RESUMO
Blood myeloid cells are known to be dysregulated in coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2. It is unknown whether the innate myeloid response differs with disease severity and whether markers of innate immunity discriminate high-risk patients. Thus, we performed high-dimensional flow cytometry and single-cell RNA sequencing of COVID-19 patient peripheral blood cells and detected disappearance of non-classical CD14LowCD16High monocytes, accumulation of HLA-DRLow classical monocytes (Human Leukocyte Antigen - DR isotype), and release of massive amounts of calprotectin (S100A8/S100A9) in severe cases. Immature CD10LowCD101-CXCR4+/- neutrophils with an immunosuppressive profile accumulated in the blood and lungs, suggesting emergency myelopoiesis. Finally, we show that calprotectin plasma level and a routine flow cytometry assay detecting decreased frequencies of non-classical monocytes could discriminate patients who develop a severe form of COVID-19, suggesting a predictive value that deserves prospective evaluation.
Assuntos
Infecções por Coronavirus , Coronavirus , Pandemias , Pneumonia Viral , Betacoronavirus , COVID-19 , Citometria de Fluxo , Humanos , Complexo Antígeno L1 Leucocitário , Monócitos , Células Mieloides , Estudos Prospectivos , SARS-CoV-2RESUMO
In response to microbial infection, the human host deploys metal-sequestering host-defense proteins, which reduce nutrient availability and thereby inhibit microbial growth and virulence. Calprotectin (CP) is an abundant antimicrobial protein released from neutrophils and epithelial cells at sites of infection. CP sequesters divalent first-row transition metal ions to limit the availability of essential metal nutrients in the extracellular space. While functional and clinical studies of CP have been pursued for decades, advances in our understanding of its biological coordination chemistry, which is central to its role in the host-microbe interaction, have been made in more recent years. In this review, we focus on the coordination chemistry of CP and highlight studies of its metal-binding properties and contributions to the metal-withholding innate immune response. Taken together, these recent studies inform our current model of how CP participates in metal homeostasis and immunity, and they provide a foundation for further investigations of a remarkable metal-chelating protein at the host-microbe interface and beyond.
Assuntos
Interações entre Hospedeiro e Microrganismos/imunologia , Interações entre Hospedeiro e Microrganismos/fisiologia , Complexo Antígeno L1 Leucocitário/imunologia , Complexo Antígeno L1 Leucocitário/metabolismo , Elementos de Transição/metabolismo , Sequência de Aminoácidos , Animais , Peptídeos Catiônicos Antimicrobianos/genética , Peptídeos Catiônicos Antimicrobianos/imunologia , Peptídeos Catiônicos Antimicrobianos/metabolismo , Humanos , Imunidade Inata , Ferro/imunologia , Ferro/metabolismo , Complexo Antígeno L1 Leucocitário/genética , Manganês/imunologia , Manganês/metabolismo , Modelos Biológicos , Modelos Moleculares , Níquel/imunologia , Níquel/metabolismo , Conformação Proteica , Homologia de Sequência de Aminoácidos , Zinco/imunologia , Zinco/metabolismoRESUMO
S100A1, a small homodimeric EF-hand Ca2+-binding protein (~21 kDa), plays an important regulatory role in Ca2+ signaling pathways involved in various biological functions including Ca2+ cycling and contractile performance in skeletal and cardiac myocytes. One key target of the S100A1 interactome is the ryanodine receptor (RyR), a huge homotetrameric Ca2+ release channel (~2.3 MDa) of the sarcoplasmic reticulum. Here, we report cryoelectron microscopy structures of S100A1 bound to RyR1, the skeletal muscle isoform, in absence and presence of Ca2+. Ca2+-free apo-S100A1 binds beneath the bridging solenoid (BSol) and forms contacts with the junctional solenoid and the shell-core linker of RyR1. Upon Ca2+-binding, S100A1 undergoes a conformational change resulting in the exposure of the hydrophobic pocket known to serve as a major interaction site of S100A1. Through interactions of the hydrophobic pocket with RyR1, Ca2+-bound S100A1 intrudes deeper into the RyR1 structure beneath BSol than the apo-form and induces sideways motions of the C-terminal BSol region toward the adjacent RyR1 protomer resulting in tighter interprotomer contacts. Interestingly, the second hydrophobic pocket of the S100A1-dimer is largely exposed at the hydrophilic surface making it prone to interactions with the local environment, suggesting that S100A1 could be involved in forming larger heterocomplexes of RyRs with other protein partners. Since S100A1 interactions stabilizing BSol are implicated in the regulation of RyR-mediated Ca2+ release, the characterization of the S100A1 binding site conserved between RyR isoforms may provide the structural basis for the development of therapeutic strategies regarding treatments of RyR-related disorders.
Assuntos
Cálcio , Microscopia Crioeletrônica , Canal de Liberação de Cálcio do Receptor de Rianodina , Proteínas S100 , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina/química , Proteínas S100/metabolismo , Proteínas S100/química , Cálcio/metabolismo , Animais , Ligação Proteica , Sítios de Ligação , Modelos Moleculares , Conformação Proteica , HumanosRESUMO
S100A11 is a small Ca2+-activatable protein known to localize along stress fibers (SFs). Analyzing S100A11 localization in HeLa and U2OS cells further revealed S100A11 enrichment at focal adhesions (FAs). Strikingly, S100A11 levels at FAs increased sharply, yet transiently, just before FA disassembly. Elevating intracellular Ca2+ levels with ionomycin stimulated both S100A11 recruitment and subsequent FA disassembly. However, pre-incubation with the non-muscle myosin II (NMII) inhibitor blebbistatin or with an inhibitor of the stretch-activatable Ca2+ channel Piezo1 suppressed S100A11 recruitment, implicating S100A11 in an actomyosin-driven FA recruitment mechanism involving Piezo1-dependent Ca2+ influx. Applying external forces on peripheral FAs likewise recruited S100A11 to FAs even if NMII activity was inhibited, corroborating the mechanosensitive recruitment mechanism of S100A11. However, extracellular Ca2+ and Piezo1 function were indispensable, indicating that NMII contraction forces act upstream of Piezo1-mediated Ca2+ influx, in turn leading to S100A11 activation and FA recruitment. S100A11-knockout cells display enlarged FAs and had delayed FA disassembly during cell membrane retraction, consistent with impaired FA turnover in these cells. Our results thus demonstrate a novel function for S100A11 in promoting actomyosin contractility-driven FA disassembly.
Assuntos
Actomiosina , Adesões Focais , Humanos , Adesões Focais/metabolismo , Actomiosina/metabolismo , Cálcio/metabolismo , Proteínas do Citoesqueleto/metabolismo , Miosina Tipo II/metabolismo , Proteínas S100/genética , Proteínas S100/metabolismoRESUMO
Metastatic cancer accounts for 90% of all cancer-related deaths and continues to be one of the toughest challenges in cancer treatment. A growing body of data indicates that S100A9, a major regulator of inflammation, plays a central role in cancer progression and metastasis, particularly in the lungs, where S100A9 forms a premetastatic niche. Thus, we developed a vaccine against S100A9 derived from plant viruses and virus-like particles. Using multiple tumor mouse models, we demonstrate the effectiveness of the S100A9 vaccine candidates in preventing tumor seeding within the lungs and outgrowth of metastatic disease. The elicited antibodies showed high specificity toward S100A9 without cross-reactivity toward S100A8, another member of the S100A family. When tested in metastatic mouse models of breast cancer and melanoma, the vaccines significantly reduced lung tumor nodules after intravenous challenge or postsurgical removal of the primary tumor. Mechanistically, the vaccines reduce the levels of S100A9 within the lungs and sera, thereby increasing the expression of immunostimulatory cytokines with antitumor function [(interleukin) IL-12 and interferonγ] while reducing levels of immunosuppressive cytokines (IL-10 and transforming growth factorß). This also correlated with decreased myeloid-derived suppressor cell populations within the lungs. This work has wide-ranging impact, as S100A9 is overexpressed in multiple cancers and linked with poor prognosis in cancer patients. The data presented lay the foundation for the development of therapies and vaccines targeting S100A9 to prevent metastasis.
Assuntos
Neoplasias Pulmonares , Vacinas Virais , Humanos , Camundongos , Animais , Calgranulina B/metabolismo , Neoplasias Pulmonares/prevenção & controle , Calgranulina A/metabolismo , Pulmão/patologia , Citocinas/metabolismoRESUMO
In various epithelial tissues, the epithelial monolayer acts as a barrier. To fulfill its function, the structural integrity of the epithelium is tightly controlled. When normal epithelial cells detach from the basal substratum and delaminate into the apical lumen, the apically extruded cells undergo apoptosis, which is termed anoikis. In contrast, transformed cells often become resistant to anoikis and able to survive and grow in the apical luminal space, leading to the formation of multilayered structures, which can be observed at the early stage of carcinogenesis. However, the underlying molecular mechanisms still remain elusive. In this study, we first demonstrate that S100A10 and ANXA2 (Annexin A2) accumulate in apically extruded, transformed cells in both various cell culture systems and murine epithelial tissues in vivo. ANXA2 acts upstream of S100A10 accumulation. Knockdown of ANXA2 promotes apoptosis of apically extruded RasV12-transformed cells and suppresses the formation of multilayered epithelia. In addition, the intracellular reactive oxygen species (ROS) are elevated in apically extruded RasV12 cells. Treatment with ROS scavenger Trolox reduces the occurrence of apoptosis of apically extruded ANXA2-knockdown RasV12 cells and restores the formation of multilayered epithelia. Furthermore, ROS-mediated p38MAPK activation is observed in apically delaminated RasV12 cells, and ANXA2 knockdown further enhances the p38MAPK activity. Moreover, the p38MAPK inhibitor promotes the formation of multilayered epithelia of ANXA2-knockdown RasV12 cells. These results indicate that accumulated ANXA2 diminishes the ROS-mediated p38MAPK activation in apically extruded transformed cells, thereby blocking the induction of apoptosis. Hence, ANXA2 can be a potential therapeutic target to prevent multilayered, precancerous lesions.
Assuntos
Anexina A2 , Animais , Camundongos , Anexina A2/genética , Apoptose , Células Epiteliais , Epitélio , Espécies Reativas de OxigênioRESUMO
Hidradenitis suppurativa (HS) is a complex inflammatory skin disease with undefined mechanistic underpinnings. Here, we investigated HS epithelial cells and demonstrated that HS basal progenitors modulate their lineage restriction and give rise to pathogenic keratinocyte clones, resulting in epidermal hyperproliferation and dysregulated inflammation in HS. When comparing to healthy epithelial stem/progenitor cells, in HS, we identified changes in gene signatures that revolve around the mitotic cell cycle, DNA damage response and repair, as well as cell-cell adhesion and chromatin remodeling. By reconstructing cell differentiation trajectory and CellChat modeling, we identified a keratinocyte population specific to HS. This population is marked by S100A7/8/9 and KRT6 family members, triggering IL1, IL10, and complement inflammatory cascades. These signals, along with HS-specific proinflammatory cytokines and chemokines, contribute to the recruitment of certain immune cells during the disease progression. Furthermore, we revealed a previously uncharacterized role of S100A8 in regulating the local chromatin environment of target loci in HS keratinocytes. Through the integration of genomic and epigenomic datasets, we identified genome-wide chromatin rewiring alongside the switch of transcription factors (TFs), which mediated HS transcriptional profiles. Importantly, we identified numerous clinically relevant inflammatory enhancers and their coordinated TFs in HS basal CD49fhigh cells. The disruption of the S100A enhancer using the CRISPR/Cas9-mediated approach or the pharmacological inhibition of the interferon regulatory transcription factor 3 (IRF3) efficiently reduced the production of HS-associated inflammatory regulators. Our study not only uncovers the plasticity of epidermal progenitor cells in HS but also elucidates the epigenetic mechanisms underlying HS pathogenesis.
Assuntos
Hidradenite Supurativa , Humanos , Hidradenite Supurativa/genética , Pele/metabolismo , Epigenômica , Epigênese Genética , Células-Tronco/metabolismo , Cromatina/metabolismoRESUMO
Fibroblast to myofibroblast transdifferentiation mediates numerous fibrotic disorders, such as idiopathic pulmonary fibrosis (IPF). We have previously demonstrated that non-muscle myosin II (NMII) is activated in response to fibrotic lung extracellular matrix, thereby mediating myofibroblast transdifferentiation. NMII-A is known to interact with the calcium-binding protein S100A4, but the mechanism by which S100A4 regulates fibrotic disorders is unclear. In this study, we show that fibroblast S100A4 is a calcium-dependent, mechanoeffector protein that is uniquely sensitive to pathophysiologic-range lung stiffness (8-25 kPa) and thereby mediates myofibroblast transdifferentiation. Re-expression of endogenous fibroblast S100A4 rescues the myofibroblastic phenotype in S100A4 KO fibroblasts. Analysis of NMII-A/actin dynamics reveals that S100A4 mediates the unraveling and redistribution of peripheral actomyosin to a central location, resulting in a contractile myofibroblast. Furthermore, S100A4 loss protects against murine in vivo pulmonary fibrosis, and S100A4 expression is dysregulated in IPF. Our data reveal a novel mechanosensor/effector role for endogenous fibroblast S100A4 in inducing cytoskeletal redistribution in fibrotic disorders such as IPF.
Assuntos
Fibrose Pulmonar Idiopática , Mecanotransdução Celular , Miofibroblastos , Proteína A4 de Ligação a Cálcio da Família S100 , Animais , Camundongos , Transdiferenciação Celular , Fibrose , Fibrose Pulmonar Idiopática/metabolismo , Fibrose Pulmonar Idiopática/patologia , Pulmão/metabolismo , Miofibroblastos/metabolismo , Miofibroblastos/patologia , Proteína A4 de Ligação a Cálcio da Família S100/genética , Proteína A4 de Ligação a Cálcio da Família S100/metabolismoRESUMO
Fungal keratitis is the foremost cause of corneal infections worldwide, of which Fusariumspp. is the common etiological agent that causes loss of vision and warrants surgical intervention. An increase in resistance to the available drugs along with severe side effects of the existing antifungals demands for new effective antimycotics. Here, we demonstrate that antimicrobial peptide S100A12 directly binds to the phospholipids of the fungal membrane, disrupts the structural integrity, and induces generation of reactive oxygen species in fungus. In addition, it inhibits biofilm formation by Fusariumspp. and exhibits antifungal property against Fusariumspp. both in vitro and in vivo. Taken together, our results delve into specific effect of S100A12 against Fusariumspp. with an aim to investigate new antifungal compounds to combat fungal keratitis.
Assuntos
Antifúngicos , Biofilmes , Membrana Celular , Fusarium , Proteína S100A12 , Antifúngicos/metabolismo , Antifúngicos/farmacologia , Biofilmes/efeitos dos fármacos , Infecções Oculares Fúngicas/microbiologia , Fusarium/efeitos dos fármacos , Ceratite/microbiologia , Proteína S100A12/metabolismo , Proteína S100A12/farmacologia , Humanos , Membrana Celular/efeitos dos fármacos , Fosfolipídeos/metabolismo , Espécies Reativas de Oxigênio/metabolismoRESUMO
Dendritic cells (DC) subsets, like Langerhans cells (LC), are immune cells involved in pathogen sensing. They express specific antimicrobial cellular factors that are able to restrict infection and limit further pathogen transmission. Here, we identify the alarmin S100A9 as a novel intracellular antiretroviral factor expressed in human monocyte-derived and skin-derived LC. The intracellular expression of S100A9 is decreased upon LC maturation and inversely correlates with enhanced susceptibility to HIV-1 infection of LC. Furthermore, silencing of S100A9 in primary human LC relieves HIV-1 restriction while ectopic expression of S100A9 in various cell lines promotes intrinsic resistance to both HIV-1 and MLV infection by acting on reverse transcription. Mechanistically, the intracellular expression of S100A9 alters viral capsid uncoating and reverse transcription. S100A9 also shows potent inhibitory effect against HIV-1 and MMLV reverse transcriptase (RTase) activity in vitro in a divalent cation-dependent manner. Our findings uncover an unexpected intracellular function of the human alarmin S100A9 in regulating antiretroviral immunity in Langerhans cells.
Assuntos
Alarminas/genética , Calgranulina B/genética , HIV-1/fisiologia , Células de Langerhans/virologia , Vírus da Leucemia Murina de Moloney/fisiologia , Infecções por Retroviridae/prevenção & controle , Animais , Linfócitos T CD4-Positivos/imunologia , Linhagem Celular , Cricetulus , HIV-1/genética , Interações Hospedeiro-Patógeno , Humanos , Células de Langerhans/imunologia , Leucemia Experimental/prevenção & controle , Camundongos , Vírus da Leucemia Murina de Moloney/genética , Transcrição Reversa , Fator de Crescimento Transformador beta/imunologia , Infecções Tumorais por Vírus/prevenção & controle , Replicação ViralRESUMO
Athletes increasingly engage in repeated sprint training consisting in repeated short all-out efforts interspersed by short recoveries. When performed in hypoxia (RSH), it may lead to greater training effects than in normoxia (RSN); however, the underlying molecular mechanisms remain unclear. This study aimed at elucidating the effects of RSH on skeletal muscle metabolic adaptations as compared to RSN. Sixteen healthy young men performed nine repeated sprint training sessions in either normoxia (FIO2 = 0.209, RSN, n = 7) or normobaric hypoxia (FIO2 = 0.136, RSH, n = 9). Before and after the training period, exercise performance was assessed by using repeated sprint ability (RSA) and Wingate tests. Vastus lateralis muscle biopsies were performed to investigate muscle metabolic adaptations using proteomics combined with western blot analysis. Similar improvements were observed in RSA and Wingate tests in both RSN and RSH groups. At the muscle level, RSN and RSH reduced oxidative phosphorylation protein content but triggered an increase in mitochondrial biogenesis proteins. Proteomics showed an increase in several S100A family proteins in the RSH group, among which S100A13 most strongly. We confirmed a significant increase in S100A13 protein by western blot in RSH, which was associated with increased Akt phosphorylation and its downstream targets regulating protein synthesis. Altogether our data indicate that RSH may activate an S100A/Akt pathway to trigger specific adaptations as compared to RSN.
Assuntos
Adaptação Fisiológica , Hipóxia , Músculo Esquelético , Proteínas S100 , Transdução de Sinais , Humanos , Masculino , Hipóxia/metabolismo , Músculo Esquelético/metabolismo , Adaptação Fisiológica/fisiologia , Transdução de Sinais/fisiologia , Adulto Jovem , Proteínas S100/metabolismo , Adulto , Proteínas Proto-Oncogênicas c-akt/metabolismo , Exercício Físico/fisiologiaRESUMO
Down syndrome (DS) is a genetic disease characterized by a supernumerary chromosome 21. Intellectual deficiency (ID) is one of the most prominent features of DS. Central nervous system defects lead to learning disabilities, motor and language delays, and memory impairments. At present, a prenatal treatment for the ID in DS is lacking. Subcutaneous administration of synthetic preimplantation factor (sPIF, a peptide with a range of biological functions) in a model of severe brain damage has shown neuroprotective and anti-inflammatory properties by directly targeting neurons and microglia. Here, we evaluated the effect of PIF administration during gestation and until weaning on Dp(16)1Yey mice (a mouse model of DS). Possible effects at the juvenile stage were assessed using behavioral tests and molecular and histological analyses of the brain. To test the influence of perinatal sPIF treatment at the adult stage, hippocampus-dependent memory was evaluated on postnatal day 90. Dp(16)1Yey pups showed significant behavioral impairment, with impaired neurogenesis, microglial cell activation and a low microglial cell count, and the deregulated expression of genes linked to neuroinflammation and cell cycle regulation. Treatment with sPIF restored early postnatal hippocampal neurogenesis, with beneficial effects on astrocytes, microglia, inflammation, and cell cycle markers. Moreover, treatment with sPIF restored the level of DYRK1A, a protein that is involved in cognitive impairments in DS. In line with the beneficial effects on neurogenesis, perinatal treatment with sPIF was associated with an improvement in working memory in adult Dp(16)1Yey mice. Perinatal treatment with sPIF might be an option for mitigating cognitive impairments in people with DS.
Assuntos
Modelos Animais de Doenças , Síndrome de Down , Neurogênese , Animais , Síndrome de Down/tratamento farmacológico , Síndrome de Down/patologia , Síndrome de Down/metabolismo , Síndrome de Down/complicações , Síndrome de Down/genética , Neurogênese/efeitos dos fármacos , Camundongos , Feminino , Gravidez , Hipocampo/metabolismo , Hipocampo/patologia , Hipocampo/efeitos dos fármacos , Microglia/metabolismo , Microglia/efeitos dos fármacos , Microglia/patologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Proteínas Tirosina Quinases/genética , Quinases Dyrk , Disfunção Cognitiva/tratamento farmacológico , Disfunção Cognitiva/metabolismo , Disfunção Cognitiva/patologia , Masculino , Transtornos Cognitivos/tratamento farmacológico , Transtornos Cognitivos/patologiaRESUMO
BACKGROUND AND AIMS: Hepatitis B virus (HBV)-associated liver cirrhosis (LC), a common condition with high incidence and mortality rates, is often associated with diabetes mellitus (DM). However, the molecular mechanisms underlying impaired glucose regulation during HBV-associated LC remain unclear. METHODS: Data from 63 patients with LC and 62 patients with LC-associated DM were analysed. Co-culture of NK cells and islet ß cell lines were used to study the glucose regulation mechanism. A mouse model of LC was used to verify the effect of S100A8/A9 on the glucose regulation. RESULTS: Higher levels of interferon (IFN)-γ derived from natural killer (NK) cells and lower levels of insulin emerged in the peripheral blood of patients with both LC and DM compared with those from patients with LC only. IFN-γ derived from NK cells facilitated ß cell necroptosis and impaired insulin production. Furthermore, S100A8/A9 elevation in patients with both LC and DM was found to upregulate IFN-γ production in NK cells. Consistently, in the mouse model for LC, mice treated with carbon tetrachloride (CCL4) and S100A8/A9 exhibited increased blood glucose, impaired insulin production, increased IFN-γ, and increased ß cells necroptosis compared with those treated with CCL4. Mechanistically, S100A8/A9 activated the p38 MAPK pathway to increase IFN-γ production in NK cells. These effects were diminished after blocking RAGE. CONCLUSION: Together, the data indicate that IFN-γ produced by NK cells induces ß cell necroptosis via the S100A8/A9-RAGE-p38 MAPK axis in patients with LC and DM. Reduced levels of S100A8/A9, NK cells, and IFN-γ could be valuable for the treatment of LC with DM. Accumulation of S100A8/A9 in patients with LC may indicate the emergence of DM.
Assuntos
Calgranulina A , Calgranulina B , Vírus da Hepatite B , Células Secretoras de Insulina , Interferon gama , Células Matadoras Naturais , Cirrose Hepática , Necroptose , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Humanos , Animais , Interferon gama/metabolismo , Calgranulina B/metabolismo , Cirrose Hepática/patologia , Cirrose Hepática/metabolismo , Cirrose Hepática/virologia , Cirrose Hepática/imunologia , Camundongos , Masculino , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Células Secretoras de Insulina/virologia , Calgranulina A/metabolismo , Camundongos Endogâmicos C57BL , Feminino , Pessoa de Meia-Idade , Hepatite B/complicações , Hepatite B/patologia , Hepatite B/metabolismo , Modelos Animais de Doenças , Tetracloreto de CarbonoRESUMO
Ubiquitin-proteasome system dysfunction triggers α-synuclein aggregation, a hallmark of neurodegenerative diseases, such as Parkinson's disease (PD). However, the crosstalk between deubiquitinating enzyme (DUBs) and α-synuclein pathology remains unclear. In this study, we observed a decrease in the level of ubiquitin-specific protease 14 (USP14), a DUB, in the cerebrospinal fluid (CSF) of PD patients, particularly females. Moreover, CSF USP14 exhibited a dual correlation with α-synuclein in male and female PD patients. To investigate the impact of USP14 deficiency, we crossed USP14 heterozygous mouse (USP14+/-) with transgenic A53T PD mouse (A53T-Tg) or injected adeno-associated virus (AAV) carrying human α-synuclein (AAV-hα-Syn) in USP14+/- mice. We found that Usp14 deficiency improved the behavioral abnormities and pathological α-synuclein deposition in female A53T-Tg or AAV-hα-Syn mice. Additionally, Usp14 inactivation attenuates the pro-inflammatory response in female AAV-hα-Syn mice, whereas Usp14 inactivation demonstrated opposite effects in male AAV-hα-Syn mice. Mechanistically, the heterodimeric protein S100A8/A9 may be the downstream target of Usp14 deficiency in female mouse models of α-synucleinopathies. Furthermore, upregulated S100A8/A9 was responsible for α-synuclein degradation by autophagy and the suppression of the pro-inflammatory response in microglia after Usp14 knockdown. Consequently, our study suggests that USP14 could serve as a novel therapeutic target in PD.
Assuntos
Calgranulina A , Calgranulina B , Camundongos Transgênicos , Doença de Parkinson , Ubiquitina Tiolesterase , alfa-Sinucleína , alfa-Sinucleína/metabolismo , alfa-Sinucleína/genética , Animais , Doença de Parkinson/metabolismo , Doença de Parkinson/genética , Doença de Parkinson/patologia , Ubiquitina Tiolesterase/metabolismo , Ubiquitina Tiolesterase/genética , Ubiquitina Tiolesterase/deficiência , Humanos , Camundongos , Feminino , Masculino , Calgranulina B/metabolismo , Calgranulina B/genética , Calgranulina A/metabolismo , Calgranulina A/genética , Modelos Animais de Doenças , Camundongos Endogâmicos C57BLRESUMO
Toll-like receptors (TLRs), especially TLR7, play an important role in systemic lupus erythematosus (SLE) pathogenesis. However, the regulatory mechanism underlying the abnormal activation of TLR pathways in patients with SLE has not been elucidated. Notably, accumulating evidence indicates that myeloid-derived suppressor cells (MDSCs) are important regulators of inflammation and autoimmune diseases. Compared with healthy control subjects, patients with SLE have a greater proportion of MDSCs among peripheral blood mononuclear cells (PBMCs); however, the effect of MDSCs on TLR7 pathway activation has not been determined. In the present study, lupus MDSCs significantly promoted TLR7 pathway activation in macrophages and dendritic cells (DCs), exacerbating the imiquimod-induced lupus model. RNA-sequencing analysis revealed significant overexpression of S100 calcium-binding protein A8 (S100A8) and S100A9 in MDSCs from diseased MRL/lpr mice. In vitro and in vivo studies demonstrated that S100A8/9 effectively promoted TLR7 pathway activation and that S100A8/9 deficiency reversed the promoting effect of MDSCs on TLR7 pathway activation in lupus. Mechanistically, MDSC-derived S100A8/9 upregulated interferon gamma (IFN-γ) secretion by macrophages and IFN-γ subsequently promoted TLR7 pathway activation in an autocrine manner. Taken together, these findings suggest that lupus MDSCs promote TLR7 pathway activation and lupus pathogenesis through the S100A8/9-IFN-γ axis. Our study identified an important target for SLE therapy.
Assuntos
Calgranulina A , Calgranulina B , Lúpus Eritematoso Sistêmico , Células Supressoras Mieloides , Animais , Camundongos , Células Dendríticas/metabolismo , Leucócitos Mononucleares/metabolismo , Lúpus Eritematoso Sistêmico/metabolismo , Lúpus Eritematoso Sistêmico/patologia , Macrófagos/metabolismo , Camundongos Endogâmicos MRL lpr , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/metabolismo , Calgranulina A/metabolismo , Calgranulina B/metabolismoRESUMO
Basal cell carcinomas (BCCs) and cutaneous squamous cell carcinomas (SCCs) are the most frequent types of cancer, and both originate from the keratinocyte transformation, giving rise to the group of tumors called keratinocyte carcinomas (KCs). The invasive behavior is different in each group of KC and may be influenced by their tumor microenvironment. The principal aim of the study is to characterize the protein profile of the tumor interstitial fluid (TIF) of KC to evaluate changes in the microenvironment that could be associated with their different invasive and metastatic capabilities. We obtained TIF from 27 skin biopsies and conducted a label-free quantitative proteomic analysis comparing seven BCCs, 16 SCCs, and four normal skins. A total of 2945 proteins were identified, 511 of them quantified in more than half of the samples of each tumoral type. The proteomic analysis revealed differentially expressed TIF proteins that could explain the different metastatic behavior in both KCs. In detail, the SCC samples disclosed an enrichment of proteins related to cytoskeleton, such as Stratafin and Ladinin-1. Previous studies found their upregulation positively correlated with tumor progression. Furthermore, the TIF of SCC samples was enriched with the cytokines S100A8/S100A9. These cytokines influence the metastatic output in other tumors through the activation of NF-kB signaling. According to this, we observed a significant increase in nuclear NF-kB subunit p65 in SCCs but not in BCCs. In addition, the TIF of both tumors was enriched with proteins involved in the immune response, highlighting the relevance of this process in the composition of the tumor environment. Thus, the comparison of the TIF composition of both KCs provides the discovery of a new set of differential biomarkers. Among them, secreted cytokines such as S100A9 may help explain the higher aggressiveness of SCCs, while Cornulin is a specific biomarker for BCCs. Finally, the proteomic landscape of TIF provides key information on tumor growth and metastasis, which can contribute to the identification of clinically applicable biomarkers that may be used in the diagnosis of KC, as well as therapeutic targets.
Assuntos
Carcinoma Basocelular , Carcinoma de Células Escamosas , Neoplasias Cutâneas , Humanos , Neoplasias Cutâneas/metabolismo , Líquido Extracelular/metabolismo , NF-kappa B/metabolismo , Proteômica , Carcinoma Basocelular/metabolismo , Carcinoma Basocelular/patologia , Carcinoma de Células Escamosas/metabolismo , Queratinócitos/metabolismo , Biomarcadores Tumorais/metabolismo , Microambiente TumoralRESUMO
BACKGROUND: Microglia is the primary source of inflammatory factors during migraine attacks. This study aims to investigate the role of microglia related genes (MRGs) in migraine attacks. METHODS: The RNA sequencing results of migraineurs and the panglaodb database were used to obtain differentially expressed genes (DEGs) in migraine related to microglia. A migraine rat model was established for validating and localizing of the MRGs, and subsequent screening for target genes was conducted. A shRNA was designed to interference the expression of target genes and administered into the trigeminal ganglion (TG) of rats. Pain sensitivity in rats was evaluated via the hot water tail-flick (HWTF) and formalin-induced pain (FIP) experiments. ELISA was used to quantify the levels of inflammatory cytokines and CGRP. WB and immunofluorescence assays were applied to detect the activation of microglia. RESULTS: A total of five DEGs in migraine related to microglia were obtained from RNA sequencing and panglaodb database. Animal experiments showed that these genes expression were heightened in the TG and medulla oblongata (MO) of migraine rats. The gene S100A8 co-localized with microglia in both TG and MO. The HWTF and FIP experiments demonstrated that interference with S100A8 alleviated the sense of pain in migraine rats. Moreover, the levels of TNFα, IL-1ß, IL-6, and CGRP in the TG and MO of rats in the model rats were increased, and the expression of microglia markers IBA-1, M1 polarization markers CD86 and iNOS was upregulated. Significantly, interference with S100A8 reversed these indicators. CONCLUSION: Interference with S100A8 in microglia increased the pain threshold during migraine attacks, and inhibited neuroinflammation and microglia activation.
RESUMO
An emerging group of spindle cell neoplasms harboring fusions involving NTRK or non-NTRK kinase genes often share characteristic S100 and/or CD34 expression; however, the diagnostic utility of immunohistochemical stains is not well established in this family owing to their lack of specificity. Recently, CD30 expression in spindle cell neoplasms with kinase gene fusions, such as NTRK, BRAF, RAF1, and RET, has been increasingly identified. We herein report a 10-year-old girl with high-grade spindle cell sarcoma of the neck. Prior to histopathological evaluation, flow cytometry (FCM) analysis and touch smear cytology of the tumor tissue revealed CD34+ and dimCD30+ spindle cell populations. Histopathologically, the case was characterized by monomorphic spindle-shaped cytomorphology with CD30, S100, and CD34 positivity and harbored close similarities with spindle cell neoplasms with NTRK or non-NTRK gene fusions. Subsequently, a comprehensive next-generation sequencing sarcoma panel identified a rare PLEKHH2::ALK fusion, and a diagnosis of ALK-rearranged spindle cell neoplasm was made. The patient showed significant tumor response to single-agent treatment with alectinib, an ALK-tyrosine kinase inhibitor. This case supports that CD30 is expressed in an ALK-rearranged mesenchymal neoplasm. The benefit of the early detection of CD30 expression by FCM for a prompt diagnosis and treatment is highlighted in the context of an aggressive clinical course. This case represents a learning experience regarding the need to the check the status of CD30 expression in these tumors and suggests the potential clinical benefits of CD30-targeted therapy.
Assuntos
Sarcoma , Neoplasias de Tecidos Moles , Feminino , Humanos , Criança , Imuno-Histoquímica , Citometria de Fluxo , Sarcoma/patologia , Neoplasias de Tecidos Moles/genética , Neoplasias de Tecidos Moles/patologia , Fusão Gênica , Receptores Proteína Tirosina Quinases/genética , Biomarcadores Tumorais/genéticaRESUMO
Parkinson's disease (PD) is a complex neurodegenerative disease with motor and non-motor symptoms. Diagnosis is complicated by lack of reliable biomarkers. To individuate peptides and/or proteins with diagnostic potential for early diagnosis, severity and discrimination from similar pathologies, the salivary proteome in 36 PD patients was investigated in comparison with 36 healthy controls (HC) and 35 Alzheimer's disease (AD) patients. A top-down platform based on HPLC-ESI-IT-MS allowed characterizing and quantifying intact peptides, small proteins and their PTMs (overall 51). The three groups showed significantly different protein profiles, PD showed the highest levels of cystatin SA and antileukoproteinase and the lowest of cystatin SN and some statherin proteoforms. HC exhibited the lowest abundance of thymosin ß4, short S100A9, cystatin A, and dimeric cystatin B. AD patients showed the highest abundance of α-defensins and short oxidized S100A9. Moreover, different proteoforms of the same protein, as S-cysteinylated and S-glutathionylated cystatin B, showed opposite trends in the two pathological groups. Statherin, cystatins SA and SN classified accurately PD from HC and AD subjects. α-defensins, histatin 1, oxidized S100A9, and P-B fragments were the best classifying factors between PD and AD patients. Interestingly statherin and thymosin ß4 correlated with defective olfactory functions in PD patients. All these outcomes highlighted implications of specific proteoforms involved in the innate-immune response and inflammation regulation at oral and systemic level, suggesting a possible panel of molecular and clinical markers suitable to recognize subjects affected by PD.
Assuntos
Doença de Alzheimer , Doenças Neurodegenerativas , Doença de Parkinson , alfa-Defensinas , Humanos , Doença de Alzheimer/diagnóstico , Doença de Alzheimer/metabolismo , Cistatina B/análise , Cistatina B/metabolismo , Proteômica/métodos , Doença de Parkinson/diagnóstico , Doença de Parkinson/metabolismo , Doenças Neurodegenerativas/metabolismo , alfa-Defensinas/análise , alfa-Defensinas/metabolismo , Saliva/química , Proteínas e Peptídeos Salivares/metabolismo , Fatores de Transcrição/metabolismo , Biomarcadores/análiseRESUMO
Solute carrier organic anion transporter family member 2A1 (SLCO2A1) is a prostaglandin (PG) transporter and serves as the osmosensitive ATP-permeable maxi-anion channel (Maxi-Cl). Since a heterotetrameric complex of annexin A2 (ANXA2) and S100A10 is obligatory for the channel activity, the present study aimed to determine if they regulate SLCO2A1-mediated PG transport. This study examined PGE2 uptake and ATP release in Anxa2 and/or S100a10 knockout (KO) murine breast C127 cells. Deletion of Slco2a1 decreased PGE2-d4 uptake by wild-type (WT) cells in an isotonic medium (290 mosmol/kgH2O). Decreased osmolarity (135 mosmol/kgH2O) stimulated ATP release but did not affect PGE2 uptake kinetics, showing Km (1,280 nM) and Vmax (10.38 pmol/15 s/mg protein) similar to those in isotonic medium (1,227 nM and 10.65 pmol/15 s/mg protein), respectively, in WT cells. Deletion of Anxa2 associated with loss of S100a10 diminished SLCO2A1-mediated ATP release and uncompetitively inhibited PGE2 uptake with lowered Km (376 nM) and Vmax (2.59 pmol/15 s/mg protein). Moreover, the immunoprecipitation assay confirmed the physical interaction of ANXA2 with SLCO2A1 in WT cells. Enforcement of ANXA2 expression to Anxa2 KO cells partially restored PGE2 uptake and increased Km (744.3 nM) and Vmax (9.07 pmol/15 s/mg protein), whereas the uptake clearance (Vmax/Km) did not change much regardless of ANXA2 expression. These results suggest that an ANXA2/S100A10 complex modulates PG transport activity but osmolality has little effect on it; therefore, the bound form of SLCO2A1, which functions as a PG transporter and Maxi-Cl, may exist regardless of changes in the cell volume.NEW & NOTEWORTHY A previous study indicated that the ANXA2/S100A10 complex represents the regulatory component of SLCO2A1-mediated Maxi-Cl channel activity. The present study showed that apparent PGE2 uptake by C127 cells was osmoinsensitive and uncompetitively inhibited by loss of ANXA2 expression, demonstrating that ANXA2 is a regulatory factor of SLCO2A1-mediated PG transport activity.