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1.
J Neuroimmunol ; 384: 578219, 2023 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-37813042

RESUMO

Small ubiquitin like modifiers (SUMO) are reversible posttranslational modifiers of intracellular proteins. In the CNS, expression of myelin genes is regulated by state of SUMOylation of their respective transcription factors. In the immune system, deSUMOylation activates innate immune responses and promotes anti-viral immunity. However, the role played by SUMO in an adaptive immune response and in the development of T cell mediated autoimmune disease has not been previously described. TAK981 is a synthetic small molecule which by forming adducts with SUMO proteins prevents SUMOylation. We examined the expression of myelin genes and their transcription factors following culture with TAK981 in Oligodendrocyte Precursor Cells (OPC). We found that myelin basic protein (MBP), a key myelin protein, is upregulated in OPC in the presence of TAK981. We also found increased expression of transcription factors Sox10 and Myrf, which engage in the expression of MBP. In the Cuprizone model of demyelination/remyelination, animals which were treated with TAK981 showed increased remyelination in areas of demyelination and an increase in the number of maturing oligodendrocytes compared to vehicle treated controls. In in vitro cultures of lymphocytes, TAK981 reduced the expression of TH17 in T cells in mice immunized with MOGp35-55. Following in vivo treatment with TAK981, there was a significant reduction in the clinical and pathological severity in mice immunized to develop experimental allergic encephalitis (EAE). The dual effects of deSUMOylation on remyelination and in regulating an autoimmune adaptive response offers a novel approach to the management of human inflammatory demyelinating diseases such as multiple sclerosis.


Assuntos
Doenças do Sistema Nervoso Central , Doenças Desmielinizantes , Remielinização , Camundongos , Humanos , Animais , Doenças Desmielinizantes/induzido quimicamente , Doenças Desmielinizantes/tratamento farmacológico , Doenças Desmielinizantes/metabolismo , Remielinização/fisiologia , Sumoilação , Interleucina-17 , Diferenciação Celular , Bainha de Mielina/patologia , Oligodendroglia/metabolismo , Cuprizona/toxicidade , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Doenças do Sistema Nervoso Central/metabolismo , Fatores de Transcrição/metabolismo , Camundongos Endogâmicos C57BL , Modelos Animais de Doenças
2.
Proc Natl Acad Sci U S A ; 117(35): 21527-21535, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32817520

RESUMO

Given the known neuroreparative actions of IL-33 in experimental models of central nervous system (CNS) injury, we predicted that compounds which induce IL-33 are likely to promote remyelination. We found anacardic acid as a candidate molecule to serve as a therapeutic agent to promote remyelination. Addition of anacardic acid to cultured oligodendrocyte precursor cells (OPCs) rapidly increased expression of myelin genes and myelin proteins, suggesting a direct induction of genes involved in myelination by anacardic acid. Also, when added to OPCs, anacardic acid resulted in the induction of IL-33. In vivo, treatment of with anacardic acid in doses which ranged from 0.025 mg/kg to 2.5 mg/kg, improved pathologic scores in experimental allergic encephalitis (EAE) and in the cuprizone model of demyelination/remyelination. Electron microscopic studies performed in mice fed with cuprizone and treated with anacardic acid showed lower g-ratio scores when compared to controls, suggesting increased remyelination of axons. In EAE, improvement in paralytic scores was seen when the drug was given prior to or following the onset of paralytic signs. In EAE and in the cuprizone model, areas of myelin loss, which are likely to remyelinate, was associated with a greater recruitment of IL-33-expressing OPCs in mice which received anacardic acid when compared to controls.


Assuntos
Ácidos Anacárdicos/farmacologia , Interleucina-33/biossíntese , Remielinização/efeitos dos fármacos , Animais , Sistema Nervoso Central/efeitos dos fármacos , Sistema Nervoso Central/metabolismo , Doenças Desmielinizantes/tratamento farmacológico , Doenças Desmielinizantes/metabolismo , Feminino , Interleucina-33/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína Básica da Mielina/metabolismo , Proteínas da Mielina/metabolismo , Bainha de Mielina/metabolismo , Células Precursoras de Oligodendrócitos/efeitos dos fármacos , Células Precursoras de Oligodendrócitos/metabolismo , Oligodendroglia/metabolismo , Remielinização/fisiologia , Células-Tronco/metabolismo
3.
Adv Biol Regul ; 76: 100694, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32019729

RESUMO

Sulfur assimilation is an essential metabolic pathway that regulates sulfation, amino acid metabolism, nucleotide hydrolysis, and organismal homeostasis. We recently reported that mice lacking bisphosphate 3'-nucleotidase (BPNT1), a key regulator of sulfur assimilation, develop iron-deficiency anemia (IDA) and anasarca. Here we demonstrate two approaches that successfully reduce metabolic toxicity caused by loss of BPNT1: 1) dietary methionine restriction and 2) overproduction of a key transcriptional regulator hypoxia inducible factor 2α (Hif-2a). Reduction of methionine in the diet reverses IDA in mice lacking BPNT1, through a mechanism of downregulation of sulfur assimilation metabolic toxicity. Gaining Hif-2a acts through a different mechanism by restoring iron homeostatic gene expression in BPNT1 deficient mouse intestinal organoids. Finally, as loss of BPNT1 impairs expression of known genetic modifiers of iron-overload, we demonstrate that intestinal-epithelium specific loss of BPNT1 attenuates hepatic iron accumulation in mice with homozygous C282Y mutations in homeostatic iron regulator (HFEC282Y), the most common cause of hemochromatosis in humans. Overall, our study uncovers genetic and dietary strategies to overcome anemia caused by defects in sulfur assimilation and identifies BPNT1 as a potential target for the treatment of hemochromatosis.


Assuntos
Anemia Ferropriva/genética , Proteína da Hemocromatose/genética , Hemocromatose/genética , Ferro/metabolismo , Nucleotidases/genética , Enxofre/metabolismo , Anemia Ferropriva/metabolismo , Anemia Ferropriva/patologia , Anemia Ferropriva/prevenção & controle , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Dieta , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica , Hemocromatose/metabolismo , Hemocromatose/patologia , Hemocromatose/prevenção & controle , Proteína da Hemocromatose/metabolismo , Homeostase/genética , Homozigoto , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Fígado/metabolismo , Fígado/patologia , Masculino , Metionina/administração & dosagem , Metionina/deficiência , Camundongos , Camundongos Knockout , Mutação , Nucleotidases/metabolismo , Organoides/metabolismo , Organoides/patologia , Transdução de Sinais
4.
Nature ; 566(7744): 398-402, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30760926

RESUMO

The human genome contains approximately 20 thousand protein-coding genes1, but the size of the collection of antigen receptors of the adaptive immune system that is generated by the recombination of gene segments with non-templated junctional additions (on B cells) is unknown-although it is certainly orders of magnitude larger. It has not been established whether individuals possess unique (or private) repertoires or substantial components of shared (or public) repertoires. Here we sequence recombined and expressed B cell receptor genes in several individuals to determine the size of their B cell receptor repertoires, and the extent to which these are shared between individuals. Our experiments revealed that the circulating repertoire of each individual contained between 9 and 17 million B cell clonotypes. The three individuals that we studied shared many clonotypes, including between 1 and 6% of B cell heavy-chain clonotypes shared between two subjects (0.3% of clonotypes shared by all three) and 20 to 34% of λ or κ light chains shared between two subjects (16 or 22% of λ or κ light chains, respectively, were shared by all three). Some of the B cell clonotypes had thousands of clones, or somatic variants, within the clonotype lineage. Although some of these shared lineages might be driven by exposure to common antigens, previous exposure to foreign antigens was not the only force that shaped the shared repertoires, as we also identified shared clonotypes in umbilical cord blood samples and all adult repertoires. The unexpectedly high prevalence of shared clonotypes in B cell repertoires, and identification of the sequences of these shared clonotypes, should enable better understanding of the role of B cell immune repertoires in health and disease.


Assuntos
Anticorpos/genética , Anticorpos/imunologia , Linfócitos B/imunologia , Células Clonais/imunologia , Receptores de Antígenos de Linfócitos B/imunologia , Adulto , Sequência de Aminoácidos , Anticorpos/química , Antígenos/imunologia , Linfócitos B/citologia , Linfócitos B/metabolismo , Sequência de Bases , Células Clonais/citologia , Células Clonais/metabolismo , Feminino , Sangue Fetal/citologia , Sangue Fetal/imunologia , Voluntários Saudáveis , Humanos , Recém-Nascido , Masculino , Receptores de Antígenos de Linfócitos B/química , Receptores de Antígenos de Linfócitos B/genética , Análise de Sequência de DNA
5.
Nat Commun ; 9(1): 2669, 2018 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-29991715

RESUMO

The high rate of antigenic drift in seasonal influenza viruses necessitates frequent changes in vaccine composition. Recent seasonal H3 vaccines do not protect against swine-origin H3N2 variant (H3N2v) strains that recently have caused severe human infections. Here, we report a human VH1-69 gene-encoded monoclonal antibody (mAb) designated H3v-47 that exhibits potent cross-reactive neutralization activity against human and swine H3N2 viruses that circulated since 1989. The crystal structure and electron microscopy reconstruction of H3v-47 Fab with the H3N2v hemagglutinin (HA) identify a unique epitope spanning the vestigial esterase and receptor-binding subdomains that is distinct from that of any known neutralizing antibody for influenza A H3 viruses. MAb H3v-47 functions largely by blocking viral egress from infected cells. Interestingly, H3v-47 also engages Fcγ receptor and mediates antibody dependent cellular cytotoxicity (ADCC). This newly identified conserved epitope can be used in design of novel immunogens for development of broadly protective H3 vaccines.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Epitopos/imunologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Antivirais/administração & dosagem , Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Reações Cruzadas/imunologia , Epitopos/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Humanos , Vírus da Influenza A Subtipo H3N2/efeitos dos fármacos , Vírus da Influenza A Subtipo H3N2/imunologia , Vírus da Influenza A Subtipo H3N2/fisiologia , Influenza Humana/imunologia , Influenza Humana/prevenção & controle , Influenza Humana/virologia , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/veterinária , Infecções por Orthomyxoviridae/virologia , Suínos , Doenças dos Suínos/imunologia , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/virologia
6.
Nat Microbiol ; 2: 16271, 2017 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-28134924

RESUMO

Respiratory syncytial virus (RSV) remains a major human pathogen, infecting the majority of infants before age two and causing re-infection throughout life. Despite decades of RSV research, there is no licensed RSV vaccine. Most candidate vaccines studied to date have incorporated the RSV fusion (F) surface glycoprotein, because the sequence of F is highly conserved among strains of RSV. To better define the human B cell response to RSV F, we isolated from a single donor 13 new neutralizing human monoclonal antibodies (mAbs) that recognize the RSV F protein in the pre-fusion conformation. Epitope binning studies showed that the majority of neutralizing mAbs targeted a new antigenic site on the globular head domain of F, designated here antigenic site VIII, which occupies an intermediate position between the previously defined major antigenic sites II and site Ø. Antibodies to site VIII competed for binding with antibodies to both of those adjacent neutralizing sites. The new mAbs exhibited unusual breadth for pre-fusion F-specific antibodies, cross-reacting with F proteins from both RSV subgroups A and B viruses. We solved the X-ray crystal structure of one site VIII mAb, hRSV90, in complex with pre-fusion RSV F protein. The structure revealed a large footprint of interaction for hRSV90 on RSV F, in which the heavy chain and light chain both have specific interactions mediating binding to site VIII, the heavy chain overlaps with site Ø, and the light chain interacts partially with site II.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Mapeamento de Epitopos , Epitopos/imunologia , Proteínas Virais de Fusão/imunologia , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/isolamento & purificação , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/isolamento & purificação , Anticorpos Antivirais/química , Anticorpos Antivirais/isolamento & purificação , Reações Cruzadas , Cristalografia por Raios X , Humanos , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Proteínas Virais de Fusão/química
7.
J Immunol ; 195(3): 1064-70, 2015 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-26101326

RESUMO

Sepsis is a major cause of neonatal mortality and morbidity worldwide. A recent report suggested that murine neonatal host defense against infection could be compromised by immunosuppressive CD71(+) erythroid splenocytes. We examined the impact of CD71(+) erythroid splenocytes on murine neonatal mortality to endotoxin challenge or polymicrobial sepsis and characterized circulating CD71(+) erythroid (CD235a(+)) cells in human neonates. Adoptive transfer or an Ab-mediated reduction in neonatal CD71(+) erythroid splenocytes did not alter murine neonatal survival to endotoxin challenge or polymicrobial sepsis challenge. Ex vivo immunosuppression of stimulated adult CD11b(+) cells was not limited to neonatal splenocytes; it also occurred with adult and neonatal bone marrow. Animals treated with anti-CD71 Ab showed reduced splenic bacterial load following bacterial challenge compared with isotype-treated mice. However, adoptive transfer of enriched CD71(+) erythroid splenocytes to CD71(+)-reduced animals did not reduce bacterial clearance. Human CD71(+)CD235a(+) cells were common among cord blood mononuclear cells and were shown to be reticulocytes. In summary, a lack of effect on murine survival to polymicrobial sepsis following adoptive transfer or diminution of CD71(+) erythroid splenocytes under these experimental conditions suggests that the impact of these cells on neonatal infection risk and progression may be limited. An unanticipated immune priming effect of anti-CD71 Ab treatment, rather than a reduction in immunosuppressive CD71(+) erythroid splenocytes, was likely responsible for the reported enhanced bacterial clearance. In humans, the well-described rapid decrease in circulating reticulocytes after birth suggests that they may have a limited role in reducing inflammation secondary to microbial colonization.


Assuntos
Antígenos CD/imunologia , Células da Medula Óssea/imunologia , Células Eritroides/imunologia , Receptores da Transferrina/imunologia , Sepse/imunologia , Transferência Adotiva , Animais , Anticorpos/imunologia , Antígeno CD11b/metabolismo , Endotoxinas/farmacologia , Feminino , Sangue Fetal/citologia , Sangue Fetal/imunologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Reticulócitos/imunologia , Baço/citologia , Baço/imunologia
8.
Immunity ; 41(3): 451-464, 2014 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-25220211

RESUMO

Innate immune responses are critical for mucosal immunity. Here we describe an innate lymphocyte population, iCD8α cells, characterized by expression of CD8α homodimers. iCD8α cells exhibit innate functional characteristics such as the capacity to engulf and kill bacteria. Development of iCD8α cells depends on expression of interleukin-2 receptor γ chain (IL-2Rγc), IL-15, and the major histocompatibility complex (MHC) class Ib protein H2-T3, also known as the thymus leukemia antigen or TL. While lineage tracking experiments indicated that iCD8α cells have a lymphoid origin, their development was independent of the transcriptional suppressor Id2, suggesting that these cells do not belong to the family of innate lymphoid cells. Finally, we identified cells with a similar phenotype in humans, which were profoundly depleted in newborns with necrotizing enterocolitis. These findings suggest a critical role of iCD8α cells in immune responses associated with the intestinal epithelium.


Assuntos
Apresentação de Antígeno/imunologia , Antígenos CD8/biossíntese , Imunidade nas Mucosas/imunologia , Mucosa Intestinal/citologia , Linfócitos/imunologia , Animais , Citrobacter rodentium/imunologia , Citocalasina D/farmacologia , Enterocolite Necrosante , Helicobacter pylori/imunologia , Antígenos de Histocompatibilidade Classe I/biossíntese , Humanos , Proteína 2 Inibidora de Diferenciação/genética , Subunidade gama Comum de Receptores de Interleucina/biossíntese , Interleucina-15/biossíntese , Interleucina-2/biossíntese , Interleucina-7/biossíntese , Mucosa Intestinal/imunologia , Ativação Linfocitária/imunologia , Linfócitos/classificação , Linfócitos/citologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fagocitose/efeitos dos fármacos , Fagocitose/imunologia
9.
PLoS One ; 9(6): e99042, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24905458

RESUMO

BACKGROUND: Gastrointestinal barrier immaturity predisposes preterm infants to necrotizing enterocolitis (NEC). Intraepithelial lymphocytes (IEL) bearing the unconventional T cell receptor (TCR) γδ (γδ IEL) maintain intestinal integrity and prevent bacterial translocation in part through production of interleukin (IL) 17. OBJECTIVE: We sought to study the development of γδ IEL in the ileum of human infants and examine their role in NEC pathogenesis. We defined the ontogeny of γδ IEL proportions in murine and human intestine and subjected tcrδ-/- mice to experimental gut injury. In addition, we used polychromatic flow cytometry to calculate percentages of viable IEL (defined as CD3+ CD8+ CD103+ lymphocytes) and the fraction of γδ IEL in surgically resected tissue from infants with NEC and gestational age matched non-NEC surgical controls. RESULTS: In human preterm infants, the proportion of IEL was reduced by 66% in 11 NEC ileum resections compared to 30 non-NEC controls (p<0.001). While γδ IEL dominated over conventional αß IEL early in gestation in mice and in humans, γδ IEL were preferential decreased in the ileum of surgical NEC patients compared to non-NEC controls (50% reduction, p<0.05). Loss of IEL in human NEC was associated with downregulation of the Th17 transcription factor retinoic acid-related orphan nuclear hormone receptor C (RORC, p<0.001). TCRδ-deficient mice showed increased severity of experimental gut injury (p<0.05) with higher TNFα expression but downregulation of IL17A. CONCLUSION: Complimentary mouse and human data suggest a role of γδ IEL in IL17 production and intestinal barrier production early in life. Specific loss of the γδ IEL fraction may contribute to NEC pathogenesis. Nutritional or pharmacological interventions to support γδ IEL maintenance in the developing small intestine could serve as novel strategies for NEC prevention.


Assuntos
Enterocolite Necrosante/imunologia , Enterocolite Necrosante/cirurgia , Recém-Nascido Prematuro/imunologia , Intestino Delgado/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Subpopulações de Linfócitos T/imunologia , Animais , Células Cultivadas , Enterocolite Necrosante/genética , Enterocolite Necrosante/patologia , Feminino , Regulação da Expressão Gênica , Humanos , Recém-Nascido , Recém-Nascido Prematuro/crescimento & desenvolvimento , Interleucina-17/genética , Interleucina-17/imunologia , Intestino Delgado/crescimento & desenvolvimento , Intestino Delgado/patologia , Intestino Delgado/cirurgia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ocludina/genética , Receptores de Antígenos de Linfócitos T gama-delta/análise , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/patologia
10.
Gut ; 62(1): 73-82, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22267598

RESUMO

BACKGROUND: Necrotising enterocolitis (NEC) is the most common gastrointestinal emergency in premature infants. Immaturity of gastrointestinal immune regulation may predispose preterm infants to NEC as FOXP3 T regulatory cells (Treg) are critical for intestinal immune homoeostasis. OBJECTIVE: To investigate the hypothesis that abnormal developmental regulation of lamina propria Treg would define premature infants with NEC. DESIGN: Lamina propria mononuclear cell populations from surgically resected ileum from 18 patients with NEC and 30 gestational age-matched non-NEC surgical controls were prospectively isolated. Polychromatic flow cytometry was performed to phenotype and analyse lamina propria T cell populations. The cytokine gene expression profile in NEC tissue was compared with that of non-NEC controls. RESULTS: The total number of Treg, CD4, or CD8 T cells in each ileum section was independent of gestational age, age or postmenstrual age and similar between patients with NEC and controls. In contrast, the ratio of Treg to CD4 T cells or Treg to CD8 T cells was significantly lower in NEC ileum than in infants without NEC (medians 2.9% vs 6.6%, p=0.001 and medians 6.6% vs 25.9%, p<0.001, respectively). For any given number of CD4 or CD8 T cells, Treg were, on average, 60% lower in NEC ileum than in controls. NEC tissue cytokine gene expression profiles were characteristic of inhibited Treg development or function. Treg/CD4 and Treg/CD8 ratios recovered between initial resection for NEC and reanastomosis. CONCLUSION: The proportion of lamina propria Treg is significantly reduced in the ileum of premature infants with NEC and may contribute to the excessive inflammatory state of this disease.


Assuntos
Linfócitos T CD8-Positivos/metabolismo , Enterocolite Necrosante/imunologia , Fatores de Transcrição Forkhead/metabolismo , Doenças do Prematuro/imunologia , Mucosa Intestinal/imunologia , Linfócitos T Reguladores/metabolismo , Biomarcadores/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Estudos de Casos e Controles , Feminino , Citometria de Fluxo , Perfilação da Expressão Gênica , Humanos , Recém-Nascido , Recém-Nascido Prematuro , Contagem de Linfócitos , Masculino , Estudos Prospectivos
11.
Pediatr Infect Dis J ; 31(4): 413-4, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22173144

RESUMO

Necrotizing enterocolitis (NEC) is one of the most common gastrointestinal emergencies in premature infants and has been linked with viral antigens in as much as 40% of cases in single-center cohorts. We examined 28 tissue sections from surgically resected ileum from 27 preterm infants with NEC from 2 separate institutions for 15 common bacterial, viral, and parasitic gastrointestinal pathogens using multiplex reverse transcriptase polymerase chain reaction amplification and suspension array detection methods. We did not detect infectious enteritis pathogens in any of the NEC tissues and conclude that gastrointestinal pathogens are a rare cause of NEC.


Assuntos
Bactérias/isolamento & purificação , Enterocolite Necrosante/microbiologia , Íleo/microbiologia , Íleo/virologia , Parasitos/isolamento & purificação , Vírus/isolamento & purificação , Animais , Feminino , Humanos , Íleo/parasitologia , Lactente , Recém-Nascido , Masculino , Reação em Cadeia da Polimerase Multiplex , Reação em Cadeia da Polimerase Via Transcriptase Reversa
12.
Pediatr Dev Pathol ; 12(6): 443-9, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19203136

RESUMO

FOXP3(+) regulatory T cells (Treg) suppress innate and adaptive immune responses and are critical for intestinal immune homeostasis. Our objective was to define the postnatal developmental regulation of Treg in relationship to other T cells in the human intestinal tract. We analyzed 41 small and 18 large intestinal paraffin-embedded tissue samples from preterm and term infants with and without necrotizing enterocolitis (NEC) for the presence of CD3(+), CD4(+), CD8(+), and FOXP3(+) cells by immunohistochemistry. We compared labeled cells against age, gestational age (GA), or (corrected) postmenstrual age (PMA). The GA ranged from 23 to 40 weeks, with a mean of 32 (standard deviation, 4.7) weeks. Independent of age, GA, or PMA, the numbers of CD4(+) cells were higher in the small intestine compared to the large intestine (P = 0.046), except in patients with NEC. FOXP3(+) cells could be detected as early as 23 weeks in GA in both large and small bowel, and similar quantities were detected at the highest GA examined (40 weeks). We saw no statistically significant effect of GA, age, or PMA on total number of FOXP3(+) cells or by comparing FOXP3(+) to CD4(+) or FOXP3(+) to CD8(+) ratios, indicating intact ontogeny of Treg in intestinal tissue early in gestation. Human infants exhibit presence of mucosal FOXP3(+) cells in the small and large intestinal mucosa at birth and as early as 23 weeks GA. The frequency of FOXP3(+) cells and the ratios of FOXP3(+) to CD4(+) or CD8(+) cells do not change with increasing intrauterine development or postnatal age.


Assuntos
Fatores de Transcrição Forkhead/imunologia , Mucosa Intestinal/imunologia , Subpopulações de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Enterocolite Necrosante/imunologia , Feminino , Fatores de Transcrição Forkhead/metabolismo , Idade Gestacional , Humanos , Imunidade nas Mucosas/fisiologia , Imuno-Histoquímica , Recém-Nascido , Mucosa Intestinal/crescimento & desenvolvimento , Intestino Grosso/crescimento & desenvolvimento , Intestino Grosso/imunologia , Intestino Delgado/crescimento & desenvolvimento , Intestino Delgado/imunologia , Masculino , Mucosa/crescimento & desenvolvimento , Mucosa/imunologia , Subpopulações de Linfócitos T/metabolismo , Linfócitos T Reguladores/metabolismo
13.
Cancer Res ; 67(19): 9380-8, 2007 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-17909047

RESUMO

Clinical and animal studies indicate a role for cyclooxygenase-2 (COX-2) and the epidermal growth factor receptor (EGFR) in the development and progression of intestinal polyps and cancers. Although this combination of enzyme inhibition has shown synergy in intestinal polyp and tumor models, the exact mechanism for these effects remains undefined. Therefore, we sought to define the molecular mechanisms through which this process occurs. We observed a significant reduction in the number and size of small intestinal polyps in APC(min+/-) mice treated with either celecoxib (a selective COX-2 inhibitor) or erlotinib (Tarceva, an EGFR inhibitor). However, in combination, there was an overall prevention in the formation of polyps by over 96%. Furthermore, we observed a 70% reduction of colorectal xenograft tumors in mice treated with the combination and microarray analysis revealed genes involved in cell cycle progression were negatively regulated. Although we did not observe significant changes in mRNAs of genes with known apoptotic function, there was a significant increase of apoptosis in tumors from animals treated with the combination. The inhibition of EGFR also induced the down-regulation of COX-2 and further inhibited prostaglandin E2 formation. We observed similar effects on the prevention of intestinal adenomas and reduction of xenograft tumor volume when nonselective COX inhibitors were used in combination with erlotinib. Together, these findings suggest that the inhibition of both COX-2 and EGFR may provide a better therapeutic strategy than either single agent through a combination of decreased cellular proliferation and prostaglandin signaling as well as increased apoptosis.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Inibidores de Ciclo-Oxigenase 2/farmacologia , Receptores ErbB/antagonistas & inibidores , Pirazóis/farmacologia , Quinazolinas/farmacologia , Sulfonamidas/farmacologia , Animais , Anti-Inflamatórios não Esteroides/administração & dosagem , Celecoxib , Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/prevenção & controle , Cloridrato de Erlotinib , Humanos , Imuno-Histoquímica , Pólipos Intestinais/tratamento farmacológico , Pólipos Intestinais/enzimologia , Pólipos Intestinais/metabolismo , Intestino Delgado/enzimologia , Intestino Delgado/patologia , Camundongos , Camundongos Endogâmicos C57BL , Análise de Sequência com Séries de Oligonucleotídeos , Prostaglandinas/biossíntese , Prostaglandinas/metabolismo , Pirazóis/administração & dosagem , Quinazolinas/administração & dosagem , Sulfonamidas/administração & dosagem , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Proc Natl Acad Sci U S A ; 103(5): 1492-7, 2006 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-16432186

RESUMO

G protein-coupled receptor ligand-dependent transactivation of growth factor receptors has been implicated in human cancer cell proliferation, migration, and cell survival. For example, prostaglandin E(2) (PGE(2))-induced transactivation of the EGF receptor (EGFR) in colorectal carcinoma cells is mediated by means of a c-Src-dependent mechanism and regulates cell proliferation and migration. Recent evidence indicates that beta-arrestin 1 may act as an important mediator in G protein-coupled receptor-induced activation of c-Src. Whether beta-arrestin 1 serves a functional role in these events is, however, unknown. We investigated the effects of PGE(2) on colorectal cancer cells expressing WT and mutant beta-arrestin 1. Here we report that PGE(2) induces the association of a prostaglandin E receptor 4/beta-arrestin 1/c-Src signaling complex resulting in the transactivation of the EGFR and downstream Akt (PKB) signaling. The interaction of beta-arrestin 1 and c-Src is critical for the regulation of colorectal carcinoma cell migration in vitro as well as metastatic spread of disease to the liver in vivo. These results show that the prostaglandin E/beta-arrestin 1/c-Src signaling complex is a crucial step in PGE(2)-mediated transactivation of the EGFR and may play a pivotal role in tumor metastasis. Furthermore, our data implicate a functional role for beta-arrestin 1 as a mediator of cellular migration and metastasis.


Assuntos
Arrestinas/fisiologia , Neoplasias Colorretais/patologia , Arrestinas/metabolismo , Western Blotting , Proteína Tirosina Quinase CSK , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Movimento Celular , Colágeno/química , Citosol/metabolismo , Densitometria , Dinoprostona/metabolismo , Progressão da Doença , Combinação de Medicamentos , Receptores ErbB/metabolismo , Humanos , Imuno-Histoquímica , Imunoprecipitação , Laminina/química , Microscopia de Fluorescência , Modelos Biológicos , Metástase Neoplásica , Fosfotirosina/química , Proteínas Tirosina Quinases/metabolismo , Proteoglicanas/química , Receptores de Prostaglandina E/metabolismo , Receptores de Prostaglandina E Subtipo EP4 , Transdução de Sinais , Fatores de Tempo , Ativação Transcricional , beta-Arrestina 1 , beta-Arrestinas , Quinases da Família src
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