Upregulation Of IFN-stimulated Genes Research Articles

Non-invasive SARS-CoV-2 RNA detection and human transcriptome analysis using skin surface lipids

There have been several reports of skin manifestations in patients with coronavirus disease 2019 (COVID-19). However, it is unclear whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA can be detected on the skin surface, including the sebum, of these patients. In this study, SARS-CoV-2 RNA was detected using real-time reverse-transcription polymerase chain reaction (RT-PCR) assay of skin surface lipids (SSLs) collected using an oil-blotting film from the faces of hospitalized patients with COVID-19. Human transcriptome analysis was also performed using the same samples. In facial SSLs of patients with COVID-19, the RT-PCR positivity rate was 84.6% (11/13 samples) within 5 days and 30.4% (7/23 samples) by 6–10 days of symptom onset. In the transcriptome analysis, the most characteristic SSL-RNA profile was the upregulation of interferon-stimulated gene (ISG)-related genes, such as ISG15, IFITM1, and MX1. This study presents an alternative technique using SSLs for non-invasive SARS-CoV-2 RNA detection and simultaneous analysis of human molecular pathogenesis in patients with COVID-19.

Interferon Stimulated Gene Expression Is a Biomarker for Primary Mitochondrial Disease.

Mitochondria are implicated in regulation of the innate immune response. We hypothesized that abnormalities in interferon signaling may contribute to pathophysiology in patients with primary mitochondrial disease (PMD). Expression of interferon stimulated genes (ISGs) was measured by real-time polymerase chain reaction (PCR) in whole blood samples from a cohort of patients with PMD. Upregulated ISG expression was observed in a high proportion (41/55, 75%) of patients with PMD on at least 1 occasion, most frequently IFI27 upregulation, seen in 50% of the samples. Some patients had extremely high IFI27 levels, similar to those seen in patients with primary interferonopathies. A statistically significant correlation was observed between elevated IFI27 gene expression and PMD, but not between IFI27 and secondary mitochondrial dysfunction, suggesting that ISG upregulation is a biomarker of PMD. In some patients with PMD, ISG abnormalities persisted on repeat measurement over several years, indicative of ongoing chronic inflammation. Subgroup analyses suggested common ISG signatures in patients with similar mitochondrial disease mechanisms and positive correlations with disease severity among patients with identical genetic diagnoses. Dysregulated interferon signaling is frequently seen in patients with PMD suggesting that interferon dysregulation is a contributor to pathophysiology. This may indicate a role for repurposing of immunomodulatory therapies for the treatment of PMDs by targeting interferon signaling. ANN NEUROL 2024.

Innate immune signatures in the nasopharynx after SARS-CoV-2 infection and links with the clinical outcome of COVID-19 in Omicron-dominant period

While severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is characterized by impaired induction of interferons (IFNs) and IFN-stimulated genes (ISGs), the IFNs and ISGs in upper airway is essential to restrict the spread of respiratory virus. Here, we identified the prominent IFN and ISG upregulation in the nasopharynx (NP) of mild and even severe coronavirus disease 2019 (COVID-19) patients (CoV2+) in Omicron era and to compare their clinical outcome depending on the level of IFNs and ISGs. Whereas the induction of IFNB was minimal, transcription of IFNA, IFNG, and IFNLs was significantly increased in the NP of CoV2 + patients. IFNs and ISGs may be more upregulated in the NP of CoV2 + patients at early phases of infection according to viral RNA levels and this is observed even in severe cases. IFN-related innate immune response might be characteristic in macrophages and monocytes at the NP and the CoV2 + patients with higher transcription of IFNs and ISGs in the NP showed a correlation with good prognosis of COVID-19. This study presents that IFNs and ISGs may be upregulated in the NP, even in severe CoV2 + patients depending on viral replication during Omicron-dominant period and the unique IFN-responsiveness in the NP links with COVID-19 clinical outcomes.

TAGLN2 induces resistance signature ISGs by activating AKT-YBX1 signal with dual pathways and mediates the IFN-related DNA damage resistance in gastric cancer

Recently, various cancer types have been identified to express a distinct subset of Interferon-stimulated genes (ISGs) that mediate therapy resistance. The mechanism through which cancer cells maintain prolonged Interferon stimulation effects to coordinate resistance remains unclear. Our research demonstrated that aberrant upregulation of TAGLN2 is associated with gastric cancer progression, and inhibiting its expression renders gastric cancer cells more susceptible to chemotherapy and radiation. We uncovered a novel role for TAGLN2 in the upregulation of resistance signature ISGs by enhancing YBX1-associated ssDNA aggregation and cGAS-STING pathway activation. TAGLN2 modulates YBX1 by recruiting c-Myc and SOX9 to YBX1 promoter region and directly interacting with AKT-YBX1, thereby enhancing YBX1 phosphorylation and nuclear translocation. Significantly, targeted downregulation of key proteins, inhibition of the TAGLN2-YBX1-AKT interaction (using Fisetin or MK2206) or disruption of the cGAS-STING pathway substantially reduced ssDNA accumulation, subsequent ISGs upregulation, and therapy resistance. The combination of Cisplatin with MK2206 displayed a synergistic effect in the higher TAGLN2-expressing xenograft tumors. Clinical analysis indicated that a derived nine-gene set effectively predicts therapeutic sensitivity and long-term prognosis in gastric cancer patients. These findings suggest that TAGLN2, YBX1 and induced ISGs are novel predictive markers for clinical outcomes, and targeting this axis is an attractive therapeutic sensitization strategy.

Loss of the DNA repair protein, polynucleotide kinase/phosphatase, activates the type 1 interferon response independent of ionizing radiation.

DNA damage has been implicated in the stimulation of the type 1 interferon (T1IFN) response. Here, we show that downregulation of the DNA repair protein, polynucleotide kinase/phosphatase (PNKP), in a variety of cell lines causes robust phosphorylation of STAT1, upregulation of interferon-stimulated genes and persistent accumulation of cytosolic DNA, all of which are indicators for the activation of the T1IFN response. Furthermore, this did not require damage induction by ionizing radiation. Instead, our data revealed that production of reactive oxygen species (ROS) synergises with PNKP loss to potentiate the T1IFN response, and that loss of PNKP significantly compromises mitochondrial DNA (mtDNA) integrity. Depletion of mtDNA or treatment of PNKP-depleted cells with ROS scavengers abrogated the T1IFN response, implicating mtDNA as a significant source of the cytosolic DNA required to potentiate the T1IFN response. The STING signalling pathway is responsible for the observed increase in the pro-inflammatory gene signature in PNKP-depleted cells. While the response was dependent on ZBP1, cGAS only contributed to the response in some cell lines. Our data have implications for cancer therapy, since PNKP inhibitors would have the potential to stimulate the immune response, and also to the neurological disorders associated with PNKP mutation.

Abstract 3242: TNG917 is a clinical-grade, potent and selective inhibitor of EHMT1/2 for the treatment of immune cold tumors

Abstract CRISPR-based functional genomics screening can be designed to identify novel cancer cell intrinsic targets that increase tumor immunogenicity. Using a FACS-based CRISPR sorting screen for PD-L1 expression, we identified Euchromatic histone-lysine-N-methyltransferase 1 and 2 (EHMT1/2) as negative modulators of the interferon signaling pathways. EHMT1 and EHMT2 are histone methyltransferases that mono- and di-methylate lysine 9 of histone H3 to repress gene transcription of defined target genes. Gene knockout or pharmacological inhibition of EHMT1/2 in cancer cells resulted in de-repression of gene promoters, upregulation of interferon-stimulated genes (ISGs), and secretion of pro-inflammatory cytokines. Here, we present the preclinical characterization of TNG917 - an oral and highly selective EHMT1/2 inhibitor with low nanomolar cellular potency, and favorable pharmacodynamic and pharmacokinetic properties. In humanized and syngeneic mouse models, treatment with TNG917 in combination with anti-PD1 promoted a T-cell-infiltrated tumor microenvironment, led to significant anti-tumor activity, and resulted in survival benefit. In summary, our in vitro and in vivo studies provide a rationale for the clinical development path of TNG917, in combination with checkpoint inhibition, in patients with immune cold tumors. Citation Format: Alvin Lu, Brian Haines, Ji Lei, Wenhai Zhang, Minjie Zhang, Douglas A. Whittington, Maria Lucia Ferdinez, Yi Yu, Samuel R. Meier, Ashley Choi, Alborz Bejnood, Madhavi Bandi, Katherine Lazarides, Xuewen Pan, Lina Gu, Alice W. Tsai, Sirimas Sudsakorn, Colin Liang, Jon Come, Brett Williams, Scott Throner, Joseph Vacca, John P. Maxwell, Jannik N. Anderson, Alan Huang, Chengyin Min, Yingnan P. Chen. TNG917 is a clinical-grade, potent and selective inhibitor of EHMT1/2 for the treatment of immune cold tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3242.

Janus kinase-inhibition modulates hepatitis E virus infection

Hepatitis E virus (HEV) usually causes a self-limiting disease, but especially immunocompromised individuals are at risk to develop a chronic and severe course of infection. Janus kinase (JAK) inhibitors (JAKi) are a novel drug class for the treatment of autoimmune inflammatory rheumatic disease (AIRD). As JAKs play a key role in innate immunity, viral infections and reactivations are frequently reported during JAKi treatment in AIRD patients. The aim of this study was to characterize the influence of JAKis on HEV replication. To this end, we evaluated liver enzymes of an AIRD patient under JAKi therapy with hepatitis E. Further, experiments with HEV (Kernow-C1 p6) were performed by infection of primary human hepatocytes (PHHs) followed by immunofluorescence staining of viral markers and transcriptomic analysis. Infection experiments in PHHs displayed an up to 50-fold increase of progeny virus production during JAKi treatment and transcriptomic analysis revealed induction of antiviral programs during infection. Upregulation of interferon-stimulated genes (ISG) was perturbed in the presence of JAKis, concomitant with elevated HEV RNA levels. The obtained results suggest that therapeutic JAK inhibition increases HEV replication by modulating the HEV-triggered immune response. Therefore, JAKi treatment and the occurrence of elevated liver enzymes requires a monitoring of potential HEV infections.

Hepatitis D virus infection, innate immune response and antiviral treatments in stem cell-derived hepatocytes.

Human pluripotent stem cell (hPSC)-derived hepatocyte-like cells (HLCs) are a valuable model to investigate host-pathogen interactions of hepatitis viruses in a mature and authentic environment. Here, we investigate the susceptibility of HLCs to the hepatitis delta virus (HDV). We differentiated hPSC into HLCs, and inoculated them with infectious HDV produced in Huh7NTCP . HDV infection and cellular response was monitored by RTqPCR and immunostaining. Cells undergoing hepatic differentiation become susceptible to HDV after acquiring expression of the viral receptor Na+ -taurocholate co-transporting polypeptide (NTCP) during hepatic specification. Inoculation of HLCs with HDV leads to detection of intracellular HDV RNA and accumulation of the HDV antigen in the cells. Upon infection, the HLCs mounted an innate immune response based on induction of the interferons IFNB and L, and upregulation of interferon-stimulated genes. The intensity of this immune response positively correlated with the level of viral replication and was dependant on both the JAK/STAT and NFκB pathway activation. Importantly, this innate immune response did not inhibit HDV replication. However, pre-treatment of the HLCs with IFNα2b reduced viral infection, suggesting that ISGs may limit early stages of infection. Myrcludex efficiently abrogated infection and blocked innate immune activation. Lonafarnib treatment of HDV mono infected HLCs on the other hand led to exacerbated viral replication and innate immune response. The HDV in vitro mono-infection model represents a new tool to study HDV replication, its host-pathogen interactions and evaluate new antiviral drugs in cells displaying mature hepatic functions.

Transcriptomics approaches reveal neuronal antiviral immune mechanisms.

Abstract Arthropod-borne viruses exist worldwide and contribute to many neurological infections. Viral infections in the central nervous system (CNS) can lead to virus- and immune-mediated damage of the brain, which can cause severe physical and cognitive defects, or even death. While antiviral responses (such as Type I interferon) have been characterized in the periphery, it is not well understood how innate immune response are coordinated by the various cell types present in the brain. Certain neurotropic arboviruses, such as La Crosse Virus (LACV), predominantly infect neurons; however, it remains unclear how neurons mediate early innate immune responses in the CNS. Upon infection of primary murine cortical neurons and human IPSC-derived cerebral organoids with LACV, we observed widespread infection. Transcriptomics approaches revealed infection of neurons results in the upregulation of interferon-stimulated genes (ISGs) that are also detectable at the protein level. Notably, we found that bystander uninfected neurons have higher ISG expression than neighboring infected cells. Future directions aim to characterize how infected neurons communicate with uninfected bystander neurons during infection. To address this, I have optimized a ProbeSeq pipeline where I infect primary cortical neurons and further characterize infected and uninfected populations using fluorescence in situ hybridization and downstream RNA-sequencing. Supported by the Penn Presidential PhD Fellowship.

Characterization of a mutant samhd1 zebrafish model implicates dysregulation of cholesterol biosynthesis in Aicardi-Goutières syndrome.

Aicardi-Goutières syndrome (AGS1-9) is a genetically determined encephalopathy that falls under the type I interferonopathy disease class, characterized by excessive type I interferon (IFN-I) activity, coupled with upregulation of IFN-stimulated genes (ISGs), which can be explained by the vital role these proteins play in self-non-self-discrimination. To date, few mouse models fully replicate the vast clinical phenotypes observed in AGS patients. Therefore, we investigated the use of zebrafish as an alternative species for generating a clinically relevant model of AGS. Using CRISPR-cas9 technology, we generated a stable mutant zebrafish line recapitulating AGS5, which arises from recessive mutations in SAMHD1. The resulting homozygous mutant zebrafish larvae possess a number of neurological phenotypes, exemplified by variable, but increased expression of several ISGs in the head region, a significant increase in brain cell death, microcephaly and locomotion deficits. A link between IFN-I signaling and cholesterol biosynthesis has been highlighted by others, but not previously implicated in the type I interferonopathies. Through assessment of neurovascular integrity and qPCR analysis we identified a significant dysregulation of cholesterol biosynthesis in the zebrafish model. Furthermore, dysregulation of cholesterol biosynthesis gene expression was also observed through RNA sequencing analysis of AGS patient whole blood. From this novel finding, we hypothesize that cholesterol dysregulation may play a role in AGS disease pathophysiology. Further experimentation will lend critical insight into the molecular pathophysiology of AGS and the potential links involving aberrant type I IFN signaling and cholesterol dysregulation.

Type I Interferonopathy due to a Homozygous Loss-of-Inhibitory Function Mutation in STAT2

PurposeSTAT2 is both an effector and negative regulator of type I interferon (IFN-I) signalling. We describe the characterization of a novel homozygous missense STAT2 substitution in a patient with a type I interferonopathy.MethodsWhole-genome sequencing (WGS) was used to identify the genetic basis of disease in a patient with features of enhanced IFN-I signalling. After stable lentiviral reconstitution of STAT2-null human fibrosarcoma U6A cells with STAT2 wild type or p.(A219V), we performed quantitative polymerase chain reaction, western blotting, immunofluorescence, and co-immunoprecipitation to functionally characterize the p.(A219V) variant.ResultsWGS identified a rare homozygous single nucleotide transition in STAT2 (c.656C > T), resulting in a p.(A219V) substitution, in a patient displaying developmental delay, intracranial calcification, and up-regulation of interferon-stimulated gene (ISG) expression in blood. In vitro studies revealed that the STAT2 p.(A219V) variant retained the ability to transduce an IFN-I stimulus. Notably, STAT2 p.(A219V) failed to support receptor desensitization, resulting in sustained STAT2 phosphorylation and ISG up-regulation. Mechanistically, STAT2 p.(A219V) showed defective binding to ubiquitin specific protease 18 (USP18), providing a possible explanation for the chronic IFN-I pathway activation seen in the patient.ConclusionOur data indicate an impaired negative regulatory role of STAT2 p.(A219V) in IFN-I signalling and that mutations in STAT2 resulting in a type I interferonopathy state are not limited to the previously reported R148 residue. Indeed, structural modelling highlights at least 3 further residues critical to mediating a STAT2-USP18 interaction, in which mutations might be expected to result in defective negative feedback regulation of IFN-I signalling.

Antiviral Effect of Manganese against Foot-and-Mouth Disease Virus Both in PK15 Cells and Mice.

Foot-and-mouth disease (FMD) is an acute contagious disease of cloven-hoofed animals such as cattle, pigs, and sheep. Current emergency FMD vaccines are of limited use for early protection because their protective effect starts 7 days after vaccination. Therefore, antiviral drugs or additives are used to rapidly stop the spread of the virus during FMD outbreaks. Manganese (Mn2+) was recently found to be an important substance necessary for the host to protect against DNA viruses. However, its antiviral effect against RNA viruses remains unknown. In this study, we found that Mn2+ has antiviral effects on the FMD virus (FMDV) both in PK15 cells and mice. The inhibitory effect of Mn2+ on FMDV involves NF-κB activation and up-regulation of interferon-stimulated genes. Animal experiments showed that Mn2+ can be highly effective in protecting C57BL/6N mice from being infected with FMDV. Overall, we suggest Mn2+ as an effective antiviral additive for controlling FMDV infection.

Activation of Interferon-Stimulated Genes following Varicella-Zoster Virus Infection in a Human iPSC-Derived Neuronal In Vitro Model Depends on Exogenous Interferon-α.

Varicella-zoster virus (VZV) infection of neuronal cells and the activation of cell-intrinsic antiviral responses upon infection are still poorly understood mainly due to the scarcity of suitable human in vitro models that are available to study VZV. We developed a compartmentalized human-induced pluripotent stem cell (hiPSC)-derived neuronal culture model that allows axonal VZV infection of the neurons, thereby mimicking the natural route of infection. Using this model, we showed that hiPSC-neurons do not mount an effective interferon-mediated antiviral response following VZV infection. Indeed, in contrast to infection with Sendai virus, VZV infection of the hiPSC-neurons does not result in the upregulation of interferon-stimulated genes (ISGs) that have direct antiviral functions. Furthermore, the hiPSC-neurons do not produce interferon-α (IFNα), a major cytokine that is involved in the innate antiviral response, even upon its stimulation with strong synthetic inducers. In contrast, we showed that exogenous IFNα effectively limits VZV spread in the neuronal cell body compartment and demonstrated that ISGs are efficiently upregulated in these VZV-infected neuronal cultures that are treated with IFNα. Thus, whereas the cultured hiPSC neurons seem to be poor IFNα producers, they are good IFNα responders. This could suggest an important role for other cells such as satellite glial cells or macrophages to produce IFNα for VZV infection control.

Pin-Pointing the Key Hubs in the IFN-γ Pathway Responding to SARS-CoV-2 Infection.

Interferon gamma (IFN-γ) may be potential adjuvant immunotherapy for COVID-19 patients. In this work, we assessed gene expression profiles associated with the IFN-γ pathway in response to SARS-CoV-2 infection. Employing a case-control study from SARS-CoV-2-positive and -negative patients, we identified IFN-γ-associated pathways to be enriched in positive patients. Bioinformatics analyses showed upregulation of MAP2K6, CBL, RUNX3, STAT1, and JAK2 in COVID-19-positive vs. -negative patients. A positive correlation was observed between STAT1/JAK2, which varied alongside the patient’s viral load. Expression of MX1, MX2, ISG15, and OAS1 (four well-known IFN-stimulated genes (ISGs)) displayed upregulation in COVID-19-positive vs. -negative patients. Integrative analyses showcased higher levels of ISGs, which were associated with increased viral load and STAT1/JAK2 expression. Confirmation of ISGs up-regulation was performed in vitro using the A549 lung cell line treated with Poly (I:C), a synthetic analog of viral double-stranded RNA; and in different pulmonary human cell lines and ferret tracheal biopsies infected with SARS-CoV-2. A pre-clinical murine model of Coronavirus infection confirmed findings displaying increased ISGs in the liver and lungs from infected mice. Altogether, these results demonstrate the role of IFN-γ and ISGs in response to SARS-CoV-2 infection, highlighting alternative druggable targets that can boost the host response.

Preventing SARS-CoV-2 Infection Using Anti-spike Nanobody-IFN-β Conjugated Exosomes.

PurposeTo inhibit the transmission of SARS-CoV-2, we developed engineered exosomes that were conjugated with anti-spike nanobodies and type I interferon β (IFN-β). We evaluated the efficacy and potency of nanobody-IFN-β conjugated exosomes to treatment of SARS-CoV-2 infection.MethodsMilk fat globule epidermal growth factor 8 (MFG-E8) is a glycoprotein that binds to phosphatidylserine (PS) exposed on the exosomes. We generated nanobody-IFN-β conjugated exosomes by fusing an anti-spike nanobody and IFN-β with MFG-E8. We used the SARS-CoV-2 pseudovirus with the spike of the D614G mutant that encodes ZsGreen to mimic the infection process of the SARS-CoV-2. The SARS-CoV-2 pseudovirus was infected with angiotensin-converting enzyme-2 (ACE2) expressing adenocarcinomic human alveolar basal epithelial cells (A549) or ACE2 expressing HEK-blue IFNα/β cells in the presence of nanobody-IFN-β conjugated exosomes. By assessing the expression of ZsGreen in target cells and the upregulation of interferon-stimulated genes (ISGs) in infected cells, we evaluated the anti-viral effects of nanobody-IFN-β conjugated exosomes.ResultsWe confirmed the anti-spike nanobody and IFN-β expressions on the exosomes. Exosomes conjugated with nanobody-hIFN-β inhibited the interaction between the spike protein and ACE2, thereby inhibiting the infection of host cells with SARS-CoV-2 pseudovirus. At the same time, IFN-β was selectively delivered to SARS-CoV-2 infected cells, resulting in the upregulation of ISGs expression.ConclusionExosomes conjugated with nanobody-IFN-β may provide potential benefits in the treatment of COVID-19 because of the cooperative anti-viral effects of the anti-spike nanobody and the IFN-β.Supplementary InformationThe online version contains supplementary material available at 10.1007/s11095-022-03400-0.

Investigation of the anti-pseudorabies virus activity of interferon lambda 3 in cultured porcine kidney epithelial cells.

It has been reported Interferon-λ (IFN-λ) has stronger antiviral effect than other interferons. IFN-λ can induce antiviral interferon stimulated genes (ISGs) in epithelia to protect against virus. Pseudorabies virus (PRV) infection in pigs resulting in fatal encephalitis in newborn piglets, respiratory disorders in finishing pigs, reproductive disorders in sows and other symptoms. Since the effect of IFN-λ on inhibiting PRV proliferation is still unknown. Inthis study, we investigate the relative contribution of porcine IFN-λ3 toward controlling the infection of PRV in vitro. Our findings may provide a new insight for the prevention and treatment of PRV. Therefore, the antiviral assay, western blot, qRT-PCR and ELISA assay were used to investigating the contribution of IFN-λ against PRV in PK-15 cells. Here, we demonstrate that the replication of PRV in PK-15 cells was inhibited after pre-treatment with IFN-λ3, and such inhibition was dose dependent. Overexpression of IFN-λ3 receptor (IFNLR) also restricted virus titre in PK-15 cells. In addition, IFN-λ3 also increased the mRNA and protein expression of interferon-stimulated genes 15 (ISG15), 2'-5'-oligoadenylate synthase 1 (OAS1), IFN-inducible transmembrane 3 (IFITM3) and myxoma resistance protein 1 (Mx1) in PRV-infected PK-15 cells. Other than modulation ISGs, IFN-λ specifically activated IFN-γ mRNA expression not IFN-α or IFN-β. IFN-λ3 had antiviral activity against PRV and the upregulation of ISGs and IFN-γ mRNA expression may be the mechanism of IFN-λ3's antiviral activities. Thus, IFN-λ3 has a decisive function that greatly limits PRV replication in PK-15 cells. Our study explores the antiviral activity of IFN-λ3 on PRV for the first time.

Intrahepatic transcriptomics reveals gene signatures in chronic hepatitis B patients responded to interferon therapy

ABSTRACT Chronic hepatitis B virus (HBV) infection remains a substantial public health burden worldwide. Alpha-interferon (IFNα) is one of the two currently approved therapies for chronic hepatitis B (CHB), to explore the mechanisms underlying IFNα treatment response, we investigated baseline and 24-week on-treatment intrahepatic gene expression profiles in 21 CHB patients by mRNA-seq. The data analyses demonstrated that PegIFNα treatment significantly induced antiviral responses. Responders who achieved HBV DNA loss and HBeAg or HBsAg seroconversion displayed higher fold change and larger number of up-regulated interferon-stimulated genes (ISGs). Interestingly, lower expression levels of certain ISGs were observed in responders in their baseline biopsy samples. In HBeAg+ patients, non-responders had relative higher baseline HBeAg levels than responders. More importantly, HBeAg− patients showed higher HBsAg loss rate than HBeAg+ patients. Although a greater fold change of ISGs was observed in HBeAg− patients than HBeAg+ patients, upregulation of ISGs in HBeAg+ responders exceeded HBeAg− responders. Notably, PegIFNα treatment increased monocyte and mast cell infiltration, but decreased CD8 T cell and M1 macrophage infiltration in both responders and non-responders, while B cell infiltration was increased only in responders. Moreover, co-expression analysis identified ribosomal proteins as critical players in antiviral response. The data also indicate that IFNα may influence the production of viral antigens associated with endoplasmic reticulum. Collectively, the intrahepatic transcriptome analyses in this study enriched our understanding of IFN-mediated antiviral effects in CHB patients and provided novel insights into the development of potential strategies to improve IFNα therapy.

Pandemic influenza A (H1N1) virus causes abortive infection of primary human T cells

ABSTRACT Influenza A virus still represents a noticeable epidemic risk to international public health at present, despite the extensive use of vaccines and anti-viral drugs. In the fight against pathogens, the immune defence lines consisting of diverse lymphocytes are indispensable for humans. However, the role of virus infection of lymphocytes and subsequent abnormal immune cell death remains to be explored. Different T cell subpopulations have distinct characterizations and functions, and we reveal the high heterogeneity of susceptibility to viral infection and biological responses such as apoptosis in various CD4+ T and CD8+ T cell subsets through single-cell transcriptome analyses. Effector memory CD8+ T cells (CD8+ TEM) that mediate protective memory are identified as the most susceptible subset to pandemic influenza A virus infection among primary human T cells. Non-productive infection is established in CD8+ TEM and naïve CD8+ T cells, which indicate the mechanism of intracellular antiviral activities for inhibition of virus replication such as abnormal viral splicing efficiency, incomplete life cycles and up-regulation of interferon-stimulated genes in human T cells. These findings provide insights into understanding lymphopenia and the infectious mechanisms of pandemic influenza A virus and broad immune host–pathogen interactional atlas in primary human T cells.

Precision treatment of Singleton Merten syndrome with ruxolitinib: a case report

BackgroundSingleton-Merten syndrome 1 (SGMRT1) is a rare type I interferonopathy caused by heterozygous mutations in the IFIH1 gene. IFIH1 encodes the pattern recognition receptor MDA5 which senses viral dsRNA and activates antiviral type I interferon (IFN) signaling. In SGMRT1, IFIH1 mutations confer a gain-of-function which causes overactivation of type I interferon (IFN) signaling leading to autoinflammation.Case presentationWe report the case of a nine year old child who initially presented with a slowly progressive decline of gross motor skill development and muscular weakness. At the age of five years, he developed osteoporosis, acro-osteolysis, alveolar bone loss and severe psoriasis. Whole exome sequencing revealed a pathogenic de novo IFIH1 mutation, confirming the diagnosis of SGMRT1. Consistent with constitutive type I interferon activation, patient blood cells exhibited a strong IFN signature as shown by marked up-regulation of IFN-stimulated genes. The patient was started on the Janus kinase (JAK) inhibitor, ruxolitinib, which inhibits signaling at the IFN-α/β receptor. Within days of treatment, psoriatic skin lesions resolved completely and the IFN signature normalized. Therapeutic efficacy was sustained and over the course muscular weakness, osteopenia and growth also improved.ConclusionsJAK inhibition represents a valuable therapeutic option for patients with SGMRT1. Our findings also highlight the potential of a patient-tailored therapeutic approach based on pathogenetic insight.

Impaired Antiviral Responses to Extracellular Double-Stranded RNA and Cytosolic DNA, but Not to Interferon-α Stimulation, in TRIM56-Deficient Cells.

The physiologic function of tripartite motif protein 56 (TRIM56), a ubiquitously expressed E3 ligase classified within the large TRIM protein family, remains elusive. Gene knockdown studies have suggested TRIM56 as a positive regulator of the type I interferon (IFN-I) antiviral response elicited via the Toll-like receptor 3 (TLR3) and cyclic GMP–AMP synthase (cGAS)–stimulator of interferon genes (STING) pathways, which detect and respond to danger signals—extracellular double-stranded (ds) RNA and cytosolic dsDNA, respectively. However, to what extent these pathways depend on TRIM56 in human cells is unclear. In addition, it is debatable whether TRIM56 plays a part in controlling the expression of IFN-stimulated genes (ISGs) resulting from IFN-I based antiviral treatment. In this study, we created HeLa-derived TRIM56 null cell lines by gene editing and used these cell models to comprehensively examine the impact of endogenous TRIM56 on innate antiviral responses. Our results showed that TRIM56 knockout severely undermined the upregulation of ISGs by extracellular dsRNA and that loss of TRIM56 weakened the response to cytosolic dsDNA. ISG induction and ISGylation following IFN-α stimulation, however, were not compromised by TRIM56 deletion. Using a vesicular stomatitis virus-based antiviral bioactivity assay, we demonstrated that IFN-α could efficiently establish an antiviral state in TRIM56 null cells, providing direct evidence that TRIM56 is not required for the general antiviral action of IFN-I. Altogether, these data ascertain the contributions of TRIM56 to TLR3- and cGAS–STING-dependent antiviral pathways in HeLa cells and add to our understanding of the roles this protein plays in innate immunity.