SLIT3 deficiency attenuates pressure overload-induced cardiac fibrosis and remodeling

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Abstract

In pulmonary hypertension and certain forms of congenital heart disease, ventricular pressure overload manifests at birth and is an obligate hemodynamic abnormality that stimulates myocardial fibrosis which leads to ventricular dysfunction and poor clinical outcomes. Thus, an attractive strategy is to attenuate the myocardial fibrosis to help preserve ventricular function. Here, by analyzing RNA-sequencing databases and comparing the transcript and protein levels of fibrillar collagen in wild-type and global knockout mice, we found that SLIT3 was predominantly present in fibrillar collagen-producing cells and that SLIT3 deficiency attenuated collagen production in the heart and other non-neuronal tissues. We then performed transverse aortic constriction or pulmonary artery banding in wild-type and knockout mice to induce left and right ventricular pressure overload, respectively. We discovered that SLIT3 deficiency abrogates fibrotic and hypertrophic changes and promotes long-term ventricular function and overall survival in both left and right ventricular pressure overload. Furthermore, we found that SLIT3 stimulated fibroblast activity and fibrillar collagen production, which coincided with the transcription and nuclear localization of the mechanotransducer YAP1. These results indicate that SLIT3 is important for regulating fibroblast activity and fibrillar collagen synthesis in an autocrine manner, making it a potential therapeutic target for fibrotic diseases, especially myocardial fibrosis and adverse remodeling induced by persistent afterload elevation.

Authors

Lianghui Gong, Shuyun Wang, Li Shen, Catherine Liu, Mena Shenouda, Baolei Li, Xiaoxiao Liu, John Shaw, Alan Wineman, Yifeng Yang, Dingding Xiong, Anne Eichmann, Sylvia M. Evans, Stephen J. Weiss, Ming-Sing Si

Targeting macrophage checkpoint inhibitor SIRPa for anticancer therapy

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Abstract

The SIRPα-CD47 interaction provides a macrophage immune checkpoint pathway that plays a critical role in cancer immune evasion across multiple cancers. Here, we report the engineering of a humanized anti-SIRPα monoclonal antibody (1H9) for antibody target cancer therapy. 1H9 has broad activity across a wide range of SIRPα variants. Binding of 1H9 to SIRPα blocks its interaction with CD47, thereby promoting macrophage-mediated phagocytosis of cancer cells. Pre-clinical studies in vitro and in vivo demonstrate that 1H9 synergizes with other therapeutic antibodies to promote phagocytosis of tumor cells and inhibit tumor growth in both syngeneic and xenograft tumor models, leading to survival benefit. Thus, 1H9 can potentially act as a universal agent to enhance therapeutic efficacy when used in combination with most tumor-targeting antibodies. We report for the first time, a comparison of anti-SIRPα and anti-CD47 antibodies in CD47/SIRPα double humanized mice, and found that 1H9 exhibits a substantially reduced antigen-sink effect due to the limited tissue distribution of SIRPα expression. Toxicokinetic studies in non-human primates show that 1H9 is well tolerated with no treatment-related adverse effects noted. These data highlight the clinical potential of 1H9 as a pan-therapeutic with the desired properties when used in combination with tumor-targeting antibodies.

Authors

Jie Liu, Seethu Xavy, Shirley Mihardja, Sharline Chen, Kavitha Sompalli, Dongdong Feng, Timothy S. Choi, Balaji Agoram, Ravindra Majeti, Irving L. Weissman, Jens-Peter Volkmer

Targeting epithelial expressed Sialyl Lewis glycans improves colonic mucosal wound healing and protects against colitis

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Abstract

Dysregulated healing of injured mucosa is a hallmark of many pathological conditions including inflammatory bowel disease. Mucosal injury and chronic inflammation including persistent neutrophil (PMN) infiltration are also associated with alterations in epithelial glycosylation. Previous studies have revealed the inflammation induced glycan sLea on epithelial CD44v6 acts as a ligand for transmigrating PMN. Furthermore, blocking sLea-mediated binding interactions with the mAb GM35 reduced PMN transepithelial migration. Here we report that robust sialylated Lewis glycan expression is induced in colonic mucosa from individuals with ulcerative colitis (UC) and Crohn’s disease (CD) as well as in colonic epithelium of mice with DSS colitis. Targeting of sialylated epithelial Lewis glycans with mAb GM35 reduced disease activity and improved mucosal integrity during DSS induced colitis in mice. Wound healing studies revealed increased epithelial proliferation and migration responses as well as improved mucosal repair following ligation of epithelial sialyl Lewis glycans. Finally, we show GM35-mediated increases in epithelial proliferation and migration are mediated through activation of kinases that signal downstream of CD44v6 (Src, FAK, Akt). These findings suggest that sialylated Lewis glycans on epithelial CD44v6 may represent targets for improved recovery of epithelial barrier function and restitution of mucosal homeostasis following intestinal inflammation or injury

Authors

Matthias Kelm, Miguel Quiros, Veronica Azcutia, Kevin Boerner, Richard D. Cummings, Asma Nusrat, Jennifer C. Brazil, Charles A. Parkos

Longitudinal analysis of naturally acquired PfEMP1 CIDR domain variant antibodies identifies associations with malaria protection

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Abstract

Background. Malaria pathogenicity is determined, in part, by the adherence of Plasmodium falciparum infected erythrocytes to the microvasculature mediated via specific interactions between PfEMP1 variant domains to host endothelial receptors. Naturally acquired antibodies against specific PfEMP1 variants can play an important role in clinical protection against malaria. Methods. We evaluated IgG responses against a repertoire of PfEMP1 CIDR domain variants to determine the rate and order of variant-specific antibody acquisition and their association with protection against febrile malaria in a prospective cohort study conducted in an area of intense, seasonal malaria transmission. Results. Using longitudinal data, we found that IgG to the pathogenic domain variants CIDRα1.7 and CIDRα1.8 were acquired the earliest. Furthermore, IgG to CIDRγ3 was associated with reduced prospective risk of febrile malaria and recurrent malaria episodes. Conclusion. This study provides evidence that acquisition of IgG antibodies to PfEMP1 variants is ordered and demonstrates that antibodies to CIDRα1 domains are acquired the earliest in children residing in an area of intense, seasonal malaria transmission. Future studies will need to validate these findings in other transmission settings and determine the functional activity of these naturally acquired CIDR variant-specific antibodies. Funding. Division of Intramural Research, National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health.

Authors

Nyamekye Obeng-Adjei, Daniel B. Larremore, Louise Turner, Aissata Ongoiba, Shanping Li, Safiatou Doumbo, Takele B. Yazew, Kassoum Kayentao, Louis H Miller, Boubacar Traore, Susan K. Pierce, Caroline O. Buckee, Thomas Lavstsen, Peter D. Crompton, Tuan M. Tran

Differential effects of PD-L1 versus PD-1 blockade on myeloid inflammation in human cancer

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Abstract

Introduction: PD-1 and PD-L1 have been studied interchangeably in the clinic as checkpoints to reinvigorate T cells in diverse tumor types. Data for biologic effects of checkpoint blockade in human premalignancy are limited. Methods: We analyzed the immunologic effects of PD-L1 blockade in a clinical trial of atezolizumab in patients with asymptomatic multiple myeloma (AMM), a precursor to clinical malignancy. Genomic signatures of PD-L1 blockade in purified monocytes and T cells in vivo were also compared to those following PD-1 blockade in lung cancer patients. Effects of PD-L1 blockade on monocyte-derived dendritic cells were analyzed to better understand its effects on myeloid antigen-presenting cells. Results: In contrast to anti-PD-1 therapy, anti-PD-L1 therapy led to a distinct inflammatory signature in CD14+ monocytes and increase in myeloid-derived cytokines (e.g. IL-18) in vivo. Treatment of AMM patients with atezolizumab led to rapid activation and expansion of circulating myeloid cells which persisted in the bone marrow. Blockade of PD-L1 on purified monocyte-derived dendritic cells (DCs) led to rapid inflammasome activation and synergized with CD40L-driven DC maturation, leading to greater antigen-specific T cell expansion. Conclusions: These data show that PD-L1 blockade leads to distinct systemic immunologic effects compared to PD-1 blockade in vivo in humans, particularly manifest as rapid myeloid activation. These findings also suggest an additional role for PD-L1 as a checkpoint for regulating inflammatory phenotype of myeloid cells and antigen-presentation in DCs, which may be harnessed to improve PD-L1-based combination therapies. Trial Registration: NCT02784483 Funding: NCI CA197603, CA238471, CA208328

Authors

Noffar Bar, Federica Costa, Rituparna Das, Alyssa Duffy, Mehmet K. Samur, Samuel S. McCachren, Scott Gettinger, Natalia Neparidze, Terri L. Parker, Jithendra Kini Bailur, Katherine E. Pendleton, Richa Bajpai, Lin Zhang, Mina L. Xu, Tara Anderson, Nicola Giuliani, Ajay K. Nooka, Hearn J. Cho, Aparna Raval, Mala Shanmugam, Kavita M. Dhodapkar, Madhav Dhodapkar

Clinical and pathological investigation of severe COVID-19 patients

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Abstract

BACKGROUND. Severe acute respiratory coronavirus 2 (SARS-CoV-2) caused coronavirus disease 2019 (COVID-19) has become a pandemic. This study addressed the clinical and immunopathological characteristics of severe COVID-19. METHODS. Sixty-nine COVID-19 patients were classified into as severe and non-severe groups to analyze their clinical and laboratory characteristics. A panel of blood cytokines was quantified over time. Biopsy specimens from two deceased cases were obtained for immunopathological, ultrastructural, and in situ hybridization examinations. RESULTS. Circulating cytokines, including IL8, IL6, TNFα, IP10, MCP1, and RANTES, were significantly elevated in severe COVID-19 patients. Dynamic IL6 and IL8 were associated with disease progression. SARS-CoV-2 was demonstrated to infect type II, type I pneumocytes and endothelial cells, leading to severe lung damage through cell pyroptosis and apoptosis. In severe cases, lymphopenia, neutrophilia, depletion of CD4+ and CD8+ T lymphocytes, and massive macrophage and neutrophil infiltrates were observed in both blood and lung tissues. CONCLUSIONS. A panel of circulating cytokines could be used to predict disease deterioration and inform clinical interventions. Severe pulmonary damage was predominantly attributed to both SARS-CoV-2 caused cytopathy and immunopathologic damage. Strategies that encourage pulmonary recruitment and overactivation of inflammatory cells by suppressing cytokine storm might improve the outcomes of severe COVID-19 patients.

Authors

Shaohua Li, Lina Jiang, Xi Li, Fang Lin, Yijin Wang, Boan Li, Tianjun Jiang, Weimin An, Shuhong Liu, Hongyang Liu, Pengfei Xu, Lihua Zhao, Lixin Zhang, Jinsong Mu, Hongwei Wang, Jiarui Kang, Yan Li, Lei Huang, Caizhong Zhu, Shousong Zhao, Jiangyang Lu, Junsheng Ji, Jingmin Zhao

TrkB-agonists prevent post-ischemic emergence of refractory neonatal seizures in mice

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Abstract

Refractory neonatal seizures do not respond to first-line anti-seizure medications (ASMs) like phenobarbital (PB), a positive allosteric modulator for GABAA receptors. GABAA receptor-mediated inhibition is dependent upon electroneutral cation-chloride transporter KCC2 which mediates neuronal chloride extrusion and its age-dependent increase, postnatally shifts GABAergic signaling from depolarizing to hyperpolarizing. BDNF-TrkB activation following excitotoxic injury recruits downstream targets like PLCγ1, leading to KCC2 hypofunction. Here, the anti-seizure efficacy of TrkB agonists LM22A-4, HIOC, and Deoxygedunin (DG), on PB-refractory seizures, and post-ischemic TrkB-pathway activation was investigated in a mouse model (CD-1, P7) of refractory neonatal seizures. LM, a BDNF loop II mimetic, rescued PB-refractory seizures in a sexually dimorphic manner. Efficacy was associated with a significant reduction in the post-ischemic phosphorylation of TrkB at Y816, a site known to mediate post-ischemic KCC2 hypofunction via PLCγ1 activation. LM rescued ischemia-induced pKCC2-S940 dephosphorylation, preserving its membrane stability. Full TrkB agonists HIOC and DG similarly rescued PB-refractoriness. Chemogenetic inactivation of TrkB significantly reduced post-ischemic neonatal seizure burdens at P7. Sex differences identified in developmental expression profiles of TrkB and KCC2 may underlie the sexually dimorphic efficacy of LM. These results support a novel role for the TrkB receptor in the emergence of age-dependent refractory neonatal seizures.

Authors

Pavel A. Kipnis, Brennan J. Sullivan, Brandon M. Carter, Shilpa Kadam

MUC1-C drives stemness in progression of colitis to colorectal cancer

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Abstract

Colitis is associated with the development of colorectal cancer (CRC) by largely undefined mechanisms that are critical for understanding the link between inflammation and cancer. Intestinal stem cells (ISCs) marked by LGR5 expression are of importance in both the inflammatory response to colitis and progression to colitis-associated colon cancer (CACC). Here, we report in human MUC1-transgenic mouse models of CACC that targeting the MUC1-C oncogenic protein, which is upregulated in inflammation, suppresses the (i) Lgr5+ ISC population, (ii) induction of Myc and core pluripotency stem cell factors, and (iii) severity and progression of colitis to dysplasia and cancer. By extension to human colon cancer cells, we demonstrate that MUC1-C drives MYC, forms a complex with MYC on the LGR5 promoter and activates LGR5 expression. We also show in CRC cells that MUC1-C induces the cancer stem cell (CSC) markers (BMI1, ALDH1, FOXA1, LIN28B) and the OCT4, SOX2 and NANOG pluripotency factors. Consistent with conferring the CSC state, targeting MUC1-C suppresses the capacity of CRC cells to promote wound healing, invasion, self-renewal and tumorigenicity. In analysis of human tissues, MUC1 expression associates with activation of inflammatory pathways, development of colitis and aggressiveness of CRCs. These results collectively indicate that MUC1-C is of importance for integrating stemness and pluripotency in colitis and CRC. Of clinical relevance, the findings further indicate that MUC1-C represents a previously unrecognized target that is druggable for treating progression of colitis and CRC.

Authors

Wei Li, Ning Zhang, Caining Jin, Mark D. Long, Hasan Rajabi, Yota Yasumizu, Atsushi Fushimi, Nami Yamashita, Masayuki Hagiwara, Rongbin Zheng, Jin Wang, Ling Kui, Harpal Singh, Surender Kharbanda, Qiang Hu, Song Liu, Donald W. Kufe

Regulation of bile duct epithelial injury by hepatic CD71+ erythroid cells

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Abstract

Extramedullary hematopoietic cells are present in the liver of normal neonates in the first few days of life and persist in infants with biliary atresia. Based on a previous report that liver genes are enriched by erythroid pathways, we examined the liver gene expression pattern at diagnosis and found the top five enriched pathways are related to erythrocyte pathobiology in children who survived with the native liver beyond 2 years of age. Using immunostaining, anti-CD71 antibodies identified CD71+ erythroid cells among extramedullary hematopoietic cells in the livers at the time of diagnosis. In mechanistic experiments, the preemptive antibody depletion of hepatic CD71+ erythroid cells in neonatal mice rendered them resistant to rotavirus (RRV)-induced biliary atresia. The depletion of CD71+ erythroid cells increased the number of effector lymphocytes and delayed the RRV infection of livers and extrahepatic bile ducts. In co-culture experiments, CD71+ erythroid cells suppressed the activation of hepatic mononuclear cells. These data uncover an immunoregulatory role for CD71+ erythroid cells in the neonatal liver.

Authors

Li Yang, Pranavkumar Shivakumar, Jeremy M. Kinder, Sing Sing Way, Bryan Donnelly, Reena Mourya, Zhenhua Luo, Jorge A. Bezerra

Identification of pathogenic TRAIL-expressing innate immune cells during HIV-1 infection in humanized mice by scRNA-seq

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Abstract

Depletion of CD4+ T cells during HIV-1 infection is mostly mediated by inflammatory cells via indirect but not clearly defined mechanisms. In this report we employed the single-cell RNA sequencing (scRNA-seq) technology to study HIV-induced transcriptomic change in innate immune cells in lymphoid organs. We performed scRNA-seq on hCD45+hCD3-hCD19- human leukocytes isolated from spleens of humanized NOD/Rag2-/-/γc-/- (NRG) mice transplanted with human CD34+ hematopoietic stem-progenitor cells (NRG-hu HSC mice). We identified major populations of innate immune cells including plasmacytoid dendritic cells (pDC), myeloid dendritic cells (mDC), macrophage, nature killer (NK) cells and innate lymphoid cells (ILC). HIV-1 infection significantly upregulated genes involved in type-I interferon inflammatory pathways in each of the innate immune subsets. Interestingly, we found that the TNF-related apoptosis-inducing ligand (TRAIL) was upregulated in the innate immune populations including pDC, mDC, macrophage, NK and ILC. We further demonstrated that blockade of TRAIL signaling pathway in NRG-hu HSC mice prevented HIV-1 induced CD4+ T cell depletion in vivo. In summary, we characterized HIV-induced transcriptomic changes of innate immune cells in the spleen at single-cell levels, identified the TRAIL+ innate immune cells and defined an important role of TRAIL signaling pathway in HIV-1 induced CD4+ T cell depletion in vivo.

Authors

Liang Cheng, Haisheng Yu, John A. Wrobel, Guangming Li, Peng Liu, Zhiyuan Hu, Xiao-Ning Xu, Lishan Su