Issue published October 8, 2021

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Loss of miR-29a/b1 promotes inflammation and fibrosis in acute pancreatitis

In this issue, Dey et al. report that depletion of miR-29a/b1 cluster exacerbates the pathogenesis of acute pancreatitis in mice due to alterations in fibroinflammatory pathways. The cover image shows the pancreas of a miR-29a/b1–KO mouse with acute pancreatitis, with staining of infiltrating macrophages (brown).

Research Articles
Abstract

Myelodysplastic syndromes (MDS) are hematopoietic stem and progenitor cell (HSPC) malignancies characterized by ineffective hematopoiesis and an increased risk of leukemia transformation. Epigenetic regulators are recurrently mutated in MDS, directly implicating epigenetic dysregulation in MDS pathogenesis. Here, we identified a tumor suppressor role of the acetyltransferase p300 in clinically relevant MDS models driven by mutations in the epigenetic regulators TET2, ASXL1, and SRSF2. The loss of p300 enhanced the proliferation and self-renewal capacity of Tet2-deficient HSPCs, resulting in an increased HSPC pool and leukemogenicity in primary and transplantation mouse models. Mechanistically, the loss of p300 in Tet2-deficient HSPCs altered enhancer accessibility and the expression of genes associated with differentiation, proliferation, and leukemia development. Particularly, p300 loss led to an increased expression of Myb, and the depletion of Myb attenuated the proliferation of HSPCs and improved the survival of leukemia-bearing mice. Additionally, we show that chemical inhibition of p300 acetyltransferase activity phenocopied Ep300 deletion in Tet2-deficient HSPCs, whereas activation of p300 activity with a small molecule impaired the self-renewal and leukemogenicity of Tet2-deficient cells. This suggests a potential therapeutic application of p300 activators in the treatment of MDS with TET2 inactivating mutations.

Authors

Na Man, Gloria Mas, Daniel L. Karl, Jun Sun, Fan Liu, Qin Yang, Miguel Torres-Martin, Hidehiro Itonaga, Concepcion Martinez, Shi Chen, Ye Xu, Stephanie Duffort, Pierre-Jacques Hamard, Chuan Chen, Beth E. Zucconi, Luisa Cimmino, Feng-Chun Yang, Mingjiang Xu, Philip A. Cole, Maria E. Figueroa, Stephen D. Nimer

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Abstract

Vascular procedures, such as stenting, angioplasty, and bypass grafting, often fail due to intimal hyperplasia (IH), wherein contractile vascular smooth muscle cells (VSMCs) dedifferentiate to synthetic VSMCs, which are highly proliferative, migratory, and fibrotic. Previous studies suggest MAPK-activated protein kinase 2 (MK2) inhibition may limit VSMC proliferation and IH, although the molecular mechanism underlying the observation remains unclear. We demonstrated here that MK2 inhibition blocked the molecular program of contractile to synthetic dedifferentiation and mitigated IH development. Molecular markers of the VSMC contractile phenotype were sustained over time in culture in rat primary VSMCs treated with potent, long-lasting MK2 inhibitory peptide nanopolyplexes (MK2i-NPs), a result supported in human saphenous vein specimens cultured ex vivo. RNA-Seq of MK2i-NP–treated primary human VSMCs revealed programmatic switching toward a contractile VSMC gene expression profile, increasing expression of antiinflammatory and contractile-associated genes while lowering expression of proinflammatory, promigratory, and synthetic phenotype–associated genes. Finally, these results were confirmed using an in vivo rabbit vein graft model where brief, intraoperative treatment with MK2i-NPs decreased IH and synthetic phenotype markers while preserving contractile proteins. These results support further development of MK2i-NPs as a therapy for blocking VSMC phenotype switch and IH associated with cardiovascular procedures.

Authors

J. William Tierney, Brian C. Evans, Joyce Cheung-Flynn, Bo Wang, Juan M. Colazo, Monica E. Polcz, Rebecca S. Cook, Colleen M. Brophy, Craig L. Duvall

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Abstract

Autoimmune disease has presented an insurmountable barrier to restoration of durable immune tolerance. Previous studies indicate that chronic therapy with metabolic inhibitors can reduce autoimmune inflammation, but it remains unknown whether acute metabolic modulation enables permanent immune tolerance to be established. In an animal model of lupus, we determined that targeting glucose metabolism with 2-deoxyglucose (2DG) and mitochondrial metabolism with metformin enables endogenous immune tolerance mechanisms to respond to tolerance induction. A 2-week course of 2DG and metformin, when combined with tolerance-inducing therapy anti-CD45RB, prevented renal deposition of autoantibodies for 6 months after initial treatment and restored tolerance induction to allografts in lupus-prone mice. The restoration of durable immune tolerance was linked to changes in T cell surface glycosylation patterns, illustrating a role for glycoregulation in immune tolerance. These findings indicate that metabolic therapy may be applied as a powerful preconditioning to reinvigorate tolerance mechanisms in autoimmune and transplant settings that resist current immune therapies.

Authors

Christopher S. Wilson, Blair T. Stocks, Emilee M. Hoopes, Jillian P. Rhoads, Kelsey L. McNew, Amy S. Major, Daniel J. Moore

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Abstract

BACKGROUND Catheterization facilitates continuous bacteriuria, for which the clinical significance remains unclear. This study aimed to determine the clinical presentation, epidemiology, and dynamics of bacteriuria in a cohort of long-term catheterized nursing home residents.METHODS Prospective urine culture, urinalysis, chart review, and assessment of signs and symptoms of infection were performed weekly for 19 study participants over 7 months. All bacteria ≥ 1 × 103 cfu/mL were cultured, isolated, identified, and tested for susceptibility to select antimicrobials.RESULTS In total, 226 of the 234 urine samples were polymicrobial (97%), with an average of 4.7 isolates per weekly specimen. A total of 228 urine samples (97%) exhibited ≥ 1 × 106 CFU/mL, 220 (94%) exhibited abnormal urinalysis, 126 (54%) were associated with at least 1 possible sign or symptom of infection, and 82 (35%) would potentially meet a standardized definition of catheter-associated urinary tract infection (CAUTI), but only 3 had a caregiver diagnosis of CAUTI. Bacterial isolates (286; 30%) were resistant to a tested antimicrobial agent, and bacteriuria composition was remarkably stable despite a combined total of 54 catheter changes and 23 weeks of antimicrobial use.CONCLUSION Bacteriuria composition was largely polymicrobial, including persistent colonization by organisms previously considered to be urine culture contaminants. Neither antimicrobial use nor catheter changes sterilized the urine, at most resulting in transient reductions in bacterial burden followed by new acquisition of resistant isolates. Thus, this patient population exhibits a high prevalence of bacteriuria coupled with potential indicators of infection, necessitating further exploration to identify sensitive markers of true infection.FUNDING This work was supported by the NIH (R00 DK105205, R01 DK123158, UL1 TR001412).

Authors

Chelsie E. Armbruster, Aimee L. Brauer, Monica S. Humby, Jiahui Shao, Saptarshi Chakraborty

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Abstract

Common variable immunodeficiency (CVID) is characterized by profound primary antibody defects and frequent infections, yet autoimmune/inflammatory complications of unclear origin occur in 50% of individuals and lead to increased mortality. Here, we show that circulating bacterial 16S rDNA belonging to gut commensals was significantly increased in CVID serum (P < 0.0001), especially in patients with inflammatory manifestations (P = 0.0007). Levels of serum bacterial DNA were associated with parameters of systemic immune activation, increased serum IFN-γ, and the lowest numbers of isotype-switched memory B cells. Bacterial DNA was bioactive in vitro and induced robust host IFN-γ responses, especially among patients with CVID with inflammatory manifestations. Patients with X-linked agammaglobulinemia (Bruton tyrosine kinase [BTK] deficiency) also had increased circulating bacterial 16S rDNA but did not exhibit prominent immune activation, suggesting that BTK may be a host modifier, dampening immune responses to microbial translocation. These data reveal a mechanism for chronic immune activation in CVID and potential therapeutic strategies to modify the clinical outcomes of this disease.

Authors

Hsi-en Ho, Lin Radigan, Gerold Bongers, Ahmed El-Shamy, Charlotte Cunningham-Rundles

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Abstract

Proline rich 11 (PRR11), a novel tumor-related gene, has been identified in different tumors. However, the relevant biological functions of PRR11 in human clear cell renal cell carcinoma (ccRCC) have not been studied. In this study, we first identified PRR11 as a biomarker of ccRCC and predictor of poor prognosis by bioinformatics. Then, we showed that PRR11 silencing substantially reduced ccRCC cell proliferation and migration in vitro and in vivo. Importantly, we found that PRR11 induced the degradation of the E2F1 protein through its interaction with E2F1, and PRR11 reduced the stability of the E2F1 protein in ccRCC cells, thereby affecting cell cycle progression. Further results indicated that the downregulation of E2F1 expression partially reversed the changes in ccRCC cell biology caused by PRR11 deletion. In addition, we showed that PRR11 was a target gene of c-Myc. The transcription factor c-Myc may have promoted the expression of PRR11 in ccRCC cells by binding to the PRR11 promoter region, thereby accelerating the progression of ccRCC. In summary, we found that PRR11 served as an oncogene in ccRCC, and PRR11 reduced the protein stability of E2F1 and could be activated by c-Myc.

Authors

Siming Chen, Zhiwen He, Tianchen Peng, Fenfang Zhou, Gang Wang, Kaiyu Qian, Lingao Ju, Yu Xiao, Xinghuan Wang

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Abstract

Mucopolysaccharidosis type II (MPS II) is a lysosomal storage disorder caused by deficiency of the iduronate-2-sulfatase (IDS) enzyme, resulting in cellular accumulation of glycosaminoglycans (GAGs) throughout the body. Treatment of MPS II remains a considerable challenge as current enzyme replacement therapies do not adequately control many aspects of the disease, including skeletal and neurological manifestations. We developed an IDS transport vehicle (ETV:IDS) that is engineered to bind to the transferrin receptor; this design facilitates receptor-mediated transcytosis of IDS across the blood-brain barrier and improves its distribution into the brain while maintaining distribution to peripheral tissues. Here we show that chronic systemic administration of ETV:IDS in a mouse model of MPS II reduced levels of peripheral and central nervous system GAGs, microgliosis, and neurofilament light chain, a biomarker of neuronal injury. Additionally, ETV:IDS rescued auricular and skeletal abnormalities when introduced in adult MPS II mice. These effects were accompanied by improvements in several neurobehavioral domains, including motor skills, sensorimotor gating, and learning and memory. Together, these results highlight the therapeutic potential of ETV:IDS for treating peripheral and central abnormalities in MPS II. DNL310, an investigational ETV:IDS molecule, is currently in clinical trials as a potential treatment for patients with MPS II.

Authors

Annie Arguello, René Meisner, Elliot R. Thomsen, Hoang N. Nguyen, Ritesh Ravi, Jeffrey Simms, Iris Lo, Jessica Speckart, Julia Holtzman, Thomas M. Gill, Darren Chan, Yuhsiang Cheng, Chi-Lu Chiu, Jason C. Dugas, Meng Fang, Isabel A. Lopez, Hilda Solanoy, Buyankhishig Tsogtbaatar, Yuda Zhu, Akhil Bhalla, Kirk R. Henne, Anastasia G. Henry, Anthony Delucchi, Simona Costanzo, Jeffrey M. Harris, Dolores Diaz, Kimberly Scearce-Levie, Pascal E. Sanchez

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Abstract

Macrophages are commonly thought to contribute to the pathophysiology of preterm labor by amplifying inflammation — but a protective role has not previously been considered to our knowledge. We hypothesized that given their antiinflammatory capability in early pregnancy, macrophages exert essential roles in maintenance of late gestation and that insufficient macrophages may predispose individuals to spontaneous preterm labor and adverse neonatal outcomes. Here, we showed that women with spontaneous preterm birth had reduced CD209+CD206+ expression in alternatively activated CD45+CD14+ICAM3– macrophages and increased TNF expression in proinflammatory CD45+CD14+CD80+HLA-DR+ macrophages in the uterine decidua at the materno-fetal interface. In Cd11bDTR/DTR mice, depletion of maternal CD11b+ myeloid cells caused preterm birth, neonatal death, and postnatal growth impairment, accompanied by uterine cytokine and leukocyte changes indicative of a proinflammatory response, while adoptive transfer of WT macrophages prevented preterm birth and partially rescued neonatal loss. In a model of intra-amniotic inflammation–induced preterm birth, macrophages polarized in vitro to an M2 phenotype showed superior capacity over nonpolarized macrophages to reduce uterine and fetal inflammation, prevent preterm birth, and improve neonatal survival. We conclude that macrophages exert a critical homeostatic regulatory role in late gestation and are implicated as a determinant of susceptibility to spontaneous preterm birth and fetal inflammatory injury.

Authors

Nardhy Gomez-Lopez, Valeria Garcia-Flores, Peck Yin Chin, Holly M. Groome, Melanie T. Bijland, Kerrilyn R. Diener, Roberto Romero, Sarah A. Robertson

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Abstract

A population genetic study identified that the asialoglycoprotein receptor 1 (ASGR1) mutation carriers had substantially lower non–HDL-cholesterol (non–HDL-c) levels and reduced risks of cardiovascular diseases. However, the mechanism behind this phenomenon remained unclear. Here, we established Asgr1-knockout mice that represented a plasma lipid profile with significantly lower non–HDL-c and triglyceride (TG) caused by decreased secretion and increased uptake of VLDL/LDL. These 2 phenotypes were linked with the decreased expression of microsomal triglyceride transfer protein and proprotein convertase subtilisin/kexin type 9, 2 key targeted genes of sterol regulatory element–binding proteins (SREBPs). Furthermore, there were fewer nuclear SREBPs (nSREBPs) on account of more SREBPs being trapped in endoplasmic reticulum, which was caused by an increased expression of insulin-induced gene 1 (INSIG1), an anchor of SREBPs. Overexpression and gene knockdown interventions, in different models, were conducted to rescue the ASGR1-deficient phenotypes, and we found that INSIG1 knockdown independently reversed the ASGR1-mutated phenotypes with increased serum total cholesterol, LDL-c, TG, and liver cholesterol content accompanied by restored SREBP signaling. ASGR1 rescue experiments reduced INSIG1 and restored the SREBP network defect as manifested by improved apolipoprotein B secretion and reduced LDL uptake. Our observation demonstrated that increased INSIG1 is a critical factor responsible for ASGR1 deficiency–associated lipid profile changes and nSREBP suppression. This finding of an ASGR1/INSIG1/SREBP axis regulating lipid hemostasis may provide multiple potential targets for lipid-lowering drug development.

Authors

Yingying Xu, Jiawang Tao, Xiaorui Yu, Yuhang Wu, Yan Chen, Kai You, Jiaye Zhang, Anteneh Getachew, Tingcai Pan, Yuanqi Zhuang, Fang Yuan, Fan Yang, Xianhua Lin, Yin-xiong Li

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Abstract

Myosin binding protein-C slow (sMyBP-C) comprises a subfamily of cytoskeletal proteins encoded by MYBPC1 that is expressed in skeletal muscles where it contributes to myosin thick filament stabilization and actomyosin cross-bridge regulation. Recently, our group described the causal association of dominant missense pathogenic variants in MYBPC1 with an early-onset myopathy characterized by generalized muscle weakness, hypotonia, dysmorphia, skeletal deformities, and myogenic tremor, occurring in the absence of neuropathy. To mechanistically interrogate the etiologies of this MYBPC1-associated myopathy in vivo, we generated a knock-in mouse model carrying the E248K pathogenic variant. Using a battery of phenotypic, behavioral, and physiological measurements spanning neonatal to young adult life, we found that heterozygous E248K mice faithfully recapitulated the onset and progression of generalized myopathy, tremor occurrence, and skeletal deformities seen in human carriers. Moreover, using a combination of biochemical, ultrastructural, and contractile assessments at the level of the tissue, cell, and myofilaments, we show that the loss-of-function phenotype observed in mutant muscles is primarily driven by disordered and misaligned sarcomeres containing fragmented and out-of-register internal membranes that result in reduced force production and tremor initiation. Collectively, our findings provide mechanistic insights underscoring the E248K-disease pathogenesis and offer a relevant preclinical model for therapeutic discovery.

Authors

Janelle Geist Hauserman, Janis Stavusis, Humberto C. Joca, Joel C. Robinett, Laurin Hanft, Jack Vandermeulen, Runchen Zhao, Joseph P. Stains, Konstantinos Konstantopoulos, Kerry S. McDonald, Christopher Ward, Aikaterini Kontrogianni-Konstantopoulos

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Abstract

The role and mechanisms for upregulating complement factor B (CFB) expression in podocyte dysfunction in diabetic kidney disease (DKD) are not fully understood. Here, analyzing Gene Expression Omnibus GSE30528 data, we identified genes enriched in mTORC1 signaling, CFB, and complement alternative pathways in podocytes from patients with DKD. In mouse models, podocyte mTOR complex 1 (mTORC1) signaling activation was induced, while blockade of mTORC1 signaling reduced CFB upregulation, alternative complement pathway activation, and podocyte injury in the glomeruli. Knocking down CFB remarkably alleviated alternative complement pathway activation and DKD in diabetic mice. In cultured podocytes, high glucose treatment activated mTORC1 signaling, stimulated STAT1 phosphorylation, and upregulated CFB expression, while blockade of mTORC1 or STAT1 signaling abolished high glucose–upregulated CFB expression. Additionally, high glucose levels downregulated protein phosphatase 2Acα (PP2Acα) expression, while PP2Acα deficiency enhanced high glucose–induced mTORC1/STAT1 activation, CFB induction, and podocyte injury. Taken together, these findings uncover a mechanism by which CFB mediates podocyte injury in DKD.

Authors

Qingmiao Lu, Qing Hou, Kai Cao, Xiaoli Sun, Yan Liang, Mengru Gu, Xian Xue, Allan Zijian Zhao, Chunsun Dai

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Abstract

Dilated cardiomyopathy (DCM) is the most common form of cardiomyopathy and main indication for heart transplantation in children. Therapies specific to pediatric DCM remain limited due to lack of a disease model. Our previous study showed that treatment of neonatal rat ventricular myocytes (NRVMs) with serum from nonfailing or DCM pediatric patients activates the fetal gene program (FGP). Here we show that serum treatment with proteinase K prevents activation of the FGP, whereas RNase treatment exacerbates it, suggesting that circulating proteins, but not circulating miRNAs, promote these pathological changes. Evaluation of the protein secretome showed that midkine (MDK) is upregulated in DCM serum, and NRVM treatment with MDK activates the FGP. Changes in gene expression in serum-treated NRVMs, evaluated by next-generation RNA-Seq, indicated extracellular matrix remodeling and focal adhesion pathways were upregulated in pediatric DCM serum and in DCM serum–treated NRVMs, suggesting alterations in cellular stiffness. Cellular stiffness was evaluated by Atomic Force Microscopy, which showed an increase in stiffness in DCM serum–treated NRVMs. Of the proteins increased in DCM sera, secreted frizzled-related protein 1 (sFRP1) was a potential candidate for the increase in cellular stiffness, and sFRP1 treatment of NRVMs recapitulated the increase in cellular stiffness observed in response to DCM serum treatment. Our results show that serum circulating proteins promoted pathological changes in gene expression and cellular stiffness, and circulating miRNAs were protective against pathological changes.

Authors

Danielle A. Jeffrey, Julie Pires Da Silva, Anastacia M. Garcia, Xuan Jiang, Anis Karimpour-Fard, Lee S. Toni, Thomas Lanzicher, Brisa Peña, Carissa A. Miyano, Karin Nunley, Armin Korst, Orfeo Sbaizero, Matthew R.G. Taylor, Shelley D. Miyamoto, Brian L. Stauffer, Carmen C. Sucharov

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Abstract

Current treatments for Parkinson’s disease (PD) provide only symptomatic relief, with no disease-modifying therapies identified to date. Repurposing FDA-approved drugs to treat PD could significantly shorten the time needed for and reduce the costs of drug development compared with conventional approaches. We developed an efficient strategy to screen for modulators of β-glucocerebrosidase (GCase), a lysosomal enzyme that exhibits decreased activity in patients with PD, leading to accumulation of the substrate glucosylceramide and oxidized dopamine and α-synuclein, which contribute to PD pathogenesis. Using a GCase fluorescent probe and affinity-based fluorescence polarization assay, we screened 1280 structurally diverse, bioactive, and cell-permeable FDA-approved drugs and found that the antipsychotic quetiapine bound GCase with high affinity. Moreover, quetiapine treatment of induced pluripotent stem cell–derived (iPSC-derived) dopaminergic neurons from patients carrying mutations in GBA1 or LRRK2 led to increased wild-type GCase protein levels and activity and partially lowered accumulation of oxidized dopamine, glucosylceramide, and α-synuclein. Similarly, quetiapine led to activation of wild-type GCase and reduction of α-synuclein in a GBA mutant mouse model (Gba1D409V/+ mice). Together, these results suggest that repurposing quetiapine as a modulator of GCase may be beneficial for patients with PD exhibiting decreased GCase activity.

Authors

Lena F. Burbulla, Jianbin Zheng, Pingping Song, Weilan Jiang, Michaela E. Johnson, Patrik Brundin, Dimitri Krainc

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Abstract

Hypomorphic RAG1 or RAG2 mutations cause primary immunodeficiencies and can lead to autoimmunity, but the underlying mechanisms are elusive. We report here a patient carrying a c.116+2T>G homozygous splice site mutation in the first intron of RAG1, which led to aberrant splicing and greatly reduced RAG1 protein expression. B cell development was blocked at both the pro-B to pre-B transition and the pre-B to immature B cell differentiation step. The patient B cells had reduced B cell receptor repertoire diversity and decreased complementarity determining region 3 lengths. Despite B cell lymphopenia, the patient had abundant plasma cells in the BM and produced large quantities of IgM and IgG Abs, including autoantibodies. The proportion of naive B cells was reduced while the frequency of IgD–CD27– double-negative (DN) B cells, which quickly differentiated into Ab-secreting plasma cells upon stimulation, was greatly increased. Immune phenotype analysis of 52 patients with primary immunodeficiency revealed a strong association of the increased proportion of DN B and memory B cells with decreased number and proportion of naive B cells. These results suggest that the lymphopenic environment triggered naive B cell differentiation into DN B and memory B cells, leading to increased Ab production.

Authors

Qing Min, Xin Meng, Qinhua Zhou, Ying Wang, Yaxuan Li, Nannan Lai, Ermeng Xiong, Wenjie Wang, Shoya Yasuda, Meiping Yu, Hai Zhang, Jinqiao Sun, Xiaochuan Wang, Ji-Yang Wang

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Abstract

The signaling mechanisms by which dietary fat and cholesterol signals regulate central pathways of glucose homeostasis are not completely understood. By using a hepatocyte-specific PKCβ-deficient (PKCβHep–/–) mouse model, we demonstrated the role of hepatic PKCβ in slowing disposal of glucose overload by suppressing glycogenesis and increasing hepatic glucose output. PKCβHep–/– mice exhibited lower plasma glucose under the fed condition, modestly improved systemic glucose tolerance and mildly suppressed gluconeogenesis, increased hepatic glycogen accumulation and synthesis due to elevated glucokinase expression and activated glycogen synthase (GS), and suppressed glucose-6-phosphatase expression compared with controls. These events were independent of hepatic AKT/GSK-3α/β signaling and were accompanied by increased HNF-4α transactivation, reduced FoxO1 protein abundance, and elevated expression of GS targeting protein phosphatase 1 regulatory subunit 3C in the PKCβHep–/– liver compared with controls. The above data strongly imply that hepatic PKCβ deficiency causes hypoglycemia postprandially by promoting glucose phosphorylation via upregulating glucokinase and subsequently redirecting more glucose-6-phosphate to glycogen via activating GS. In summary, hepatic PKCβ has a unique and essential ability to induce a coordinated response that negatively affects glycogenesis at multiple levels under physiological postprandial conditions, thereby integrating nutritional fat intake with dysregulation of glucose homeostasis.

Authors

Yaoling Shu, Faizule Hassan, Michael C. Ostrowski, Kamal D. Mehta

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Abstract

Repetitive mild traumatic brain injuries (mTBI) disrupt CNS barriers, the erosion of which has been linked to long-term neurodegenerative and psychiatric conditions. Although much attention has been devoted to CNS vasculature following mTBI, little is known about the glia limitans superficialis — a barrier of surface-associated astrocytes that helps protect the CNS parenchyma and maintain homeostasis. Here, we identify the glia limitans superficialis as a crucial barrier surface whose breakdown after acute repeat mTBI facilitates increased cell death and recruitment of peripheral myelomonocytic cells. Using intravital microscopy, we show that brain-resident microglia fortify this structure after a single mTBI, yet they fail to do so following secondary injury, which triggers massive recruitment of myelomonocytic cells from the periphery that contribute to further destruction of the glia limitans superficialis but not cortical cell death. We demonstrate, instead, that reactive oxygen species (ROS) generated in response to repetitive head injury are largely responsible for enhanced cortical cell death, and therapeutic administration of the antioxidant glutathione markedly reduces this cell death, preserves the glia limitans, and prevents myelomonocytic cells from entering the brain parenchyma. Collectively, our findings underscore the importance of preserving the glia limitans superficialis after brain injury and offer a therapeutic means to protect this structure and the underlying cortex.

Authors

Hannah D. Mason, Alexis M. Johnson, Nicole A. Mihelson, Panagiotis Mastorakos, Dorian B. McGavern

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Abstract

MicroRNA-29 (miR-29) is a critical regulator of fibroinflammatory processes in human diseases. In this study, we found a decrease in miR-29a in experimental and human chronic pancreatitis, leading us to investigate the regulatory role of the miR-29a/b1 cluster in acute pancreatitis (AP) utilizing a conditional miR-29a/b1–KO mouse model. miR-29a/b1-sufficient (WT) and -deficient (KO) mice were administered supramaximal caerulein to induce AP and characterized at different time points, utilizing an array of IHC and biochemical analyses for AP parameters. In caerulein-induced WT mice, miR-29a remained dramatically downregulated at injury. Despite high-inflammatory milieu, fibrosis, and parenchymal disarray in the WT mice during early AP, the pancreata fully restored during recovery. miR-29a/b1–KO mice showed significantly greater inflammation, lymphocyte infiltration, macrophage polarization, and ECM deposition, continuing until late recovery with persistent parenchymal disorganization. The increased pancreatic fibrosis was accompanied by enhanced TGFβ1 coupled with persistent αSMA+ PSC activation. Additionally, these mice exhibited higher circulating IL-6 and inflammation in lung parenchyma. Together, this collection of studies indicates that depletion of miR-29a/b1 cluster impacts the fibroinflammatory mechanisms of AP, resulting in (a) aggravated pathogenesis and (b) delayed recovery from the disease, suggesting a protective role of the molecule against AP.

Authors

Shatovisha Dey, Lata M. Udari, Primavera RiveraHernandez, Jason J. Kwon, Brandon Willis, Jeffrey J. Easler, Evan L. Fogel, Stephen Pandol, Janaiah Kota

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Abstract

Obesity, a major health care issue, is characterized by metabolic abnormalities in multiple tissues, including the skeletal muscle. Although dysregulation of skeletal muscle metabolism can strongly influence the homeostasis of systemic energy, the underlying mechanism remains unclear. We found promoter hypermethylation and decreased gene expression of fibroblast growth factor 6 (FGF6) in the skeletal muscle of individuals with obesity using high-throughput sequencing. Reduced binding of the cyclic AMP responsive element binding protein-1 (CREB1) to the hypermethylated cyclic AMP response element, which is a regulatory element upstream of the transcription initiation site, partially contributed to the downregulation of FGF6 in patients with obesity. Overexpression of Fgf6 in mouse skeletal muscle stimulated protein synthesis, activating the mammalian target of rapamycin pathway, and prevented the increase in weight and the development of insulin resistance in high-fat diet–fed mice. Thus, our findings highlight the role played by Fgf6 in regulating skeletal muscle hypertrophy and whole-body metabolism, indicating its potential in strategies aimed at preventing and treating metabolic diseases.

Authors

Bo Xu, Caizhi Liu, Hong Zhang, Rong Zhang, Mengyang Tang, Yan Huang, Li Jin, Lingyan Xu, Cheng Hu, Weiping Jia

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Abstract

Herein, we report that Shroom3 knockdown, via Fyn inhibition, induced albuminuria with foot process effacement (FPE) without focal segmental glomerulosclerosis (FSGS) or podocytopenia. Interestingly, knockdown mice had reduced podocyte volumes. Human minimal change disease (MCD), where podocyte Fyn inactivation was reported, also showed lower glomerular volumes than FSGS. We hypothesized that lower glomerular volume prevented the progression to podocytopenia. To test this hypothesis, we utilized unilateral and 5/6th nephrectomy models in Shroom3-KD mice. Knockdown mice exhibited less glomerular and podocyte hypertrophy after nephrectomy. FYN-knockdown podocytes had similar reductions in podocyte volume, implying that Fyn was downstream of Shroom3. Using SHROOM3 or FYN knockdown, we confirmed reduced podocyte protein content, along with significantly increased phosphorylated AMPK, a negative regulator of anabolism. AMPK activation resulted from increased cytoplasmic redistribution of LKB1 in podocytes. Inhibition of AMPK abolished the reduction in glomerular volume and induced podocytopenia in mice with FPE, suggesting a protective role for AMPK activation. In agreement with this, treatment of glomerular injury models with AMPK activators restricted glomerular volume, podocytopenia, and progression to FSGS. Glomerular transcriptomes from MCD biopsies also showed significant enrichment of Fyn inactivation and Ampk activation versus FSGS glomeruli. In summary, we demonstrated the important role of AMPK in glomerular volume regulation and podocyte survival. Our data suggest that AMPK activation adaptively regulates glomerular volume to prevent podocytopenia in the context of podocyte injury.

Authors

Khadija Banu, Qisheng Lin, John M. Basgen, Marina Planoutene, Chengguo Wei, Anand C. Reghuvaran, Xuefei Tian, Hongmei Shi, Felipe Garzon, Aitor Garzia, Nicholas Chun, Arun Cumpelik, Andrew D. Santeusanio, Weijia Zhang, Bhaskar Das, Fadi Salem, Li Li, Shuta Ishibe, Lloyd G. Cantley, Lewis Kaufman, Kevin V. Lemley, Zhaohui Ni, John Cijiang He, Barbara Murphy, Madhav C. Menon

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Abstract

A role for hereditary influences in the susceptibility for chronic obstructive pulmonary disease (COPD) is widely recognized. Cytotoxic lymphocytes are implicated in COPD pathogenesis, and functions of these leukocytes are modulated by interactions between their killer cell Ig-like receptors (KIR) and human leukocyte antigen–Class I (HLA–Class I) molecules on target cells. We hypothesized HLA–Class I and KIR inheritance affect risks for COPD. HLA–Class I alleles and KIR genotypes were defined by candidate gene analyses in multiple cohorts of patients with COPD (total n = 392) and control smokers with normal spirometry (total n = 342). Compared with controls, patients with COPD had overrepresentations of HLA-C*07 and activating KIR2DS1, with underrepresentations of HLA-C*12. Particular HLA-KIR permutations were synergistic; e.g., the presence of HLA-C*07 + KIR2DS1 + HLA-C12null versus HLAC*07null + KIR2DS1null + HLA-C12 was associated with COPD, especially among HLA-C1 allotype homozygotes. Cytotoxicity of COPD lymphocytes was more enhanced by KIR stimulation than those of controls and was correlated with lung function. These data show HLA-C and KIR polymorphisms strongly influence COPD susceptibility and highlight the importance of lymphocyte-mediated cytotoxicity in COPD pathogenesis. Findings here also indicate that HLA-KIR typing could stratify at-risk patients and raise possibilities that HLA-KIR axis modulation may have therapeutic potential.

Authors

Takudzwa Mkorombindo, Thi K. Tran-Nguyen, Kaiyu Yuan, Yingze Zhang, Jianmin Xue, Gerard J. Criner, Young-il Kim, Joseph M. Pilewski, Amit Gaggar, Michael H. Cho, Frank C. Sciurba, Steven R. Duncan

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Abstract

IgA nephropathy is caused by deposition of circulatory IgA1 in the kidney. Hypogalactosylated IgA1 has the propensity to form poly-IgA aggregates that are prone to deposition. Herein, we purified poly-IgA from the plasma of patients with IgA nephropathy and showed that the complex is susceptible to reducing conditions, suggesting intermolecular disulfide connections between IgA units. We sought to find the cysteine residue(s) that form intermolecular disulfide. Naturally assembled dimeric IgA, also known as secretory IgA, involves a J chain subunit connected with 2 IgA1 molecules via their penultimate cysteine-471 residue on a “tailpiece” segment of IgA heavy chain. It is plausible that, with the absence of J chain, the cysteine residue of mono-IgA1 might aberrantly form a disulfide bond in poly-IgA formation. Mutagenesis confirmed that cysteine-471 is capable of promoting IgA aggregation. These discoveries prompted us to test thiol-based drugs for stabilizing cysteine. Specifically, the cystine-reducing drug cysteamine used for treatment of cystinosis showed a remarkable potency in preventing self-aggregation of IgA. When administrated to rat and mouse models of IgA nephropathy, cysteamine significantly reduced glomerular IgA deposition. Collectively, our results reveal a potentially novel molecular mechanism for aberrant formation of IgA aggregates, to which the repurposed cystinosis drug cysteamine was efficacious in preventing renal IgA deposition.

Authors

Xinfang Xie, Li Gao, Pan Liu, Jicheng Lv, Wan-Hong Lu, Hong Zhang, Jing Jin

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Abstract

BACKGROUND SARS-CoV-2 infection induces mucin overexpression, further promoting disease. Given that mucins are critical components of innate immunity, unraveling their expression profiles that dictate the course of disease could greatly enhance our understanding and management of COVID-19.METHODS Using validated RT-PCR assays, we assessed mucin mRNA expression in the blood of patients with symptomatic COVID-19 compared with symptomatic patients without COVID-19 and healthy controls and correlated the data with clinical outcome parameters. Additionally, we analyzed mucin expression in mucus and lung tissue from patients with COVID-19 and investigated the effect of drugs for COVID-19 treatment on SARS-CoV-2–induced mucin expression in pulmonary epithelial cells.RESULTS We identified a dynamic blood mucin mRNA signature that clearly distinguished patients with symptomatic COVID-19 from patients without COVID-19 based on expression of MUC1, MUC2, MUC4, MUC6, MUC13, MUC16, and MUC20 (AUCROC of 91.8%; sensitivity and specificity of 90.6% and 93.3%, respectively) and that discriminated between mild and critical COVID-19 based on the expression of MUC16, MUC20, and MUC21 (AUCROC of 89.1%; sensitivity and specificity of 90.0% and 85.7%, respectively). Differences in the transcriptional landscape of mucins in critical cases compared with mild cases identified associations with COVID-19 symptoms, respiratory support, organ failure, secondary infections, and mortality. Furthermore, we identified different mucins in the mucus and lung tissue of critically ill COVID-19 patients and showed the ability of baricitinib, tocilizumab, favipiravir, and remdesivir to suppress expression of SARS-CoV-2–induced mucins.CONCLUSION This multifaceted blood mucin mRNA signature showed the potential role of mucin profiling in diagnosing, estimating severity, and guiding treatment options in patients with COVID-19.FUNDING The Antwerp University Research and the Research Foundation Flanders COVID-19 funds.

Authors

Annemieke Smet, Tom Breugelmans, Johan Michiels, Kevin Lamote, Wout Arras, Joris G. De Man, Leo Heyndrickx, Anne Hauner, Manon Huizing, Surbhi Malhotra-Kumar, Martin Lammens, An Hotterbeekx, Samir Kumar-Singh, Aline Verstraeten, Bart Loeys, Veronique Verhoeven, Rita Jacobs, Karolien Dams, Samuel Coenen, Kevin K. Ariën, Philippe G. Jorens, Benedicte Y. De Winter

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Abstract

BACKGROUND This study systematically investigated circulating and retinal tissue lipid determinants of human diabetic retinopathy (DR) to identify underlying lipid alterations associated with severity of DR.METHODS Retinal tissues were retrieved from postmortem human eyes, including 19 individuals without diabetes, 20 with diabetes but without DR, and 20 with diabetes and DR, for lipidomic study. In a parallel study, serum samples from 28 American Indians with type 2 diabetes from the Gila River Indian Community, including 12 without DR, 7 with mild nonproliferative DR (NPDR), and 9 with moderate NPDR, were selected. A mass-spectrometry–based lipidomic platform was used to measure serum and tissue lipids.RESULTS In the postmortem retinas, we found a graded decrease of long-chain acylcarnitines and longer-chain fatty acid ester of hydroxyl fatty acids, diacylglycerols, triacylglycerols, phosphatidylcholines, and ceramide(NS) in central retina from individuals with no diabetes to those with diabetes with DR. The American Indians’ sera also exhibited a graded decrease in circulating long-chain acylcarnitines and a graded increase in the intermediate-length saturated and monounsaturated triacylglycerols from no DR to moderate NPDR.CONCLUSION These findings suggest diminished synthesis of complex lipids and impaired mitochondrial β-oxidation of fatty acids in retinal DR, with parallel changes in circulating lipids.TRIAL REGISTRATION ClinicalTrials.gov NCT00340678.FUNDING This work was supported by NIH grants R24 DK082841, K08DK106523, R03DK121941, P30DK089503, P30DK081943, P30DK020572, P30 EY007003; The Thomas Beatson Foundation; and JDRF Center for Excellence (5-COE-2019-861-S-B).

Authors

Patrice E. Fort, Thekkelnaycke M. Rajendiran, Tanu Soni, Jaeman Byun, Yang Shan, Helen C. Looker, Robert G. Nelson, Matthias Kretzler, George Michailidis, Jerome E. Roger, Thomas W. Gardner, Steven F. Abcouwer, Subramaniam Pennathur, Farsad Afshinnia

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Abstract

Avian H7N9 influenza viruses cause sporadic outbreaks of human infections and threaten to cause a major pandemic. The breadth of B cell responses to natural infection and the dominant antigenic sites recognized during first exposure to H7 HA following infection are incompletely understood. Here, we studied the B cell response to H7 HA of 2 individuals who had recovered from natural H7N9 virus infection. We used competition binding, hydrogen-deuterium mass spectrometry, and single-particle negative stain electron microscopy to identify the patterns of molecular recognition of the antibody responses to H7 HA. We found that circulating H7-reactive B cells recognized a diverse antigenic landscape on the HA molecule, including HA head domain epitopes in antigenic sites A and B and in the trimer interface-II region and epitopes in the stem region. Most H7 antibodies exhibited little heterosubtypic breadth, but many recognized a wide diversity of unrelated H7 strains. We tested the antibodies for functional activity and identified clones with diverse patterns of inhibition, including neutralizing, hemagglutination- or egress-inhibiting, or HA trimer–disrupting activities. Thus, the human B cell response to primary H7 natural infection is diverse, highly functional, and broad for recognition of diverse H7 strains.

Authors

Iuliia M. Gilchuk, Sandhya Bangaru, Nurgun Kose, Robin G. Bombardi, Andrew Trivette, Sheng Li, Hannah L. Turner, Robert H. Carnahan, Andrew B. Ward, James E. Crowe Jr.

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Abstract

Tregs play vital roles in suppressing atherogenesis. Pathological conditions reshape Tregs and increase Treg-weakening plasticity. It remains unclear how Tregs preserve their function and how Tregs switch into alternative phenotypes in the environment of atherosclerosis. In this study, we observed a great induction of CD4+Foxp3+ Tregs in the spleen and aorta of ApoE–/– mice, accompanied by a significant increase of plasma IL-35 levels. To determine if IL-35 devotes its role in the rise of Tregs, we generated IL-35 subunit P35–deficient (IL-35P35–deficient) mice on an ApoE–/– background and found Treg reduction in the spleen and aorta compared with ApoE–/– controls. In addition, our RNA sequencing data show the elevation of a set of chemokine receptor transcripts in the ApoE–/– Tregs, and we have validated higher CCR5 expression in ApoE–/– Tregs in the presence of IL-35 than in the absence of IL-35. Furthermore, we observed that CCR5+ Tregs in ApoE–/– have lower Treg-weakening AKT-mTOR signaling, higher expression of inhibitory checkpoint receptors TIGIT and PD-1, and higher expression of IL-10 compared with WT CCR5+ Tregs. In conclusion, IL-35 counteracts hyperlipidemia in maintaining Treg-suppressive function by increasing 3 CCR5-amplified mechanisms, including Treg migration, inhibition of Treg weakening AKT-mTOR signaling, and promotion of TIGIT and PD-1 signaling.

Authors

Ying Shao, William Y. Yang, Fatma Saaoud, Charles Drummer IV, Yu Sun, Keman Xu, Yifan Lu, Huimin Shan, Ethan M. Shevach, Xiaohua Jiang, Hong Wang, Xiaofeng Yang

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Abstract

COVID-19, caused by SARS-CoV-2, has spread worldwide with dire consequences. To urgently investigate the pathogenicity of COVID-19 and develop vaccines and therapeutics, animal models that are highly susceptible to SARS-CoV-2 infection are needed. In the present study, we established an animal model highly susceptible to SARS-CoV-2 via the intratracheal tract infection in CAG promoter–driven human angiotensin-converting enzyme 2–transgenic (CAG-hACE2) mice. The CAG-hACE2 mice showed several severe symptoms of SARS-CoV-2 infection, with definitive weight loss and subsequent death. Acute lung injury with elevated cytokine and chemokine levels was observed at an early stage of infection in CAG-hACE2 mice infected with SARS-CoV-2. Analysis of the hACE2 gene in CAG-hACE2 mice revealed that more than 15 copies of hACE2 genes were integrated in tandem into the mouse genome, supporting the high susceptibility to SARS-CoV-2. In the developed model, immunization with viral antigen or injection of plasma from immunized mice prevented body weight loss and lethality due to infection with SARS-CoV-2. These results indicate that a highly susceptible model of SARS-CoV-2 infection in CAG-hACE2 mice via the intratracheal tract is suitable for evaluating vaccines and therapeutic medicines.

Authors

Masamitsu N. Asaka, Daichi Utsumi, Haruhiko Kamada, Satoshi Nagata, Yutaka Nakachi, Tomokazu Yamaguchi, Yoshihiro Kawaoka, Keiji Kuba, Yasuhiro Yasutomi

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Abstract

The mechanisms that link visceral mechanosensation to the perception of internal organ status (i.e., interoception) remain elusive. In response to bladder filling, the urothelium releases ATP, which is hypothesized to stimulate voiding function by communicating the degree of bladder fullness to subjacent tissues, including afferent nerve fibers. To determine if PIEZO channels function as mechanosensors in these events, we generated conditional urothelial Piezo1-, Piezo2-, and dual Piezo1/2-knockout (KO) mice. While functional PIEZO1 channels were expressed in all urothelial cell layers, Piezo1-KO mice had a limited phenotype. Piezo2 expression was limited to a small subset of superficial umbrella cells, yet male Piezo2-KO mice exhibited incontinence (i.e., leakage) when their voiding behavior was monitored during their active dark phase. Dual Piezo1/2-KO mice had the most affected phenotype, characterized by decreased urothelial responses to mechanical stimulation, diminished ATP release, bladder hypoactivity in anesthetized Piezo1/2-KO females but not males, and urinary incontinence in both male and female Piezo1/2-KO mice during their dark phase but not inactive light one. Our studies reveal that the urothelium functions in a sex- and circadian rhythm–dependent manner to link urothelial PIEZO1/2 channel–driven mechanotransduction to normal voiding function and behavior, and in the absence of these signals, bladder dysfunction ensues.

Authors

Marianela G. Dalghi, Wily G. Ruiz, Dennis R. Clayton, Nicolas Montalbetti, Stephanie L. Daugherty, Jonathan M. Beckel, Marcelo D. Carattino, Gerard Apodaca

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Abstract

Understanding the presence and durability of antibodies against SARS-CoV-2 in the airways is required to provide insights on the ability of individuals to neutralize the virus locally and prevent viral spread. Here, we longitudinally assessed both systemic and airway immune responses upon SARS-CoV-2 infection in a clinically well-characterized cohort of 147 infected individuals representing the full spectrum of COVID-19 severity; from asymptomatic infection to fatal disease. In addition, we evaluated how SARS-CoV-2 vaccination influenced the antibody responses in a subset of these individuals during convalescence as compared to naïve individuals. Not only systemic but also airway antibody responses correlated with the degree of COVID-19 disease severity. However, while systemic IgG levels were durable for up to 8 months, airway IgG and IgA had declined significantly within 3 months. After vaccination, there was an increase in both systemic and airway antibodies, in particular IgG, often exceeding the levels found during acute disease. In contrast, naïve individuals showed low airway antibodies after vaccination. In the former COVID-19 patients, airway antibody levels were significantly elevated after the boost vaccination, highlighting the importance of prime and boost vaccination also for previously infected individuals to obtain optimal mucosal protection.

Authors

Alberto Cagigi, Meng Yu, Björn Österberg, Julia Svensson, Sara Falck-Jones, Sindhu Vangeti, Eric Åhlberg, Lida Azizmohammadi, Anna Warnqvist, Ryan Falck-Jones, Pia C. Gubisch, Mert Ödemis, Farangies Ghafoor, Mona Eisele, Klara Lenart, Max Bell, Niclas Johansson, Jan Albert, Jörgen Sälde, Deleah D. Pettie, Michael P. Murphy, Lauren Carter, Neil P. King, Sebastian Ols, Johan Normark, Clas Ahlm, Mattias N. Forsell, Anna Färnert, Karin Loré, Anna Smed-Sörensen

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Abstract

BACKGROUND. Recessive dystrophic epidermolysis bullosa (RDEB) is a rare, devastating, and life-threatening inherited skin fragility disorder due to a lack of functional type VII collagen, for which no effective therapy exists. ABCB5-positive dermal mesenchymal stem cells (ABCB5+ MSCs) possess immunomodulatory capacities, a favorable skin homing potential and the ability to secrete type VII collagen. In a COL7A1–/– mouse model of RDEB, treatment with ABCB5+ MSCs markedly extended the animals’ lifespans. METHODS. In this international, multicentric, single-arm, phase I/IIa clinical trial, 16 patients (aged 4–36 years) enrolled into four age cohorts received three intravenous infusions of 2×106 ABCB5+ MSCs/kg on days 0, 17 and 35. Patients were followed up for 12 weeks regarding efficacy and 12 months regarding safety. RESULTS. At 12 weeks, statistically significant median (IQR) reductions in the Epidermolysis Bullosa Disease Activity and Scarring Index activity (EBDASI activity) score of 13.0% (2.9%-30%; P = 0.049) and the Instrument for Scoring Clinical Outcome of Research for Epidermolysis Bullosa clinician (iscorEB c) score of 18.2% (4.1%-41.7%; P = 0.037) were observed. Reductions in itch and pain numerical rating scale scores were greatest on day 35, amounting to 37.5% (0.0%-42.9%; P = 0.033) and 25.0% (-8.4%-46.4%; P = 0.168), respectively. Three adverse events were considered related to the cell product, one mild lymphadenopathy and two hypersensitivity reactions. The latter two were serious but resolved without sequelae shortly after withdrawal of treatment. CONCLUSION. This trial demonstrates good tolerability, manageable safety and potential efficacy of intravenous ABCB5+ MSCs as a readily available disease-modifying therapy for RDEB and provides a rationale for further clinical evaluation. TRIAL REGISTRATION. clinicaltrials.gov NCT03529877; EudraCT 2018-001009-98 FUNDING. The trial was sponsored by RHEACELL GmbH & Co. KG, Heidelberg, Germany. Contributions by NY Frank and MH Frank to this work were supported by the National Institutes of Health (NIH)/National Eye Institute (NEI) grants RO1EY025794 and R24EY028767.

Authors

Dimitra Kiritsi, Kathrin Dieter, Elke Niebergall-Roth, Silvia Fluhr, Cristina Daniele, Jasmina Esterlechner, Samar Sadeghi, Seda Ballikaya, Leoni Erdinger, Franziska Schauer, Stella Gewert, Martin Laimer, Johann W. Bauer, Alain Hovnanian, Giovanna Zambruno, May El Hachem, Emmanuelle Bourrat, Maria Papanikolaou, Gabriela Petrof, Sophie Kitzmüller, Christen L. Ebens, Markus H. Frank, Natasha Y. Frank, Christoph Ganss, Anna E. Martinez, John A. McGrath, Jakub Tolar, Mark A. Kluth

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Abstract

Lung allograft rejection results in the accumulation of low molecular weight hyaluronic acid (LMW-HA), which further propagates inflammation and tissue injury. We have previously shown that therapeutic lymphangiogenesis in a murine model of lung allograft rejection reduced tissue LMW-HA and was associated with improved transplant outcomes. Herein we investigated the use of 4-Methylumbelliferone, a known inhibitor of HA synthesis, to alleviate acute allograft rejection in a murine model of lung transplantation. We found that treating mice with 4MU from day 20-30 post-transplant was sufficient to significantly improve outcomes, characterized by a reduction in T-cell mediated lung inflammation, LMW-HA content, and improved pathology scores. In vitro, 4MU directly attenuated activation, proliferation, and differentiation of naïve CD4+ T-cells into Th1 cells. As 4MU has already been demonstrated to be safe for human use, we believe examining 4MU for the treatment of acute lung allograft rejection may be of clinical significance.

Authors

Jewel Imani, Kaifeng Liu, Ye Cui, Jean-Pierre Assaker, Junwen Han, Auyon J. Ghosh, Julie Ng, Shikshya Shrestha, Anthony M. Lamattina, Pierce H. Louis, Anne Hentschel, Anthony J. Esposito, Ivan O. Rosas, Xiaoli Liu, Mark A. Perrella, Jamil Azzi, Gary Visner, Souheil El-Chemaly

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Abstract

Patients with acute leukemia who are unable to achieve complete remission prior to allogeneic hematopoietic stem cell transplantation (SCT) have dismal outcomes with relapse rates well in excess of 60%. Haplo-identical SCT (haplo-SCT) may allow enhanced graft-versus-leukemia (GVL) effects by virtue of HLA class I/II donor-host disparities but typically requires intensive immune-suppression with post-transplant cyclophosphamide (PT-Cy) to prevent lethal graft-versus-host disease (GVHD). Here we demonstrate in preclinical models that glucocorticoid administration from day -1 to +5 inhibits alloantigen presentation by professional recipient antigen presenting cells in the gastrointestinal tract and prevents donor T-cell priming and subsequent expansion therein. In contrast, direct glucocorticoid signaling of donor T-cells promotes chemokine and integrin signatures permissive of preferential circulation and migration into the bone marrow, promoting donor T-cell residency. This results in significant reductions in GVHD whilst promoting potent GVL effects (relapse in recipients receiving glucocorticoids, vehicle or PT-Cy was 12%, 56% and 100% respectively). Intriguingly, patients with acute myeloid leukemia not in remission that received unmanipulated haplo-SCT and peri-transplant glucocorticoids also had an unexpectedly low relapse rate at 1 year (32%: 95% CI, 18%-47%) with high overall survival at 3 years (58%: 95% CI, 38-74%). These data highlight a potentially simple and effective approach to prevent relapse in patients with otherwise incurable leukemia that could be studied in prospective randomized trials.

Authors

Takayuki Inoue, Motoko Koyama, Katsuji Kaida, Kazuhiro Ikegame, Kathleen S. Ensbey, Luke Samson, Shuichiro Takahashi, Ping Zhang, Simone A. Minnie, Satoshi Maruyama, Shinichi Ishii, Takashi Daimon, Takahiro Fukuda, Hirohisa Nakamae, Takahide Ara, Yumiko Maruyama, Ken Ishiyama, Tatsuo Ichinohe, Yoshiko Atsuta, Bruce R. Blazar, Scott N. Furlan, Hiroyasu Ogawa, Geoffrey R. Hill

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Abstract

Genetic variants in Granulin (GRN), which encodes the secreted glycoprotein Progranulin (PGRN), are associated with several neurodegenerative diseases including frontotemporal lobar degeneration, neuronal ceroid lipofuscinosis, and Alzheimer’s disease. These genetic alterations manifest in pathological changes due to a reduction of PGRN expression; therefore, identifying factors that can modulate PGRN levels in vivo would enhance our understanding of PGRN in neurodegeneration, and could reveal novel potential therapeutic targets. Here, we report that modulation of the endocytosis-lysosomal pathway via reduction of Nemo-like kinase (Nlk) in microglia, and not neurons, can alter total brain Pgrn levels in mice. We demonstrate that Nlk reduction promotes Pgrn degradation by enhancing its trafficking through endocytosis-lysosomal pathway, specifically in microglia. Furthermore, genetic interaction studies in mice showed that Nlk heterozygosity in Grn haploinsufficient mice further reduces Pgrn levels and induces neuropathological phenotypes associated with PGRN deficiency. Our results reveal a new mechanism for Pgrn level regulation in the brain through the active catabolism by microglia and provide insights into the pathophysiology of PGRN-associated diseases.

Authors

Tingting Dong, Leon Tejwani, Youngseob Jung, Hiroshi Kokubu, Kimberly Luttik, Terri M. Driessen, Janghoo Lim

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