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The increasing application of single-cell RNA sequencing (scRNA-seq) technology in life science and biomedical research has significantly increased our understanding of the cellular heterogeneities in immunology, oncology and developmental biology. This review will summarize the development of various scRNA-seq technologies; primarily discussing the application of scRNA-seq on infectious diseases, and exploring the current development, challenges, and potential applications of scRNA-seq technology in the future.

Although immunotherapy has emerged as an effective therapeutic strategy for various cancers including head and neck squamous cell carcinomas (HNSCCs), only a subset of patients can benefit from such therapy. Hence, it is pressing to discover predictive biomarkers for cancer immunotherapy response. TP53 and HRAS mutations frequently occur in HNSCC and correlate with a worse prognosis in HNSCC. We extensively characterized the associations of TP53 mutations and HRAS mutations with HNSCC immunity based on multiple cancer genomics datasets. We compared the enrichment levels of 20 immune signatures between TP53-mutated and TP53-wildtype HNSCCs, and between HRAS-mutated and HRAS-wildtype HNSCCs, and found that TP53 mutations were associated with depressed immune signatures while HRAS mutations were associated with enhanced immune signatures in HNSCC. Moreover, we found multiple p53- and RAS-mediated pathways showing significant correlations with HNSCC immunity. Furthermore, we demonstrated that the association between TP53 mutation and tumor immunity was independent of the human papillomavirus (HPV) infection and smoking status in HNSCC. These data suggest that p53 and RAS may play important roles in regulating HNSCC immunity and that the TP53 and HRAS mutation status could be useful biomarkers for stratifying HNSCC patients responsive to immunotherapy., Graphical abstract TP53-mutations result in the deregulation of p53-mediated cell cycle, apoptosis, and genome stability thereby contributing to the depressed immune activity in HNSCC.Unlabelled Image

Although previous studies have shown that the host immune response is crucial in determining clinical outcomes in COVID-19 patients, the association between host immune signatures and COVID-19 patient outcomes remains unclear. Based on the enrichment levels of 11 immune signatures (eight immune-inciting and three immune-inhibiting signatures) in leukocytes of 100 COVID-19 patients, we identified three COVID-19 subtypes: Im-C1, Im-C2, and Im-C3, by clustering analysis. Im-C1 had the lowest immune-inciting signatures and high immune-inhibiting signatures. Im-C2 had medium immune-inciting signatures and high immune-inhibiting signatures. Im-C3 had the highest immune-inciting signatures while the lowest immune-inhibiting signatures. Im-C3 and Im-C1 displayed the best and worst clinical outcomes, respectively, suggesting that antiviral immune responses alleviated the severity of COVID-19 patients. We further demonstrated that the adaptive immune response had a stronger impact on COVID-19 outcomes than the innate immune response. The patients in Im-C3 were younger than those in Im-C1, indicating that younger persons have stronger antiviral immune responses than older persons. Nevertheless, we did not observe a significant association between sex and immune responses in COVID-19 patients. In addition, we found that the type II IFN response signature was an adverse prognostic factor for COVID-19. Our identification of COVID-19 immune subtypes has potential clinical implications for the management of COVID-19 patients.

The sudden outbreak of COVID-19 has once again shrouded people in the enormous threat of RNA virus. Extracellular vesicles (EVs), eukaryotic cells-derived small bi-layer vesicles mainly consisting of exosomes and microvesicles, share many properties with RNA viruses including structure, size, generation, and uptake. Emerging evidence has implicated the involvement of EVs in the pathogenesis of infectious diseases induced by RNA viruses. EVs can transfer viral receptors (e.g., ACE2 and CD9) to recipient cells to facilitate viral infection, directly transport infectious viral particles to adjacent cells for virus spreading, and mask viruses with a host structure to escape immune surveillance. Here, we examine the current status of EVs to summarize their roles in mediating RNA virus infection, together with a comprehensive discussion of the underlying mechanisms.

The COVID-19-, SARS- and MERS-related coronaviruses share many genomic and structural similarities. However, the SARS-CoV-2 is less pathogenic than SARS-CoV and MERS-CoV. Despite some differences in the cytokine patterns, it seems that the cytokine storm plays a crucial role in the pathogenesis of COVID-19-, SARS- and MERS. Monocytes and macrophages may be infected by SARS-CoV-2 through ACE2-dependent and ACE2-independent pathways. SARS-CoV-2 can effectively suppress the anti-viral IFN response in monocytes and macrophages. Since macrophages and dendritic cells (DCs) act as antigen presenting cells (APCs), the infection of these cells by SARS-CoV-2 impairs the adaptive immune responses against the virus. Upon infection, monocytes migrate to the tissues where they become infected resident macrophages, allowing viruses to spread through all organs and tissues. The SARS-CoV-2-infected monocytes and macrophages can produce large amounts of numerous types of pro-inflammatory cytokines and chemokines, which contribute to local tissue inflammation and a dangerous systemic inflammatory response called cytokine storm. Both local tissue inflammation and the cytokine storm play a fundamental role in the development of COVID-19-related complications, such as acute respiratory distress syndrome (ARDS), which is a main cause of death in COVID-19 patients. Here, we describe the monocytes and macrophage responses during severe coronavirus infections, while highlighting potential therapeutic interventions to attenuate macrophage-related inflammatory reactions in possible approaches for COVID-19 treatment., Graphical abstract Unlabelled Image, Highlights • A subset of monocytes and macrophages may be infected by SARS-CoV-2. • Coronavirus-infected monocyte can migrate to tissue and become infected macrophage. • Coronavirus-infected monocyte/macrophage produces high level of inflammatory factors. • Monocyte/macrophage-related cytokines promote organ inflammation and cytokine storm. • Local tissue inflammation and cytokine storm cause multi-organ failure in COVID-19.

Mitochondrial reactive oxygen species (mtROS) generated continuously under physiological conditions have recently emerged as critical players in the regulation of immune signaling pathways. In this study we have investigated the regulation of antiviral signaling by increased mtROS production in plasmacytoid dendritic cells (pDCs), which, as major producers of type I interferons (IFN), are the key coordinators of antiviral immunity. The early phase of type I IFN production in pDCs is mediated by endosomal Toll-like receptors (TLRs), whereas the late phase of IFN response can also be triggered by cytosolic retinoic acid-inducible gene-I (RIG-I), expression of which is induced upon TLR stimulation. Therefore, pDCs provide an ideal model to study the impact of elevated mtROS on the antiviral signaling pathways initiated by receptors with distinct subcellular localization. We found that elevated level of mtROS alone did not change the phenotype and the baseline cytokine profile of resting pDCs. Nevertheless increased mtROS levels in pDCs lowered the TLR9-induced secretion of pro-inflammatory mediators slightly, whereas reduced type I IFN production markedly via blocking phosphorylation of interferon regulatory factor 7 (IRF7), the key transcription factor of the TLR9 signaling pathway. The TLR9-induced expression of RIG-I in pDCs was also negatively regulated by enhanced mtROS production. On the contrary, elevated mtROS significantly augmented the RIG-I-stimulated expression of type I IFNs, as well as the expression of mitochondrial antiviral-signaling (MAVS) protein and the phosphorylation of Akt and IRF3 that are essential components of RIG-I signaling. Collectively, our data suggest that increased mtROS exert diverse immunoregulatory functions in pDCs both in the early and late phase of type I IFN responses depending on which type of viral sensing pathway is stimulated., Graphical abstract fx1, Highlights • MtROS have versatile role in fine-tuning the type I IFN responses of pDCs. • MtROS affect the TLR- and RIG-I-mediated type I IFN responses in pDCs oppositely. • MtROS decrease the TLR-induced early phase of type I IFN production in pDCs. • MtROS enhance the RIG-I-mediated late phase of type I IFN production in pDCs.

Wilms' tumors (WT), which accountfor 6% of all childhood cancers, arise from dysregulated differentiation of nephrogenic progenitor cells from embryonic kidneys. Though there is an improvement in the prognosis of WT, still 10% of patients with WT die due to recurrence. Thus more effective treatment approaches are necessary. We previously characterized the inflammatory microenvironment in human WT and observed the robust expression of COX-2. The aim of this study was to extend our studies to analyze the role of COX-2 pathway components in WT progression using a mouse model of WT. Herein, COX-2 pathway components such as COX-2, HIF1-α, p-ERK1/2, and p-STAT3 were upregulated in mouse and human tumor tissues. In our RPPA analysis, COX-2 was up-regulated in M15 cells after Wt1 gene was knocked down. Flow cytometry analysis showed the increased infiltration of immune suppressive inflammatory cells such as pDC's and Treg cells in tumors. The chemotactic chemokines responsible for the infiltration of these cells were also induced in CCR5 and CXCR4 dependent manner respectively. The immunosuppressive cytokines IL-10, TGF-β, and TNF-α were also up-regulated. Furthermore, more pronounced Th2 and Treg induced cytokine response was observed than Th1 response in tumors. Basing on all these evidences it is speculated that COX-2 pathway may be a beneficial target for the treatment of WT. It may be most effective as an adjuvant therapy together with other inhibitors. Thus, our current study provides a good rationale for initiating animal studies to confirm the efficacy of COX-2 inhibitors in decreasing tumor cell growth in vivo.

Plasmacytoid dendritic cells (pDCs) have been shown to both mediate and prevent autoimmunity, and the regulation of their immunogenic versus tolerogenic functions remains incompletely understood. Here we demonstrate that, compared to other cells, pDCs are the major expressors of Indoleamine-2,3-dioxygenase (IDO) in steady-state lymph nodes (LNs). IDO expression by LN pDCs was closely dependent on MHCII-mediated, antigen-dependent, interactions with Treg. We further established that IDO production by pDCs was necessary to confer suppressive function to Tregs. During EAE development, IDO expression by pDCs was required for the generation of Tregs capable of dampening the priming of encephalitogenic T cell and disease severity. Thus, we describe a novel crosstalk between pDCs and Tregs: Tregs shape tolerogenic functions of pDCs prior to inflammation, such that pDCs in turn, promote Treg suppressive functions during autoimmunity., Graphical abstract, Highlights • IDO expression by LN pDCs is closely dependent on MHCII-mediated, antigen-dependent, interactions with Tregs. • pDCs are the predominant source of IDO in both steady-state and inflamed lymph nodes. • IDO production by pDCs is necessary to confer suppressive function to Tregs in EAE.

Graphical abstract, Highlights • Induction and action of IFN during acute and chronic HIV-1 infection. • Host and viral factors influencing IFN response in HIV-1 infected patients. • Type I IFN and IFNα subtypes signatures and their antiviral activity during HIV-1 infection. • The microbiome and intestinal IFN responses relationship in HIV-1 infection and disease., Type I interferon (IFN) response initially limits HIV-1 spread and may delay disease progression by stimulating several immune system components. Nonetheless, persistent exposure to type I IFN in the chronic phase of HIV-1 infection is associated with desensitization and/or detrimental immune activation, thereby hindering immune recovery and fostering viral persistence. This review provides a basis for understanding the complexity and function of IFN pleiotropic activity in HIV-1 infection. In particular, the dichotomous role of the IFN response in HIV-1 immunopathogenesis will be discussed, highlighting recent advances in the dynamic modulation of IFN production in acute versus chronic infection, expression signatures of IFN subtypes, and viral and host factors affecting the magnitude of IFN response during HIV-1 infection. Lastly, the review gives a forward-looking perspective on the interplay between microbiome compositions and IFN response.

Highlights • Type I interferon represents the first line of innate host defense against a number of invading pathogens that also instructs adaptive immunity. • Type I interferon induction downstream germ-line-encoded pattern recognition receptors triggers a signaling pathway that results in the expression of hundreds of IFN-stimulated genes with both direct anti-pathogen as well as immunomodulatory activities. • An inverse interference to escape at every step the IFN system is used by pathogens to eventually promote disease progression or establish a chronic infection. • Learning from bacterial and viral escape mechanisms may help in understanding how to cheat the pathogen and fit out the immune system to shift the balance in favor of the host., Type I interferon (IFN) comprises a class of cytokines first discovered more than 50 years ago and initially characterized for their ability to interfere with viral replication and restrict locally viral propagation. As such, their induction downstream of germ-line encoded pattern recognition receptors (PRRs) upon recognition of pathogen-associated molecular patterns (PAMPs) is a hallmark of the host antiviral response. The acknowledgment that several PAMPs, not just of viral origin, may induce IFN, pinpoints at these molecules as a first line of host defense against a number of invading pathogens. Acting in both autocrine and paracrine manner, IFN interferes with viral replication by inducing hundreds of different IFN-stimulated genes with both direct anti-pathogenic as well as immunomodulatory activities, therefore functioning as a bridge between innate and adaptive immunity. On the other hand an inverse interference to escape the IFN system is largely exploited by pathogens through a number of tactics and tricks aimed at evading, inhibiting or manipulating the IFN pathway, that result in progression of infection or establishment of chronic disease. In this review we discuss the interplay between the IFN system and some selected clinically important and challenging viruses and bacteria, highlighting the wide array of pathogen-triggered molecular mechanisms involved in evasion strategies.

There is increased production of plasmacytoid dendritic cells (pDCs) in the bone marrow (BM) of multiple myeloma (MM) patients and these favor Th22 cell differentiation. Here, we found that the frequency of interleukin (IL)-22+IL-17-IL-13+ T cells is significantly increased in peripheral blood (PB) and BM of stage III and relapsed/refractory MM patients compared with healthy donors and patients with asymptomatic or stage I/II disease. Th22 cells cloned from the BM of MM patients were CCR6+CXCR4+CCR4+CCR10- and produced IL-22 and IL-13 but not IL-17. Furthermore, polyfunctional Th22-Th2 and Th22-Th1 clones were identified based on the co-expression of additional chemokine receptors and cytokines (CRTh2 or CXCR3 and IL-5 or interferon gamma [IFNγ], respectively). A fraction of MM cell lines and primary tumors aberrantly expressed the IL-22RA1 and IL-22 induced STAT-3 phosphorylation, cell growth, and resistance to drug-induced cell death in MM cells. IL-13 treatment of normal BM mesenchymal stromal cells (MSCs) induced STAT-6 phosphorylation, adhesion molecule upregulation, and increased IL-6 production and significantly favored MM cell growth compared with untreated BM MSCs. Collectively, our data show that increased frequency of IL-22+IL-17-IL-13+ T cells correlates with poor prognosis in MM through IL-22 and IL-13 protumor activity and suggest that interference with IL-22 and IL-13 signaling pathways could be exploited for therapeutic intervention.

Virus infection is sensed by the innate immune system which then rapidly initiates biosynthesis of type I interferon (IFN). The IFN signaling systems produce a broadly effective innate antiviral response by creating an antiviral state in both an autocrine and paracrine manner in cells and by activating innate and adaptive immunity. Plasmacytoid dendritic cells (pDCs) have the unique ability to produce very high levels of type I IFN following viral infection in vivo. Most recent research has focused on oligonucleotide-mediated induction of type I IFN production, implicating viral genome and replication intermediates as the stimulus for this response. However there are additional viral ligands which can potentially induce type I IFN production in pDCs, such as envelope glycoproteins, viral glycolipids, tegument, capsid or nuclear proteins. This area of viral immunology, which has been neglected in the literature, will be discussed here.

To develop CpG oligodeoxynucleotides (CpG ODNs) based therapy for prevention and treatment of severe acute respiratory syndrome (SARS), we selected a novel CpG ODN (BW001), which displays B-type CpG ODN structure feature at the 5' and A-type CpG ODN structure feature at the 3', and tested for its anti-SARS-CoV activity. We found that the supernatants of human PBMCs stimulated by BW001 significantly protected Vero cells from SARS-CoV infection. BW001 could stimulate human PBMCs and pDCs to secrete high level of IFN-alpha and promote human PBMCs and B cells to proliferate. Furthermore, we demonstrated that BW001 could activate CD19+ B cells and CD56+ NK cells in human PBMCs. In addition, BW001 could enhance NK cytotoxicity and IFN-gamma secretion in human PBMCs. Together, BW001 represents a novel type of CpG ODN and may have potential for the development of treatment and prevention for SARS as well as other viral associated diseases.

Several studies have demonstrated a strong link between Chlamydia pneumoniae (Cp) infection and atherosclerosis progression/exacerbation. Here, we try to understand whether a single administration of Cp could exacerbate atherosclerosis. Apoe(-/-) mice were intranasally infected with Cp followed by a high fat diet. Mice were sacrificed at different time points after Cp infection to monitor the development of the atheroma. Cp infection increased lipid content in the aortic sinus of Apoe(-/-) mice starting from 8 weeks. This was associated with increased numbers of active myeloid dendritic cells and plasmacytoid DCs which were co-localized with T-cells in the atherosclerotic plaque. The serum levels of IFN-γ showed a Th1-like environment typical of atherosclerosis. In conclusion, we demonstrate that one dose of Cp could exacerbate atherosclerotic lesion development, triggering innate immune cell accumulation early on that allowed the involvement of Th1-like cells in the exacerbation of the atherosclerotic plaque at later time points.