Intermittent low-dose far-UVC irradiation inhibits growth of common mold below threshold limit value.

Mold infestations in buildings pose significant challenges to human health, affecting both private residences and hospitals. While molds commonly trigger asthma and allergies in the immunocompetent, they can cause life-threatening diseases in the immunocompromised. Currently, there is an unmet need for new strategies to reduce or prevent mold infestations. Far-UVC technology can inactivate microorganisms while remaining safe for humans. This study investigates the inhibitory efficacy of far-UVC light at 222 nm on the growth of common mold-producing fungi, specifically Penicillium candidum, when delivered in low-dose on-off duty cycles, a configuration consistent with its use in real-world settings. The inhibitory effect of the low-dose duty cycles was assessed on growth induced by i) an adjacent spore-producing P. candidum donor and ii) P. candidum spores seeded directly onto agar plates. In both setups, the far-UVC light significantly inhibited both vertical and horizontal growth of P. candidum, even when the UV doses were below the Threshold Value Limit of 23 mJ/cm2. These results suggest that far-UVC light holds the potential to improve indoor air quality by reducing or preventing mold growth, also when people are present.

Dual intron-targeted CRISPR-Cas9-mediated disruption of the AML RUNX1-RUNX1T1 fusion gene effectively inhibits proliferation and decreases tumor volume in vitro and in vivo.

Oncogenic fusion drivers are common in hematological cancers and are thus relevant targets of future CRISPR-Cas9-based treatment strategies. However, breakpoint-location variation in patients pose a challenge to traditional breakpoint-targeting CRISPR-Cas9-mediated disruption strategies. Here we present a new dual intron-targeting CRISPR-Cas9 treatment strategy, for targeting t(8;21) found in 5-10% of de novo acute myeloid leukemia (AML), which efficiently disrupts fusion genes without prior identification of breakpoint location. We show in vitro growth rate and proliferation reduction by 69 and 94% in AML t(8;21) Kasumi-1 cells, following dual intron-targeted disruption of RUNX1-RUNX1T1 compared to a non t(8;21) AML control. Furthermore, mice injected with RUNX1-RUNX1T1-disrupted Kasumi-1 cells had in vivo tumor growth reduction by 69 and 91% compared to controls. Demonstrating the feasibility of RUNX1-RUNX1T1 disruption, these findings were substantiated in isolated primary cells from a patient diagnosed with AML t(8;21). In conclusion, we demonstrate proof-of-principle of a dual intron-targeting CRISPR-Cas9 treatment strategy in AML t(8;21) without need for precise knowledge of the breakpoint location.

Plasmacytoid dendritic cells during COVID-19: Ally or adversary?

Plasmacytoid dendritic cells (pDCs) are specialized cells of the immune system that are thought to be the main cellular source of type I interferon alpha (IFNα) in response to viral infections. IFNs are powerful antivirals, whereas defects in their function or induction lead to impaired resistance to virus infections, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19. IFN production needs to be controlled, because sustained IFN production can also have detrimental effects on disease outcome. As such, pDCs are likely important for acute antiviral protection against SARS-CoV-2 infection but could potentially also contribute to chronic IFN levels. Here, we provide a historical overview of pDC biology and summarize existing literature addressing their involvement and importance during viral infections of the airways. Furthermore, we outline recent reports focused on the potential role of pDCs during SARS-CoV-2 infection, as well as the potential for this cellular subset to impact COVID-19 disease outcome.

Influenza A induces lactate formation to inhibit type I IFN in primary human airway epithelium.

Pathogenic viruses induce metabolic changes in host cells to secure the availability of biomolecules and energy to propagate. Influenza A virus (IAV) and severe acute respiratory syndrome corona virus 2 (SARS-CoV-2) both infect the human airway epithelium and are important human pathogens. The metabolic changes induced by these viruses in a physiologically relevant human model and how this affects innate immune responses to limit viral propagation are not well known. Using an ex vivo model of pseudostratified primary human airway epithelium, we here demonstrate that infection with both IAV and SARS-CoV-2 resulted in distinct metabolic changes including increases in lactate dehydrogenase A (LDHA) expression and LDHA-mediated lactate formation. Interestingly, LDHA regulated both basal and induced mitochondrial anti-viral signaling protein (MAVS)-dependent type I interferon (IFN) responses to promote IAV, but not SARS-CoV-2, replication. Our data demonstrate that LDHA and lactate promote IAV but not SARS-CoV-2 replication by inhibiting MAVS-dependent induction of type I IFN in primary human airway epithelium.

Cellular Metabolites Regulate Central Nucleic Acid Sensing Pathways.

Detection of pathogen-derived DNA or RNA species by cellular nucleic acid sensors prompts release of anti-microbial interferons and cytokines. In contrast to their protective anti-microbial functions, inappropriate or excessive activation of nucleic acid sensors can cause inflammatory diseases. Nucleic acid sensing is therefore tightly controlled by regulatory factors acting through both transcriptional and post-transcriptional mechanisms. Recently, it has become clearer that metabolic pathways-previously thought to be unconnected with immune responses-can influence nucleic acid sensing. This regulation can be observed when immune system cells undergo metabolic reprogramming in response to stimulation with pathogen-associated molecular patterns such as lipopolysaccharide from gram negative bacteria. Metabolic reprogramming leads to accumulation and secretion of metabolites, which have been mostly viewed as end-products of processes providing cellular energy and building blocks. However, metabolites have now been identified as important regulators of nucleic acid sensing. This mini-review aims to outline current knowledge on regulation of central nucleic acid sensing pathways by metabolites during metabolic reprogramming.

Virus-induced metabolic reprogramming and innate sensing hereof by the infected host.

To make new infectious particles, all viruses must manipulate host cell metabolism to secure sufficient availability of biomolecules and energy-a phenomenon now known as metabolic reprogramming. Numerous observations of this has already been made for a range of viruses with each type of virus seemingly applying its own unique tactics to accomplish this unifying goal. In this light, metabolic reprogramming of the infected cell is largely beneficial to the virus and not to the host. On the other hand, virus-induced metabolic reprogramming represents a transformed self with distorted cellular and extracellular levels of distinct metabolites and metabolic by-products. This review briefly outlines current knowledge of virus-induced metabolic reprogramming, discusses how this could be sensed by the infected host to initiate anti-viral programs, and presents examples of innate anti-viral mechanisms of the host that target the availability of biomolecules to block viral replication.

SARS-CoV-2 evades immune detection in alveolar macrophages.

Respiratory infections, like the current COVID-19 pandemic, target epithelial cells in the respiratory tract. Alveolar macrophages (AMs) are tissue-resident macrophages located within the lung. They play a key role in the early phases of an immune response to respiratory viruses. AMs are likely the first immune cells to encounter SARS-CoV-2 during an infection, and their reaction to the virus will have a profound impact on the outcome of the infection. Interferons (IFNs) are antiviral cytokines and among the first cytokines produced upon viral infection. In this study, AMs from non-infectious donors are challenged with SARS-CoV-2. We demonstrate that challenged AMs are incapable of sensing SARS-CoV-2 and of producing an IFN response in contrast to other respiratory viruses, like influenza A virus and Sendai virus, which trigger a robust IFN response. The absence of IFN production in AMs upon challenge with SARS-CoV-2 could explain the initial asymptotic phase observed during COVID-19 and argues against AMs being the sources of pro-inflammatory cytokines later during infection.

Nrf2 Negatively Regulates Type I Interferon Responses and Increases Susceptibility to Herpes Genital Infection in Mice.

Herpes simplex virus-2 (HSV-2) is a leading cause of sexually transmitted infections for which no effective vaccines or prophylactic treatment currently exist. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor involved in the detoxification of reactive oxygen species (ROS) and has been more recently shown to regulate inflammatory and antiviral responses. Here, we evaluated the importance of Nrf2 in the control of HSV-2 genital infection, and its role in the regulation of HSV-induced innate antiviral immunity. Comparison of antiviral gene expression profile by RNA-sequencing analysis of wild type and Nrf2-mutant (Nrf2AY/AY ) murine macrophages showed an upregulation at the basal level of the type I interferon-associated gene network. The same basal increased antiviral profile was also observed in the spleen of Nrf2-/- mice. Interestingly, the lack of Nrf2 in murine cells was sufficient to increase the responsiveness to HSV-derived dsDNA and protect cells from HSV-2 infection in vitro. Surprisingly, there was no indication of an alteration in STING expression in murine cells as previously reported in cells of human origin. Additionally, genetic activation of Nrf2 in Keap1-/- mouse embryonic fibroblasts increased HSV-2 infectivity and replication. Finally, using an in vivo vaginal herpes infection model, we showed that Nrf2 controlled early innate immune responses to HSV-2 without affecting STING expression levels. Nrf2-/- mice exhibited reduced viral replication that was associated with higher level of type I interferons in vaginal washes. Nrf2-/- mice also displayed reduced weight loss, lower disease scores, and higher survival rates than wild type animals. Collectively, these data identify Nrf2 as a negative regulator of the interferon-driven antiviral response to HSV-2 without impairing STING mRNA and protein expression levels in murine cells.

Vaginal HSV-2 Infection and Tissue Analysis.

The vaginal murine HSV-2 infection model is very useful in studying mucosal immunity against HSV-2 ( Overall et al., 1975 ; Renis et al., 1976 ; Parr and Parr, 2003). Histologically, the vagina of Depo-Provera-treated mice is lined by a single layer of mucus secreting columnar epithelial cells overlying two to three layers of proliferative cells. Even though this is morphologically different from the human vagina, it closely resembles the endocervical epithelium, which is thought to be the primary site of HSV-2 infection in women ( Parr et al., 1994 ; Kaushic et al., 2011). In the protocol presented here, mice are pre-treated with Depo-Provera before intra-vaginal inoculation with HSV-2. The virus replicates in the mucosal epithelium from where it spreads to and replicates in the CNS including the spinal cord, brain stem, cerebrum and cerebellum. Cytokine responses can be detected in vaginal washings using ELISA or in vaginal tissues using qPCR. Further, the recruitment of leukocytes to the vagina can be determined by flow cytometry. The model is suitable for research of both innate and adaptive immunity to HSV-2 infection.

Overexpression of microRNA-155 increases IL-21 mediated STAT3 signaling and IL-21 production in systemic lupus erythematosus.

INTRODUCTION: Interleukin (IL)-21 is a key cytokine in autoimmune diseases such as systemic lupus erythematosus (SLE) by its regulation of autoantibody production and inflammatory responses. The objective of this study is to investigate the signaling capacity of IL-21 in T and B cells and assess its possible regulation by microRNA (miR)-155 and its target gene suppressor of cytokine signaling 1 (SOCS1) in SLE. METHODS: The signaling capacity of IL-21 was quantified by stimulating peripheral blood mononuclear cells (PBMCs) with IL-21 and measuring phosphorylation of STAT3 (pSTAT3) in CD4+ T cells, B cells, and natural killer cells. Induction of miR-155 by IL-21 was investigated by stimulating purified CD4+ T cells with IL-21 and measuring miR-155 expression levels. The functional role of miR-155 was assessed by overexpressing miR-155 in PBMCs from SLE patients and healthy controls (HCs) and measuring its effects on STAT3 and IL-21 production in CD4+ and CD8+ T cells. RESULTS: Induction of pSTAT3 in CD4+ T cells in response to IL-21 was significantly decreased in SLE patients compared to HCs (p < 0.0001). Further, expression levels of miR-155 were significantly decreased and SOCS1 correspondingly increased in CD4+ T cells from SLE patients. Finally, overexpression of miR-155 in CD4+ T cells increased STAT3 phosphorylation in response to IL-21 treatment (p < 0.01) and differentially increased IL-21 production in SLE patients compared to HCs (p < 0.01). CONCLUSION: We demonstrate that SLE patients have reduced IL-21 signaling capacity, decreased miR-155 levels, and increased SOCS1 levels compared to HCs. The reduced IL-21 signaling in SLE could be rescued by overexpression of miR-155, suggesting an important role for miR-155 in the reduced IL-21 signaling observed in SLE.

NKT cell activation by local α-galactosylceramide administration decreases susceptibility to HSV-2 infection.

NKT cells are a subgroup of T cells, which express a restricted TCR repertoire and are critical for the innate immune responses to viral infections. Activation of NKT cells depends on the major histocompatibility complex-related molecule CD1d, which presents bioactive lipids to NKT cells. The marine sponge derived lipid αGalCer has recently been demonstrated as a specific agonist for activation of human and murine NKT cells. In the present study we investigated the applicability of αGalCer pre-treatment for immune protection against intra-vaginal HSV-2 infection. We found that C57BL/6 WT mice that received local pre-treatment with αGalCer prior to intra-vaginal HSV-2 infection had a lower mean disease score, mortality and viral load in the vagina following infection, compared to mice that did not receive αGalCer pre-treatment. Further, we found increased numbers of CD45 and NK1.1 positive cells in vaginal tissue and elevated levels of IFN-γ in the vaginal tissue and in vaginal fluids 24h after αGalCer pre-treatment. Collectively our data demonstrate a protective effect of αGalCer induced activation of NKT cells in the innate immune protection against viral infection.

Increased plasma levels of IL-21 and IL-23 in spondyloarthritis are not associated with clinical and MRI findings.

We have investigated the role of the Th17-related cytokines interleukin-17A (IL-17A), IL-21, and IL-23 in spondyloarthritis (SpA) by examining their association with disease activity and magnetic resonance imaging (MRI) findings in patients with SpA (n = 80). Furthermore, to investigate the cellular origins of the cytokines, paired mononuclear cells from blood and synovial fluid were examined for the expression of IL-17A, IL-21, and IL-23R using multicolor flow cytometry. Both IL-21 and IL-23 levels were increased in plasma from SpA patients compared with healthy volunteers (P < 0.05), whereas IL-17A was not. A significant correlation was observed between individual levels of IL-21 and IL-23 (r = 0.7, P < 0.001). No association between individual levels of IL-17A, IL-21, and IL-23 with C-reactive protein (CRP), MRI changes, and clinical scoring (BASMI, BASFI, and BASDAI) were observed. The frequency of CD4+CD45RO+ T cells expressing IL-21 and IL-23R was increased in the inflamed SpA joint compared to peripheral blood (P < 0.05). This study demonstrate that the plasma levels of the Th17-related cytokines IL-21 and IL-23, but not IL-17A, are increased in SpA patients, but we did not find evidence that the level of these cytokines reflect disease activity in SpA.

Increased interleukin 21 (IL-21) and IL-23 are associated with increased disease activity and with radiographic status in patients with early rheumatoid arthritis.

OBJECTIVE: To investigate the levels of the T helper (Th)17-related cytokines interleukin 17A (IL-17A), IL-21, and IL-23 and their association with disease activity in rheumatoid arthritis (RA). METHODS: In a longitudinal sample set from patients with early RA (< 6 months; n = 40), we measured the plasma cytokine levels of IL-17A, IL-21, and IL-23 and analyzed for correlation with disease activity in 28 joints (Disease Activity Score 28-joint count; DAS28), C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), and total Sharp score (TSS). In a transverse sample set of patients with chronic RA (> 8 years), using paired peripheral blood mononuclear cells and synovial fluid mononuclear cells, we investigated the cellular expression of IL-17A, IL-21, and IL-23R. RESULTS: Patients with early-stage RA had significantly increased plasma levels of IL-21 and IL-23, but not IL-17A, compared to patients with chronic RA and healthy volunteer controls. Plasma levels of IL-21 and IL-23 after 12 months of treatment correlated with DAS28 and ESR, but not to TSS. Changes in IL-23 plasma levels from time of diagnosis to 12 months correlated with change in DAS28 and with TSS scores at 2 years. The numbers of CD4+ T cells producing IL-21 were significantly increased in the synovial fluid of patients with chronic RA, with only marginal coexpression of IL-21 and IL-17A. CONCLUSION: Our results show a significant association between plasma levels of IL-21 and IL-23 and disease activity in RA, supporting the hypothesis that IL-21 and IL-23 are important pathogenic factors of this disease.