RESUMO
Chronic granulomatous disease (CGD) is characterized by recurrent infections and granuloma formation. In addition, we have observed a number of diverse autoimmune conditions in our CGD population, suggesting that patients with CGD are at an elevated risk for development of autoimmune disorders. In this report, we describe antiphospholipid syndrome, recurrent pericardial effusion, juvenile idiopathic arthritis, IgA nephropathy, cutaneous lupus erythematosus, and autoimmune pulmonary disease in the setting of CGD. The presence and type of autoimmune disease have important treatment implications for patients with CGD.
Assuntos
Doenças Autoimunes/etiologia , Doença Granulomatosa Crônica/complicações , Pneumopatias/etiologia , Derrame Pericárdico/etiologia , Adolescente , Adulto , Doenças Autoimunes/patologia , Doenças Autoimunes/terapia , Feminino , Doença Granulomatosa Crônica/patologia , Doença Granulomatosa Crônica/terapia , Humanos , Pneumopatias/patologia , Pneumopatias/terapia , Masculino , Derrame Pericárdico/patologia , Derrame Pericárdico/terapia , Fatores de RiscoAssuntos
4-Butirolactona/intoxicação , Tratamento de Emergência/métodos , Transtornos Relacionados ao Uso de Substâncias/complicações , Transtornos Relacionados ao Uso de Substâncias/diagnóstico , Inconsciência/induzido quimicamente , Adolescente , Europa (Continente)/epidemiologia , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Masculino , Prevalência , Detecção do Abuso de Substâncias , Transtornos Relacionados ao Uso de Substâncias/epidemiologia , Transtornos Relacionados ao Uso de Substâncias/terapiaRESUMO
P-glycoprotein (Pgp) is a well-defined ATP-binding cassette (ABC) protein and a close relative of cystic fibrosis transmembrane conductance regulator (CFTR), whose dysfunction causes cystic fibrosis (CF). It is postulated that Pgp can complement deficient CFTR functions because of structural and functional homologies. One of the most relevant functions appears to be the regulation of ATP release, which influences mucociliary clearance in respiratory epithelia by nucleotide receptor stimulation. However, mechanisms involved in ATP secretion remain a controversial issue. In the present study, CF epithelial cells (sigmaCFTE29ó) were transduced with the retroviral vector MP1m encoding Pgp, and thus, a stable Pgp-overexpressing CF cell line (sigmaCFTE29óPgp) was established and used for studies of hypothesized CFTR complementation. In addition, overexpression of native Pgp in sigmaCFTE29ó could also be achieved by long-term treatment with colchicine, a drug, which may be of great interest in CF therapy. We confirmed that overexpression of Pgp causes a significant increase in cellular ATP release, which could even be enhanced by stimulation with hypoosmolar medium. A potential clinical benefit is discussed.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Trifosfato de Adenosina/metabolismo , Fibrose Cística/metabolismo , Líquido Extracelular/metabolismo , Expressão Gênica , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/biossíntese , Células Cultivadas , Colchicina/farmacologia , Fibrose Cística/patologia , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Citometria de Fluxo , Expressão Gênica/efeitos dos fármacos , Vetores Genéticos , Humanos , Técnicas In Vitro , RNA Mensageiro/biossíntese , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Retroviridae/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Traqueia/patologia , TransfecçãoRESUMO
Major limitations to gene therapy using HSCs are low gene transfer efficiency and the inability of most therapeutic genes to confer a selective advantage on the gene-corrected cells. One approach to enrich for gene-modified cells in vivo is to include in the retroviral vector a drug resistance gene, such as the P140K mutant of the DNA repair enzyme O6-methylguanine-DNA methyltransferase (MGMT*). We transplanted 5 rhesus macaques with CD34+ cells transduced with lentiviral vectors encoding MGMT* and a fluorescent marker, with or without homeobox B4 (HOXB4), a potent stem cell self-renewal gene. Transgene expression and common integration sites in lymphoid and myeloid lineages several months after transplantation confirmed transduction of long-term repopulating HSCs. However, all animals showed only a transient increase in gene-marked lymphoid and myeloid cells after O6-benzylguanine (BG) and temozolomide (TMZ) administration. In 1 animal, cells transduced with MGMT* lentiviral vectors were protected and expanded after multiple courses of BG/TMZ, providing a substantial increase in the maximum tolerated dose of TMZ. Additional cycles of chemotherapy using 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU) resulted in similar increases in gene marking levels, but caused high levels of nonhematopoietic toxicity. Inclusion of HOXB4 in the MGMT* vectors resulted in no substantial increase in gene marking or HSC amplification after chemotherapy treatment. Our data therefore suggest that lentivirally mediated gene transfer in transplanted HSCs can provide in vivo chemoprotection of progenitor cells, although selection of long-term repopulating HSCs was not seen.
Assuntos
Antineoplásicos Alquilantes/farmacologia , Dacarbazina/análogos & derivados , Terapia Genética , Lentivirus/genética , O(6)-Metilguanina-DNA Metiltransferase/genética , Animais , Carmustina/farmacologia , Dacarbazina/farmacologia , Vetores Genéticos , Guanina/análogos & derivados , Guanina/farmacologia , Transplante de Células-Tronco Hematopoéticas , Proteínas de Homeodomínio/fisiologia , Macaca mulatta , TemozolomidaRESUMO
WHIM(warts, hypogammaglobulinemia, recurrent bacterial infection, and myelokathexis) syndrome is a rare immunodeficiency caused in many cases by autosomal dominant C-terminal truncation mutations in the chemokine receptor CXCR4. A prominent and unexplained feature of WHIM is myelokathexis (hypercellularity with apoptosis of mature myeloid cells in bone marrow and neutropenia). We transduced healthy human CD34(+) peripheral blood-mobilized stem cells (PBSCs) with retrovirus vector encoding wild-type (wt) CXCR4 or WHIM-type mutated CXCR4 and studied these cells ex vivo in culture and after engraftment in a nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mouse xenograft model. Neither wt CXCR4 nor mutated CXCR4 transgene expression itself enhanced apoptosis of neutrophils arising in transduced PBSC cultures even with stimulation by a CXCR4 agonist, stromal cell-derived factor-1 (SDF-1 [CXCL12]). Excess wt CXCR4 expression by transduced human PBSCs enhanced marrow engraftment, but did not affect bone marrow (BM) apoptosis or the release of transduced leukocytes into PB. However, mutated CXCR4 transgene expression further enhanced BM engraftment, but was associated with a significant increase in apoptosis of transduced cells in BM and reduced release of transduced leukocytes into PB. We conclude that increased apoptosis of mature myeloid cells in WHIM is secondary to a failure of marrow release and progression to normal myeloid cell senescence, and not a direct effect of activation of mutated CXCR4.
Assuntos
Agamaglobulinemia/genética , Infecções Bacterianas/etiologia , Células da Medula Óssea/patologia , Síndromes de Imunodeficiência/genética , Neutropenia/genética , Receptores CXCR4/fisiologia , Verrugas/genética , Substituição de Aminoácidos , Animais , Apoptose , Células da Medula Óssea/metabolismo , Sinalização do Cálcio , Movimento Celular , Quimiocina CXCL12 , Quimiocinas CXC/fisiologia , Ensaio de Unidades Formadoras de Colônias , Suscetibilidade a Doenças , Dosagem de Genes , Sobrevivência de Enxerto , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Mutação de Sentido Incorreto , Transplante de Células-Tronco de Sangue Periférico , Mutação Puntual , Quimera por Radiação , Receptores CXCR4/genética , Proteínas Recombinantes de Fusão/fisiologia , Recidiva , Transgenes , Transplante HeterólogoRESUMO
Retroviral gene therapy can restore immunity to infants with X-linked severe combined immunodeficiency (XSCID) caused by mutations in the IL2RG gene encoding the common gamma chain (gammac) of receptors for interleukins 2 (IL-2), -4, -7, -9, -15, and -21. We investigated the safety and efficacy of gene therapy as salvage treatment for older XSCID children with inadequate immune reconstitution despite prior bone marrow transplant from a parent. Subjects received retrovirus-transduced autologous peripherally mobilized CD34(+) hematopoietic cells. T-cell function significantly improved in the youngest subject (age 10 years), and multilineage retroviral marking occurred in all 3 children.
Assuntos
Terapia Genética/métodos , Imunidade/efeitos dos fármacos , Receptores de Interleucina-2/administração & dosagem , Doenças por Imunodeficiência Combinada Ligada ao Cromossomo X/imunologia , Doenças por Imunodeficiência Combinada Ligada ao Cromossomo X/terapia , Transplante de Células-Tronco Hematopoéticas/métodos , Humanos , Lactente , Mutação , Receptores de Interleucina-2/genética , Retroviridae/genética , Linfócitos T/imunologia , Transdução Genética , Transplante Autólogo , Doenças por Imunodeficiência Combinada Ligada ao Cromossomo X/genéticaRESUMO
A critical role for eosinophils in remodeling of allergic airways was observed in vivo upon disruption of the dblGATA enhancer that regulates expression of GATA-1, which resulted in an eosinophil-deficient phenotype in the DeltadblGATA mouse. We demonstrate here that bone marrow progenitors isolated from DeltadblGATA mice can differentiate into mature eosinophils when subjected to cytokine stimulation ex vivo. Cultured DeltadblGATA eosinophils contain cytoplasmic granules with immunoreactive major basic protein and they express surface Siglec F and transcripts encoding major basic protein, eosinophil peroxidase, and GATA-1, -2, and -3 to an extent indistinguishable from cultured wild-type eosinophils. Fibroblast coculture and bone marrow cross-transplant experiments indicate that the in vivo eosinophil deficit is an intrinsic progenitor defect, and remains unaffected by interactions with stromal cells. Interestingly, and in contrast to those from the wild type, a majority of the GATA-1 transcripts from cultured DeltadblGATA progenitors express a variant GATA-1 transcript that includes a first exon (1E(B)), located approximately 3700 bp downstream to the previously described first exon found in hemopoietic cells (1E(A)) and approximately 42 bp upstream to another variant first exon, 1E(C). These data suggest that cultured progenitors are able to circumvent the effects of the DeltadblGATA ablation by using a second, more proximal, promoter and use this mechanism to generate quantities of GATA-1 that will support eosinophil growth and differentiation.
Assuntos
Células da Medula Óssea/imunologia , Diferenciação Celular/genética , Linhagem da Célula/genética , Elementos Facilitadores Genéticos , Eosinófilos/metabolismo , Fator de Transcrição GATA1/genética , Células-Tronco Hematopoéticas/imunologia , Regiões Promotoras Genéticas/imunologia , Animais , Sítios de Ligação/genética , Sítios de Ligação/imunologia , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Diferenciação Celular/imunologia , Linhagem da Célula/imunologia , Proliferação de Células , Células Cultivadas , Técnicas de Cocultura , Eosinófilos/citologia , Eosinófilos/imunologia , Eosinófilos/transplante , Fator de Transcrição GATA1/fisiologia , Fator de Transcrição GATA2/genética , Fator de Transcrição GATA3/genética , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Interleucina-5/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Mutantes , Células Th2/imunologiaRESUMO
UNLABELLED: Interleukin-5 (IL-5) promotes signal transduction and expansion of eosinophil colonies in bone marrow via interactions with its heterodimeric receptor (IL-5R). Two variants encoding soluble forms of the alpha subunit (sIL-5R alpha) have been described, although the signals promoting and/or limiting differential transcription remain to be clarified. OBJECTIVES: Our intent was to explore the role of IL-5 in regulating differential transcription of these splice variants in vivo. METHODS: We have designed a quantitative reverse transcriptase-polymerase chain reaction assay to detect transcripts encoding the transmembrane, soluble 1 and 2 forms of IL-5R alpha in two strains of wild-type (BALB/c and C57BL/6) and corresponding IL-5 gene-deleted mice. Wild-type mice respond to S. mansoni infection with a gradual increase in serum IL-5 and eosinophilia, which is not observed in IL-5 gene-deleted mice. RESULTS AND CONCLUSIONS: We find that IL-5 is not necessary for differential splicing to occur in vivo, as all three forms of the IL-5R alpha are detected in both strains of IL-5 gene-deleted mice, with ratios of transcript expression (transmembrane : soluble 1 : soluble 2) that were indistinguishable from their wild-type counterparts. Differential splicing does vary markedly between strains, potentially because of local effects of strain-specific polymorphisms.
Assuntos
Interleucina-5/metabolismo , Receptores de Interleucina/biossíntese , Receptores de Interleucina/genética , Processamento Alternativo , Animais , Sequência de Bases , Medula Óssea/patologia , DNA Complementar/genética , Eosinófilos/patologia , Éxons , Interleucina-5/sangue , Interleucina-5/deficiência , Interleucina-5/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Dados de Sequência Molecular , Isoformas de Proteínas/biossíntese , Isoformas de Proteínas/genética , Receptores de Interleucina-5 , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Esquistossomose mansoni/genética , Esquistossomose mansoni/imunologia , Esquistossomose mansoni/patologia , Solubilidade , Especificidade da Espécie , Transcrição GênicaRESUMO
In addition to increased susceptibility to infections in patients with chronic granulomatous disease (CGD), a higher incidence of sterile inflammatory disorders in these patients has been noted. However, sarcoidosis has not been reported previously in CGD. In this report, we describe two patients who have CGD and a disorder consistent with sarcoidosis on the basis of unequivocal clinical-radiographic presentations, their responses to treatment, and serum angiotensin-converting enzyme levels. Serum angiotensin-converting enzyme levels were measured in 26 other patients with CGD to establish an appropriate reference range. A possible relationship between CGD and sarcoidosis is discussed.
Assuntos
Doença Granulomatosa Crônica/complicações , Sarcoidose/complicações , Adolescente , Doenças do Sistema Nervoso Central/complicações , Doenças do Sistema Nervoso Central/diagnóstico , Criança , Feminino , Doença Granulomatosa Crônica/diagnóstico , Humanos , Masculino , Sarcoidose/diagnóstico , Sarcoidose Pulmonar/complicações , Sarcoidose Pulmonar/diagnósticoRESUMO
X-linked severe combined immunodeficiency (XSCID) is characterized by profound immunodeficiency and early mortality, the only potential cure being hematopoietic stem cell (HSC) transplantation or gene therapy. Current clinical gene therapy protocols targeting HSCs are based upon ex vivo gene transfer, potentially limited by the adequacy of HSC harvest, transduction efficiencies of repopulating HSCs, and the potential loss of their engraftment potential during ex vivo culture. We demonstrate an important proof of principle by showing achievement of durable immune reconstitution in XSCID dogs following intravenous injection of concentrated RD114-pseudotyped retrovirus vector encoding the corrective gene, the interleukin-2 receptor gamma chain (gamma c). In 3 of 4 dogs treated, normalization of numbers and function of T cells were observed. Two long-term-surviving animals (16 and 18 months) showed significant marking of B lymphocytes and myeloid cells, normalization of IgG levels, and protective humoral immune response to immunization. There were no adverse effects from in vivo gene therapy, and in one dog that reached sexual maturity, sparing of gonadal tissue from gene transfer was demonstrated. This is the first demonstration that in vivo gene therapy targeting HSCs can restore both cellular and humoral immunity in a large-animal model of a fatal immunodeficiency.