bluuebunny/crossref_metadata_2025 huggingface dataset:
Short-chain fatty acids regulate regulatory T cells and intestinal pathology during oral mucosal infection
Authors: Bhaskaran Natarajan , Schneider Elizabeth , Pandiyan Pushpa
Published: January 2023

Complex interactions between the microbial flora and the host exert sophisticated means of immune tolerance and regulation mechanisms. One mechanism is by inducing the accumulation of regulatory T (Treg) cells. Here we show that the depletion of resident bacteria using antibiotics (Abx) causes oral and gut immunopathology during Oropharyngeal Candidiasis (OPC) infection. Abx treatment causes decrease in the frequency of Foxp3+ regulatory cells (Tregs) and IL-17A producing T cells, with a concomitant increase in oral tissue pathology. Although oral C. albicans (CA) is commonly controlled in the oral cavity, Abx treatment led to CA dependent oral and gut inflammation. The combination of short chain fatty acids (SCFA) partially controlled the pathology in Abx treated mice, correlating to an increase in the frequency of Foxp3+, IL-17A+, and Foxp3+IL-17A+ double positive (Treg17) cells in tongue and oral draining lymph nodes. SCFA enabled the restoration of Th17 cells and Treg cells and oral infection clearance, but did not reverse weight loss. Because SCFA treatment did not fully reverse the gut inflammation, it is evident that resident microbiota have SCFA independent homeostatic mechanisms in gut mucosa. We also found that SCFA potently induce Foxp3 and IL-17A expression in CD4+ T cells, depending on the cytokine milieu in vitro. Taken together, our data reveal that SCFA derived from resident bacteria play a critical role in controlling gut immunopathology by regulating T cell cytokines during oral mucosal infections.
Recently alloactivated CD4+CD8−CD25+T regulatory cells express CD8alpha and are potent suppressor cells
Authors: Hall Bruce Milne , Verma Nirupama Darshan , Robinson Catherine Margaret , Wang Chaunmin , Sharland Alexandra , Tran Giang , Wilcox Paul , Hodgkinson Suzanne Jean
Published: January 2023

Therapy with antigen-specific CD4+CD25+ T regulatory cells (Treg) for induction of transplant tolerance is desirable, as naïve thymic Treg (tTreg) are non-antigen specific and weak suppressor cells. Naïve tTreg from DA rats cultured with fully allogeneic PVG stimulator cells and IL-2 express IFN-gamma receptor (IFNGR) and IL-12 receptor beta2 (IL-12Rb2) and are more potent alloantigen-specific regulators that we call Ts1 cells.

This study examined other markers that could identify the activated alloantigen-specific Treg as a subpopulation within the CD4+CD25+Foxp3+Treg. After culture of naïve DA CD4+CD8−CD25+T cells with rIL-2 and PVG alloantigen, CD8a was expressed on 10–20% and CD8b on <5% of these cells. RT-PCR showed the CD8+ cells had increased Ifngr that characterizes Ts1 cells as well was Irf4, a transcription factor induced by TCR activation. Proliferation induced by re-culture with rIL-12 and alloantigen was greater with CD4+CD8+CD25+Treg consistent with the CD8a+ cells expressing IL-12R. In MLC, the CD8+ fraction had greater suppression than the mixed Ts1 population, whereas the CD4+CD8−CD25+T cells had less suppression. In an adoptive transfer assay, rIL-2 and alloantigen activated Treg suppress rejection at a ratio of 1:10 with naïve effector cells, whereas alloantigen and rIL-2 activated tTreg depleted of the CD8a cells did not prevent rejection.

This study showed the expression of CD8alpha by rIL-2 and alloantigen activation of CD4+CD8−CD25+Foxp3+T cells was a marker of alloantigen-specific Treg and may be of use to enrich alloantigen-specific Treg.
A protective role of IFN-γ in T cell-mediated colitis by regulation of Treg/Th17 via induction of indoleamine-2,3-deoxygenase
Authors: Zheng Song Guo , Xu Zhenjian , Wang Julie
Published: January 2023

IFN-γ is an important Th1 type cytokine and has a paradoxical effect on the pathogenesis of several chronic inflammatory disorders, including inflammatory bowel disease. Here we demonstrate that IFN-γ plays a protective effect on the model of T cell-mediated colitis. Compare with wild-type control, adoptive transfer of CD4+CD62L+ naïve T cells isolated from IFN-γ−/−mice into Rag1−/−mice induced a more severe wasting disease, correlating with increased expression of Th17 cells but decreased expression of Foxp3+ T regulatory cells in colon tissue and peripheral lymphoid organs. Additionally, infusion of CD4+CD62L+ naïve T cells isolated from IFN-γ receptor knock-out mice into recipient Rag1−/−mice also elicited a more severe wasting disease when compared to infusion of naïve T cells isolated from wild-type mice. IFN-γ induced indoleamine-2,3-dioxgenase (IDO), an enzyme in the metabolism of tryptophan, plays a critical roles in immune responses regulating the balance between Treg and Th17 cells. Our data suggest that IFN-γ provides a protective function in T cell-mediated intestinal inflammation by regulation of Treg and Th17 via the induction of IDO.
Regulatory T cells limit unconventional memory to preserve the capacity to mount protective CD8 memory responses to pathogens
Authors: Graham Jessica B , Da Costa Andreia S. , Swarts Jessica L , Lund Jennifer M
Published: January 2023

Immunological memory exists so that following infection, an expanded population of pathogen-specific lymphocytes can rapidly and efficiently control infection in the case of re-exposure. However, in the case of CD8+ T lymphocytes, a population of unconventional CD44+CD122+ virtual memory T cells (TVM) has been described that possesses many, though not all, features of “true memory” T cells, without the requirement of first encountering cognate antigen. Here, we demonstrate a novel role for regulatory T cell-mediated restraint of TVM through limiting IL-15 trans-presentation by CD11b+ dendritic cells. Further, we show that keeping TVM in check ensures development of functional, antigen-specific “true” memory phenotype CD8 T cells that can assist in pathogen control upon re-exposure.
IL-34 differentiation of immunosuppressive macrophage
Authors: Kelly-Scumpia Kindra , Shirazi Roksana , Modlin Robert
Published: January 2023

Myeloid cells are necessary for effective immune responses in infection. Myeloid cells enter the skin differentiate into effector myeloid cells, such as macrophages (MΦ). The local and systemic factors responsible for this phenomena are poorly understood. Leprosy provides a human model to study myeloid cells in the skin as exposure of patients to the same pathogen, Mycobacterium leprae, results in distinct disease manifestations. In self-limited, tuberculoid leprosy (T-lep), appropriate monocyte activation results in differentiation into effector M1 MΦ that produce antimicrobial peptides and control M. leprae. In lepromatous leprosy (L-lep), inadequate immune responses fail to clear M. leprae. IL-15 and IL-10 were previously found to promote effector M1 MΦ in T-lep and immunosuppressive M2 MΦ in L-lep patients, respectively, but whether other immunologic factors affect MΦ differentiation in leprosy has not been investigated. Herein, we find IL-34, a cytokine known to drive M2 MΦ development in vitro, is increased in L-lep lesions. Differentiation of monocyte derived MΦ (MDM) with IL-34 results in increased IL-10 production following stimulation with M. leprae when compared to M-CSF derived MDM. Similar to IL-10 derived MDM, IL-34 derived MDM demonstrate increased phagocytic activity, but surprisingly, are able to activate a vitamin D dependent antimicrobial response similar to IL-15 derived MDM. These data indicate that IL-34 drives an alternative pathway of M2 MΦ development. IL-34 MDM produce anti-inflammatory cytokines, but retain the ability to induce antimicrobial responses. These findings raise the potential of IL-34 or IL-34 MDM to treat inflammatory skin disorders while retaining the ability to fight infection.
Sympathetic nervous tone influences anti-tumor immunity by mediating accumulation of myeloid-derived suppressor cells and regulatory T-cells
Authors: Nevin James Timothy , Corwin William L. , Srivastava Pramod K.
Published: January 2023

Sympathetic nerves innervate primary and secondary lymphoid organs, providing a means by which nervous system signaling can affect cellular immunity. For example, signaling by sympathetic nerves that innervate the spleen and bone marrow can regulate the development of myeloid cells. In cancer, aberrant myeloid development has been shown to drive accumulation of myeloid-derived suppressor cells (MDSCs) that suppress the anti-tumor immune response and are associated with poor prognosis. Given the known regulatory effect of sympathetic nervous system (SNS) signaling on myelopoiesis and the consequences of aberrant myeloid development in cancer, we hypothesized that the SNS plays an important role in regulating the immune response to cancer by influencing accumulation of MDSCs. Here, we demonstrate that inhibition of tonic SNS signaling compromises the anti-tumor immune response and results in increased tumor growth. We show that chemical ablation of the SNS by peripheral administration of 6-hydroxydopamine disrupts normal myeloid development in the bone marrow and results in expansion of CD11b+Ly6G+ polymorphonuclear MDSC and CD11b+ Ly6Chi monocytic MDSC in tumor-bearing mice. We show that these MDSC are able to potently suppress T-cell proliferation and mediate the expansion of regulatory T-cells that further inhibit the anti-tumor immune response. These data demonstrate how SNS-mediated regulation of immune development and function plays an important role in instructing the anti-tumor immune response. These findings have the potential to improve our understanding of the interplay between the nervous and immune systems, and better inform treatment of cancer and other immune-mediated pathologies.
PI3K activated tumors evade tumor immunity by promoting an inhibitory myeloid microenvironment
Authors: Collins Natalie B , Abosy Rose Al , Miller Brian , Bi Kevin , Manguso Robert , Yates Kathleen , Haining W. Nicholas
Published: January 2023

Oncogenes act in a cell-intrinsic way to promote tumorigenesis. Whether oncogenes also have a cell-extrinsic effect on suppressing the immune response to cancer is less well understood. Here we use an in vivo screen of known cancer-associated somatic mutations in immunocompetent mouse tumor models treated with checkpoint blockade to identify oncogenes that confer immune evasion. We found that a catalytically active mutation in Phospho-Inositol 3 Kinase (PI3K), PIK3CA c.3140A>G (H1047R) confers selective growth advantage after immunotherapy. Pharmacologic PI3K inhibition resensitizes mutant tumors to immunotherapy with anti-PD-1. The tumor microenvironment (TME) in PIK3CA H1047R-expressing tumors has fewer infiltrating CD8+ T cells after immunotherapy but is enriched for immune inhibitory myeloid cells. Inhibition of myeloid infiltration in PIK3CA H1047R tumors results in increased sensitivity to PD-1 checkpoint blockade. Thus PI3K has a role in tumor immune evasion mediated by establishment of an inhibitory myeloid microenvironment in addition to its well-described, cell-intrinsic oncogenic role. Activating mutations in PI3K may be useful as a biomarker of poor response to immunotherapy. More generally, our data suggest a rationale to combine PI3K inhibition with immunotherapy of PI3KCA mutant tumors.
Myeloid responders to programmed cell death
Authors: Messmer Michelle N , Snyder Annelise G , Gerner Michael Y , Oberst Andrew
Published: January 2023

Necroptosis is a programmed form of cell death that more efficiently primes CD8 T cell responses when compared to apoptosis. This efficiency has been associated with key signaling effects downstream of RIP kinase activation. However, mechanisms whereby these cell-intrinsic signals translate into enhanced adaptive immunity remain unclear. We hypothesized that dying cells would attract and engage with distinct myeloid populations based on which cell death pathway is active, and that these interactions would determine T cell priming ability. By introducing dying cells loaded with non-degradable ‘antigen’ into naïve mice, we have tracked myeloid cells that interacted with necroptotic or apoptotic cell debris up to 72 hrs after injection, corresponding to early detectable differences in T cell priming. Migratory dendritic cells equivalently took up antigen following either necroptosis or apoptosis. However, necroptosis resulted in a greater number of antigen positive macrophages and lymph node resident dendritic cells. Understanding how the immune system recognizes and responds to distinct forms of cell death will benefit investigations into cytotoxic and immunotherapeutic drug combinations.
Autophagy-based unconventional secretion of alarmin HMGB1 by keratinocytes plays a pivotal role in psoriatic skin inflammation
Authors: WANG ZHEN , ZHOU HONG , ZHENG HUAPING , TENG XIU , WEI XIAOQIONG , LI JIONG
Published: January 2023

The precise mechanism by which autophagy affects psoriasis is poorly understood. Here we show that keratinocytes (KCs) autophagy was activated in psoriatic lesions of patients and mice model, positively correlating with psoriatic severity, and could be inhibited by MAPK family (ERK, p38, JNK) inactivation, implying autophagy was associated with psoriatic inflammation. Indeed, impaired autophagy flux, caused by autophagy inhibitors or KC-specific deletion of Atg5, alleviated psoriasisform inflammation, demonstrating autophagy positively regulated psoriatic inflammation. We then found an autophagy-based unconventional secretory pathway (autosecretion), depended on Atg5 and GRASP55, promoted the psoriasiform KCs inflammation. Moreover, alarmin HMGB1 was more effective than other autosecretory proteins, IL-1β and IL-18, to regulate the psoriasiform cutaneous inflammation. HMGB1 neutralization in autophagy-efficient KCs vanished the psoriasiform inflammation differences between autophagy-efficient KCs and autophagy-deficient KCs. Conversely, recombinant HMGB1 almost completely restored the psoriasiform inflammation in autophagy-deficient KCs in vivo. These results suggested that HMGB1-associated autosecretion, not other intracellular autophagy pathways, played a pivotal role in cutaneous inflammation. Finally, we demonstrated that KC-specific HMGB1 deficient mice, but not DC- or myeloid cell-specific HMGB1 deficient mice, displayed attenuated psoriatic inflammation, due to the essential crosstalk between KC-specific HMGB1-associated autosecretion and γδT cells. Thus, we uncover a novel autophagy mechanism in psoriasis pathogenesis, and imply its clinical significance in psoriasis treatment.
Exploiting altered enhancer landscapes to decode pathogenic changes in gene expression of diverse hepatic macrophages
Authors: Troutman Ty Dale , Seidman Jason S. , Sakai Mashito , Gola Anita , Ouyang Zhengyu , Spann Nathanael J. , Bruni Cassi M. , Bennett Hunter , Ego Kaori , Vu BaoChau , Sun Xiaoli , Pasillas Martina , Witztum Joseph L. , Germain Ronald N. , Glass Christopher K.
Published: January 2023

Kupffer cells have specialized roles supporting the environment of the liver during homeostasis and disease. However, the key regulatory elements governing these behaviors are unknown. Using scRNA-seq, we found diversification of Kupffer cells and recruitment of additional macrophage subtypes during nonalcoholic steatohepatitis (NASH). A significant source of macrophage heterogeneity during NASH was traced to Cx3cr1 expressing monocytes. Further, macrophage subsets were localized in distinct niches, suggesting environmental specification as a key determinant of macrophage heterogeneity. We profiled chromatin accessibility of the major NASH associated macrophage populations to identify transcription factors governing their environmental specification. These results predict greater NFκB, RUNX, and AP1 activity in recruited hepatic macrophages compared to Kupffer cells. Surprisingly, we found minimal significant chromatin accessibility changes comparing Kupffer cells from healthy mice to mice with NASH, even though several thousand genes were differentially expressed. Instead, NASH led to altered chromatin activity, as measured by H3K27ac ChIP-seq, at Kupffer cell enhancer regions. A binding element for LXR (liver X receptor) was the top transcription factor motif identified in Kupffer cell enhancers with reduced activity during NASH. Furthermore, LXRα was required to maintain expression of a unique gene signature defining healthy Kupffer cells. Thus, our studies establish for the first time gene regulatory events controlling diverse hepatic macrophages during homeostasis and NASH.
Ptpn6 inhibits Caspase-8- and Ripk3/Mlkl-dependent inflammation
Authors: Croker Ben A , Speir Mary , Nowell Cameron J , Chen Alyce A , O’Donnell Joanne A , D’Cruz Akshay A , Bliss-Moreau Meghan , Wang Shu , Kelliher Michelle A , Hakem Razq , Gerlic Motti
Published: January 2023

Neutrophilic dermatoses are a group of inflammatory skin disorders characterized by sterile infiltrates of neutrophils. These syndromes include pyoderma gangrenosum (PG) and Sweet’s syndrome (SS), and they are associated with an increased incidence of inflammatory bowel disease, rheumatoid arthritis, and acute myeloid leukemia. IL-1β is detectable in skin lesions, and IL-1 neutralizing therapies have met with success in some patients. Splicing variants and promoter region deletions of protein tyrosine phosphatase-6 (PTPN6) are a feature of SS and PG. Mice lacking Ptpn6 develop a cutaneous inflammatory disease that is dependent on the IL-1 receptor, G-CSF, and neutrophils, but the source of IL-1 and the mechanisms of IL-1 release remain unclear. Here, we investigated the mechanisms controlling IL-1α/β release specifically from neutrophils (Ptpn6ΔPMN) by inhibiting Caspase-8-dependent apoptosis and Ripk1/Ripk3/Mlkl-regulated necroptosis. Loss of Ripk1 from neutrophils accelerated disease onset, whereas combined deletion of caspase-8 and either Ripk3 or Mlkl strongly protected Ptpn6ΔPMN mice. Ptpn6ΔPMN neutrophils displayed increased p38-dependent Ripk1-independent IL-1 and TNF production, and were prone to cell death. Together, these data emphasize dual functions for Ptpn6 in the negative regulation of p38 MAP kinase activation to control TNF and IL-1α/β transcription, and in maintaining Ripk1 function to prevent caspase-8- and Ripk3/Mlkl-dependent cell death and concomitant IL-1α/β release. These findings implicate neutrophils as the dominant producers of IL-1 in neutrophilic dermatoses, and identify novel therapeutic targets for the treatment of these skin disorders.
Extracellular calcium-phosphate nanoparticles activate the NLRP3 inflammasome through CaSR signaling
Authors: Jäger Elisabeth , Stäubert Claudia , Sungur Pelin , Höppener Stephanie , Strobel Sarah , Murthy Supriya , Rossol Manuela , Wagner Ulf
Published: January 2023

Calcium and phosphate are well known as the main minerals in bone ossification. But they also have an immunomodulatory potential as components of calciprotein particles. Such particles precipitate spontaneously in body fluids if the calcium and phosphate homeostasis is disturbed. An abnormally increased mineral load could lead to soft tissue calcification. In our study we investigated the cellular response of freshly isolated peripheral blood monocytes in serum-containing medium under precipitating (high calcium and high phosphate content) conditions. Using dynamic mass redistribution analyses with the EPIC® system and IL-1β measurements as readout, we identified phosphate as a necessary co-modulator of calcium-triggered overall cellular responses and NLRP3 inflammasome activation. Moreover, flow cytometry and ImageStream® analyses revealed the uptake of spontaneously formed nanoparticles under stimulating conditions, which depends on the activation of the G protein-coupled receptor Calcium-sensing receptor (CaSR). Our study unraveled a GPCR-mediated mechanism by which monocytes detect imbalances in calcium phosphate homeostasis in the extracellular space resulting in an enormous pro-inflammatory cellular response.
A sustained small increase in NOD1 expression promotes ligand-independent oncogenic activity
Authors: Rommereim Leah , Akhade Ajay Suresh , Dutta Bhaskar , Hutcheon Carolyn , Lounsbury Nicolas , Rostomily Clifford , Savan Ram , Fraser Iain , Germain Ronald , Subramanian Naeha
Published: January 2023

Small genetically-determined differences in transcription are implicated in complex disease but the mechanisms by which small changes in gene expression impact the origin of complex disease are unknown. Recent GWAS fine-mapping studies show that many causal alleles for complex immune-related diseases lie in non-coding regions of genes, generating expression quantitative trait loci (eQTLs), with the extent of expression variation between susceptible and resistant genotypes often in the 1.5–3 fold range. Concurrently, systems-level studies across several cancer types suggest a regulatory organization in which disease-promoting genomic mutations lie upstream of functional master regulator proteins, a change in whose expression by 1.5–3 fold is responsible for propagating a tumor cell state. Here we show that a persistent small (~1.5 fold) increase in expression of the innate sensor NOD1 (Nucleotide-binding Oligomerization Domain-containing protein 1) precipitates large cancer-promoting changes in cell state in the absence of ligand-driven activity. A ~1.2–1.4-fold increase in NOD1 protein concentration by loss of microRNA regulation sensitizes cells to ligand-induced inflammation with an additional slight increase leading to ligand-independent NOD1 activation that upregulates the expression of proto-oncogenes and is linked to poor prognosis in gastric cancer. Our data show that tight expression regulation of NOD1 prevents this sensor from exceeding a physiological switching checkpoint that promotes persistent inflammation and oncogene expression and reveal the impact of a single small quantitative change in cell state on cancer.
γδ T cell activation in response to a keratinocyte wound signature
Authors: Johnson Margarete Daisy , Higginson Cody , Tekkam Srinivas , Crooke Stephen , Lloyd Jessica , Witherden Deborah , Finn M.G. , Havran Wendy L
Published: January 2023

Epidermal resident γδ T cells are key players in the wound healing response, providing growth factors and cytokines that promote keratinocyte proliferation following recognition of damaged self. These cells are known to be unresponsive in chronic wounds and as such represent a novel target for new wound healing therapies. In this study we seek to modulate the γδ T cell response in chronic wounds through the identification, characterization, and delivery of keratinocyte-expressed proteins that comprise a “wound signature” allowing γδ T cells to recognize and respond to epithelial damage. We have identified a set of target genes though RNA-seq analysis which are upregulated following wounding in mice and correspond to membrane-bound and secreted proteins available for interactions with neighboring cells. Candidate gene expression was measured following wounding of human skin tissue, and ICAM1, CRISPLD2, and HSPA8 were shown to also be significantly upregulated in human skin wound samples. Hspa8 and ICAM-1 have been validated through in vitro assays to activate γδ T cells, and these proteins have been chosen for in vivo delivery. In order to deliver these candidate proteins into in vivo wound models we have developed a hydrogel-based delivery system with a highly tunable time frame of release. This system improves the retention of protein cargo at the wounds site post-wounding in mice and is being used to administer our candidate proteins into mouse models of non-healing wounds to assess effects on acceleration of tissue repair.
Tracking the in vivo history and fate of IL-2 producing CD8 T cells during memory differentiation
Authors: Yuzefpolskiy Yevgeniy , Sarkar Surojit , Kalia Vandana
Published: January 2023

IL-2 is a key driver of T cell responses critical for the conversion of resting antigen specific T cells into glycolytic powerhouses that undergo massive expansion and elicit cytolytic function to clear infections. Recent studies have used ex vivo antigen challenge combined with somatic IL-2 deletion to show that both paracrine and autocrine IL-2 signals are indispensable for the formation of highly functional memory CD8 T cells. Despite this critical role of autocrine IL-2 for memory T cell fate, the in vivo history and fate of autocrine IL-2 producing CD8 T cells remains unknown. We generated a mouse model in which autocrine IL-2 production leads to transcription of TdTomato, leading to an indelibly marked T cell. Following acute infection, a proportion of antigen specific CD8 T cells made autocrine IL-2, however, more than half of the T cell response had no history of IL-2 production. Furthermore, contrary to their ex vivo functionality, memory precursor and terminal effector T cells had similar history of in vivo IL-2 production. Despite autocrine IL-2 production being a benchmark of highly functional memory T cells, we observed no increase in the ex vivo polyfunctionality of T cells that produced in vivo IL-2 during infection. Finally, while strong TCR engagement is critical for the recruitment of robust T cell responses, we saw that in vivo autocrine IL-2 production is more effectively elicited by lower affinity TCR signals. Thus, we see that while proportion of T cells rely on autocrine IL-2 during infection, the ability of a CD8 T cell to recruit autocrine IL-2 function heavily depends on the avidity of the TCR to the peptide-MHC complex. Together these data provide the first insights into the in vivo function of IL-2 in antigen specific CD8 T cells.
Delineating the functions of RIPK1 in vivo through CRISPR/Cas9-mediated gene editing in mice
Authors: Zhang Xuhua , Dowling John P. , Zhang Jianke
Published: January 2023

RIPK1 is originally identified as a protein associated with Fas and TNFR1, which can trigger apoptosis. However, initial studies indicated that RIPK1 is dispensable for apoptotic signaling. Instead, cells lacking RIPK1 are defective in TNFR1-induced activation of NF-kB, a critical player involved in pro-survival signaling. Recent studies indicate that a certain form of necrosis-like death, necroptosis, requires RIPK1. In contrast, the Fas-associated death domain (FADD) protein appears to play an obligatory role in apoptosis induced by Fas and TNFR1. Fadd−/− mice die in utero due to RIPK1 mediated necroptosis. More recently, studies including ours demonstrated that only when both FADD and RIPK3 were deleted will Ripk1−/− mice survive to adulthood. This result indicates perinatal lethality in Ripk1−/− mice is due to not only FADD-mediated apoptosis but also RIPK3-dependent necrosis. Interestingly, we found that in adult T cells RIPK1 only suppresses apoptosis. To facilitate the dissection of the multiple functions of RIPK1, we performed in vivo gene editing using the CRISPR/Cas9 technology. This allows selective interruption of one functional domain/motif of RIPK1. Using these novel mouse models, our data reveals a previously unappreciated function for RIPK1 in promoting apoptosis.
The NF-kappa B regulator Bcl-3 is critical factor for memory CD8+ T cell generation
Authors: Jaiswal Hemant , Siebenlist Ulrich
Published: January 2023

Bcl-3 is an atypical member of the IκB family. Unlike classical IκBs, Bcl-3 is not degraded in response to activation signals, but instead its expression is often induced and it acts in the nucleus to modulate transcription of NF-κB target genes by associating with p50/NF-κB1 and p52/NF-κB2 homodimers. The latter complexes do not trans-activate autonomously, and association with Bcl-3 may promote or inhibit transcription in a highly gene- and context-dependent manner. Bcl-3 has been shown in mouse models to have important biological roles in promoting adaptive and innate immune responses to infectious agents as well as autoimmunity; however, the mechanisms have not been clearly defined. Here we investigate the role of Bcl-3 in primary and secondary T cell responses to lymphocytic choriomeningitis virus (LCMV) infection. Acutely infected mixed bone marrow chimeric mice generated from WT and Bcl-3 KO mice had defective LCMV antigen-specific CD8+ T cell recall responses in the absence of Bcl-3 upon challenge with LCMV (clone 13), despite having comparable primary CD8+ T cell responses at day 8 post primary infection (acute). Bcl-3 deficient CD8+ T cells instead exhibited significantly reduced proportions of LCMV epitope specific cells after 8 weeks of acute infection. Single cell RNA sequencing analysis of sorted LCMV epitope (GP33) specific CD8+ T cells at day 8 after acute infection showed a reduced proportion of memory precursor effector cells (MPECs) in the absence of Bcl-3. This suggests a crucial role of Bcl-3 in memory CD8+ T cell generation by regulating the formation or maintenance of MPECs during the primary response.
Signaling from individual Opioid Receptors differentially modulates the functional and phenotypic potential of resting and activated human CD8+ T cells
Authors: Mazahery Claire , Valadkhan Saba , Levine Alan D
Published: January 2023

Opioid use disorder is a worldwide epidemic. Endogenous opioids are released at sites of injury, and their cognate GPCR opioid receptors (OR) are expressed on immune cells; drugs of misuse appropriate these ORs. Conflicting published data show that opioid users have elevated risk of infection, opioids activate innate immune cells, and opioids attenuate murine T cell autoimmunity models. Intravenous drug use transmits blood borne pathogens, particularly viruses, making the study of CD8+ T cells pertinent to the crisis. Sequencing studies on leukocytes from heroin users show an altered transcriptome and a higher frequency of a SNP in a gene regulating long noncoding RNA. We demonstrate functional ORs (μ-, κ-, δ-subclasses) on human CD8+ T cells and employed unbiased analyses (flow cytometry with tSNE and transcriptome mapping) to study the effects of DPDPE (δ-OR agonist) and morphine (μ-OR biased agonist) on phenotype and activation status. Both agonists decrease activation markers on resting T cells, with δ-OR affecting naïve cells and μ-OR affecting naïve and memory cells. Pre-exposure to opioids changes the response to T cell receptor stimulation. Within 24 h, pre-exposure to either OR agonist decreases CD3, CD69, and CD25 expression. At 48 h, pre-exposure to μ-OR agonist downregulates CCR7 and CD27, while δ-OR agonist sustains CD3 expression. Regulation does not appear to be mediated by signaling cascades or calcium flux, suggesting a transcriptional mechanism: opioids dramatically modify the protein-coding and non-coding transcriptome of resting and activated T cells, with hundreds of non-coding RNAs showing increased expression. Selective engagement of OR represents a possible pathway to fine tune an immune response.
Immunology as a basis for STEM related teaching that is culturally appropriate and engages underrepresented High School students
Authors: Sika-Paotonu Dianne , Cockburn Alana
Published: January 2023

Indigenous Māori and Pacific Peoples in New Zealand (NZ) remain overrepresented both in adverse health statistics and education outcomes. To improve education and health outcomes for Māori and Pacific communities, various strategies and interventions have been proposed. This includes efforts to increase the Māori and Pacific health workforce populations figures more reflective of the make-up of the NZ population. These health careers require participation in science and health related subject matter throughout high school and at higher education learning levels. Māori and Pacific students remain underrepresented amongst these groups. The purpose of this work was to support improved interest and participation in science related learning that was Immunologically focused, amongst high school students from underrepresented groups that included Māori and Pacific students. A visit was carried out by a Biomedical Scientist to an all-girls high school class comprised of students who were of predominantly Māori and Pacific heritage to teach and share about Immunology research work that was Cancer based. The female Biomedical Scientist was of Pacific heritage and was an important consideration in the application of the role modelling concept for the Pacific students within this interaction. Verbal and written feedback highlighted the session was well-received and enjoyed by those in attendance. An important observation was the positive impact and enthusiasm of the students in response to the emphasis on the Immunologically focused Cancer based examples. This work demonstrated the importance of cultural considerations blended with scientific knowledge communication and interactions in the classroom.
The new paradigm shift in microbiology and immunology education in Azerbaijan Medical University Baku Azerbaijan
Authors: Fazal Nadeem , Ali Shamsideen , Sheriff Dauda , Brewer David , Childs Porscha
Published: January 2023

Medical Schools in USA and the rest of western world are launching their first major redesign of its early curriculum since the 1980s, integrating multiple disciplines into single courses and introducing earlier clinical immersion and flipped classrooms. It will require medical students to learn more actively, rather than cram and memorize material, and that it seeks to reflect how medicine has changed over the last four decades. Medical curricular change is part of a broader shift in medical education from asking students to memorize material to emphasizing its application. The new paradigm is driven by new innovations in medicine as well as new needs in medicine. Azerbaijan is a central Asian country at the cross roads of Western and Eastern cultures. As a former member of the Soviet Union, Azerbaijan science and education developed in last century on the basis of German-Russian model. However, in recent years, scientific, medical, and education fields are also drawing on American, British, French and Turkish models in developing new research/education standards. In this study we used innovative teaching techniques and modern web-based tools to advance student’s learning and comprehension. These parameters were evaluated in proposing a new paradigm shift in medical curriculum design at Azerbaijan Medical University via discussions, dialogues and planning, envision and strategize future efforts to slowly mold towards short-term and long-term goals. A pilot online course in Medical Microbiology and Immunology was tested for students registered in medicine, dental, pharmacy and public health faculties. Data from student and faculty surveys showed a marked acceptance of the need for a curricular redesign and welcoming paradigm shift.
Just in Time Teaching (JiTT) Strategies: From Implementation to Student Buy-in
Authors: Brown Deborah M , Brazeal Kathleen R. , Couch Brian A.
Published: January 2023

The Vision and Change in Undergraduate Biology Education calls for a transformation in instructional practices toward a more student-centered learning environment. Students in immunology courses can benefit from these strategies and one such approach, known as Just-in-Time Teaching (JiTT), is an inquiry based, active learning tactic that has been shown to enhance student retention of complex concepts. Prior to class, students read assignments and answer specific conceptual questions that involve linking new material or ideas with previously learned concepts. Importantly, an open-ended, reflective question is also asked about what students found most interesting, confusing, or difficult about the assignment. JiTT assignments have a positive impact on learning by allowing students to prepare ahead and use real life applications to better understand the material. Accordingly, the most successful JiTT assignments are questions based on the Case Studies in Immunology, 7th edition (Raif Geha and Luigi Notarangelo), that indicate which concepts are being discussed for each case. Overall, the JiTT approach has been well-received for upper level Immunology students. Results from formative assessment surveys generated by discipline based educational researchers (DBER) in our department in 2017 and 2018 indicate that most students understand the value of JiTT to enhance their learning, however, some students believe that assessments should always come after lecture, not before. Students agree that JiTT questions give them feedback on what material is important and what they still need to learn. Utilizing these surveys will help improve the messaging, learning intentions and overall student buy-in for future iterations of the course.
Post-traumatic pulmonary infection: the innate immune response protects the lung at the expense of the healing cutaneous wound
Authors: Crane Meredith , Xu Yun , Monaghan Sean F. , Hall Benjamin M. , Tran Holly L. , Henry William L , Albina Jorge E. , Jamieson Amanda M.
Published: January 2023

Hospitalized trauma patients have an increased risk of pulmonary infection. The acute response to injury, which relies on the coordinated activity of recruited leukocytes, may affect the early, innate immune-mediated control of lung infection. We hypothesized that a cutaneous wound would inhibit the ability of innate leukocytes to respond to a subsequent pulmonary bacterial infection. To address this, we developed a murine model of post-injury pneumonia. Mice were wounded by the dorsal subcutaneous implantation of polyvinyl alcohol sponges, which models the acute stages of wound repair, and a cohort was infected intranasally with the opportunistic bacterium Klebsiella oxytoca. For comparison, a control group was only infected. Wounding did not alter the early control of lung bacterial infection, the infiltration of innate leukocytes into the infected lung, or cytokine and chemokine levels in the bronchoalveolar lavage fluid. In contrast, pulmonary infection suppressed wound healing, caused decreased monocyte and neutrophil trafficking and lower cytokine and chemokine concentrations in the wound. Exogenous delivery of CCL2 and CXCL1 to the wound increased the number of neutrophils and accelerated healing. However, this led to increased pulmonary bacterial burden, highlighting the balance of cellular responses required to protect the lung. These data suggest that the innate immune system prioritizes the protection of one site above the other when faced with competing inflammatory insults. Here, early control of lung infection took priority over the healing wound. This work aims to elucidate mechanisms by which the innate immune system responds to multiple insults, which may be broadly applicable to inflammatory conditions.
Role of pulmonary innate immune memory leukocytes in the repeated respiratory infection of Influenza A virus
Authors: li nan
Published: January 2023

In recent years, innate immune memory and inflammation have become the frontier hot research fields in the host anti-infection field, especially in the respiratory infection of respirovirus, such as the influenza A virus. However, the fundamental and important scientific issues in the field of innate immune memory and inflammation remains poorly understood, such as the mechanism of pathogen-host interaction under acute and chronic infection in the infected upper respiratory track and lower respiratory track, and the way by which innate immune cells ( dendritic cells, monocytes/macrophages, NK cells, etc. ) acquire immune memory. Platelet chemokines CXCL4/5 play roles in mouse respiratory infection of bacteria or influenza A virus and also have regulatory effects on immune/inflammatory response in the infected lungs. Here studies on exploring the function and molecular mechanism of CXCL4/5 in inflammatory memory under IAV primary and repeat infection were conducted using CXCL4/5 knockout mice and wild-type mice by means of FACS, ATAC-seq, immunohistochemistry, mass cytometry and viral quantification. Data revealed that NK cells, NKT cells and macrophages have different inflammatory memory functions in repeated infection, and can be rapidly activated to play the function of anti-virus infection to protect the body. This memory function is due to epigenetic modification of genetic chromosome of the above leukocytes. Our results indicate that influenza-induced innate immune memory of NKT cells and macrophages play critical roles in anti-influenza repeat infections.
HIV infection associated oxidation of innate immune proteins in the human lung mucosa alters the host response to Mycobacterium tuberculosis infection
Authors: Akhter Anwari , Olmo-Fontánez Angélica , Moliva Juan , Azad Abul , Kelley Holden , Scordo Julia , Gavrilin Mikhail , Wewers Mark , Torrelles Jordi
Published: January 2023

Tuberculosis (TB) is the leading killer of people with HIV infection. Limited studies have considered HIV infection associated changes to the soluble components of the innate immune system in the alveolar lining fluid (ALF) as a contributing factor to increased susceptibility to pulmonary infections. We hypothesize that altered function of soluble innate immune components in human ALF of HIV infected subjects, drives susceptibility to primary M.tb infection. Here, we analyzed levels and functions of various soluble innate immune molecules present in human ALF from both healthy and HIV infected subjects. We also examined the effects of ALF components from HIV positive subjects on M.tb infection of human macrophages in vitro. Our results indicate an increased oxidation of innate proteins within ALF from HIV positive subjects not treated with anti-retroviral therapy (ART). We also found that complement components and some Th1 and Th2 cytokines/chemokines significantly differed between ALF from HIV positive and healthy subjects. Moreover, we found a deficiency of binding of surfactant protein-D to M.tb in ALF from HIV positive subjects, suggesting a defective innate immune function in the lung with HIV infection. Macrophages infected with M.tb exposed to ALF from HIV positive subjects without ART were less capable of controlling the infection. Our studies provide new information on how HIV infection could modify lung mucosal innate immune responses, thereby impacting M.tb infection and potentially other pulmonary diseases in the HIV infected population without ART treatment. Our data suggest that M.tb may benefit from the declining host innate defense function of the lung mucosa during HIV infection.
Age and HIV perturb the TB Immunity in Regulatory and Cytotoxic Subpopulations of NKT cells
Authors: Nishiguchi Tomoki , Mtetwa Godwin , Kay Alexander , Maphalala Gugu , Mace Emily , Mandalakas Anna M , DiNardo Andrew R
Published: January 2023

In the pre-antibiotic era, ~20% of Tuberculosis (TB) patients overcame the disease without therapy, implying host immunity can sometimes control disease. The risk of disease progression is higher among HIV-infected individuals and children, therefore we evaluate how HIV and age alter the balance of cytotoxic versus immune regulatory NKT immune phenotype in individuals with TB and latent Mycobacterium tuberculosis infection (LTBi). We evaluated the NKT immune response from 49 individuals with TB (25 HIV-infected) and 48 with asymptomatic LTBi (14 HIV-infected). Using flow cytometry based multi-dimensional immune profiling, we analyzed the expression level of previously described determinants of NKT cytotoxicity (CD3dim) and immune regulatory (CD3bright) phenotype and function. CD3bright NKT cell population was correlated with age during children TB (p = 0.002). This cell type had increased in adults with TB (p = 0.034), but not in children. Also, high level of perforin producing CD3bright NKT cells had increased in adults with TB compare to adults with LTBi (p = 0.004). On the other hand, CD3dim NKT cell had no age correlation nor difference between TB and LTBi participants. While HIV-infected LTBi individuals had decreased perforin production in CD3dim NKT cells (p = 0.012). CD3bright and CD3dim NKT sub-populations previously reported influence regulatory versus cytotoxic function. We demonstrate that age and HIV status of participants have effected to regulatory and cytotoxic function, respectively. This suggested that these two factors have different mechanisms, decreasing perforin high producing CD3bright NKT cells and decreasing perforin production in CD3dim NKT, which promote TB disease progression.
Ketogenic diet activates protective γδ T cell responses against influenza virus infection
Authors: Goldberg Emily Lauren , Molony Ryan , Sidorov Sviatoslav , Kudo Eriko , Dixit Vishwa Deep , Iwasaki Akiko
Published: January 2023

Influenza A virus (IAV) infection-associated morbidity and mortality are a key global healthcare concern, necessitating the identification of novel therapies capable of reducing the severity of IAV infections. In this study, we show that the consumption of a low-carbohydrate, high-fat ketogenic diet (KD) protects mice from lethal IAV infection and disease. KD feeding resulted in an expansion of γδ T cells in the lung that improved barrier functions, thereby enhancing anti-viral resistance. Expansion of these protective γδ T cells required metabolic adaptation to a ketogenic diet, as neither feeding mice a high-fat high-carbohydrate diet nor providing chemical ketone body substrate that bypasses hepatic ketogenesis protected against infection. Therefore, KD mediated immune-metabolic integration represents a viable avenue towards preventing or alleviating influenza disease.
Gamma Herpesvirus manipulates HVEM regulated IL-1β pathway to evade immune responses
Authors: Lin Wai W , Chapell Elizabeth , Brar Gurpreet , Wu Ting-Ting , Sun Ren , Baaten Bas , Ware Carl F.
Published: January 2023

The herpes virus entry mediator (HVEM) a member of the TNFR superfamily, is a key immunoregulator with both proinflammatory and inhibitory signaling functions. Both α and β-herpes viruses exploit HVEM to establish persistent infections, however little is known about whether the oncogenic γ-herpesviruses modulate HVEM. Here, we investigate the role of HVEM in a mouse model of γ-herpesvirus (MHV68) infection. Control C57BL/6 and HVEM-deficient (Tnfrsf14−/−) mice were intranasally infected with MHV68. We found that virus replication in the lungs of Tnfrsf14−/− mice increased significantly indicating HVEM expression is required to control MHV68 infection. We found that alveolar macrophages were drastically reduced in Tnfrsf14−/− mice compared to C57/BL6 controls suggesting their survival requires HVEM signaling. Importantly, we showed that IL-1β levels in the lungs of MHV68 infected Tnfrsf14−/−mice were significantly higher than C57BL/6 controls. Bone marrow-derived macrophage from Tnfrsf14−/− showed that HVEM is necessary to prevent IL-1β production in response to inflammasome activator. Antibodies that block IL-1R or IL-1β drastically increased the viral load in the lungs suggested that IL-1β is necessary to control viral replication. Taken together, our results highlight the role for HVEM in innate immune response evasion of MHV68 that could involve the inflammasome.