Background
Disease progression in the absence of therapy varies significantly in HIV-infected individuals. Most patients experience progressive CD4
+ T cell loss and develop AIDS [
1]. However, a small proportion of HIV-infected patients remain clinically and/or immunologically stable for years, including long-term nonprogressors (LTNPs) who maintain normal CD4
+ T cell counts for prolonged periods (>10 years) and elite controllers (ECs) who have undetectable viremia (<50 copies/ml) [
2‐
4]. In past decades, numerous studies have been conducted searching for the cause for this lack of progression [
5‐
7]. Among them, high-throughput techniques, such as microarray analysis, have contributed to the understanding of the complex host-virus interactions associated with the delayed disease progression in nonprogressors [
8‐
12].
Using various microarray techniques and various types of samples, including whole blood, PBMC, CD4
+ T cells, CD8
+ T cells and monocytes, previous studies have provided valuable information on transcriptomic profiles in nonprogressors (including LTNPs and ECs, referred to as “nonprogressors” below). Several transcriptomic analyses of T cells have highlighted the role of reduced interferon-stimulated genes (ISGs) associated with the nonprogressing status in LTNPs and ECs [
8,
10,
13]. T cell transcriptomic studies have also revealed the enhancing pathways in LTNPs, including the APK, WNT, and AKT pathways, contributing to cell survival and antiviral responses [
14], cytokine–cytokine receptor interaction, a negative control of apoptosis or regulation of actin cytoskeleton [
15]. A PBMC transcriptomic study demonstrated that cell death/proapoptotic genes were mostly downregulated and cell survival/antiapoptotic genes or genes belonging to the canonical Wnt/beta-catenin signaling pathway were upregulated in LTNPs [
16]. A monocyte transcriptomic study revealed upregulation of Toll-like receptor (TLR) signaling with subsequent downregulation of MAPK, NF-kB, JAK-STAT, and the IRF cascades in LTNPs [
17]. A transcriptomic study on whole blood from LTNPs and progressors identified a novel ISG gene, LY6E, which restrains the hyperactivation of monocytes during HIV-1 infection [
11].
Although the accumulating transcriptome data provide useful information in studying the host protective immune responses in nonprogressors, the identification of key genes and pathways from these studies was restricted due to the limited sample size in the independent study. A transcriptome meta-analysis can incorporate high-throughput data from multiple independent studies and overcome the aforementioned limitations. In addition, different cells have unique functional activities in the immune responses of nonprogressors, but the differences in the transcriptomic profiles between the major cell types of nonprogressors have not been elucidated. In the present study, we performed a meta-analysis of five independent microarray datasets to identify differentially expressed genes (DEGs) in whole blood, CD4+ and CD8+ T cells from HIV-infected nonprogressors compared with progressors. Our study provides a comprehensive evaluation of the gene expression profiles in the major blood types of nonprogressors, which will provide new insights in the understanding of HIV pathogenesis and developing strategies to delay HIV-1 disease progression.
Discussion
Identification of the most relevant genes and pathways involved in HIV-infected nonprogressors is important in understanding the molecular and cellular processes determining the cause for the nonprogression status in these patients. In this study, we performed a meta-analysis of multiple public microarray datasets to investigate the transcriptomic profiles of HIV-infected nonprogressors, including LTNPs and ECs. Through transcriptomic meta-analysis, we identified DEGs in various types of samples in the comparison of nonprogressors and progressors, including whole blood, CD4+ and CD8+ T cells.
We found that the expression of multiple important ISG genes, CD38 and LAG-3 was significantly downregulated in HIV-infected nonprogressors compared with that of progressors in all the three sample types. The downregulation of important ISG genes, including LY6E, IFI27, ISG15 and IFI44L, was observed in our study. Type-1 interferons (IFNs) is of critical importance for its potent antiviral effects [
29]. However, it is well established that type-1 IFNs exert their full antiviral effect at very low concentration and their expression is required locally [
30,
31]. Emerging lines of evidence reveal that high level and sustained type-1 IFNs expression is associated with hyper-immune activation and disease progression in persistent infections [
31‐
34]. Dynamics of type-1 IFNs distinguishes Simian immunodeficiency virus (SIV) infection of natural hosts, that do not develop AIDS, from pathogenic SIV infections [
35‐
38]. Natural hosts rapidly mute their type-1 IFNs responses after acute SIV infection whereas disease-susceptible macaque species maintain type-1 IFNs signaling indefinitely, which triggers hyper-activation of immune system and contributes to provide an environment that favors progression to AIDS [
29,
36‐
39]. As the primary producers of type-1 IFNs, plasmacytoid dendritic cells (pDCs) in natural hosts of SIV have attenuated recruitment to lymphoid tissues compared with non-natural host [
39‐
41]. This reduces pDCs exposure to sites of high level virus replication, and limits the pathogenic T-cell activation and chronic inflammation driven by activated pDCs [
39,
41,
42]. In HIV infected human, although pDCs are depleted in blood during chronic infection, they accumulate in lymph nodes [
39,
41‐
43], which may directly contribute to triggering sustain pathogenic immune activation [
41‐
45]. High levels of IFN-α in the serum of chronically HIV-infected patients, as well as in tonsils during both chronic and acute HIV infection were observed [
46,
47], which positively correlated with markers of immune activation [
48,
49]. The expression of ISGs is elevated in HIV-infected rapid progressors rather than in nonprogressors [
9‐
11,
13,
14]. Our results indicated that reduced expression of ISGs, which leads to reduced activation of immune system, was a key factor affecting disease progression. This was confirmed by our further finding that CD38 was downregulated in HIV-infected nonprogressors. CD38 is an activation marker and a nicotinamide adenine dinucleotide consuming enzyme (NADases) [
15,
50]. Previous studies showed that the elevated expression of CD38 has a strong relationship with activation and cell aging [
15,
50]. Our results showed that nonprogressors have lower expression of CD38 in different types of cells, which was beneficial in maintaining the survival of the cell. In addition, we found a lower expression of LAG-3, an important coinhibitory molecule in the immune system, in HIV-infected nonprogressors. It has been reported that LAG-3 is associated with immune dysfunction/exhaustion of T cells [
51]. Taken together, our data revealed that HIV-infected nonprogressors have lower expression of ISGs, CD38 and LAG-3, which was beneficial in maintaining a healthy status of the immune system and contributed to the nonprogression of the disease.
Gene ontology analysis and pathway enrichment analysis offered insight into the possible roles of DEGs in the pathogenesis of nonprogressors. Consistent with our data on DEGs, we found that the type I interferon signaling pathway was involved in the most significantly enriched terms shared by the three sample types. Furthermore, we found an upregulated PI3K-Akt signaling pathway in whole blood and MAPK pathway in CD8
+ T cells in nonprogressors. Both pathways contribute to cell survival and a successful immune response depends upon the ability of T lymphocytes to respond to antigenic stimulation [
14,
15,
52]. In addition, the MAPK pathway plays an important role in T cell homeostasis [
53], which is required for the cytotoxic activity of most CD8
+ T cells [
54]. Wu et al. reported that the nonprogressing status in HIV-infected LTNPs was associated with the MAPK, WNT, and AKT pathways. Our results are consistent with their study, indicating that the upregulated signaling pathways in nonprogressors is helpful for cell survival and the development of effector functions of the immune system.
In the past several years, a wealth of evidence has emerged illustrating how metabolism supports many aspects of immune system biology [
55]. The field of cellular immunometabolism has made big strides over the past decade, becoming one of the hottest areas of research in immunology [
56]. Through KEGG Pathway analysis, we found OXPHOS was downregulated in all kinds of samples, including whole blood, CD4
+ and CD8
+ T cells. Through further analysis of metabolic pathways, we found the number of downregulated DEGs was more than upregulated DEGs in different metabolic processes, including carbohydrate metabolic process, cellular amino acid metabolic process, lipid metabolic process and protein metabolic process, and almost all the DEGs in OXPHOS and the TCA cycle were downregulated in the three sample types in nonprogressors compared with progressors. Previous studies have indicated that HIV itself and/or ART can damage the mitochondria, affecting the normal functioning of the cell [
57‐
59]. Two recent studies showed that HIV infection caused the upregulation of components of OXPHOS, TCA cycle, amino acid metabolism, and fatty acid metabolism in human CD4
+ T cell lines at the protein level, which may be compensation for the mitochondria dysfunction [
60,
61]. Previous studies have demonstrated that the LTNPs had slighter mitochondrial impairment and lower frequencies of cells with decreased mitochondrial membrane potential; this correlates may result in suppression of spontaneous apoptosis and higher CD4
+ T cells counts when compared to HIV infected individuals [
44,
45]. Compared with viremic patients in the HAART (VIR) group, LTNP downregulated the OXPHOS pathway and the TCA cycle in CD4
+ and CD8
+ T cells [
14,
62]. Our results revealed that nonprogressors have lower levels of the OXPHOS pathway and the TCA cycle, which may be due to the milder mitochondrial impairment in nonprogressors than progressors. Generating more effective adoptive cellular immunotherapies to rescue the abnormal metabolic profiles in progressors is valuable for delaying disease progression.