DJ-1 Upregulation Research Articles

A novel compound alleviates oxidative stress via PKA/CREB1-mediated DJ-1 upregulation.

Oxidative stress is one of the major culprits causing dopaminergic neuron loss in Parkinson's disease (PD). DJ-1 is a protein with multiple actions against oxidative stress, apoptosis, neuroinflammation, etc. DJ-1 expression is decreased in sporadic PD, therefore increasing DJ-1 expression might be beneficial in PD treatment. However, drugs known to upregulate DJ-1 are still lacking. In this study, we identified a novel DJ-1-elevating compound called ChemJ through luciferase assay-based high-throughput compound screening in SH-SY5Y cells and confirmed that ChemJ upregulated DJ-1 in SH-SY5Y cell line and primary cortical neurons. DJ-1 upregulation by ChemJ alleviated MPP+-induced oxidative stress. In exploring the underlying mechanisms, we found that the transcription factor CREB1 bound to DJ-1 promoter and positively regulated its expression under both unstressed and 1-methyl-4-phenylpyridinium-induced oxidative stress conditions and that ChemJ promoted DJ-1 expression via activating PKA/CREB1 pathway in SH-SY5Y cells. Our results demonstrated that ChemJ alleviated the MPP+-induced oxidative stress through a PKA/CREB1-mediated regulation of DJ-1 expression, thus offering a novel and promising avenue for PD treatment.

DJ-1 protects cell death from a mitochondrial oxidative stress due to GBA1 deficiency.

GBA1 mutations are the most common genetic risk factor for development of Parkinson's disease (PD). The loss of catalytic activity in GBA1, as well as the reduction of the GBA1 protein in certain cellular compartment, may increase disease progression. However, the mechanisms underlying cellular dysfunction caused by GBA1 deficiency are still mostly unknown. In this study, we focus on the genetic interaction between GBA1 deficiency and PD-causing genes, such as DJ-1, in mitochondrial dysfunction. GBA1 knockout (KO) SH-SY5Y cells were used to assess DJ-1 functions against oxidative stress in vitro. The levels of cellular reactive oxygen species were monitored with MitoSOX reagent. The expression of the PARK7 gene was analyzed using the quantitative real-time PCR (qRT-PCR). To understand the mechanism underlying DJ-1 upregulation in GBA1 KO cells, we assess ROS levels, antioxidant protein, and cell viability in GBA1 KO cells with treatment of ROS inhibitor N-acetyl-cysteine or miglustat, which is an inhibitor of glucosylceramide synthase. Dopaminergic degeneration was assessed from Gba1 L444P heterozygous mice mated with Park7 knockout mice. We find that DJ-1 is significantly upregulated in GBA1 KO cells. Elevated levels of DJ-1 are attributed to the transcriptional expression of PARK7 mRNA, but not the inhibition of DJ-1 protein degradation. Because DJ-1 expression is highly linked to oxidative stress, we observe cellular reactive oxygen species (ROS) in GBA1 KO cells. Moreover, several antioxidant gene expressions and protein levels are increased in GBA1 KO cells. To this end, GBA1 KO cells are more susceptible to H2O2-induced cell death. Importantly, there is a significant reduction in dopaminergic neurons in the midbrain from Gba1 L444P heterozygous mice mated with Park7 knockout mice, followed by mild motor dysfunction. Taken together, our results suggest that DJ-1 upregulation due to GBA1 deficiency has a protective role against oxidative stress. It may be supposed that mutations or malfunctions in the DJ-1 protein may have disadvantages in the survival of dopaminergic neurons in the brains of patients harboring GBA1 mutations.

DJ-1-mediated p62 degradation delays intervertebral disc degeneration by inhibiting apoptosis of nucleus pulposus cells.

Intervertebral disc degeneration (IDD) is the most important pathological basis of degenerative spinal diseases, for which effective interventions are still lacking. Oxidative stress is considered to be one of the leading pathological mechanisms contributing to IDD. However, the exact role of DJ-1 as an essential member of the antioxidant defense system in IDD is still unclear. Therefore, the aim of this study was to investigate the role played by DJ-1 in IDD and to reveal its potential molecular mechanisms. Western blot and immunohistochemical staining assays were performed to detect the expression of DJ-1 in degenerative nucleus pulposus cells (NPCs). After overexpression of DJ-1 in NPCs by lentiviral transfection, DCFH-DA and MitoSOX fluorescent probes were used to evaluate the levels of reactive oxygen species (ROS); while western blot, TUNEL staining, and Caspase-3 activity were used to assess apoptosis. Immunofluorescence staining was used to demonstrate the relationship between DJ-1 and p62. After inhibition of lysosomal degradation function with chloroquine, p62 degradation and apoptosis in DJ-1 overexpressing NPCs were further examined. In vivo, we assessed the therapeutic effect of upregulated DJ-1 on IDD by X-ray, MRI and Safranin O-Fast green staining. The protein expression of DJ-1 was significantly decreased in degenerated NPCs, accompanied by increased apoptosis. However, overexpression of DJ-1 significantly inhibited the elevated ROS levels and apoptosis in NPCs under oxidative stress. Mechanistically, our results showed that upregulation of DJ-1 promoted p62 degradation via the autophagic lysosomal pathway and that the protective effect of DJ-1 on NPCs under oxidative stress was partially mediated by promoting lysosomal pathway degradation of p62. Moreover, intradiscal injection of adeno-associated virus for overexpression of DJ-1 mitigated the progression of IDD in rats. This study reveals that DJ-1 maintains the homeostasis of NPCs by promoting the degradation of p62 through the autophagic lysosomal pathway, suggesting that DJ-1 is a promising new target for IDD intervention.

Upregulation of DJ-1 in Dopaminergic Neurons by a Physically-Modified Saline: Implications for Parkinson’s Disease

Parkinson's disease (PD) is the second most common neurodegenerative disorder in human and loss-of-functions DJ-1 mutations are associated with a familial form of early onset PD. Functionally, DJ-1 (PARK7), a neuroprotective protein, is known to support mitochondria and protect cells from oxidative stress. Mechanisms and agents by which the level of DJ-1 could be increased in the CNS are poorly described. RNS60 is a bioactive aqueous solution created by exposing normal saline to Taylor-Couette-Poiseuille flow under high oxygen pressure. Recently we have described neuroprotective, immunomodulatory and promyelinogenic properties of RNS60. Here we delineate that RNS60 is also capable of increasing the level of DJ-1 in mouse MN9D neuronal cells and primary dopaminergic neurons, highlighting another new neuroprotective effect of RNS60. While investigating the mechanism we found the presence of cAMP response element (CRE) in DJ-1 gene promoter and stimulation of CREB activation in neuronal cells by RNS60. Accordingly, RNS60 treatment increased the recruitment of CREB to the DJ-1 gene promoter in neuronal cells. Interestingly, RNS60 treatment also induced the enrollment of CREB-binding protein (CBP), but not the other histone acetyl transferase p300, to the promoter of DJ-1 gene. Moreover, knockdown of CREB by siRNA led to the inhibition of RNS60-mediated DJ-1 upregulation, indicating an important role of CREB in DJ-1 upregulation by RNS60. Together, these results indicate that RNS60 upregulates DJ-1 in neuronal cells via CREB-CBP pathway. It may be of benefit for PD and other neurodegenerative disorders.

DJ-1 Alleviates Neuroinflammation and the Related Blood-Spinal Cord Barrier Destruction by Suppressing NLRP3 Inflammasome Activation via SOCS1/Rac1/ROS Pathway in a Rat Model of Traumatic Spinal Cord Injury.

DJ-1 has been shown to play essential roles in neuronal protection and anti-inflammation in nervous system diseases. This study aimed to explore how DJ-1 regulates neuroinflammation after traumatic spinal cord injury (t-SCI). The rat model of spinal cord injury was established by the clamping method. The Basso, Beattie, Bresnahan (BBB) score and the inclined plane test (IPT) were used to evaluate neurological function. Western blot was then applied to test the levels of DJ-1, NLRP3, SOCS1, and related proinflammatory factors (cleaved caspase 1, IL-1β and IL-18); ROS level was also examined. The distribution of DJ-1 was assessed by immunofluorescence staining (IF). BSCB integrity was assessed by the level of MMP-9 and tight junction proteins (Claudin-5, Occludin and ZO-1). We found that DJ-1 became significantly elevated after t-SCI and was mainly located in neurons. Knockdown of DJ-1 with specific siRNA aggravated NLRP3 inflammasome-related neuroinflammation and strengthened the disruption of BSCB integrity. However, the upregulation of DJ-1 by Sodium benzoate (SB) reversed these effects and improved neurological function. Furthermore, SOCS1-siRNA attenuated the neuroprotective effects of DJ-1 and increased the ROS, Rac1 and NLRP3. In conclusion, DJ-1 may alleviate neuroinflammation and the related BSCB destruction after t-SCI by suppressing NLRP3 inflammasome activation by SOCS1/Rac1/ROS pathways. DJ-1 shows potential as a feasible target for mediating neuroinflammation after t-SCI.

DJ-1-Nrf2 axis is activated upon murine β-coronavirus infection in the CNS

Coronaviruses have emerged as alarming pathogens owing to their inherent ability of genetic variation and cross-species transmission. Coronavirus infection burdens the endoplasmic reticulum (ER.), causes reactive oxygen species production and induces host stress responses, including unfolded protein response (UPR) and antioxidant system. In this study, we have employed a neurotropic murine β-coronavirus (M-CoV) infection in the Central Nervous System (CNS) of experimental mice model to study the role of host stress responses mediated by interplay of DJ-1 and XBP1. DJ-1 is an antioxidant molecule with established functions in neurodegeneration. However, its regulation in virus-induced cellular stress response is less explored. Our study showed that M-CoV infection activated the glial cells and induced antioxidant and UPR genes during the acute stage when the viral titer peaks. As the virus particles decreased and acute neuroinflammation diminished at day ten p.i., a significant up-regulation in UPR responsive XBP1, antioxidant DJ-1, and downstream signaling molecules, including Nrf2, was recorded in the brain tissues. Additionally, preliminary in silico analysis of the binding between the DJ-1 promoter and a positively charged groove of XBP1 is also investigated, thus hinting at a mechanism behind the upregulation of DJ-1 during MHV-infection. The current study thus attempts to elucidate a novel interplay between the antioxidant system and UPR in the outcome of coronavirus infection.

Red Light Irradiation In Vivo Upregulates DJ-1 in the Retinal Ganglion Cell Layer and Protects against Axotomy-Related Dendritic Pruning.

Retinal ganglion cells (RGCs) undergo dendritic pruning in a variety of neurodegenerative diseases, including glaucoma and autosomal dominant optic atrophy (ADOA). Axotomising RGCs by severing the optic nerve generates an acute model of RGC dendropathy, which can be utilized to assess the therapeutic potential of treatments for RGC degeneration. Photobiomodulation (PBM) with red light provided neuroprotection to RGCs when administered ex vivo to wild-type retinal explants. In the current study, we used aged (13–15-month-old) wild-type and heterozygous B6;C3-Opa1Q285STOP (Opa1+/−) mice, a model of ADOA exhibiting RGC dendropathy. These mice were pre-treated with 4 J/cm2 of 670 nm light for five consecutive days before the eyes were enucleated and the retinas flat-mounted into explant cultures for 0-, 8- or 16-h ex vivo. RGCs were imaged by confocal microscopy, and their dendritic architecture was quantified by Sholl analysis. In vivo 670 nm light pretreatment inhibited the RGC dendropathy observed in untreated wild-type retinas over 16 h ex vivo and inhibited dendropathy in ON-center RGCs in wild-type but not Opa1+/− retinas. Immunohistochemistry revealed that aged Opa1+/− RGCs exhibited increased nitrosative damage alongside significantly lower activation of NF-κB and upregulation of DJ-1. PBM restored NF-κB activation in Opa1+/− RGCs and enhanced DJ-1 expression in both genotypes, indicating a potential molecular mechanism priming the retina to resist future oxidative insult. These data support the potential of PBM as a treatment for diseases involving RGC degeneration.

Tanshinone IIA protects against dopaminergic neuron degeneration via regulation of DJ-1 and Nrf2/HO-1 pathways in a rodent model of Parkinson’s disease

Purpose: To study the potential neuroprotective effects of tanshinone IIA, a diterpene quinone, in an experimental model of 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced Parkinson disease (PD).
 Methods: Mice (C57BL/6) were administered freshly-prepared MPTP at a dose of 20 mg/kg body weight intraperitoneally, 4 times at 2-h intervals, to induce PD. Doses of 12.5, 25 and 50 mg/kg tanshinone IIA were administered to the mice as treatments for PD. Pole and Rota-rod tests were carried out to assess muscular coordination and bradykinesia. Protein expressions, reactive oxygen species (ROS) and malonaldehyde and other parameters were evaluated.
 Results: Tanshinone IIA at doses of 12.5, 25 and 50 mg/kg reduced deficits in muscular coordination and improved learning ability of MPTP-treated mice. It also reduced loss of tyrosine hydroxylase (TH)- positive neurons following MPTP-induction. Tanshinone IIA regulated apoptotic pathway proteins, i.e., Bax and Bcl-2, and inhibited the translocation of Cyt C to the mitochondria. Oxidative stress induced by MPTP was significantly inhibited by tanshinone IIA via up-regulation of DJ-1/Nrf2 /HO-1 expression and reduction of ROS and MDA levels. Brain tissue total glutathione content was increased by tanshinone IIA treatment.
 Conclusion: Tanshinone IIA effectively improves antioxidant status and reduces neuronal loss following MPTP treatment. These results indicate that tanshinone IIA exerts protective effects in MPTPinduced PD in mice. Thus, tanshinone IIA has a good potential for use as a therapy for PD.

Ischemic Postconditioning-Mediated DJ-1 Activation Mitigate Intestinal Mucosa Injury Induced by Myocardial Ischemia Reperfusion in Rats Through Keap1/Nrf2 Pathway.

Intestinal mucosal barrier dysfunction induced by myocardial ischemia reperfusion (IR) injury often leads to adverse cardiovascular outcomes after myocardial infarction. Early detection and prevention of remote intestinal injury following myocardial IR may help to estimate and improve prognosis after acute myocardial infarction (AMI). This study investigated the protective effect of myocardial ischemic postconditioning (IPo) on intestinal barrier injury induced by myocardial IR and the underlying cellular signaling mechanisms with a focus on the DJ-1. Adult SD rats were subjected to unilateral myocardial IR with or without ischemic postconditioning. After 30 min of ischemia and 120 min of reperfusion, heart tissue, intestine, and blood were collected for subsequent examination. The outcome measures were (i) intestinal histopathology, (ii) intestinal barrier function and inflammatory responses, (iii) apoptosis and oxidative stress, and (iv) cellular signaling changes. IPo significantly attenuated intestinal injury induced by myocardial IR. Furthermore, IPo significantly increased DJ-1, nuclear Nrf2, NQO1, and HO-1 expression in the intestine and inhibited IR-induced apoptosis and oxidative stress. The protective effect of IPo was abolished by the knockdown of DJ-1. Conversely, the overexpression of DJ-1 provided a protective effect similar to that of IPo. Our data indicate that IPo protects the intestine against myocardial IR, which is likely mediated by the upregulation of DJ-1/Nrf2 pathway.

DJ-1 alleviates anoxia and hypoglycemia injury in cardiac microvascular via AKT and GSH

Cardiac microvascular damage, which is often caused by anoxia and hypoglycemia, is associated with the development of cardiac injury. DJ-1 encodes a peptidase C56 protein family related protein, is has been linked to oxidative stress in various cells such as neurons, COPD epithelial cells, and macrophages. However, the effect of DJ-1 towards oxidative stress caused by anoxia and hypoglycemia of cardiac microvascular endothelial cells (CMEC) remains unclear. In this study, we investigated the role and underlying molecular mechanism of DJ-1 in CMEC with anoxia/hypoglycemic (A/H) injury. We found that the mRNA and the protein expression of DJ-1 in CMEC with A/H injury were significantly downregulated. DJ-1 overexpression by pcDNA.3.1-DJ-1 transfection elevated cell viability while it inhibited LDH leakage, cell apoptosis, caspase-3 activity, ROS level, and MDA contents, while knockdown of DJ-1 has the opposite results. In addition, tube formation was increased in DJ-1 overexpression, while it was decreased in DJ-1 knockdown CMEC with A/H injury. In addition, our results indicated that DJ-1 can regulate glutathione (GSH) levels by modulating AKT activity in CMEC with A/H injury. The downregulation of AKT and GSH may remove the protective role of DJ-1 against A/H injury in CMEC. Taken together, this study showed that DJ-1 upregulation protected CMEC against A/H injury via the AKT/GSH signaling pathway.

Increased miR-142 and decreased DJ-1 enhance the sensitivity of pancreatic cancer cell to adriamycin.

Phosphatidylinositol-3 kinase (PI3K)/protein kinase B (AKT) signaling pathway is related to tumorigenesis by up-regulating survivin. Phosphatase and tensin homologue deleted on chromosome ten (PTEN) can suppress PI3K/AKT signaling pathway, while DJ-1 is the negative regulator of PTEN. DJ-1 up-regulation is closely correlated with the occurrence, progression, and drug resistance of pancreatic cancer. MicroRNA-142 (MiR-142) is significantly declined in pancreatic cancer tissue. Bioinformatics analysis demonstrated that complementary binding site exists between miR-142 and DJ-1. This investigation, therefore, aimed to study the role of miR-142 in the regulation of DJ-1-PTEN/PI3K/AKT/Survivin signaling pathway as well as in pancreatic cancer cell proliferation, apoptosis, and adriamycin (ADM) resistance. Dual luciferase assay was performed to assess the targeted relationship between miR-142 and DJ-1. MiR-142, DJ-1, and PTEN expressions in SW1990 cells and drug-resistant SW1990/ADM cells were compared. SW1990/ADM cells were divided into five groups, including mimic-NC, miR-142 mimic, small interfere normal control (si-NC), si-DJ-1, and miR-142 mimic + si-DJ-1 groups. DJ-1, PTEN, phosphorylated-AKT (p-AKT), and Survivin expressions were tested. Cell apoptosis was determined by flow cytometry. Cell proliferation was evaluated by EdU staining. MiR-142 targeted inhibited DJ-1 expression. MiR-142, PTEN, and cell apoptosis significantly down-regulated, while DJ-1, p-AKT, Survivin, and cell proliferation significantly elevated in SW1990/ADM cells compared with SW1990 cells. MiR-142 mimics and/or si-DJ-1 transfection markedly reduced DJ-1, p-AKT, and Survivin expressions enhanced PTEN level, attenuated cell proliferation, enhanced cell apoptosis, and weakened ADM resistance. MiR-142 over-expression weakened ADM resistance in pancreatic cancer cells by targeting DJ-1 to enhance PTEN expression and attenuate PI3K/AKT signaling pathway activity.

Protocatechuic acid-mediated DJ-1/PARK7 activation followed by PI3K/mTOR signaling pathway activation as a novel mechanism for protection against ketoprofen-induced oxidative damage in the gastrointestinal mucosa

Oxidative stress contributes to the progression of non-steroidal anti-inflammatory drug (NSAID)-induced gastrointestinal (GI) cell apoptosis. In our previous study, we reported that nuclear factor erythroid 2-related factor 2 (Nrf2) plays a protective role against ketoprofen-induced GI mucosal oxidative injury. Recent reports suggest that Nrf2 could exhibit antioxidative and antiapoptosis responses through up-regulation of DJ-1 (PARK7). In the current study, we proposed that induction of DJ-1 expression by protocatechuic acid (PCA) might provide a potential therapeutic approach for treating oxidative stress-associated GI ulcer diseases. The results indicated that PCA increased mRNA expression of glutathione peroxidase and heme oxygenase-1 through up-regulation of DJ-1 followed by Nrf2 translocation. Furthermore, PCA protected Int-407 cells against ketoprofen-induced oxidative stress by regulating the DJ-1, PI3K, and mTOR pathways. Pretreatment with PCA inhibited mitochondrial ROS generation, up-regulated the mitochondrial membrane potential, and down-regulated pro-apoptotic Bax as well as downstream caspase-8, caspase-9, and caspase-3 activity, and reversed impaired DJ-1 and anti-apoptotic Bcl-2 protein expression in Int-407 cells induced by ketoprofen. Similar to the in vitro results, SD rats treated with PCA before administration of ketoprofen exhibited decreased caspase-3 protein expression as well as oxidative damage, and impairment of the antioxidant system and DJ-1 protein expression in the GI mucosa were reversed. The administration of lansoprazole, a type of proton pump inhibitor (PPI), strongly inhibited ketoprofen-induced GI mucosal injuries via up-regulation of DJ-1, indicating that DJ-1 is essential for the dietary antioxidant- and PPI drug-mediated mechanism of ulcer therapy. These results suggest that DJ-1 could be a novel target for protection against ketoprofen-induced GI ulcers due to its antioxidant and anti-apoptosis characteristics.

Astroglial DJ-1 over-expression up-regulates proteins involved in redox regulation and is neuroprotective in vivo

DJ-1, a Parkinson's disease-associated protein, is strongly up-regulated in reactive astrocytes in Parkinson's disease. This is proposed to represent a neuronal protective response, although the mechanism has not yet been identified. We have generated a transgenic zebrafish line with increased astroglial DJ-1 expression driven by regulatory elements from the zebrafish GFAP gene. Larvae from this transgenic line are protected from oxidative stress-induced injuries as caused by MPP+, a mitochondrial complex I inhibitor shown to induce dopaminergic cells death. In a global label-free proteomics analysis of wild type and transgenic larvae exposed to MPP+, 3418 proteins were identified, in which 366 proteins were differentially regulated. In particular, we identified enzymes belonging to primary metabolism to be among proteins affected by MPP+ in wild type animals, but not affected in the transgenic line. Moreover, by performing protein profiling on isolated astrocytes we showed that an increase in astrocytic DJ-1 expression up-regulated a large group of proteins associated with redox regulation, inflammation and mitochondrial respiration. The majority of these proteins have also been shown to be regulated by Nrf2. These findings provide a mechanistic insight into the protective role of astroglial up-regulation of DJ-1 and show that our transgenic zebrafish line with astrocytic DJ-1 over-expression can serve as a useful animal model to understand astrocyte-regulated neuroprotection associated with oxidative stress-related neurodegenerative disease.

Pretreatment with Sodium Phenylbutyrate Alleviates Cerebral Ischemia/Reperfusion Injury by Upregulating DJ-1 Protein.

Oxidative stress and mitochondrial dysfunction play critical roles in ischemia/reperfusion (I/R) injury. DJ-1 is an endogenous antioxidant that attenuates oxidative stress and maintains mitochondrial function, likely acting as a protector of I/R injury. In the present study, we explored the protective effect of a possible DJ-1 agonist, sodium phenylbutyrate (SPB), against I/R injury by protecting mitochondrial dysfunction via the upregulation of DJ-1 protein. Pretreatment with SPB upregulated the DJ-1 protein level and rescued the I/R injury-induced DJ-1 decrease about 50% both in vivo and in vitro. SPB also improved cellular viability and mitochondrial function and alleviated neuronal apoptosis both in cell and animal models; these effects of SPB were abolished by DJ-1 knockdown with siRNA. Furthermore, SPB improved the survival rate about 20% and neurological functions, as well as reduced about 50% of the infarct volume and brain edema, of middle cerebral artery occlusion mice 23 h after reperfusion. Therefore, our findings demonstrate that preconditioning of SPB possesses a neuroprotective effect against cerebral I/R injury by protecting mitochondrial function dependent on the DJ-1 upregulation, suggesting that DJ-1 is a potential therapeutic target for clinical ischemic stroke.

Hypoxic preconditioning up-regulates DJ-1 protein expression in rat heart-derived H9c2 cells through the activation of extracellular-regulated kinase 1/2 pathway

Myocardial preconditioning is a powerful phenomenon that can attenuate ischemia/reperfusion-induced oxidant stress and elicit delayed cardioprotection. Its mechanisms involve activation of intracellular signaling pathways and up-regulation of the protective antioxidant proteins. DJ-1 protein, as a multifunctional intracellular protein, plays an important role in attenuating oxidant stress and promoting cell survival. In the present study, we investigated whether DJ-1 is up-regulated during the late phase of hypoxic preconditioning (HP) and the up-regulation of DJ-1 is mediated by extracellular-regulated kinase 1/2 (ERK1/2) signaling pathway. Rat heart-derived H9c2 cells were exposed to HP. Twenty-four hours later cells were subjected to hypoxia/reoxygenation (H/R) and then cell viability, lactate dehydrogenase (LDH), intracellular reactive oxygen species (ROS), ERK1/2 phosphorylation, and DJ-1 protein were measured appropriately. The results showed that HP efficiently attenuated H/R-induced viability loss and LDH leakage. In addition, HP promoted ERK1/2 activation, up-regulated DJ-1 protein expression, inhibited H/R induced the elevation of ROS. However, when ERK1/2 phosphorylation was specifically inhibited by U0126, the increase in DJ-1 expression occurring during HP was almost completely abolished and, as a result, the delayed cardioprotection induced by HP was abolished, and the inhibitory effect of HP on H/R-induced oxidant stress was also reversed. Furthermore, knocking down DJ-1 by siRNA attenuated the delayed cardioprotection induced by HP. Our data indicate that HP can up-regulate DJ-1 protein expression through the ERK1/2-dependent signaling pathway. Importantly, DJ-1 might be involved in the delayed cardioprotective effect of HP against H/R injury.

2′-Benzoyloxycinnamaldehyde-Mediated DJ-1 Upregulation Protects MCF-7 Cells from Mitochondrial Damage

2'-Benzoyloxycinnamaldehyde (BCA) is a promising antitumor agent which induces cancer cells apoptosis via reactive oxygen species (ROS) generation. BCA shows more effective antiproliferation in MDA-MB-435 than in MCF-7 breast cancer cells. DJ-1 has been known to protect cells against oxidative stress as an antioxidant because of its cysteine residues sensitive to oxidative stress. In the present study, we evaluated the mechanism of DJ-1 for cell protection from oxidative stress after BCA treatment in MCF-7 cell. BCA upregulates the expression of DJ-1 in MCF-7 cells. However, DJ-1 expression decreased continuously for 24 h after BCA treatment in MDA-MB-435 cells. DJ-1 knockdown sensitized MCF-7 cells to BCA, on the contrary, DJ-1 overexpression induced MDA-MB-435 cells less sensitive to BCA. Confocal microscopic observation showed that only in MCF-7 cells BCA increased the overlapped signal between mitochondria and DJ-1 protein. Mitochondrial membrane potential (MMP) was decreased in MDA-MB-435 cells by BCA, and DJ-1 overexpression inhibited BCA-induced MMP decrease in these cells. On the contrary, DJ-1 knockdown in MCF-7 induced MMP perturbation by BCA. These findings suggest that DJ-1 upregulation protects MCF-7 cells from BCA via inhibiting mitochondrial damage.

Consequences of DJ-1 upregulation following p53 loss and cell transformation

p53 is a tumor suppressor that responds to various stress signals by initiating cell-cycle arrest, senescence and apoptosis. Mutations of the p53 gene are found in over 50% of human tumors, highlighting the importance of p53 in tumor suppression. Numerous studies have reported on the interactions between p53, IGF-1-AKT and mTOR pathways as potentially explaining some of the tumor suppressive activities of p53. To further understand the basis of these interactions, we analyzed the involvement of DJ-1, an oncogene known to drive AKT-mediated cell survival, in the p53-AKT axis. In this study, we show that DJ-1 and p53 are tightly 'linked': p53 prevents the accumulation of DJ-1 protein, whereas loss of p53 leads to stabilization and enhancement of DJ-1 expression. Interestingly, this increase in DJ-1 level is only observed when p53 loss is accompanied by transformation of cells. Moreover, DJ-1 seems to be required for the enhanced activation of AKT observed in p53-deficient cells. Such observation confers a new property to DJ-1 associated to transforming-process to its oncogenic ability to drive AKT activation. We also show that DJ-1 is necessary for p53 activation following oxidative stress, suggesting the existence of a finely regulated loop between these two proteins in transformed cells. Finally, we demonstrate that in the absence of p53, DJ-1 is stabilized by ROS accumulation, and surprisingly seems to be required for this high intracellular ROS production. These data offer new insights into the regulation of DJ-1 and suggest that DJ-1 is a target of p53. Importantly, our study highlights that during transformation, DJ-1 is having a key role in the p53-regulated AKT pathway and p53-driven oxidative-stress response.

Myostatin inactivation induces a similar muscle molecular signature in double-muscled cattle as in mice

Myostatin (MSTN), a member of the TGF-β superfamily, is a negative regulator of skeletal muscle mass. We have previously shown that the cell survival/apoptosis pathway is a downstream target of MSTN loss-of-function in mice through the regulation of the expression or abundance of many survival and apoptotic factors. In this study, we used western-blot and quantitative PCR (qPCR) analyses to validate these novel downstream targets of MSTN in double-muscled (DM) cattle v. their controls including 260-day-old foetuses and adult cows from the INRA95 strain. MSTN loss-of-function in DM foetuses and DM cows resulted in a glycolytic shift of the muscles (e.g. upregulation of H-MyBP, PGM1 and SNTA1 and downregulation of H-FABP), activation of cell survival pathway through regulation of some components of the PI3K/Akt pathway (e.g. upregulation of DJ-1 and Gsk-3βser9/Gsk-3βtotal ratio and downregulation of PTEN) and upregulation of cell survival factors translationally controlled tumour protein (14-3-3E, Pink1). We also found a lower abundance of pro-apoptotic transcripts and/or proteins (Caspase-3, caspase-8, caspase-9, BID, ID2 and Daxx) and a higher expression of anti-apoptotic transcripts (Traf2 and Bcl2l2) in DM muscles. All together, these results are in favour of activation of the cell survival pathway and loss of apoptosis pathway within the muscles of DM animals. Alteration of both pathways may increase myonuclear or satellite cell survival, which is crucial for protein synthesis. This could contribute to muscle hypertrophy in DM foetuses and DM cows.

Regulation of astrocyte inflammatory responses by the Parkinson's disease‐associated gene DJ–1

The Parkinson's disease (PD)-associated gene DJ-1 mediates direct neuroprotection. The up-regulation of DJ-1 in reactive astrocytes also suggests a role in glia. Here we show that DJ-1 regulates proinflammatory responses in mouse astrocyte-rich primary cultures. When treated with a Toll-like receptor 4 agonist, the bacterial endotoxin lipopolysaccharide (LPS), Dj-1-knockout astrocytes generated >10 times more nitric oxide (NO) than littermate controls. Lentiviral reintroduction of DJ-1 restored the NO response to LPS. The enhanced NO production in Dj-1(-/-) astrocytes was mediated by a signaling pathway involving reactive oxygen species leading to specific hyperinduction of type II NO synthase [inducible NO synthase (iNOS)]. These effects coincided with significantly increased phosphorylation of p38 mitogen-activated protein kinase (MAPK), and p38(MAPK) inhibition suppressed NO production and iNOS mRNA and protein induction. Dj-1(-/-) astrocytes also induced the proinflammatory mediators cyclooxygenase-2 and interleukin-6 significantly more strongly, but not nerve growth factor. Finally, primary neuron cultures grown on Dj-1(-/-) astrocytes became apoptotic in response to LPS in an iNOS-dependent manner, directly demonstrating the neurotoxic potential of astrocytic DJ-1 deficiency. These findings identify DJ-1 as a regulator of proinflammatory responses and suggest that loss of DJ-1 contributes to PD pathogenesis by deregulation of astrocytic neuroinflammatory damage.

DJ-1 protects against dopamine toxicity

Parkinson's disease (PD) is a slowly progressive neurodegenerative disorder characterized by the loss of dopaminergic neurons. Dopamine is a highly toxic compound leading to generation of reactive oxygen species (ROS). DJ-1 mutations lead to early-onset inherited PD. Here, we show that DJ-1 protects against dopamine toxicity. Dopamine-exposure led to upregulation of DJ-1. Overexpression of DJ-1 increased cell resistance to dopamine toxicity and reduced intracellular ROS. Contrary effects were achieved when DJ-1 levels were reduced by siRNA. Similarly, in vivo striatal administration of 6-hydroxydopamine led to upregulation of DJ-1. Upregulation of DJ-1 was mediated by the MAP kinases pathway through activation of ERK 1, 2 in vitro and in vivo. Hence, oxidative stress, generated by free cytoplasmic dopamine, leads to upregulation of DJ-1 through the MAP kinases pathway. This mechanism elucidates how mutations in DJ-1 prompt PD and imply that modulation of DJ-1 may serve as a novel neuroprotective modality.