死亡率[18], 这些结果表明, 药物抑制HIF2α是治疗PAH患者严重的肺血管重构和右心衰竭的一种有前途的新治疗策略。

5 结论和展望

近年来, 对于PH治疗方面的研究有较大进步, 但这种难治性的疾病的预后仍然比许多癌症差, 3年存活率仅为68%~70%[114]。目前, 对于PH的药物治疗仍以扩张血管为主, 而不是增殖性血管重塑, 随着对PH发病机制研究的深入, 现在公认, 必须通过抑制增殖和激活抗增殖机制来解决血管重塑问题。其中RTKs、Ras/Raf/MEK、PI3K/Akt/mTOR、Notch-Hes、FOXs、PPAR-γ、Wnt和HIF等信号通路、转录因子通过调控上下游靶基因的表达, 参与了肺血管细胞的增殖、迁移和凋亡, 促进PVR和肺动脉高压的发生。现有的研究表明, 通过肺血管增殖信号通路抑制剂的干预, 对肺血管细胞增殖所致的重塑有较好的逆转作用。了解靶向调控肺血管细胞增殖、迁移和血管生成的信号通路是开发有效治疗和预防PH药物的关键方向。但是, 靶向信号通路的抑制剂仍处于临床前实验阶段, 在治疗PH方面的应用有限。目前, 研究中只有一些针对增殖信号通路的抑制剂被用于抑制PASMCs的增殖和逆转PVR, 然而, 由于其严重的不良反应, 它们的临床应用受到限制, 进入临床开发研究的药物中约有50%因疗效差而失败, 25%因不良反应而失败[115]。在将抑制剂转化为临床应用之前, 还需要更多的验证, 以期为PH靶向药物的研发提供依据。

作者贡献: 张小楠负责查阅文献、撰写文章; 李占强负责文章选题、指导撰写思路; 芦殿香负责指导撰写思路、文章修改。

利益冲突: 所有作者均声明不存在利益冲突。

参考文献
[1]
Pulmonary Embolism and Pulmonary Vascular Disease Group R and Society CMA. Chinese guidelines for the diagnosis and treatment of pulmonary hypertension (the 2021 version)[J]. Natl Med J China (中华医学杂志), 2021, 101: 11-51.
[2]
Simonneau G, Montani D, Celermajer DS, et al. Haemodynamic definitions and updated clinical classification of pulmonary hypertension[J]. Eur Respir J, 2019, 53: 1801913. DOI:10.1183/13993003.01913-2018
[3]
Rafikova O, Al Ghouleh I, Rafikov R. Focus on early events: pathogenesis of pulmonary arterial hypertension development[J]. Antioxid Redox Signal, 2019, 31: 933-953. DOI:10.1089/ars.2018.7673
[4]
Humbert M, Guignabert C, Bonnet S, et al. Pathology and pathobiology of pulmonary hypertension: state of the art and research perspectives[J]. Eur Respir J, 2019, 53: 1801887. DOI:10.1183/13993003.01887-2018
[5]
Schermuly RT, Ghofrani HA, Wilkins MR, et al. Mechanisms of disease: pulmonary arterial hypertension[J]. Nat Rev Cardiol, 2011, 8: 443-455. DOI:10.1038/nrcardio.2011.87
[6]
Guignabert C, Tu L, Le Hiress M, et al. Pathogenesis of pulmonary arterial hypertension: lessons from cancer[J]. Eur Respir Rev, 2013, 22: 543-551. DOI:10.1183/09059180.00007513
[7]
Boucherat O, Vitry G, Trinh I, et al. The cancer theory of pulmonary arterial hypertension[J]. Pulm Cir, 2017, 7: 285-299. DOI:10.1177/2045893217701438
[8]
Merklinger SL, Jones PL, Martinez EC, et al. Epidermal growth factor receptor blockade mediates smooth muscle cell apoptosis and improves survival in rats with pulmonary hypertension[J]. Circulation, 2005, 112: 423-431. DOI:10.1161/CIRCULATIONAHA.105.540542
[9]
Dahal BK, Cornitescu T, Tretyn A, et al. Role of epidermal growth factor inhibition in experimental pulmonary hypertension[J]. Am Respir Crit Care Med, 2010, 181: 158-167. DOI:10.1164/rccm.200811-1682OC
[10]
Izikki M, Guignabert C, Fadel E, et al. Endothelial-derived FGF2 contributes to the progression of pulmonary hypertension in humans and rodents[J]. J Clin Invest, 2009, 119: 512-523. DOI:10.1172/JCI35070
[11]
Goncharova EA, Ammit AJ, Irani C, et al. PI3K is required for proliferation and migration of human pulmonary vascular smooth muscle cells[J]. Am J Physio Lung Cell Mol Physiol, 2002, 283: L354-L363. DOI:10.1152/ajplung.00010.2002
[12]
Garat CV, Crossno JT, Sullivan TM, et al. Inhibition of phosphatidylinositol 3-kinase/Akt signaling attenuates hypoxia-induced pulmonary artery remodeling and suppresses CREB depletion in arterial smooth muscle cells[J]. J Cardiovasc Pharmacol, 2013, 62: 539-548. DOI:10.1097/FJC.0000000000000014
[13]
Seyfarth HJ, Hammerschmidt S, Halank M, et al. Everolimus in patients with severe pulmonary hypertension: a safety and efficacy pilot trial[J]. Pulm Circ, 2013, 3: 632-638. DOI:10.1086/674311
[14]
Zhang Y, Xie X, Zhu Y, et al. Inhibition of Notch3 prevents monocrotaline-induced pulmonary arterial hypertension[J]. Exp Lung Res, 2015, 41: 435-443. DOI:10.3109/01902148.2015.1060545
[15]
Savai R, Al-Tamari HM, Sedding D, et al. Pro-proliferative and inflammatory signaling converge on FoxO1 transcription factor in pulmonary hypertension[J]. Nat Med, 2014, 20: 1289-1300. DOI:10.1038/nm.3695
[16]
Abud EM, Maylor J, Undem C, et al. Digoxin inhibits development of hypoxic pulmonary hypertension in mice[J]. Proc Natl Acad Sci U S A, 2012, 109: 1239-1244. DOI:10.1073/pnas.1120385109
[17]
Hu CJ, Poth JM, Zhang H, et al. Suppression of HIF2 signalling attenuates the initiation of hypoxia-induced pulmonary hypertension[J]. Eur Respir J, 2019, 54: 1900378.
[18]
Dai Z, Zhu MM, Peng Y, et al. Therapeutic targeting of vascular remodeling and right heart failure in pulmonary arterial hypertension with a HIF-2α inhibitor[J]. Am J Respir Crit Care Med, 2018, 198: 1423-1434. DOI:10.1164/rccm.201710-2079OC
[19]
Thenappan T, Ormiston ML, Ryan JJ, et al. Pulmonary arterial hypertension: pathogenesis and clinical management[J]. BMJ, 2018, 360: j5492.
[20]
Zhang H, Wang D, Li M, et al. Metabolic and proliferative state of vascular adventitial fibroblasts in pulmonary hypertension is regulated through a microRNA-124/ptbp1 (polypyrimidine tract binding protein 1) /pyruvate kinase muscle axis[J]. Circulation, 2017, 136: 2468-2485. DOI:10.1161/CIRCULATIONAHA.117.028069
[21]
Weiss A, Neubauer MC, Yerabolu D, et al. Targeting cyclin-dependent kinases for the treatment of pulmonary arterial hypertension[J]. Nat Commun, 2019, 10: 2204. DOI:10.1038/s41467-019-10135-x
[22]
Thompson AAR, Lawrie A. Targeting vascular remodeling to treat pulmonary arterial hypertension[J]. Trends Mol Med, 2017, 23: 31-45. DOI:10.1016/j.molmed.2016.11.005
[23]
Pullamsetti SS, Savai R, Seeger W, et al. Translational advances in the field of pulmonary hypertension. From cancer biology to new pulmonary arterial hypertension therapeutics. Targeting cell growth and proliferation signaling hubs[J]. Am J Respir Crit Care Med, 2017, 195: 425-437. DOI:10.1164/rccm.201606-1226PP
[24]
Hume S, Dianov GL, Ramadan K. A unified model for the G1/S cell cycle transition[J]. Nucleic Acids Res, 2020, 48: 12483-12501. DOI:10.1093/nar/gkaa1002
[25]
Dick FA, Rubin SM. Molecular mechanisms underlying Rb protein function[J]. Nat Rev Mol Cell Biol, 2013, 14: 297-306. DOI:10.1038/nrm3567
[26]
Goel S, DeCristo MJ, McAllister SS, et al. Cdk4/6 inhibition in cancer: beyond cell cycle arrest[J]. Trends Cell Biol, 2018, 28: 911-925. DOI:10.1016/j.tcb.2018.07.002
[27]
Dyson NJ. Rb1: a prototype tumor suppressor and an enigma[J]. Genes Dev, 2016, 30: 1492-1502. DOI:10.1101/gad.282145.116
[28]
Roskoski R. Cyclin-dependent protein kinase inhibitors including palbociclib as anticancer drugs[J]. Pharmacol Res, 2016, 107: 249-275. DOI:10.1016/j.phrs.2016.03.012
[29]
Malumbres M, Barbacid M. Cell cycle, cdks and cancer: a changing paradigm[J]. Nat Rev Cancer, 2009, 9: 153-166. DOI:10.1038/nrc2602
[30]
Montani D, Chaumais MC, Guignabert C, et al. Targeted therapies in pulmonary arterial hypertension[J]. Pharmacol Ther, 2014, 141: 172-191.
[31]
Nakamura K, Akagi S, Ogawa A, et al. Pro-apoptotic effects of imatinib on PDGF-stimulated pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension[J]. Int J Cardiol, 2012, 159: 100-106. DOI:10.1016/j.ijcard.2011.02.024
[32]
Vantler M, Karikkineth BC, Naito H, et al. PDGF-BB protects cardiomyocytes from apoptosis and improves contractile function of engineered heart tissue[J]. J Mol Cell Cardiol, 2010, 48: 1316-1323. DOI:10.1016/j.yjmcc.2010.03.008
[33]
Schermuly RT, Dony E, Ghofrani HA, et al. Reversal of experimental pulmonary hypertension by PDGF inhibition[J]. J Clin Invest, 2005, 115: 2811-2821. DOI:10.1172/JCI24838
[34]
Hoeper MM, Barst RJ, Bourge RC, et al. Imatinib mesylate as add-on therapy for pulmonary arterial hypertension: results of the randomized impres study[J]. Circulation, 2013, 127: 1128-1138. DOI:10.1161/CIRCULATIONAHA.112.000765
[35]
Maurer B, Reich N, Juengel A, et al. FRA-2 transgenic mice as a novel model of pulmonary hypertension associated with systemic sclerosis[J]. Ann Rheum Dis, 2012, 71: 1382-1387. DOI:10.1136/annrheumdis-2011-200940
[36]
Guignabert C, Phan C, Seferian A, et al. Dasatinib induces lung vascular toxicity and predisposes to pulmonary hypertension[J]. Br J Clin Invest, 2016, 126: 3207-3218. DOI:10.1172/JCI86249
[37]
Özgür Yurttaş N, Eşkazan AE. Dasatinib-induced pulmonary arterial hypertension[J]. Br J Clin Pharmacol, 2018, 84: 835-845. DOI:10.1111/bcp.13508
[38]
Ciuclan L, Bonneau O, Hussey M, et al. A novel murine model of severe pulmonary arterial hypertension[J]. Am J Respir Crit Care Med, 2011, 184: 1171-1182. DOI:10.1164/rccm.201103-0412OC
[39]
Degirmenci U, Wang M, Hu J. Targeting aberrant RAS/RAF/MEK/ERK signaling for cancer therapy[J]. Cells, 2020, 9: 198. DOI:10.3390/cells9010198
[40]
De Luca A, Maiello MR, D'Alessio A, et al. The RAS/RAF/MEK/ERK and the PI3K/AKT signalling pathways: role in cancer pathogenesis and implications for therapeutic approaches[J]. Expert Opin Ther Targets, 2012, 16 Suppl 2: S17-S27.
[41]
Awad KS, Elinoff JM, Wang S, et al. RAF/ERK drives the prolife-rative and invasive phenotype of BMPR2-silenced pulmonary artery endothelial cells[J]. Am J Physiol Lung Cell Mol Physiol, 2016, 310: L187-L201. DOI:10.1152/ajplung.00303.2015
[42]
Lane KB, Blackwell TR, Runo J, et al. Aberrant signal transduction in pulmonary hypertension[J]. Chest, 2005, 128: 564s-565s.
[43]
Hopper RK, Feinstein JA, Manning MA, et al. Neonatal pulmonary arterial hypertension and Noonan syndrome: two fatal cases with a specific RAF1 mutation[J]. Am J Med Genet A, 2015, 167A: 882-885.
[44]
Zhou C, Chen Y, Kang W, et al. miR-455-3p-1 represses FGF7 expression to inhibit pulmonary arterial hypertension through inhibiting the RAS/ERK signaling pathway[J]. J Mol Cell Cardiol, 2019, 130: 23-35. DOI:10.1016/j.yjmcc.2019.03.002
[45]
Moreno-Vinasco L, Gomberg-Maitland M, Maitland ML, et al. Genomic assessment of a multikinase inhibitor, sorafenib, in a rodent model of pulmonary hypertension[J]. Physiol Genomics, 2008, 33: 278-291. DOI:10.1152/physiolgenomics.00169.2007
[46]
Kojonazarov B, Sydykov A, Pullamsetti SS, et al. Effects of multikinase inhibitors on pressure overload-induced right ventricular remodeling[J]. Int J Cardiol, 2013, 167: 2630-2637. DOI:10.1016/j.ijcard.2012.06.129
[47]
Gomberg-Maitland M, Maitland ML, Barst RJ, et al. A dosing/cross-development study of the multikinase inhibitor sorafenib in patients with pulmonary arterial hypertension[J]. Clin Pharmacol Ther, 2010, 87: 303-310. DOI:10.1038/clpt.2009.217
[48]
Kimura G, Kataoka M, Inami T, et al. Sorafenib as a potential strategy for refractory pulmonary arterial hypertension[J]. Pulm Pharmacol Ther, 2017, 44: 46-49. DOI:10.1016/j.pupt.2017.03.009
[49]
Dienstmann R, Rodon J, Serra V, et al. Picking the point of inhibition: a comparative review of PI3K/AKT/MTOR pathway inhibitors[J]. Mol Cancer Ther, 2014, 13: 1021-1031. DOI:10.1158/1535-7163.MCT-13-0639
[50]
Bilanges B, Posor Y, Vanhaesebroeck B. PI3K isoforms in cell signalling and vesicle trafficking[J]. Nat Rev Mol Cell Biol, 2019, 20: 515-534. DOI:10.1038/s41580-019-0129-z
[51]
Houssaini A, Abid S, Mouraret N, et al. Rapamycin reverses pulmonary artery smooth muscle cell proliferation in pulmonary hypertension[J]. Am J Respir Cell Mol Biol, 2013, 48: 568-577. DOI:10.1165/rcmb.2012-0429OC
[52]
Guo Y, Liu X, Zhang Y, et al. 3-Bromopyruvate ameliorates pulmonary arterial hypertension by improving mitochondrial metabolism[J]. Life Sci, 2020, 256: 118009. DOI:10.1016/j.lfs.2020.118009
[53]
Liu P, Gu Y, Luo J, et al. Inhibition of src activation reverses pulmonary vascular remodeling in experimental pulmonary arterial hypertension via AKT/MTOR/HIF-1 signaling pathway[J]. Exp Cell Res, 2019, 380: 36-46. DOI:10.1016/j.yexcr.2019.02.022
[54]
Ogawa A, Firth AL, Yao W, et al. Inhibition of mTor attenuates store-operated Ca2+ entry in cells from endarterectomized tissues of patients with chronic thromboembolic pulmonary hypertension[J]. Am J Physiol Lung Cell Mol Physiol, 2009, 297: L666-L676. DOI:10.1152/ajplung.90548.2008
[55]
Wang AP, Li XH, Yang YM, et al. A critical role of the mTor/eif2α pathway in hypoxia-induced pulmonary hypertension[J]. PLoS One, 2015, 10: e0130806. DOI:10.1371/journal.pone.0130806
[56]
Yang Y, Yin L, Zhu M, et al. Protective effects of dioscin on vascular remodeling in pulmonary arterial hypertension via adjusting GRB2/ERK/PI3K-AKT signal[J]. Biomed Pharmacother, 2021, 133: 111056. DOI:10.1016/j.biopha.2020.111056
[57]
Li J, Kim SG, Blenis J. Rapamycin: one drug, many effects[J]. Cell Metab, 2014, 19: 373-379. DOI:10.1016/j.cmet.2014.01.001
[58]
Nishimura T, Faul JL, Berry GJ, et al. 40-O- (2-Hydroxyethyl)-rapamycin attenuates pulmonary arterial hypertension and neointimal formation in rats[J]. Am J Respir Crit Care Med, 2001, 163: 498-502. DOI:10.1164/ajrccm.163.2.2006093
[59]
Paddenberg R, Stieger P, von Lilien AL, et al. Rapamycin attenuates hypoxia-induced pulmonary vascular remodeling and right ventricular hypertrophy in mice[J]. Respir Res, 2007, 8: 15. DOI:10.1186/1465-9921-8-15
[60]
Goncharov DA, Kudryashova TV, Ziai H, et al. Mammalian target of rapamycin complex 2 (mTORC2) coordinates pulmonary artery smooth muscle cell metabolism, proliferation, and survival in pulmonary arterial hypertension[J]. Circulation, 2014, 129: 864-874. DOI:10.1161/CIRCULATIONAHA.113.004581
[61]
Artavanis-Tsakonas S, Rand MD, Lake RJ. Notch signaling: cell fate control and signal integration in development[J]. Science, 1999, 284: 770-776. DOI:10.1126/science.284.5415.770
[62]
Li MX, Gu SL, Ying MF, et al. Advances in research on Notch signaling pathway in pulmonary vascular remodeling of pulmonary arterial hypertension[J]. Chin Pharmacol Bull (中国药理学通报), 2020, 36: 5-8.
[63]
Kopan R, Ilagan MX. The canonical notch signaling pathway: unfolding the activation mechanism[J]. Cell, 2009, 137: 216-233. DOI:10.1016/j.cell.2009.03.045
[64]
Choy L, Hagenbeek TJ, Solon M, et al. Constitutive Notch3 signaling promotes the growth of basal breast cancers[J]. Cancer Res, 2017, 77: 1439-1452. DOI:10.1158/0008-5472.CAN-16-1022
[65]
Gu ZY, Wang L, Gao CJ. Abnormal Notch-Hes signaling pathways and acute leukemia[J]. J Exp Hematol (中国实验血液学杂志), 2017, 25: 240-243.
[66]
Baeten JT, Lilly B. Notch signaling in vascular smooth muscle cells[J]. Adv Pharmacol, 2017, 78: 351-382.
[67]
Song Y, Zhang Y, Jiang H, et al. Activation of Notch3 promotes pulmonary arterial smooth muscle cells proliferation via Hes1/p27KIP1 signaling pathway[J]. FEBS Open Bio, 2015, 5: 656-660. DOI:10.1016/j.fob.2015.08.007
[68]
Li XD, Zhang X, Leathers R, et al. Notch3 signaling promotes the development of pulmonary arterial hypertension[J]. Nat Med, 2009, 15: 1289-1297. DOI:10.1038/nm.2021
[69]
Chen X, Zhou W, Hu Q, et al. Exploration of the Notch3-Hes5 signal pathway in monocrotaline-induced pulmonary hypertension using rat model[J]. Congenit Heart Dis, 2019, 14: 396-402. DOI:10.1111/chd.12733
[70]
Wang W, Liu J, Ma A, et al. Mtorc1 is involved in hypoxia-induced pulmonary hypertension through the activation of Notch3[J]. J Cell Physiol, 2014, 229: 2117-2125. DOI:10.1002/jcp.24670
[71]
Lai YJ, Chang GJ, Yeh YH, et al. Propylthiouracil attenuates experimental pulmonary hypertension via suppression of pen-2, a key component of gamma-secretase[J]. PLoS One, 2015, 10: e0137426. DOI:10.1371/journal.pone.0137426
[72]
Takebe N, Nguyen D, Yang SX. Targeting Notch signaling pathway in cancer: clinical development advances and challenges[J]. Pharmacol Ther, 2014, 141: 140-149. DOI:10.1016/j.pharmthera.2013.09.005
[73]
Ghahhari NM, Babashah S. Interplay between micrornas and Wnt/β-catenin signalling pathway regulates epithelial-mesenchymal transition in cancer[J]. Eur J Cancer, 2015, 51: 1638-1649. DOI:10.1016/j.ejca.2015.04.021
[74]
Pataki CA, Couchman JR, Brábek J. Wnt signaling cascades and the roles of syndecan proteoglycans[J]. J Histochem Cytochem, 2015, 63: 465-480. DOI:10.1369/0022155415586961
[75]
De A. Wnt/Ca2+ signaling pathway: a brief overview[J]. Acta Biochim Biophys Sin, 2011, 43: 745-756. DOI:10.1093/abbs/gmr079
[76]
Kahn M. Can we safely target the wnt pathway?[J]. Nat Rev Drug Discov, 2014, 13: 513-532. DOI:10.1038/nrd4233
[77]
Chiurillo MA. Role of the Wnt/β-catenin pathway in gastric cancer: an in-depth literature review[J]. World J Exp Med, 2015, 5: 84-102. DOI:10.5493/wjem.v5.i2.84
[78]
de Jesus Perez V, Yuan K, Alastalo TP, et al. Targeting the Wnt signaling pathways in pulmonary arterial hypertension[J]. Drug Discov Today, 2014, 19: 1270-1276. DOI:10.1016/j.drudis.2014.06.014
[79]
Quasnichka H, Slater SC, Beeching CA, et al. Regulation of smooth muscle cell proliferation by beta-catenin/t-cell factor signaling involves modulation of cyclin D1 and p21 expression[J]. Circ Res, 2006, 99: 1329-1337. DOI:10.1161/01.RES.0000253533.65446.33
[80]
Sklepkiewicz P, Schermuly RT, Tian X, et al. Glycogen synthase kinase 3beta contributes to proliferation of arterial smooth muscle cells in pulmonary hypertension[J]. PLoS One, 2011, 6: e18883. DOI:10.1371/journal.pone.0018883
[81]
Yu XM, Wang L, Li JF, et al. Wnt5a inhibits hypoxia-induced pulmonary arterial smooth muscle cell proliferation by downregulation of β-catenin[J]. Am J Physiol Lung Cell Mol Physiol, 2013, 304: L103-L111. DOI:10.1152/ajplung.00070.2012
[82]
Yuan K, Shamskhou EA, Orcholski ME, et al. Loss of endothelium-derived Wnt5a is associated with reduced pericyte recruitment and small vessel loss in pulmonary arterial hypertension[J]. Circulation, 2019, 139: 1710-1724. DOI:10.1161/CIRCULATIONAHA.118.037642
[83]
Zhou J, Wang JX. Advances in the role and mechanism of FOXO in promoting tumor genesis and development[J]. Oncol Prog (癌症进展), 2020, 18: 343-346.
[84]
Laissue P. The forkhead-box family of transcription factors: key molecular players in colorectal cancer pathogenesis[J]. Mol Cancer, 2019, 18: 5. DOI:10.1186/s12943-019-0938-x
[85]
Weng XK, Sun F, Yu JP. Recent advances of FOXO in cancer[J]. Oncol Prog (癌症进展), 2021, 19: 541-544.
[86]
O'Regan RM, Nahta R. Targeting forkhead box M1 transcription factor in breast cancer[J]. Biochem Pharmacol, 2018, 154: 407-413. DOI:10.1016/j.bcp.2018.05.019
[87]
Raghavan A, Zhou G, Zhou Q, et al. Hypoxia-induced pulmonary arterial smooth muscle cell proliferation is controlled by forkhead box M1[J]. Am J Respir Cell Mol Biol, 2012, 46: 431-436. DOI:10.1165/rcmb.2011-0128OC
[88]
Dai J, Zhou Q, Tang H, et al. Smooth muscle cell-specific foxm1 controls hypoxia-induced pulmonary hypertension[J]. Cell Signal, 2018, 51: 119-129. DOI:10.1016/j.cellsig.2018.08.003
[89]
Bourgeois A, Lambert C, Habbout K, et al. FOXM1 promotes pulmonary artery smooth muscle cell expansion in pulmonary arterial hypertension[J]. J Mol Med (Berl), 2018, 96: 223-235. DOI:10.1007/s00109-017-1619-0
[90]
Dai Z, Zhu MM, Peng Y, et al. Endothelial and smooth muscle cell interaction via FOXM1 signaling mediates vascular remodeling and pulmonary hypertension[J]. Am J Respir Crit Care Med, 2018, 198: 788-802. DOI:10.1164/rccm.201709-1835OC
[91]
Gu L, Liu HM. Forkhead box M1 transcription factor: a novel target for pulmonary arterial hypertension therapy[J]. World J Pediatr, 2020, 16: 113-119. DOI:10.1007/s12519-019-00271-1
[92]
Calvier L, Boucher P, Herz J, et al. LRP1 deficiency in vascular SMC leads to pulmonary arterial hypertension that is reversed by PPARγ activation[J]. Circ Res, 2019, 124: 1778-1785. DOI:10.1161/CIRCRESAHA.119.315088
[93]
Tseng V, Sutliff RL, Hart CM. Redox biology of peroxisome proliferator-activated receptor-γ in pulmonary hypertension[J]. Antioxid Redox Signal, 2019, 31: 874-897. DOI:10.1089/ars.2018.7695
[94]
Prins KW, Thenappan T, Weir EK, et al. Repurposing medications for treatment of pulmonary arterial hypertension: what's old is new again[J]. J Am Heart Assoc, 2019, 8: e011343. DOI:10.1161/JAHA.118.011343
[95]
Rashid J, Nozik-Grayck E, McMurtry IF, et al. Inhaled combination of sildenafil and rosiglitazone improves pulmonary hemodynamics, cardiac function, and arterial remodeling[J]. Am J Physiol Lung Cell Mol Physiol, 2019, 316: L119-L130. DOI:10.1152/ajplung.00381.2018
[96]
Jiang F, Xing X, Wang X, et al. Drugs and drug delivery stra-tegies for treatment of pulmonary arterial hypertension[J]. Acta Pharm Sin (药学学报), 2021, 56: 1332-1342.
[97]
Heidbreder M, Fröhlich F, Jöhren O, et al. Hypoxia rapidly activates HIF-3alpha mRNA expression[J]. FASEB J, 2003, 17: 1541-1543.
[98]
Clerici C, Planès C. Gene regulation in the adaptive process to hypoxia in lung epithelial cells[J]. Am J Physiol Lung Cell Mol Physiol, 2009, 296: L267-L274. DOI:10.1152/ajplung.90528.2008
[99]
Pullamsetti SS, Mamazhakypov A, Weissmann N, et al. Hypoxia-inducible factor signaling in pulmonary hypertension[J]. J Clin Invest, 2020, 130: 5638-5651. DOI:10.1172/JCI137558
[100]
Semenza GL. Oxygen sensing, hypoxia-inducible factors, and disease pathophysiology[J]. Annu Rev Pathol, 2014, 9: 47-71. DOI:10.1146/annurev-pathol-012513-104720
[101]
Veith C, Schermuly RT, Brandes RP, et al. Molecular mechanisms of hypoxia-inducible factor-induced pulmonary arterial smooth muscle cell alterations in pulmonary hypertension[J]. J Physiol, 2016, 594: 1167-1177. DOI:10.1113/JP270689
[102]
Brusselmans K, Compernolle V, Tjwa M, et al. Heterozygous deficiency of hypoxia-inducible factor-2alpha protects mice against pulmonary hypertension and right ventricular dysfunction during prolonged hypoxia[J]. J Clin Invest, 2003, 111: 1519-1527. DOI:10.1172/JCI15496
[103]
Shimoda LA, Manalo DJ, Sham JS, et al. Partial HIF-1alpha deficiency impairs pulmonary arterial myocyte electrophysio-logical responses to hypoxia[J]. Am J Physiol Lung Cell Mol Physiol, 2001, 281: L202-L208. DOI:10.1152/ajplung.2001.281.1.L202
[104]
Yu AY, Shimoda LA, Iyer NV, et al. Impaired physiological responses to chronic hypoxia in mice partially deficient for hypoxia-inducible factor 1alpha[J]. J Clin Invest, 1999, 103: 691-696. DOI:10.1172/JCI5912
[105]
Ball MK, Waypa GB, Mungai PT, et al. Regulation of hypoxia-induced pulmonary hypertension by vascular smooth muscle hypoxia-inducible factor-1α[J]. Am J Respir Crit Care Med, 2014, 189: 314-324. DOI:10.1164/rccm.201302-0302OC
[106]
Malczyk M, Veith C, Fuchs B, et al. Classical transient receptor potential channel 1 in hypoxia-induced pulmonary hypertension[J]. Am J Respir Crit Care Med, 2013, 188: 1451-1459. DOI:10.1164/rccm.201307-1252OC
[107]
Veith C, Zakrzewicz D, Dahal BK, et al. Hypoxia- or PDGF-BB-dependent paxillin tyrosine phosphorylation in pulmonary hypertension is reversed by HIF-1α depletion or imatinib treatment[J]. Thromb Haemost, 2014, 112: 1288-1303. DOI:10.1160/th13-12-1031
[108]
Chen W, Hill H, Christie A, et al. Targeting renal cell carcinoma with a HIF-2 antagonist[J]. Nature, 2016, 539: 112-117. DOI:10.1038/nature19796
[109]
Cowburn AS, Crosby A, Macias D, et al. HIF2α-arginase axis is essential for the development of pulmonary hypertension[J]. Proc Natl Acad Scie U S A, 2016, 113: 8801-8806.
[110]
Dai Z, Li M, Wharton J, et al. Prolyl-4 hydroxylase 2 (PHD2) deficiency in endothelial cells and hematopoietic cells induces obliterative vascular remodeling and severe pulmonary arterial hypertension in mice and humans through hypoxia-inducible factor-2α[J]. Circulation, 2016, 133: 2447-2458.
[111]
Tang H, Babicheva A, McDermott KM, et al. Endothelial HIF-2α contributes to severe pulmonary hypertension due to endothelial-to-mesenchymal transition[J]. Am J Physiol Lung Cell Mol Physiol, 2018, 314: L256-L275.
[112]
Zhang HF, Qian DZ, Tan YS, et al. Digoxin and other cardiac glycosides inhibit HIF-1 synthesis and block tumor growth[J]. Proc Natl Acad Sci U S A, 2008, 105: 19579-19586.
[113]
Ji L, Li ZQ, Lu DX. Recent advances of HIF-2α in pulmonary hypertension[J]. Acta Pharm Sin (药学学报), 2022, 57: 277-286.
[114]
Farber HW, Miller DP, Poms AD, et al. Five-year outcomes of patients enrolled in the reveal registry[J]. Chest, 2015, 148: 1043-1054.
[115]
Lythgoe MP, Rhodes CJ, Ghataorhe P, et al. Why drugs fail in clinical trials in pulmonary arterial hypertension, and strategies to succeed in the future[J]. Pharmacol Ther, 2016, 164: 195-203.