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1.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 38(9): 880-883, 2021 Sep 10.
Artigo em Chinês | MEDLINE | ID: mdl-34487536

RESUMO

OBJECTIVE: To explore the genetic etiology of a fetus with autosomal recessive polycystic kidney disease (ARPKD). METHODS: Prenatal ultrasonography has revealed oligohydramnios and abnormal structure of fetal kidneys. After careful counseling, the couple opted induced abortion. With informed consent, genomic DNA was extracted from the muscle sample of the abortus and peripheral blood samples of the couple. High throughput whole exome sequencing was carried out to detect potential variants in relation with the disease. Suspected variants were verified by Sanger sequencing. RESULTS: Prenatal ultrasound revealed increased size of fetal kidneys, with multiple hyperechos from the right kidney, and multiple hyperechos with anechoic masses within the left kidney. DNA sequencing revealed that the fetus has carried heterozygous variants of the PKHD1 gene, including c.7994T>C inherited from its father, and two heterozygous variants of the PKHD1 gene c.5681G>A from its mother. CONCLUSION: The compound heterozygous c.7994T>C and c.5681G>A variants of the PKHD1 gene probably underlay the pathogenesis of ARPKD in this fetus. Above results can provide guidance for subsequent pregnancies of the couple.


Assuntos
Rim Policístico Autossômico Recessivo , Feminino , Feto , Testes Genéticos , Humanos , Mutação , Rim Policístico Autossômico Recessivo/diagnóstico por imagem , Rim Policístico Autossômico Recessivo/genética , Gravidez , Receptores de Superfície Celular/genética
2.
Int J Mol Sci ; 22(17)2021 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-34502134

RESUMO

The current spreading coronavirus SARS-CoV-2 is highly infectious and pathogenic. In this study, we screened the gene expression of three host receptors (ACE2, DC-SIGN and L-SIGN) of SARS coronaviruses and dendritic cells (DCs) status in bulk and single cell transcriptomic datasets of upper airway, lung or blood of COVID-19 patients and healthy controls. In COVID-19 patients, DC-SIGN gene expression was interestingly decreased in lung DCs but increased in blood DCs. Within DCs, conventional DCs (cDCs) were depleted while plasmacytoid DCs (pDCs) were augmented in the lungs of mild COVID-19. In severe cases, we identified augmented types of immature DCs (CD22+ or ANXA1+ DCs) with MHCII downregulation. In this study, our observation indicates that DCs in severe cases stimulate innate immune responses but fail to specifically present SARS-CoV-2. It provides insights into the profound modulation of DC function in severe COVID-19.


Assuntos
COVID-19/imunologia , Moléculas de Adesão Celular/genética , Células Dendríticas/imunologia , Regulação da Expressão Gênica/imunologia , Lectinas Tipo C/genética , Receptores de Superfície Celular/genética , SARS-CoV-2/imunologia , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/metabolismo , COVID-19/diagnóstico , COVID-19/patologia , COVID-19/virologia , Moléculas de Adesão Celular/metabolismo , Conjuntos de Dados como Assunto , Células Dendríticas/metabolismo , Estudo de Associação Genômica Ampla , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata , Lectinas Tipo C/metabolismo , Pulmão/imunologia , Pulmão/patologia , Pulmão/virologia , Análise da Randomização Mendeliana , Nasofaringe/imunologia , Nasofaringe/patologia , Nasofaringe/virologia , RNA-Seq , Receptores de Superfície Celular/metabolismo , Índice de Gravidade de Doença , Análise de Célula Única
3.
Int J Mol Sci ; 22(16)2021 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-34445398

RESUMO

Gibberellins (GAs) are an important group of phytohormones associated with diverse growth and developmental processes, including cell elongation, seed germination, and secondary growth. Recent genomic and genetic analyses have advanced our knowledge of GA signaling pathways and related genes in model plant species. However, functional genomics analyses of GA signaling pathways in Panax ginseng, a perennial herb, have rarely been carried out, despite its well-known economical and medicinal importance. Here, we conducted functional characterization of GA receptors and investigated their physiological roles in the secondary growth of P. ginseng storage roots. We found that the physiological and genetic functions of P. ginseng gibberellin-insensitive dwarf1s (PgGID1s) have been evolutionarily conserved. Additionally, the essential domains and residues in the primary protein structure for interaction with active GAs and DELLA proteins are well-conserved. Overexpression of PgGID1s in Arabidopsis completely restored the GA deficient phenotype of the Arabidopsis gid1a gid1c (atgid1a/c) double mutant. Exogenous GA treatment greatly enhanced the secondary growth of tap roots; however, paclobutrazol (PCZ), a GA biosynthetic inhibitor, reduced root growth in P. ginseng. Transcriptome profiling of P. ginseng roots revealed that GA-induced root secondary growth is closely associated with cell wall biogenesis, the cell cycle, the jasmonic acid (JA) response, and nitrate assimilation, suggesting that a transcriptional network regulate root secondary growth in P. ginseng. These results provide novel insights into the mechanism controlling secondary root growth in P. ginseng.


Assuntos
Perfilação da Expressão Gênica/métodos , Giberelinas/farmacologia , Panax/crescimento & desenvolvimento , Receptores de Superfície Celular/genética , Evolução Molecular , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Mutação com Perda de Função , Panax/genética , Proteínas de Plantas/genética , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Domínios Proteicos , Receptores de Superfície Celular/química , Análise de Sequência de RNA , Transdução de Sinais/efeitos dos fármacos , Triazóis/farmacologia
4.
Int J Mol Sci ; 22(15)2021 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-34361063

RESUMO

BACKGROUND: Induced tooth movement during orthodontic therapy requires mechano-induced bone remodeling. Besides various cytokines and growth-factors, neuronal guidance molecules gained attention for their roles in bone homeostasis and thus, potential roles during tooth movement. Several neuronal guidance molecules have been implicated in the regulation of bone remodeling. Amongst them, Semaphorin 3A is particular interesting as it concurrently induces osteoblast differentiation and disturbs osteoclast differentiation. METHODS: Mechano-regulation of Sema3A and its receptors PlexinA1 and Neuropilin (RT-qPCR, WB) was evaluated by applying compressive and tension forces to primary human periodontal fibroblasts (hPDLF) and alveolar bone osteoblasts (hOB). The association of the transcription factor Osterix (SP7) and SEMA3A was studied by RT-qPCR. Mechanisms involved in SEMA3A-mediated osteoblast differentiation were assessed by Rac1GTPase pull-downs, ß-catenin expression analyses (RT-qPCR) and nuclear translocation assays (IF). Osteogenic markers were analyzed by RT-qPCR. RESULTS: SEMA3A, PLXNA1 and NRP1 were differentially regulated by tension or compressive forces in hPDLF. Osterix (SP7) displayed the same pattern of regulation. Recombinant Sema3A induced the activation of Rac1GTPase, the nuclear translocation of ß-catenin and the expression of osteogenic marker genes. CONCLUSION: Sema3A, its receptors and Osterix are regulated by mechanical forces in hPDLF. SEMA3A upregulation was associated with Osterix (SP7) modulation. Sema3A-enhanced osteogenic marker gene expression in hOB might be dependent on a pathway involving Rac1GTPase and ß-catenin. Thus, Semaphorin 3A might contribute to bone remodeling during induced tooth movement.


Assuntos
Fibroblastos/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Neuropilinas/metabolismo , Osteoblastos/fisiologia , Ligamento Periodontal/fisiologia , Receptores de Superfície Celular/metabolismo , Semaforina-3A/metabolismo , Técnicas de Movimentação Dentária/métodos , Adolescente , Adulto , Remodelação Óssea , Diferenciação Celular , Células Cultivadas , Criança , Fibroblastos/citologia , Humanos , Proteínas do Tecido Nervoso/genética , Neuropilinas/genética , Osteoblastos/citologia , Osteogênese , Ligamento Periodontal/citologia , Receptores de Superfície Celular/genética , Semaforina-3A/genética , Adulto Jovem
5.
Nat Commun ; 12(1): 4871, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34381056

RESUMO

The heteromeric complex between PKD1L3, a member of the polycystic kidney disease (PKD) protein family, and PKD2L1, also known as TRPP2 or TRPP3, has been a prototype for mechanistic characterization of heterotetrametric TRP-like channels. Here we show that a truncated PKD1L3/PKD2L1 complex with the C-terminal TRP-fold fragment of PKD1L3 retains both Ca2+ and acid-induced channel activities. Cryo-EM structures of this core heterocomplex with or without supplemented Ca2+ were determined at resolutions of 3.1 Å and 3.4 Å, respectively. The heterotetramer, with a pseudo-symmetric TRP architecture of 1:3 stoichiometry, has an asymmetric selectivity filter (SF) guarded by Lys2069 from PKD1L3 and Asp523 from the three PKD2L1 subunits. Ca2+-entrance to the SF vestibule is accompanied by a swing motion of Lys2069 on PKD1L3. The S6 of PKD1L3 is pushed inward by the S4-S5 linker of the nearby PKD2L1 (PKD2L1-III), resulting in an elongated intracellular gate which seals the pore domain. Comparison of the apo and Ca2+-loaded complexes unveils an unprecedented Ca2+ binding site in the extracellular cleft of the voltage-sensing domain (VSD) of PKD2L1-III, but not the other three VSDs. Structure-guided mutagenic studies support this unconventional site to be responsible for Ca2+-induced channel activation through an allosteric mechanism.


Assuntos
Canais de Cálcio/química , Cálcio/metabolismo , Receptores de Superfície Celular/química , Canais de Cátion TRPP/química , Aminoácidos , Animais , Sítios de Ligação , Cálcio/química , Canais de Cálcio/genética , Canais de Cálcio/metabolismo , Microscopia Crioeletrônica , Ativação do Canal Iônico , Camundongos , Mutagênese , Conformação Proteica , Domínios Proteicos , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Canais de Cátion TRPP/genética , Canais de Cátion TRPP/metabolismo
6.
Int J Mol Sci ; 22(12)2021 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-34204582

RESUMO

Autosomal recessive polycystic kidney disease (ARPKD) is a rare disorder and one of the most severe forms of polycystic kidney disease, leading to end-stage renal disease (ESRD) in childhood. PKHD1 is the gene that is responsible for the vast majority of ARPKD. However, some cases have been related to a new gene that was recently identified (DZIP1L gene), as well as several ciliary genes that can mimic a ARPKD-like phenotypic spectrum. In addition, a number of molecular pathways involved in the ARPKD pathogenesis and progression were elucidated using cellular and animal models. However, the function of the ARPKD proteins and the molecular mechanism of the disease currently remain incompletely understood. Here, we review the clinics, treatment, genetics, and molecular basis of ARPKD, highlighting the most recent findings in the field.


Assuntos
Suscetibilidade a Doenças , Rim Policístico Autossômico Recessivo/etiologia , Rim Policístico Autossômico Recessivo/metabolismo , Animais , Biomarcadores , Ensaios Clínicos como Assunto , Terapia Combinada , Diagnóstico Diferencial , Gerenciamento Clínico , Estudos de Associação Genética/métodos , Predisposição Genética para Doença , Humanos , Mutação , Fenótipo , Rim Policístico Autossômico Recessivo/patologia , Locos de Características Quantitativas , Receptores de Superfície Celular/química , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Relação Estrutura-Atividade , Resultado do Tratamento
7.
J Immunol ; 207(3): 837-848, 2021 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-34282004

RESUMO

Dendritic cells (DCs) are critical for pathogen recognition and Ag processing/presentation. Human monocyte-derived DCs (moDCs) have been extensively used in experimental studies and DC-based immunotherapy approaches. However, the extent of human moDC and peripheral DCs heterogeneity and their interrelationship remain elusive. In this study, we performed single-cell RNA sequencing of human moDCs and blood DCs. We identified seven subtypes within moDCs: five corresponded to type 2 conventional DCs (cDC2s), and the other two were CLEC10A+CD127+ cells with no resemblance to any peripheral DC subpopulations characterized to date. Moreover, we defined five similar subtypes in human cDC2s, revealed the potential differentiation trajectory among them, and unveiled the transcriptomic differences between moDCs and cDC2s. We further studied the transcriptomic changes of each moDC subtype during maturation, demonstrating SLAMF7 and IL15RA as maturation markers and CLEC10A and SIGLEC10 as markers for immature DCs. These findings will enable more accurate functional/developmental analyses of human cDC2s and moDCs.


Assuntos
Células Dendríticas/fisiologia , Monócitos/fisiologia , Análise de Célula Única/métodos , Adulto , Diferenciação Celular/genética , Células Cultivadas , Citocinas/metabolismo , Feminino , Perfilação da Expressão Gênica , Humanos , Lectinas/genética , Lectinas Tipo C/genética , Masculino , Receptores de Superfície Celular/genética , Receptores de Interleucina-15/genética , Análise de Sequência de RNA , Família de Moléculas de Sinalização da Ativação Linfocitária/genética , Células Th2/imunologia , Adulto Jovem
8.
Nat Microbiol ; 6(8): 991-999, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34294905

RESUMO

More than one-third of the world's population is exposed to Plasmodium vivax malaria, mainly in Asia1. P. vivax preferentially invades reticulocytes (immature red blood cells)2-4. Previous work has identified 11 parasite proteins involved in reticulocyte invasion, including erythrocyte binding protein 2 (ref. 5) and the reticulocyte-binding proteins (PvRBPs)6-10. PvRBP2b binds to the transferrin receptor CD71 (ref. 11), which is selectively expressed on immature reticulocytes12. Here, we identified CD98 heavy chain (CD98), a heteromeric amino acid transporter from the SLC3 family (also known as SLCA2), as a reticulocyte-specific receptor for the PvRBP2a parasite ligand using mass spectrometry, flow cytometry, biochemical and parasite invasion assays. We characterized the expression level of CD98 at the surface of immature reticulocytes (CD71+) and identified an interaction between CD98 and PvRBP2a expressed at the merozoite surface. Our results identify CD98 as an additional host membrane protein, besides CD71, that is directly associated with P. vivax reticulocyte tropism. These findings highlight the potential of using PvRBP2a as a vaccine target against P. vivax malaria.


Assuntos
Eritrócitos/parasitologia , Cadeia Pesada da Proteína-1 Reguladora de Fusão/metabolismo , Malária Vivax/metabolismo , Plasmodium vivax/metabolismo , Antígenos CD , Antígenos de Protozoários/genética , Antígenos de Protozoários/metabolismo , Eritrócitos/metabolismo , Cadeia Pesada da Proteína-1 Reguladora de Fusão/genética , Interações Hospedeiro-Parasita , Humanos , Malária Vivax/sangue , Malária Vivax/genética , Plasmodium vivax/genética , Ligação Proteica , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores da Transferrina , Reticulócitos/metabolismo , Reticulócitos/parasitologia
9.
Front Immunol ; 12: 611673, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34220793

RESUMO

In tuberculosis, T cell-mediated immunity is extensively studied whilst B cells received limited attention in human and mice. Of interest, Mycobacterium tuberculosis (Mtb) does increase IL-4 Receptor-alpha (IL4Rα) expression in murine B cells. To better understand the role of IL4Rα signalling in B cells, we compared wild type mice with B cell-specific IL4Rα deficient mice (mb1creIL-4Rα-/lox mice). Chronic Mtb aerosol infection in mb1creIL-4Rα-/lox mice reduced lung and spleen bacterial burdens, compared to littermate (IL-4Rα-/lox) control animals. Consequently, lung pathology, inflammation and inducible nitric oxide synthase (iNOS) expression were reduced in the lungs of mb1creIL-4Rα-/lox mice, which was also accompanied by increased lung IgA and decreased IgG1 levels. Furthermore, intratracheal adoptive transfer of wild-type B cells into B cell-specific IL4Rα deficient mice reversed the protective phenotype. Moreover, constitutively mCherry expressing Mtb showed decreased association with B cells from mb1creIL-4Rα-/lox mice ex vivo. In addition, supernatants from Mtb-exposed B cells of mb1creIL-4Rα-/lox mice also increased the ability of macrophages to produce nitric oxide, IL-1ß, IL-6 and TNF. Together, this demonstrates that IL-4-responsive B cells are detrimental during the chronic phase of tuberculosis in mice with perturbed antibody profiles, inflammatory cytokines and tnf and stat1 levels in the lungs.


Assuntos
Linfócitos B/imunologia , Imunoglobulina A/metabolismo , Interleucina-4/metabolismo , Pulmão/metabolismo , Macrófagos/patologia , Mycobacterium tuberculosis/fisiologia , Tuberculose/imunologia , Animais , Doença Crônica , Citocinas/metabolismo , Modelos Animais de Doenças , Humanos , Mediadores da Inflamação/metabolismo , Pulmão/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Receptores de Superfície Celular/genética , Transdução de Sinais
10.
Int J Mol Sci ; 22(11)2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-34205973

RESUMO

The Arabidopsis AtCRK5 protein kinase is involved in the establishment of the proper auxin gradient in many developmental processes. Among others, the Atcrk5-1 mutant was reported to exhibit a delayed gravitropic response via compromised PIN2-mediated auxin transport at the root tip. Here, we report that this phenotype correlates with lower superoxide anion (O2•-) and hydrogen peroxide (H2O2) levels but a higher nitric oxide (NO) content in the mutant root tips in comparison to the wild type (AtCol-0). The oxidative stress inducer paraquat (PQ) triggering formation of O2•- (and consequently, H2O2) was able to rescue the gravitropic response of Atcrk5-1 roots. The direct application of H2O2 had the same effect. Under gravistimulation, correct auxin distribution was restored (at least partially) by PQ or H2O2 treatment in the mutant root tips. In agreement, the redistribution of the PIN2 auxin efflux carrier was similar in the gravistimulated PQ-treated mutant and untreated wild type roots. It was also found that PQ-treatment decreased the endogenous NO level at the root tip to normal levels. Furthermore, the mutant phenotype could be reverted by direct manipulation of the endogenous NO level using an NO scavenger (cPTIO). The potential involvement of AtCRK5 protein kinase in the control of auxin-ROS-NO-PIN2-auxin regulatory loop is discussed.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Ácidos Indolacéticos/metabolismo , Proteínas Serina-Treonina Quinases/genética , Receptores de Superfície Celular/genética , Arabidopsis/crescimento & desenvolvimento , Transporte Biológico/genética , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Gravitação , Gravitropismo/genética , Peróxido de Hidrogênio/farmacologia , Meristema/genética , Meristema/crescimento & desenvolvimento , Óxido Nítrico/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Paraquat/farmacologia , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Espécies Reativas de Oxigênio/metabolismo
11.
Int J Mol Sci ; 22(11)2021 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-34198929

RESUMO

The molecular mechanisms of insect resistance to Cry toxins generated from the bacterium Bacillus thuringiensis (Bt) urgently need to be elucidated to enable the improvement and sustainability of Bt-based products. Although downregulation of the expression of midgut receptor genes is a pivotal mechanism of insect resistance to Bt Cry toxins, the underlying transcriptional regulation of these genes remains elusive. Herein, we unraveled the regulatory mechanism of the downregulation of the ABC transporter gene PxABCG1 (also called Pxwhite), a functional midgut receptor of the Bt Cry1Ac toxin in Plutella xylostella. The PxABCG1 promoters of Cry1Ac-susceptible and Cry1Ac-resistant strains were cloned and analyzed, and they showed clear differences in activity. Subsequently, a dual-luciferase reporter assay, a yeast one-hybrid (Y1H) assay, and RNA interference (RNAi) experiments demonstrated that a cis-mutation in a binding site of the Hox transcription factor Antennapedia (Antp) decreased the promoter activity of the resistant strain and eliminated the binding and regulation of Antp, thereby enhancing the resistance of P. xylostella to the Cry1Ac toxin. These results advance our knowledge of the roles of cis- and trans-regulatory variations in the regulation of midgut Cry receptor genes and the evolution of Bt resistance, contributing to a more complete understanding of the Bt resistance mechanism.


Assuntos
Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Toxinas de Bacillus thuringiensis/genética , Proteínas de Insetos/genética , Resistência a Inseticidas/genética , Receptores de Superfície Celular/genética , Animais , Bacillus thuringiensis/genética , Endotoxinas/genética , Lepidópteros/efeitos dos fármacos , Lepidópteros/genética , Mutação/genética , Regiões Promotoras Genéticas/genética
12.
Sci Rep ; 11(1): 14015, 2021 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-34234248

RESUMO

Venous thromboembolism is the third common cardiovascular disease and is composed of two entities, deep vein thrombosis (DVT) and its potential fatal form, pulmonary embolism (PE). While PE is observed in ~ 40% of patients with documented DVT, there is limited biomarkers that can help identifying patients at high PE risk. To fill this need, we implemented a two hidden-layers artificial neural networks (ANN) on 376 antibodies and 19 biological traits measured in the plasma of 1388 DVT patients, with or without PE, of the MARTHA study. We used the LIME algorithm to obtain a linear approximate of the resulting ANN prediction model. As MARTHA patients were typed for genotyping DNA arrays, a genome wide association study (GWAS) was conducted on the LIME estimate. Detected single nucleotide polymorphisms (SNPs) were tested for association with PE risk in MARTHA. Main findings were replicated in the EOVT study composed of 143 PE patients and 196 DVT only patients. The derived ANN model for PE achieved an accuracy of 0.89 and 0.79 in our training and testing sets, respectively. A GWAS on the LIME approximate identified a strong statistical association peak (rs1424597: p = 5.3 × 10-7) at the PLXNA4 locus. Homozygote carriers for the rs1424597-A allele were then more frequently observed in PE than in DVT patients from the MARTHA (2% vs. 0.4%, p = 0.005) and the EOVT (3% vs. 0%, p = 0.013) studies. In a sample of 112 COVID-19 patients known to have endotheliopathy leading to acute lung injury and an increased risk of PE, decreased PLXNA4 levels were associated (p = 0.025) with worsened respiratory function. Using an original integrated proteomics and genetics strategy, we identified PLXNA4 as a new susceptibility gene for PE whose exact role now needs to be further elucidated.


Assuntos
Predisposição Genética para Doença/genética , Redes Neurais de Computação , Proteômica , Embolia Pulmonar/sangue , Embolia Pulmonar/genética , Receptores de Superfície Celular/sangue , Receptores de Superfície Celular/genética , Adulto , COVID-19/complicações , Feminino , Estudo de Associação Genômica Ampla , Humanos , Masculino , Fenótipo , Polimorfismo de Nucleotídeo Único , Embolia Pulmonar/complicações , Embolia Pulmonar/metabolismo
13.
Nat Commun ; 12(1): 3516, 2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-34112782

RESUMO

Profiling studies have revealed considerable phenotypic heterogeneity in cancer-associated fibroblasts (CAFs) present within the tumour microenvironment, however, functional characterisation of different CAF subsets is hampered by the lack of specific markers defining these populations. Here we show that genetic deletion of the Endo180 (MRC2) receptor, predominantly expressed by a population of matrix-remodelling CAFs, profoundly limits tumour growth and metastasis; effects that can be recapitulated in 3D co-culture assays. This impairment results from a CAF-intrinsic contractility defect and reduced CAF viability, which coupled with the lack of phenotype in the normal mouse, demonstrates that upregulated Endo180 expression by a specific, potentially targetable CAF subset is required to generate a supportive tumour microenvironment. Further, characterisation of a tumour subline selected via serial in vivo passage for its ability to overcome these stromal defects provides important insight into, how tumour cells adapt to a non-activated stroma in the early stages of metastatic colonisation.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Fibroblastos Associados a Câncer/metabolismo , Glicoproteínas de Membrana/metabolismo , Receptores de Superfície Celular/metabolismo , Esferoides Celulares/metabolismo , Células Estromais/metabolismo , Microambiente Tumoral/genética , Animais , Neoplasias da Mama/patologia , Neoplasias da Mama/secundário , Fibroblastos Associados a Câncer/citologia , Proliferação de Células/genética , Sobrevivência Celular/genética , Células Cultivadas , Técnicas de Cocultura , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Feminino , Humanos , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Células NIH 3T3 , Metástase Neoplásica , Receptores de Superfície Celular/genética , Ensaio Tumoral de Célula-Tronco
14.
Cell Mol Life Sci ; 78(14): 5569-5585, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34089345

RESUMO

The macrophage mannose receptor (CD206, MR) is an endocytic lectin receptor which plays an important role in homeostasis and innate immunity, however, the endogenous glycan and glycoprotein ligands recognized by its C-type lectin domains (CTLD) have not been well studied. Here we used the murine MR CTLD4-7 coupled to the Fc-portion of human IgG (MR-Fc) to investigate the MR glycan and glycoprotein recognition. We probed 16 different cancer and control tissues using the MR-Fc, and observed cell- and tissue-specific binding with varying intensity. All cancer tissues and several control tissues exhibited MR-Fc ligands, intracellular and/or surface-located. We further confirmed the presence of ligands on the surface of cancer cells by flow cytometry. To characterize the fine specificity of the MR for glycans, we screened a panel of glycan microarrays. Remarkably, the results indicate that the CTLD4-7 of the MR is highly selective for specific types of pauci- and oligomannose N-glycans among hundreds of glycans tested. As lung cancer tissue and the lung cancer cell line A549 showed intense MR-Fc binding, we further investigated the MR glycoprotein ligands in those cells by immunoprecipitation and glycoproteomic analysis. All enriched glycoproteins, of which 42 were identified, contained pauci- or oligomannose N-glycans, confirming the microarray results. Our study demonstrates that the MR CTLD4-7 is highly selective for pauci- and oligomannosidic N-glycans, structures that are often elevated in tumor cells, and suggest a potential role for the MR in tumor biology.


Assuntos
Glicoproteínas/metabolismo , Lectinas Tipo C/metabolismo , Neoplasias Pulmonares/patologia , Lectinas de Ligação a Manose/metabolismo , Oligossacarídeos/metabolismo , Polissacarídeos/metabolismo , Receptores de Superfície Celular/metabolismo , Células A549 , Glicoproteínas/genética , Glicosilação , Humanos , Lectinas Tipo C/genética , Ligantes , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Lectinas de Ligação a Manose/genética , Modelos Moleculares , Receptores de Superfície Celular/genética
15.
Arch Microbiol ; 203(7): 4653-4662, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34173855

RESUMO

Yersinia enterocolitica is an important zoonotic pathogen, which seriously endangers food-safety risk. In this study, the recombinant outer membrane protein OmpF and its antibody were prepared and coupled with immunomagnetic beads (IMBs) to capture Y. enterocolitica in food samples, combining the quantitative PCR detection with primers of virulence factor gene foxA for Yersinia enterocolitica contamination. The results showed that the capture efficiency of approximately 80% using anti-OmpF antibody-immunomagnetic beads and linearly dependent capture under 101-105 CFU/mL Y. enterocolitica compared with less than 10% capture of other bacteria. The detection limit of 64 CFU/mL was obtained based on foxA gene PCR detection combined with capture of the anti-OmpF antibody-immunomagnetic beads to detect Yersinia enterocolitica in artificially contaminated milk and pork samples. Compared to the culture method, the developed IMBs-qPCR method has higher consistency, was less time consuming, which taken together provides an effective alternative method for rapid detection of Y. enterocolitica in food.


Assuntos
Proteínas da Membrana Bacteriana Externa , Microbiologia de Alimentos , Reação em Cadeia da Polimerase em Tempo Real , Receptores de Superfície Celular , Yersinia enterocolitica , Proteínas da Membrana Bacteriana Externa/genética , Microbiologia de Alimentos/métodos , Separação Imunomagnética , Reação em Cadeia da Polimerase em Tempo Real/normas , Receptores de Superfície Celular/genética , Sensibilidade e Especificidade , Yersinia enterocolitica/genética
16.
Clin Immunol ; 229: 108784, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34126239

RESUMO

OBJECTIVE: Osteoarthritis (OA), the leading cause of joint failure, is characterized by breakdown of articular cartilage and remodeling of subchondral bone in synovial joints. Despite the high prevalence and debilitating effects of OA, no disease-modifying drugs exist. Increasing evidence, including genetic variants of the interleukin 4 (IL-4) and IL-4 receptor genes, implicates a role for IL-4 in OA, however, the mechanism underlying IL-4 function in OA remains unknown. Here, we investigated the role of IL-4 in OA pathogenesis. METHODS: Il4-, myeloid-specific-Il4ra-, and Stat6-deficient and control mice were subjected to destabilization of the medial meniscus to induce OA. Macrophages, osteoclasts, and synovial explants were stimulated with IL-4 in vitro, and their function and expression profiles characterized. RESULTS: Mice lacking IL-4, IL-4Ra in myeloid cells, or STAT6 developed exacerbated cartilage damage and osteophyte formation relative to WT controls. In vitro analyses revealed that IL-4 downregulates osteoarthritis-associated genes, enhances macrophage phagocytosis of cartilage debris, and inhibits osteoclast differentiation and activation via the type I receptor. CONCLUSION: Our findings demonstrate that IL-4 protects against osteoarthritis in a myeloid and STAT6-dependent manner. Further, IL-4 can promote an immunomodulatory microenvironment in which joint-resident macrophages polarize towards an M2 phenotype and efficiently clear pro-inflammatory debris, and osteoclasts maintain a homeostatic level of activity in subchondral bone. These findings support a role for IL-4 modulation of myeloid cell types in maintenance of joint health and identify a pathway that could provide therapeutic benefit for osteoarthritis.


Assuntos
Interleucina-4/imunologia , Macrófagos/imunologia , Osteoartrite/prevenção & controle , Osteoclastos/imunologia , Animais , Cartilagem Articular/imunologia , Cartilagem Articular/patologia , Modelos Animais de Doenças , Humanos , Inflamação/imunologia , Inflamação/patologia , Inflamação/prevenção & controle , Interleucina-4/deficiência , Interleucina-4/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Osteoartrite/imunologia , Osteoartrite/patologia , Osteoclastos/patologia , Fagocitose , Receptores de Superfície Celular/deficiência , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/imunologia , Fator de Transcrição STAT6/deficiência , Fator de Transcrição STAT6/genética , Fator de Transcrição STAT6/imunologia , Transdução de Sinais/imunologia
17.
Nat Commun ; 12(1): 3626, 2021 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-34131117

RESUMO

Platelet aggregation at the site of atherosclerotic vascular injury is the underlying pathophysiology of myocardial infarction and stroke. To build upon prior GWAS, here we report on 16 loci identified through a whole genome sequencing (WGS) approach in 3,855 NHLBI Trans-Omics for Precision Medicine (TOPMed) participants deeply phenotyped for platelet aggregation. We identify the RGS18 locus, which encodes a myeloerythroid lineage-specific regulator of G-protein signaling that co-localizes with expression quantitative trait loci (eQTL) signatures for RGS18 expression in platelets. Gene-based approaches implicate the SVEP1 gene, a known contributor of coronary artery disease risk. Sentinel variants at RGS18 and PEAR1 are associated with thrombosis risk and increased gastrointestinal bleeding risk, respectively. Our WGS findings add to previously identified GWAS loci, provide insights regarding the mechanism(s) by which genetics may influence cardiovascular disease risk, and underscore the importance of rare variant and regulatory approaches to identifying loci contributing to complex phenotypes.


Assuntos
Plaquetas/metabolismo , Mapeamento Cromossômico , Sequenciamento Completo do Genoma , Sequência de Bases , Proteínas de Ligação ao GTP , Estudo de Associação Genômica Ampla , Células HEK293 , Humanos , Células K562 , Fenótipo , Agregação Plaquetária , Testes de Função Plaquetária , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Proteínas RGS/genética , Proteínas RGS/metabolismo , Receptores de Superfície Celular/genética , Trombose/genética
18.
Transfusion ; 61(7): 2169-2178, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34181769

RESUMO

BACKGROUND: Despite the significant adverse clinical consequences of RBC alloimmunization, our understanding of the signals that induce immune responses to transfused RBCs remains incomplete. Though RBC storage has been shown to enhance alloimmunization in the hen egg lysozyme, ovalbumin, and human Duffy (HOD) RBC alloantigen mouse model, the molecular signals leading to immune activation in this system remain unclear. Given that the nonclassical major histocompatibility complex (MHC) Class I molecule CD1D can bind to multiple different lysophospholipids and direct immune activation, we hypothesized that storage of RBCs increases lysophospholipids known to bind CD1D, and further that recipient CD1D recognition of these altered lipids mediates storage-induced alloimmunization responses. STUDY DESIGN AND METHODS: We used a mass spectrometry-based approach to analyze the changes in lysophospholipids that are induced during storage of mouse RBCs. CD1D knockout (CD1D-KO) and wild-type (WT) control mice were transfused with stored HOD RBCs to measure the impact of CD1D deficiency on RBC alloimmunization. RESULTS: RBC storage results in alterations in multiple lysophospholipid species known to bind to CD1D and activate the immune system. Prior to transfusion, CD1D-deficient mice had lower baseline levels of polyclonal immunoglobulin (IgG) relative to WT mice. In response to stored RBC transfusion, CD1D-deficient mice generated similar levels of anti-HOD IgM and anti-HOD IgG. CONCLUSION: Although storage of RBCs leads to alteration of several lysophospholipids known to be capable of binding CD1D, storage-induced RBC alloimmunization responses are not impacted by recipient CD1D deficiency.


Assuntos
Antígenos CD1d/imunologia , Preservação de Sangue , Transfusão de Sangue , Eritrócitos/imunologia , Isoanticorpos/biossíntese , Isoantígenos/imunologia , Lisofosfolipídeos/sangue , Reação Transfusional/imunologia , Alarminas/sangue , Alarminas/imunologia , Animais , Especificidade de Anticorpos , Antígenos CD1d/genética , Antígenos CD1d/metabolismo , Sistema do Grupo Sanguíneo Duffy/genética , Sistema do Grupo Sanguíneo Duffy/imunologia , Feminino , Imunização , Imunoglobulina G/biossíntese , Imunoglobulina G/imunologia , Imunoglobulina M/biossíntese , Imunoglobulina M/imunologia , Isoanticorpos/imunologia , Lisofosfolipídeos/metabolismo , Masculino , Espectrometria de Massas , Camundongos , Camundongos Endogâmicos , Camundongos Knockout , Camundongos Transgênicos , Muramidase/imunologia , Ovalbumina/imunologia , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/imunologia
19.
BMC Plant Biol ; 21(1): 291, 2021 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-34167462

RESUMO

Brassinosteroids (BRs) play important roles in plant growth and development. Although BR receptors have been intensively studied in Arabidopsis, those in foxtail millet remain largely unknown. Here, we show that the BR signaling function of BRASSINOSTEROID INSENSITIVE 1 (BRI1) is conserved between Arabidopsis and foxtail millet, a new model species for C4 and Panicoideae grasses. We identified four putative BR receptor genes in the foxtail millet genome: SiBRI1, SiBRI1-LIKE RECEPTOR KINASE 1 (SiBRL1), SiBRL2 and SiBRL3. Phylogenetic analysis was used to classify the BR receptors in dicots and monocots into three branches. Analysis of their expression patterns by quantitative real-time PCR (qRT-PCR) showed that these receptors were ubiquitously expressed in leaves, stems, dark-grown seedlings, roots and non-flowering spikelets. GFP fusion experiments verified that SiBRI1 localized to the cell membrane. We also explored the SiBRI1 function in Arabidopsis through complementation experiments. Ectopic overexpression of SiBRI1 in an Arabidopsis BR receptor loss-of-function mutant, bri1-116, mostly reversed the developmental defects of the mutant. When SiBRI1 was overexpressed in foxtail millet, the plants showed a drooping leaf phenotype and root development inhibition, lateral root initiation inhibition, and the expression of BR synthesis genes was inhibited. We further identified BRI1-interacting proteins by immunoprecipitation (IP)-mass spectrometry (MS). Our results not only demonstrate that SiBRI1 plays a conserved role in BR signaling in foxtail millet but also provide insight into the molecular mechanism of SiBRI1.


Assuntos
Brassinosteroides/metabolismo , Genes de Plantas/genética , Proteínas de Plantas/genética , Receptores de Superfície Celular/genética , Setaria (Planta)/genética , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/fisiologia , Evolução Molecular , Filogenia , Proteínas de Plantas/metabolismo , Proteínas de Plantas/fisiologia , Raízes de Plantas/crescimento & desenvolvimento , Proteínas Quinases/genética , Proteínas Quinases/fisiologia , Receptores de Superfície Celular/metabolismo , Receptores de Superfície Celular/fisiologia , Setaria (Planta)/metabolismo
20.
Life Sci ; 280: 119671, 2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34087284

RESUMO

AIMS: Focal segmental glomerulosclerosis (FSGS) is the common cause of chronic renal disease worldwide. Although there are many etiologic factors which have common theme of podocyte injury conclusive etiology is not clearly understood. In this study, we aimed to explore the role of autophagy in the pathogenesis of podocyte injury, which is the key point in disease progression, and the roles of intrarenal microRNAs and the prorenin receptor (PRR) in the 5/6 nephrectomy and adriamycin nephropathy models of FSGS. MAIN METHODS: For experimental FSGS model, 5/6 nephrectomy and adriamycin nephropathy models were created and characterized in adult Sprague Dawley rats. Microarray analysis was performed on FSGS and control groups that was confirmed by q-RT-PCR. Beclin1, LC3B, PRR, ATG7 and ATG5 expression were evaluated by western blotting and immunohistochemistry. Also, Beclin1 and PRR expression were measured by ELISA. Glomerular podocyte isolation was performed and autophagic activity was evaluated in podocytes before and after transfection with miRNA mimic and antagonists. KEY FINDINGS: Glomerular expression of Beclin1, LC3B, PRR, ATG7 and ATG5 were significantly lower in the 5/6 nephrectomy than adriamycin nephropathy group and in both groups lower when compared to control groups. Western blot results were consistent with immunohistochemical data. Electron microscopy revealed signs of impaired autophagy in FSGS. Autophagic activity decreased significantly after miR-214, miR-132 and miR-34c mimics and increased after transfection with antagonists. SIGNIFICANCE: These results showed that the role of autophagic activity and decreased expression of PRR in FSGS pathogenesis and miR-34c, miR-132 and miR-214 could be a potential treatment strategy by regulating autophagy.


Assuntos
Glomerulosclerose Segmentar e Focal/genética , MicroRNAs/genética , Receptores de Superfície Celular/genética , Animais , Autofagia , Células Cultivadas , Regulação da Expressão Gênica , Glomerulosclerose Segmentar e Focal/patologia , Masculino , Ratos , Ratos Sprague-Dawley
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