Fig 2 - available via license: Creative Commons Attribution 4.0 International
Content may be subject to copyright.
AngII and 6-OHDA increase ROS production in neuronal cell lines over time. Neuronal cells were loaded with probe DCFDA and the effect of different treatments on the ROS production in response to different treatments was measured in a plate reader for 120 min. Graphs show an increase in DCFDA fluorescence over time when dopaminergic Mes23.5 cells are treated with AngII, 6-OHDA or a combination of both (A). In dopaminergic N27 cells, treatment with 6OHDA induced an increase in DCFDA fluorescence signal, but no significant effect on ROS production could be measured in the plate reader in response to AngII treatment (B) SH-SY5Y cells are sensitive to both AngII and 6-OHDA-induced ROS production, increasing DCFDA fluorescence levels over time (C) Data are represented as fold increase in DCFDA fluorescence over control at time 15 min. þ p o 0.05 AngII vs control, þ þp o 0.001 AngII vs control; †p o 0.05 6-OHDA vs control, † †p o 0.001 6-OHDA vs control; ‡p o 0.05 AngIIþ 6-OHDA vs control, ‡ ‡p o 0.001 AngII þ6-OHDA vs control; #p o 0.001 AngII þ 6-OHDA vs 6-OHDA, ##p o 0.001 AngII þ6-OHDA vs 6-OHDA. Ang II, Angiotensin II; DCFDA, 2 0 ,7 0-dichlorofluorescin-diacetate; NAC, N-Acetyl-LCysteine; H 2 O 2 , hydrogen peroxide; ROS, reactive oxygen species.
Source publication
This article describes the effect of the oxidative stress inducers Angiotensin II and 6-hydroxydopamine (6-OHDA) on different cell lines. The levels of expression Angiotensin type 1 and type 2 receptors in different dopaminergic cell lines are shown. The data indicate that treatment with Angiotensin II and 6-OHDA increases the production of reactiv...
Context in source publication
Context 1
... data presented herein contain gene expression levels of Angiotensin II receptors in dopami- nergic cell lines Mes23.5 and N27 (Fig. 1). We also present data on the fluorescence intensity of DCFDA (i.e. ROS generation) over time in these lines and in the human neuroblastoma SH-SY5Y cell line (Figs. 2, 6). We analyzed ROS generation induced by Angiotensin II and/or 6-OHDA, and the corresponding positive and negative controls. Furthermore, we provide data of mRNA expression analysis of genes related to the NRF2 pathway at different time points (Figs. 4-5). We also show the effects of NRF2 overexpression on the viability of neuronal cell ...
Citations
... In order to understand the effect of AngII and 6-OHDA in the NRF2 signaling pathway, the mRNA expression of Hmox1 and Nqo1 was analyzed by real-time quantitative RT-PCR. Based on previous data [49], the expression of these genes was estimated 24 h after treatment. Treatment of Mes23.5 with AngII or 6-OHDA induced a significant increase in mRNA levels of Hmox1 and Nqo1, and maximal expression was obtained with combination of AngII+ 6-OHDA ( Fig. 2A,B). ...
... The above mentioned results suggest that AngII increases the generation of ROS in dopaminergic neurons, and this increase causes both cell damage and the activation of the NRF2 pathway. The neuroprotective role of NRF2 is supported by the increase in survival of Mes23.5 cell line infected with a NRF2 expression vector [49]. and SH-SY5Y cells (E,F). ...
... Previous studies have suggested that the expression of the transcription factor KLF9 may be regulated by NRF2 and affect cell survival. To verify this effect in our cells, we analyzed the expression of KLF9 by RT-PCR in the same conditions that induced NRF2-pathway activation [49]. In Mes23.5, treatment with AngII induced an increase in the levels of KLF9 compared to untreated cells (2.2 ± 0.3 fold increase relative to control, p ≤ 0.001; Fig. 3A). ...
Nuclear factor-E2-related factor 2 (NRF2) is a transcription factor that activates the antioxidant cellular defense in response to oxidative stress, leading to neuroprotective effects in Parkinson's disease (PD) models. We have previously shown that Angiotensin II (AngII) induces an increase in reactive oxygen species (ROS) via AngII receptor type 1 and NADPH oxidase (NOX), which may activate the NRF2 pathway. However, controversial data suggest that AngII induces a decrease in NRF2 signaling leading to an increase in oxidative stress. We analyzed the effect of AngII and the dopaminergic neurotoxin 6-hydroxydopamine (6-OHDA) in culture and in vivo, and examined the effects on the expression of NRF2-related genes. Treatment of neuronal cell lines Mes23.5, N27 and SH-SY5Y with AngII, 6-OHDA or a combination of both increased ROS production and reduced cell viability. Simultaneously, these treatments induced an increase in expression in the NRF2-regulated genes heme oxygenase 1 (Hmox1), NAD(P)H quinone dehydrogenase 1 (Nqo1) and Kruppel like factor 9 (Klf9). Moreover, overexpression of KLF9 transcription factor caused a reduction in the production of ROS induced by treatment with AngII or 6-OHDA and improved the survival of these neuronal cells. Rats treated with AngII, 6-OHDA or a combination of both also showed an increased expression of NRF2 related genes and KLF9. In conclusion, our data indicate that AngII induces a damaging effect in neuronal cells, but also acts as a signaling molecule to activate NRF2 and KLF9 neuroprotective pathways in cellular and animal models of PD.
