The use of broad-spectrum antibiotics in empirical antimicrobial therapy is a lifesaving strategy for patients in intensive care. At the same time, antibiotics dramatically increase the risk for nosocomial infections, such as hospital‑acquired pneumonia caused by Pseudomonas aeruginosa, and other antibiotic-resistant bacteria. In this issue of the JCI, Robak and colleagues identified a mechanism by which depletion of resident gut and lung microbiota by antibiotic treatment results in secondary IgA deficiency and impaired anti–P. aeruginosa host defense. Impaired defenses could be improved by substitution of polyclonal IgA via the intranasal route in a mouse model of pneumonia. Importantly, antibiotic treatment caused lung IgA deficiency that involved reduced TLR-dependent production of a proliferation-inducing ligand (APRIL) and B cell–activating factor (BAFF) in intensive care unit patients. These patients might therefore benefit from future strategies to increase pulmonary IgA levels.
Juergen Lohmeyer, Rory E. Morty, Susanne Herold
BACKGROUND. Recombinant leptin (metreleptin) ameliorates hyperphagia and metabolic abnormalities in leptin-deficient humans with lipodystrophy. We aimed to determine whether metreleptin improves glucose and lipid metabolism in humans when food intake is held constant. METHODS. Patients with lipodystrophy were hospitalized for 19 days, with food intake held constant by a controlled diet in an inpatient metabolic ward. In a nonrandomized, crossover design, patients previously treated with metreleptin (n = 8) were continued on metreleptin for 5 days and then taken off metreleptin for the next 14 days (withdrawal cohort). This order was reversed in metreleptin-naive patients (n = 14), who were reevaluated after 6 months of metreleptin treatment on an ad libitum diet (initiation cohort). Outcome measurements included insulin sensitivity by hyperinsulinemic-euglycemic clamp, fasting glucose and triglyceride levels, lipolysis measured using isotopic tracers, and liver fat by magnetic resonance spectroscopy. RESULTS. With food intake constant, peripheral insulin sensitivity decreased by 41% after stopping metreleptin for 14 days (withdrawal cohort) and increased by 32% after treatment with metreleptin for 14 days (initiation cohort). In the initiation cohort only, metreleptin decreased fasting glucose by 11% and triglycerides by 41% and increased hepatic insulin sensitivity. Liver fat decreased from 21.8% to 18.7%. In the initiation cohort, changes in lipolysis were not independent of food intake, but after 6 months of metreleptin treatment on an ad libitum diet, lipolysis decreased by 30% (palmitate turnover) to 35% (glycerol turnover). CONCLUSION. Using lipodystrophy as a human model of leptin deficiency and replacement, we show that metreleptin improves insulin sensitivity and decreases hepatic and circulating triglycerides and that these improvements are independent of its effects on food intake. TRIAL REGISTRATION. ClinicalTrials.gov NCT01778556 FUNDING. This research was supported by the intramural research program of the NIDDK.
Rebecca J. Brown, Areli Valencia, Megan Startzell, Elaine Cochran, Peter J. Walter, H. Martin Garraffo, Hongyi Cai, Ahmed M. Gharib, Ronald Ouwerkerk, Amber B. Courville, Shanna Bernstein, Robert J. Brychta, Kong Y. Chen, Mary Walter, Sungyoung Auh, Phillip Gorden
Resolution of inflammation is a critical process that is facilitated by specialized proresolving mediators (SPMs). In this issue, Bang et al. show that the G protein–coupled receptor GPR37 is a receptor for one such SPM, neuroprotectin D1. They also show that GPR37 activation in macrophages enhances phagocytosis, shifts cytokine release toward an antiinflammatory profile, and thereby helps to reverse inflammatory pain.
Lintao Qu, Michael J. Caterina
Donna M. Martin, W. Kimryn Rathmell, Sohail F. Tavazoie
The biological activity of 24R,25-dihydroxyvitamin D3 [24R,25(OH)2D3] remains controversial, but it has been suggested that it contributes to fracture healing. Cyp24a1–/– mice, synthesizing no 24R,25(OH)2D3, show suboptimal endochondral ossification during fracture repair, with smaller callus and reduced stiffness. These defects were corrected by 24R,25(OH)2D3 treatment, but not by 1,25-dihydroxyvitamin D3. Microarrays with Cyp24a1–/– callus mRNA identified FAM57B2 as a mediator of the 24R,25(OH)2D3 effect. FAM57B2 produced lactosylceramide (LacCer) upon specific binding of 24R,25(OH)2D3. Fam57b inactivation in chondrocytes (Col2-Cre Fam57bfl/fl) phenocopied the callus formation defect of Cyp24a1–/– mice. LacCer or 24R,25(OH)2D3 injections restored callus volume, stiffness, and mineralized cartilage area in Cyp24a1-null mice, but only LacCer rescued Col2-Cre Fam57bfl/fl mice. Gene expression in callus tissue suggested that the 24R,25(OH)2D3/FAM57B2 cascade affects cartilage maturation. We describe a previously unrecognized pathway influencing endochondral ossification during bone repair through LacCer production upon binding of 24R,25(OH)2D3 to FAM57B2. Our results identify potential new approaches to ameliorate fracture healing.
Corine Martineau, Roy Pascal Naja, Abdallah Husseini, Bachar Hamade, Martin Kaufmann, Omar Akhouayri, Alice Arabian, Glenville Jones, René St-Arnaud
Chromatin remodeler Brahma related gene 1 (BRG1) is silenced in approximately 10% of human pancreatic ductal adenocarcinomas (PDAs). We previously showed that BRG1 inhibits the formation of intraductal pancreatic mucinous neoplasm (IPMN) and that IPMN-derived PDA originated from ductal cells. However, the role of BRG1 in pancreatic intraepithelial neoplasia–derived (PanIN-derived) PDA that originated from acinar cells remains elusive. Here, we found that exclusive elimination of Brg1 in acinar cells of Ptf1a-CreER; KrasG12D; Brg1fl/fl mice impaired the formation of acinar-to-ductal metaplasia (ADM) and PanIN independently of p53 mutation, while PDA formation was inhibited in the presence of p53 mutation. BRG1 bound to regions of the Sox9 promoter to regulate its expression and was critical for recruitment of upstream regulators, including PDX1, to the Sox9 promoter and enhancer in acinar cells. SOX9 expression was downregulated in BRG1-depleted ADMs/PanINs. Notably, Sox9 overexpression canceled this PanIN-attenuated phenotype in KBC mice. Furthermore, Brg1 deletion in established PanIN by using a dual recombinase system resulted in regression of the lesions in mice. Finally, BRG1 expression correlated with SOX9 expression in human PDAs. In summary, BRG1 is critical for PanIN initiation and progression through positive regulation of SOX9. Thus, the BRG1/SOX9 axis is a potential target for PanIN-derived PDA.
Motoyuki Tsuda, Akihisa Fukuda, Nilotpal Roy, Yukiko Hiramatsu, Laura Leonhardt, Nobuyuki Kakiuchi, Kaja Hoyer, Satoshi Ogawa, Norihiro Goto, Kozo Ikuta, Yoshito Kimura, Yoshihide Matsumoto, Yutaka Takada, Takuto Yoshioka, Takahisa Maruno, Yuichi Yamaga, Grace E. Kim, Haruhiko Akiyama, Seishi Ogawa, Christopher V. Wright, Dieter Saur, Kyoichi Takaori, Shinji Uemoto, Matthias Hebrok, Tsutomu Chiba, Hiroshi Seno
Purinergic receptor-7 (P2X7R) signaling controls Th17 and Th1 generation/differentiation, while NOD-like receptor P3 (NLRP3) acts as a Th2 transcriptional factor. Here, we demonstrated the existence of a P2X7R/NLRP3 pathway in T cells that is dysregulated by a P2X7R intracellular region loss-of-function mutation, leading to NLRP3 displacement and to excessive Th17 generation due to abrogation of the NLRP3-mediated Th2 program. This ultimately resulted in poor outcomes in cardiac-transplanted patients carrying the mutant allele, who showed abnormal Th17 generation. Transient NLRP3 silencing in nonmutant T cells or overexpression in mutant T cells normalized the Th profile. Interestingly, IL-17 blockade reduced Th17 skewing of human T cells in vitro and abrogated the severe allograft vasculopathy and abnormal Th17 generation observed in preclinical models in which P2X7R was genetically deleted. This P2X7R intracellular region mutation thus impaired the modulatory effects of P2X7R on NLRP3 expression and function in T cells and led to NLRP3 dysregulation and Th17 skewing, delineating a high-risk group of cardiac-transplanted patients who may benefit from personalized therapy.
Francesca D’Addio, Andrea Vergani, Luciano Potena, Anna Maestroni, Vera Usuelli, Moufida Ben Nasr, Roberto Bassi, Sara Tezza, Sergio Dellepiane, Basset El Essawy, Maria Iascone, Attilio Iacovoni, Laura Borgese, Kaifeng Liu, Gary Visner, Sirano Dhe-Paganon, Domenico Corradi, Reza Abdi, Randall C. Starling, Franco Folli, Gian Vincenzo Zuccotti, Mohamed H. Sayegh, Peter S. Heeger, Anil Chandraker, Francesco Grigioni, Paolo Fiorina
Brandon M. Fox, Alexander J. Adami, Travis D. Hull
Brown et al. report that two weeks of exogenous leptin administration to leptin-naive individuals with lipodystrophy resulted in increased energy expenditure and lipolysis, decreased ectopic liver fat, improved hepatic and peripheral insulin sensitivity, and attenuated dyslipidemia. Leptin withdrawal in individuals with lipodystrophy did not produce reciprocal effects on these phenotypes and resulted in significant improvements only in hepatic insulin sensitivity. This asymmetry in responses to leptin initiation and cessation is consistent with the other aspects of leptin biology that are dependent on the metabolic context in which this adipocyte-derived hormone functions.
Michael Rosenbaum, Rudolph L. Leibel
In type 1 diabetes, cytotoxic CD8+ T cells with specificity for β cell autoantigens are found in the pancreatic islets, where they are implicated in the destruction of insulin-secreting β cells. In contrast, the disease relevance of β cell–reactive CD8+ T cells that are detectable in the circulation, and their relationship to β cell function, are not known. Here, we tracked multiple, circulating β cell–reactive CD8+ T cell subsets and measured β cell function longitudinally for 2 years, starting immediately after diagnosis of type 1 diabetes. We found that change in β cell–specific effector memory CD8+ T cells expressing CD57 was positively correlated with C-peptide change in subjects below 12 years of age. Autoreactive CD57+ effector memory CD8+ T cells bore the signature of enhanced effector function (higher expression of granzyme B, killer-specific protein of 37 kDa, and CD16, and reduced expression of CD28) compared with their CD57– counterparts, and network association modeling indicated that the dynamics of β cell–reactive CD57+ effector memory CD8+ T cell subsets were strongly linked. Thus, coordinated changes in circulating β cell–specific CD8+ T cells within the CD57+ effector memory subset calibrate to functional insulin reserve in type 1 diabetes, providing a tool for immune monitoring and a mechanism-based target for immunotherapy.
Lorraine Yeo, Alyssa Woodwyk, Sanjana Sood, Anna Lorenc, Martin Eichmann, Irma Pujol-Autonell, Rosella Melchiotti, Ania Skowera, Efthymios Fidanis, Garry M. Dolton, Katie Tungatt, Andrew K. Sewell, Susanne Heck, Alka Saxena, Craig A. Beam, Mark Peakman
Mutations in superoxide dismutase 1 (SOD1) are responsible for 20% of familial ALS. Given the gain of toxic function in this dominantly inherited disease, lowering SOD1 mRNA and protein is predicted to provide therapeutic benefit. An early generation antisense oligonucleotide (ASO) targeting SOD1 was identified and tested in a phase I human clinical trial, based on modest protection in animal models of SOD1 ALS. Although the clinical trial provided encouraging safety data, the drug was not advanced because there was progress in designing other, more potent ASOs for CNS application. We have developed next-generation SOD1 ASOs that more potently reduce SOD1 mRNA and protein and extend survival by more than 50 days in SOD1G93A rats and by almost 40 days in SOD1G93A mice. We demonstrated that the initial loss of compound muscle action potential in SOD1G93A mice is reversed after a single dose of SOD1 ASO. Furthermore, increases in serum phospho-neurofilament heavy chain levels, a promising biomarker for ALS, are stopped by SOD1 ASO therapy. These results define a highly potent, new SOD1 ASO ready for human clinical trial and suggest that at least some components of muscle response can be reversed by therapy.
Alex McCampbell, Tracy Cole, Amy J. Wegener, Giulio S. Tomassy, Amy Setnicka, Brandon J. Farley, Kathleen M. Schoch, Mariah L. Hoye, Mark Shabsovich, Linhong Sun, Yi Luo, Mingdi Zhang, Sai Thankamony, David W. Salzman, Merit Cudkowicz, Danielle L. Graham, C. Frank Bennett, Holly B. Kordasiewicz, Eric E. Swayze, Timothy M. Miller
Broad-spectrum antibiotics are widely used with patients in intensive care units (ICUs), many of whom develop hospital-acquired infections with Pseudomonas aeruginosa. Although preceding antimicrobial therapy is known as a major risk factor for P. aeruginosa–induced pneumonia, the underlying mechanisms remain incompletely understood. Here we demonstrate that depletion of the resident microbiota by broad-spectrum antibiotic treatment inhibited TLR-dependent production of a proliferation-inducing ligand (APRIL), resulting in a secondary IgA deficiency in the lung in mice and human ICU patients. Microbiota-dependent local IgA contributed to early antibacterial defense against P. aeruginosa. Consequently, P. aeruginosa–binding IgA purified from lamina propria culture or IgA hybridomas enhanced resistance of antibiotic-treated mice to P. aeruginosa infection after transnasal substitute. Our study provides a mechanistic explanation for the well-documented risk of P. aeruginosa infection following antimicrobial therapy, and we propose local administration of IgA as a novel prophylactic strategy.
Oliver H. Robak, Markus M. Heimesaat, Andrey A. Kruglov, Sandra Prepens, Justus Ninnemann, Birgitt Gutbier, Katrin Reppe, Hubertus Hochrein, Mark Suter, Carsten J. Kirschning, Veena Marathe, Jan Buer, Mathias W. Hornef, Markus Schnare, Pascal Schneider, Martin Witzenrath, Stefan Bereswill, Ulrich Steinhoff, Norbert Suttorp, Leif E. Sander, Catherine Chaput, Bastian Opitz
The mechanisms of pain induction by inflammation have been extensively studied. However, the mechanisms of pain resolution are not fully understood. Here, we report that GPR37, expressed by macrophages (MΦs) but not microglia, contributes to the resolution of inflammatory pain. Neuroprotectin D1 (NPD1) and prosaptide TX14 increase intracellular Ca2+ (iCa2+) levels in GPR37-transfected HEK293 cells. NPD1 and TX14 also bind to GPR37 and cause GPR37-dependent iCa2+ increases in peritoneal MΦs. Activation of GPR37 by NPD1 and TX14 triggers MΦ phagocytosis of zymosan particles via calcium signaling. Hind paw injection of pH-sensitive zymosan particles not only induces inflammatory pain and infiltration of neutrophils and MΦs, but also causes GPR37 upregulation in MΦs, phagocytosis of zymosan particles and neutrophils by MΦs in inflamed paws, and resolution of inflammatory pain in WT mice. Mice lacking Gpr37 display deficits in MΦ phagocytic activity and delayed resolution of inflammatory pain. Gpr37-deficient MΦs also show dysregulations of proinflammatory and antiinflammatory cytokines. MΦ depletion delays the resolution of inflammatory pain. Adoptive transfer of WT but not Gpr37-deficient MΦs promotes the resolution of inflammatory pain. Our findings reveal a previously unrecognized role of GPR37 in regulating MΦ phagocytosis and inflammatory pain resolution.
Sangsu Bang, Ya-Kai Xie, Zhi-Jun Zhang, Zilong Wang, Zhen-Zhong Xu, Ru-Rong Ji
Clarin-1, a tetraspan-like membrane protein defective in Usher syndrome type IIIA (USH3A), is essential for hair bundle morphogenesis in auditory hair cells. We report a new synaptic role for clarin-1 in mouse auditory hair cells elucidated by characterization of Clrn1 total (Clrn1ex4–/–) and postnatal hair cell–specific conditional (Clrn1ex4fl/fl Myo15-Cre+/–) knockout mice. Clrn1ex4–/– mice were profoundly deaf, whereas Clrn1ex4fl/fl Myo15-Cre+/– mice displayed progressive increases in hearing thresholds, with, initially, normal otoacoustic emissions and hair bundle morphology. Inner hair cell (IHC) patch-clamp recordings for the 2 mutant mice revealed defective exocytosis and a disorganization of synaptic F-actin and CaV1.3 Ca2+ channels, indicative of a synaptopathy. Postsynaptic defects were also observed, with an abnormally broad distribution of AMPA receptors associated with a loss of afferent dendrites and defective electrically evoked auditory brainstem responses. Protein-protein interaction assays revealed interactions between clarin-1 and the synaptic CaV1.3 Ca2+ channel complex via the Cavβ2 auxiliary subunit and the PDZ domain–containing protein harmonin (defective in Usher syndrome type IC). Cochlear gene therapy in vivo, through adeno-associated virus–mediated Clrn1 transfer into hair cells, prevented the synaptic defects and durably improved hearing in Clrn1ex4fl/fl Myo15-Cre+/– mice. Our results identify clarin-1 as a key organizer of IHC ribbon synapses, and suggest new treatment possibilities for USH3A patients.
Didier Dulon, Samantha Papal, Pranav Patni, Matteo Cortese, Philippe F.Y. Vincent, Margot Tertrais, Alice Emptoz, Abdelaziz Tlili, Yohan Bouleau, Vincent Michel, Sedigheh Delmaghani, Alain Aghaie, Elise Pepermans, Olinda Alegria-Prevot, Omar Akil, Lawrence Lustig, Paul Avan, Saaid Safieddine, Christine Petit, Aziz El-Amraoui
Cancer progression is associated with alterations of intra- and extramedullary hematopoiesis to support a systemic tumor-promoting myeloid response. However, the functional specialty, mechanism, and clinical relevance of extramedullary hematopoiesis (EMH) remain unclear. Here, we showed that the heightened splenic myelopoiesis in tumor-bearing hosts was not only characterized by the accumulation of myeloid precursors, but also associated with profound functional alterations of splenic early hematopoietic stem/progenitor cells (HSPCs). With the distinct capability to produce and respond to granulocyte-macrophage CSF (GM-CSF), these splenic HSPCs were “primed” and committed to generating immunosuppressive myeloid cells. Mechanistically, the CCL2/CCR2 axis–dependent recruitment and the subsequent local education by the splenic stroma were critical for eliciting this splenic HSPC response. Selective abrogation of this splenic EMH was sufficient to synergistically enhance the therapeutic efficacy of immune checkpoint blockade. Clinically, patients with different types of solid tumors exhibited increased splenic HSPC levels associated with poor survival. These findings reveal a unique and important role of splenic hematopoiesis in tumor-associated myelopoiesis.
Chong Wu, Huiheng Ning, Mingyu Liu, Jie Lin, Shufeng Luo, Wenjie Zhu, Jing Xu, Wen-Chao Wu, Jing Liang, Chun-Kui Shao, Jiaqi Ren, Bin Wei, Jun Cui, Min-Shan Chen, Limin Zheng
The motor neuron disease spinal muscular atrophy (SMA) is caused by recessive, loss-of-function mutations of the survival motor neuron 1 gene (SMN1). Alone, such mutations are embryonically lethal, but SMA patients retain a paralog gene, SMN2, that undergoes alternative pre-mRNA splicing, producing low levels of SMN protein. By mechanisms that are not well understood, reduced expression of the ubiquitously expressed SMN protein causes an early-onset motor neuron disease that often results in infantile or childhood mortality. Recently, striking clinical improvements have resulted from two novel treatment strategies to increase SMN protein by (a) modulating the splicing of existing SMN2 pre-mRNAs using antisense oligonucleotides, and (b) transducing motor neurons with self-complementary adeno-associated virus 9 (scAAV9) expressing exogenous SMN1 cDNA. We review the recently published clinical trial results and discuss the differing administration, tissue targeting, and potential toxicities of these two therapies. We also focus on the challenges that remain, emphasizing the many clinical and biologic questions that remain open. Answers to these questions will enable further optimization of these remarkable SMA treatments as well as provide insights that may well be useful in application of these therapeutic platforms to other diseases.
Charlotte J. Sumner, Thomas O. Crawford
Neuronatin (Nnat) is an imprinted gene implicated in human obesity and widely expressed in neuroendocrine and metabolic tissues in a hormone- and nutrient-sensitive manner. However, its molecular and cellular functions and precise role in organismal physiology remain only partly defined. Here we demonstrate that mice lacking Nnat globally or specifically in β cells display impaired glucose-stimulated insulin secretion leading to defective glucose handling under conditions of nutrient excess. In contrast, we report no evidence for any feeding or body weight phenotypes in global Nnat-null mice. At the molecular level neuronatin augments insulin signal peptide cleavage by binding to the signal peptidase complex and facilitates translocation of the nascent preprohormone. Loss of neuronatin expression in β cells therefore reduces insulin content and blunts glucose-stimulated insulin secretion. Nnat expression, in turn, is glucose-regulated. This mechanism therefore represents a novel site of nutrient-sensitive control of β cell function and whole-animal glucose homeostasis. These data also suggest a potential wider role for Nnat in the regulation of metabolism through the modulation of peptide processing events.
Steven J. Millership, Gabriela Da Silva Xavier, Agharul I. Choudhury, Sergio Bertazzo, Pauline Chabosseau, Silvia M.A. Pedroni, Elaine E. Irvine, Alex Montoya, Peter Faull, William R. Taylor, Julie Kerr-Conte, Francois Pattou, Jorge Ferrer, Mark Christian, Rosalind M. John, Mathieu Latreille, Ming Liu, Guy A. Rutter, James Scott, Dominic J. Withers
Crescentic glomerulonephritis, a complication of severe immune glomerular injury, is the pathological correlate of rapidly progressive glomerulonephritis, mediated by both humoral and cellular effectors. In the current issue of the JCI, Chen et al. have implicated Bowman’s capsule in functionally isolating potentially immune effectors, specifically antigen-specific CD8+ T lymphocytes, from podocytes. They suggest that, in crescentic glomerulonephritis, immune-mediated glomerular endothelial injury results in inside-out injury to the glomerulus, with subsequent leukocyte migration through a weakened or ruptured Bowman’s capsule, resulting in outside-in injury. Effector T cells then recognize nephritogenic antigens presented by podocytes or other cells within the urinary space, enhancing injury and crescent formation.
A. Richard Kitching, Maliha A. Alikhan
T cells play a key role in immune-mediated glomerulonephritis, but how cytotoxic T cells interact with podocytes remains unclear. To address this, we injected EGFP-specific CD8+ T cells from just EGFP death inducing (Jedi) mice into transgenic mice with podocyte-specific expression of EGFP. In healthy mice, Jedi T cells could not access EGFP+ podocytes. Conversely, when we induced nephrotoxic serum nephritis (NTSN) and injected Jedi T cells, EGFP+ podocyte transgenic mice showed enhanced proteinuria and higher blood urea levels. Morphometric analysis showed greater loss of EGFP+ podocytes, which was associated with severe crescentic and necrotizing glomerulonephritis. Notably, only glomeruli with disrupted Bowman’s capsule displayed massive CD8+ T cell infiltrates that were in direct contact with EGFP+ podocytes, causing their apoptosis. Thus, under control conditions with intact Bowman’s capsule, podocytes are not accessible to CD8+ T cells. However, breaches in Bowman’s capsule, as also noted in human crescentic glomerulonephritis, allow access of CD8+ T cells to the glomerular tuft and podocytes, resulting in their destruction. Through these mechanisms, a potentially reversible glomerulonephritis undergoes an augmentation process to a rapidly progressive glomerulonephritis, leading to end-stage kidney disease. Translating these mechanistic insights to human crescentic nephritis should direct future therapeutic interventions at blocking CD8+ T cells, especially in progressive stages of rapidly progressive glomerulonephritis.
Anqun Chen, Kyung Lee, Vivette D. D’Agati, Chengguo Wei, Jia Fu, Tian-Jun Guan, John Cijiang He, Detlef Schlondorff, Judith Agudo