Functional Comparison of Interferon-α Subtypes Reveals Potent Hepatitis B Virus Suppression by a Concerted Action of Interferon-α and Interferon-γ Signaling.

BACKGROUND AND AIMS: Interferon (IFN)-α, composed of numerous subtypes, plays a crucial role in immune defense. As the most studied subtype, IFN-α2 has been used for treating chronic hepatitis B virus (HBV) infection, with advantages of finite treatment duration and sustained virologic response, but its efficacy remains relatively low. This study aimed to screen for IFN-α subtypes with the highest anti-HBV potency and to characterize mechanisms of IFN-α-mediated HBV restriction. APPROACH AND RESULTS: Using cell culture-based HBV infection systems and a human-liver chimeric mouse model, IFN-α subtype-mediated antiviral response and signaling activation were comprehensively analyzed. IFN-α14 was identified as the most effective subtype in suppression of HBV covalently closed circular DNA transcription and HBV e antigen/HBV surface antigen production, with median inhibitory concentration values approximately 100-fold lower than those of the conventional IFN-α2. IFN-α14 alone elicited IFN-α and IFN-γ signaling crosstalk in a manner similar to the combined use of IFN-α2 and IFN-γ, inducing multiple potent antiviral effectors, which synergistically restricted HBV replication. Guanylate binding protein 5, one of the most differentially expressed genes between IFN-α14-treated and IFN-α2-treated liver cells, was identified as an HBV restriction factor. A strong IFN-α-IFN-α receptor subunit 1 interaction determines the anti-HBV activity of IFN-α. The in vivo anti-HBV activity of IFN-α14 and treatment-related transcriptional patterns were further confirmed, and few adverse effects were observed. CONCLUSIONS: A concerted IFN-α and IFN-γ response in liver, which could be efficiently elicited by IFN-α subtype 14, is associated with potent HBV suppression. These data deepen the understanding of the divergent activities of IFN-α subtypes and the mechanism underlying the synergism between IFN-α and IFN-γ signaling, with implications for improved IFN therapy and HBV curative strategies.

Cytotoxic Nitrogenous Terpenoids from Two South China Sea Nudibranchs Phyllidiella pustulosa, Phyllidia coelestis, and Their Sponge-Prey Acanthella cavernosa.

A detailed chemical investigation of two South China Sea nudibranchs Phyllidiella pustulosa and Phyllidia coelestis, as well as their possible sponge-prey Acanthella cavernosa, led to the isolation of one new nitrogenous cadinane-type sesquiterpenoid xidaoisocyanate A (1), one new naturally occurring nitrogen-containing kalihinane-type diterpenoid bisformamidokalihinol A (16), along with 17 known nitrogenous terpenoids (2⁻15, 17⁻19). The structures of all the isolates were elucidated by detailed spectroscopic analysis and by the comparison of their spectroscopic data with those reported in the literature. In addition, the absolute stereochemistry of the previously reported axiriabiline A (5) was determined by X-ray diffraction (XRD) analysis. In a bioassay, the bisabolane-type sesquiterpenoids 8, 10, and 11 exhibited cytotoxicity against several human cancer cell lines.

Identification of Retinoic Acid Receptor Agonists as Potent Hepatitis B Virus Inhibitors via a Drug Repurposing Screen.

Currently available therapies for chronic hepatitis B virus (HBV) infection can efficiently reduce viremia but induce hepatitis B surface antigen (HBsAg) loss in very few patients; also, these therapies do not greatly affect the viral covalently closed circular DNA (cccDNA). To discover new agents with complementary anti-HBV effects, we performed a drug repurposing screen of 1,018 Food and Drug Administration (FDA)-approved compounds using HBV-infected primary human hepatocytes (PHH). Several compounds belonging to the family of retinoic acid receptor (RAR) agonists were identified that reduced HBsAg levels in a dose-dependent manner without significant cytotoxicity. Among them, tazarotene exhibited the most potent anti-HBV effect, with a half-maximal inhibitory concentration (IC50) for HBsAg of less than 30 nM in PHH. The inhibitory effect was also observed in HBV-infected differentiated HepaRG (dHepaRG) models, but not in HepG2.215 cells, and HBV genotypes A to D were similarly inhibited. Tazarotene was further demonstrated to repress HBV cccDNA transcription, as determined by the levels of HBV cccDNA and RNAs and the activation of HBV promoters. Moreover, RNA sequence analysis showed that tazarotene did not induce an interferon response but altered the expression of a number of genes associated with RAR and metabolic pathways. Inhibition of RARß, but not RARα, by a specific antagonist significantly attenuated the anti-HBV activity of tazarotene, suggesting that tazarotene inhibits HBV in part through RARß. Finally, a synergistic effect of tazarotene and entecavir on HBV DNA levels was observed. Therefore, RAR agonists as represented by tazarotene were identified as potential novel anti-HBV agents.

[Roles of astrocytes in cerebral infarction and related therapeutic strategies].

Astrocytes are the most abundant cells in the central nervous system and play significant roles in normal brain. With cerebral infarction, astrocytes are activated as reactive astrocytes and form glial scars, which play an essential part in brain injury. According to their roles in neuroprotection after cerebral infarction, regulation of scar formation, nerve regeneration, maintenance of blood-brain barrier, promotion of angiogenesis and immune response, scholars have proposed a variety of therapeutic strategies based on targeting astrocytes. This article reviews the research progress on the changes in astrocyte signaling pathways before and after cerebral infarction and the related therapeutic strategies.

An extracellular humanized IFNAR immunocompetent mouse model for analyses of human interferon alpha and subtypes.

Type I interferons (IFN-Is) have key roles in immune defense and treatments for various diseases, including chronic hepatitis B virus (HBV) infection. All IFN-Is signal through a shared IFN-I heterodimeric receptor complex comprising IFN-α receptor 1 (IFNAR1) and IFNAR2 subunits, but differences in antiviral and immunomodulatory responses among IFN-I subtypes remain largely unknown. Because the IFN-IFNAR interactions are species-specific, mice exhibit weak responses to human IFN-I. To more fully characterize the actions of human IFN-α and its subtypes in vivo, a gene targeting strategy was employed to generate gene knock-in mice with extracellular-humanized IFNAR1/2 (IFNAR-hEC) in the C57BL/6N strain. IFNAR-hEC mice actively responded to human IFN-I, and endogenous mouse IFN-I signalling remained active in heterozygous mice (IfnarhEC/+). Analyses of IFNAR-hEC mice and isolated cells showed that human IFN-α2 and α14 subtypes exerted differential effect on the activation of JAK-STAT signalling and immune responses. Compared with IFN-α2, IFN-α14 induced greater activation of STAT1/2 and IFN-stimulated genes, synergistically elicited IFN-α and -γ signalling, and induced higher numbers of antigen-specific CD8+ T cells. Moreover, IFNAR-hEC mice with HBV replication displayed long-term viral suppression upon treatment with the clinically-used PEGylated hIFN-α2. These results indicate that IFNAR-hEC mice may be useful for elucidating antiviral and immunomodulatory functions of human IFN-Is and for conducting preclinical studies. A better understanding of the distinct activities of IFN-α subtypes can provide insights concerning the development of improved IFN-based therapy.

Genome-Wide Extrachromosomal Circular DNA Profiling of Paired Hepatocellular Carcinoma and Adjacent Liver Tissues.

Hepatocellular carcinoma (HCC) develops through multiple mechanisms. While recent studies have shown the presence of extrachromosomal circular DNA (eccDNA) in most cancer types, the eccDNA expression pattern and its association with HCC remain obscure. We aimed to investigate this problem. The genome-wide eccDNA profiles of eight paired HCC and adjacent non-tumor tissue samples were comprehensively elucidated based on Circle-seq, and they were further cross-analyzed with the RNA sequencing data to determine the association between eccDNA expression and transcriptome dysregulation. A total of 60,423 unique eccDNA types were identified. Most of the detected eccDNAs were smaller than 1 kb, with a length up to 182,363 bp and a mean sizes of 674 bp (non-tumor) and 813 bp (tumor), showing a greater association with gene-rich rather than with gene-poor regions. Although there was no statistical difference in length and chromosome distribution, the eccDNA patterns between HCC and adjacent non-tumor tissues showed significant differences at both the chromosomal and single gene levels. Five of the eight HCC tissues showed significantly higher amounts of chromosome 22-derived eccDNA expression compared to the non-tumor tissue. Furthermore, two genes, SLC16A3 and BAIAP2L2, with a higher transcription level in tumor tissues, were related to eccDNAs exclusively detected in three HCC samples and were negatively associated with survival rates in HCC cohorts from public databases. These results indicate the existence and massive heterogeneity of eccDNAs in HCC and adjacent liver tissues, and suggest their potential association with dysregulated gene expression.

Long-Term Hepatitis B Virus Infection Induces Cytopathic Effects in Primary Human Hepatocytes, and Can Be Partially Reversed by Antiviral Therapy.

Chronic infection of hepatitis B virus (HBV) remains a major health burden worldwide. While the immune response has been recognized to play crucial roles in HBV pathogenesis, the direct cytopathic effects of HBV infection and replication on host hepatocytes and the HBV-host interactions are only partially defined due to limited culture systems. Here, based on our recently developed 5 chemical-cultured primary human hepatocytes (5C-PHHs) model that supports long-term HBV infection, we performed multiplexed quantitative analysis of temporal changes of host proteome and transcriptome on PHHs infected by HBV for up to 4 weeks. We showed that metabolic-, complement-, cytoskeleton-, mitochondrial-, and oxidation-related pathways were modulated at transcriptional or posttranscriptional levels during long-term HBV infection, which led to cytopathic effects and could be partially rescued by early, rather than late, nucleot(s)ide analog (NA) administration and could be significantly relieved by blocking viral antigens with RNA interference (RNAi). Overexpression screening of the dysregulated proteins identified a series of host factors that may contribute to pro- or anti-HBV responses of the infected hepatocytes. In conclusion, our results suggest that long-term HBV infection in primary human hepatocytes leads to cytopathic effects through remodeling the proteome and transcriptome and early antiviral treatment may reduce the extent of such effects, indicating a role of virological factors in HBV pathogenesis and a potential benefit of early administration of antiviral treatment. IMPORTANCE Global temporal quantitative proteomic and transcriptomic analysis using long-term hepatitis B virus (HBV)-infected primary human hepatocytes uncovered extensive remodeling of the host proteome and transcriptome and revealed cytopathic effects of long-term viral replication. Metabolic-, complement-, cytoskeleton-, mitochondrial-, and oxidation-related pathways were modulated at transcriptional or posttranscriptional levels, which could be partially rescued by early, rather than late, NA therapy and could be relieved by blocking viral antigens with RNAi. Overexpression screening identified a series of pro- or anti-HBV host factors. These data have deepened the understanding of the mechanisms of viral pathogenesis and HBV-host interactions in hepatocytes, with implications for therapeutic intervention.

Interferon and Hepatitis B: Current and Future Perspectives.

Chronic hepatitis B virus (HBV) infection remains a major health burden worldwide for which there is still no effective curative treatment. Interferon (IFN) consists of a group of cytokines with antiviral activity and immunoregulatory and antitumor effects, that play crucial roles in both innate and adaptive immune responses. IFN-α and its pegylated form have been used for over thirty years to treat chronic hepatitis B (CHB) with advantages of finite treatment duration and sustained virologic response, however, the efficacy is limited and side effects are common. Here, we summarize the status and unique advantages of IFN therapy against CHB, review the mechanisms of IFN-α action and factors affecting IFN response, and discuss the possible improvement of IFN-based therapy and the rationale of combinations with other antiviral agents in seeking an HBV cure.