ABSTRACT
CONTEXT: Human mastocytosis is a rare disease, in which the serotonergic system may be involved. OBJECTIVE: The objective of the present study was to examine the possible presence of serotonin (5-HT) and its 5-HT1A receptor (R) in the skin of patients with mastocytosis. In addition, the effect of the 5-HT1AR was tested on human mastocytosis cells, cultured in vitro. MATERIALS AND METHODS: The expression of 5-HT and 5-HT1AR in patients with urticaria pigmentosa and mastocytoma was studied using immunohistochemistry. The effects of 8-OH-DPAT, an agonist of 5-HT1AR, on the proliferation (cell number), viability, apoptosis, spontaneous release of histamine, as well as a possible 5-HT metabolism, in the human HMC-1 mast cell line, were investigated. RESULTS: Both 5-HT and 5-HT1AR were expressed in the mast cells in biopsies of mastocytoma and urticaria pigmentosa, as well as in HMC-1 cells. However, no metabolism of 5-HT by the cell line could be detected by the methodology used. The 5-HT1AR agonist had no significant effect on the viability and number of HMC-1 cells, and was without effect on the apoptosis. At concentrations of 10â»6 mol/L and 10â»8-10⻹° mol/L (i.e. also at physiological concentrations), the agonist inhibited histamine release by these cells by as much as 30%. CONCLUSION: These findings indicate that 5-HT and its 5-HT1AR are expressed in human mastocytosis and that an agonist of the 5-HT1AR might be of value in the treatment of these patients.
Subject(s)
Gene Expression Regulation, Neoplastic/immunology , Neoplasm Proteins/immunology , Receptor, Serotonin, 5-HT1A/immunology , Serotonin/immunology , Skin Neoplasms/immunology , Skin/immunology , 8-Hydroxy-2-(di-n-propylamino)tetralin/pharmacology , Adult , Cell Line , Female , Gene Expression Regulation, Neoplastic/drug effects , Humans , Male , Mast Cells/immunology , Mast Cells/metabolism , Mast Cells/pathology , Mastocytosis, Cutaneous , Middle Aged , Neoplasm Proteins/biosynthesis , Receptor, Serotonin, 5-HT1A/biosynthesis , Serotonin/metabolism , Serotonin Receptor Agonists/pharmacology , Skin/metabolism , Skin/pathology , Skin Neoplasms/metabolism , Skin Neoplasms/pathology , Tumor Cells, CulturedABSTRACT
Genetic association of programmed cell death-1 (PDCD1) has been implicated in several autoimmune inflammatory disorders. Hence, in this study, our main objective is to evaluate the association of PDCD1 gene to Wegener's granulomatosis (WG). We, thus, analyzed three single nucleotide polymorphisms (SNPs) in PDCD1 gene among WG patients and controls. Further, we quantified circulating serum levels of soluble (s) PD-1 in patients and controls. The methodologies used were ABI Taqman allelic discrimination and restriction fragment length polymorphism for genotyping and in-house ELISA for quantifying sPD-1. Statistical relevance was analyzed by Fischer's exact test. As a result, reduced AA homozygote for SNP in intron-1 was observed, among the patients. However, no association was demonstrated after Bonferroni correction. Also, no differences in genotype and allele frequency were elucidated for SNPs in intron-4 and exon-5. Moreover, we could not demonstrate circulating sPD-1. In conclusion, we show no association of selected SNPs in PDCD1 gene with WG.
Subject(s)
Antigens, CD/genetics , Apoptosis Regulatory Proteins/genetics , Genetic Predisposition to Disease , Granulomatosis with Polyangiitis/genetics , Polymorphism, Single Nucleotide , Adult , Aged , Aged, 80 and over , Alleles , Case-Control Studies , Exons , Gene Frequency , Genotype , Haplotypes , Homozygote , Humans , Introns , Linkage Disequilibrium , Middle Aged , Polymorphism, Restriction Fragment Length , Programmed Cell Death 1 Receptor , Solubility , SwedenABSTRACT
Myasthenia gravis (MG) is an autoimmune disease characterized by muscle weakness induced by autoantibodies against the acetylcholine receptor. CTLA-4 (CD152) plays an inhibitory role in the immune response and has been suggested to be involved in the pathophysiology of MG. In this study, we focused on alternative CTLA-4 mRNA expression in PBMCs from MG patients. We defined two new isoforms of CTLA-4 mRNA that arise due to alternative splicing. By semi-quantitative RT-PCR analysis, we observed a lower expression of sCTLA-4 mRNA relative to the membrane form in MG patients. In addition, the MG patients had lower levels of sCTLA-4 mRNA in PBMCs compared to healthy controls, as assessed by real-time PCR. One of the spliced isoforms (LCTLA-4) was more prevalent in MG patients compared to healthy controls. The alternative splicing was not associated with sex, thymectomy, serum levels of anti-AChR, immunosuppressive treatment or the four CTLA-4 gene polymorphisms analyzed. This study reveals an abnormal spectrum of mRNA expression of CTLA-4 in MG patients, which marks the importance of studying gene expression of alternative splicing.
Subject(s)
Alternative Splicing , Antigens, CD/genetics , Leukocytes, Mononuclear/immunology , Myasthenia Gravis/genetics , Myasthenia Gravis/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Antigens, CD/blood , CTLA-4 Antigen , Female , Humans , Leukocytes, Mononuclear/metabolism , Male , Middle Aged , Myasthenia Gravis/metabolism , Protein Isoforms/genetics , Protein Isoforms/immunology , RNA, Messenger/genetics , RNA, Messenger/metabolismABSTRACT
Inhibitory regulatory functions of B7-H3 and B7-H4 regarding T-cell activation have been reported recently. Little is known about the significance of human B7-H3 and B7-H4 expression in non-small-cell lung cancer (NSCLC), and we conducted the present study to address this issue in cell lines and tumor tissue from 70 patients. B7-H3 is over-expressed by all six NSCLC cell lines on both mRNA and protein level. B7-H4 is only transcripted by one cell line in which an alternatively spliced form was discovered. In tumor tissues, expression of B7-H3 and B7-H4 was found both on the cell membrane and in the cytoplasm. Thirty-seven percent of the specimens expressed B7-H3 and 43% B7-H4. The number of T infiltrating lymphoid cells (TILs) in the tumor tissues that expressed B7-H3 or B7-H4 was much lower than those who did not. There was a significant positive relation between the expression of B7-H3 and B7-H4, and high B7-H3 or B7-H4 expression was significantly more common in cases with lymph node metastasis. These observations suggest the contribution of B7-H3 and B7-H4 to immune dysfunction and tumor progression in NSCLC patients. B7-H3 and B7-H4 may be an important target for diagnosis and/or therapy of NSCLC.
Subject(s)
B7-1 Antigen/biosynthesis , Carcinoma, Non-Small-Cell Lung/metabolism , Carcinoma, Non-Small-Cell Lung/pathology , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Adenocarcinoma/metabolism , Adenocarcinoma/pathology , Alternative Splicing , Antigens, CD , B7 Antigens , Carcinoma, Squamous Cell/metabolism , Carcinoma, Squamous Cell/pathology , Cell Line, Tumor , Female , Flow Cytometry , Humans , Immunohistochemistry , Leukocyte Common Antigens/biosynthesis , Lymphocytes, Tumor-Infiltrating/metabolism , Male , Middle Aged , RNA, Messenger/metabolism , Receptors, Immunologic , Reverse Transcriptase Polymerase Chain Reaction , V-Set Domain-Containing T-Cell Activation Inhibitor 1Subject(s)
Antigens, CD/genetics , Myasthenia Gravis/genetics , Polymorphism, Single Nucleotide/genetics , Receptors, Immunologic/genetics , Alleles , B7 Antigens , Case-Control Studies , Chi-Square Distribution , Electrophoresis, Agar Gel , Ethidium/metabolism , Female , Gene Frequency , Humans , Male , Polymerase Chain Reaction , Promoter Regions, GeneticABSTRACT
Cytolytic T lymphocyte-associated antigen-4 (CTLA-4) plays a critical role in the down-regulation of antigen-activated immune responses. The aberrant CTLA-4 expression is characterized by low surface and intracellular levels of CTLA-4 protein, impaired up-regulation of CTLA-4 in T cells in response to ConA stimulation and high levels of soluble CTLA-4 (sCTLA-4) in serum. The serum levels of sCTLA-4 are positively correlated with the serum concentration of antibodies against the acetylcholine receptor. The (AT)(n) polymorphism in the 3'-untranslated region contributes to decreased mRNA stability and, hence, to reduced expression of CTLA-4.
Subject(s)
AT Rich Sequence/genetics , Antigens, Differentiation/blood , Antigens, Differentiation/genetics , Immunoconjugates , Myasthenia Gravis/blood , Myasthenia Gravis/genetics , Polymorphism, Genetic/genetics , T-Lymphocytes/immunology , Up-Regulation/genetics , Abatacept , Adult , Alleles , Antigens, CD , Antigens, Differentiation/immunology , CD28 Antigens/genetics , CD28 Antigens/immunology , CD3 Complex/immunology , CTLA-4 Antigen , Cells, Cultured , Concanavalin A , DNA/analysis , DNA/genetics , Female , Gene Expression Regulation/immunology , HLA-DR Antigens/genetics , HLA-DR Antigens/immunology , Humans , Immunosuppressive Agents/pharmacology , Male , Middle Aged , Myasthenia Gravis/immunology , RNA, Messenger/metabolism , Receptors, Nicotinic/immunology , T-Lymphocytes/cytology , Up-Regulation/immunologyABSTRACT
OBJECTIVE: To analyze the association of Wegener's granulomatosis (WG) with 2 single nucleotide polymorphisms (SNP), a +49 A/G polymorphism in coding sequence (CDS) 1 and a C/T base exchange in the promoter region at position -318. METHODS: Restriction enzyme digestion of PCR amplified genomic DNA was used to analyze the CTLA-4 SNP in 32 patients with WG and 100-122 ethnically matched healthy controls. RESULTS: Patients were more often heterozygous for C/T in the promoter region (31% of the patients vs 14% of controls; p < 0.05). Homozygosity for C was less frequent in patients (69% of patients vs 86% of controls; p < 0.05). There was no association with the A/G SNP in CDS 1. There was a linkage disequilibrium between allele A of CDS 1 and the shortest allele, 86 bp, in the (AT)n of the 3' untranslated region in controls but not in patients. CONCLUSION: The CTLA-4 SNP in the promoter region at position -318 is associated with WG. The loss of linkage disequilibrium between allele A of CDS 1 and the short 86 bp in the (AT)n in patients indicates that the promoter SNP and the (AT)n polymorphism are independent genetic risk factors.
Subject(s)
Antigens, Differentiation/genetics , Granulomatosis with Polyangiitis/genetics , Immunoconjugates , Polymorphism, Single Nucleotide , Abatacept , Adult , Aged , Aged, 80 and over , Antigens, CD , CTLA-4 Antigen , Genetic Predisposition to Disease/epidemiology , Granulomatosis with Polyangiitis/epidemiology , Homozygote , Humans , Linkage Disequilibrium , Middle Aged , Promoter Regions, Genetic/genetics , Risk FactorsABSTRACT
OBJECTIVE: Wegener's granulomatosis (WG) is a necrotizing vasculitis characterized by clonal expansions of T cells and production of antibodies against proteinase 3. The disease is associated with expanded dinucleotide repeats in the cytotoxic T lymphocyte antigen 4 (CTLA-4) gene, suggesting that genetic variation(s) in T cell related gene(s) could contribute to the T cell hyperactivity in WG. We investigated the polymorphisms in the genes of 2 cytokines, interleukin 4 (IL-4) and IL-10, which are essential for the polarization of T cells towards Th2 development and for the Ig production by B cells. METHODS: Polymorphisms in the genes coding for IL-10 and IL-4 were analyzed in 32-36 Swedish Caucasian patients and 109 ethnically matched healthy individuals. RESULTS: There was no association with the IL-4 gene. A CA repeat polymorphism in IL-10 gene, IL-10.G, was associated with the disease. This polymorphism has earlier been associated with high autoantibody production. CONCLUSION: Our results indicate that the IL-10 gene may influence the disease, perhaps by influencing the production of autoantibodies.
Subject(s)
Granulomatosis with Polyangiitis/genetics , Interleukin-10/genetics , Interleukin-4/genetics , Adult , Aged , Aged, 80 and over , DNA/analysis , Female , Gene Frequency , Genotype , Humans , Male , Microsatellite Repeats , Middle Aged , Polymorphism, GeneticABSTRACT
En el presente artículo se reportan las características del reconocimiento del anticuerpo monoclonal (AcM) IOR-T2 sobre células mononucleares normales activadas con Concanavalina A (Con A), células tímicas y líneas celulares de cultivo mediante la técnica de inmunofluorescencia indirecta; además, se realizó la determinación de la subclase de inmunoglobulina y del punto isoeléctrico de este AcM. Los resultados muestran que el AcM IOR-T2 es una IgG2b que no reconoce a los antígenos que se expresan durante el proceso de estimulación con Con A de las células linfoides normales. A diferencia de lo observado en las células mononucleares de la sangre periférica normal, el AcM IOR-T2 reconoce el 59 ñ 3 % de los timocitos fetales y el 16 ñ 3 % de las células de los timos pediátricos. El AcM IOR-T2 parece identificar a un antígeno de diferenciación de las células tímicas, el cual se expresa en un alto porcentaje de las células tumorales de la sangre periférica de pacientes con linfomas T cutáneos
Subject(s)
Humans , Antibodies, Monoclonal/immunology , Concanavalin A , Immunoglobulin G/isolation & purification , Thymus Gland/cytology , Lymphoma , Skin NeoplasmsABSTRACT
En el presente artículo se reportan las características del reconocimiento del anticuerpo monoclonal (AcM) IOR-T2 sobre células mononucleares normales activadas con Concanavalina A (Con A), células tímicas y líneas celulares de cultivo mediante la técnica de inmunofluorescencia indirecta; además, se realizó la determinación de la subclase de inmunoglobulina y del punto isoeléctrico de este AcM. Los resultados muestran que el AcM IOR-T2 es una IgG2b que no reconoce a los antígenos que se expresan durante el proceso de estimulación con Con A de las células linfoides normales. A diferencia de lo observado en las células mononucleares de la sangre periférica normal, el AcM IOR-T2 reconoce el 59 ñ 3
de los timocitos fetales y el 16 ñ 3
de las células de los timos pediátricos. El AcM IOR-T2 parece identificar a un antígeno de diferenciación de las células tímicas, el cual se expresa en un alto porcentaje de las células tumorales de la sangre periférica de pacientes con linfomas T cutáneos
