Abstract Natural killer (NK) cells are critical innate immune cells involved in the clearance of virally infected and malignant cells. Human NK cells are distinguished by their surface expression of CD56 and a lack of CD3. While CD56 expression and cell surface density has long been used as the prototypic marker to characterize primary human NK cell functional subsets, the exact functional role of CD56 in primary human NK cells is still not fully understood. Here, we eliminated the expression of CD56 in human ex vivo expanded NK cells (CD56bright) using CRISPR/Cas9 in order to assess the function of CD56 in this highly activated and cytotoxic NK cell population. We show that the expression of CD56 has no effect on NK cell proliferative capacity or expression of various activation and inhibitory markers. Further, CD56 does not contribute to NK cell–mediated cytotoxicity, inflammatory cytokine production, or the ability of NK cells to control tumor engraftment in vivo. We also found that while deletion of CD56 did not impact NK cell glycolytic metabolism, it did increase NK cell reliance on oxidative phosphorylation. Last, CD56 does not alter expanded NK cell in vivo tissue trafficking. Our results indicate that while CD56 expression could be used to indicate a hyperfunctional state of NK cells, it does not directly influence the antitumor functions of expanded NK cells.
Mouse models of Alzheimer's disease (AD) show progression through stages reflective of human pathology. Proteomics identification of temporal and sex-linked factors driving AD-related pathways can be used to dissect initiating and propagating events of AD stages to develop biomarkers or design interventions. In the present study, we conducted label-free proteome measurements of mouse hippocampus tissue with variables of time (3, 6, and 9 months), genetic background (5XFAD versus WT), and sex (equal males and females). These time points are associated with well-defined phenotypes with respect to the following: Aβ42 plaque deposition, memory deficits, and neuronal loss, allowing correlation of proteome-based molecular signatures with the mouse model stages. Our data show 5XFAD mice exhibit increases in known human AD biomarkers as amyloid-beta peptide, APOE, GFAP, and ITM2B are upregulated across all time points/stages. At the same time, 23 proteins are here newly associated with Alzheimer's pathology as they are also dysregulated in 5XFAD mice. At a pathways level, the 5XFAD-specific upregulated proteins are significantly enriched for DNA damage and stress-induced senescence at 3-month only, while at 6-month, the AD-specific proteome signature is altered and significantly enriched for membrane trafficking and vesicle-mediated transport protein annotations. By 9-month, AD-specific dysregulation is also characterized by significant neuroinflammation with innate immune system, platelet activation, and hyper-reactive astrocyte-related enrichments. Aside from these temporal changes, analysis of sex-linked differences in proteome signatures uncovered novel sex and AD-associated proteins. Pathway analysis revealed sex-linked differences in the 5XFAD model to be involved in the regulation of well-known human AD-related processes of amyloid fibril formation, wound healing, lysosome biogenesis, and DNA damage. Verification of the discovery results by Western blot and parallel reaction monitoring confirm the fundamental conclusions of the study and poise the 5XFAD model for further use as a molecular tool for understanding AD.
Abstract Regulation of immune responses during viral infection is critical to preventing the development immunopathology that impairs host survival. Natural killer (NK) cells are well known for their antiviral functions that promote viral clearance; however, their roles in limiting immune-mediated pathology are still unclear. Using a mouse model for genital herpes simplex virus type 2 infection, we find that NK cell-derived interferon-γ directly counteracts interleukin-6-mediated matrix metalloproteases (MMPs) activity in macrophages to limit MMP-mediated tissue damage. Our findings uncover a key immunoregulatory function of NK cells during host-pathogen interactions that highlight the potential of NK cell therapy for treatment of severe viral infections.
SummaryBackgroundThe use of cannabis during pregnancy is rising following its widespread legalization. Cannabidiol (CBD) is gaining popularity due to the public perception that it is safer than the psychoactive cannabis component Δ9-tetrahydrocannabinol (THC). However, while evidence underpins the harm of THC and cannabis smoke on fetal development, there is minimal research on the safety of CBD and oral cannabis. The current study aims to decipher the safety of oral CBD and THC use during pregnancy.MethodsUsing a mouse model, we directly compared the effects of oral CBD and THC oil exposure (20 mg/kg body weight) from early to mid-gestation on implantation site remodelling and fetal growth. We examined offspring behaviour and metabolic activity using both traditional and automated cage systems. Lastly, using human and mouse immune cells we assessed how CBD and THC influence angiogenic factor production.FindingsWe observed impaired maternal spiral artery remodelling in cannabis exposed mice and found that CBD and THC disrupt immune cell angiogenic factor production. Oral consumption of THC or CBD oil also resulted in significant fetal growth impairment and led to long-lasting sex-dependent consequences as male offspring exhibited altered aggression and metabolic activity while females had impaired spatial learning.InterpretationOur results show that oral consumption of either CBD or THC oil during pregnancy in mice results in harm to the developing fetus and causes behavioural changes after birth.FundingThe Michael G. DeGroote Centre for Medicinal Cancer Research, the Canadian Institutes of Health Research, and the Canadian Foundation for Innovation.
Herpes simplex virus type 2 (HSV-2) infection is one of the most prevalent sexually transmitted infections that disproportionately impacts women worldwide. Currently, there are no vaccines or curative treatments, resulting in life-long infection. The mucosal environment of the female reproductive tract (FRT) is home to a complex array of local immune defenses that must be carefully coordinated to protect against genital HSV-2 infection, while preventing excessive inflammation to prevent disease symptoms. Crucial to the defense against HSV-2 infection in the FRT are three classes of highly related and integrated cytokines, type I, II, and III interferons (IFN). These three classes of cytokines control HSV-2 infection and reduce tissue damage through a combination of directly inhibiting viral replication, as well as regulating the function of resident immune cells. In this review, we will examine how interferons are induced and their critical role in how they shape the local immune response to HSV-2 infection in the FRT.
ABSTRACT Elevated replication stress is evident at telomeres of about 10-15% of cancer cells, which maintain their telomeres via a homologous recombination (HR)-based mechanism, referred to as alternative lengthening of telomeres (ALT). How ALT cells resolve replication stress to support their growth remains incompletely characterized. Here, we report that CSB (also known as ERCC6) promotes recruitment of HR repair proteins (MRN, BRCA1, BLM and RPA32) and POLD3 to ALT telomeres, a process that requires the ATPase activity of CSB and is controlled by ATM- and CDK2-dependent phosphorylation. Loss of CSB stimulates telomeric recruitment of MUS81 and SLX4, components of the structure-specific MUS81-EME1-SLX1-SLX4 (MUS-SLX) endonuclease complex, suggesting that CSB restricts MUS-SLX-mediated processing of stalled forks at ALT telomeres. Loss of CSB coupled with depletion of SMARCAL1, a chromatin remodeler implicated in catalyzing regression of stalled forks, synergistically promotes not only telomeric recruitment of MUS81 but also the formation of fragile telomeres, the latter of which is reported to arise from fork stalling. These results altogether suggest that CSB-mediated HR repair and SMARCAL1-mediated fork regression cooperate to prevent stalled forks from being processed into fragile telomeres in ALT cells.
As highlighted by the COVID-19 global pandemic, elderly individuals comprise the majority of cases of severe viral infection outcomes and death. A combined inability to control viral replication and exacerbated inflammatory immune activation in elderly patients causes irreparable immune-mediated tissue pathology in response to infection. Key to these responses are type I, II, and III interferons (IFNs), which are involved in inducing an antiviral response, as well as controlling and suppressing inflammation and immunopathology. IFNs support monocyte/macrophage-stimulated immune responses that clear infection and promote their immunosuppressive functions that prevent excess inflammation and immune-mediated pathology. The timing and magnitude of IFN responses to infection are critical towards their immunoregulatory functions and ability to prevent immunopathology. Aging is associated with multiple defects in the ability of macrophages and dendritic cells to produce IFNs in response to viral infection, leading to a dysregulation of inflammatory immune responses. Understanding the implications of aging on IFN-regulated inflammation will give critical insights on how to treat and prevent severe infection in vulnerable individuals. In this review, we describe the causes of impaired IFN production in aging, and the evidence to suggest that these impairments impact the regulation of the innate and adaptive immune response to infection, thereby causing disease pathology.
Abstract Although many viral infections are linked to the development of neurological disorders, the mechanism governing virus-induced neuropathology remains poorly understood, particularly when the virus is not directly neuropathic. Using a mouse model of Zika virus (ZIKV) infection, we found that the severity of neurological disease did not correlate with brain ZIKV titers, but rather with infiltration of bystander activated NKG2D + CD8 + T cells. Antibody depletion of CD8 or blockade of NKG2D prevented ZIKV-associated paralysis, suggesting that CD8 + T cells induce neurological disease independent of TCR signaling. Furthermore, spleen and brain CD8 + T cells exhibited antigen-independent cytotoxicity that correlated with NKG2D expression. Finally, viral infection and inflammation in the brain was necessary but not sufficient to induce neurological damage. We demonstrate that CD8 + T cells mediate virus-induced neuropathology via antigen-independent, NKG2D-mediated cytotoxicity, which may serve as a therapeutic target for treatment of virus-induced neurological disease.
