Donor NKG2C homozygosity contributes to CMV clearance after haploidentical transplantation

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Abstract

Cytomegalovirus (CMV) infection remains an important cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Several investigators have reported that adaptive NKG2C+ NK cells persistently expand during CMV reactivation. In our study, two cohorts were enrolled to explored the relationships among the NKG2C genotype, NKG2C+ NK cell reconstitution, and CMV infection. Multivariate analysis showed that donor NKG2C gene deletion was an independent prognostic factor for CMV reactivation and refractory CMV reactivation. Furthermore, the quantitative, qualitative reconstitution and anti-CMV function of adaptive NKG2C+ NK cells after transplantation was significantly lower in patients grafted with NKG2Cwt/del donor cells than in those grafted with NKG2Cwt/wt donor cells. The quantitative reconstitution of NKG2C+ NK cells at day 30 after transplantation was significantly lower in patients with treatment-refractory CMV reactivation than in those in the no-CMV-reactivation and CMV-reactivation groups. In humanized CMV-infected mice, we found that compared with those from NKG2Cwt/del donors, adaptive NKG2C+ NK cells from NKG2Cwt/wt donors induced earlier and stronger expansion of NKG2C+ NK cells and earlier and stronger CMV clearance in vivo. In conclusion, donor NKG2C homozygosity contributes to CMV clearance by promoting the quantitative and qualitative reconstruction of adaptive NKG2C+ NK cells after haploidentical allo-HSCT.

Authors

Xing-Xing Yu, Qian-Nan Shang, Xue-Fei Liu, Mei He, Xu-Ying Pei, Xiao-Dong Mo, Meng Lv, Ting-Ting Han, Ming-Rui Huo, Xiaosu Zhao, Ying-Jun Chang, Yu Wang, Xiao-Hui Zhang, Lan-Ping Xu, Kai-Yan Liu, Xiangyu Zhao, Xiaojun Huang

Fibrotic lung disease inhibits innate immune responses to Staphylococcal pneumonia via impaired neutrophil and macrophage function

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Abstract

Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal disease characterized by collagen deposition within the lung interstitium. Bacterial infection is associated with increased morbidity and more rapid mortality in IPF patient populations and pathogens such as methicillin-resistant Staphylococcus aureus (MRSA) are commonly isolated from the lungs of hospitalized IPF patients. Despite this, the effects of fibrotic lung injury on critical immune responses to infection remain unknown. In the present study, we show that, like human IPF, fibrotic mice infected with MRSA exhibit increased morbidity and mortality compared to uninfected fibrotic mice. We determine that fibrosis confers a defect in MRSA clearance compared to non-fibrotic mice, resulting from blunted innate immune responses. We show that fibrosis inhibits neutrophil intracellular killing of MRSA through impaired neutrophil elastase (NE) release and oxidative radical production. Additionally, we demonstrate that lung macrophages from fibrotic mice have impaired phagocytosis of MRSA. Our study describes potentially novel impairments to antimicrobial responses upon the development of pulmonary fibrosis and our findings suggest a possible mechanism for why IPF patients are at greater risk of morbidity and mortality related to infection.

Authors

Helen I. Warheit-Niemi, Summer J. Edwards, Shuvasree SenGupta, Carole A. Parent, Xiaofeng Zhou, David N. O'Dwyer, Bethany B. Moore

DDR1 contributes to kidney inflammation and fibrosis by promoting the phosphorylation of BCR and STAT3

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Abstract

Discoidin domain receptor 1 (DDR1), a receptor tyrosine kinase activated by collagen, contributes to chronic kidney disease (CKD). However, its role in acute kidney injury (AKI) and subsequent development of kidney fibrosis is not clear. Thus, we performed a model of severe ischemia-reperfusion-induced AKI that progresses to kidney fibrosis in wild-type and Ddr1-null mice. We show that Ddr1-null mice had reduced acute tubular injury, inflammation, and tubulointerstitial fibrosis with overall decreased renal monocyte chemoattractant protein (MCP-1) levels and STAT3 activation. We identified breakpoint cluster region (BCR) protein as a phosphorylated target of DDR1 that controls MCP-1 production in renal proximal tubule epithelial cells. DDR1-induced BCR phosphorylation or BCR downregulation increased MCP-1 secretion, suggesting that BCR negatively regulates the levels of MCP-1. Mechanistically, phosphorylation or downregulation of BCR increases β-catenin activity and in turn MCP-1 production. Finally, we show that DDR1-mediated STAT3 activation is required to stimulate the secretion of TGF-β. Thus, DDR1 contributes to acute and chronic kidney injury by regulating BCR and STAT3 phosphorylation and in turn the production of MCP-1 and TGF-β. These findings identify DDR1 an attractive therapeutic target for ameliorating both pro-inflammatory and pro-fibrotic signaling in kidney disease.

Authors

Corina M. Borza, Gema Bolas, Fabian Bock, Xiuqi Zhang, Favour C. Akabogu, Ming-Zhi Zhang, Mark de Caestecker, Min Yang, Hai-chun Yang, Ethan Lee, Leslie Gewin, Agnes B. Fogo, W. Hayes McDonald, Roy Zent, Ambra Pozzi

RB expression confers sensitivity to CDK4/6 inhibitor-mediated radiosensitization across breast cancer subtypes

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Abstract

Standard radiation (RT) therapy does not reliably provide locoregional control for women with multi-node positive and triple-negative (TNBC) breast cancers. We hypothesized that CDK4/6 inhibition (CDK4/6i) would increase the radiosensitivity not only of estrogen receptor positive (ER+) cells, but also TNBC that express retinoblastoma (RB) protein. We found that CDK4/6i radiosensitized RB wild-type TNBC (n=4, rER 1.49 – 2.22), but failed to radiosensitize RB-null TNBC (n=3, rER: 0.84 – 1.00). RB expression predicted response to CDK4/6i + RT (R2=0.84), and radiosensitization was lost in ER+/TNBC cells (rER: 0.88 – 1.13) after RB1 knockdown in isogenic and non-isogenic models. CDK4/6i suppressed homologous recombination (HR) in RB wild-type cells, but not in RB-null cells or isogenic models of RB1 loss; HR competency was rescued with RB re-expression. Radiosensitization was independent of non-homologous end joining and the known effects of CDK4/6i on cell cycle arrest. Mechanistically, RB and RAD51 interact in vitro to promote HR repair. CDK4/6i produced RB-dependent radiosensitization in TNBC xenografts, but not in isogenic RB1-null xenografts. Our data provide the preclinical rationale for a clinical trial expanding the use of CDK4/6i + RT to difficult to control RB-intact breast cancers (including TNBC) and nominate RB status as a predictive biomarker of therapeutic efficacy.

Authors

Andrea M. Pesch, Nicole H. Hirsh, Anna R. Michmerhuizen, Kassidy M. Jungles, Kari Wilder-Romans, Benjamin C. Chandler, Meilan Liu, Lynn M. Lerner, Charles A. Nino, Connor Ward, Erin F. Cobain, Theodore S. Lawrence, Lori J. Pierce, James M. Rae, Corey W. Speers

Stromal architecture directs early dissemination in pancreatic ductal adenocarcinoma

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Abstract

Pancreatic ductal adenocarcinoma (PDA) is an extremely metastatic and lethal disease. Here in both murine and human PDA we demonstrate that extracellular matrix architecture regulates cell extrusion and subsequent invasion from intact ductal structures through Tumor-Associated Collagen Signatures (TACS). This results in early dissemination from histologically pre-malignant lesions and continual invasion from well-differentiated disease, and suggests TACS as a biomarker to aid in the pathologic assessment of early disease. Furthermore, we show that pancreatitis results in invasion-conducive architectures, thus priming the stroma prior to malignant disease. Analysis in novel microfluidics-derived microtissues and in vivo demonstrates decreased extrusion and invasion following focal adhesion kinase (FAK) inhibition, consistent with decreased metastasis. Thus, data suggest that targeting FAK or strategies to re-engineer and normalize tumor microenvironments, may have a role not only in also in very early disease but also for limiting continued dissemination from unresectable disease. Likewise, it may be beneficial to employ stroma targeting strategies to resolve precursor diseases such as pancreatitis in order to remove stromal architectures that increase risk for early dissemination.

Authors

Arja Ray, Mackenzie K. Callaway, Nelson J. Rodríguez-Merced, Alexandra L. Crampton, Marjorie Carlson, Kenneth B. Emme, Ethan A. Ensminger, Alexander A. Kinne, Jonathan H. Schrope, Haley R. Rasmussen, Hong Jiang, David G. DeNardo, David K. Wood, Paolo P. Provenzano

Epac1-/- and Epac2-/- mice exhibit deficient epithelial Na⁺ channel regulation and impaired urinary Na⁺ conservation

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Abstract

Exchange proteins directly activated by cAMP (Epacs) are abundantly expressed in the renal tubules. We used genetic and pharmacological tools in combination with balance, electrophysiological and biochemical approaches to examine the role of Epac1 and Epac2 in renal sodium handling. We demonstrate that Epac1-/- and Epac2-/- mice exhibit a delayed anti-natriuresis to dietary sodium restriction despite augmented aldosterone levels. This was associated with a significantly lower response to the epithelial Na+ channel (ENaC) blocker amiloride, reduced ENaC activity in split-opened collecting ducts, and defective posttranslational processing of α and γENaC subunits in the knockout mice fed with Na+ deficient diet. Concomitant deletion of both isoforms led to a marginally greater natriuresis but further increased aldosterone levels. Epac2 blocker, ESI-05 and Epac1&2 blocker, ESI-09 decreased ENaC activity in EpacWT mice kept on Na+ deficient diet but not on the regular diet. ESI-09 injections led to natriuresis in EpacWT mice on Na+ deficient diet, which was caused by ENaC inhibition. In summary, our results demonstrate non-redundant actions of Epac1 and Epac2 in stimulation of ENaC activity during variations in dietary salt intake. We speculate that inhibition of Epac signaling could be instrumental in treatment of hypertensive states associated with ENaC over-activation.

Authors

Victor N. Tomilin, Kyrylo Pyrshev, Anna Stavniichuk, Naghmeh Hassanzadeh Khayyat, Guohui Ren, Oleg Zaika, Sherif Khedr, Alexander Staruschenko, Fang C. Mei, Xiaodong Cheng, Oleh Pochynyuk

Hypoxic vasodilatory defect and pulmonary hypertension in mice lacking hemoglobin β-cysteine93 S-nitrosylation

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Abstract

Systemic hypoxia is characterized by peripheral vasodilation and pulmonary vasoconstriction. However, the system-wide mechanism for signaling hypoxia remains unknown. Accumulating evidence suggests that hemoglobin in RBCs may serve as an O2 sensor and O2-responsive NO signal transducer to regulate systemic and pulmonary vascular tone, but this remains unexamined at the integrated system level. One residue invariant in mammalian hemoglobins (Hb), β-globin Cys93 (βCys93), carries NO as vasorelaxant S-nitrosothiol (SNO) to autoregulate blood flow during oxygen delivery. βCys93Ala mutant mice thus exhibit systemic hypoxia despite transporting oxygen normally. Here we show that βCys93Ala mutant mice have reduced S-nitrosohemoglobin (SNO-Hb) at baseline and upon targeted SNO repletion, and that hypoxic vasodilation by RBCs is impaired in vitro and in vivo, recapitulating hypoxic pathophysiology. Notably, βCys93Ala mutant mice show marked impairment of hypoxic peripheral vasodilation and develop signs of pulmonary hypertension with age. Mutant mice also die prematurely with cor pulmonale (pulmonary hypertension with right ventricular dysfunction) when living under low oxygen. Altogether, we identify a major role for RBC-SNO in clinically-relevant vasodilatory responses attributed previously to endothelial NO. We conclude that SNO-Hb transduces the integrated, system-wide response to hypoxia in the mammalian respiratory cycle, expanding a core physiological principle.

Authors

Rongli Zhang, Alfred Hausladen, Zhaoxia Qian, Xudong Liao, Richard T. Premont, Jonathan S. Stamler

Peripheral ablation of type Ⅲ adenylyl cyclase induces hyperalgesia and eliminates KOR-mediated analgesia in mice

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Abstract

Ca2+/calmodulin-stimulated group Ⅰ adenylyl cyclase (AC) isoforms AC1 and AC8 have been involved in nociceptive processing and morphine responses. However, whether AC3, another member of group I ACs, is involved in nociceptive transmission and regulates opioid receptor signaling remain elusive. Here we report that conditional knockout of AC3 (AC3CKO) in L3 and L4 DRGs robustly facilitates the mouse nociceptive responses, decreases voltage-gated potassium (Kv) channel currents and increases neuronal excitability. Also, AC3CKO eliminates the analgesic effect of κ opioid receptor (KOR) agonist and its inhibition on Kv channel by classical Gαi/o signaling or nonclassical direct interaction of KOR and AC3 proteins. Interestingly, significantly upregulated AC1 level and cAMP concentration are detected in AC3 deficient DRGs. Inhibition of AC1 completely reversed cAMP upregulation, neuronal excitability enhancement and nociceptive behavioral hypersensitivity in AC3CKO mice. Our findings suggest a crucial role of peripheral AC3 in nociceptive modulation and KOR opioid analgesia.

Authors

Wen-Wen Zhang, Hong Cao, Yang Li, Xian-Jun Fu, Yu-Qiu Zhang

Ceramide-mediated gut dysbiosis enhances cholesterol esterification and promotes colorectal tumorigenesis in mice

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Abstract

Colorectal cancer (CRC) very severely threatens human health and lifespan. Effective therapeutic strategy has not been established because of without clearly knowing its pathogenesis. Here we report that ceramide and SOAT1 have the roles on both spontaneous and chemical-induced intestine cancers. It was first found by us that miR-148a deficiency dramatically increased mouse gut dysbiosis through upregulating Cers5 expression, which promoted ceramide synthesis afterward. The newly generated ceramide further promoted both AOM/DSS-induced and ApcMin/+ spontaneous intestine tumorigenesis via increasing mouse gut dysbiosis. Meanwhile, increased level of ceramide correlated with the significant enhancements on both β-catenin activity and colorectal tumorigenesis in a fashion of TLR4-dependent. Next, it was found that a direct binding of β-catenin to SOAT1 promoter to activate transcriptional expression of SOAT1, which further induced cholesterol esterification and colorectal tumorigenesis. In human patients of CRC, the same CERS5-TLR4-β-catenin-SOAT1 axis was also found with dysregulation. Finally, the SOAT1 inhibitor (Avasimibe) showed the significant levels of therapeutic effects on both AOM/DSS-induced and ApcMin/+ spontaneous intestine cancer. Our study clarified that ceramide promoted CRC development through increasing gut dysbiosis, further resulting in the increase of cholesterol esterification in a special way of SOAT1-dependent. The treatment through Avasimibe to specifically decrease cholesterol esterification could be considered as a clinical strategy for effective CRC therapy in future study.

Authors

Yahui Zhu, Li Gu, Xi Lin, Jinmiao Zhang, Yi Tang, Xinyi Zhou, Bingjun Lu, Xingrong Lin, Cheng Liu, Edward V. Prochownik, Youjun Li

The RNA binding protein IMP2 drives a stromal-Th17 cell circuit in autoimmune neuroinflammation

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Abstract

Stromal cells are emerging as key drivers of autoimmunity, in part by producing inflammatory chemokines that orchestrate inflammation. Chemokine expression is regulated transcriptionally but also through post-transcriptional mechanisms, the specific drivers of which are still incompletely defined. CCL2 (MCP1) is a multifunctional chemokine that drives myeloid cell recruitment. During experimental autoimmune encephalomyelitis (EAE), an IL-17-driven model of multiple sclerosis, CCL2 produced by lymph node (LN) stromal cells is essential for immunopathology. Here, we show that Ccl2 mRNA upregulation in human stromal fibroblasts in response to IL-17 requires the RNA binding protein (RBP) insulin like growth factor 2 mRNA binding protein 2 (IGF2BP2, IMP2), which is expressed almost exclusively in non-hematopoietic cells. IMP2 binds directly to CCL2 mRNA, markedly extending its transcript half-life and thus required for efficient CCL2 secretion. Consistent with this, Imp2−/− mice showed reduced CCL2 production in LN during EAE, causing impairments in monocyte recruitment and Th17 cell polarization. Imp2-/- mice were fully protected from CNS inflammation. Moreover, deletion of IMP2 after EAE onset was sufficient to mitigate disease severity. These data show that posttranscriptional control of Ccl2 in stromal cells by IMP2 is required to permit IL-17-driven progression of EAE pathogenesis.

Authors

Rami Bechara, Nilesh Amatya, Saikat Majumder, Chunsheng Zhou, Yang Li, Qixing Liu, Mandy J. McGeachy, Sarah L. Gaffen