Tat-DJ-1 enhances cell survival by inhibition of oxidative stress, NF-κB and MAPK activation in HepG2 cells.

OBJECTIVES: To identify the protective effect of DJ-1 protein against oxidative stress-induced HepG2 cell death, we used cell-permeable wild type (WT) and a mutant (C106A Tat-DJ-1) protein. RESULTS: By using western blotting and fluorescence microscopy, we observed WT and C106A Tat-DJ-1 proteins were efficiently transduced into HepG2 cells. Transduced WT Tat-DJ-1 proteins increased cell survival and protected against DNA fragmentation and intracellular ROS generation levels in H2O2-exposed HepG2 cells. At the same time, transduced WT Tat-DJ-1 protein significantly inhibited NF-κB and MAPK (JNK and p38) activation as well as regulated the Bcl-2 and Bax expression levels. However, C106A Tat-DJ-1 protein did not show any protective effect against cell death responses in H2O2-exposed HepG2 cells. CONCLUSIONS: Oxidative stress-induced HepG2 cell death was significantly reduced by transduced WT Tat-DJ-1 protein, not by C106A Tat-DJ-1 protein. Thus, transduction of WT Tat-DJ-1 protein could be a novel strategy for promoting cell survival in situations of oxidative stress-induced HepG2 cell death.

Tat-DJ-1 inhibits oxidative stress-mediated RINm5F cell death through suppression of NF-κB and MAPK activation.

Oxidative stress is highly involved in the development of diabetes mellitus by destruction of pancreatic ß-cells. DJ-1 is an antioxidant protein and DJ-1 expression levels are known to be reduced in diabetes mellitus. Thus, we examined the effects of DJ-1 protein against oxidative stress-induced pancreatic ß-cell (RINm5F) death using cell permeable wild-type and mutant-type (C106A) Tat-DJ-1 proteins, which both efficiently transduced into RINm5F cells. Intracellular stability of wild-type Tat-DJ-1 persisted two times longer than C106A Tat-DJ-1. Wild-type Tat-DJ-1 protein markedly protected cells from hydrogen peroxide-induced toxicities such as cell death, reactive oxygen species generation, and DNA fragmentation. Further, wild-type Tat-DJ-1 protein significantly inhibited hydrogen peroxide-induced activation of mitogen-activated protein kinases and NF-κB signaling. On the other hand, C106A Tat-DJ-1 protein did not show the same protective effects. These results indicate that wild-type Tat-DJ-1 inhibits oxidative stress-induced cellular toxicity and activation of mitogen-activated protein kinases and NF-κB signals in RINm5F cells. These results suggest that wild-type Tat-DJ-1 protein may be a potential therapeutic agent against diabetes mellitus or toward the prevention of pancreatic ß-cell destruction.

Tat-antioxidant 1 protects against stress-induced hippocampal HT-22 cells death and attenuate ischaemic insult in animal model.

Oxidative stress-induced reactive oxygen species (ROS) are responsible for various neuronal diseases. Antioxidant 1 (Atox1) regulates copper homoeostasis and promotes cellular antioxidant defence against toxins generated by ROS. The roles of Atox1 protein in ischaemia, however, remain unclear. In this study, we generated a protein transduction domain fused Tat-Atox1 and examined the roles of Tat-Atox1 in oxidative stress-induced hippocampal HT-22 cell death and an ischaemic injury animal model. Tat-Atox1 effectively transduced into HT-22 cells and it protected cells against the effects of hydrogen peroxide (H2O2)-induced toxicity including increasing of ROS levels and DNA fragmentation. At the same time, Tat-Atox1 regulated cellular survival signalling such as p53, Bad/Bcl-2, Akt and mitogen-activate protein kinases (MAPKs). In the animal ischaemia model, transduced Tat-Atox1 protected against neuronal cell death in the hippocampal CA1 region. In addition, Tat-Atox1 significantly decreased the activation of astrocytes and microglia as well as lipid peroxidation in the CA1 region after ischaemic insult. Taken together, these results indicate that transduced Tat-Atox1 protects against oxidative stress-induced HT-22 cell death and against neuronal damage in animal ischaemia model. Therefore, we suggest that Tat-Atox1 has potential as a therapeutic agent for the treatment of oxidative stress-induced ischaemic damage.

PEP-1-PEA-15 protects against toxin-induced neuronal damage in a mouse model of Parkinson's disease.

BACKGROUND: PEA-15 is abundantly expressed in both neurons and astrocytes throughout the brain. It is a multifunctional protein with the ability to increase cell survival via anti-apoptotic and anti-proliferative properties. However, the function of PEA-15 in neuronal diseases such as Parkinson's disease (PD) remains unclear. In this study, we investigated the protective effects of PEA-15 on neuronal damage induced by MPP(+) in neuroblastoma SH-SY5Y and BV2 microglia cells and in a MPTP-induced PD mouse model using cell-permeable PEP-1-PEA-15. METHODS: PEP-1-PEA-15 was purified using affinity chromatography. Cell viability and DNA fragmentation were examined by MTT assay and TUNEL staining. Dopaminergic neuronal cell death in the animal model was examined by immunohistochemistry. RESULTS: PEP-1-PEA-15 transduced into the SH-SY5Y and BV2 cells in a time- and dose-dependent manner. Transduced PEP-1-PEA-15 protected against MPP(+)-induced toxicity by inhibiting intracellular ROS levels and DNA fragmentation. Further, it enhanced the expression levels of Bcl-2 and caspase-3 while reducing the expression levels of Bax and cleaved caspase-3. We found that PEP-1-PEA-15 transduced into the substantia nigra and prevented dopaminergic neuronal cell death in a MPTP-induced PD mouse. Also, we showed the neuroprotective effects in the model by demonstrating that treatment with PEP-1-PEA-15 ameliorated MPTP-induced behavioral dysfunctions and increased dopamine levels in the striatum. CONCLUSIONS: PEP-1-PEA-15 can efficiently transduce into cells and protects against neurotoxin-induced neuronal cell death in vitro and in vivo. GENERAL SIGNIFICANCE: These results demonstrate the potential for PEP-1-PEA-15 to provide a new strategy for protein therapy treatment of a variety of neurodegenerative diseases including PD.

Tat-CBR1 inhibits inflammatory responses through the suppressions of NF-κB and MAPK activation in macrophages and TPA-induced ear edema in mice.

Human carbonyl reductase 1 (CBR1) plays a crucial role in cell survival and protects against oxidative stress response. However, its anti-inflammatory effects are not yet clearly understood. In this study, we examined whether CBR1 protects against inflammatory responses in macrophages and mice using a Tat-CBR1 protein which is able to penetrate into cells. The results revealed that purified Tat-CBR1 protein efficiently transduced into Raw 264.7 cells and inhibited lipopolysaccharide (LPS)-induced cyclooxygenase-2 (COX-2), nitric oxide (NO) and prostaglandin E2 (PGE2) expression levels. In addition, Tat-CBR1 protein leads to decreased pro-inflammatory cytokine expression through suppression of nuclear transcription factor-kappaB (NF-κB) and mitogen activated protein kinase (MAPK) activation. Furthermore, Tat-CBR1 protein inhibited inflammatory responses in 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced skin inflammation when applied topically. These findings indicate that Tat-CBR1 protein has anti-inflammatory properties in vitro and in vivo through inhibition of NF-κB and MAPK activation, suggesting that Tat-CBR1 protein may have potential as a therapeutic agent against inflammatory diseases.

Anti-inflammatory effects of Tat-Annexin protein on ovalbumin-induced airway inflammation in a mouse model of asthma.

Chronic airway inflammation is a key feature of bronchial asthma. Annexin-1 (ANX1) is an anti-inflammatory protein that is an important modulator and plays a key role in inflammation. Although the precise action of ANX1 remains unclear, it has emerged as a potential drug target for inflammatory diseases such as asthma. To examine the protective effects of ANX1 protein on ovalbumin (OVA)-induced asthma in animal models, we used a cell-permeable Tat-ANX1 protein. Mice sensitized and challenged with OVA antigen had an increased amount of cytokines and eosinophils in their bronchoalveolar lavage (BAL) fluid. However, administration of Tat-ANX1 protein before OVA challenge significantly decreased the levels of cytokines (interleukin (IL)-4, IL-5, and IL-13) and BAL fluid in lung tissues. Furthermore, OVA significantly increased the activation of mitogen-activated protein kinase (MAPK) in lung tissues, whereas Tat-ANX1 protein markedly reduced phosphorylation of MAPKs such as extracellular signal-regulated protein kinase, p38, and stress-activated protein kinase/c-Jun N-terminal kinase. These results suggest that transduced Tat-ANX1 protein may be a potential protein therapeutic agent for the treatment of lung disorders including asthma.

Effects of silk solution against laminectomy-induced dural adhesion formation and inflammation in a rat model.

The authors investigated the effects of a silk solution against laminectomy-induced dural adhesion formation and inflammation in a rat model. They found that it significantly reduced postlaminectomy dural adhesion formation and inflammation. Dural adhesion formation, thought to be an inevitable consequence of laminectomy, is one of the most common complications following spinal surgery, and the authors' results indicate that the silk solution might be a potential novel therapeutic agent for dural adhesion formation.

PEP-1-PEA15 suppresses inflammatory responses by regulation of MAPK in macrophages and animal models.

Phosphoprotein enriched in astrocytes 15 (PEA15) plays a multi-functional role in neuronal cell survival, however the effects of PEA15 against inflammation have not been investigated yet. To examine the effects of PEP-1-PEA15 protein against lipopolysaccharide (LPS)-induced inflammatory responses in Raw 264.7 cells and in a 12-O-tetradecanoylphobol 13-acetate (TPA)-induced mouse model, we constructed and purified PEP-1-PEA15 protein, which can transduce into cells or tissues. PEP-1-PEA15 inhibited LPS-induced damage in cells including that caused by reactive oxygen species (ROS) production and DNA fragmentation. PEP-1-PEA15 also significantly suppressed activation of mitogen activated protein kinases (MAPKs), pro-inflammatory mediator proteins and various cytokines. In a TPA-induced mouse ear edema model, PEP-1-PEA15 significantly reduced ear weight and thickness as well as MAPK activation as well as the expression levels of COX-2, iNOS, IL-6, IL-1ß, and TNF-α. These results demonstrated that PEP-1-PEA15 showed anti-inflammatory effect in cells and animal model suggesting that this fusion protein protects cells or skin tissues from inflammatory response.

PEP­1­glutaredoxin 1 protects against hippocampal neuronal cell damage from oxidative stress via regulation of MAPK and apoptotic signaling pathways.

Oxidative stress is known to be a primary risk factor for neuronal diseases. Glutaredoxin (GLRX)­1, a redox­regulator of the thioredoxin superfamily, is known to exhibit an important role in cell survival via various cellular functions. However, the precise roles of GLRX1 in brain ischemia are still not fully understood. The present study investigated whether transduced PEP­1­GLRX1 protein has protective effects against oxidative stress in cells and in an animal model. Transduced PEP­1­GLRX1 protein increased HT­22 cell viability under oxidative stress and this fusion protein significantly reduced intracellular reactive oxygen species and levels of DNA damage. In addition, PEP­1­GLRX1 protein regulated RAC­a serine/threonine­protein kinase and mitogen­activated protein kinase signaling, in addition to apoptotic signaling including B cell lymphoma (Bcl)­2, Bcl­2 associated X, apoptosis regulator, pro­caspase­9 and p53 expression levels. In an ischemic animal model, it was verified that PEP­1­GLRX1 transduced into the Cornu Ammonis 1 region of the animal brain, where it markedly protected against ischemic injury. These results indicate that PEP­1­GLRX1 attenuates neuronal cell death resulting from oxidative stress in vitro and in vivo. Therefore, PEP­1­GLRX1 may exhibit a beneficial role in the treatment of neuronal disorders, including ischemic injury.

Tat-HSP22 inhibits oxidative stress-induced hippocampal neuronal cell death by regulation of the mitochondrial pathway.

Oxidative stress plays an important role in the progression of various neuronal diseases including ischemia. Heat shock protein 22 (HSP22) is known to protect cells against oxidative stress. However, the protective effects and mechanisms of HSP22 in hippocampal neuronal cells under oxidative stress remain unknown. In this study, we determined whether HSP22 protects against hydrogen peroxide (H2O2)-induced oxidative stress in HT-22 using Tat-HSP22 fusion protein. We found that Tat-HSP22 transduced into HT-22 cells and that H2O2-induced cell death, oxidative stress, and DNA damage were significantly reduced by Tat-HSP22. In addition, Tat-HSP22 markedly inhibited H2O2-induced mitochondrial membrane potential, cytochrome c release, cleaved caspase-3, and Bax expression levels, while Bcl-2 expression levels were increased in HT-22 cells. Further, we showed that Tat-HSP22 transduced into animal brain and inhibited cleaved-caspase-3 expression levels as well as significantly inhibited hippocampal neuronal cell death in the CA1 region of animals in the ischemic animal model. In the present study, we demonstrated that transduced Tat-HSP22 attenuates oxidative stress-induced hippocampal neuronal cell death through the mitochondrial signaling pathway and plays a crucial role in inhibiting neuronal cell death, suggesting that Tat-HSP22 protein may be used to prevent oxidative stress-related brain diseases including ischemia.

Effects of PEP-1-FK506BP on cyst formation in polycystic kidney disease.

Polycystic kidney disease (PKD) is one of the most common inherited disorders, involving progressive cyst formation in the kidney that leads to renal failure. FK506 binding protein 12 (FK506BP) is an immunophilin protein that performs multiple functions, including regulation of cell signaling pathways and survival. In this study, we determined the roles of PEP-1-FK506BP on cell proliferation and cyst formation in PKD cells. Purified PEP-1-FK506BP transduced into PKD cells markedly inhibited cell proliferation. Also, PEP-1-FK506BP drastically inhibited the expression levels of p-Akt, p-p70S6K, p-mTOR, and p-ERK in PKD cells. In a 3D-culture system, PEP-1-FK506BP significantly reduced cyst formation. Furthermore, the combined effects of rapamycin and PEP-1-FK506BP on cyst formation were markedly higher than the effects of individual treatments. These results suggest that PEP-1-FK506BP delayed cyst formation and could be a new therapeutic strategy for renal cyst formation in PKD. [BMB Reports 2017; 50(9): 460-465].

Transduced Tat-DJ-1 protein inhibits cytokines-induced pancreatic RINm5F cell death.

Loss of pancreatic ß-cells by oxidative stress or cytokines is associated with diabetes mellitus (DM). DJ-1 is known to as a multifunctional protein, which plays an important role in cell survival. We prepared cell permeable wild type (WT) and mutant type (M26I) Tat-DJ-1 proteins to investigate the effects of DJ-1 against combined cytokines (IL-1ß, IFN-γ and TNF-α)-induced RINm5F cell death. Both Tat-DJ-1 proteins were transduced into RINm5F cells. WT Tat-DJ-1 proteins significantly protected against cell death from cytokines by reducing intracellular toxicities. Also, WT Tat-DJ-1 proteins markedly regulated cytokines-induced pro- and anti-apoptosis proteins. However, M26I Tat-DJ-1 protein showed relatively low protective effects, as compared to WT Tat-DJ-1 protein. Our experiments demonstrated that WT Tat-DJ-1 protein protects against cytokine-induced RINm5F cell death by suppressing intracellular toxicities and regulating apoptosisrelated protein expression. Thus, WT Tat-DJ-1 protein could potentially serve as a therapeutic agent for DM and cytokine related diseases. [BMB Reports 2016; 49(5): 297-302].

Protective effects of Tat-NQO1 against oxidative stress-induced HT-22 cell damage, and ischemic injury in animals.

Oxidative stress is closely associated with various diseases and is considered to be a major factor in ischemia. NAD(P)H:quinone oxidoreductase 1 (NQO1) protein is a known antioxidant protein that plays a protective role in various cells against oxidative stress. We therefore investigated the effects of cell permeable Tat-NQO1 protein on hippocampal HT-22 cells, and in an animal ischemia model. The Tat-NQO1 protein transduced into HT-22 cells, and significantly inhibited against hydrogen peroxide (H2O2)-induced cell death and cellular toxicities. Tat-NQO1 protein inhibited the Akt and mitogen activated protein kinases (MAPK) activation as well as caspase-3 expression levels, in H2O2 exposed HT-22 cells. Moreover, Tat-NQO1 protein transduced into the CA1 region of the hippocampus of the animal brain and drastically protected against ischemic injury. Our results indicate that Tat-NQO1 protein exerts protection against neuronal cell death induced by oxidative stress, suggesting that Tat-NQO1 protein may potentially provide a therapeutic agent for neuronal diseases. [BMB Reports 2016; 49(11): 617-622].

PEP-1-GSTpi protein enhanced hippocampal neuronal cell survival after oxidative damage.

Reactive oxygen species generated under oxidative stress are involved in neuronal diseases, including ischemia. Glutathione S-transferase pi (GSTpi) is a member of the GST family and is known to play important roles in cell survival. We investigated the effect of GSTpi against oxidative stress-induced hippocampal HT-22 cell death, and its effects in an animal model of ischemic injury, using a cell-permeable PEP-1-GSTpi protein. PEP-1-GSTpi was transduced into HT-22 cells and significantly protected against H2O2-treated cell death by reducing the intracellular toxicity and regulating the signal pathways, including MAPK, Akt, Bax, and Bcl-2. PEP-1-GSTpi transduced into the hippocampus in animal brains, and markedly protected against neuronal cell death in an ischemic injury animal model. These results indicate that PEP-1-GSTpi acts as a regulator or an antioxidant to protect against oxidative stressinduced cell death. Our study suggests that PEP-1-GSTpi may have potential as a therapeutic agent for the treatment of ischemia and a variety of oxidative stress-related neuronal diseases. [BMB Reports 2016; 49(7): 382-387].

Tat-PRAS40 prevent hippocampal HT-22 cell death and oxidative stress induced animal brain ischemic insults.

Proline rich Akt substrate (PRAS40) is a component of mammalian target of rapamycin complex 1 (mTORC1) and is known to play an important role against reactive oxygen species-induced cell death. However, the precise function of PRAS40 in ischemia remains unclear. Thus, we investigated whether Tat-PRAS40, a cell-permeable fusion protein, has a protective function against oxidative stress-induced hippocampal neuronal (HT-22) cell death in an animal model of ischemia. We showed that Tat-PRAS40 transduced into HT-22 cells, and significantly protected against cell death by reducing the levels of H2O2 and derived reactive species, and DNA fragmentation as well as via the regulation of Bcl-2, Bax, and caspase 3 expression levels in H2O2 treated cells. Also, we showed that transduced Tat-PARS40 protein markedly increased phosphorylated RRAS40 expression levels and 14-3-3σ complex via the Akt signaling pathway. In an animal ischemia model, Tat-PRAS40 effectively transduced into the hippocampus in animal brain and significantly protected against neuronal cell death in the CA1 region. We showed that Tat-PRAS40 protein effectively transduced into hippocampal neuronal cells and markedly protected against neuronal cell damage. Therefore, we suggest that Tat-PRAS40 protein may be used as a therapeutic protein for ischemia and oxidative stress-induced brain disorders.

Protective effects of PEP-1-Catalase on stress-induced cellular toxicity and MPTP-induced Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disability caused by a decrease of dopaminergic neurons in the substantia nigra (SN). Although the etiology of PD is not clear, oxidative stress is believed to lead to PD. Catalase is antioxidant enzyme which plays an active role in cells as a reactive oxygen species (ROS) scavenger. Thus, we investigated whether PEP-1-Catalase protects against 1-methyl-4-phenylpyridinium (MPP+) induced SH-SY5Y neuronal cell death and in a 1-methyl- 4-phenyl-1,2,3,6-trtrahydropyridine (MPTP) induced PD animal model. PEP-1-Catalase transduced into SH-SY5Y cells significantly protecting them against MPP+-induced death by decreasing ROS and regulating cellular survival signals including Akt, Bax, Bcl-2, and p38. Immunohistochemical analysis showed that transduced PEP-1-Catalase markedly protected against neuronal cell death in the SN in the PD animal model. Our results indicate that PEP-1-Catalase may have potential as a therapeutic agent for PD and other oxidative stress related diseases.

Effects of low doses of Tat-PIM2 protein against hippocampal neuronal cell survival.

Oxidative stress is considered a major factor in various neuronal diseases including ischemia-reperfusion injury. Proviral Integration Moloney 2 (PIM2) proteins, one of the families of PIM kinases, play crucial roles in cell survival. However, the functions of PIM2 protein against ischemia are not understood. Therefore, the protective effects of PIM2 against oxidative stress-induced hippocampal HT22 cell death and brain ischemic injury were evaluated using Tat-PIM2, a cell permeable fusion protein. Tat-PIM2 protein transduced into hippocampal HT22 cells. Low doses of transduced Tat-PIM2 protein protected against oxidative stress-induced cell death including DNA damage and markedly inhibited the activation of mitogen activated protein kinase (MAPKs), NF-κB and the expression levels of Bax protein. Furthermore, Tat-PIM2 protein transduced into the CA1 region of the hippocampus and significantly prevented neuronal cell death in an ischemic insult animal model. These results indicated that low doses of Tat-PIM2 protein protects against oxidative stress-induced neuronal cell death, suggesting low doses of Tat-PIM2 protein provides a potential therapeutic agent against oxidative stress-induced neuronal diseases including ischemia.

Tat-biliverdin reductase A inhibits inflammatory response by regulation of MAPK and NF-κB pathways in Raw 264.7 cells and edema mouse model.

Reactive oxygen species (ROS) accumulation induces oxidative stress and cell damage, which then activates several signaling pathways and triggers inflammatory response. Biliverdin is a natural product of heme metabolism which is converted to bilirubin by the enzyme biliverdin reductase A (BLVRA) which also plays a role in antioxidant activity via the ROS scavenging activity of bilirubin. In this study, we examined the anti-inflammatory and anti-apoptotic effects of Tat-BLVRA protein on lipopolysaccharide (LPS)-induced inflammation in Raw 264.7 macrophage cells. Transduction of Tat-BLVRA protein into Raw 264.7 cells and mice ear tissue was tested by Western blot analysis and immunohistochemical analysis. Tat-BLVRA protein was effective in inhibiting mitogen activated protein kinases (MAPKs), Akt and NF-κB activation, intracellular ROS production and DNA fragmentation. Also, Tat-BLVRA protein significantly inhibited the expression of cytokines, COX-2, and iNOS. In a 12-O-tetradecanoylphobol 13-acetate (TPA)-induced mouse model, mice ears treated with Tat-BLVRA protein showed decreased ear thickness and weight, as well as inhibited MAPKs activation and cytokine expression. Thus we suggested that Tat-BLVRA protein may provide an effective therapeutic agent for inflammatory skin diseases.

Transduced PEP-1-PON1 proteins regulate microglial activation and dopaminergic neuronal death in a Parkinson's disease model.

Parkinson's disease (PD) is an oxidative stress-mediated neurodegenerative disorder caused by selective dopaminergic neuronal death in the midbrain substantia nigra. Paraoxonase 1 (PON1) is a potent inhibitor of low-density lipoprotein (LDL) and high-density lipoprotein (HDL) against oxidation by destroying biologically active phospholipids with potential protective effects against oxidative stress-induced inflammatory disorders. In a previous study, we constructed protein transduction domain (PTD) fusion PEP-1-PON1 protein to transduce PON1 into cells and tissue. In this study, we examined the role of transduced PEP-1-PON1 protein in repressing oxidative stress-mediated inflammatory response in microglial BV2 cells after exposure to lipopolysaccharide (LPS). Moreover, we identified the functions of transduced PEP-1-PON1 proteins which include, mitigating mitochondrial damage, decreasing reactive oxidative species (ROS) production, matrix metalloproteinase-9 (MMP-9) expression and protecting against 1-methyl-4-phenylpyridinium (MPP(+))-induced neurotoxicity in SH-SY5Y cells. Furthermore, transduced PEP-1-PON1 protein reduced MMP-9 expression and protected against dopaminergic neuronal cell death in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mice model. Taken together, these results suggest a promising therapeutic application of PEP-1-PON1 proteins against PD and other inflammation and oxidative stress-related neuronal diseases.

Tat-glyoxalase protein inhibits against ischemic neuronal cell damage and ameliorates ischemic injury.

Methylglyoxal (MG), a metabolite of glucose, is the major precursor of protein glycation and induces apoptosis. MG is associated with neurodegeneration, including oxidative stress and impaired glucose metabolism, and is efficiently metabolized to S-D-lactoylglutathione by glyoxalase (GLO). Although GLO has been implicated as being crucial in various diseases including ischemia, its detailed functions remain unclear. Therefore, we investigated the protective effect of GLO (GLO1 and GLO2) in neuronal cells and an animal ischemia model using Tat-GLO proteins. Purified Tat-GLO protein efficiently transduced into HT-22 neuronal cells and protected cells against MG- and H2O2-induced cell death, DNA fragmentation, and activation of caspase-3 and mitogen-activated protein kinase. In addition, transduced Tat-GLO protein increased D-lactate in MG- and H2O2-treated cells whereas glycation end products (AGE) and MG levels were significantly reduced in the same cells. Gerbils treated with Tat-GLO proteins displayed delayed neuronal cell death in the CA1 region of the hippocampus compared with a control. Furthermore, the combined neuroprotective effects of Tat-GLO1 and Tat-GLO2 proteins against ischemic damage were significantly higher than those of each individual protein. Those results demonstrate that transduced Tat-GLO protein protects neuronal cells by inhibiting MG- and H2O2-mediated cytotoxicity in vitro and in vivo. Therefore, we suggest that Tat-GLO proteins could be useful as a therapeutic agent for various human diseases related to oxidative stress including brain diseases.