RESUMO
Parkinson's disease (PD) is one of the most common neurodegenerative diseases worldwide. Currently applied therapeutic protocols are limited to improve the motor functions of patients. Therefore, seeking alternative regimes with better therapeutic impact is crucial. This study aims to validate the therapeutic impact of mesenchymal stem cell injection using two delivery methods, intracranial administration and intravenous administration, on rotenone (ROT)-induced PD model in rats. Our work included behavioral, biochemical, histological, and molecular investigations. Open field test (OFT) and rotarod tests were applied. Important oxidative stress, antioxidant and proinflammatory markers were monitored. Substantia Nigra and Striatum tissues were examined histologically and the molecular expression of DOPA decarboxylase, Tyrosine hydroxylase, and α-synuclein in neurons in these tissues were investigated. Our results showed that MSC grafting improved motor and memory impairments and oxidative stress status that were observed after ROT administration. Additionally, BM-MSCs application restored SOD and CAT activities and the levels of DA, L-Dopa, IL6, IL1ß, and TNFα. Moreover, MSC grafting overwhelmed the pathological changes induced by ROT and normalized the expression of Tyrosine hydroxylase, DOPA decarboxylase, and α-synuclein towards the control values in the Nigral and Striatal tissues of male rats. Conclusively, both administration routes improved motor function, protection of the nigrostriatal system, and improved striatal dopamine release. The observed beneficial effect of applying MSCs suggests potential benefits in clinical applications. No significant differences in the outcomes of the treatment would favor a certain way of MSC application over the other. However, the intravenous delivery method seems to be safer and more feasible compared to the intrastriatal method.
Assuntos
Células-Tronco Mesenquimais , Doença de Parkinson , Transtornos Parkinsonianos , Humanos , Ratos , Masculino , Animais , alfa-Sinucleína/metabolismo , Transtornos Parkinsonianos/terapia , Transtornos Parkinsonianos/tratamento farmacológico , Doença de Parkinson/metabolismo , Substância Negra/metabolismo , Rotenona/farmacologia , Dopa Descarboxilase/metabolismo , Células-Tronco Mesenquimais/metabolismo , Administração Intravenosa , Modelos Animais de DoençasRESUMO
Object Exclusively dopamine-producing pheochromocytoma/paraganglioma (PPGL) is an extremely rare subtype. In this condition, intratumoral dopamine ß-hydroxylase (DBH), which controls the conversion of norepinephrine from dopamine, is impaired, resulting in suppressed norepinephrine and epinephrine production. However, the rarity of this type of PPGL hampers the understanding of its pathophysiology. We therefore conducted genetic and immunohistological analyses of a patient with an exclusively dopamine-producing paraganglioma. Methods Paraganglioma samples from a 52-year-old woman who presented with a 29.6- and 41.5-fold increase in plasma and 24-h urinary dopamine, respectively, but only a minor elevation in the plasma norepinephrine level was subjected to immunohistological and gene expression analyses of catecholamine synthases. Three tumors carrying known somatic PPGL-related gene variants (HRAS, EPAS1) were used as controls. Whole-exome sequencing (WES) was also performed using the patient's blood and tumor tissue. Results Surprisingly, the protein expression of DBH was not suppressed, and its mRNA expression was clearly higher in the patient than in the controls. Furthermore, dopa decarboxylase (DDC), which governs the conversion of 3,4-dihydroxyphenyl-L-alanine (L-DOPA) to dopamine, was downregulated at the protein and gene levels. In addition, melanin, which is synthesized by L-DOPA, accumulated in the tumor. WES revealed no PPGL-associated pathogenic germline variants, but a missense somatic variant (c.1798G>T) in CSDE1 was identified. Conclusion Although pre-operative plasma L-DOPA was not measured, our histological and gene expression analyses suggest that L-DOPA, rather than dopamine, might have been overproduced in the tumor. This raises the possibility of pathophysiological heterogeneity in exclusively dopamine-producing PPGL.
Assuntos
Neoplasias das Glândulas Suprarrenais , Paraganglioma , Feocromocitoma , Feminino , Humanos , Pessoa de Meia-Idade , Dopamina/metabolismo , Dopa Descarboxilase/genética , Dopa Descarboxilase/metabolismo , Melaninas/genética , Melaninas/metabolismo , Dopamina beta-Hidroxilase/genética , Dopamina beta-Hidroxilase/metabolismo , Regulação para Cima , Paraganglioma/genética , Norepinefrina , Feocromocitoma/genética , Levodopa , Neoplasias das Glândulas Suprarrenais/genética , Neoplasias das Glândulas Suprarrenais/patologia , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a RNARESUMO
L-Dopa decarboxylase (DDC) is the most significantly co-expressed gene with ACE2, which encodes for the SARS-CoV-2 receptor angiotensin-converting enzyme 2 and the interferon-inducible truncated isoform dACE2. Our group previously showed the importance of DDC in viral infections. We hereby aimed to investigate DDC expression in COVID-19 patients and cultured SARS-CoV-2-infected cells, also in association with ACE2 and dACE2. We concurrently evaluated the expression of the viral infection- and interferon-stimulated gene ISG56 and the immune-modulatory, hypoxia-regulated gene EPO. Viral load and mRNA levels of DDC, ACE2, dACE2, ISG56 and EPO were quantified by RT-qPCR in nasopharyngeal swab samples from COVID-19 patients, showing no or mild symptoms, and from non-infected individuals. Samples from influenza-infected patients were analyzed in comparison. SARS-CoV-2-mediated effects in host gene expression were validated in cultured virus-permissive epithelial cells. We found substantially higher gene expression of DDC in COVID-19 patients (7.6-fold; p = 1.2e-13) but not in influenza-infected ones, compared to non-infected subjects. dACE2 was more elevated (2.9-fold; p = 1.02e-16) than ACE2 (1.7-fold; p = 0.0005) in SARS-CoV-2-infected individuals. ISG56 (2.5-fold; p = 3.01e-6) and EPO (2.6-fold; p = 2.1e-13) were also increased. Detected differences were not attributed to enrichment of specific cell populations in nasopharyngeal tissue. While SARS-CoV-2 virus load was positively associated with ACE2 expression (r≥0.8, p<0.001), it negatively correlated with DDC, dACE2 (r≤-0.7, p<0.001) and EPO (r≤-0.5, p<0.05). Moreover, a statistically significant correlation between DDC and dACE2 expression was observed in nasopharyngeal swab and whole blood samples of both COVID-19 and non-infected individuals (r≥0.7). In VeroE6 cells, SARS-CoV-2 negatively affected DDC, ACE2, dACE2 and EPO mRNA levels, and induced cell death, while ISG56 was enhanced at early hours post-infection. Thus, the regulation of DDC, dACE2 and EPO expression in the SARS-CoV-2-infected nasopharyngeal tissue is possibly related with an orchestrated antiviral response of the infected host as the virus suppresses these genes to favor its propagation.
Assuntos
Enzima de Conversão de Angiotensina 2/metabolismo , COVID-19/patologia , Dopa Descarboxilase/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adulto , Idoso , Enzima de Conversão de Angiotensina 2/genética , Área Sob a Curva , Descarboxilases de Aminoácido-L-Aromático , COVID-19/virologia , Dopa Descarboxilase/genética , Regulação para Baixo , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Eritropoetina/genética , Eritropoetina/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Nasofaringe/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Curva ROC , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Regulação para Cima , Carga ViralRESUMO
Pheochromocytomas (PHEOs) are relatively rare catecholamine-producing tumors derived from adrenal medulla. Tumor microenvironment (TME) including neoangiogenesis has been explored in many human neoplasms but not necessarily in PHEOs. Therefore, in this study, we examined tumor infiltrating lymphocytes (CD4 and CD8), tumor associated macrophages (CD68 and CD163), sustentacular cells (S100p), and angiogenic markers (CD31 and areas of intratumoral hemorrhage) in 39 cases of PHEOs in the quantitative fashion. We then compared the results with pheochromocytoma of the adrenal gland scaled score (PASS), grading system for pheochromocytoma and paraganglioma (GAPP) and the status of intra-tumoral catecholamine-synthesizing enzymes (TH, DDC, and PNMT) as well as their clinicopathological factors. Intratumoral CD8 (p = 0.0256), CD31 (p = 0.0400), and PNMT (p = 0.0498) status was significantly higher in PHEOs with PASS <4 than PASS â§4. In addition, intratumoral CD8+ lymphocytes were also significantly more abundant in well-than moderately differentiated PHEO according to GAPP score (p = 0.0108) and inversely correlated with tumor size (p = 0.0257). Intratumoral CD68+ cells were significantly higher in PHEOs with regular or normal histological patterns than those not (p = 0.0370) and inversely correlated with tumor size (p = 0.0457). The status of CD163 was significantly positively correlated with that of CD8 positive cells (p = 0.0032). The proportion of intratumoral hemorrhage areas was significantly higher in PHEOs with PASS â§4 (p = 0.0172). DDC immunoreactivity in tumor cells was significantly positively correlated with PASS score (p = 0.0356) and TH status was significantly higher in PHEOs harboring normal histological patterns (p = 0.0236) and cellular monotony (p = 0.0219) than those not. Results of our present study did demonstrate that abundant CD8+ and CD68+ cells could represent a histologically low-scored tumor. In particular, PHEOs with increased intratumoral hemorrhage should be considered rather malignant. In addition, abnormal catecholamine-producing status of tumor cells such as deficient PNMT and TH and increased DDC could also represent more aggressive PHEOs.
Assuntos
Neoplasias das Glândulas Suprarrenais/irrigação sanguínea , Neoplasias das Glândulas Suprarrenais/imunologia , Neovascularização Patológica , Feocromocitoma/irrigação sanguínea , Feocromocitoma/imunologia , Microambiente Tumoral/imunologia , Neoplasias das Glândulas Suprarrenais/epidemiologia , Neoplasias das Glândulas Suprarrenais/patologia , Adulto , Idoso , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Biomarcadores Tumorais/análise , Linfócitos T CD8-Positivos/imunologia , Catecolaminas/metabolismo , Dopa Descarboxilase/metabolismo , Feminino , Hemorragia , Humanos , Imuno-Histoquímica , Japão/epidemiologia , Linfócitos do Interstício Tumoral/imunologia , Masculino , Pessoa de Meia-Idade , Feocromocitoma/epidemiologia , Feocromocitoma/patologia , Macrófagos Associados a Tumor/imunologia , Tirosina 3-Mono-Oxigenase/deficiênciaRESUMO
Mn3O4 nanoparticles (NPs) are used increasingly in various fields due to their excellent physiochemical properties. Previous studies have documented that Mn-based nanomaterials resulted in excess reactive oxygen species (ROS) generation and dopamine (DA) reduction both in vivo and in vitro experiments. However, little is known about the mechanism of ROS production and DA decrease induced by Mn-based nanomaterials. The present study was carried out to elucidate the mechanism of the co-incubation model of dopaminergic neuron PC12â¯cells and the synthesized Mn3O4 NPs. The results demonstrated that exposure to Mn3O4 NPs reduced cell viability, increased level of lactate dehydrogenase (LDH), triggered oxidative stress and induced apoptosis. Notably, the level of ROS was remarkably increased (>10-fold) with Mn3O4 NPs exposure. We also found that mitochondrial calcium Ca2+ uniporter (MCU) was up-regulated and the mitochondrial Ca2+ concentration ([Ca2+]mito) increased induced by Mn3O4 NPs in PC12â¯cells. Furthermore, the MCU inhibitor RuR significantly attenuated Mn3O4 NPs-induced [Ca2+]mito, ROS production and apoptosis. In PC12â¯cells, the decrease of DA content was mainly due to the downregulation of DOPA decarboxylase (DDC) expression caused by Mn3O4 NPs treatment. The expression of proteins related to DA storage system was not significantly affected by treatment.
Assuntos
Apoptose/efeitos dos fármacos , Neurônios Dopaminérgicos/efeitos dos fármacos , Nanopartículas Metálicas/toxicidade , Óxidos/toxicidade , Animais , Canais de Cálcio/genética , Canais de Cálcio/metabolismo , Sobrevivência Celular , Dopa Descarboxilase/genética , Dopa Descarboxilase/metabolismo , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/patologia , Compostos de Manganês/química , Nanopartículas Metálicas/química , Estresse Oxidativo/efeitos dos fármacos , Óxidos/química , Células PC12 , Ratos , Espécies Reativas de Oxigênio/metabolismoRESUMO
Tissue fibrosis is characterized by uncontrolled deposition and diminished clearance of fibrous connective tissue proteins, ultimately leading to organ scarring. Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ) have recently emerged as pivotal drivers of mesenchymal cell activation in human fibrosis. Therapeutic strategies inhibiting YAP and TAZ have been hindered by the critical role that these proteins play in regeneration and homeostasis in different cell types. Here, we find that the Gαs-coupled dopamine receptor D1 (DRD1) is preferentially expressed in lung and liver mesenchymal cells relative to other resident cells of these organs. Agonism of DRD1 selectively inhibits YAP/TAZ function in mesenchymal cells and shifts their phenotype from profibrotic to fibrosis resolving, reversing in vitro extracellular matrix stiffening and in vivo tissue fibrosis in mouse models. Aromatic l-amino acid decarboxylase [DOPA decarboxylase (DDC)], the enzyme responsible for the final step in biosynthesis of dopamine, is decreased in the lungs of subjects with idiopathic pulmonary fibrosis, and its expression inversely correlates with disease severity, consistent with an endogenous protective role for dopamine signaling that is lost in pulmonary fibrosis. Together, these findings establish a pharmacologically tractable and cell-selective approach to targeting YAP/TAZ via DRD1 that reverses fibrosis in mice.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas de Ciclo Celular/antagonistas & inibidores , Fibroblastos/patologia , Cirrose Hepática/patologia , Fibrose Pulmonar/patologia , Receptores de Dopamina D1/agonistas , Receptores de Dopamina D1/metabolismo , Transativadores/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Bleomicina , Proteínas de Ciclo Celular/metabolismo , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Dopa Descarboxilase/metabolismo , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Feminino , Fibroblastos/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/patologia , Humanos , Pulmão/efeitos dos fármacos , Pulmão/patologia , Lesão Pulmonar/patologia , Masculino , Camundongos Endogâmicos C57BL , Fenantridinas/farmacologia , Fenótipo , Transporte Proteico/efeitos dos fármacos , Interferência de RNA , Transativadores/metabolismo , Proteínas de Sinalização YAPRESUMO
l-dopa decarboxylase (DDC) that catalyzes the biosynthesis of bioactive amines, such as dopamine and serotonin, is expressed in the nervous system and peripheral tissues, including the liver, where its physiological role remains unknown. Recently, we reported a physical and functional interaction of DDC with the major signaling regulator phosphoinosite-3-kinase (PI3K). Here, we provide compelling evidence for the involvement of DDC in viral infections. Studying dengue (DENV) and hepatitis C (HCV) virus infection in hepatocytes and HCV replication in liver samples of infected patients, we observed a negative association between DDC and viral replication. Specifically, replication of both viruses reduced the levels of DDC mRNA and the ~120 kDa SDS-resistant DDC immunoreactive functional complex, concomitant with a PI3K-dependent accumulation of the ~50 kDa DDC monomer. Moreover, viral infection inhibited PI3K-DDC association, while DDC did not colocalize with viral replication sites. DDC overexpression suppressed DENV and HCV RNA replication, while DDC enzymatic inhibition enhanced viral replication and infectivity and affected DENV-induced cell death. Consistently, we observed an inverse correlation between DDC mRNA and HCV RNA levels in liver biopsies from chronically infected patients. These data reveal a novel relationship between DDC and Flaviviridae replication cycle and the role of PI3K in this process.
Assuntos
Dengue/metabolismo , Dopa Descarboxilase/metabolismo , Hepatite C/metabolismo , Fígado/enzimologia , Replicação Viral , Animais , Linhagem Celular Tumoral , Chlorocebus aethiops , Dengue/virologia , Vírus da Dengue/patogenicidade , Vírus da Dengue/fisiologia , Dopa Descarboxilase/genética , Hepacivirus/patogenicidade , Hepacivirus/fisiologia , Hepatite C/virologia , Humanos , Fígado/virologia , Fosfatidilinositol 3-Quinases/metabolismo , Células VeroRESUMO
BACKGROUND: L-Dopa decarboxylase (DDC) expression has been implicated in the biochemistry of several human cancers. Docetaxel and Mitoxantrone are two widely used anticancer agents. Docetaxel is a semi-synthetic analogue of Paclitaxel, an extract from the bark of the rare Pacific yew tree Taxus brevifolia, and Mitoxantrone is an anthracenedione anticancer agent. OBJECTIVE: The purpose of the present study was to investigate the effect of chemotherapeutic agents on the expression of human DDC in human prostate and human breast cancer cell lines. Furthermore, the study focused on the effect of chemotherapeutics - particularly Docetaxel and Mitoxantrone - on the viability of mammalian cells expressing human DDC protein isoforms. METHODS: We investigated the effect of Docetaxel and Mitoxantrone on the expression of DDC in DU- 145 (androgen-independent prostate cancer cell line) and MCF-7 (human breast adenocarcinoma cell line). In order to gain insight into the effect of DDC on cell viability following chemotherapeutic agent treatment, we investigated the cytotoxicity and apoptosis levels on CHO cells expressing different human DDC protein isoforms. RESULTS: Our obtained data indicated that exposure of DU-145 and MCF-7 cells to Docetaxel and Mitoxantrone enhances the expression of neural type DDC mRNA isoforms. Interestingly, DDC protein levels were not affected, despite the cytotoxic events elicited by the chemotherapeutic agent treatment. Moreover, expression of DDC and its alternative protein isoforms, appear to enhance the cytotoxic and apoptotic events conferred by exposure to Docetaxel and Mitoxantrone. CONCLUSION: This study suggests the possible involvement of DDC expression in Docetaxel and Mitoxantrone- induced cytotoxicity and apoptosis.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Docetaxel/farmacologia , Dopa Descarboxilase/metabolismo , Mitoxantrona/farmacologia , Animais , Apoptose/genética , Neoplasias da Mama/enzimologia , Neoplasias da Mama/patologia , Células CHO , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Cricetinae , Cricetulus , Dopa Descarboxilase/genética , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Células MCF-7 , Masculino , Neoplasias da Próstata/enzimologia , Neoplasias da Próstata/patologia , Isoformas de ProteínasRESUMO
BACKGROUND: The identification of a DNA variant in pyridoxal kinase (Pdxk) associated with increased risk to Parkinson disease (PD) gene led us to study the inhibition of this gene in the Dopa decarboxylase (Ddc)-expressing neurons of the well-studied model organism Drosophila melanogaster. The multitude of biological functions attributable to the vitamers catalysed by this kinase reveal an overabundance of possible links to PD, that include dopamine synthesis, antioxidant activity and mitochondrial function. Drosophila possesses a single homologue of Pdxk and we used RNA interference to inhibit the activity of this kinase in the Ddc-Gal4-expressing neurons. We further investigated any association between this enhanced disease risk gene with the established PD model induced by expression of α-synuclein in the same neurons. We relied on the pro-survival functions of Buffy, an anti-apoptotic Bcl-2 homologue, to rescue the Pdxk-induced phenotypes. RESULTS: To drive the expression of Pdxk RNA interference in DA neurons of Drosophila, we used Ddc-Gal4 which drives expression in both dopaminergic and serotonergic neurons, to result in decreased longevity and compromised climbing ability, phenotypes that are strongly associated with Drosophila models of PD. The inhibition of Pdxk in the α-synuclein-induced Drosophila model of PD did not alter longevity and climbing ability of these flies. It has been previously shown that deficiency in vitamers lead to mitochondrial dysfunction and neuronal decay, therefore, co-expression of Pdxk-RNAi with the sole pro-survival Bcl-2 homologue Buffy in the Ddc-Gal4-expressing neurons, resulted in increased survival and a restored climbing ability. In a similar manner, when we inhibited Pdxk in the developing eye using GMR-Gal4, we found that there was a decrease in the number of ommatidia and the disruption of the ommatidial array was more pronounced. When Pdxk was inhibited with the α-synuclein-induced developmental eye defects, the eye phenotypes were unaltered. Interestingly co-expression with Buffy restored ommatidia number and decreased the severity of disruption of the ommatidial array. CONCLUSIONS: Though Pdxk is not a confirmed Parkinson disease gene, the inhibition of this kinase recapitulated the PD-like symptoms of decreased lifespan and loss of locomotor function, possibly producing a new model of PD.
Assuntos
Proteínas de Drosophila/genética , Drosophila/genética , Doença de Parkinson/enzimologia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Piridoxal Quinase/genética , Animais , Modelos Animais de Doenças , Dopa Descarboxilase/genética , Dopa Descarboxilase/metabolismo , Evolução Molecular , Locomoção , Longevidade/genética , Neurônios/metabolismo , Doença de Parkinson/genética , Piridoxal Quinase/antagonistas & inibidores , Especificidade da Espécie , Fatores de Transcrição/genética , Vitamina B 6/metabolismo , Complexo Vitamínico B/metabolismo , alfa-Sinucleína/biossínteseRESUMO
The simultaneous role of neuroprotective estrogen and neurodegenerative inflammation during the progression of Parkinson's disease (PD) is still remaining elusive. The novel importance of the present study in MPTP mediated mouse model of Parkinson's disease (PD) is-to investigate the status of neuronal and glial cells in a time chase experiment; to explore which pathway of NF-kappaB exist to proceed the neuroinflammation; to investigate the status of estrogen and the activation pattern of nuclear or cytosolic estrogen receptors in either sexes of Swiss albino mice during MPTP mediated progressive neurodegeneration in the substantia nigra. After MPTP intoxication, the nigral molecular anatomy was changed differently in separate time interval during the progression of neurodegeneration with/without association of glial cells and functional (via its nuclear and cytosolic receptors) estrogen level. Both the canonical and/or non-canonical pathways of NF-kappaB exist in the substantia nigra of both the sexes after MPTP treatment that is why inspite of presence of estrogen, neuroinflammation progresses. The homodimeric or heterodimeric form of ER-beta binds with NF-kappaB molecules p65 and RelB differently, but the canonical or non-canonical pathways of NF-kappaB molecules could not be stopped or may be promoted.
Assuntos
1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina , NF-kappa B/metabolismo , Doença de Parkinson/metabolismo , Receptores de Estrogênio/metabolismo , Substância Negra/metabolismo , Animais , Astrócitos/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Contagem de Células , Modelos Animais de Doenças , Dopa Descarboxilase/metabolismo , Estradiol/metabolismo , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/metabolismo , Feminino , Proteína Glial Fibrilar Ácida/metabolismo , Inflamação/metabolismo , Masculino , Camundongos , Proteínas dos Microfilamentos/metabolismo , Microglia/metabolismo , Neurônios/enzimologia , Neurônios/patologia , Doença de Parkinson/etiologia , Fatores Sexuais , Fator de Transcrição RelA/metabolismo , Fator de Transcrição RelB/metabolismoRESUMO
The enzyme DOPA decarboxylase (aromatic-L-amino-acid decarboxylase, DDC) plays an important role in the dopaminergic system and participates in the uptake and decarboxylation of amine precursors in the peripheral tissues. Apart from catecholamines, DDC catalyses the biosynthesis of serotonin and trace amines. It has been shown that the DDC amino acid sequence is highly evolutionarily conserved across many species. The activity of holoenzyme is regulated by stimulation/blockade of membrane receptors, phosphorylation of serine residues, and DDC interaction with regulatory proteins. A single gene codes for DDC both in neuronal and non-neuronal tissue, but synthesized isoforms of mRNA differ in the 5' UTR and in the presence of alternative exons. Tissue-specific expression of the DDC gene is controlled by two spatially distinct promoters - neuronal and non-neuronal. Several consensus sequences recognized by the HNF and POU family proteins have been mapped in the neuronal DDC promoter. Since DDC is located close to the imprinted gene cluster, its expression can be subjected to tightly controlled epigenetic regulation. Perturbations in DDC expression result in a range of neurodegenerative and psychiatric disorders and correlate with neoplasia. Apart from the above issues, the role of DDC in prostate cancer, bipolar affective disorder, Parkinson's disease and DDC deficiency is discussed in our review. Moreover, novel and prospective clinical treatments based on gene therapy and stem cells for the diseases mentioned above are described.
Assuntos
Dopa Descarboxilase/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Descarboxilases de Aminoácido-L-Aromático/deficiência , Descarboxilases de Aminoácido-L-Aromático/metabolismo , Catecolaminas/biossíntese , Dopa Descarboxilase/química , Dopa Descarboxilase/genética , Dopa Descarboxilase/fisiologia , Feminino , Regulação da Expressão Gênica , Humanos , Masculino , Doença de Parkinson/metabolismo , Neoplasias da Próstata/metabolismo , Conformação Proteica , Isoformas de Proteínas/metabolismo , Serotonina/biossínteseRESUMO
Infection by the neurotropic agent Toxoplasma gondii alters rodent behavior and can result in neuropsychiatric symptoms in humans. Little is understood regarding the effects of infection on host neural processes but alterations to dopaminergic neurotransmission are implicated. We have previously reported elevated levels of dopamine (DA) in infected dopaminergic cells however the involvement of the host enzymes and fate of the produced DA were not defined. In order to clarify the effects of infection on host DA biosynthetic enzymes and DA packaging we examined enzyme levels and activity and DA accumulation and release in T. gondii-infected neurosecretory cells. Although the levels of the host tyrosine hydroxylase (TH) and DOPA decarboxylase and AADC (DDC) did not change significantly in infected cultures, DDC was found within the parasitophorous vacuole (PV), the vacuolar compartment where the parasites reside, as well as in the host cytosol in infected dopaminergic cells. Strikingly, DDC was found within the intracellular parasite cysts in infected brain tissue. This finding could provide some explanation for observations of DA within tissue cysts in infected brain as a parasite-encoded enzyme with TH activity was also localized within tissue cysts. In contrast, cellular DA packaging appeared unchanged in single-cell microamperometry experiments and only a fraction of the increased DA was accessible to high potassium-induced release. This study provides some understanding of how this parasite produces elevated DA within dopaminergic cells without the toxic ramifications of free cytosolic DA. The mechanism for synthesis and packaging of DA by T. gondii-infected dopaminergic cells may have important implications for the effects of chronic T. gondii infection on humans and animals.
Assuntos
Encéfalo/parasitologia , Dopamina/biossíntese , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/parasitologia , Toxoplasmose/metabolismo , Animais , Descarboxilases de Aminoácido-L-Aromático/metabolismo , Encéfalo/metabolismo , Dopa Descarboxilase/metabolismo , Neurônios Dopaminérgicos/enzimologia , Células PC12 , Ratos , Vesículas Sinápticas/metabolismo , Toxoplasmose/enzimologia , Tirosina 3-Mono-Oxigenase/metabolismo , Proteínas Vesiculares de Transporte de Monoamina/metabolismoRESUMO
Rotenone (RT) produces reactive oxygen species (ROS) by inhibiting the mitochondrial electron transport chain; causing dopaminergic (DA) cell death in the substantia nigra (SN) and simulates other models of induced Parkinson's disease (PD). There is a sincere dearth of knowledge regarding the status of glial cells, neuroprotective estrogen and the status of neuroinflammatory TNF-α in the different brain regions in either sex during healthy, as well as during PD conditions. In the present study of RT-induced mouse model of PD, we have selected the frontal cortex (FC), hippocampus (HC) and SN from either sex of Swiss albino mice as these are the major regions involved during PD pathogenesis. During non pathogenic conditions, the ROS-scavenging enzyme activity varied among the brain regions and also in between genders. The number of DOPA decarboxylase-positive cells, astrocytes and microglia was similar in the respective regions of the brain in both the sexes. The level of proinflammatory cytokine TNF-α was same in the respective FC and HC in either sex except that of SN. The expression level of estrogen and its receptors varied among the three brain regions. During RT treatment, ROS-scavenging enzyme activities increased, DOPA decarboxylase-positive neurons and fibers in DA as well as in norepinephrinergic (NE) systems become degenerated, number of astrocytes decreased and microglial cells increased in those specific brain regions in either of the sexes except in the SN region of males where astrocyte number remained unaltered and microglial cell percentage decreased. TNF-α increased in the FC and SN but remained unaltered in the HC of both sexes. Estradiol level decreased in the HC and SN but the level unevenly varied in the FC. Similarly, the estrogen bound and nuclear-cytosolic receptor α and ß also varied differentially among the brain regions of the two sexes. Therefore our present study depicts that there exists a clear variation of neuronal and astroglial cell population, estrogen and its receptor levels in different brain regions of both the sexes during control and RT-treated pathogenic condition and these variations have major implication in PD pathogenesis and progression.
Assuntos
Encéfalo/fisiopatologia , Estrogênios/metabolismo , Neuroglia/fisiologia , Neurônios/fisiologia , Transtornos Parkinsonianos/fisiopatologia , Caracteres Sexuais , Animais , Aromatase/metabolismo , Encéfalo/patologia , Núcleo Celular/fisiologia , Citosol/fisiologia , Dopa Descarboxilase/metabolismo , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/metabolismo , Feminino , Masculino , Camundongos , Neuroglia/patologia , Neuroimunomodulação/fisiologia , Neurônios/patologia , Transtornos Parkinsonianos/patologia , Distribuição Aleatória , Rotenona , Fator de Necrose Tumoral alfa/metabolismoRESUMO
l-DOPA decarboxylase (DDC) is a multiply-regulated gene which encodes the enzyme that catalyzes the biosynthesis of dopamine in humans. MicroRNAs comprise a novel class of endogenously transcribed small RNAs that can post-transcriptionally regulate the expression of various genes. Given that the mechanism of microRNA target recognition remains elusive, several genes, including DDC, have not yet been identified as microRNA targets. Nevertheless, a number of specifically designed bioinformatic algorithms provide candidate miRNAs for almost every gene, but still their results exhibit moderate accuracy and should be experimentally validated. Motivated by the above, we herein sought to discover a microRNA that regulates DDC expression. By using the current algorithms according to bibliographic recommendations we found that miR-145 could be predicted with high specificity as a candidate regulatory microRNA for DDC expression. Thus, a validation experiment followed by firstly transfecting an appropriate cell culture system with a synthetic miR-145 sequence and sequentially assessing the mRNA and protein levels of DDC via real-time PCR and Western blotting, respectively. Our analysis revealed that miR-145 had no significant impact on protein levels of DDC but managed to dramatically downregulate its mRNA expression. Overall, the experimental and bioinformatic analysis conducted herein indicate that miR-145 has the ability to regulate DDC mRNA expression and potentially this occurs by recognizing its mRNA as a target.
Assuntos
Descarboxilases de Aminoácido-L-Aromático/genética , Biologia Computacional/métodos , Dopa Descarboxilase/genética , MicroRNAs/metabolismo , Neoplasias da Próstata/genética , RNA Mensageiro/metabolismo , Algoritmos , Descarboxilases de Aminoácido-L-Aromático/metabolismo , Linhagem Celular Tumoral , Dopa Descarboxilase/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Neoplasias da Próstata/metabolismo , Estudos de Validação como AssuntoRESUMO
Insect molting is an important developmental process of metamorphosis, which is initiated by molting hormone. Molting includes the activation of dermal cells, epidermal cells separation, molting fluid secretion, the formation of new epidermis and old epidermis shed and other series of continuous processes. Polyphenol oxidases, dopa decarboxylase and acetyltransferase are necessary enzymes for this process. Traditionally, the dopa decarboxylase (BmDdc) was considered as an enzyme for epidermal layer's tanning and melanization. This work suggested that dopa decarboxylase is one set of the key enzymes in molting, which closely related with the regulation of ecdysone at the time of biological molting processes. The data showed that the expression peak of dopa decarboxylase in silkworm is higher during molting stage, and decreases after molting. The significant increase in the ecdysone levels of haemolymph was also observed in the artificially fed silkworm larvae with ecdysone hormone. Consistently, the dopa decarboxylase expression was significantly elevated compared to the control. BmDdc RNAi induced dopa decarboxylase expression obviously declined in the silkworm larvae, and caused the pupae appeared no pupation or incomplete pupation. BmDdc was mainly expressed and stored in the peripheral plasma area near the nucleus in BmN cells. In larval, BmDdc was mainly located in the brain and epidermis, which is consisted with its function in sclerotization and melanization. Overall, the results described that the dopa decarboxylase expression is regulated by the molting hormone, and is a necessary enzyme for the silkworm molting.
Assuntos
Bombyx/enzimologia , Dopa Descarboxilase/genética , Ecdisona/farmacologia , Animais , Western Blotting , Bombyx/efeitos dos fármacos , Bombyx/genética , Bombyx/crescimento & desenvolvimento , Dopa Descarboxilase/metabolismo , Ecdisona/administração & dosagem , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Larva/efeitos dos fármacos , Larva/genética , Especificidade de Órgãos/efeitos dos fármacos , Especificidade de Órgãos/genética , Transporte Proteico/efeitos dos fármacos , RNA de Cadeia Dupla/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
BACKGROUND: Sodium and water transport across renal proximal tubules is regulated by diverse hormones such as dopamine and urodilatin. We have previously reported that urodilatin stimulates extraneuronal dopamine uptake in external renal cortex by activation of the type A natriuretic peptide receptor, coupled to cyclic guanylate monophosphate signaling and protein kinase G. Moreover, urodilatin enhances dopamine-induced inhibition of Na+, K+-ATPase activity in renal tubules. The aim of the present study was to evaluate whether urodilatin could also alter renal dopamine synthesis, release, catabolism and turnover. METHODS: The effects of urodilatin on dopamine synthesis, release, catabolism and turnover were measured in samples of renal cortex from Sprague Dawley rats. RESULTS: The results indicate that urodilatin increases L-DOPA decarboxylase activity and decreases catechol-o-methyl transferase and monoamine oxidase activity. Moreover, urodilatin does not affect either dopamine basal secretion or potassium chloride-induced dopamine release in external renal cortex, and reduces amine turnover. CONCLUSIONS: Both the present results and previous findings show that urodilatin modifies dopamine metabolism in external renal cortex of rats by enhancing dopamine uptake and synthesis and by decreasing catechol-o-methyl transferase and monoamine oxidase activity and dopamine turnover. Those effects taken together may favor dopamine accumulation in renal cells and increase its endogenous content and availability. This would permit D1 receptor recruitment and stimulation and, in turn, overinhibition of Na+, K+-ATPase activity, which results in decreased sodium reabsorption. Therefore, urodilatin and dopamine enhance natriuresis and diuresis through a common pathway.
Assuntos
Fator Natriurético Atrial/fisiologia , Catecol O-Metiltransferase/metabolismo , Dopa Descarboxilase/metabolismo , Dopamina/metabolismo , Rim/metabolismo , Monoaminoxidase/metabolismo , Animais , Diurese , Túbulos Renais Proximais/metabolismo , Natriurese , Fragmentos de Peptídeos/fisiologia , Ratos , Ratos Sprague-Dawley , Receptores de Dopamina D1/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismoRESUMO
PURPOSE: Rats that are overfed during lactation exhibit neonatal hyperleptinemia and higher visceral adiposity, hypertension, higher liver oxidative stress and insulin resistance in the liver as adults. Previously, we demonstrated that neonatal hyperleptinemia is associated with adrenal medullary hyperfunction, hypertension and liver steatosis in adulthood. Therefore, we hypothesised that adrenal and liver functions are altered in adult obese rats that were overfed during lactation, which would underlie their hypertension and liver alterations. METHODS: The litter size was reduced from ten to three male pups on the third day of lactation until weaning (SL) to induce early overfeeding in Wistar rats. The control group had ten rats per litter (NL). Rats had free access to standard diet, and water after weaning until the rats were 180 days old. RESULTS: The SL group exhibited higher adrenal catecholamine content (absolute: +35% and relative: +40%), tyrosine hydroxylase (+31%) and DOPA decarboxylase (+90%) protein contents and basal catecholamine secretion in vitro (+57%). However, the hormones of the hypothalamic-pituitary-adrenal cortex axis were unchanged. ß3-adrenergic receptor content in visceral adipose tissue was unchanged in SL rats, but the ß2-adrenergic receptor content in the liver was lower in this group (-45%). The SL group exhibited higher glycogen and triglycerides contents in the liver (+79 and +49%, respectively), which suggested microesteatosis. CONCLUSIONS: Neonatal overfeeding led to higher adrenomedullary function, but the liver ß2-adrenergic receptor content was reduced. These results may contribute to the hepatic dysfunction characteristic of liver obesity complications.
Assuntos
Glândulas Suprarrenais/metabolismo , Catecolaminas/metabolismo , Comportamento Alimentar , Insuficiência Hepática/etiologia , Hiperfagia/fisiopatologia , Fígado/fisiopatologia , Regulação para Cima , Glândulas Suprarrenais/patologia , Animais , Animais Recém-Nascidos , Comportamento Animal , Dopa Descarboxilase/metabolismo , Regulação para Baixo , Hiperfagia/metabolismo , Hiperfagia/patologia , Hipertensão/etiologia , Fígado/metabolismo , Fígado/patologia , Glicogênio Hepático/metabolismo , Masculino , Obesidade/etiologia , Obesidade/fisiopatologia , Ratos , Receptores Adrenérgicos beta 2/metabolismo , Triglicerídeos/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) represents one of the most commonly diagnosed malignancies worldwide. The DDC gene encodes L-DOPA decarboxylase, an enzyme catalyzing the decarboxylation of L-DOPA to dopamine. We have recently shown that DDC mRNA is a significant predictor of patients' prognosis in colorectal adenocarcinoma and prostate cancer. The aim of the current study was to analyze the DDC mRNA expression in HNSCC patients. METHODS: 53 malignant tumors were resected from the larynx, pharynx, tongue, buccal mucosa, parotid glands, and nasal cavity, as well as from 34 adjacent non-cancerous tissues of HNSCC patients, and were homogenized. Total RNA was isolated and converted into first-strand cDNA. An ultrasensitive real-time PCR method based on the SYBR Green chemistry was used for DDC mRNA quantification in head and neck tissue specimens. Relative quantification was performed using the comparative Ct (2-ddCt) method. RESULTS: DDC mRNA levels were lower in squamous cell carcinomas (SCCs) of the larynx and tongue than in adjacent non-cancerous tissue specimens. Furthermore, low DDC mRNA expression was noticed in laryngeal and tongue tumors of advanced TNM stage or bigger size, compared to early-stage or smaller tumors, respectively. No statistically significant differences were observed between SCCs resected from pharynx, buccal mucosa, or nasal cavity, and their normal counterparts. CONCLUSION: This is the first study examining the DDC mRNA expression in HNSCC. According to our results, DDC mRNA expression may constitute a potential prognostic biomarker in tongue and/or larynx SCCs, which principally represent the overwhelming majority of HNSCC cases.
Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Dopa Descarboxilase/genética , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/patologia , RNA Mensageiro/biossíntese , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/análise , Biomarcadores Tumorais/genética , Carcinoma de Células Escamosas/enzimologia , Carcinoma de Células Escamosas/metabolismo , Dopa Descarboxilase/biossíntese , Dopa Descarboxilase/metabolismo , Neoplasias de Cabeça e Pescoço/enzimologia , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Pessoa de Meia-Idade , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reprodutibilidade dos Testes , Estudos Retrospectivos , Carcinoma de Células Escamosas de Cabeça e Pescoço , Estatísticas não ParamétricasRESUMO
Parkinson's disease (PD) is one of the most common neurodegenerative disorders characterized by decreased levels of the neurotransmitter dopamine (DA) in the striatum of the brain, as a result of degeneration of DA neurons. Levodopa (L-Dopa) crosses the blood-brain barrier and its administration replenishes the loss of DA in dopaminergic neurons in PD patients. Despite the evident beneficial effects, L-Dopa use may cause side effects and its toxicity found in in vitro assays has been attributed to the generation of reactive oxygen species (ROS): L-Dopa is converted to DA and its metabolism and autoxidation gives rise to quinones, semiquinones, and hydrogen peroxide. Despite this evidence, L-Dopa in some in vivo and in vitro experiments showed no toxic effects, or even antioxidant effects. Two major peripheral L-Dopa metabolic pathways, driven by the enzymes Aromatic L-amino acid decarboxylase (AADC) and catechol-O-methyl transferase (COMT), significantly deplete the amount of L-Dopa reaching the brain. The low bioavailability of L-Dopa may cause a wide variation in clinical response between patients. Strategies addressing to improve the bioavailability of L-Dopa include coadministering L-Dopa with carbidopa, a decarboxylase inhibitor, as multiple daily doses. We utilized catecholaminergic human neuroblastoma cells to study DNA damage and ROS production after L-Dopa and carbidopa treatments. Our data lead us to confirm that L-Dopa may have a protective effect on dopaminergic cells especially at certain concentrations, in particular, toward the production of ROS and their toxic effects on DNA. Furthermore in the combined treatment, with induction of ROS following administration of H(2)O(2), carbidopa is effective in reducing the damage caused by reactive oxygen intermediates both alone and in combination with L-Dopa.
Assuntos
Carbidopa/farmacologia , Dopamina/metabolismo , Levodopa/farmacologia , Antiparkinsonianos/metabolismo , Antiparkinsonianos/farmacologia , Inibidores das Descarboxilases de Aminoácidos Aromáticos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ensaio Cometa , Dano ao DNA/efeitos dos fármacos , Dopa Descarboxilase/metabolismo , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Inibidores Enzimáticos/farmacologia , Humanos , Peróxido de Hidrogênio/farmacologia , Levodopa/metabolismo , Neuroblastoma/genética , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Oxidantes/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Fatores de TempoRESUMO
Recently it has been demonstrated that catecholamines are produced and used by macrophages and mediate immune response. The aim of this study is to verify whether endothelial cells (ECs), which are of myeloid origin, can produce catecholamines. We demonstrated that genes coding for tyrosine hydroxylase, Dopa decarboxylase, dopamine ß hydroxylase (DßH), and phenylethanolamine-N-methyl transferase, enzymes involved in the synthesis of catecholamines, are all expressed in basal conditions in bovine aorta ECs, and their expression is enhanced in response to hypoxia. Moreover, hypoxia enhances catecholamine release. To evaluate the signal transduction pathway that regulates catecholamine synthesis in ECs, we overexpressed in bovine aorta ECs either protein kinase A (PKA) or the transcription factor cAMP response element binding, because PKA/cAMP response element binding activation induces tyrosine hydroxylase transcription and activity in response to stress. Both cAMP response element binding and PKA overexpression enhance DßH and phenylethanolamine-N-methyl transferase gene expression and catecholamine release, whereas H89, inhibitor of PKA, exerts the opposite effect, evidencing the role of PKA/cAMP response element binding transduction pathway in the regulation of catecholamine release in bovine aorta ECs. We then evaluated by immunohistochemistry the expression of tyrosine hydroxylase, Dopa decarboxylase, DßH, and phenylethanolamine-N-methyl transferase in femoral arteries from hindlimbs of C57Bl/6 mice 3 days after removal of the common femoral artery to induce chronic ischemia. Ischemia evokes tyrosine hydroxylase, Dopa decarboxylase, DßH, and phenylethanolamine-N-methyl transferase expression in the endothelium. Finally, the pharmacological inhibition of catecholamine release by fusaric acid, an inhibitor of DßH, reduces the ability of ECs to form network-like structures on Matrigel matrix. In conclusion, our study demonstrates for the first time that ECs are able to synthesize and release catecholamines in response to ischemia.