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The Lancet. Haematology May 2024Diamond-Blackfan anaemia (DBA), first described over 80 years ago, is a congenital disorder of erythropoiesis with a predilection for birth defects and cancer. Despite...
Diamond-Blackfan anaemia (DBA), first described over 80 years ago, is a congenital disorder of erythropoiesis with a predilection for birth defects and cancer. Despite scientific advances, this chronic, debilitating, and life-limiting disorder continues to cause a substantial physical, psychological, and financial toll on patients and their families. The highly complex medical needs of affected patients require specialised expertise and multidisciplinary care. However, gaps remain in effectively bridging scientific discoveries to clinical practice and disseminating the latest knowledge and best practices to providers. Following the publication of the first international consensus in 2008, advances in our understanding of the genetics, natural history, and clinical management of DBA have strongly supported the need for new consensus recommendations. In 2014 in Freiburg, Germany, a panel of 53 experts including clinicians, diagnosticians, and researchers from 27 countries convened. With support from patient advocates, the panel met repeatedly over subsequent years, engaging in ongoing discussions. These meetings led to the development of new consensus recommendations in 2024, replacing the previous guidelines. To account for the diverse phenotypes including presentation without anaemia, the panel agreed to adopt the term DBA syndrome. We propose new simplified diagnostic criteria, describe the genetics of DBA syndrome and its phenocopies, and introduce major changes in therapeutic standards. These changes include lowering the prednisone maintenance dose to maximum 0·3 mg/kg per day, raising the pre-transfusion haemoglobin to 9-10 g/dL independent of age, recommending early aggressive chelation, broadening indications for haematopoietic stem-cell transplantation, and recommending systematic clinical surveillance including early colorectal cancer screening. In summary, the current practice guidelines standardise the diagnostics, treatment, and long-term surveillance of patients with DBA syndrome of all ages worldwide.
Topics: Humans; Anemia, Diamond-Blackfan; Consensus; Disease Management; Hematopoietic Stem Cell Transplantation
PubMed: 38697731
DOI: 10.1016/S2352-3026(24)00063-2 -
Immunity May 2024The ubiquitin-binding endoribonuclease N4BP1 potently suppresses cytokine production by Toll-like receptors (TLRs) that signal through the adaptor MyD88 but is...
The ubiquitin-binding endoribonuclease N4BP1 potently suppresses cytokine production by Toll-like receptors (TLRs) that signal through the adaptor MyD88 but is inactivated via caspase-8-mediated cleavage downstream of death receptors, TLR3, or TLR4. Here, we examined the mechanism whereby N4BP1 limits inflammatory responses. In macrophages, deletion of N4BP1 prolonged activation of inflammatory gene transcription at late time points after TRIF-independent TLR activation. Optimal suppression of inflammatory cytokines by N4BP1 depended on its ability to bind polyubiquitin chains, as macrophages and mice-bearing inactivating mutations in a ubiquitin-binding motif in N4BP1 displayed increased TLR-induced cytokine production. Deletion of the noncanonical IκB kinases (ncIKKs), Tbk1 and Ikke, or their adaptor Tank phenocopied N4bp1 deficiency and enhanced macrophage responses to TLR1/2, TLR7, or TLR9 stimulation. Mechanistically, N4BP1 acted in concert with the ncIKKs to limit the duration of canonical IκB kinase (IKKα/β) signaling. Thus, N4BP1 and the ncIKKs serve as an important checkpoint against over-exuberant innate immune responses.
Topics: Animals; Mice; Signal Transduction; Inflammation; Toll-Like Receptors; Macrophages; I-kappa B Kinase; Protein Serine-Threonine Kinases; Mice, Knockout; Endoribonucleases; Ubiquitin; Cytokines; Mice, Inbred C57BL; Adaptor Proteins, Signal Transducing
PubMed: 38697117
DOI: 10.1016/j.immuni.2024.04.004 -
Molecular Neurobiology May 2024Heterozygous loss-of-function mutations in the progranulin gene (GRN) are a major cause of frontotemporal dementia due to progranulin haploinsufficiency; complete...
Heterozygous loss-of-function mutations in the progranulin gene (GRN) are a major cause of frontotemporal dementia due to progranulin haploinsufficiency; complete deficiency of progranulin causes neuronal ceroid lipofuscinosis. Several progranulin-deficient mouse models have been generated, including both knockout mice and knockin mice harboring a common patient mutation (R493X). However, the Grn mouse model has not been characterized completely. Additionally, while homozygous Grn and Grn knockout mice have been extensively studied, data from heterozygous mice is still limited. Here, we performed more in-depth characterization of heterozygous and homozygous Grn knockin mice, which includes biochemical assessments, behavioral studies, and analysis of fluid biomarkers. In the brains of homozygous Grn mice, we found increased phosphorylated TDP-43 along with increased expression of lysosomal genes, markers of microgliosis and astrogliosis, pro-inflammatory cytokines, and complement factors. Heterozygous Grn mice did not have increased TDP-43 phosphorylation but did exhibit limited increases in lysosomal and inflammatory gene expression. Behavioral studies found social and emotional deficits in Grn mice that mirror those observed in Grn knockout mouse models, as well as impairment in memory and executive function. Overall, the Grn knockin mouse model closely phenocopies Grn knockout models. Lastly, in contrast to homozygous knockin mice, heterozygous Grn mice do not have elevated levels of fluid biomarkers previously identified in humans, including neurofilament light chain (NfL) and glial fibrillary acidic protein (GFAP) in both plasma and CSF. These results may help to inform pre-clinical studies that use this Grn knockin mouse model and other Grn knockout models.
PubMed: 38696065
DOI: 10.1007/s12035-024-04190-9 -
Plant Communications Apr 2024The crosstalk between clathrin-mediated endocytosis (CME) and the autophagy pathway has been reported in mammals; however, the interconnection of CME with autophagy has...
The crosstalk between clathrin-mediated endocytosis (CME) and the autophagy pathway has been reported in mammals; however, the interconnection of CME with autophagy has not been established in plants. Here, we report that the Arabidopsis CLATHRIN LIGHT CHAIN (CLC) subunit 2 and 3 double mutant, clc2-1 clc3-1, phenocopies Arabidopsis AUTOPHAGY-RELATED GENE (ATG) mutants in both autoimmunity and nutrient sensitivity. Accordingly, the autophagy pathway is significantly compromised in the clc2-1 clc3-1 mutant. Interestingly, multiple assays demonstrate that CLC2 directly interacts with ATG8h/ATG8i in a domain-specific manner. As expected, both GFP-ATG8h/GFP-ATG8i and CLC2-GFP are subjected to autophagic degradation, and degradation of GFP-ATG8h is significantly reduced in the clc2-1 clc3-1 mutant. Notably, simultaneous knockout of ATG8h and ATG8i by CRISPR-Cas9 results in enhanced resistance against Golovinomyces cichoracearum, supporting the functional relevance of the CLC2-ATG8h/8i interactions. In conclusion, our results reveal a link between the function of CLCs and the autophagy pathway in Arabidopsis.
PubMed: 38693694
DOI: 10.1016/j.xplc.2024.100937 -
Nature Communications May 2024In diabetes, macrophages and inflammation are increased in the islets, along with β-cell dysfunction. Here, we demonstrate that galectin-3 (Gal3), mainly produced and...
In diabetes, macrophages and inflammation are increased in the islets, along with β-cell dysfunction. Here, we demonstrate that galectin-3 (Gal3), mainly produced and secreted by macrophages, is elevated in islets from both high-fat diet (HFD)-fed and diabetic db/db mice. Gal3 acutely reduces glucose-stimulated insulin secretion (GSIS) in β-cell lines and primary islets in mice and humans. Importantly, Gal3 binds to calcium voltage-gated channel auxiliary subunit gamma 1 (CACNG1) and inhibits calcium influx via the cytomembrane and subsequent GSIS. β-Cell CACNG1 deficiency phenocopies Gal3 treatment. Inhibition of Gal3 through either genetic or pharmacologic loss of function improves GSIS and glucose homeostasis in both HFD-fed and db/db mice. All animal findings are applicable to male mice. Here we show a role of Gal3 in pancreatic β-cell dysfunction, and Gal3 could be a therapeutic target for the treatment of type 2 diabetes.
Topics: Animals; Humans; Male; Mice; Calcium; Calcium Channels; Diabetes Mellitus, Type 2; Diet, High-Fat; Galectin 3; Glucose; Insulin; Insulin Secretion; Insulin-Secreting Cells; Macrophages; Mice, Inbred C57BL; Mice, Knockout
PubMed: 38693121
DOI: 10.1038/s41467-024-47959-1 -
American Journal of Physiology.... Jul 2024Pancreatic endocrine cells employ a sophisticated system of paracrine and autocrine signals to synchronize their activities, including glutamate, which controls hormone...
Hyperglycemia impairs EAAT2 glutamate transporter trafficking and glutamate clearance in islets of Langerhans: implications for type 2 diabetes pathogenesis and treatment.
Pancreatic endocrine cells employ a sophisticated system of paracrine and autocrine signals to synchronize their activities, including glutamate, which controls hormone release and β-cell viability by acting on glutamate receptors expressed by endocrine cells. We here investigate whether alteration of the excitatory amino acid transporter 2 (EAAT2), the major glutamate clearance system in the islet, may occur in type 2 diabetes mellitus and contribute to β-cell dysfunction. Increased EAAT2 intracellular localization was evident in islets of Langerhans from T2DM subjects as compared with healthy control subjects, despite similar expression levels. Chronic treatment of islets from healthy donors with high-glucose concentrations led to the transporter internalization in vesicular compartments and reduced [H]-d-glutamate uptake (65 ± 5% inhibition), phenocopying the findings in T2DM pancreatic sections. The transporter relocalization was associated with decreased Akt phosphorylation protein levels, suggesting an involvement of the phosphoinositide 3-kinase (PI3K)/Akt pathway in the process. In line with this, PI3K inhibition by a 100-µM LY294002 treatment in human and clonal β-cells caused the transporter relocalization in intracellular compartments and significantly reduced the glutamate uptake compared to control conditions, suggesting that hyperglycemia changes the trafficking of the transporter to the plasma membrane. Upregulation of the glutamate transporter upon treatment with the antibiotic ceftriaxone rescued hyperglycemia-induced β-cells dysfunction and death. Our data underscore the significance of EAAT2 in regulating islet physiology and provide a rationale for potential therapeutic targeting of this transporter to preserve β-cell survival and function in diabetes. The glutamate transporter SLC1A2/excitatory amino acid transporter 2 (EAAT2) is expressed on the plasma membrane of pancreatic β-cells and controls islet glutamate clearance and β-cells survival. We found that the EAAT2 membrane expression is lost in the islets of Langerhans from type 2 diabetes mellitus (T2DM) patients due to hyperglycemia-induced downregulation of the phosphoinositide 3-kinase/Akt pathway and modification of its intracellular trafficking. Pharmacological rescue of EAAT2 expression prevents β-cell dysfunction and death, suggesting EAAT2 as a new potential target of intervention in T2DM.
Topics: Excitatory Amino Acid Transporter 2; Humans; Diabetes Mellitus, Type 2; Glutamic Acid; Hyperglycemia; Islets of Langerhans; Male; Middle Aged; Female; Protein Transport; Insulin-Secreting Cells; Proto-Oncogene Proteins c-akt; Aged; Adult; Animals; Phosphatidylinositol 3-Kinases
PubMed: 38690938
DOI: 10.1152/ajpendo.00069.2024 -
Molecular Cancer Therapeutics Apr 2024Oncogene-driven expression and activation of receptor tyrosine kinases (RTK) promotes tumorigenesis and contributes to drug resistance. Increased expression of the...
Oncogene-driven expression and activation of receptor tyrosine kinases (RTK) promotes tumorigenesis and contributes to drug resistance. Increased expression of the kinases DDR2 (Discoid Domain Receptor 2), RET, PDGFRA, KIT, MET, and ALK (Anaplastic Lymphoma Kinase) independently correlate with decreased overall survival (OS) and event free survival (EFS) of pediatric neuroblastoma. The multikinase inhibitor sitravatinib targets DDR2, RET, PDGFRA, KIT and MET with low nanomolar activity and we therefore tested its efficacy against orthotopic and syngeneic tumor models. Sitravatinib markedly reduced cell proliferation and migration in vitro independently of MYCN (N-Myc proto-oncogene), ALK, or MYC (c-Myc proto-oncogene) status, and inhibited proliferation and metastasis of human orthotopic xenografts. Oral administration of sitravatinib to homozygous Th-MYCN transgenic mice (Th-MYCN+/+) after tumor initiation completely arrested further tumor development with no mice dying of disease while maintained on sitravatinib treatment (control cohort 57 days median time to sacrifice). Among these top kinases, DDR2 expression has the strongest correlation with poor survival and high stage at diagnosis, and the highest sensitivity to the drug. We confirmed on-target inhibition of collagen-mediated activation of DDR2. Genetic knockdown of DDR2 partially phenocopies Sitravatinib treatment, limiting tumor development and metastasis across tumor models. Analysis of single cell sequencing data demonstrated that DDR2 is restricted to mesenchymal-type tumor subpopulations and is enriched in Schwann Cell Precursor (SCP) subpopulations found in high-risk disease. These data define an unsuspected role for sitravatinib as a therapeutic agent in neuroblastoma and reveal a novel function for DDR2 as a driver of tumor growth and metastasis.
PubMed: 38670553
DOI: 10.1158/1535-7163.MCT-23-0741 -
Science Advances Apr 2024While the kinesin-2 motors KIF3A and KIF3B have essential roles in ciliogenesis and Hedgehog (HH) signal transduction, potential role(s) for another kinesin-2 motor,...
While the kinesin-2 motors KIF3A and KIF3B have essential roles in ciliogenesis and Hedgehog (HH) signal transduction, potential role(s) for another kinesin-2 motor, KIF17, in HH signaling have yet to be explored. Here, we investigated the contribution of KIF17 to HH-dependent cerebellar development, where is expressed in both HH-producing Purkinje cells and HH-responding cerebellar granule neuron progenitors (CGNPs). Germline deletion in mice results in cerebellar hypoplasia due to reduced CGNP proliferation, a consequence of decreased HH pathway activity mediated through decreased Sonic HH (SHH) protein. Notably, Purkinje cell-specific deletion partially phenocopies germline mutants. Unexpectedly, CGNP-specific deletion results in the opposite phenotype-increased CGNP proliferation and HH target gene expression due to altered GLI transcription factor processing. Together, these data identify KIF17 as a key regulator of HH-dependent cerebellar development, with dual and opposing roles in HH-producing Purkinje cells and HH-responding CGNPs.
Topics: Animals; Kinesins; Cerebellum; Hedgehog Proteins; Mice; Purkinje Cells; Signal Transduction; Cell Proliferation; Mice, Knockout; Gene Expression Regulation, Developmental; Neural Stem Cells; Zinc Finger Protein GLI1; Developmental Disabilities; Nervous System Malformations
PubMed: 38669326
DOI: 10.1126/sciadv.ade1650 -
BioRxiv : the Preprint Server For... Apr 2024Astrocytes play critical roles in supporting structural and metabolic homeostasis in the central nervous system (CNS). Inflammatory conditions bring about a range of...
Astrocytes play critical roles in supporting structural and metabolic homeostasis in the central nervous system (CNS). Inflammatory conditions bring about a range of poorly understood, heterogeneous, reactive phenotypes in astrocytes. Finding ways to manipulate the phenotype of reactive astrocytes, and leveraging a pro-recovery phenotype, holds promise in treating CNS injury. Previous studies have shown that the protein transglutaminase 2 (TG2) plays a significant role in determining the phenotype of reactive astrocytes. Recently it has been demonstrated that ablation of TG2 from astrocytes improves injury outcomes both and . Excitingly, in an mouse model, pharmacological inhibition of TG2 with the irreversible inhibitor VA4 phenocopies the neurosupportive effects of TG2 deletion in astrocytes. The focus of this study was to provide insights into the mechanisms by which TG2 deletion or inhibition of TG2 with VA4 result in a more neurosupportive astrocytic phenotype. Using a neuron-astrocyte co-culture model of neurite outgrowth, we show that VA4 treatment improves the ability of astrocytes to support neurite outgrowth on an injury-relevant matrix, further validating the ability of VA4 to phenocopy astrocytic TG2 deletion. VA4 treatment of neurons alone had no effect on neurite outgrowth. VA4 covalently binds to active site residues of TG2 that are exposed in its open conformation and are critical for its enzymatic function, and prevents TG2 from taking on a closed conformation, which interferes with its protein scaffolding function. To begin to understand how pharmacologically altering TG2's conformation affects its ability to regulate reactive astrocyte phenotypes, we assayed the impact of VA4 on TG2's interaction with Zbtb7a, a transcription factor that we have previously identified as a TG2 interactor, and whose functional outputs are significantly regulated by TG2. The results of these studies demonstrated that VA4 significantly decreases the interaction of TG2 and Zbtb7a. Further, previous findings indicate that TG2 may act as an epigenetic regulator, through its nuclear protein-protein interactions, to modulate gene expression. Since both TG2 and Zbtb7a interact with members of the Sin3a chromatin repressor complex, we assayed the effect of TG2 deletion and VA4 treatment on histone acetylation and found significantly greater acetylation with TG2 deletion or inhibition with VA4. Overall, this work points toward a possible epigenetic mechanism by which genetic deletion or acute inhibition of TG2 leads to enhanced astrocytic support of neurons.
PubMed: 38659783
DOI: 10.1101/2024.04.15.589192 -
Cell Death Discovery Apr 2024Pancreatic cancer is one of the most fatal cancers in the world. A growing number of studies have begun to demonstrate that mitochondria play a key role in...
Pancreatic cancer is one of the most fatal cancers in the world. A growing number of studies have begun to demonstrate that mitochondria play a key role in tumorigenesis. Our previous study reveals that NDUFS2 (NADH: ubiquinone oxidoreductase core subunit S2), a core subunit of the mitochondrial respiratory chain complex I, is upregulated in Pancreatic adenocarcinoma (PAAD). However, its role in the development of PAAD remains unknown. Here, we showed that NDUFS2 played a critical role in the survival, proliferation and migration of pancreatic cancer cells by inhibiting mitochondrial cell death. Additionally, protein mass spectrometry indicated that the NDUFS2 was interacted with a deubiquitinase, OTUB1. Overexpression of OTUB1 increased NDUFS2 expression at the protein level, while knockdown of OTUB1 restored the effects in vitro. Accordingly, overexpression and knockdown of OTUB1 phenocopied those of NDUFS2 in pancreatic cancer cells, respectively. Mechanically, NDUFS2 was deubiquitinated by OTUB1 via K48-linked polyubiquitin chains, resulted in an elevated protein stability of NDUFS2. Moreover, the growth of OTUB1-overexpressed pancreatic cancer xenograft tumor was promoted in vivo, while the OTUB1-silenced pancreatic cancer xenograft tumor was inhibited in vivo. In conclusion, we revealed that OTUB1 increased the stability of NDUFS2 in PAAD by deubiquitylation and this axis plays a pivotal role in pancreatic cancer tumorigenesis and development.
PubMed: 38653740
DOI: 10.1038/s41420-024-01948-x