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Living Donor Liver Transplantation (LDLT) is a promising approach to treating end-stage liver diseases, however, some post-operatory complications such as pneumonia, bacteremia, urinary tract infections, and hepatic dysfunction have been reported. In murine models using partial hepatectomy (PHx), a model that emulates LDLT, it has been determined that the synthesis of hepatic cell proliferation factors that are associated with noradrenaline synthesis are produced in locus coeruleus (LC). In addition, studies have shown that PHx decreases GABA and 5-HT2A receptors, promotes loss of dendritic spines, and favors microgliosis in rat hippocampus. The GABA and serotonin-altered circuits suggest that catecholaminergic neurons such as dopamine and noradrenaline neurons, which are highly susceptible to cellular stress, can also be damaged. To understand post-transplant affections and to perform well-controlled studies it is necessary to know the potential causes that explain as a liver surgical procedure can produce brain damage. In this paper, we review several cellular processes that could induce gliosis in LC after rat PHx.
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Bromodeoxyuridine (BrdU) is used in studies related to cell proliferation and neurogenesis. The multiple intraperitoneal injections of this molecule could favor liver function profile changes. In this study, we evaluate the systemic and hepatocellular impact of BrdU in male adult Wistar rats in 30 %-partial hepatectomy (PHx) model. The rats received BrdU 50 mg/Kg by intraperitoneal injection at 0.5, 1, 2, 3, 6, 9 and 16 days after 30 %-PH. The rats were distributed into four groups as follows, control, sham, PHx/BrdU(-) and PHx/BrdU(+). On day 16, we evaluated hepatocellular nuclei and analyzed histopathological features by haematoxylin-eosin stain and apoptotic profile was qualified by caspase-3 presence. The systemic effect was evaluated by liver markers such as alanine transferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), alkaline phosphatase (AP), bilirubin, total proteins and serum albumin content. The statistical analysis consisted of a student t-test and one-way ANOVA. BrdU did not induce apoptosis or hepatocellular damage in male rats. Multiple administrations of BrdU in male rats did not induce significant decrease body weight, but increased serum ALT and LDH levels were found. Our results show that the BrdU does not produce hepatocellular damage.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Ratos , Masculino , Animais , Ratos Wistar , Bromodesoxiuridina/farmacologia , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/patologia , Fígado/patologia , Alanina Transaminase/metabolismo , Alanina Transaminase/farmacologia , Aspartato Aminotransferases/metabolismo , Aspartato Aminotransferases/farmacologiaRESUMO
PURPOSE: The main goal of this study was to determine the relationship of cleaved-caspase-3 (C3)-related apoptosis and hepatic proliferation, during the liver repopulation in a living liver donor rat model. MATERIAL/METHODS: Thirty-three animals were randomized into eleven groups and evaluated on postoperative from 3 âh until 384 âh after 30%-partial hepatectomy (30%-PHx). Liver sections (5 âµm) were processed by hematoxylin-eosin, and immunostaining for C3, accompanied by hepatic function test. C3 content and the hepatic lobule enlargement were analyzed by optical density, followed by cell counting. RESULTS: Transient variations of alanine transferase (ALT), aspartate aminotransferase (AST) and lactate dehydrogenase (LDH) were found. Significant increase in the C3 levels, and cell nuclei number, were detected at 12 âh and 48 âh after 30%-PHx, evidencing a correlation of p â= â-0.3679. CONCLUSION: In the 30%-PHx rat model, C3-related apoptosis prevents proliferative pathological conditions during the hepatic lobule re-modeling.
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Apoptose , Caspase 3/metabolismo , Proliferação de Células , Hepatectomia/métodos , Regeneração Hepática , Fígado/patologia , Animais , Caspase 3/genética , Doadores Vivos/estatística & dados numéricos , Masculino , Ratos , Ratos WistarRESUMO
Neurotensin (NTS)-polyplex is a multicomponent nonviral vector that enables gene delivery via internalization of the neurotensin type 1 receptor (NTSR1) to dopaminergic neurons and cancer cells. An approach to improving its therapeutic safety is replacing the viral karyophilic component (peptide KPSV40; MAPTKRKGSCPGAAPNKPK), which performs the nuclear import activity, by a shorter synthetic peptide (KPRa; KMAPKKRK). We explored this issue and the mechanism of plasmid DNA translocation through the expression of the green fluorescent protein or red fluorescent protein fused with KPRa and internalization assays and whole-cell patch-clamp configuration experiments in a single cell together with importin α/ß pathway blockers. We showed that KPRa electrostatically bound to plasmid DNA increased the transgene expression compared with KPSV40 and enabled nuclear translocation of KPRa-fused red fluorescent proteins and plasmid DNA. Such translocation was blocked with ivermectin or mifepristone, suggesting importin α/ß pathway mediation. KPRa also enabled NTS-polyplex-mediated expression of reporter or physiological genes such as human mesencephalic-derived neurotrophic factor (hMANF) in dopaminergic neurons in vivo. KPRa is a synthetic monopartite peptide that showed nuclear import activity in NTS-polyplex vector-mediated gene delivery. KPRa could also improve the transfection of other nonviral vectors used in gene therapy.
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Portadores de Fármacos/síntese química , Técnicas de Transferência de Genes , Vetores Genéticos/administração & dosagem , Neurotensina/administração & dosagem , Fragmentos de Peptídeos/síntese química , Transporte Ativo do Núcleo Celular , Animais , Linhagem Celular , Núcleo Celular/metabolismo , Neurônios Dopaminérgicos/citologia , Neurônios Dopaminérgicos/metabolismo , Terapia Genética/métodos , Vetores Genéticos/genética , Masculino , Camundongos , Modelos Animais , Nanopartículas/química , Neurotensina/genética , Neurotensina/farmacocinética , Técnicas de Patch-Clamp , Plasmídeos/genética , Ratos , Receptores de Neurotensina/metabolismo , Análise de Célula Única , Técnicas EstereotáxicasRESUMO
Lipopolysaccharide (LPS) is a potent immunogen when administered locally and/or systemically. The peripheral immunization with LPS could contribute to the progression of neurological diseases because a strong link between neuroinflammation and dopaminergic degeneration has been found. The switch between the survival and neuronal death in substantia nigra could be related to M1 (neurotoxic) and M2 (neuroprotective) microglia phenotypes. In this review, we present the current findings about microglia roles, biomarkers, and natural or synthetic immune modulators determined in the LPS-based murine model.
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Modelos Animais de Doenças , Inflamação/imunologia , Lipopolissacarídeos/farmacologia , Microglia/imunologia , Animais , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Humanos , Inflamação/induzido quimicamente , Microglia/efeitos dos fármacosRESUMO
The peripheral inflammatory stimulus could induce cell damage in peripheral organs and activate microglial cells in the brain. One such stimulus was given to adult male Wistar rats by injecting different concentrations of lipopolysaccharide (LPS; 50, 300, 500 ïg/kg and 5 mg/kg i.p.). To verify the systemic effect of the LPS administration, the serum content of C-reactive protein (CRP), the variation of body weight and cellular changes in the spleen, liver and kidney were determined. Motor impairment was evaluated by rotarod and open field tests. Microglia activation and dopaminergic degeneration was confirmed by immunolabelling for CD11b/c (microglia) and tyrosine hydroxylase (TH), respectively. The cell counting was performed in substantia nigra pars compacta (SNpc), microglial activation was explored in SNpc, substantia nigra pars reticulata (SNpr), substantia nigra pars compacta dorsal (SNcd) and the ventral tegmental area (VTA). For the statistical analysis, one-way ANOVA followed by Tukey post hoc test (p ≤ 0.05) was used. On day 7 post intraperitoneal administration of LPS, cellular atrophy was detected in the liver, kidney and spleen at 5 mg/kg, without significant changes in CRP levels. Body weight loss and motor impairment was present only on day 1 post LPS administration. The dosage of 500 ïg/kg and 5 mg/kg of LPS caused the loss of dopaminergic neurons (40%) in SNpc and microglia migration in a dose-dependent manner in SNcd, SNpc and SNpr. LPS-induced endotoxemia favours damage to the peripheral organs and microglial migration in a dose-dependent manner in rat substantia nigra.
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Endotoxemia/patologia , Lipopolissacarídeos/toxicidade , Microglia/patologia , Substância Negra/patologia , Animais , Movimento Celular , Neurônios Dopaminérgicos/patologia , Endotoxemia/induzido quimicamente , Masculino , Ratos , Ratos WistarRESUMO
OBJECTIVES: Parkinson's disease (PD) is characterized by motor and cognitive dysfunctions. The progressive degeneration of dopamine-producing neurons that are present in the substantia nigra pars compacta (SNpc) has been the main focus of study and PD therapies since ages. MATERIALS AND METHODS: In this manuscript, a systematic revision of experimental and clinical evidence of PD-associated cell process was conducted. RESULTS: Classically, the damage in the dopaminergic neuronal circuits of SNpc is favored by reactive oxidative/nitrosative stress, leading to cell death. Interestingly, the therapy for PD has only focused on avoiding the symptom progression but not in finding a complete reversion of the disease. Recent evidence suggests that the renin-angiotensin system imbalance and neuroinflammation are the main keys in the progression of experimental PD. CONCLUSION: The progression of neurodegeneration in SNpc is due to the complex interaction of multiple processes. In this review, we analyzed the main contribution of four cellular processes and discussed in the perspective of novel experimental approaches.
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BACKGROUND: The neurotrophin Brain-Derived Neurotrophic Factor (BDNF) influences nigral dopaminergic neurons via autocrine and paracrine mechanisms. The reduction of BDNF expression in Parkinson's disease substantia nigra (SN) might contribute to the death of dopaminergic neurons because inhibiting BDNF expression in the SN causes parkinsonism in the rat. This study aimed to demonstrate that increasing BDNF expression in dopaminergic neurons of rats with one week of 6-hydroxydopamine lesion recovers from parkinsonism. The plasmids phDAT-BDNF-flag and phDAT-EGFP, coding for enhanced green fluorescent protein, were transfected using neurotensin (NTS)-polyplex, which enables delivery of genes into the dopaminergic neurons via neurotensin-receptor type 1 (NTSR1) internalization. RESULTS: Two weeks after transfections, RT-PCR and immunofluorescence techniques showed that the residual dopaminergic neurons retain NTSR1 expression and susceptibility to be transfected by the NTS-polyplex. phDAT-BDNF-flag transfection did not increase dopaminergic neurons, but caused 7-fold increase in dopamine fibers within the SN and 5-fold increase in innervation and dopamine levels in the striatum. These neurotrophic effects were accompanied by a significant improvement in motor behavior. CONCLUSIONS: NTS-polyplex-mediated BDNF overexpression in dopaminergic neurons has proven to be effective to remit hemiparkinsonism in the rat. This BDNF gene therapy might be helpful in the early stage of Parkinson's disease.