Immune cells express an array of inhibitory checkpoint receptors that are upregulated upon activation and limit tissue damage associated with excessive response to pathogens or allergens. Mouse leukocyte immunoglobulin like receptor B4 (LILRB4), also known as glycoprotein 49B (gp49B), is an inhibitory checkpoint receptor constitutively expressed in myeloid cells and upregulated in B cells, T cells, and NK cells upon activation. Here, we report that expression of LILRB4, which binds Zika virus (ZIKV), was increased in microglia and myeloid cells infiltrating the brains of neonatal mice with ZIKV-associated meningoencephalitis. Importantly, while C57BL/6 mice developed transient neurological symptoms but survived infection, mice lacking LILRB4/gp49B (LILRB4 KO) exhibited more severe signs of neurological disease and succumbed to disease. Their brains showed increased cellular infiltration but reduced control of viral burden. The reduced viral clearance was associated with altered NK cell function in the absence of LILRB4/gp49B. In naive animals, this manifested as reduced granzyme B responses to stimulation, but in ZIKV-infected animals, NK cells showed phenotypic changes that suggested altered maturation, diminished glucose consumption, reduced IFN-γ and granzyme B production, and impaired cytotoxicity. Together, our data reveal LILRB4/gp49B as an important regulator of NK cell function during viral infections.
Ha-Na Lee, Mohanraj Manangeeswaran, Aaron P. Lewkowicz, Kaliroi Engel, Monica Chowdhury, Mamatha Garige, Michael A. Eckhaus, Carole Sourbier, Derek D.C. Ireland, Daniela Verthelyi
Cells recognize both foreign and host-derived double-stranded RNA (dsRNA) via a signaling pathway that is usually studied in the context of viral infection. It has become increasingly clear that the sensing and handling of endogenous dsRNA is also critical for cellular differentiation and development. The adenosine RNA deaminase, ADAR1, has been implicated as a central regulator of the dsRNA response, but how regulation of the dsRNA response might mediate cell fate during injury and whether such signaling is cell intrinsic remain unclear. Here, we show that the ADAR1-mediated response to dsRNA was dramatically induced in 2 distinct injury models of gastric metaplasia. Mouse organoid and in vivo genetic models showed that ADAR1 coordinated a cell-intrinsic, epithelium-autonomous, and interferon signaling–independent dsRNA response. In addition, dsRNA accumulated within a differentiated epithelial population (chief cells) in mouse and human stomachs as these cells reprogrammed to a proliferative, reparative (metaplastic) state. Finally, chief cells required ADAR1 to reenter the cell cycle during metaplasia. Thus, cell-intrinsic ADAR1 signaling is critical for the induction of metaplasia. Because metaplasia increases cancer risk, these findings support roles for ADAR1 and the response to dsRNA in oncogenesis.
José B. Sáenz, Nancy Vargas, Charles J. Cho, Jason C. Mills
CD8+ tumor-infiltrating lymphocytes (TILs) are associated with improved survival in triple-negative breast cancer (TNBC) yet have no association with survival in estrogen receptor–positive (ER+) BC. The basis for these contrasting findings remains elusive. We identified subsets of BC tumors infiltrated by CD8+ T cells with characteristic features of exhausted T cells (TEX). Tumors with abundant CD8+ TEX exhibited a distinct tumor microenvironment marked by amplified interferon-γ signaling–related pathways and higher programmed death ligand 1 expression. Paradoxically, higher levels of tumor-infiltrating CD8+ TEX associated with decreased overall survival of patients with ER+ BC but not patients with TNBC. Moreover, high tumor expression of a CD8+ TEX signature identified dramatically reduced survival in premenopausal, but not postmenopausal, patients with ER+ BC. Finally, we demonstrated the value of a tumor TEX signature score in identifying high-risk premenopausal ER+ BC patients among those with intermediate Oncotype DX Breast Recurrence Scores. Our data highlight the complex relationship between CD8+ TILs, interferon-γ signaling, and ER status in BC patient survival. This work identifies tumor-infiltrating CD8+ TEX as a key feature of reduced survival outcomes in premenopausal patients with early-stage ER+ BC.
Colt A. Egelston, Weihua Guo, Jiayi Tan, Christian Avalos, Diana L. Simons, Min Hui Lim, Yinghui J. Huang, Michael S. Nelson, Arnab Chowdhury, Daniel B. Schmolze, John H. Yim, Laura Kruper, Laleh Melstrom, Kim Margolin, Joanne E. Mortimer, Yuan Yuan, James R. Waisman, Peter P. Lee
Treatment with anti–PD-1 and anti-PD-L1 therapies has shown durable clinical benefit in non–small cell lung cancer (NSCLC). However, patients with NSCLC with epidermal growth factor receptor (EGFR) mutations do not respond as well to treatment as patients without an EGFR mutation. We show that EGFR-mutated NSCLC expressed higher levels of CD73 compared with EGFR WT tumors and that CD73 expression was regulated by EGFR signaling. EGFR-mutated cell lines were significantly more resistant to T cell killing compared with WT cell lines through suppression of T cell proliferation and function. In a xenograft mouse model of EGFR-mutated NSCLC, neither anti–PD-L1 nor anti-CD73 antibody alone inhibited tumor growth compared with the isotype control. In contrast, the combination of both antibodies significantly inhibited tumor growth, increased the number of tumor-infiltrating CD8+ T cells, and enhanced IFN-γ and TNF-α production of these T cells. Consistently, there were increases in gene expression that corresponded to inflammation and T cell function in tumors treated with the combination of anti–PD-L1 and anti-CD73. Together, these results further support the combination of anti-CD73 and anti–PD-L1 therapies in treating EGFR-mutated NSCLC, while suggesting that increased T cell activity may play a role in response to therapy.
Eric Tu, Kelly McGlinchey, Jixin Wang, Philip Martin, Steven L.K. Ching, Nicolas Floc’h, James Kurasawa, Jacqueline H. Starrett, Yelena Lazdun, Leslie Wetzel, Barrett Nuttall, Felicia S.L. Ng, Karen T. Coffman, Paul D. Smith, Katerina Politi, Zachary A. Cooper, Katie Streicher
Aortic dissection and rupture are triggered by decreased vascular wall strength and/or increased mechanical loads. We investigated the role of mTOR signaling in aortopathy using a well-described model of angiotensin II–induced dissection, aneurysm, or rupture of the suprarenal abdominal aorta in Apoe-deficient mice. Although not widely appreciated, nonlethal hemorrhagic lesions present as pseudoaneurysms without significant dissection in this model. Angiotensin II–induced aortic tears result in free rupture, contained rupture with subadventitial hematoma (forming pseudoaneurysms), dilatation, or healing, while the media invariably thickens regardless of mural tears. Medial thickening results from smooth muscle cell hypertrophy and extracellular matrix accumulation, including matricellular proteins. Angiotensin II activates mTOR signaling in vascular wall cells, and inhibition of mTOR signaling by rapamycin prevents aortic rupture but promotes dissection. Decreased aortic rupture correlates with decreased inflammation and metalloproteinase expression, whereas extensive dissection correlates with induction of matricellular proteins that modulate adhesion of vascular cells. Thus, mTOR activation in vascular wall cells determines whether aortic tears progress to dissection or rupture. Previous mechanistic studies of aortic aneurysm and dissection by angiotensin II in Apoe-deficient mice should be reinterpreted as clinically relevant to pseudoaneurysms, and mTOR inhibition for aortic disease should be explored with caution.
Changshun He, Bo Jiang, Mo Wang, Pengwei Ren, Sae-Il Murtada, Alexander W. Caulk, Guangxin Li, Lingfeng Qin, Roland Assi, Constantinos J. Lovoulos, Martin A. Schwartz, Jay D. Humphrey, George Tellides
Norrie disease is caused by mutation of the NDP gene, presenting as congenital blindness followed by later onset of hearing loss. Protecting patients from hearing loss is critical for maintaining their quality of life. This study aimed to understand the onset of pathology in cochlear structure and function. By investigating patients and juvenile Ndp-mutant mice, we elucidated the sequence of onset of physiological changes (in auditory brainstem responses, distortion product otoacoustic emissions, endocochlear potential, blood-labyrinth barrier integrity) and determined the cellular, histological, and ultrastructural events leading to hearing loss. We found that cochlear vascular pathology occurs earlier than previously reported and precedes sensorineural hearing loss. The work defines a disease mechanism whereby early malformation of the cochlear microvasculature precedes loss of vessel integrity and decline of endocochlear potential, leading to hearing loss and hair cell death while sparing spiral ganglion cells. This provides essential information on events defining the optimal therapeutic window and indicates that early intervention is needed. In an era of advancing gene therapy and small-molecule technologies, this study establishes Ndp-mutant mice as a platform to test such interventions and has important implications for understanding the progression of hearing loss in Norrie disease.
Dale Bryant, Valda Pauzuolyte, Neil J. Ingham, Aara Patel, Waheeda Pagarkar, Lucy A. Anderson, Katie E. Smith, Dale A. Moulding, Yeh C. Leong, Daniyal J. Jafree, David A. Long, Amina Al-Yassin, Karen P. Steel, Daniel J. Jagger, Andrew Forge, Wolfgang Berger, Jane C. Sowden, Maria Bitner-Glindzicz
The fibrous annulus of the mitral valve plays an important role in valvular function and cardiac physiology, while normal variation in the size of cardiovascular anatomy may share a genetic link with common and rare disease. We derived automated estimates of mitral valve annular diameter in the 4-chamber view from 32,220 MRI images from the UK Biobank at ventricular systole and diastole as the basis for GWAS. Mitral annular dimensions corresponded to previously described anatomical norms, and GWAS inclusive of 4 population strata identified 10 loci, including possibly novel loci (GOSR2, ERBB4, MCTP2, MCPH1) and genes related to cardiac contractility (BAG3, TTN, RBFOX1). ATAC-Seq of primary mitral valve tissue localized multiple variants to regions of open chromatin in biologically relevant cell types and rs17608766 to an algorithmically predicted enhancer element in GOSR2. We observed strong genetic correlation with measures of contractility and mitral valve disease and clinical correlations with heart failure, cerebrovascular disease, and ventricular arrhythmias. Polygenic scoring of mitral valve annular diameter in systole was predictive of risk mitral valve prolapse across 4 cohorts. In summary, genetic and clinical studies of mitral valve annular diameter revealed genetic determinants of mitral valve biology, while highlighting clinical associations. Polygenic determinants of mitral valve annular diameter may represent an independent risk factor for mitral prolapse. Overall, computationally estimated phenotypes derived at scale from medical imaging represent an important substrate for genetic discovery and clinical risk prediction.
Mengyao Yu, Catherine Tcheandjieu, Adrien Georges, Ke Xiao, Helio Tejeda, Christian Dina, Thierry Le Tourneau, Madalina Fiterau, Renae Judy, Noah L. Tsao, Dulguun Amgalan, Chad J. Munger, Jesse M. Engreitz, Scott M. Damrauer, Nabila Bouatia-Naji, James R. Priest
The duodenum is a major site of HIV persistence during suppressive antiretroviral therapy despite harboring abundant tissue-resident memory (Trm) CD8+ T cells. The role of duodenal Trm CD8+ T cells in viral control is still not well defined. We examined the spatial localization, phenotype, and function of CD8+ T cells in the human duodenal tissue from people living with HIV (PLHIV) and healthy controls. We found that Trm (CD69+CD103hi) cells were the predominant CD8+ T cell population in the duodenum. Immunofluorescence imaging of the duodenal tissue revealed that CD103+CD8+ T cells were localized in the intraepithelial region, while CD103–CD8+ T cells and CD4+ T cells were mostly localized in the lamina propria (LP). Furthermore, HIV-specific CD8+ T cells were enriched in the CD69+CD103–/lo population. However, the duodenal HIV-specific CD8+ Trm cells rarely expressed canonical molecules for potent cytolytic function (perforin and granzyme B) but were more polyfunctional than those from peripheral blood. Taken together, our results show that duodenal CD8+ Trm cells possess limited perforin-mediated cytolytic potential and are spatially separated from HIV-susceptible LP CD4+ T cells. This could contribute to HIV persistence in the duodenum and provides critical information for the design of cure therapies.
Leonard Mvaya, Trevor Khaba, Agness E. Lakudzala, Thandeka Nkosi, Ndaru Jambo, Innocent Kadwala, Anstead Kankwatira, Priyanka D. Patel, Melita A. Gordon, Tonney S. Nyirenda, Kondwani C. Jambo, Zaza M. Ndhlovu
There is a high prevalence of ventricular arrhythmias related to sudden cardiac death in patients with chronic kidney disease (CKD). To explored the possible mechanism of CKD-related ventricular arrhythmias, a CKD rat model was created, and indoxyl sulfate (IS) was further used in vivo and in vitro. This project used the following methods: patch clamp, electrocardiogram, and some molecular biology experimental techniques. IS was found to be significantly elevated in the serum of CKD rats. Interestingly, the expression levels of the fast transient outward potassium current–related (Ito,f-related) proteins (Kv4.2, Kv4.3, and KChIP2) in the heart of CKD rats and rats treated with IS decreased. IS dose-dependently reduced Ito,f density, accompanied by the decreases in Kv4.2, Kv4.3, and KChIP2 proteins in vitro. IS also prolonged the action potential duration and QT interval, and paroxysmal ventricular tachycardia could be induced by IS. In-depth studies have shown that ROS/p38MAPK, ROS–p44/42 MAPK, and NF-κB signaling pathways play key roles in the reduction of Ito,f density and Ito,f-related proteins caused by IS. These data suggest that IS reduces Ito,f-related proteins and Ito,f density by activating ROS/MAPK and NF-κB signaling pathways, and the action potential duration and QT interval are subsequently prolonged, which contributes to increasing the susceptibility to arrhythmia in CKD.
Jing Yang, Hongxia Li, Chi Zhang, Yafeng Zhou
Tetralogy of Fallot (TOF) is the most common cyanotic heart defect, yet the underlying genetic mechanisms remain poorly understood. Here, we performed whole-genome sequencing analysis on 146 nonsyndromic TOF parent-offspring trios of Chinese ethnicity. Comparison of de novo variants and recessive genotypes of this data set with data from a European cohort identified both overlapping and potentially novel gene loci and revealed differential functional enrichment between cohorts. To assess the impact of these mutations on early cardiac development, we integrated single-cell and spatial transcriptomics of early human heart development with our genetic findings. We discovered that the candidate gene expression was enriched in the myogenic progenitors of the cardiac outflow tract. Moreover, subsets of the candidate genes were found in specific gene coexpression modules along the cardiomyocyte differentiation trajectory. These integrative functional analyses help dissect the pathogenesis of TOF, revealing cellular hotspots in early heart development resulting in cardiac malformations.
Clara Sze Man Tang, Mimmi Mononen, Wai-Yee Lam, Sheng Chih Jin, Xuehan Zhuang, Maria-Mercè Garcia-Barcelo, Qiongfen Lin, Yujia Yang, Makoto Sahara, Elif Eroglu, Kenneth R. Chien, Haifa Hong, Paul Kwong Hang Tam, Peter J. Gruber
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