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Gaucher disease (GD) is caused by biallelic GBA1/Gba1 mutations that encode defective glucocerebrosidase (GCase). Progranulin (PGRN, encoded by GRN/Grn) is a modifier of GCase, but the interplay between PGRN and GCase, specifically GBA1/Gba1 mutations, contributing to GD severity is unclear. Mouse models were developed with various dosages of Gba1 D409V mutation against the PGRN deficiency (Grn-/-) [Grn-/-;Gba1D409V/WT (PG9Vwt), Grn-/-;Gba1D409V/D409V (PG9V), Grn-/-;Gba1D409V/Null (PG9VN)]. Disease progression in those mouse models was characterized by biochemical, pathological, transcriptomic, and neurobehavioral analyses. Compared to PG9Vwt, Grn-/-;Gba1WT/Null and Grn-/- mice that had a higher level of GCase activity and undetectable pathologies, homozygous or hemizygous D409V in PG9V or PG9VN, respectively, resulted in profound inflammation and neurodegeneration. PG9VN mice exhibited much earlier onset, shorter life span, tissue fibrosis, and more severe phenotypes than PG9V mice. Glycosphingolipid accumulation, inflammatory responses, lysosomal-autophagy dysfunction, microgliosis, retinal gliosis, as well as α-Synuclein increases were much more pronounced in PG9VN mice. Neurodegeneration in PG9VN was characterized by activated microglial phagocytosis of impaired neurons and programmed cell death due to necrosis and, possibly, pyroptosis. Brain transcriptomic analyses revealed the intrinsic relationship between D409V dosage, and the degree of altered gene expression related to lysosome dysfunction, microgliosis, and neurodegeneration in GD, suggesting the disease severity is dependent on a GCase activity threshold related to Gba1 D409V dosage and loss of PGRN. These findings contribute to a deeper understanding of GD pathogenesis by elucidating additional underlying mechanisms of interplay between PGRN and Gba1 mutation dosage in modulating GCase function and disease severity in GD and GBA1-associated neurodegenerative diseases.
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The inhibition of tumor necrosis factor (TNF)-α trimer formation renders it inactive for binding to its receptors, thus mitigating the vicious cycle of inflammation. We designed a peptide (PIYLGGVFQ) that simulates a sequence strand of human TNFα monomer using a series of in silico methods, such as active site finding (Acsite), protein-protein interaction (PPI), docking studies (GOLD and Flex-X) followed by molecular dynamics (MD) simulation studies. The MD studies confirmed the intermolecular interaction of the peptide with the TNFα. Fluorescence-activated cell sorting and fluorescence microscopy revealed that the peptide effectively inhibited the binding of TNF to the cell surface receptors. The cell culture assays showed that the peptide significantly inhibited the TNFα-mediated cell death. In addition, the nuclear translocation of the nuclear factor kappa B (NFκB) was significantly suppressed in the peptide-treated A549 cells, as observed in immunofluorescence and gel mobility-shift assays. Furthermore, the peptide protected against joint damage in the collagen-induced arthritis (CIA) mouse model, as revealed in the micro focal-CT scans. In conclusion, this TNFα antagonist would be helpful for the prevention and repair of inflammatory bone destruction and subsequent loss in the mouse model of CIA as well as human rheumatoid arthritis (RA) patients. This calls upon further clinical investigation to utilize its potential effect as an antiarthritic drug.
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Péptidos , Factor de Necrosis Tumoral alfa , Humanos , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Animales , Ratones , Péptidos/farmacología , Péptidos/química , Artritis Experimental/tratamiento farmacológico , Artritis Experimental/metabolismo , Artritis Experimental/patología , Simulación del Acoplamiento Molecular , Células A549 , Simulación de Dinámica Molecular , FN-kappa B/metabolismo , FN-kappa B/antagonistas & inhibidores , Masculino , Antirreumáticos/farmacología , Antirreumáticos/química , Antirreumáticos/uso terapéutico , Unión Proteica , Modelos Animales de EnfermedadRESUMEN
Obesity has a multifactorial etiology and is known to be a state of chronic low-grade inflammation, known as meta-inflammation. This state is associated with the development of metabolic disorders such as glucose intolerance and nonalcoholic fatty liver disease. Pyruvate is a glycolytic metabolite and a crucial node in various metabolic pathways. However, its role and molecular mechanism in obesity and associated complications are obscure. In this study, we reported that pyruvate substantially inhibited adipogenic differentiation in vitro and its administration significantly prevented HFD-induced weight gain, white adipose tissue inflammation, and metabolic dysregulation. To identify the target proteins of pyruvate, drug affinity responsive target stability was employed with proteomics, cellular thermal shift assay, and isothermal drug response to detect the interactions between pyruvate and its molecular targets. Consequently, we identified cytosolic phospholipase A2 (cPLA2) as a novel molecular target of pyruvate and demonstrated that pyruvate restrained diet-induced obesity, white adipose tissue inflammation, and hepatic steatosis in a cPLA2-dependent manner. Studies with global ablation of cPLA2 in mice showed that the protective effects of pyruvate were largely abrogated, confirming the importance of pyruvate/cPLA2 interaction in pyruvate attenuation of inflammation and obesity. Overall, our study not only establishes pyruvate as an antagonist of cPLA2 signaling and a potential therapeutic option for obesity, but it also sheds light on the mechanism of its action. Pyruvate's prior clinical use indicates that it can be considered a safe and viable alternative for obesity, whether consumed as a dietary supplement or as part of a regular diet.
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BACKGROUND: In recent years, the implementation of the first case of pig-to-human heart xenotransplantation and the report of three cases of pig-to-brain-dead human recipient kidney transplantation indicate that xenotransplantation is getting closer to clinical application. In the near future, China may also launch clinical trials of kidney xenotransplantation. Therefore, it is necessary to investigate the level of knowledge and acceptance of xenotransplantation among kidney transplant recipients and candidates in China. This study aims to investigate the level of comprehension and acceptance of kidney xenotransplantation in kidney transplant recipients and explore related factors, providing a reference for promoting the application and clinical trials of xenotransplantation in the near future. METHODS: A questionnaire was completed by 211 kidney transplant recipients and 21 candidates. Answers to the questionnaires were self-administered by the respondents. Scores were compared using nonparametric tests, as well as using Chi-square test or Fisher's exact test to compare differences in answers. RESULTS: Respondents demonstrated a high score of 75 (out of 100) on knowledge and acceptance of kidney xenotransplantation. The sector "Knowledge and Attitude" received the overall highest score from respondents (85.0 out of 100), while "Risks and Concerns" received the lowest score (50 out of 100). Interestingly, respondents paid more attention to infection risks but showed less concern about rejection or unknown risks. Furthermore, 191 respondents (82.3%) expected that pig kidney xenografts could function for at least 5 years or more. The scores were statistically significant in terms of age, gender, level of education, level of knowledge on the case of xenotransplantation clinical trial, and willingness to donate organs. cognition CONCLUSIONS: The awareness rates of xenotransplantation are high among kidney transplant recipients and candidates, for which the majority showed a generally favorable attitude towards this procedure. Respondents did not have a comprehensive understanding of the specific knowledge of xenotransplantation and expressed more concern about the risk of infection compared to the risk of rejection and other unknown risks, while also expecting long-term survival similar to allograft transplantation for pig kidney xenografts.
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Trasplante de Riñón , Animales , Humanos , China , Xenoinjertos , Riñón , Porcinos , Trasplante Heterólogo , Ensayos Clínicos como AsuntoRESUMEN
BACKGROUND: Tumor-infiltrating T cells enter an exhausted or dysfunctional state, which limits antitumor immunity. Among exhausted T cells, a subset of cells with features of progenitor or stem-like cells has been identified as TCF1+ CD8+ T cells that respond to immunotherapy. In contrast to the finding that TCF1 controls epigenetic and transcriptional reprogramming in tumor-infiltrating stem-like T cells, little is known about the regulation of TCF1. Emerging data show that elevated body mass index is associated with outcomes of immunotherapy. However, the mechanism has not been clarified. METHODS: We investigated the proliferation of splenic lymphocytes or CD8+ T cells induced by CD3/CD28 stimulation in vitro. We evaluated the effects of low-density lipoprotein (LDL) and LRP11 inhibitors, as well as MAPK13 inhibitors. Additionally, we used shRNA technology to validate the roles of LRP11 and MAPK13. In an in vivo setting, we employed male C57BL/6J injected with B16 cells or MC38 cells to build a tumor model to assess the effects of LDL and LRP11 inhibitors, LRP11 activators, MAPK13 inhibitors on tumor growth. Flow cytometry was used to measure cell proportions and activation status. Molecular interactions and TCF1 status were examined using Western blotting. Moreover, we employed RNA sequencing to investigate the effects of LDL stimulation and MAPK13 inhibition in CD8+ T cells. RESULTS: By using a tumor-bearing mouse model, we found that LDL-induced tumor-infiltrating TCF1+PD1+CD8+ T cells. Using a cell-based chimeric receptor screening system, we showed that LRP11 interacted with LDL and activated TCF1. LRP11 activation enhanced TCF1+PD1+CD8+ T-cell-mediated antitumor immunity, consistent with LRP11 blocking impaired T-cell function. Mechanistically, LRP11 activation induces MAPK13 activation. Then, MAPK13 phosphorylates TCF1, leading to increase of stem-like T cells. CONCLUSIONS: LRP11-MAPK13-TCF1 enhanced antitumor immunity and induced tumor-infiltrating stem-like T cells.
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Linfocitos T CD8-positivos , Melanoma Experimental , Masculino , Ratones , Animales , Fosforilación , Receptor de Muerte Celular Programada 1 , Ratones Endogámicos C57BL , InmunoterapiaRESUMEN
Osteoarthritis (OA) is the most common joint disease. Currently there are no effective methods that simultaneously prevent joint degeneration and reduce pain1. Although limited evidence suggests the existence of voltage-gated sodium channels (VGSCs) in chondrocytes2, their expression and function in chondrocytes and in OA remain essentially unknown. Here we identify Nav1.7 as an OA-associated VGSC and demonstrate that human OA chondrocytes express functional Nav1.7 channels, with a density of 0.1 to 0.15 channels per µm2 and 350 to 525 channels per cell. Serial genetic ablation of Nav1.7 in multiple mouse models demonstrates that Nav1.7 expressed in dorsal root ganglia neurons is involved in pain, whereas Nav1.7 in chondrocytes regulates OA progression. Pharmacological blockade of Nav1.7 with selective or clinically used pan-Nav channel blockers significantly ameliorates the progression of structural joint damage, and reduces OA pain behaviour. Mechanistically, Nav1.7 blockers regulate intracellular Ca2+ signalling and the chondrocyte secretome, which in turn affects chondrocyte biology and OA progression. Identification of Nav1.7 as a novel chondrocyte-expressed, OA-associated channel uncovers a dual target for the development of disease-modifying and non-opioid pain relief treatment for OA.
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Condrocitos , Canal de Sodio Activado por Voltaje NAV1.7 , Osteoartritis , Bloqueadores del Canal de Sodio Activado por Voltaje , Animales , Humanos , Ratones , Calcio/metabolismo , Señalización del Calcio/efectos de los fármacos , Condrocitos/efectos de los fármacos , Condrocitos/metabolismo , Progresión de la Enfermedad , Ganglios Espinales/citología , Ganglios Espinales/metabolismo , Canal de Sodio Activado por Voltaje NAV1.7/deficiencia , Canal de Sodio Activado por Voltaje NAV1.7/genética , Canal de Sodio Activado por Voltaje NAV1.7/metabolismo , Neuronas/metabolismo , Osteoartritis/complicaciones , Osteoartritis/tratamiento farmacológico , Osteoartritis/genética , Osteoartritis/metabolismo , Dolor/complicaciones , Dolor/tratamiento farmacológico , Dolor/metabolismo , Bloqueadores del Canal de Sodio Activado por Voltaje/farmacología , Bloqueadores del Canal de Sodio Activado por Voltaje/uso terapéuticoRESUMEN
Autoimmune hepatitis is a serious hepatic disorder with unknown nosogenesis, and natural products have been deemed to be one of the most significant sources of new drugs against this disease. Prenyllongnols A-D (1-4), four undescribed prenylated acylphloroglucinols, were isolated from Hypericum longistylum. Compounds 1-4 exhibited remarkable immunosuppressive activities in murine splenocyte proliferation under the induction of concanavalin A (Con A), and IC50 values ranged from 2.98 ± 0.21 to 6.34 ± 0.72 µM. Furthermore, in a Con A-challenged autoimmune hepatitis mouse model, the mice in the group that were pretreated with isolate 2 significantly ameliorated liver injury and decreased proinflammatory cytokine production. Notably, natural product 2 was the first prenylated acylphloroglucinol to protect against concanavalin A-induced autoimmune hepatitis. This finding underscores the potential of prenylated acylphloroglucinol-type metabolites as promising candidates for designing novel immunosuppressors in the quest for new antiautoimmune hepatitis drugs.
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Hepatitis Autoinmune , Hypericum , Animales , Ratones , Concanavalina A , Hepatitis Autoinmune/tratamiento farmacológico , Hepatitis Autoinmune/etiología , Floroglucinol/farmacología , InmunosupresoresRESUMEN
Hepatic ischemia/reperfusion injury (IRI) is a major factor contributing to the failure of hepatic resection and liver transplantation. As part of our ongoing investigation into bioactive compounds derived from fungi, we isolated eight indole alkaloids (1-8) from the endophytic fungus Aspergillus amoenus TJ507. Among these alkaloids, one previously undescribed compound, amoenamide D (1), was identified. The planar structure of 1 was elucidated by extensive spectroscopic analysis, including HRESIMS and NMR spectra. The absolute configuration of 1 was elucidated by using electronic circular dichroism calculations. Notably, in the CoCl2-induced hepatocyte damage model, notoamide Q (3) exhibited significant anti-hypoxia injury activity. Furthermore, in a murine hepatic ischemia/reperfusion injury model, treatment with 3 prevents IRI-induced liver damage and hepatocellular apoptosis. Consequently, 3 might serve as a potential lead compound to prevent hepatic ischemia/reperfusion injury.
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Hepatopatías , Daño por Reperfusión , Ratones , Animales , Hígado , Hongos , Alcaloides Indólicos/química , Daño por Reperfusión/tratamiento farmacológico , IsquemiaRESUMEN
Objective: This study is aimed to determine the metabolomic effects of the hybrid medicine formula Yi-Qi-Bu-Shen (YQBS) on the neurotransmitter aspects of cognitive impairment in diabetic rats. Methods: In the current study, streptozotocin (STZ) was used to induce diabetic animal model in male Sprague Dawley (SD) rats. After successful establishment of diabetic SD rats' model, age-matched healthy SD rats and diabetic SD rats were treated with low and high doses of YQBS, and then tested for learning memory ability and analyzed for pathological changes. In addition, neurotransmitter metabolic changes in hippocampal subdivisions of rats from different treated groups were analyzed using liquid chromatography-mass spectrometry (LC-MS) technique. Results: YQBS could significantly improve memory-cognitive impairment in diabetic rats as evidenced by the shortening of latency to target and the reduction of latency first entrance to target. Moreover, YQBS also improved the pathological alterations in the hippocampal region in the brains of diabetic rats. Metabolomic analysis showed that the expression of noradrenaline hydrochloride was down-regulated and the expressions of levodopa and 5-hydroxytryptophan were up-regulated in the hippocampal tissues of diabetic rats treated with YQBS. Conclusion: These findings demonstrate that YQBS has protective effects against diabetic cognitive dysfunction, which might act through alteration in tyrosine and tryptophan metabolism.
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Hepatic ischemia/reperfusion injury is a major cause of hypohepatia after surgical procedures such as hypovolemic shock, transplantation, and so on. In our continuous study of bioactive natural products from fungus, eight ergosterol-type sterides (1-8), including two undescribed compounds, sterolaspers A (1) and B (2), were isolated from Aspergillus sp. TJ507. Structure elucidation was accomplished by extensive spectroscopic analysis and comparison with the reported NMR data as well as X-Ray single crystal diffraction tests. Activity screen of these isolates showed 5α-stigmast-3,6-dione (3) possessing anti-hypoxia injury effects against CoCl2-induced hypoxia damage in hepatocytes. More importantly, compound 3 could improve liver function, alleviate liver damage, and restrain the hepatocellular apoptosis in hepatic ischemia/reperfusion injury murine model. As such, this ergosterol-type steride, 5α-stigmast-3,6-dione (3), might serve as lead structure for the development of novel hepatoprotective agents in the clinical treatment of hepatic ischemia/reperfusion injury.
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Hígado , Daño por Reperfusión , Ratones , Animales , Hepatocitos , Daño por Reperfusión/tratamiento farmacológico , Apoptosis , Isquemia/complicaciones , AspergillusRESUMEN
Extracellular regulated protein kinases (ERK) signaling is a master regulator of cell behavior, life, and fate. Although ERK pathway is shown to be involved in T-cell activation, little is known about its role in the development of allograft rejection. Here, it is reported that ERK signaling pathway is activated in allograft-infiltrating T cells. On the basis of surface plasmon resonance technology, lycorine is identified as an ERK-specific inhibitor. ERK inhibition by lycorine significantly prolongs allograft survival in a stringent mouse cardiac allotransplant model. As compared to untreated mice, lycorine-treated mice show a decrease in the number and activation of allograft-infiltrated T cells. It is further confirmed that lycorine-treated mouse and human T cells are less responsive to stimulation in vitro, as indicated by their low proliferative rates and decreased cytokine production. Mechanistic studies reveal that T cells treated with lycorine exhibit mitochondrial dysfunction, resulting in metabolic reprogramming upon stimulation. Transcriptome analysis of lycorine-treated T cells reveals an enrichment in a series of downregulated terms related to immune response, the mitogen-activated protein kinase cascade, and metabolic processes. These findings offer new insights into the development of immunosuppressive agents by targeting the ERK pathway involved in T-cell activation and allograft rejection.
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Alcaloides de Amaryllidaceae , Linfocitos T , Ratones , Humanos , Animales , Proteínas Quinasas/metabolismo , Alcaloides de Amaryllidaceae/metabolismo , Proteínas/metabolismo , AloinjertosRESUMEN
Gaucher disease (GD), one of the most common lysosomal storage diseases, is caused by GBA1 mutations resulting in defective glucocerebrosidase (GCase) and consequent accumulation of its substrates ß-glucosylceramide (ß-GlcCer). We reported progranulin (PGRN), a secretary growth factor-like molecule and an intracellular lysosomal protein was a crucial co-factor of GCase. PGRN binds to GCase and recruits Heat Shock Protein 70 (Hsp70) to GCase through its C-terminal Granulin (Grn) E domain, termed as ND7. In addition, both PGRN and ND7 are therapeutic against GD. Herein we found that both PGRN and its derived ND7 still displayed significant protective effects against GD in Hsp70 deficient cells. To delineate the molecular mechanisms underlying PGRN's Hsp70-independent regulation of GD, we performed a biochemical co-purification and mass spectrometry with His-tagged PGRN and His-tagged ND7 in Hsp70 deficient cells, which led to the identification of ERp57, also referred to as protein disulfide isomerase A3 (PDIA3), as a protein that binds to both PGRN and ND7. Within type 2 neuropathic GD patient fibroblasts L444P, bearing GBA1 L444P mutation, deletion of ERp57 largely abolished the therapeutic effects of PGRN and ND7, as manifested by loss of effects on lysosomal storage, GCase activity, and ß-GlcCer accumulation. Additionally, recombinant ERp57 effectively restored the therapeutic effects of PGRN and ND7 in ERp57 knockout L444P fibroblasts. Collectively, this study reports ERp57 as a previously unrecognized binding partner of PGRN that contributes to PGRN regulation of GD.
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Enfermedad de Gaucher , Progranulinas , Humanos , Enfermedad de Gaucher/tratamiento farmacológico , Enfermedad de Gaucher/genética , Enfermedad de Gaucher/metabolismo , Glucosilceramidasa/genética , Glucosilceramidasa/metabolismo , Glucosilceramidasa/uso terapéutico , Lisosomas/metabolismo , Mutación , Progranulinas/genética , Progranulinas/metabolismo , Progranulinas/uso terapéutico , Proteína Disulfuro Isomerasas/genética , Proteína Disulfuro Isomerasas/metabolismo , Proteína Disulfuro Isomerasas/uso terapéuticoRESUMEN
Concanavalin A (Con A) is known to be a T-cell mitogen and has been shown to induce hepatitis in mice through the triggering of conventional T cells and NKT cells. However, it remains unknown whether Con A itself can directly induce rapid hepatocyte death in the absence of a functional immune system. Here, by using an immunodeficient mouse model, we found Con A rapidly induced liver injury in vivo despite a lack of immunocyte involvement. We further observed in vitro that hepatocytes underwent a dose-dependent but caspase-independent apoptosis in response to Con A stimulation in vitro. Moreover, transcriptome RNA-sequencing analysis revealed that apoptosis pathways were activated in both our in vivo and in vitro models. We conclude that Con A can directly induce rapid but non-classical apoptosis in hepatocytes without the participation of immunocytes. These findings provide new insights into the mechanism of Con A-induced hepatitis.
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Enfermedad Hepática Inducida por Sustancias y Drogas , Hepatitis , Animales , Ratones , Caspasas/metabolismo , Concanavalina A/farmacología , Concanavalina A/metabolismo , Hepatocitos , Apoptosis , Hígado , Hepatitis/metabolismo , Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismoRESUMEN
Mutations in GBA1, encoding glucocerebrosidase (GCase), cause Gaucher disease (GD) and are also genetic risks in developing Parkinson's disease (PD). Currently, the approved therapies are only effective for directly treating visceral symptoms, but not for primary neuronopathic involvement in GD (nGD). Progranulin (PGRN), encoded by GRN, is a novel modifier of GCase, but the impact of PGRN in GBA1 mutation-associated pathologies in vivo remains unknown. Herein, Grn-/- mice crossed into Gba9v/9v mice, a Gba1 mutant line homozygous for the Gba1 D409V mutation, generating Grn-/-Gba9v/9v (PG9V) mice. PG9V mice exhibited neurobehavioral deficits, early onset, and more severe GD phenotypes compared to Grn-/- and Gba9v/9v mice. Moreover, PG9V mice also displayed PD-like phenotype. Mechanistic analysis revealed that PGRN deficiency caused severe neuroinflammation with microgliosis and astrogliosis, along with impaired autophagy associated with the Gba1 mutation. A PGRN-derived peptide, termed ND7, ameliorated the disease phenotype in GD patient fibroblasts ex vivo. Unexpectedly, ND7 penetrated the blood-brain barrier (BBB) and effectively ameliorated the nGD manifestations and PD pathology in Gba9v/null and PG9V mice. Collectively, this study not only provides the first line of in vivo but also ex vivo evidence demonstrating the crucial role of PGRN in GBA1/Gba1 mutation-related pathologies, as well as a clinically relevant mouse model for mechanistic and potential therapeutics studies for nGD and PD. Importantly, a BBB penetrant PGRN-derived biologic was developed that may provide treatment for rare lysosomal storage diseases and common neurodegenerative disorders, particularly nGD and PD.
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Enfermedad de Gaucher , Enfermedad de Parkinson , Progranulinas , Animales , Ratones , Encéfalo/metabolismo , Enfermedad de Gaucher/genética , Glucosilceramidasa/genética , Glucosilceramidasa/metabolismo , Lisosomas/metabolismo , Mutación , Enfermedad de Parkinson/genética , Progranulinas/genética , Ratones NoqueadosRESUMEN
Neurodegenerative diseases are debilitating impairments that affect millions of people worldwide and are characterized by progressive degeneration of structure and function of the central or peripheral nervous system. Effective biomarkers for neurodegenerative diseases can be used to improve the diagnostic workup in the clinic as well as facilitate the development of effective disease-modifying therapies. Progranulin (PGRN) has been reported to be involved in various neurodegenerative disorders. Hence, in the current study we systematically compared the inflammation and accumulation of typical neurodegenerative disease markers in the brain tissue between PGRN knockout (PGRN KO) and wildtype (WT) mice. We found that PGRN deficiency led to significant neuron loss as well as activation of microglia and astrocytes in aged mice. Several characteristic neurodegenerative markers, including α-synuclein, TAR DNA-binding protein 43 (TDP-43), Tau, and ß-amyloid, were all accumulated in the brain of PGRN-deficient mice as compared to WT mice. Moreover, higher aggregation of lipofuscin was observed in the brain tissue of PGRN-deficient mice compared with WT mice. In addition, the autophagy was also defective in the brain of PGRN-deficient mice, indicated by the abnormal expression level of autophagy marker LC3-II. Collectively, comprehensive assays support the idea that PGRN plays an important role during the development of neurodegenerative disease, indicating that PGRN might be a useful biomarker for neurodegenerative diseases in clinical settings.
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Envejecimiento/patología , Biomarcadores/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Progranulinas/deficiencia , Péptidos beta-Amiloides/metabolismo , Animales , Astrocitos/metabolismo , Astrocitos/patología , Autofagia , Encéfalo/patología , Encéfalo/ultraestructura , Proteínas de Unión al ADN/metabolismo , Lipofuscina/metabolismo , Ratones Noqueados , Microglía/metabolismo , Microglía/patología , Neuronas/metabolismo , Neuronas/patología , Fosforilación , Progranulinas/metabolismo , Agregado de Proteínas , alfa-Sinucleína/metabolismo , Proteínas tau/metabolismoRESUMEN
Inflammatory Bowel Disease (IBD) is an autoimmune condition with complicated pathology and diverse clinical signs. TNFα is believed to play a crucial role in the pathogenesis of IBD. We recently identified fexofenadine, a well-known antagonist of histamine H1 receptor, as a novel inhibitor of TNFα signaling. Additionally, cytosolic phospholipase A2 (cPLA2) was isolated as a binding target of fexofenadine, and fexofenadine-mediated anti-TNF activity relied on cPLA2 in vitro. The objective of this study is to determine whether fexofenadine is therapeutic against chemically-induced murine IBD model and whether cPLA2 and/or histamine H1 receptor is important for fexofenadine's anti-inflammatory activity in vivo by leveraging various genetically modified mice and chemically induced murine IBD models. Both dextran sulfate sodium- and 2, 4, 6-trinitrobenzene sulfonic acid-induced murine IBD models revealed that orally delivered fexofenadine was therapeutic against IBD, evidenced by mitigated clinical symptoms, decreased secretions of the proinflammatory cytokine IL-6 and IL-1ß, lowered intestinal inflammation, and reduced p-p65 and p-IĸBα. Intriguingly, Fexofenadine-mediated protective effects against IBD were lost in cPLA2 deficient mice but not in histamine H1 receptor-deficient mice. Collectively, these findings demonstrate the therapeutic effects of over-the-counter drug Fexofenadine in treating DSS-induced IBD murine and provide first in vivo evidence showing that cPLA2 is required for fexofenadine's therapeutic effects in murine IBD model and probably other inflammatory and autoimmune diseases as well.
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Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Fosfolipasas A2 Citosólicas/fisiología , Terfenadina/análogos & derivados , Animales , Biomarcadores Farmacológicos , Modelos Animales de Enfermedad , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fosfolipasas A2 Citosólicas/genética , Terfenadina/uso terapéuticoRESUMEN
Progranulin (PGRN) is a key regulator of lysosomes, and its deficiency has been linked to various lysosomal storage diseases (LSDs), including Gaucher disease (GD), one of the most common LSD. Here, we report that PGRN plays a previously unrecognized role in autophagy within the context of GD. PGRN deficiency is associated with the accumulation of LC3-II and p62 in autophagosomes of GD animal model and patient fibroblasts, resulting from the impaired fusion of autophagosomes and lysosomes. PGRN physically interacted with Rab2, a critical molecule in autophagosome-lysosome fusion. Additionally, a fragment of PGRN containing the Grn E domain was required and sufficient for binding to Rab2. Furthermore, this fragment significantly ameliorated PGRN deficiency-associated impairment of autophagosome-lysosome fusion and autophagic flux. These findings not only demonstrate that PGRN is a crucial mediator of autophagosome-lysosome fusion but also provide new evidence indicating PGRN's candidacy as a molecular target for modulating autophagy in GD and other LSDs in general. KEY MESSAGES : PGRN acts as a crucial factor involved in autophagosome-lysosome fusion in GD. PGRN physically interacts with Rab2, a molecule in autophagosome-lysosome fusion. A 15-kDa C-terminal fragment of PGRN is required and sufficient for binding to Rab2. This PGRN derivative ameliorates PGRN deficiency-associated impairment of autophagy. This study provides new insights into autophagy and may develop novel therapy for GD.
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Autofagosomas , Enfermedad de Gaucher/metabolismo , Lisosomas , Progranulinas/metabolismo , Proteína de Unión al GTP rab2/metabolismo , Alérgenos/administración & dosificación , Animales , Células Cultivadas , Humanos , Ratones Noqueados , Ovalbúmina/administración & dosificación , Progranulinas/genética , Proteína de Unión al GTP rab2/genéticaRESUMEN
CD26, acting as a costimulator of T cell activation, plays an important role in the immune system. However, the role of CD26 in the differentiation of T cell subsets, especially of new paradigms of T cells, such as Th17 and Tregs, is not fully clarified. In the present study, the role of CD26 in T cell differentiation was investigated in vitro. CD26 expression was analyzed in the different subsets of human peripheral blood T lymphocytes after solid-phase immobilized specific anti-CD3 mAb stimulation. Here, the percentage of CD4+ cells significantly increased and most of these cells were coexpressed with CD26, suggesting a close correlation of CD26 expression with the proliferation of CD4+ cells. Subsequently, after immobilized anti-CD3 mAb stimulation, CD26 high-expressing cells (CD26high) were separated from CD26 low-expressing cells (CD26low) by magnetic cell sorting. We found that the percentages of cells secreting Th1 typical cytokines (IL-2, IFN-γ) and Th17 typical cytokines (IL-6, IL-17, and IL-22) or expressing Th17 typical biomarkers (IL-23R, CD161, and CD196) in the CD26high group were markedly higher than in those in the CD26low group. In addition, a coexpression of CD26 with IL-2, IFN-γ, IL-17, IL-22, and IL-23R in lymphocytes was demonstrated by fluorescence microscopy. These results provide direct evidence that the high expression of CD26 is accompanied by the differentiation of T lymphocytes into Th1 and Th17, indicating that CD26 plays a crucial role in regulating the immune response.
Asunto(s)
Diferenciación Celular , Dipeptidil Peptidasa 4/metabolismo , Células TH1/citología , Células TH1/metabolismo , Células Th17/citología , Células Th17/metabolismo , Biomarcadores , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Citocinas/metabolismo , Expresión Génica , Humanos , Inmunofenotipificación , Activación de Linfocitos/genética , Activación de Linfocitos/inmunología , Recuento de Linfocitos , Subgrupos de Linfocitos T/citología , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Células TH1/inmunología , Células Th17/inmunologíaRESUMEN
OBJECTIVE: Tumour necrosis factor alpha (TNF-α) signalling plays a central role in the pathogenesis of various autoimmune diseases, particularly inflammatory arthritis. This study aimed to repurpose clinically approved drugs as potential inhibitors of TNF-α signalling in treatment of inflammatory arthritis. METHODS: In vitro and in vivo screening of an Food and Drug Administration (FDA)-approved drug library; in vitro and in vivo assays for examining the blockade of TNF actions by fexofenadine: assays for defining the anti-inflammatory activity of fexofenadine using TNF-α transgenic (TNF-tg) mice and collagen-induced arthritis in DBA/1 mice. Identification and characterisation of the binding of fexofenadine to cytosolic phospholipase A2 (cPLA2) using drug affinity responsive target stability assay, proteomics, cellular thermal shift assay, information field dynamics and molecular dynamics; various assays for examining fexofenadine inhibition of cPLA2 as well as the dependence of fexofenadine's anti-TNF activity on cPLA2. RESULTS: Serial screenings of a library composed of FDA-approved drugs led to the identification of fexofenadine as an inhibitor of TNF-α signalling. Fexofenadine potently inhibited TNF/nuclear factor kappa-light-chain-enhancer of activated B cells (NF-ĸB) signalling in vitro and in vivo, and ameliorated disease symptoms in inflammatory arthritis models. cPLA2 was isolated as a novel target of fexofenadine. Fexofenadine blocked TNF-stimulated cPLA2 activity and arachidonic acid production through binding to catalytic domain 2 of cPLA2 and inhibition of its phosphorylation on Ser-505. Further, deletion of cPLA2 abolished fexofenadine's anti-TNF activity. CONCLUSION: Collectively, these findings not only provide new insights into the understanding of fexofenadine action and underlying mechanisms but also provide new therapeutic interventions for various TNF-α and cPLA2-associated pathologies and conditions, particularly inflammatory rheumatic diseases.