Blockade of monocyte-endothelial trafficking by transduced Tat-superoxide dismutase protein

PARK, SIN-HYE,SHIN, MIN JAE,KIM, DAE WON,PARK, JINSEU,CHOI, SOO YOUNG,KANG, YOUNG-HEE D.A. Spandidos 2016 INTERNATIONAL JOURNAL OF MOLECULAR MEDICINE Vol.37 No.2

<P>It has previously been suggested that reactive oxygen species (ROS) are involved in the pathogenesis of chronic inflammatory diseases, which entails the initial activation of pro-inflammatory cytokines to facilitate leukocyte transmigration. The present study investigated whether intracellular superoxide dismutase (SOD) suppressed monocyte endothelial trafficking and transmigration. Human umbilical vein endothelial cells (HUVECs) and THP-1 monocytes were activated by the cytokine tumor necrosis factor-α (TNF-α) in the absence and presence of cell-permeable transactivator of transcription (Tat)-SOD protein. External stimulation with SOD was conducted using endothelial cells and monocytes. Purified cell-permeable Tat-SOD, but not non-targeted SOD, at 1–3 <I>µ</I>M was transduced into endothelial cells in a time- and dose-dependent manner. Non-toxic Tat-SOD at ≤0.5 <I>µ</I>M, but not 1 <I>µ</I>M SOD, blocked the monocyte-endothelium interactions by inhibiting the TNF-α-induced stimulation of vascular cell adhesion molecule-1 (VCAM-1) in HUVECs and integrin β1 in THP-1 cells. Endothelial VCAM-1 induction by TNF-α was responsible for superoxide anion production being quenched by N-acetyl-cysteine and Tat-SOD. SOD treatment markedly inhibited superoxide anion production induced by TNF-α, but no inhibition of endothelial transmigration was noted. Tat-SOD prevented transendothelial monocyte migration by firmly localizing occludin-1, platelet/endothelial cell adhesion molecule-1 (PECAM-1) and vascular endothelial-cadherin present in paracellular junctions and inhibiting endothelial induction and activation of matrix-degrading membrane type-1 (MT-1) matrix metalloproteinase (MMP), MMP-2 and MMP-9. By contrast, treatment with 1 <I>µ</I>M SOD did not have such effects. Furthermore, transduced Tat-SOD hindered nuclear transactivation of nuclear factor-κB (NF-κB), modulating the induction of paracellular junction proteins and matrix-degrading MMP in TNF-α-stimulated HUVECs. Transduced Tat-SOD, but not external SOD, impeded cytokine-induced endothelial adhesion and the transmigration of monocytes. Thus, we suggest that transduced Tat-SOD qualifies as an atheroprotective agent against oxidation-driven and inflammation-associated atherosclerosis.</P>

Exoenzyme Tat-C3 Inhibits Association of Zymosan Particles, Phagocytosis, Adhesion, and Complement Binding in Macrophage Cells

최수영,Jinseu Park,김준섭,Kyeong-Cheon Jung,Hak-Ju Lee,Jong-Il Kim,김재봉,이재용,Jae-Bong Park 한국분자세포생물학회 2003 Molecules and cells Vol.16 No.2

Expression of Human Immunodeficiency Virus Type 1 Tat Proteins in Escherichia coli and Application to Study Tat Functions

Choi, Soo Young,Park, Jinseu,Kang, Young Hee,Lee, Yoon,Lee, Hangyu,Rhim, Hyangshuk The Korea Science and Technology Center 2000 BMB Reports Vol.33 No.4

The human immunodeficiency virus type 1 (HIV-1), transactivator of transcription (Tat), is one of the viral gene products that is essential for HIV-1 replication. The HIV-1 Tat protein regulates transcription from an HIV-1 long terminal repeat (LTR) and affects the gene expression of cellular proteins during infection. In order to develop an expression system to overexpress and simply purify HIV-1 Tat proteins, the HIV-1 Tat coding sequences that contain one or two exons were amplified using PCR and cloned into a pET vector, which contains a consecutive stretch of six histidine residues at the amino-terminus. The reconstituted vectors were overexpressed in the E. coli strain and the soluble recombinant proteins were purified to be homogeneity in a single step by Ni??-nitrilotriacetic acid Sepharose chromatography under nondenaturing conditions. Recombinant HIV-1 LTR promoter in a dose-dependent manner when introduced into mammalian cells. In addition, treatment of human endothelial cells with purified Tat proteins resulted in a significant increase in the level of vascular cell adhesion molecule-1 (VCAM-1) expression. These results indicate that the recombinant HIV-1 Tat proteins are active in transactivating viral and cellular described in this study will facilitate in characterizing the biological functions of the Tat proteins.

Transduction of Cu, Zn-superoxide dismutase mediated by an HIV-1 Tat protein basic domain into human chondrocytes

Kim, Hyun Ah,Won Kim, Dae,Park, Jinseu,Choi, Soo Young BioMed Central 2006 ARTHRITIS RESEARCH AND THERAPY Vol.8 No.4

<P>This study was performed to investigate the transduction of a full-length superoxide dismutase (SOD) protein fused to transactivator of transcription (Tat) into human chondrocytes, and to determine the regulatory function of transduced Tat-SOD in the inflammatory cytokine induced catabolic pathway. The pTat-SOD expression vector was constructed to express the basic domain of HIV-1 Tat as a fusion protein with Cu, Zn-SOD. We also purified histidine-tagged SOD without an HIV-1 Tat and Tat-GFP as control proteins. Cartilage samples were obtained from patients with osteoarthritis (OA) and chondrocytes were cultured in both a monolayer and an explant. For the transduction of fusion proteins, cells/explants were treated with a variety of concentrations of fusion proteins. The transduced protein was detected by fluorescein labeling, western blotting and SOD activity assay. Effects of transduced Tat-SOD on the regulation of IL-1 induced nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) mRNA expression was assessed by the Griess reaction and reverse transcriptase PCR, respectively. Tat-SOD was successfully delivered into both the monolayer and explant cultured chondrocytes, whereas the control SOD was not. The intracellular transduction of Tat-SOD into cultured chondrocytes was detected after 1 hours, and the amount of transduced protein did not change significantly after further incubation. SOD enzyme activity increased in a dose-dependent manner. NO production and iNOS mRNA expression, in response to IL-1 stimulation, was significantly down-regulated by pretreatment with Tat-SOD fusion proteins. This study shows that protein delivery employing the Tat-protein transduction domain is feasible as a therapeutic modality to regulate catabolic processes in cartilage. Construction of additional Tat-fusion proteins that can regulate cartilage metabolism favorably and application of this technology in <I>in vivo </I>models of arthritis are the subjects of future studies.</P>

Brain Succinic Semialdehyde Dehydrogenase : Reaction of Arginine Residues Connected with Catalytic Activities

Choi, Soo Young,Bahn, Jae Hoon,Park, Jinseu,Jin, Li Hua,Lee, Byung Ryong,Kim, Chung Kwon,Cho, Sung-Woo,Jeon, Seong Gyu,Cho, Yong Joon,Jang, Joong Sik,Kwon, Oh-Shin The Korea Science and Technology Center 2000 BMB Reports Vol.33 No.4

The succinic semialdehyde dehydrogenase from bovine brain was inactivated by treatment with phenylglyoxal, a reagent that specifically modifies arginine residues. The inhibition at various phenylglyoxal ocncentrations shows pseudo-first-order kinetics with an apparent second-order rate constant of 30 M-¹min-¹ for inactivation. Partial pretection against inactivation was provided by the coenzyme NAD+, but not by the substrate succinic semialdehyde. Spectrophotoetric studies indicated that complete inactivation of the enzyme resulted from the binding of 2 mol phenylglyoxal per mol of enzyme. These results suggest that essential arginine residues, located at or near the coenzyme-bindig site, are connected with the catalytic activity of brain succinic semialdehyde dehydrogenase.

Tat-indoleamine 2,3-dioxygenase 1 elicits neuroprotective effects on ischemic injury

박정환,Dae Won Kim,Min Jea Shin,Jinseu Park,한규형,이근욱,Jong Kook Park,Yeon Joo Choi,여현지,여은지,Eun Jeong Sohn,Hyoung-Chun Kim,Eun-Joo Shin,Eun-Joo Shin,김덕수,조용준,Won Sik Eum,최수영 생화학분자생물학회 2020 BMB Reports Vol.53 No.11

It is well known that oxidative stress participates in neuronal cell death caused production of reactive oxygen species (ROS). The increased ROS is a major contributor to the development of ischemic injury. Indoleamine 2,3-dioxygenase 1 (IDO-1) is involved in the kynurenine pathway in tryptophan metabolism and plays a role as an anti-oxidant. However, whether IDO-1 would inhibit hippocampal cell death is poorly known. Therefore, we explored the effects of cell permeable Tat-IDO-1 protein against oxidative stress-induced HT-22 cells and in a cerebral ischemia/reperfusion injury model. Transduced Tat-IDO-1 reduced cell death, ROS production, and DNA fragmentation and inhibited mitogen-activated protein kinases (MAPKs) activation in H2O2 exposed HT-22 cells. In the cerebral ischemia/ reperfusion injury model, Tat-IDO-1 transduced into the brain and passing by means of the blood-brain barrier (BBB) significantly prevented hippocampal neuronal cell death. These results suggest that Tat-IDO-1 may present an alternative strategy to improve from the ischemic injury.

Effects of long-term post-ischemic treadmill exercise on gliosis in the aged gerbil hippocampus induced by transient cerebral ischemia

Ahn, Ji Hyeon,Shin, Myoung Cheol,Park, Joon Ha,Kim, In Hye,Cho, Jeong-Hwi,Lee, Tae-Kyeong,Lee, Jae-Chul,Chen, Bai Hui,Shin, Bich Na,Tae, Hyun-Jin,Park, Jinseu,Choi, Soo Young,Lee, Yun Lyul,Kim, Dae Wo SPANDIDOS PUBLICATIONS 2017 MOLECULAR MEDICINE REPORTS Vol.15 No.6

<P>Therapeutic exercise is an integral component of the rehabilitation of patients who have suffered a stroke. The objective of the present study was to use immunohistochemistry to investigate the effects of post-ischemic exercise on neuronal damage or death and gliosis in the aged gerbil hippocampus following transient cerebral ischemia. Aged gerbils (male; age, 22–24 months) underwent ischemia and were subjected to treadmill exercise for 1 or 4 weeks. Neuronal death was detected in the stratum pyramidale of the hippocampal CA1 region and in the polymorphic layer of the dentate gyrus using cresyl violet and Fluoro-Jade B histofluorescence staining. No significant difference in neuronal death was identified following 1 or 4 weeks of post-ischemic treadmill exercise. However, post-ischemic treadmill exercise affected gliosis (the activation of astrocytes and microglia). Glial fibrillary acidic protein-immunoreactive astrocytes and ionized calcium binding adaptor molecule 1-immunoreactive microglia were activated in the CA1 and polymorphic layer of the dentate gyrus of the group without treadmill exercise. Conversely, 4 weeks of treadmill exercise significantly alleviated ischemia-induced astrocyte and microglial activation; however, 1 week of treadmill exercise did not alleviate gliosis. These findings suggest that long-term post-ischemic treadmill exercise following transient cerebral ischemia does not influence neuronal protection; however, it may effectively alleviate transient cerebral ischemia-induced astrocyte and microglial activation in the aged hippocampus.</P>

Enhanced Uptake of a Heterologous Protein with an HIV-1 Tat Protein Transduction Domains (PTD) at Both Termini

최수영,Jiyoon Ryu,Kyuhyung Han,Jinseu Park 한국분자세포생물학회 2003 Molecules and cells Vol.16 No.3

E. coli DNA 절단과 mutagenicity에 대한 동충하초의 영향

이완희(Wanhee Lee),권혁일(Hyeok Yil Kwon),박진서(Jinseu Park),최수영(Soo Young Choi),이길수(Kil Soo Lee) 한국독성학회 1999 Toxicological Research Vol.15 No.3

Cordyceps militaris is one of the parasitic fungi Cordyceps sp. and used for Chinese traditional herbal medicine. Some components of Cordyceps sinensis extract such as cordycepin, certain polysaccharides and even it's crude extract are significantly effective on the immune system, enzyme inhibition, reveal anticancer and antifungal activity in animal experimental system. Protecting activity of Cordyceps militaris extract (CME) against DNA damage and antimutagenic activity of CME were investigated in microbial test system and in DNA fragmentation experiment. CME inhibited not only chromosomal DNA damage in E. coli, but also daunorubicin-induced plasmid DNA damage. It was observed that DNA damage inhibition in vitro by CME resulted from the inactivation of daunorubicin by CME, but not from cellular DNA protection by CME. In vivo mutagenicity test system in E. coli, CME did not repair DNA damage induced by 4-nitroquinoline-n-oxide (4NQO), but it caused the 4NQO inactivation which leads to inhibit 4NQO-induced β-galactosidase synthesis in SOS-chromotest. From the correlation between the inhibition of in vitro DNA fragmentation with the inhibition of cellular DNA damage in vivo, it is suggested that CME has an inhibitory effect on the DNA damaging action of daunorubicin and on the mutagenicity of 4NQO in E. coli test system.

PEP-1-paraoxonase 1 fusion protein prevents cytokine-induced cell destruction and impaired insulin secretion in rat insulinoma cells

( Su Jin Lee ),( Hyung Kyung Kang ),( Yeon Joo Choi ),( Won Sik Eum ),( Jinseu Park ),( Soo Young Choi ),( Hyeok Yil Kwon ) 생화학분자생물학회(구 한국생화학분자생물학회) 2018 BMB Reports Vol.51 No.10

Pancreatic beta cell destruction and dysfunction induced by cytokines is a major cause of type 1 diabetes. Paraoxonase 1 (PON1), an arylesterase with antioxidant activity, has been shown to play an important role in preventing the development of diabetes in transgenic mice. However, no studies have examined the anti-diabetic effect of PON1 delivered to beta cells using protein transduction. In this study, we expressed the cell-permeable PON1 fused with PEP-1 protein transduction domain (PEP-1-PON1) to investigate whether transduced PEP-1-PON1 protects beta cells against cytokine-induced cytotoxicity. PEP-1-PON1 was effectively delivered to INS-1 cells and prevented cytokine-induced cell destruction in a dose-dependent manner. Transduced PEP-1-PON1 significantly reduced the levels of reactive oxygen species (ROS) and nitric oxide (NO), DNA fragmentation, and expression of inflammatory mediators, endoplasmic reticulum (ER) stress proteins, and apoptosis-related proteins in cytokine-treated cells. Moreover, transduced PEP-1-PON1 restored the decrease in basal and glucose-stimulated insulin secretion induced by cytokines. These data indicate that PEP-1-PON1 protects beta cells from cytokine-induced cytotoxicity by alleviating oxidative/nitrosative stress, ER stress, and inflammation. Thus, PEP-1-mediated PON1 transduction might be an effective method to reduce the extent of destruction and dysfunction of pancreatic beta cells in autoimmune diabetes. [BMB Reports 2018; 51(10): 538-543]