RESUMO
It is well known that monocytes may play an active role in thrombogenesis, since they may express on their surface tissue factor, the major initiator of the clotting cascade. The results of this investigation demonstrate beta-2-glycoprotein I (beta2-GPI) mRNA expression by human peripheral blood monocytes, indicating that these cells synthesize beta2-GPI. In addition, we show beta2-GPI expression on cell surface of these cells by flow cytometric analysis, and the presence of this protein in cell lysate by Western blot. Interestingly, beta2-GPI expression on monocytes is significantly increased in patients with anti-phospholipid syndrome (APS) or systemic lupus erythematosus (SLE) as against healthy blood donors and correlates with tissue factor expression on monocytes. These findings support the view that monocytes are able to synthesize beta2-GPI and suggest that patients with APS may have increased beta2-GPI exposure on cell surface, which leads to persistently high monocyte tissue factor expression and consequently to a prothrombotic diathesis.
Assuntos
Síndrome Antifosfolipídica/sangue , Glicoproteínas/sangue , Monócitos/metabolismo , Tromboplastina/metabolismo , Adolescente , Adulto , Síndrome Antifosfolipídica/imunologia , Autoanticorpos/sangue , Western Blotting , Criança , Fragmentação do DNA , Feminino , Citometria de Fluxo , Técnica Indireta de Fluorescência para Anticorpo , Expressão Gênica , Glicoproteínas/genética , Humanos , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/imunologia , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , beta 2-Glicoproteína IRESUMO
There is increasing interest in the identification of biological markers for the early diagnosis of Parkinson's disease (PD). Previous studies indicate changes of dopamine content, tyrosine hydroxylase immunoreactivity and dopamine receptors in peripheral blood lymphocytes (PBL) in PD. Here we demonstrate a reduction of dopamine transporter immunoreactivity in PBL in the early clinical stages of the disease. These findings contribute to our understanding of the peripheral dopamine system in PD.
Assuntos
Dopamina/sangue , Linfócitos/imunologia , Glicoproteínas de Membrana , Proteínas de Membrana Transportadoras/imunologia , Proteínas do Tecido Nervoso , Doença de Parkinson/imunologia , Análise de Variância , Biomarcadores/sangue , Proteínas da Membrana Plasmática de Transporte de Dopamina , Humanos , Linfócitos/sangue , Proteínas de Membrana Transportadoras/sangue , Pessoa de Meia-Idade , Doença de Parkinson/sangueRESUMO
The early clinical symptoms of Parkinson's disease (PD) may be difficult to perceive and are frequently overlooked. Thus, interest has focused on the identification of biological or instrumental markers that may contribute to the early diagnosis of PD, with the aim of introducing neuroprotective therapies at the very start of illness. Impairment of nigrostriatal dopamine transmission can be visualized in vivo by functional imaging techniques, but these are rather complex and expensive examinations, available only in selected institutions. Here we show that dopamine content and tyrosine hydroxylase immunoreactivity are reduced in peripheral blood lymphocytes (PBL) in the early stages of PD. These data suggest that PBL may represent a simple and useful tool with which to identify precociously dopamine impairment in PD.
Assuntos
Dopamina/sangue , Linfócitos/metabolismo , Doença de Parkinson/metabolismo , Idoso , Feminino , Humanos , Imuno-Histoquímica , Linfócitos/enzimologia , Masculino , Pessoa de Meia-Idade , Doença de Parkinson/sangue , Doença de Parkinson/enzimologia , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
1. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to investigate the effects of haloperidol treatment (1 mg/kg/day for 2, 7 or 21 consecutive days) on the expression of D1B and D3 dopamine receptor mRNAs in the rat lymphocytes. 2. The expression of D1B receptor mRNA was significantly decreased after 2 days of treatment and progressively returned toward basal values at the end of treatment. Conversely, haloperidol failed to modify the expression of lymphocyte D3 receptor mRNA. 3. These results indicate short-lasting dynamic changes of expression of lymphocyte D1B dopamine receptor mRNA by haloperidol and suggest that the effects of dopamine and dopaminergic drugs on the immune system might be mediated, at least in part, by direct interaction of these substances with dopamine receptors on lymphocyte membrane.
Assuntos
Antipsicóticos/farmacologia , Haloperidol/farmacologia , Linfócitos/metabolismo , RNA Mensageiro/biossíntese , Receptores Dopaminérgicos/biossíntese , Animais , Catalepsia/induzido quimicamente , Primers do DNA , Linfócitos/efeitos dos fármacos , Masculino , Ratos , Ratos Sprague-Dawley , Receptores de Dopamina D1/biossíntese , Receptores de Dopamina D2/biossíntese , Receptores de Dopamina D3 , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de TempoRESUMO
Dopamine D1-like and D2-like receptors on peripheral blood lymphocytes (PBL) were assayed in 50 de novo patients with idiopathic Parkinson's disease (PD), in 36 neurologic control subjects (multiple-system atrophy, n = 16; essential tremor, n = 10; other neurodegenerative diseases, n = 10), and in 26 healthy control subjects by radioligand binding assay techniques using [3H]SCH 23390 and [3H]7OH-DPAT as ligands. Patients with PD revealed a higher density (Bmax) of dopamine D1-like (p <0.001) and D2-like (p <0.00001) receptors on PBL than either neurologic or healthy control subjects, whereas no differences in Bmax were observed among patients affected by other neurologic diseases and healthy control subjects. The affinity (Kd) of both radioligands was similar in the groups investigated. The pharmacologic profile of [3H]SCH 23390 and [3H]7OH-DPAT binding was consistent with the labeling of dopamine D5 and D3 receptor subtypes, respectively. Twenty-five of the 50 patients with PD were retested after 3 months of therapy with levodopa or bromocriptine. Both treatments reduced the density of D1-like (p <0.001) and D2-like (p <0.001) receptors on PBL to values comparable to those of control subjects. The increased density of D1-like and D2-like receptors on PBL in de novo PD patients may represent an upregulation mechanism resulting from the diffuse impairment of the dopaminergic system in PD.
Assuntos
Linfócitos/metabolismo , Doença de Parkinson/metabolismo , Receptores Dopaminérgicos/metabolismo , Idoso , Antiparkinsonianos/uso terapêutico , Ligação Competitiva/fisiologia , Encéfalo/patologia , Contagem de Células , Agonistas de Dopamina/farmacocinética , Feminino , Humanos , Levodopa/uso terapêutico , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Doença de Parkinson/diagnóstico , Doença de Parkinson/tratamento farmacológico , Ensaio Radioligante , Sensibilidade e Especificidade , Resultado do TratamentoRESUMO
We report here the expression of beta2-GPI mRNA by cell types involved in the pathophysiology of the anti-phospholipid syndrome (APS), i.e. endothelial cells as a target of autoantibodies in the APS, astrocytes and neurones involved in APS of the central nervous system (CNS). Lymphocytes were also included in the study, as it has been demonstrated that patients with systemic lupus erythematosus-associated CNS diseases have serum anti-lymphocyte antibodies cross-reacting with brain antigens, and intrathecally synthesized anti-neurone antibodies. Reverse transcriptase-polymerase chain reaction followed by restriction enzyme digestion of the product obtained demonstrated the presence of beta2-GPI mRNA in all cell types here tested, cultured both in presence and absence of fetal calf serum. In both culture conditions, the same cell types were immunoreactive to an anti-beta2-GPI MoAb, as determined by indirect immunofluorescence technique. Taken together, these results indicate a direct cell synthesis of beta2-GPI, suggesting an antigenic function of beta2-GPI in the APS, including the CNS disease that occurs in this syndrome.
Assuntos
Síndrome Antifosfolipídica/genética , Síndrome Antifosfolipídica/patologia , Glicoproteínas/genética , Actinas/genética , Linhagem Celular/metabolismo , Meios de Cultivo Condicionados , Expressão Gênica , Humanos , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais Cultivadas/metabolismo , beta 2-Glicoproteína IRESUMO
This study was performed on a family of CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leucoencephalopathy) subjects. Neuropathological alterations of small arteries consisting in thickening, reduplication and fragmentation of the internal elastic lamella, and granular periodic acid-Schiff-positive material deposited in the arterial media were demonstrated in 1 autopsy case by histochemistry and electron microscopy. This material reacted with a monoclonal antibody anti-elastin (aE), as demonstrated by immunohistochemistry and immunoelectron microscopy. Significant increases of aE-immunoreactivity and elastin mRNA expression were found in cultured skin fibroblasts from 5 family members genetically affected by CADASIL, but not genetically and clinically healthy members. These results suggest that alterations of the elastic apparatus are associated with CADASIL genotype and related to the clinical expression of the disease.
Assuntos
Encéfalo/patologia , Doenças Arteriais Cerebrais/patologia , Infarto Cerebral/patologia , Elastina/análise , Leucoencefalopatia Multifocal Progressiva/patologia , Pele/patologia , Adulto , Análise de Variância , Biópsia por Agulha , Células Cultivadas/metabolismo , Doenças Arteriais Cerebrais/genética , Doenças Arteriais Cerebrais/metabolismo , Artérias Cerebrais/ultraestrutura , Infarto Cerebral/genética , Infarto Cerebral/metabolismo , Colágeno/ultraestrutura , Elastina/biossíntese , Elastina/genética , Feminino , Fibroblastos/metabolismo , Fibronectinas/análise , Seguimentos , Humanos , Imuno-Histoquímica , Ataque Isquêmico Transitório/genética , Ataque Isquêmico Transitório/metabolismo , Ataque Isquêmico Transitório/patologia , Leucoencefalopatia Multifocal Progressiva/genética , Leucoencefalopatia Multifocal Progressiva/metabolismo , Masculino , Microscopia Eletrônica , Pessoa de Meia-Idade , RNA Mensageiro/análise , Valores de Referência , Pele/metabolismo , SíndromeRESUMO
Sera from 20 patients with antiphospholipid syndrome (APS), primary or secondary to systemic lupus erythematosus (SLE), or with SLE, were assayed by immunoblot analysis for anti-beta2-glycoprotein I antibodies (abeta2-GPI), and by indirect immunofluorescence (IIF) technique for reactivity with astrocyte and neuron cell lines and with histological sections of human brain biopsies and monkey cerebellum. Six sera from healthy donors were studied as a control. Eleven out of the 20 patient sera contained abeta2-GPI and were immunoreactive with astrocytes and neurons, both in culture and in the histological sections, and with the endotheliocytes of the microvessels present in the histological sections. Cell localization and the pattern of immune reaction were similar to those obtained with a monoclonal antibody abeta2-GPI. Eight of the remaining patient sera, found abeta2-GPI-, did not react with the nervous substrates (and the control sera), while one exhibited immunoreactivity analogous to the abeta2-GPI+ sera. The interference of anticardiolipin antibodies (aCL) in the immunoreactivity with the nervous substrates was excluded since aCL were present in all patient sera and no immune reaction was observed in the histological sections incubated with a monoclonal aCL. Therefore, the binding of abeta2-GPI from patients to cells of the central nervous system (CNS) occurs independently from aCL. This issue may be relevant to further evaluate the potential pathogenetic role of abeta2-GPI in the CNS damage of APS-like conditions.
Assuntos
Síndrome Antifosfolipídica/imunologia , Autoanticorpos/sangue , Sistema Nervoso Central/imunologia , Glicoproteínas/imunologia , Animais , Anticorpos Anticardiolipina/sangue , Síndrome Antifosfolipídica/complicações , Astrócitos/imunologia , Estudos de Casos e Controles , Linhagem Celular , Endotélio Vascular/imunologia , Haplorrinos , Humanos , Imuno-Histoquímica , Técnicas In Vitro , Lúpus Eritematoso Sistêmico/complicações , Lúpus Eritematoso Sistêmico/imunologia , Neurônios/imunologia , beta 2-Glicoproteína IRESUMO
Glial mitogenic effect was investigated in sera from the following groups of subjects: group (1) 31 patients clinically and genetically affected by Neurofibromatosis type 1 (NF1) belonging to different families; group (2) 42 patients without family history of NF1 affected by sporadic neoplasms of the same histogenetic origin as the proliferative lesions that are present in NF1; group (3) 51 healthy volunteers without family history of NF1 nor of neoplastic disease; group (4) 54 clinically healthy relatives of the NF1 patients included in the first group. All NF1 patients and 3/54 healthy relatives had alterations of exons 31 or 32 of NF1 gene. Glial proliferation, measured by [3H]thymidine incorporation, was significantly increased by sera from all NF1 patients and from 23/54 of clinically healthy relatives, as compared to sera from healthy volunteers. This serum mitogenic activity strongly suggests the existence of soluble glial proliferating molecules in NF1 families. The molecular weight (3-30 kDa), the heat- and freeze-stability and the specificity for glial cells, suggest that the molecules responsible for this mitogenic effect are different from the growth factors previously described in NF1-associated tumor extracts and from lymphokines. Within each NF1 family, the maximal serum dilution stimulating glial proliferation was similar both in affected members and in their clinically healthy relatives. Since none of the clinically healthy relatives showing serum mitogenic activity was positive for the NF1 mutation analysis and, conversely, those having altered exons 31 or 32 of NF1 gene did not show any mitogenic activity; these results suggest that the phenotype expression of NF1 might depend not only on the NF1 mutations per se, but also on other genetic or epigenetic factors, such as serum glial proliferating molecules.
Assuntos
Mitógenos/sangue , Mitógenos/fisiologia , Mitose/fisiologia , Neurofibromatose 1/sangue , Neurofibromatose 1/patologia , Neuroglia/patologia , Animais , Relação Dose-Resposta a Droga , Glioma , Humanos , Linfócitos , Mitose/efeitos dos fármacos , Peso Molecular , Neurofibromatose 1/genética , Neuroglia/efeitos dos fármacos , Ratos , Timidina/análise , Timidina/metabolismo , Trítio , Células Tumorais CultivadasRESUMO
It has been suggested that dopamine might play a role in the regulation of the immune system. In this study, reverse transcriptase-polymerase chain reaction (RT-PCR) was used to investigate the expression of mRNA for the different subtypes of dopamine receptors in the rat lymphocytes. D1, D3 and D5 receptor mRNAs were identified. These results provide further evidence for the interaction of dopamine systems and the immune system, and suggest to further investigate whether the immunosuppressive actions of dopamine and dopaminergic drugs might depend on a direct interaction with dopamine receptors on the lymphocyte membrane. Moreover, they suggest the suitability of this animal species to further investigate the correlation between changes in the expression of central and peripheral dopamine receptors produced by manipulations of the dopamine systems.
Assuntos
Linfócitos/química , RNA Mensageiro/análise , Receptores Dopaminérgicos/genética , Animais , Corpo Estriado/química , Masculino , Reação em Cadeia da Polimerase , DNA Polimerase Dirigida por RNA , Ratos , Ratos Sprague-Dawley , Receptores de Dopamina D1/genética , Receptores de Dopamina D2/genética , Receptores de Dopamina D3 , Receptores de Dopamina D4 , Receptores de Dopamina D5RESUMO
The effects of testosterone (T), dihydrotestosterone (DHT) and methyltrienolone (R 1881) on cell proliferation of eight human pituitary tumors in culture wre assessed by [3H]thymidine incorporation and compared to those of progesterone (Pg) and 17 beta-estradiol. Receptors for androgens (AR), estrogens (ER) and progesterone (PgR) were characterized. AR had a significant inhibitory effect on all AR-positive tumors, whatever their hormonal content. Inhibitory effects of either T and DHT < R1881 < Pg were observed in tumors co-expressing AR and PgR. The inhibitory effect of R 1881 on a PgR-positive/AR-negative tumor suggested that R 1881 action was partially PgR-mediated. The effects of either T or the nonaromatizable DHT and R 1881 were unrelated to ER expression. We conclude that AR can modulate the growth of human pituitary tumors through direct receptor-mediated intracellular pathways which may be common to various pituitary cell types.