A positive cytokine/chemokine feedback loop establishes plasmacytoid dendritic cell-driven autoimmune pancreatitis in IgG4-related disease.

The pathogenesis of the murine model of autoimmune pancreatitis associated with IgG4-related disease (AIP/IgG4-RD) induced by administration of polyinosinic-polycytidylic acid, is incompletely understood. While it is known that murine and human AIP/IgG4-RD is driven by plasmacytoid dendritic cells (pDCs) producing IFN-α, the origin of these cells and their relation to effector T cells is not known. Here we show that murine AIP was initiated by TLR3-bearing conventional DCs in the uninflamed pancreas whose activation by TLR3 ligand (polyinosinic-polycytidylic acid) caused IFN-α, CXCL9, and CXCL10 secretion. This, in turn, induced pancreatic recruitment of CXCR3+ T cells and these T cells, via their secretion of CCL25, facilitated migration of pDCs bearing CCR9 into the pancreas. This established a feedback loop anchored by the now dominant pDC production of IFN-α and the continued CXCR3+ T cell facilitation of pDC migration. Remarkably, the interaction between CXCR3+ T cells and pDCs also existed at the functional levels since this interaction enhanced the production of CCL25 and IFN-α by CXCR3+ T cells and pDCs, respectively. Evidence presented here that a similar disease mechanism was present in human AIP/IgG4-RD creates new avenues of disease treatment.

Intratumoral delivery of a highly active form of XCL1 enhances antitumor CTL responses through recruitment of CXCL9-expressing conventional type-1 dendritic cells.

Conventional type 1 dendritic cells (cDC1s) play a crucial role in antitumor immunity through the induction and activation of tumor-specific CD8+ cytotoxic T cells (CTLs). The chemokine XCL1 is a major chemotactic factor for cDC1s and its receptor XCR1 is selectively expressed on cDC1s. Here, we investigated the effect of intratumoral delivery of a highly active form of murine XCL1 (mXCL1-V21C/A59C) on cDC1-mediated antitumor immunity using a hydrophilic gel patch. The hydrophilic gel patch containing mXCL1-V21C/A59C increased cDC1 accumulation in the tumor masses and promoted their migration to the regional lymph nodes, resulting in enhanced induction of tumor-specific CTLs. Tumor-infiltrating cDC1s not only expressed XCR1 but also produced CXCL9, a ligand for CXCR3 which is highly expressed on CTLs and NK cells. Consequently, CTLs and NK cells were increased in the tumor masses of mice treated with mXCL1-V21C/A59C, while immunosuppressive cells such as monocyte-derived suppressive cells and regulatory T cells were decreased. We also confirmed that anti-CXCL9 treatment decreased the tumor infiltration of CTLs. The intratumoral delivery of mXCL1-V21C/A59C significantly decreased tumor growth and prolonged survival in E.G7-OVA and B16-F10 tumor-bearing mice. Furthermore, the antitumor effect of mXCL1-V21CA59C was enhanced in combination with anti-programmed cell death protein 1 treatment. Finally, using The Cancer Genome Atlas database, we found that XCL1 expression was positively correlated with tumor-infiltrating cDC1s and a better prognosis in melanoma patients. Collectively, our findings provide a novel therapeutic approach to enhance tumor-specific CTL responses through the selective recruitment of CXCL9-expressing cDC1s into the tumor masses.

BCL6 fine-tunes long-term tumor control.

Sun et al. provide comprehensive evidence that the transcription factor BCL6 functions as a gatekeeper for CD8+ progenitor cell function in tumors and prevents their excessive terminal differentiation, thereby preserving this stem-like population for long-term tumor control.

Activation of the aryl hydrocarbon receptor inhibits the development of experimental autoimmune pancreatitis through IL-22-mediated signaling pathways.

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor expressed in hematopoietic and non-hematopoietic cells. Activation of the AhR by xenobiotics, microbial metabolites, and natural substances induces immunoregulatory responses. Autoimmune pancreatitis (AIP) is a chronic fibroinflammatory disorder of the pancreas driven by autoimmunity. Although AhR activation generally suppresses pathogenic autoimmune responses, the roles played by the AhR in AIP have been poorly defined. In this study, we examined how AhR activation affected the development of experimental AIP caused by the activation of plasmacytoid dendritic cells producing IFN-α and IL-33. Experimental AIP was induced in MRL/MpJ mice by repeated injections of polyinosinic-polycytidylic acid. Activation of the AhR by indole-3-pyruvic acid and indigo naturalis, which were supplemented in the diet, inhibited the development of experimental AIP, and these effects were independent of the activation of plasmacytoid dendritic cells producing IFN-α and IL-33. Interaction of indole-3-pyruvic acid and indigo naturalis with AhRs robustly augmented the production of IL-22 by pancreatic islet α cells. The blockade of IL-22 signaling pathways completely canceled the beneficial effects of AhR ligands on experimental AIP. Serum IL-22 concentrations were elevated in patients with type 1 AIP after the induction of remission with prednisolone. These data suggest that AhR activation suppresses chronic fibroinflammatory reactions that characterize AIP via IL-22 produced by pancreatic islet α cells.

THE CRITICAL ROLE OF THE HISTONE MODIFICATION ENZYME SETDB2 IN THE PATHOGENESIS OF ACUTE RESPIRATORY DISTRESS SYNDROME.

ABSTRACT: Introduction: Acute respiratory distress syndrome (ARDS) is a severe hypoxemic respiratory failure with a high in-hospital mortality. However, the molecular mechanisms underlying ARDS remain unclear. Recent findings have indicated that the onset of severe inflammatory diseases, such as sepsis, is regulated by epigenetic changes. We investigated the role of epigenetic changes in ARDS pathogenesis using mouse models and human samples. Methods: Acute respiratory distress syndrome was induced in a mouse model (C57BL/6 mice, myeloid cell or vascular endothelial cell [VEC]-specific SET domain bifurcated 2 [Setdb2]-deficient mice [Setdb2 ff Lyz2 Cre+ or Setdb2 ff Tie2 Cre+ ], and Cre - littermates) by intratracheal administration of lipopolysaccharide (LPS). Analyses were performed at 6 and 72 h after LPS administration. Sera and lung autopsy specimens from ARDS patients were examined. Results: In the murine ARDS model, we observed high expression of the histone modification enzyme SET domain bifurcated 2 ( Setdb2 ) in the lungs. In situ hybridization examination of the lungs revealed Setdb2 expression in macrophages and VECs. The histological score and albumin level of bronchoalveolar lavage fluid were significantly increased in Setdb2 ff Tie2 Cre+ mice following LPS administration compared with Setdb2 ff Tie2 Cre- mice, whereas there was no significant difference between the control and Setdb2 ff Lyz2 Cre+ mice. Apoptosis of VECs was enhanced in Setdb2 ff Tie2 Cre+ mice. Among the 84 apoptosis-related genes, the expression of TNF receptor superfamily member 10b ( Tnfrsf10b ) was significantly higher in Setdb2 ff Tie2 Cre+ mice than in control mice. Acute respiratory distress syndrome patients' serum showed higher SETDB2 levels than those of healthy volunteers. SETDB2 levels were negatively correlated with the partial pressure of oxygen in arterial blood/fraction of inspiratory oxygen concentration ratio. Conclusion: Acute respiratory distress syndrome elevates Setdb2 , apoptosis of VECs, and vascular permeability. Elevation of histone methyltransferase Setdb2 suggests the possibility to histone change and epigenetic modification. Thus, Setdb2 may be a novel therapeutic target for controlling the pathogenesis of ARDS.

Histopathological subtyping of high-grade serous ovarian cancer using whole slide imaging.

OBJECTIVE: We have established 4 histopathologic subtyping of high-grade serous ovarian cancer (HGSOC) and reported that the mesenchymal transition (MT) type has a worse prognosis than the other subtypes. In this study, we modified the histopathologic subtyping algorithm to achieve high interobserver agreement in whole slide imaging (WSI) and to characterize the tumor biology of MT type for treatment individualization. METHODS: Four observers performed histopathological subtyping using WSI of HGSOC in The Cancer Genome Atlas data. As a validation set, cases from Kindai and Kyoto Universities were independently evaluated by the 4 observers to determine concordance rates. In addition, genes highly expressed in MT type were examined by gene ontology term analysis. Immunohistochemistry was also performed to validate the pathway analysis. RESULTS: After algorithm modification, the kappa coefficient, which indicates interobserver agreement, was greater than 0.5 (moderate agreement) for the 4 classifications and greater than 0.7 (substantial agreement) for the 2 classifications (MT vs. non-MT). Gene expression analysis showed that gene ontology terms related to angiogenesis and immune response were enriched in the genes highly expressed in the MT type. CD31 positive microvessel density was higher in the MT type compared to the non-MT type, and tumor groups with high infiltration of CD8/CD103 positive immune cells were observed in the MT type. CONCLUSION: We developed an algorithm for reproducible histopathologic subtyping classification of HGSOC using WSI. The results of this study may be useful for treatment individualization of HGSOC, including angiogenesis inhibitors and immunotherapy.

Alveolar macrophages instruct CD8+ T cell expansion by antigen cross-presentation in lung.

Lung CD8+ memory T cells play central roles in protective immunity to respiratory viruses, such as influenza A virus (IAV). Here, we find that alveolar macrophages (AMs) function as antigen-presenting cells that support the expansion of lung CD8+ memory T cells. Intranasal antigen administration to mice subcutaneously immunized with antigen results in a rapid expansion of antigen-specific CD8+ T cells in the lung, which is dependent on antigen cross-presentation by AMs. AMs highly express interleukin-18 (IL-18), which mediates subsequent formation of CD103+CD8+ resident memory T (TRM) cells in the lung. In a mouse model of IAV infection, AMs are required for expansion of virus-specific CD8+ T cells and CD103+CD8+ TRM cells and inhibiting virus replication in the lungs during secondary infection. These results suggest that AMs instruct a rapid expansion of antigen-specific CD8+ T cells in lung, which protect the host from respiratory virus infection.

Disruption of the intestinal barrier exacerbates experimental autoimmune pancreatitis by promoting the translocation of Staphylococcus sciuri into the pancreas.

Autoimmune pancreatitis (AIP) and IgG4-related disease (IgG4-RD) are new disease entities characterized by enhanced IgG4 antibody responses and involvement of multiple organs, including the pancreas and salivary glands. Although the immunopathogenesis of AIP and IgG4-RD is poorly understood, we previously reported that intestinal dysbiosis mediates experimental AIP through the activation of IFN-α- and IL-33-producing plasmacytoid dendritic cells (pDCs). Because intestinal dysbiosis is linked to intestinal barrier dysfunction, we explored whether the latter affects the development of AIP and autoimmune sialadenitis in MRL/MpJ mice treated with repeated injections of polyinosinic-polycytidylic acid [poly (I:C)]. Epithelial barrier disruption was induced by the administration of dextran sodium sulfate (DSS) in the drinking water. Mice co-treated with poly (I:C) and DSS, but not those treated with either agent alone, developed severe AIP, but not autoimmune sialadenitis, which was accompanied by the increased accumulation of IFN-α- and IL-33-producing pDCs. Sequencing of 16S ribosomal RNA revealed that Staphylococcus sciuri translocation from the gut to the pancreas was preferentially observed in mice with severe AIP co-treated with DSS and poly (I:C). The degree of experimental AIP, but not of autoimmune sialadenitis, was greater in germ-free mice mono-colonized with S. sciuri and treated with poly (I:C) than in germ-free mice treated with poly (I:C) alone, which was accompanied by the increased accumulation of IFN-α- and IL-33-producing pDCs. Taken together, these data suggest that intestinal barrier dysfunction exacerbates AIP through the activation of pDCs and translocation of S. sciuri into the pancreas.

Reduced Number and Immune Dysfunction of CD4+ T Cells in Obesity Accelerate Colorectal Cancer Progression.

Obesity, a known risk factor for various types of cancer, reduces the number and function of cytotoxic immune cells in the tumor immune microenvironment (TIME). However, the impact of obesity on CD4+ T cells remains unclear. Therefore, this study aimed to clarify the impact of obesity on CD4+ T cells in the TIME. A tumor-bearing obese mouse model was established by feeding with 45% high-fat diet (HFD), followed by inoculation with a colon cancer cell line MC38. Tumor growth was significantly accelerated compared to that in mice fed a control diet. Tumor CD4+ T cells showed a significant reduction in number and an increased expression of programmed death-1 (PD-1), and decreased CD107a expression and cytokine such as IFN-γ and TNF-α production, indicating dysfunction. We further established CD4+ T cell-depleted HFD-fed model mice, which showed reduced tumor infiltration, increased PD-1 expression in CD8+ T cells, and obesity-induced acceleration of tumor growth in a CD4+ T cell-dependent manner. These findings suggest that the reduced number and dysfunction of CD4+ T cells due to obesity led to a decreased anti-tumor response of both CD4+ and CD8+ T cells to ultimately accelerate the progression of colorectal cancer. Our findings may elucidate the pathogenesis for poor outcomes of colorectal cancer associated with obesity.

Long-term maintenance of lung resident memory T cells is mediated by persistent antigen.

Tissue-resident memory T cells (TRM) in the lungs are pivotal for protection against repeated infection with respiratory viruses. However, the gradual loss of these cells over time and the associated decline in clinical protection represent a serious limit in the development of efficient T cell based vaccines against respiratory pathogens. Here, using an adenovirus expressing influenza nucleoprotein (AdNP), we show that CD8 TRM in the lungs can be maintained for at least 1 year post vaccination. Our results reveal that lung TRM continued to proliferate in situ 8 months after AdNP vaccination. Importantly, this required airway vaccination and antigen persistence in the lung, as non-respiratory routes of vaccination failed to support long-term lung TRM maintenance. In addition, parabiosis experiments show that in AdNP vaccinated mice, the lung TRM pool is also sustained by continual replenishment from circulating memory CD8 T cells that differentiate into lung TRM, a phenomenon not observed in influenza-infected parabiont partners. Concluding, these results demonstrate key requirements for long-lived cellular immunity to influenza virus, knowledge that could be utilized in future vaccine design.

Vaccine Based on Dendritic Cells Electroporated with an Exogenous Ovalbumin Protein and Pulsed with Invariant Natural Killer T Cell Ligands Effectively Induces Antigen-Specific Antitumor Immunity.

(1) Background: Cancer vaccines are administered to induce cytotoxic CD8+ T cells (CTLs) specific for tumor antigens. Invariant natural killer T (iNKT) cells, the specific T cells activated by α-galactosylceramide (α-GalCer), play important roles in this process as they are involved in both innate and adaptive immunity. We developed a new cancer vaccine strategy in which dendritic cells (DCs) were loaded with an exogenous ovalbumin (OVA) protein by electroporation (EP) and pulsed with α-GalCer. (2) Methods: We generated bone marrow-derived DCs from C57BL/6 mice, loaded full-length ovalbumin proteins to the DCs by EP, and pulsed them with α-GalCer (OVA-EP-galDCs). The OVA-EP-galDCs were intravenously administered to C57BL/6 mice as a vaccine. We then investigated subsequent immune responses, such as the induction of iNKT cells, NK cells, intrinsic DCs, and OVA-specific CD8+ T cells, including tissue-resident memory T (TRM) cells. (3) Results: The OVA-EP-galDC vaccine efficiently rejected subcutaneous tumors in a manner primarily dependent on CD8+ T cells. In addition to the OVA-specific CD8+ T cells both in early and late phases, we observed the induction of antigen-specific TRM cells in the skin. (4) Conclusions: The OVA-EP-galDC vaccine efficiently induced antigen-specific antitumor immunity, which was sustained over time, as shown by the TRM cells.

Divergence of Tissue-Memory T Cells: Distribution and Function-Based Classification.

Tissue-resident memory T cells (Trm) comprise the majority of memory cells in nonlymphoid tissues and play a predominant role in immunity at barrier surfaces. A better understanding of Trm cell maintenance and function is essential for the development of vaccines that confer frontline protection. However, it is currently challenging to precisely distinguish Trm cells from other T cells, and this has led to confusion in the literature. Here we highlight gaps in our understanding of tissue memory and discuss recent advances in the classification of Trm cell subsets based on their distribution and functional characteristics.

Impact of multiple hits with cognate antigen on memory CD8+ T-cell fate.

Antigen-driven activation of CD8+ T cells results in the development of a robust anti-pathogen response and ultimately leads to the establishment of long-lived memory T cells. During the primary response, CD8+ T cells interact multiple times with cognate antigen on distinct types of antigen-presenting cells. The timing, location and context of these antigen encounters significantly impact the differentiation programs initiated in the cells. Moderate re-activation in the periphery promotes the establishment of the tissue-resident memory T cells that serve as sentinels at the portal of pathogen entry. Under some circumstances, moderate re-activation of T cells in the periphery can result in the excessive expansion and accumulation of circulatory memory T cells, a process called memory inflation. In contrast, excessive re-activation stimuli generally impede conventional T-cell differentiation programs and can result in T-cell exhaustion. However, these conditions can also elicit a small population of exhausted T cells with a memory-like signature and self-renewal capability that are capable of responding to immunotherapy, and restoration of functional activity. Although it is clear that antigen re-encounter during the primary immune response has a significant impact on memory T-cell development, we still do not understand the molecular details that drive these fate decisions. Here, we review our understanding of how antigen encounters and re-activation events impact the array of memory CD8+ T-cell subsets subsequently generated. Identification of the molecular programs that drive memory T-cell generation will advance the development of new vaccine strategies that elicit high-quality CD8+ T-cell memory.

Environmental cues regulate epigenetic reprogramming of airway-resident memory CD8+ T cells.

Tissue-resident memory T cells (TRM cells) are critical for cellular immunity to respiratory pathogens and reside in both the airways and the interstitium. In the present study, we found that the airway environment drove transcriptional and epigenetic changes that specifically regulated the cytolytic functions of airway TRM cells and promoted apoptosis due to amino acid starvation and activation of the integrated stress response. Comparison of airway TRM cells and splenic effector-memory T cells transferred into the airways indicated that the environment was necessary to activate these pathways, but did not induce TRM cell lineage reprogramming. Importantly, activation of the integrated stress response was reversed in airway TRM cells placed in a nutrient-rich environment. Our data defined the genetic programs of distinct lung TRM cell populations and show that local environmental cues altered airway TRM cells to limit cytolytic function and promote cell death, which ultimately leads to fewer TRM cells in the lung.

Pulmonary monocytes interact with effector T cells in the lung tissue to drive TRM differentiation following viral infection.

Lung resident memory CD8 T cells (TRM) are critical for protection against respiratory viruses, but the cellular interactions required for their development are poorly understood. Herein we describe the necessity of classical monocytes for the establishment of lung TRM following influenza infection. We find that, during the initial appearance of lung TRM, monocytes and dendritic cells are the most numerous influenza antigen-bearing APCs in the lung. Surprisingly, depletion of DCs after initial T cell priming did not impact lung TRM development or maintenance. In contrast, a monocyte deficient pulmonary environment in CCR2-/- mice results in significantly less lung TRM following influenza infection, despite no defect in the antiviral effector response or in the peripheral memory pool. Imaging shows direct interaction of antigen-specific T cells with antigen-bearing monocytes in the lung, and pulmonary classical monocytes from the lungs of influenza infected mice are sufficient to drive differentiation of T cells in vitro. These data describe a novel role for pulmonary monocytes in mediating lung TRM development through direct interaction with T cells in the lung.

U3-1402 sensitizes HER3-expressing tumors to PD-1 blockade by immune activation.

Immunotherapy targeting programmed cell death-1 (PD-1) induces durable antitumor efficacy in many types of cancer. However, such clinical benefit is limited because of the insufficient reinvigoration of antitumor immunity with the drug alone; therefore, rational therapeutic combinations are required to improve its efficacy. In our preclinical study, we evaluated the antitumor effect of U3-1402, a human epidermal growth factor receptor 3-targeting (HER3-targeting) antibody-drug conjugate, and its potential synergism with PD-1 inhibition. Using a syngeneic mouse tumor model that is refractory to anti-PD-1 therapy, we found that treatment with U3-1402 exhibited an obvious antitumor effect via direct lysis of tumor cells. Disruption of tumor cells by U3-1402 enhanced the infiltration of innate and adaptive immune cells. Chemotherapy with exatecan derivative (Dxd, the drug payload of U3-1402) revealed that the enhanced antitumor immunity produced by U3-1402 was associated with the induction of alarmins, including high-mobility group box-1 (HMGB-1), via tumor-specific cytotoxicity. Notably, U3-1402 significantly sensitized the tumor to PD-1 blockade, as a combination of U3-1402 and the PD-1 inhibitor significantly enhanced antitumor immunity. Further, clinical analyses indicated that tumor-specific HER3 expression was frequently observed in patients with PD-1 inhibitor-resistant solid tumors. Overall, U3-1402 is a promising candidate as a partner of immunotherapy for such patients.

Interstitial-resident memory CD8+ T cells sustain frontline epithelial memory in the lung.

Populations of CD8+ lung-resident memory T (TRM) cells persist in the interstitium and epithelium (airways) following recovery from respiratory virus infections. While it is clear that CD8+ TRM cells in the airways are dynamically maintained via the continuous recruitment of new cells, there is a vigorous debate about whether tissue-circulating effector memory T (TEM) cells are the source of these newly recruited cells. Here we definitively demonstrate that CD8+ TRM cells in the lung airways are not derived from TEM cells in the circulation, but are seeded continuously by TRM cells from the lung interstitium. This process is driven by CXCR6 that is expressed uniquely on TRM cells but not TEM cells. We further demonstrate that the lung interstitium CD8+ TRM cell population is also maintained independently of TEM cells via a homeostatic proliferation mechanism. Taken together, these data show that lung memory CD8+ TRM cells in the lung interstitium and airways are compartmentally separated from TEM cells and clarify the mechanisms underlying their maintenance.

CXCR6 regulates localization of tissue-resident memory CD8 T cells to the airways.

Resident memory T cells (TRM cells) are an important first-line defense against respiratory pathogens, but the unique contributions of lung TRM cell populations to protective immunity and the factors that govern their localization to different compartments of the lung are not well understood. Here, we show that airway and interstitial TRM cells have distinct effector functions and that CXCR6 controls the partitioning of TRM cells within the lung by recruiting CD8 TRM cells to the airways. The absence of CXCR6 significantly decreases airway CD8 TRM cells due to altered trafficking of CXCR6-/- cells within the lung, and not decreased survival in the airways. CXCL16, the ligand for CXCR6, is localized primarily at the respiratory epithelium, and mice lacking CXCL16 also had decreased CD8 TRM cells in the airways. Finally, blocking CXCL16 inhibited the steady-state maintenance of airway TRM cells. Thus, the CXCR6/CXCL16 signaling axis controls the localization of TRM cells to different compartments of the lung and maintains airway TRM cells.