Active immunization with the apolipoprotein B-100 (ApoB-100) peptide P210 reduces experimental atherosclerosis. To advance this immunization strategy to future clinical testing, we explored the possibility of delivering P210 as an antigen using nanoparticles, given this approach has been used clinically. We first characterized the responses of T cells to P210 using PBMCs from patients with atherosclerotic cardiovascular disease (ASCVD). We then investigated the use of P210 in self-assembling peptide amphiphile micelles (P210-PAMs) as a vaccine formulation to reduce atherosclerosis in B6.129P2-Apoetm1Unc/J (ApoE–/–) mice and P210’s potential mechanisms of action. We also generated and characterized a humanized mouse model with chimeric HLA-A*02:01/Kb in ApoE–/– background to test the efficacy of P210-PAM immunization as a bridge to future clinical testing. P210 provoked T cell activation and memory response in PBMCs of patients with ASCVD. Dendritic cell uptake of P210-PAM and its costaining with MHC-I molecules supported its use as a vaccine formulation. In ApoE–/– mice, immunization with P210-PAMs dampened P210-specific CD4+ T cell proliferative response and CD8+ T cell cytolytic response, modulated macrophage phenotype, and significantly reduced aortic atherosclerosis. Potential clinical relevance of P210-PAM immunization was demonstrated by reduced atherosclerosis in the humanized ApoE–/– mouse model. Our data support experimental and translational use of P210-PAM as a potential vaccine candidate against human ASCVD.
Kuang-Yuh Chyu, Xiaoning Zhao, Jianchang Zhou, Paul C. Dimayuga, Nicole W.M. Lio, Bojan Cercek, Noah T. Trac, Eun Ji Chung, Prediman K. Shah
Positive regulatory domain 1 (PRDM1) encodes B lymphocyte–induced maturation protein 1 (BLIMP1), also known as a master regulator of T cell homeostasis. We observed a negative relationship between Blimp-1 and IL-21 based on our previous data that Blimp-1 overexpression in T cells suppresses autoimmune diabetes while Blimp-1–deficient T cells contribute to colitis in NOD mice. Reanalysis of published data sets also revealed an inverse correlation between PRDM1 and IL21 in Crohn’s disease. Here, we illustrate that Blimp-1 repressed IL-21 by reducing chromatin accessibility and evicting an IL-21 activator, c-Maf, from the Il21 promoter. Moreover, Blimp-1 overexpression–mediated reduction in permissive chromatin structures at the Il21 promoter could override IL-21–accelerated autoimmune diabetogenesis in small ubiquitin-like modifier–defective c-Maf–transgenic mice. An autoregulatory feedback loop to harness IL-21 expression was unveiled by the evidence that IL-21 addition induced time-dependent Blimp-1 expression and subsequently enriched its binding to the Il21 promoter to suppress IL-21 overproduction. Furthermore, intervention of this feedback loop by IL-21 blockade, with IL-21R.Fc administration or IL-21 receptor deletion, attenuated Blimp-1 deficiency–mediated colitis and reinforced the circuit between Blimp-1 and IL-21 in the regulation of autoimmunity. We highlight the translation of Blimp-1–based epigenetic and transcriptomic profiles applicable to a personalized medicine approach in autoimmune diseases.
Yu-Wen Liu, Shin-Huei Fu, Ming-Wei Chien, Chao-Yuan Hsu, Ming-Hong Lin, Jia-Ling Dong, Rita Jui-Hsien Lu, Yi-Jing Lee, Pao-Yang Chen, Chih-Hung Wang, Huey-Kang Sytwu
Vertical sleeve gastrectomy (VSG) results in an increase in the number of hormone-secreting enteroendocrine cells (EECs) in the intestinal epithelium; however, the mechanism remains unclear. Notably, the beneficial effects of VSG are lost in a mouse model lacking the nuclear bile acid receptor farnesoid X receptor (FXR). FXR is a nuclear transcription factor that has been shown to regulate intestinal stem cell (ISC) function in cancer models. Therefore, we hypothesized that the VSG-induced increase in EECs is due to changes in intestinal differentiation driven by an increase in bile acid signaling through FXR. To test this, we performed VSG in mice that express EGFP in ISC/progenitor cells and performed RNA-Seq on GFP-positive cells sorted from the intestinal epithelia. We also assessed changes in EEC number (marked by glucagon-like peptide-1, GLP-1) in mouse intestinal organoids following treatment with bile acids, an FXR agonist, and an FXR antagonist. RNA-Seq of ISCs revealed that bile acid receptors are expressed in ISCs and that VSG explicitly alters expression of several genes that regulate EEC differentiation. Mouse intestinal organoids treated with bile acids and 2 different FXR agonists increased GLP-1–positive cell numbers, and administration of an FXR antagonist blocked these effects. Taken together, these data indicate that VSG drives ISC fate toward EEC differentiation through bile acid signaling.
Ki-Suk Kim, Bailey C.E. Peck, Yu-Han Hung, Kieran Koch-Laskowski, Landon Wood, Priya H. Dedhia, Jason R. Spence, Randy J. Seeley, Praveen Sethupathy, Darleen A. Sandoval
Fibrolamellar carcinoma (FLC) is an aggressive liver cancer primarily afflicting adolescents and young adults. Most patients with FLC harbor a heterozygous deletion on chromosome 19 that leads to the oncogenic gene fusion, DNAJB1-PRKACA. There are currently no effective therapeutics for FLC. To address that, it is critical to gain deeper mechanistic insight into FLC pathogenesis. We assembled a large sample set of FLC and nonmalignant liver tissue (n = 52) and performed integrative multiomic analysis. Specifically, we carried out small RNA sequencing to define altered microRNA expression patterns in tumor samples and then coupled this analysis with RNA sequencing and chromatin run-on sequencing data to identify candidate master microRNA regulators of gene expression in FLC. We also evaluated the relationship between DNAJB1-PRKACA and microRNAs of interest in several human and mouse cell models. Finally, we performed loss-of-function experiments for a specific microRNA in cells established from a patient-derived xenograft (PDX) model. We identified miR-10b-5p as the top candidate pro-proliferative microRNA in FLC. In multiple human cell models, overexpression of DNAJB1-PRKACA led to significant upregulation of miR-10b-5p. Inhibition of miR-10b in PDX-derived cells increased the expression of several potentially novel target genes, concomitant with a significant reduction in metabolic activity, proliferation, and anchorage-independent growth. This study highlights a potentially novel proliferative axis in FLC and provides a rich resource for further investigation of FLC etiology.
Adam B. Francisco, Matt Kanke, Andrew P. Massa, Timothy A. Dinh, Ramja Sritharan, Khashayar Vakili, Nabeel Bardeesy, Praveen Sethupathy
Epileptic seizures are common sequelae of stroke, acute brain injury, and chronic neurodegenerative diseases, including Alzheimer’s disease (AD), and cannot be effectively controlled in approximately 40% of patients, necessitating the development of novel therapeutic agents. Activation of the A1 receptor (A1R) by endogenous adenosine is an intrinsic mechanism to self-terminate seizures and protect neurons from excitotoxicity. However, targeting A1R for neurological disorders has been hindered by side effects associated with its broad expression outside the nervous system. Here we aim to target the neural-specific A1R/neurabin/regulator of G protein signaling 4 (A1R/neurabin/RGS4) complex that dictates A1R signaling strength and response outcome in the brain. We developed a peptide that blocks the A1R-neurabin interaction to enhance A1R activity. Intracerebroventricular or i.n. administration of this peptide shows marked protection against kainate-induced seizures and neuronal death. Furthermore, in an AD mouse model with spontaneous seizures, nasal delivery of this blocking peptide reduces epileptic spike frequency. Significantly, the anticonvulsant and neuroprotective effects of this peptide are achieved through enhanced A1R function in response to endogenous adenosine in the brain, thus, avoiding side effects associated with A1R activation in peripheral tissues and organs. Our study informs potentially new anti-seizure therapy applicable to epilepsy and other neurological illness with comorbid seizures.
Shalini Saggu, Yunjia Chen, Liping Chen, Diana Pizarro, Sandipan Pati, Wen Jing Law, Lori McMahon, Kai Jiao, Qin Wang
In many solid cancers, tumor-associated macrophages (TAM) represent the predominant myeloid cell population. Antigen (Ag) cross-presentation leading to tumor Ag–directed cytotoxic CD8+ T cell responses is crucial for antitumor immunity. However, the role of recruited monocyte-derived macrophages, including TAM, as potential cross-presenting cells is not well understood. Here, we show that primary human as well as mouse CD206+ macrophages are effective in functional cross-presentation of soluble self-Ag and non–self-Ag, including tumor-associated Ag (TAA), as well as viral Ag. To confirm the presence of cross-presenting TAM in vivo, we performed phenotypic and functional analysis of TAM from B16-F10 and CT26 syngeneic tumor models and have identified CD11b+F4/80hiCD206+ TAM to effectively cross-present TAA. We show that CD11b+CD206+ TAM represent the dominant tumor-infiltrating myeloid cell population, expressing a unique cell surface repertoire, promoting Ag cross-presentation and Ag-specific CD8+ T cell activation comparable with cross-presenting CLEC9A+ DCs (cDC1). The presence of cross-presenting CD206+ TAM is associated with reduced tumor burden in mouse syngeneic tumor models and with improved overall survival in cutaneous melanoma patients. Therefore, the demonstration of effective Ag cross-presentation capabilities of CD206+ TAM, including their clinical relevance, expands our understanding of TAM phenotypic diversity and functional versatility.
Madhura Modak, Ann-Kathrin Mattes, Daniela Reiss, Wioletta Skronska-Wasek, Rebecca Langlois, Nicolas Sabarth, Renate Konopitzky, Fidel Ramirez, Katharina Lehr, Tobias Mayr, David Kind, Coralie Viollet, Lee Kim Swee, Jutta Petschenka, Karim C. El Kasmi, Elfriede Noessner, Kerstin Kitt, Stefan Pflanz
The placenta is the primary organ for immune regulation, nutrient delivery, gas exchange, protection against environmental toxins, and physiologic perturbations during pregnancy. Placental inflammation and vascular dysfunction during pregnancy are associated with a growing list of prematurity-related complications. The goal of this study was to identify differences in gene expression profiles in fetal monocytes — cells that persist and differentiate postnatally — according to distinct placental histologic domains. Here, by using bulk RNA-Seq, we report that placental lesions are associated with gene expression changes in fetal monocyte subsets. Specifically, we found that fetal monocytes exposed to acute placental inflammation upregulate biological processes related to monocyte activation, monocyte chemotaxis, and platelet function, while monocytes exposed to maternal vascular malperfusion lesions downregulate these processes. Additionally, we show that intermediate monocytes might be a source of mitogens, such as HBEGF, NRG1, and VEGFA, implicated in different outcomes related to prematurity. This is the first study to our knowledge to show that placental lesions are associated with unique changes in fetal monocytes and monocyte subsets. As fetal monocytes persist and differentiate into various phagocytic cells following birth, our study may provide insight into morbidity related to prematurity and ultimately potential therapeutic targets.
Abhineet M. Sharma, Robert Birkett, Erika T. Lin, Linda M. Ernst, William A. Grobman, Suchitra Swaminathan, Hiam Abdala-Valencia, Alexander V. Misharin, Elizabeth T. Bartom, Karen K. Mestan
Insight into processes that determine CD8+ T cell memory formation has been obtained from infection models. These models are biased toward an inflammatory milieu and often use high-avidity CD8+ T cells in adoptive-transfer procedures. It is unclear whether these conditions mimic the differentiation processes of an endogenous repertoire that proceed upon noninflammatory conditions prevailing in premalignant tumor lesions. We examined the role of cytolytic capacity on CD8+ T cell fate decisions when primed by tumor cells or by minor histocompatibility antigen–mismatched leukocytes. CD8+ memory commitment was analyzed in Ebag9-deficient mice that exhibited enhanced tumor cell lysis. This property endowed Ebag9–/– mice with extended control of Tcl-1 oncogene–induced chronic lymphocytic leukemia progression. In Ebag9–/– mice, an expanded memory population was obtained for anti-HY and anti–SV-40 T antigen–specific T cells, despite unchanged effector frequencies in the primary response. By comparing the single-cell transcriptomes of CD8+ T cells responding to tumor cell vaccination, we found differential distribution of subpopulations between Ebag9+/+ and Ebag9–/– T cells. In Ebag9–/– cells, these larger clusters contained genes encoding transcription factors regulating memory cell differentiation and anti-apoptotic gene functions. Our findings link EBAG9-controlled cytolytic activity and the commitment to the CD8+ memory lineage.
Armin Rehm, Anthea Wirges, Dana Hoser, Cornelius Fischer, Stefanie Herda, Kerstin Gerlach, Sascha Sauer, Gerald Willimsky, Uta E. Höpken
BACKGROUND COVID-19 remains a global health emergency with limited treatment options, lagging vaccine rates, and inadequate healthcare resources in the face of an ongoing calamity. The disease is characterized by immune dysregulation and cytokine storm. Cyclosporine A (CSA) is a calcineurin inhibitor that modulates cytokine production and may have direct antiviral properties against coronaviruses.METHODS To test whether a short course of CSA was safe in patients with COVID-19, we treated 10 hospitalized, oxygen-requiring, noncritically ill patients with CSA (starting at a dose of 9 mg/kg/d). We evaluated patients for clinical response and adverse events, measured serum cytokines and chemokines associated with COVID-19 hyperinflammation, and conducted gene-expression analyses.RESULTS Five participants experienced adverse events, none of which were serious; transaminitis was most common. No participant required intensive care unit–level care, and all patients were discharged alive. CSA treatment was associated with significant reductions in serum cytokines and chemokines important in COVID-19 hyperinflammation, including CXCL10. Following CSA administration, we also observed a significant reduction in type I IFN gene expression signatures and other transcriptional profiles associated with exacerbated hyperinflammation in the peripheral blood cells of these patients.CONCLUSION Short courses of CSA appear safe and feasible in patients with COVID-19 who require oxygen and may be a useful adjunct in resource-limited health care settings.TRIAL REGISTRATION This trial was registered on ClinicalTrials.gov (Investigational New Drug Application no. 149997; ClinicalTrials.gov NCT04412785).FUNDING This study was internally funded by the Center for Cellular Immunotherapies.
Emily A. Blumberg, Julia Han Noll, Pablo Tebas, Joseph A. Fraietta, Ian Frank, Amy Marshall, Anne Chew, Elizabeth A. Veloso, Alison Carulli, Walter Rogal, Avery L. Gaymon, Aliza H. Schmidt, Tiffany Barnette, Renee Jurek, Rene Martins, Briana M. Hudson, Kalyan Chavda, Christina M. Bailey, Sarah E. Church, Hooman Noorchashm, Wei-Ting Hwang, Carl H. June, Elizabeth O. Hexner
DNA damage and genomic instability contribute to non–small cell lung cancer (NSCLC) etiology and progression. However, their therapeutic exploitation is disappointing. CTC-derived explants (CDX) offer systems for mechanistic investigation of CTC metastatic potency and may provide rationale for biology-driven therapeutics. Four CDX models and 3 CDX-derived cell lines were established from NSCLC CTCs and recapitulated patient tumor histology and response to platinum-based chemotherapy. CDX (GR-CDXL1, GR-CDXL2, GR-CDXL3, GR-CDXL4) demonstrated considerable mutational landscape similarity with patient tumor biopsy and/or single CTCs. Truncal alterations in key DNA damage response (DDR) and genome integrity–related genes were prevalent across models and assessed as therapeutic targets in vitro, in ovo, and in vivo. GR-CDXL1 presented homologous recombination deficiency linked to biallelic BRCA2 mutation and FANCA deletion, unrepaired DNA lesions after mitosis, and olaparib sensitivity, despite resistance to chemotherapy. SLFN11 overexpression in GR-CDXL4 led to olaparib sensitivity and was in coherence with neuroendocrine marker expression in patient tumor biopsy, suggesting a predictive value of SLFN11 in NSCLC histological transformation into small cell lung cancer (SCLC). Centrosome clustering promoted targetable chromosomal instability in GR-CDXL3 cells. These CDX unravel DDR and genome integrity–related defects as a central mechanism underpinning metastatic potency of CTCs and provide rationale for their therapeutic targeting in metastatic NSCLC.
Tala Tayoun, Vincent Faugeroux, Marianne Oulhen, Olivier Déas, Judith Michels, Laura Brulle-Soumare, Stefano Cairo, Jean-Yves Scoazec, Virginie Marty, Agathe Aberlenc, David Planchard, Jordi Remon, Santiago Ponce, Benjamin Besse, Patricia L. Kannouche, Jean-Gabriel Judde, Patrycja Pawlikowska, Françoise Farace
Preterm infants are susceptible to bloodstream infection by coagulase-negative staphylococci (CONS) that can lead to sepsis. Glucose-rich parenteral nutrition is commonly used to support the infants’ growth and energy expenditure but may exceed endogenous regulation during infection, causing dysregulated immune response and clinical deterioration. Using a preterm piglet model of neonatal CONS sepsis induced by Staphylococcus epidermidis (S. epidermidis) infection, we demonstrate the delicate interplay between immunity and glucose metabolism to regulate the host infection response. Circulating glucose levels, glycolysis, and inflammatory response to infection are closely connected across the states of tolerance, resistance, and immunoparalysis. Furthermore, high parenteral glucose provision during infection induces hyperglycemia, elevated glycolysis, and inflammation, leading to metabolic acidosis and sepsis, whereas glucose-restricted individuals are clinically unaffected with increased gluconeogenesis to maintain moderate hypoglycemia. Finally, standard glucose supply maintaining normoglycemia or pharmacological glycolysis inhibition enhances bacterial clearance and dampens inflammation but fails to prevent sepsis. Our results uncover how blood glucose and glycolysis control circulating immune responses, in turn determining the clinical fate of preterm infants infected with CONS. Our findings suggest further refinements of the current practice of parenteral glucose supply for preterm infants during infection.
Tik Muk, Anders Brunse, Nicole L. Henriksen, Karoline Aasmul-Olsen, Duc Ninh Nguyen
Follicular Th (Tfh) cells are key CD4+ Th cells involved in regulating B cell differentiation in the germinal center. Mice with conditional KO (CKO) of B lymphocyte-induced maturation protein-1 (BLIMP-1) expression in CD11chi DCs (Prdm1 CKO) spontaneously develop a lupus-like disease. We found an increase in the number of Tfh cells in secondary lymphoid organs in lupus-prone Prdm1-CKO mice. B cells stimulated with Tfh cells become Ab-secreting cells (ASCs); there was an increase in ASC differentiation mediated by Tfh cells from Prdm1-CKO mice compared with Tfh cells from control mice. This was mainly due to the increased expression of molecules within the IL-23/IL-17 pathway in Tfh cells from Prdm1-CKO mice. There was an increased frequency of IL-17A+ Tfh cells in Prdm1-CKO mice. These Tfh cells secrete IL-17A and an increased amount of IL-21. Furthermore, neutralizing IL-17A and IL-21 reduced ASC differentiation induced by Tfh cells. Lastly, we found the expression of IL-1β and IL-6 was increased in BLIMP-1–deficient DCs, which are key for Th17 induction. Altogether, the lack of BLIMP-1 expression in DCs induces not only an increased number of Tfh cells, but also functionally distinct Tfh cells that lead to increased ASC differentiation.
Vera Kim, Kyungwoo Lee, Hong Tian, Su Hwa Jang, Betty Diamond, Sun Jung Kim
SARS-CoV-2 has been confirmed in over 450 million confirmed cases since 2019. Although several vaccines have been certified by the WHO and people are being vaccinated on a global scale, it has been reported that multiple SARS-CoV-2 variants can escape neutralization by antibodies, resulting in vaccine breakthrough infections. Bacillus Calmette-Guérin (BCG) is known to induce heterologous protection based on trained immune responses. Here, we investigated whether BCG-induced trained immunity protected against SARS-CoV-2 in the K18-hACE2 mouse model. Our data demonstrate that i.v. BCG (BCG-i.v.) vaccination induces robust trained innate immune responses and provides protection against WT SARS-CoV-2, as well as the B.1.617.1 and B.1.617.2 variants. Further studies suggest that myeloid cell differentiation and activation of the glycolysis pathway are associated with BCG-induced training immunity in K18-hACE2 mice. Overall, our study provides the experimental evidence that establishes a causal relationship between BCG-i.v. vaccination and protection against SARS-CoV-2 challenge.
Bao-Zhong Zhang, Huiping Shuai, Hua-Rui Gong, Jing-Chu Hu, Bingpeng Yan, Terrence Tsz-Tai Yuen, Ye-Fan Hu, Chaemin Yoon, Xiao-Lei Wang, Yuxin Hou, Xuansheng Lin, Xiner Huang, Renhao Li, Yee Man Au-Yeung, Wenjun Li, Bingjie Hu, Yue Chai, Ming Yue, Jian-Piao Cai, Guang Sheng Ling, Ivan Fan-Ngai Hung, Kwok-Yung Yuen, Jasper Fuk-Woo Chan, Jian-Dong Huang, Hin Chu
Inherited kidney diseases are the fifth most common cause of end-stage renal disease (ESRD). Mitochondrial dysfunction plays a vital role in the progression of inherited kidney diseases, while mitochondrial-transfer RNA (mt-tRNA) variants and their pathogenic contributions to kidney disease remain largely unclear. In this study, we identified the pathogenic mt-tRNAPhe 616T>C mutation in 3 families and documented that m.616T>C showed a high pathogenic threshold, with both heteroplasmy and homoplasmy leading to isolated chronic kidney disease and hyperuricemia without hematuria, proteinuria, or renal cyst formation. Moreover, 1 proband with homoplamic m.616T>C presented ESRD as a child. No symptoms of nervous system evolvement were observed in these families. Lymphoblast cells bearing m.616T>C exhibited swollen mitochondria, underwent active mitophagy, and showed respiratory deficiency, leading to reduced mitochondrial ATP production, diminished membrane potential, and overproduction of mitochondrial ROS. Pathogenic m.616T>C abolished a highly conserved base pair (A31-U39) in the anticodon stem-loop which altered the structure of mt-tRNAPhe, as confirmed by a decreased melting temperature and slower electrophoretic mobility of the mutant tRNA. Furthermore, the unstable structure of mt-tRNAPhe contributed to a shortage of steady-state mt-tRNAPhe and enhanced aminoacylation efficiency, which resulted in impaired mitochondrial RNA translation and a significant decrease in mtDNA–encoded polypeptides. Collectively, these findings provide potentially new insights into the pathogenesis underlying inherited kidney disease caused by mitochondrial variants.
Chengxian Xu, Lingxiao Tong, Jia Rao, Qing Ye, Yuxia Chen, Yingying Zhang, Jie Xu, Xiaoting Mao, Feilong Meng, Huijun Shen, Zhihong Lu, Xiaohui Cang, Haidong Fu, Shugang Wang, Weiyue Gu, En-Yin Lai, Min-Xin Guan, Pingping Jiang, Jianhua Mao
COVID-19 infection causes collapse of glomerular capillaries and loss of podocytes, culminating in a severe kidney disease called COVID-19–associated nephropathy (COVAN). The underlying mechanism of COVAN is unknown. We hypothesized that cytokines induced by COVID-19 trigger expression of pathogenic APOL1 via JAK/STAT signaling, resulting in podocyte loss and COVAN phenotype. Here, based on 9 biopsy-proven COVAN cases, we demonstrated for the first time, to the best of our knowledge, that APOL1 protein was abundantly expressed in podocytes and glomerular endothelial cells (GECs) of COVAN kidneys but not in controls. Moreover, a majority of patients with COVAN carried 2 APOL1 risk alleles. We show that recombinant cytokines induced by SARS-CoV-2 acted synergistically to drive APOL1 expression through the JAK/STAT pathway in primary human podocytes, GECs, and kidney micro-organoids derived from a carrier of 2 APOL1 risk alleles, but expression was blocked by a JAK1/2 inhibitor, baricitinib. We demonstrate that cytokine-induced JAK/STAT/APOL1 signaling reduced the viability of kidney organoid podocytes but was rescued by baricitinib. Together, our results support the conclusion that COVID-19–induced cytokines are sufficient to drive COVAN-associated podocytopathy via JAK/STAT/APOL1 signaling and that JAK inhibitors could block this pathogenic process. These findings suggest JAK inhibitors may have therapeutic benefits for managing cytokine-induced, APOL1-mediated podocytopathy.
Sarah E. Nystrom, Guojie Li, Somenath Datta, Karen L. Soldano, Daniel Silas, Astrid Weins, Gentzon Hall, David B. Thomas, Opeyemi A. Olabisi
Respiratory failure in COVID-19 is characterized by widespread disruption of the lung’s alveolar gas exchange interface. To elucidate determinants of alveolar lung damage, we performed epithelial and immune cell profiling in lungs from 24 COVID-19 autopsies and 43 uninfected organ donors ages 18–92 years. We found marked loss of type 2 alveolar epithelial (T2AE) cells and increased perialveolar lymphocyte cytotoxicity in all fatal COVID-19 cases, even at early stages before typical patterns of acute lung injury are histologically apparent. In lungs from uninfected organ donors, there was also progressive loss of T2AE cells with increasing age, which may increase susceptibility to COVID-19–mediated lung damage in older individuals. In the fatal COVID-19 cases, macrophage infiltration differed according to the histopathological pattern of lung injury. In cases with acute lung injury, we found accumulation of CD4+ macrophages that expressed distinctly high levels of T cell activation and costimulation genes and strongly correlated with increased extent of alveolar epithelial cell depletion and CD8+ T cell cytotoxicity. Together, our results show that T2AE cell deficiency may underlie age-related COVID-19 risk and initiate alveolar dysfunction shortly after infection, and we define immune cell mediators that may contribute to alveolar injury in distinct pathological stages of fatal COVID-19.
Michael Chait, Mine M. Yilmaz, Shanila Shakil, Amy W. Ku, Pranay Dogra, Thomas J. Connors, Peter A. Szabo, Joshua I. Gray, Steven B. Wells, Masaru Kubota, Rei Matsumoto, Maya M.L. Poon, Mark E. Snyder, Matthew R. Baldwin, Peter A. Sims, Anjali Saqi, Donna L. Farber, Stuart P. Weisberg
K-ras–mutant lung adenocarcinoma (KM-LUAD) is associated with abysmal prognosis and is tightly linked to tumor-promoting inflammation. A human mAb, canakinumab, targeting the proinflammatory cytokine IL-1β, significantly decreased the risk of lung cancer in the Canakinumab Anti-inflammatory Thrombosis Outcomes Study. Interestingly, we found high levels of IL-1β in the lungs of mice with K-rasG12D–mutant tumors (CC-LR mice). Here, we blocked IL-1β using an anti–IL-1β mAb in cohorts of 6- or 14-week-old CC-LR mice to explore its preventive and therapeutic effect, respectively. IL-1β blockade significantly reduced lung tumor burden, which was associated with reprogramming of the lung microenvironment toward an antitumor phenotype characterized by increased infiltration of cytotoxic CD8+ T cells (with high IFN-γ and granzyme B expression but low programmed cell death 1 [PD-1] expression) while suppressing neutrophils and polymorphonuclear (PMN) myeloid-derived suppressor cells. When querying the Cancer Genome Atlas data set, we found positive correlations between IL1B expression and infiltration of immunosuppressive PMNs and expression of their chemoattractant, CXCL1, and PDCD1 expressions in patients with KM-LUAD. Our data provide evidence that IL-1β blockade may be a preventive strategy for high-risk individuals and an alternative therapeutic approach in combination with currently available treatments for KM-LUAD.
Bo Yuan, Michael J. Clowers, Walter V. Velasco, Stephen Peng, Qian Peng, Yewen Shi, Marco Ramos-Castaneda, Melody Zarghooni, Shuanying Yang, Rachel L. Babcock, Seon Hee Chang, John V. Heymach, Jianjun Zhang, Edwin J. Ostrin, Stephanie S. Watowich, Humam Kadara, Seyed Javad Moghaddam
Understanding the immune response to dengue virus (DENV) is essential for developing a dengue vaccine that is protective against all 4 DENV serotypes. We evaluated the immune response after vaccination (live attenuated tetravalent dengue vaccine TV005 or trivalent admixture) and after challenge with DEN2Δ30 (Tonga/74) to better understand the importance of homotypic immunity in vaccine protection. Significant increases in IP-10 expression were observed following receipt of either the trivalent or tetravalent vaccine. After challenge, a large increase in IP-10 expression was observed in the placebo and trivalent admixture groups but not in the tetravalent vaccine group. MCP-1, IL-1RA, and MIP-1β exhibited a similar pattern as IP-10. These results demonstrate protective effects of trivalent and tetravalent vaccines against DENV and suggest that the tetravalent vaccine has a better protective effect compared with the trivalent admixture. We also explored the postvaccination and postchallenge immune response differences between Black and White participants. White participants responded to vaccine differently than Black participants; Black participants receiving trivalent and tetravalent vaccines responded strongly and White participants responded only transiently in trivalent group. In response to challenge, White participants elicited a stronger response than Black participants. These results may explain why White participants may have a more vigorous DENV immune response than Black participants, as reported in literature.
Ruixue Hou, Lewis E. Tomalin, Jessica Pintado Silva, Seunghee Kim-Schulze, Stephen S. Whitehead, Ana Fernandez-Sesma, Anna P. Durbin, Mayte Suárez-Fariñas
Studies have demonstrated the phenotypic heterogeneity of vascular endothelial cells (ECs) within a vascular bed; however, little is known about how distinct endothelial subpopulations in a particular organ respond to an inflammatory stimulus. We performed single-cell RNA-Seq of 35,973 lung ECs obtained during baseline as well as postinjury time points after inflammatory lung injury induced by LPS. Seurat clustering and gene expression pathway analysis identified 2 major subpopulations in the lung microvascular endothelium, a subpopulation enriched for expression of immune response genes such as MHC genes (immuneEC) and another defined by increased expression of vascular development genes such as Sox17 (devEC). The presence of immuneEC and devEC subpopulations was also observed in nonhuman primate lungs infected with SARS-CoV-2 and murine lungs infected with H1N1 influenza virus. After the peak of inflammatory injury, we observed the emergence of a proliferative lung EC subpopulation. Overexpression of Sox17 prevented inflammatory activation in ECs. Thus, there appeared to be a “division of labor” within the lung microvascular endothelium in which some ECs showed propensity for inflammatory signaling and others for endothelial regeneration. These results provide underpinnings for the development of targeted therapies to limit inflammatory lung injury and promote regeneration.
Lianghui Zhang, Shang Gao, Zachary White, Yang Dai, Asrar B. Malik, Jalees Rehman
Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a chronic, debilitating, multisystem illness of unknown etiology for which no cure and no diagnostic tests are available. Despite increasing evidence implicating EBV and human herpesvirus 6A (HHV-6A) as potential causative infectious agents in a subset of patients with ME/CFS, few mechanistic studies address a causal relationship. In this study we examined a large ME/CFS cohort and controls and demonstrated a significant increase in activin A and IL-21 serum levels, which correlated with seropositivity for antibodies against the EBV and HHV-6 protein deoxyuridine triphosphate nucleotidohydrolase (dUTPases) but no increase in CXCL13. These cytokines are critical for T follicular helper (TFH) cell differentiation and for the generation of high-affinity antibodies and long-lived plasma cells. Notably, ME/CFS serum was sufficient to drive TFH cell differentiation via an activin A–dependent mechanism. The lack of simultaneous CXCL13 increase with IL-21 indicates impaired TFH function in ME/CFS. In vitro studies revealed that virus dUTPases strongly induced activin A secretion while in vivo, EBV dUTPase induced the formation of splenic marginal zone B and invariant NKTFH cells. Together, our data indicate abnormal germinal center (GC) activity in participants with ME/CFS and highlight a mechanism by which EBV and HHV6 dUTPases may alter GC and extrafollicular antibody responses.
Brandon S. Cox, Khaled Alharshawi, Irene Mena-Palomo, William P. Lafuse, Maria Eugenia Ariza
Atrial fibrillation (AF), the most common sustained cardiac arrhythmia and a major risk factor for stroke, often arises through ectopic electrical impulses derived from the pulmonary veins (PVs). Sequence variants in enhancers controlling expression of the transcription factor PITX2, which is expressed in the cardiomyocytes (CMs) of the PV and left atrium (LA), have been implicated in AF predisposition. Single nuclei multiomic profiling of RNA and analysis of chromatin accessibility combined with spectral clustering uncovered distinct PV- and LA-enriched CM cell states. Pitx2-mutant PV and LA CMs exhibited gene expression changes consistent with cardiac dysfunction through cell type–distinct, PITX2-directed, cis-regulatory grammars controlling target gene expression. The perturbed network targets in each CM were enriched in distinct human AF predisposition genes, suggesting combinatorial risk for AF genesis. Our data further reveal that PV and LA Pitx2-mutant CMs signal to endothelial and endocardial cells through BMP10 signaling with pathogenic potential. This work provides a multiomic framework for interrogating the basis of AF predisposition in the PVs of humans.
Jeffrey D. Steimle, Francisco J. Grisanti Canozo, Minjun Park, Zachary A. Kadow, Md. Abul Hassan Samee, James F. Martin
Inflammation of the esophageal epithelium is a hallmark of eosinophilic esophagitis (EoE), an emerging chronic allergic disease. Herein, we probed human esophageal epithelial cells at single-cell resolution during homeostasis and EoE. During allergic inflammation, the epithelial differentiation program was blocked, leading to loss of KRT6hi differentiated populations and expansion of TOP2hi proliferating, DSPhi transitioning, and SERPINB3hi transitioning populations; however, there was stability of the stem cell–enriched PDPNhi basal epithelial compartment. This differentiation program blockade was associated with dysregulation of transcription factors, including nuclear receptor signalers, in the most differentiated epithelial cells and altered NOTCH-related cell-to-cell communication. Each epithelial population expressed genes with allergic disease risk variants, supporting their functional interplay. The esophageal epithelium differed notably between EoE in histologic remission and controls, indicating that remission is a transitory state poised to relapse. Collectively, our data uncover the dynamic nature of the inflamed human esophageal epithelium and provide a framework to better understand esophageal health and disease.
Mark Rochman, Ting Wen, Michael Kotliar, Phillip J. Dexheimer, Netali Ben-Baruch Morgenstern, Julie M. Caldwell, Hee-Woong Lim, Marc E. Rothenberg
Dominant infectious tolerance explains how brief tolerance-inducing therapies result in lifelong tolerance to donor antigens and “linked” third-party antigens, while recipient sensitization and ensuing immunological memory prevent the successful induction of transplant tolerance. In this study, we juxtapose these 2 concepts to test whether mechanisms of dominant infectious tolerance can control a limited repertoire of memory T and B cells. We show that sensitization to a single donor antigen is sufficient to prevent stable transplant tolerance, rendering it unstable. Mechanistic studies revealed that recall antibody responses and memory CD8+ T cell expansion were initially controlled, but memory CD4+Foxp3– T cell (Tconv) responses were not. Remarkably, naive donor-specific Tconvs at tolerance induction also acquired a resistance to tolerance, proliferating and acquiring a phenotype similar to memory Tconvs. This phenomenon of “linked sensitization” underscores the challenges of reprogramming a primed immune response toward tolerance and identifies a potential therapeutic checkpoint for synergizing with costimulation blockade to achieve transplant tolerance in the clinic.
Michael S. Andrade, James S. Young, Jared M. Pollard, Dengping Yin, Maria-Luisa Alegre, Anita S. Chong
Secondary infections are frequent complications of viral respiratory infections, but the potential consequence of SARS-CoV-2 coinfection with common pulmonary pathogens is poorly understood. We report that coinfection of human ACE2–transgenic mice with sublethal doses of SARS-CoV-2 and Streptococcus pneumoniae results in synergistic lung inflammation and lethality. Mortality was observed regardless of whether SARS-CoV-2 challenge occurred before or after establishment of sublethal pneumococcal infection. Increased bacterial levels following coinfection were associated with alveolar macrophage depletion, and treatment with murine GM-CSF reduced numbers of lung bacteria and pathology and partially protected from death. However, therapeutic targeting of IFNs, an approach that is effective against influenza coinfections, failed to increase survival. Combined vaccination against both SARS-CoV-2 and pneumococci resulted in 100% protection against subsequent coinfection. The results indicate that when seasonal respiratory infections return to prepandemic levels, they could lead to an increased incidence of lethal COVID-19 superinfections, especially among the unvaccinated population.
Tarani Kanta Barman, Amit K. Singh, Jesse L. Bonin, Tanvir Noor Nafiz, Sharon L. Salmon, Dennis W. Metzger
BACKGROUND Limited information is available on the impact of immunosuppressants on COVID-19 vaccination in patients with immune-mediated inflammatory diseases (IMID).METHODS This observational cohort study examined the immunogenicity of SARS-CoV-2 mRNA vaccines in adult patients with inflammatory bowel disease, rheumatoid arthritis, ankylosing spondylitis, or psoriatic disease, with or without maintenance immunosuppressive therapies. Ab and T cell responses to SARS-CoV-2, including neutralization against SARS-CoV-2 variants, were determined before and after 1 and 2 vaccine doses.RESULTS We prospectively followed 150 subjects, 26 healthy controls, 9 patients with IMID on no treatment, 44 on anti-TNF, 16 on anti-TNF with methotrexate/azathioprine (MTX/AZA), 10 on anti–IL-23, 28 on anti–IL-12/23, 9 on anti–IL-17, and 8 on MTX/AZA. Ab and T cell responses to SARS-CoV-2 were detected in all participants, increasing from dose 1 to dose 2 and declining 3 months later, with greater attrition in patients with IMID compared with healthy controls. Ab levels and neutralization efficacy against variants of concern were substantially lower in anti-TNF–treated patients than in healthy controls and were undetectable against Omicron by 3 months after dose 2.CONCLUSIONS Our findings support the need for a third dose of the mRNA vaccine and for continued monitoring of immunity in these patient groups.FUNDING Funded by a donation from Juan and Stefania Speck and by Canadian Institutes of Health (CIHR)/COVID-Immunity Task Force (CITF) grants VR-1 172711 and VS1-175545 (to THW and ACG), CIHR FDN-143250 (to THW), GA2-177716 (to VC, ACG, and THW), and GA1-177703 (to ACG) and the CIHR rapid response network to SARS-CoV-2 variants, CoVaRR-Net (to ACG).
Roya M. Dayam, Jaclyn C. Law, Rogier L. Goetgebuer, Gary Y.C. Chao, Kento T. Abe, Mitchell Sutton, Naomi Finkelstein, Joanne M. Stempak, Daniel Pereira, David Croitoru, Lily Acheampong, Saima Rizwan, Klaudia Rymaszewski, Raquel Milgrom, Darshini Ganatra, Nathalia V. Batista, Melanie Girard, Irene Lau, Ryan Law, Michelle W. Cheung, Bhavisha Rathod, Julia Kitaygorodsky, Reuben Samson, Queenie Hu, W. Rod Hardy, Nigil Haroon, Robert D. Inman, Vincent Piguet, Vinod Chandran, Mark S. Silverberg, Anne-Claude Gingras, Tania H. Watts