THE ROLE OF PERIVASCULAR CELL (PERICYTE) DYSFUNCTION IN HEPATIC COMPLICATIONS OF SEVERE DENGUE

世界卫生组织将登革热列为2019年全球公共卫生面临的十大威胁之一，与气候变化和抗菌素耐药性不相上下，也是名单上仅有的三种传染病之一。世界上一半的人口面临感染登革热病毒的风险，其中大部分在低收入和中等收入国家。登革热通常表现为发热综合征，但在一些患者中，疾病发展到严重阶段，表现为广泛和急性的血液和液体从血管泄漏，可导致全身休克和死亡。由于目前的疫苗不适合在所有环境下使用，没有特定的抗病毒药物，也没有预后标记物来识别将发展为严重疾病的患者，登革热给受影响的国家和个人带来了巨大的健康、经济和社会负担。严重登革热患者的肝脏经常受损，导致腹部积液和局部出血；实际上，肝功能障碍的生物标志物与登革热疾病的严重程度和预后不良相关。严重登革热疾病影响肝脏的方式，特别是其血管系统，在机制层面上尚未完全了解，并可能提供重要线索，以识别患有严重疾病的高风险患者，并提供开发药物以改善患者预后的新方法。该项目将重点研究受登革热出血影响的血管的生物学，以了解严重登革热患者肝脏血管功能障碍的发展。肝血管由两种细胞类型组成，血管内表面的内皮细胞和包裹内皮管的血管周围细胞(肝星状细胞)。周细胞的功能是密切调节内皮细胞的功能，调节血管的完整性。我们自己的研究首次表明，一种名为“NS1”的病毒蛋白的存在，这种蛋白在严重登革热患者的血液中浓度很高，阻止了周细胞执行这一调节功能。事实上，我们的实验表明，NS1蛋白破坏了内皮细胞和周细胞之间的相互作用，使内皮血管裸露和渗漏，因为它们缺乏周细胞。在这项拨款中，我们建议研究登革病毒可能导致肝脏星状细胞功能障碍从而引发血管渗漏的三种不同方式：(1)患者血液中存在的NS1蛋白的影响，(2)登革病毒直接感染肝星状细胞，以及(3)受感染的肝细胞(肝细胞)释放炎性物质。在研究这些机制时，我们将重点研究它们如何通过导致肝星状细胞分化为不同类型的细胞来改变其身份，以及它们如何改变肝星状细胞与血管的关系，影响血管屏障的完整性。此外，我们将调查所有四种变种登革热病毒(血清型)的行为，因为据报道，严重症状的表现因每次爆发的主要变种而不同。

The World Health Organisation listed dengue as one of its top ten threats to global public health in 2019, comparable to climate change and antimicrobial resistance, and one of only three infectious diseases on the list. Half of the world's population is at risk of dengue virus infection, mostly in low- and middle-income countries. Dengue disease normally manifest as a febrile syndrome, but in some patients the disease progresses into a severe phase presenting with generalised and acute blood and fluid leakage from blood vessels that can lead to systemic shock and death. With the current vaccine unsuitable for use in all settings, no specific antiviral drugs, and no prognostic markers to identify patients that will progress to severe disease, dengue poses an enormous health, economic and social burden on the affected countries and individuals.The liver is often compromised in severe dengue patients, leading to fluid accumulation in the abdomen and local haemorrhage; indeed, biomarkers of liver dysfunction correlate with dengue disease severity and a poor prognostic outcome. The ways in which severe dengue disease affects the liver, and in particular its vasculature, are not entirely understood at a mechanistic level and might provide important clues to identify patients at high risk of developing severe disease, and new ways of developing drugs to improve patient outcomes. This project will focus on understanding the development of blood vessel malfunction in the liver of severe dengue patients by focusing on the biology of the blood vessels affected by dengue haemorrhage. Liver blood vessels are formed of two cell types, endothelial cells lining the inner surface of the vessel and perivascular cells (hepatic stellate cells) enveloping the endothelial tubes. The function of pericytes is to closely regulate the function of endothelial cells and modulate the integrity of blood vessel. Our own research has shown for the first time that the presence of a viral protein called 'NS1', which is present in high concentration in the blood of severe dengue patients, prevents pericytes from performing this regulatory function. Indeed, our experiments showed that the NS1 protein disrupts the interaction between endothelial cells and pericytes, leaving the endothelial vessels naked and leaky because they lack pericytes. In this grant, we propose to study three separate ways in which dengue virus might be causing the dysfunction of liver hepatic stellate cells, and therefore provoking vessel leakage: (1) the impact of the NS1 protein present in patient blood, (2) direct infection of hepatic stellate cells by dengue virus, and (3) the release of inflammatory substances by infected liver cells (hepatocytes). While investigating these mechanisms, we will focus on how they change the identity of the hepatic stellate cells by causing their differentiation into different cell type and how they modify the hepatic stellate cells' relationship with blood vessels, affecting the integrity of the blood vessel barrier. Furthermore, we will investigate the behaviour of all four variants dengue viruses (serotypes) as the presentation of severe symptoms has been reported to differ depending on the predominant variant in each outbreak.