RESUMO
Parkinson's disease (PD) is characterized by the loss of dopaminergic neurons in the substantia nigra pars compact (SNpc), and no effective treatment has yet been established to prevent PD. Neurotrophic factors, such as cerebral dopamine neurotrophic factor (CDNF), have shown a neuroprotective effect on dopaminergic neurons. Previously, we developed a cell-penetrating-peptide-based delivery system that includes Asn194Lys mutation in the rabies virus glycoprotein-9R peptide (mRVG9R), which demonstrated a higher delivery rate than the wild-type. In this study, using a mouse PD-like model, we evaluated the intrastriatal mRVG9R-KP-CDNF gene therapy through motor and cognitive tests and brain cell analysis. The mRVG9R-KP-CDNF complex was injected into the striatum on days 0 and 20. To induce the PD-like model, mice were intraperitoneally administered Paraquat (PQ) twice a week for 6 weeks. Our findings demonstrate that mRVG9R-KP-CDNF gene therapy effectively protects brain cells from PQ toxicity and prevents motor and cognitive dysfunction in mice. We propose that the mRVG9R-KP-CDNF complex inhibits astrogliosis and microglia activation, safeguarding dopaminergic neurons and oligodendrocytes from PQ-induced damage. This study presents an efficient CDNF delivery system, protecting neurons and glia in the nigrostriatal pathway from PQ-induced damage, which is known to lead to motor and cognitive dysfunction in neurodegenerative diseases such as PD.
Assuntos
Doença de Parkinson , Animais , Doença de Parkinson/terapia , Doença de Parkinson/metabolismo , Fatores de Crescimento Neural/genética , Fatores de Crescimento Neural/metabolismo , Substância Negra , Modelos Animais de Doenças , Neurônios DopaminérgicosRESUMO
The Cerebral Dopamine Neurotrophic Factor (CDNF) is a neurotrophic factor that has a protective effect in cell and animal models of several neurodegenerative diseases. The molecular mechanism of the protective effect of CDNF is unclear. Many neurodegenerative diseases have been related to a proteostasis dysregulation in the endoplasmic reticulum (ER). A failure of proteostasis produces ER stress, triggering the unfolded protein response (UPR) and, in the long-term, induces cell death. An adaptive UPR solves ER stress by attenuating protein synthesis, inducing chaperones expression, and degradation of misfolded proteins. Since CDNF is an ER resident protein, we investigated whether the role of CDNF is to regulate ER proteostasis. To this end, we determined the effect of CDNF in thapsigargin-induced ER stress in HEK293-T cells and cultured hippocampal neurons. Our results show that CDNF improved the viability of HEK293-T cells exposed to thapsigargin. CDNF increased levels of protective proteins of the early UPR, such as BiP, ATF4, ATF6, and XBP-1 in both HEK293-T cells and neurons. Conversely, expression of CDNF attenuated ER stress-induced apoptotic proteins, CHOP and cleaved caspase-3 in HEK293-T cells and neurons. A mutant CDNF lacking the ER retention sequence failed to protect against ER stress. In conclusion, CDNF regulates proteostasis in the ER by inducing the adaptive UPR response and inhibiting apoptotic pathways triggered by ER stress. We propose that neuroprotection induced by CDNF is mediated by regulating ER proteostasis.
Assuntos
Morte Celular , Estresse do Retículo Endoplasmático , Retículo Endoplasmático/metabolismo , Fatores de Crescimento Neural/metabolismo , Resposta a Proteínas não Dobradas , Animais , Apoptose/efeitos dos fármacos , Biologia Computacional/métodos , Retículo Endoplasmático/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Células HEK293 , Humanos , Fatores de Crescimento Neural/farmacologia , Células Piramidais/metabolismo , RatosRESUMO
Neurotrophic factors comprise essential secreted proteins that have several functions in neural and non-neural tissues, mediating the development, survival and maintenance of peripheral and central nervous system. Therefore, neurotrophic factor issue has been extensively investigated into the context of neurodegenerative diseases. Alzheimer's disease and Parkinson's disease show changes in the regulation of specific neurotrophic factors and their receptors, which appear to be critical for neuronal degeneration. Indeed, neurotrophic factors prevent cell death in degenerative processes and can enhance the growth and function of affected neurons in these disorders. Based on recent reports, this review discusses the main findings related to the neurotrophic factor support - mainly brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor - in the survival, proliferation and maturation of affected neurons in Alzheimer's disease and Parkinson's disease as well as their putative application as new therapeutic approach for these diseases management.
RESUMO
CDNF is a recently described evolutionary conserved neurotrophic factor reported to be of relevance for the treatment of Parkinson's disease. Treatment with recombinant CDNF showed neurorestorative and neuroprotective effects on dopaminergic neurons in Parkinsonian animal models. Similar results are obtained using adeno-associated viral (AAV) vectors for CDNF expression in these animal models; however, the extent of the transduced brain tissue is difficult to assess due to the lack of reporter genes in the vectors used. Here, we describe two bicistronic lentiviral plasmids based on the Δ1D/2A and IRES elements for the expression of EGFP and rat CDNF, in order to track the transduced cells expressing CDNF with EGFP fluorescence. Transfected heterologous cells or transduced neurons with these vectors are easily identified by EGFP fluorescence and CDNF expression results in its recruitment to the endoplasmic reticulum (ER) by both bicistronic vectors. CDNF immunostaining is also observed in the Golgi apparatus when expressed in heterologous cells or hippocampal neuronal cultures; however, colocalization with a dense core secretory vesicle marker was scarce. Additionally, we showed that the expression of CDNF inhibited dendrite formation in hypothalamic neurons, suggesting that CDNF expressed by these bicistronic lentiviral vectors is functional and could have a role in neuronal morphology. The bicistronic lentiviral plasmids developed here could be of use to study the effect of rat CDNF at the cellular level or to better delineate the perikarya of neurons transduced with lentiviral vectors in animal models of Parkinson's disease.