Dynamics of IL-15/IL-15R-α expression in response to HSV-1 infection reveal a novel mode of viral immune evasion counteracted by iNKT cells.

Herpes simplex virus type 1 (HSV-1) infects and persists in most of the human population. Interleukin-15 (IL-15) has an important role in the activation of cell-mediated immune responses and acts in complex with IL-15 receptor alpha (IL-15R-α) through cell surface transpresentation. Here, we have examined the IL-15/IL-15R-α complex response dynamics during HSV-1 infection in human keratinocytes. Surface expression of the IL-15/IL-15R-α complex rapidly increased in response to HSV-1, reaching a peak around 12 h after infection. This response was dependent on detection of viral replication by TLR3, and enhancement of IL15 and IL15RA gene expression. Beyond the peak of expression, levels of IL-15 and IL-15R-α gradually declined, reaching a profound loss of surface expression beyond 24 h of infection. This involved the loss of IL15 and IL15RA transcription. Interestingly, invariant natural killer T (iNKT) cells inhibited the viral interference with IL-15/IL-15R-α complex expression in an IFNγ-dependent manner. These results indicate that rapid upregulation of the IL-15/IL-15R-α complex occurs in HSV-1 infected keratinocytes, and that this response is targeted by viral interference. Shutdown of the IL-15 axis represents a novel mode of HSV-1 immune evasion, which can be inhibited by the host iNKT cell response.

NK cell frequencies, function and correlates to vaccine outcome in BNT162b2 mRNA anti-SARS-CoV-2 vaccinated healthy and immunocompromised individuals.

Adaptive immune responses have been studied extensively in the course of mRNA vaccination against COVID-19. Considerably fewer studies have assessed the effects on innate immune cells. Here, we characterized NK cells in healthy individuals and immunocompromised patients in the course of an anti-SARS-CoV-2 BNT162b2 mRNA prospective, open-label clinical vaccine trial. See trial registration description in notes. Results revealed preserved NK cell numbers, frequencies, subsets, phenotypes, and function as assessed through consecutive peripheral blood samplings at 0, 10, 21, and 35 days following vaccination. A positive correlation was observed between the frequency of NKG2C+ NK cells at baseline (Day 0) and anti-SARS-CoV-2 Ab titers following BNT162b2 mRNA vaccination at Day 35. The present results provide basic insights in regards to NK cells in the context of mRNA vaccination, and have relevance for future mRNA-based vaccinations against COVID-19, other viral infections, and cancer.Trial registration: The current study is based on clinical material from the COVAXID open-label, non-randomized prospective clinical trial registered at EudraCT and clinicaltrials.gov (no. 2021-000175-37). Description: https://clinicaltrials.gov/ct2/show/NCT04780659?term=2021-000175-37&draw=2&rank=1 .

Tissue-resident MAIT cell populations in human oral mucosa exhibit an activated profile and produce IL-17.

Mucosa-associated invariant T (MAIT) cells are unconventional T lymphocytes defined by their innate-like characteristics and broad antimicrobial responsiveness. Whether MAIT cells are part of the tissue-resident defense in the oral mucosal barrier is unknown. Here, we found MAIT cells present in the buccal mucosa, with a tendency to cluster near the basement membrane, and located in both epithelium and the underlying connective tissue. Overall MAIT cell levels were similar in the mucosa compared to peripheral blood, in contrast to conventional T cells that showed an altered representation of CD4+ and CD8+ subsets. The major mucosal MAIT cell subset displayed a tissue-resident and activated profile with high expression of CD69, CD103, HLA-DR, and PD-1, as well as a skewed subset distribution with higher representation of CD4- /CD8- double-negative cells and CD8αα+ cells. Interestingly, tissue-resident MAIT cells had a specialized polyfunctional response profile with higher IL-17 levels, as assessed by polyclonal stimulus and compared to tissue nonresident and circulating populations. Furthermore, resident buccal MAIT cells were low in perforin. Together, these data indicate that MAIT cells form a part of the oral mucosal T cell compartment, where they exhibit a tissue-resident-activated profile biased toward IL-17 production.

Clinical Microbial Identification of Severe Oral Infections by MALDI-TOF Mass Spectrometry in Stockholm County: an 11-Year (2010 to 2020) Epidemiological Investigation.

Growing evidence suggests that oral infections can modify the course of systemic diseases. To date, epidemiological data on microbial oral infections are scarce. Here, we performed a comprehensive analysis of the trend and microbial diversity in oral infection specimens referred for clinical microbiology analysis from 2010 to 2020. The microbes were isolated by culture and were identified via matrix-assisted laser desorption ionization-time of flight mass spectrometry technology (MALDI-TOF MS) throughout the study period. A total of 1,014 referred samples from dental clinics in Stockholm County with dentoalveolar abscesses and jaw osteomyelitis being the main reason were identified. Overall, the microbial composition was dominated by Firmicutes (51%), followed by Bacteroidetes (19%), Proteobacteria (12%), and Actinobacteria (5%). At the genus level, Streptococcus spp. (36%), Prevotella spp. (18%), and Staphylococcus spp. (11%) were among the most frequently reported. Interestingly, a strong increase in trend was noted for Streptococcus anginosus, Streptococcus mitis, Streptococcus sanguinis, Eikenella corrodens, Actinomyces spp., Aggregatibacter aphrophilus, Staphylococcus epidermidis, and Granulicatella adiacens during the study time (R = 0.66 to 0.89, P < 0.05), and a minor increase was noted for Enterococcus faecalis and Klebsiella spp., whereas steady levels were noted for most of the others. The present study shows the diversity of bacteria that have been involved in dental infections during the last decade in the capital of Sweden, as well as the emerging oral microbiota trend, with clear clinical implications on the oral-systemic link. IMPORTANCE Oral diseases and associated microbes are a risk factor for systemic diseases and can change the courses of these diseases. To date, epidemiological data on microbial oral infections are scarce, and longitudinal reports are lacking. We present for the first time the microbial composition of severe oral bacterial infections determined via the MALDI-TOF mass spectrometry technique in a comprehensive study between 2010 and 2020 (11 years) in Stockholm County. The trend and microbial diversity of oral infections were analyzed on referred clinical microbiological samples and were processed by standardized protocols. Trend increase was noted for Streptococcus anginosus, Streptococcus mitis, Streptococcus sanguinis, Eikenella corrodens, Actinomyces spp., Aggregatibacter aphrophilus, Staphylococcus epidermidis, Granulicatella adiacens, Enterococcus faecalis, and Klebsiella spp. Our results provide new insights into the diversity and trend of oral microbiota that were involved in serious oral infections over the past decade in the capital of Sweden and may influence the oral-systemic link.

Expansion of donor-unrestricted MAIT cells with enhanced cytolytic function suitable for TCR redirection.

Progress in our understanding of MR1-restricted mucosa-associated invariant T (MAIT) cells has raised interest in harnessing these cells for immunotherapy. The innate-like response characteristics, abundance in the blood, donor-unrestricted nature, and tropism for tissues make MAIT cells suitable candidates for adoptive cell transfer therapies. However, reliable methods and tools to utilize MAIT cells in such approaches are lacking. Here, we established methodology for efficient expansion of human MAIT cells in culture with high purity and yield, while preserving their functional response toward their natural ligand and increasing their cytotoxic potential. The cultured MAIT cells retained their effector memory characteristics without signs of terminal differentiation and expressed a more diverse set of chemokine receptors, potentially widening their already broad tissue tropism. To investigate the potential of MAIT cells in a context outside their main role in controlling bacterial infection, we engineered cultured MAIT cells with a new TCR specificity to mediate effective antiviral HLA class I-restricted effector function. In summary, we developed robust and effective methodology for the expansion of human MAIT cells with enhanced cytolytic capacity and for their engineering with a new specificity. These findings form a basis for the development of MAIT cells as a platform for adoptive immunotherapy.

Human MAIT cells endowed with HBV specificity are cytotoxic and migrate towards HBV-HCC while retaining antimicrobial functions.

BACKGROUND & AIMS: Virus-specific T cell dysfunction is a common feature of HBV-related hepatocellular carcinoma (HBV-HCC). Conventional T (ConT) cells can be redirected towards viral antigens in HBV-HCC when they express an HBV-specific receptor; however, their efficacy can be impaired by liver-specific physical and metabolic features. Mucosal-associated invariant T (MAIT) cells are the most abundant innate-like T cells in the liver and can elicit potent intrahepatic effector functions. Here, we engineered ConT and MAIT cells to kill HBV expressing hepatoma cells and compared their functional properties. METHODS: Donor-matched ConT and MAIT cells were engineered to express an HBV-specific T cell receptor (TCR). Cytotoxicity and hepatocyte homing potential were investigated using flow cytometry, real-time killing assays, and confocal microscopy in 2D and 3D HBV-HCC cell models. Major histocompatibility complex (MHC) class I-related molecule (MR1)-dependent and MR1-independent activation was evaluated in an Escherichia coli THP-1 cell model and by IL-12/IL-18 stimulation, respectively. RESULTS: HBV TCR-MAIT cells demonstrated polyfunctional properties (CD107a, interferon [IFN] γ, tumour necrosis factor [TNF], and IL-17A) with strong HBV target sensitivity and liver-homing chemokine receptor expression when compared with HBV TCR-ConT cells. TCR-mediated lysis of hepatoma cells was comparable between the cell types and augmented in the presence of inflammation. Coculturing with HBV+ target cells in a 3D microdevice mimicking aspects of the liver microenvironment demonstrated that TCR-MAIT cells migrate readily towards hepatoma targets. Expression of an ectopic TCR did not affect the ability of the MAIT cells to be activated via MR1-presented bacterial antigens or IL-12/IL-18 stimulation. CONCLUSIONS: HBV TCR-MAIT cells demonstrate anti-HBV functions without losing their endogenous antimicrobial mechanisms or hepatotropic features. Our results support future exploitations of MAIT cells for liver-directed immunotherapies. LAY SUMMARY: Chronic HBV infection is a leading cause of liver cancer. T cell receptor (TCR)-engineered T cells are patients' immune cells that have been modified to recognise virus-infected and/or cancer cells. Herein, we evaluated whether mucosal-associated invariant T cells, a large population of unconventional T cells in the liver, could recognise and kill HBV infected hepatocytes when engineered with an HBV-specific TCR. We show that their effector functions may exceed those of conventional T cells currently used in the clinic, including antimicrobial properties and chemokine receptor profiles better suited for targeting liver tumours.

In Situ Detection of MAIT Cells and MR1-Expressing Cells in Tissue Biopsies Utilizing Immunohistochemistry.

The mucosa-associated invariant T (MAIT) cells are innate-like T cells that recognize microbial vitamin B2 metabolites presented via MR1, a MHC-I-related protein. MAIT cells are abundant in blood and mucosa, where they display a broad range of functions. Spatial distribution of cells and their proximity to other cells, including infected cells and antigen presenting cells, are crucial components of cell-mediated immunity. Here we describe techniques to detect MAIT cells and MR1-expressing cells in situ, which enable the visualization, distribution, and localization of these cells within their histological context. We provide specific protocols and describe potential advantages and limitations for each of the presented methodologies for studying MAIT cells in human tissues.

Chronic Viral Liver Diseases: Approaching the Liver Using T Cell Receptor-Mediated Gene Technologies.

Chronic infection with viral hepatitis is a major risk factor for liver injury and hepatocellular carcinoma (HCC). One major contributing factor to the chronicity is the dysfunction of virus-specific T cell immunity. T cells engineered to express virus-specific T cell receptors (TCRs) may be a therapeutic option to improve host antiviral responses and have demonstrated clinical success against virus-associated tumours. This review aims to give an overview of TCRs identified from viral hepatitis research and discuss how translational lessons learned from cancer immunotherapy can be applied to the field. TCR isolation pipelines, liver homing signals, cell type options, as well as safety considerations will be discussed herein.

Circulating and Salivary Antibodies to Fusobacterium nucleatum Are Associated With Cystic Pancreatic Neoplasm Malignancy.

Objectives: Intraductal papillary mucinous neoplasms (IPMNs) are cystic precursor lesions to pancreatic cancer. The presence of oral microbes in pancreatic tissue or cyst fluid has been associated with high-grade dysplasia (HGD) and cancer. The present study aims at investigating if humoral immunity to pancreas-associated oral microbes reflects IPMN severity. Design: Paired plasma (n = 109) and saliva (n = 65) samples were obtained from IPMN pancreatic cystic tumor cases and controls, for anti-bacterial antibody analysis and DNA quantification by enzyme-linked immunosorbent assay (ELISA) and qPCR, respectively. Tumor severity was graded by histopathology, laboratory, and clinical data. Circulating plasma and salivary antibody reactivity to a pancreas-associated oral microbe panel were measured by ELISA and correlated to tumor severity. Results: The patient group with high-risk cystic tumors (HGD and/or associated invasive cancer) shows ample circulating IgG reactivity to Fusobacterium nucleatum (F. nucleatum) but not to Granulicatella adiacens (G. adiacens), which is independent of the salivary bacteria DNA levels. This group also shows higher salivary IgA reactivity to F. nucleatum, Fap2 of F. nucleatum, and Streptococcus gordonii (S. gordonii) compared to low-risk IPMN and controls. The salivary antibody reactivity to F. nucleatum and Fap2 are found to be highly correlated, and cross-competition assays further confirm that these antibodies appear cross-reactive. Conclusion: Our findings indicate that humoral reactivity against pancreas-associated oral microbes may reflect IPMN severity. These findings are beneficial for biomarker development.

Mucosal-associated invariant T cells and oral microbiome in persistent apical periodontitis.

Opportunistic bacteria in apical periodontitis (AP) may pose a risk for systemic dissemination. Mucosal-associated invariant T (MAIT) cells are innate-like T cells with a broad and potent antimicrobial activity important for gut mucosal integrity. It was recently shown that MAIT cells are present in the oral mucosal tissue, but the involvement of MAIT cells in AP is unknown. Here, comparison of surgically resected AP and gingival tissues demonstrated that AP tissues express significantly higher levels of Vα7.2-Jα33, Vα7.2-Jα20, Vα7.2-Jα12, Cα and tumour necrosis factor (TNF), interferon (IFN)-γ and interleukin (IL)-17A transcripts, resembling a MAIT cell signature. Moreover, in AP tissues the MR1-restricted MAIT cells positive for MR1-5-OP-RU tetramer staining appeared to be of similar levels as in peripheral blood but consisted mainly of CD4+ subset. Unlike gingival tissues, the AP microbiome was quantitatively impacted by factors like fistula and high patient age and had a prominent riboflavin-expressing bacterial feature. When merged in an integrated view, the examined immune and microbiome data in the sparse partial least squares discriminant analysis could identify bacterial relative abundances that negatively correlated with Vα7.2-Jα33, Cα, and IL-17A transcript expressions in AP, implying that MAIT cells could play a role in the local defence at the oral tissue barrier. In conclusion, we describe the presence of MAIT cells at the oral site where translocation of oral microbiota could take place. These findings have implications for understanding the immune sensing of polymicrobial-related oral diseases.