The impact of Vitamin D supplementation on immune responses to respiratory pathogens in older children in Zambia with perinatally-acquired HIV-1

该项目的目的是评估补充维生素D是否能改善撒哈拉以南非洲艾滋病毒感染儿童（CWH）对呼吸道病原体的免疫反应。这项研究将被纳入赞比亚正在进行的一项更大规模的维生素D干预试验。撒哈拉以南非洲儿童受到全球艾滋病毒流行病的严重影响。在全球200万艾滋病毒感染儿童中，90%居住在撒哈拉以南非洲。尽管抗逆转录病毒疗法（ART）的推出，艾滋病毒及其相关的合并症仍然是撒哈拉以南非洲儿童死亡的主要原因之一。虽然抗逆转录病毒疗法大大提高了这一群体进入青春期的存活率，但这些人继续患有慢性合并症，其中主要是严重和复发性呼吸道感染。据认为，造成这一问题的主要原因是维生素D缺乏。撒哈拉以南非洲的CWH往往维生素D水平很低，这对健康有多效性影响，对免疫功能有重大不利影响。此外，在其他环境中，维生素D缺乏已被确定为呼吸道感染不良结局的公认因素。因此，有人假设，在CWH中补充维生素D（与ART一起）将具有广泛的健康益处，包括降低对呼吸道感染的易感性。这一假设正在EDCTP资助的VITALITY试验中进行测试，这是一项正在进行的大型研究，旨在对赞比亚和津巴布韦围产期获得HIV-1的青少年每周补充高剂量维生素D3。拟议的项目将在VITALITY内形成一个子研究，测试维生素D3补充剂改善对呼吸道病原体的免疫反应的假设。为此，将测量以下参数：1.可溶性和细胞免疫标志物VITALITY试验的样本将同时评价细胞和可溶性免疫标志物。T细胞免疫检测将包括T细胞亚群（如TH 1、TH 2、TH 17和T调节细胞）的流式细胞术评估，以及CD 8+和CD 4 + T细胞活化变化的评价。将使用多重方法研究可溶性标志物，以鉴定患者样本中的促炎标志物。将进行体外刺激试验，以检测病原体对细菌病原体（如肺炎链球菌和结核分枝杆菌）和病毒病原体（如甲型流感病毒、RSV和选定病例中的SARS-CoV-2）的特异性反应。2.维生素D途径中的等位基因变异体已经表明，维生素D途径中的等位基因变异体与呼吸道感染风险增加有关。因此，将按基因型对患者样本（n=420）进行追溯分层，以确定对治疗的反应是否根据维生素D相关基因的变异而变化。这些信息将进一步用于确定基因型对维生素D治疗临床反应的影响。这项工作的结果将证明维生素D补充剂是否能改善这一高危人群对呼吸道疾病的保护性免疫反应。可能的机制将被阐明，从而更好地了解介质，如维生素D调节免疫力。鉴于持续的呼吸道流行病和新型呼吸道病毒大流行病（如COVID-19），本研究的结果可以通过提供廉价且易于扩展的治疗来解决未满足的临床需求，以改善CWH的健康结果。通过这个项目，学生将发展一系列技能，包括实验室技能，统计技能和相关的计算技能，转化研究的经验，以及在赞比亚卢萨卡的大型临床试验现场工作的经验。

The purpose of this project is to assess whether vitamin D supplementation improves immune responses to respiratory pathogens in children living with HIV (CWH) in Sub-Saharan Africa. This study will be embedded in a larger ongoing vitamin D intervention trial in Zambia.Sub-Saharan African children are disproportionately affected by the global HIV epidemic. Of the two million children living with HIV globally, 90% reside in Sub-Saharan Africa. Despite the roll-out of antiretroviral therapy (ART), HIV and its associated comorbidities remains one of the leading causes of death among children in Sub-Saharan Africa. While ART has greatly improved this cohort's survival into adolescence, these individuals continue to suffer from chronic comorbidities, principal among which are severe and recurrent respiratory infections. It is believed that a major contributor to this problem is vitamin D deficiency. CWH in Sub-Saharan Africa tend to have very low levels of vitamin D, and this has pleiotropic effects on health with major adverse effects on immune function. Additionally, in other settings, vitamin D deficiency has been established as a well-recognised contributor to poor outcomes in respiratory infections. It has therefore been hypothesised that vitamin D supplementation in CWH (alongside ART) will have wide-ranging health benefits, including a reduction in susceptibility to respiratory infection. This hypothesis is being tested in the EDCTP-funded VITALITY trial, a large ongoing study of weekly high-dose vitamin D3 supplementation in adolescents with perinatally acquired HIV-1 in Zambia and Zimbabwe. The proposed project will form a sub-study within VITALITY, testing the hypothesis that vitamin D3 supplementation improves immune responses to respiratory pathogens. To do this, the following parameters will be measured:1. Soluble and Cellular Markers of ImmunitySamples from the VITALITY trial will be evaluated for both cellular and soluble markers of immunity. Tests of T-cell immunity will include flow cytometric assessment of T-cell subsets such as TH1, TH2, TH17 and T-Regulatory cells, and evaluation of changes in CD8+ and CD4+ T-Cell activation. Soluble markers will be investigated using a multiplexed approach to identify proinflammatory markers in patient samples. In vitro stimulation assays will be conducted to test pathogen-specific responses to bacterial pathogens such as Streptococcus pneumoniae and Mycobacterium tuberculosis, and viral pathogens such as Influenza A, RSV and in selected cases SARS-CoV-2. 2. Allelic Variants in the Vitamin D PathwayIt has been shown that allelic variants in the vitamin D pathway are associated with increased risk to respiratory infection. As a result, patient samples (n=420) will be retroactively stratified by genotype to determine if response to treatment varies according to variants at vitamin D associated genes. This information will furthermore be used to determine the impact of genotype on clinical response to vitamin D therapy. The findings of this work will demonstrate whether vitamin D supplementation improves protective immune responses to respiratory diseases in this at-risk population. Likely mechanisms will be elucidated leading to a better understanding of how mediators such as vitamin D modulate immunity. Given ongoing respiratory epidemics, and novel respiratory virus pandemics such as COVID-19, the findings of this study could address an unmet clinical need by providing an inexpensive and easily scalable treatment to improve health outcomes for CWH. Through this project the student will develop a range of skills including lab-based skills, statistical skills and associated computational skills, experience in translational research, and the experience of working on-site at a large clinical trial in Lusaka, Zambia.