Long-term use of clopidogrel versus ticagrelor or prasugrel in patients with acute myocardial infarction after percutaneous coronary intervention.

BACKGROUND AND OBJECTIVES: To compare the long-term clinical outcomes of dual antiplatelet therapy (DAPT) with clopidogrel and DAPT with ticagrelor or prasugrel in patients with acute myocardial infarction (AMI) who underwent coronary intervention. METHODS: Between November 2011 and December 2015, a total of 13,104 patients with AMI were enrolled in the Korea Acute Myocardial Infarction Registry-National Institutes of Health (KAMIR-NIH) registry. Among them, 4,696 patients who received DAPT for more than 24 months were categorized into two groups: the clopidogrel group (n = 4,053) and ticagrelor or prasugrel group (n = 643). Propensity score matching (PSM) was used to reduce the bias due to confounding variables. Following PSM, the impacts of P2Y12 inhibitors on the clinical outcomes in both groups were compared during a 36-month clinical follow-up period. RESULTS: There were no significant differences in clinical outcomes in terms of cardiac death (7.1% vs. 9.7%, p = 0.101), stroke (1.4% vs. 1.0%, p = 0.436), major bleeding (0.5% vs. 0.8%, p = 0.478), major adverse cardiac events (MACE) (21.6% vs. 20.5%, p = 0.626), and net adverse cardiac event (NACE) (22.1% vs. 21.3%, p = 0.731) between the groups. The ticagrelor or prasugrel group had a lower incidence of recurrent percutaneous coronary intervention (PCI) (12.2% vs. 7.6%, p = 0.006) than the clopidogrel group. However, no differences were observed in the cumulative incidences of 3-year NACE between the ticagrelor or prasugrel and clopidogrel groups. CONCLUSIONS: Cumulative incidences of long-term NACE did not differ between the two groups. Therefore, the type and duration of DAPT should be customized for each patient with AMI.

Impact of a screening protocol for blood pressure level for hypertension in the Korean community health survey.

PURPOSE: A community program is an efficient model for improving the management of chronic diseases such as hypertension, diabetes, and dyslipidemia. A specific blood pressure (BP) measurement protocol was developed for community settings in which BP was measured by the interviewer at the interviewee's home. MATERIALS AND METHODS: In the 2018 Korean Community Health Survey, BP was measured twice at a five-minute interval after a five-minute resting period at the beginning of the survey. In 2019, BP was measured at the end of the survey after a two-minute rest and was obtained as three measurements at one-minute intervals. As factors related to BP level, stressful stimuli within 30 min before BP measurement such as smoking, caffeine, and/or exercise; duration of rest; and survey year were analysed. RESULTS: The mean age of participants was 55.2 years, and females accounted for 55.4% of the participants (n = 399,838). Stressful stimuli were observed in 21.9% of the participants in 2018 (n = 188,440) and 11.3% in 2019 (n = 211,398). Duration of rest was 0 min (2.1%), two minutes (55.0%), and five minutes (47.9%). When adjusted for age, sex, body mass index, antihypertensive medication, the arm of measurement, survey year (beta= -4.092), stressful stimuli (beta = 0.834), and resting time (beta = -1.296 per one minute of rest) were significant factors for mean systolic BP. A two-minute rest was not a significant factor in mean BP. The differences in adjusted mean systolic BPs were significant for rest times of five minutes vs. two minutes (3.1 mmHg, p < 0.0001), for stressful stimuli (0.8 mmHg, p < 0.0001), and for survey year (127.8 ± 0.2 mmHg vs. 122.2 ± 0.3 mmHg for 2018 vs. 2019, p < 0.0001). CONCLUSION: For the community-based home visit survey, avoidance of stressful stimuli, five-minute rest, and allocation of BP measurement in the last part of the survey was useful for obtaining a stable BP level.

Super-Strong Carbon Nanotube Fibers Achieved by Engineering Gas Flow and Postsynthesis Treatment.

Carbon nanotube fibers (CNTFs) are directly spun from a floating-catalyst chemical vapor deposition apparatus using gas-phase carbon and an iron nanocatalyst. The essential synthesis and post-treatment factors that affect the strength of CNTFs are investigated to obtain CNTFs with greater strength than those of any previously reported high-performance fibers. The key factors optimized included the degree of rotational flow inside the reactor, the ratio of the starting materials, and the postsynthesis treatment conditions. The formation of rotational gas flow inside the reactor was confirmed by computational fluid dynamics simulations, and the feed ratio of the starting materials was optimized through response surface methodology. In addition, a reproducible and highly efficient postsynthesis treatment method was established. Pristine CNTFs with a high specific strength (SS) (average 2.2 N/tex, max. 2.3 N/tex) were synthesized through decreased rotational flow and optimization of the CNTF synthesis conditions. To improve the SS of the CNTFs further, we adopted an acid wet-stretching method that included washing and heat treatment. This drastically increased the SS of the CNTFs (average 5.5 N/tex, max. 6.4 N/tex) because of the decrease in the volume of the pores between the CNT bundles.

The Current Status of Percutaneous Coronary Intervention in Korea: Based on Year 2014 & 2016 Cohort of Korean Percutaneous Coronary Intervention (K-PCI) Registry.

BACKGROUND AND OBJECTIVES: In this second report from Korean percutaneous coronary intervention (K-PCI) registry, we sought to describe the updated information of PCI practices and Korean practice pattern of PCI (KP3). METHODS: In addition to K-PCI registry of 2014, new cohort of 2016 from 92 participating centers was appended. Demographic and procedural information, as well as in-hospital outcomes, of PCI was collected using a web-based reporting system. KP3 class C was defined as any strategy with less evidence from randomized trials and more aggressive for PCI than medical therapy or bypass-surgery. RESULTS: In 2016, total 48,823 PCI procedures were performed at 92 participating centers. Mean age of the patients was 65.7±11.6 years, and 71.7% were males. Overall patient characteristics and PCI practices in 2016 were similar to those in 2014. The biggest change was the decrease in the in-hospital occurrence of myocardial infarction (MI;1.6%→0.7%, p<0.001). Many associations between PCI volumes and demographic/procedural characteristics observed in 2014 have disappeared. The median of door-to-balloon time was 62 minutes, and 83.3% of ST-elevation MI patients received primary PCI within 90 minutes, while the median of total ischemic time was 168 minutes and patients who had total ischemic time within 120 and 180 minutes were 29.1% and 54.1%, respectively. The proportion of KP3 class C cases in non-acute coronary syndrome patients decreased from 13.5% in 2014 to 12.1% in 2016 (p<0.001). CONCLUSIONS: In this second report from K-PCI registry, we described the current practices of PCI and changes from 2014 to 2016 in Korea.

Hypoxia/Reoxygenation-Preconditioned Human Bone Marrow-Derived Mesenchymal Stromal Cells Rescue Ischemic Rat Cortical Neurons by Enhancing Trophic Factor Release.

Bone marrow-derived mesenchymal stromal cells (BM-MSCs) represent a promising tool for stem cell-based therapies. However, the majority of MSCs fail to reach the injury site and have only minimal therapeutic effect. In this study, we assessed whether hypoxia/reoxygenation (H/R) preconditioning of human BM-MSCs could increase their functional capacity and beneficial effect on ischemic rat cortical neurons. Human BM-MSCs were cultured under hypoxia (1% O2) and with long-term reoxygenation for various times to identify the optimal conditions for increasing their viability and proliferation. The effects of H/R preconditioning on the BM-MSCs were assessed by analyzing the expression of prosurvival genes, trophic factors, and cell migration assays. The functionally improved BM-MSCs were cocultured with ischemic rat cortical neurons to compare with normoxic cultured BM-MSCs. Although the cell viability and proliferation of BM-MSCs were reduced after 1 day of hypoxic culture (1% O2), when this was followed by 5-day reoxygenation, the BM-MSCs recovered and multiplied extensively. The immunophenotype and trilineage differentiation of BM-MSCs were also maintained under this H/R preconditioning. In addition, the preconditioning enhanced the expression of prosurvival genes, the messenger RNA (mRNA) levels of various trophic factors and migration capacity. Finally, coculture with the H/R-preconditioned BM-MSCs promoted the survival of ischemic rat cortical neurons. H/R preconditioning of BM-MSCs increases prosurvival signals, trophic factor release, and cell migration and appears to increase their ability to rescue ischemic cortical neurons. This optimized H/R preconditioning procedure could provide the basis for a new strategy for stem cell therapy in ischemic stroke patients.

Sphingomonas kyeonggiense sp. nov., isolated from soil of a ginseng field.

A Gram-staining negative bacterium, THG-DT81(T), which was isolated from soil of a ginseng field, was investigated using a polyphasic taxonomic approach. Cells were oxidase- and catalase-positive, aerobic, rod-shaped and motile with one polar flagellum. Strain THG-DT81(T) grew optimally at pH 7.0 and in the absence of NaCl on trypticase soy agar. Its optimum growth temperature was 25-28 °C. Phylogenetic analysis based on 16S rRNA gene sequence showed that strain THG-DT81(T) belongs to the family Sphingomonadaceae and was related to Sphingomonas pituitosa EDIV(T) (98.0 % similarity), S. leidyi ATCC 15260(T) (97.8 %), S. trueperi LMG 2142(T) (97.1 %), S. azotifigens NBRC 15497(T) (97.1 %), S. koreensis JSS26 (T) (97.1 %) and S. dokdonensis DS-4(T) (97.0 %). Strain THG-DT81(T) contained Q-10 as the predominant ubiquinone and C18:1 ω7c and C16:0 as the major fatty acids. The G+C content of the genomic DNA was determined to be 66.8 mol %. The major component in the polyamine pattern was identified as sym-homospermidine. The major polar lipids detected in strain THG-DT81(T) were sphingoglycolipid, phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol and phosphatidylcholine. The DNA-DNA relatedness values of the strain THG-DT81(T) and its closest phylogenetically neighbors were below 21 %. The phenotypic characteristics and genotypic data demonstrated the affiliation of strain THG-DT81(T) to the genus Sphingomonas. On the basis of the polyphasic taxonomic data presented, strain THG-DT81(T) is described as a novel species of genus Sphingomonas, for which the name Sphingomonas kyeonggiense sp. nov. is proposed. The type strain is THG-DT81(T) (= KACC 17173(T) = JCM 18825(T)).

Taibaiella koreensis sp. nov., isolated from soil of a ginseng field.

A Gram-staining-negative, strictly aerobic, motile (by gliding), non-spore-forming and rod-shaped bacterial strain, designated THG-DT86(T), was isolated from soil of a ginseng field of Pocheon province in the Republic of Korea and its taxonomic position was investigated by a polyphasic approach. Growth occurred at 10-35 °C, at pH 6.5-8.5 and with 0-1.5 % (w/v) NaCl on trypticase soy agar. Flexirubin-type pigments were found to be present. On the basis of 16S rRNA gene sequence similarity, strain THG-DT86(T) was shown to belong to the genus Taibaiella and was related to Taibaiella smilacinae PTJT-5(T) (95.3 %). The G+C content of the genomic DNA was 50.1 mol%. The only isoprenoid quinone detected in strain THG-DT86(T) was menaquinone-7 (MK-7) and the only polyamine was homospermidine. The predominant fatty acids were iso-C15 : 0, iso-C17 : 0 3-OH, C16 : 0, iso-C15 : 1 G and iso-C17 : 0, and the major polar lipids were phosphatidylethanolamine, an unidentified aminophosphoglycolipid and an unidentified aminophospholipid. Phenotypic data and phylogenetic inference supported the affiliation of strain THG-DT86(T) to the genus Taibaiella, and a number of biochemical tests differentiated strain THG-DT86(T) from the recognized species of the genus Taibaiella. Therefore, the novel isolate represents a novel species, for which the name Taibaiella koreensis sp. nov. is proposed, with THG-DT86(T) as the type strain ( = KACC 17171(T) = JCM 18823(T)).

Dyella kyungheensis sp. nov., isolated from soil of a cornus fruit field.

A Gram-reaction-negative, aerobic, motile by one polar flagellum, yellow-pigmented, rod-shaped bacterium, designated strain THG-B117(T), was isolated from soil of a cornus fruit field of Hoengseong province in South Korea and its taxonomic position was investigated by a polyphasic study. Strain THG-B117(T) grew well at 25-30 °C and at pH 6.0-8.0 in the absence of NaCl on nutrient agar. On the basis of 16S rRNA gene sequence similarity, strain THG-B117(T) was shown to belong to the family Xanthomonadaceae and be related to Dyella japonica XD53(T) (98.7 % similarity), Dyella terrae JS14-6(T) (98.0 %), Dyella koreensis BB4(T) (96.9 %), Dyella soli JS12-10(T) (96.9 %) and Dyella thiooxydans ATSB10(T) (96.7 %). DNA-DNA hybridization experiments showed that DNA relatedness between strain THG-B117(T) and its phylogenetically closest neighbours was below 45.1 %. The G+C content of the genomic DNA of strain THG-B117(T) was 64.8 mol%. The major polar lipids were phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, unidentified aminolipids, unidentified aminophospholipids and unidentified phospholipids. Phenotypic and chemotaxonomic data (major ubiquinone was Q-8, and major fatty acids were iso-C15 : 0, iso-C16 : 0, iso-C17 : 0 and iso-C17 : 1ω9c) supported the affiliation of strain THG-B117(T) with the genus Dyella. The results of physiological and biochemical tests suggested that strain THG-B117(T) was different genotypically and phenotypically from recognized species of the genus Dyella, and represents a novel species of this genus. The name Dyella kyungheensis sp. nov. is proposed, with the type strain THG-B117(T) ( = KACC 16981(T) = JCM 18747(T)).

Hair growth activity of Crataegus pinnatifida on C57BL/6 mouse model.

Crataegus pinnatifida has a long history of use in traditional oriental herbal medicine to stimulating digestion and improving blood circulation. Based on nutrition of hair, the present study was conducted to assess the effect of C. pinnatifida extract on hair growth using mouse model and its mechanisms of action. The C. pinnatifida extract containing the contents of total polyphenol of 5.88□0.82 g gallic acid/100 g extract and proanthocyanidin of 9.15□1.58 mg cyaniding chloride/100 g extract was orally administered daily at a dosage of 50 mg/kg weight to the 7-week-old C57BL/6 mice in telogen. The C. pinnatifida extract promoted hair growth by inducing anagen phase in mice in telogen, reflected by color of skin, thickness of hair shaft, and density of hair. The ratio of anagento telogen was determined by shape of hair follicles in vertically sectioned slide and increased by oral administration of C. pinnatifida extract. The number and the size of hair follicles were also enlarged, indicating anagen phase induction. The proliferation of human dermal papilla cells (hDPC) was accelerated by addition of C. pinnatifida extract, which activated the signaling of mitogen-activated protein kinases (Erk, p-38, and JNK) and Akt. Moreover, the ratio of Bcl-2/Bax as the determinant of cell fate was also raised in skin. These results suggest that the C. pinnatifida extract promotes hair growth by inducing anagen phase, which might be mediated by the activation of cellular signalings that enhance the survival of cultured hDPC and the increase of the ratio of Bcl-2 to Bax that protects cells against cell death.

Gene expression profiling of murine hepatic steatosis induced by tamoxifen.

Tamoxifen is an antiestrogenic agent used widely in the treatment of estrogen receptor-positive breast cancer. However, hepatic steatosis has been reported during clinical trials of tamoxifen. To explore the mechanism responsible for this tamoxifen-induced hepatic steatosis, we used microarray analysis to profile the gene expression pattern of mouse liver after tamoxifen treatment. Tamoxifen was administered orally as a single dose of 10mg/kg (low dose), 50mg/kg (medium dose), or 100mg/kg (high dose) to C57BL/6 mice, and the livers were removed 2h, 4h, 8h, and 24h later. From microarray data obtained from the liver samples, 414 genes were selected as tamoxifen-responsive genes (P<0.05, two-way ANOVA; cutoff ≥ 1.5-fold response). These genes were classified into three groups: 308 of the 414 genes showed a time-dependent response, nine genes showed a dose-dependent response, and 97 genes showed a time- and dose-dependent response. Most of the 308 time-dependent-responsive genes were associated predominantly with the biological processes involved in lipid metabolism. Overrepresented transcription factor binding site analysis showed that the following nuclear receptors that are important in lipid and carbohydrate metabolism were overrepresented: the androgen receptor (AR), nuclear receptor subfamily 2 group F member 1 (NR2F1), hepatocyte nuclear factor 4α (HNF4α), and retinoic acid receptor-related orphan receptor alpha 1 (RORα1). Reporter gene analysis further revealed that tamoxifen repressed the 5α-dihydrotestosterone-induced activation of the AR and the intrinsic transactivation function of RORα1, HNF4α, and NR2F1. Taken together, these data provide a better understanding of the molecular mechanism underlying tamoxifen-induced steatogenic hepatotoxicity and useful information for predicting steatogenic hepatotoxicity.

Comparison of growth factor and cytokine expression in patients with degenerated disc disease and herniated nucleus pulposus.

OBJECTIVES: This study was conducted to investigate the expression of cytokines and growth factors in disc specimens obtained from patients with herniated nucleus pulposus (HNP) and degenerated disc disease (DDD). DESIGN AND METHODS: MRI and Western blot analyses were performed to evaluate the levels of disc degeneration and the expression levels of cytokines and growth factors. RESULTS: The levels of TNF-alpha and IL-8 were significantly greater in the DDD group than in the HNP group, but no statistical differences were observed in the expression of IL-1beta, IL-6 and IL-12 between the HNP and DDD groups. In addition, the expression of TGF beta, VEGF and NGF was significantly higher in the DDD group than in the HNP group. CONCLUSION: The greater levels of cytokine and growth factor expression in the DDD group than in the HNP explain why discogenic patients usually have more severe back pain than patients with herniated discs.

Gene expression profiles of murine fatty liver induced by the administration of methotrexate.

Methotrexate (MTX) is used to treat a variety of chronic inflammatory and neoplastic diseases. However, it can induce hepatotoxicity such as microvesicular steatosis and necrosis. To explore the mechanisms of MTX-induced hepatic steatosis, we used microarray analysis to profile the gene expression patterns of mouse liver after MTX treatment. MTX was administered orally as a single dose of 10mg/kg (low dose) or 100 mg/kg (high dose) to ICR mice, and the livers were obtained 6 h, 24 h, and 72 h after treatment. Serum alanine aminotransferase, aspartate aminotransferase and triacylglycerol levels were not significantly altered in the experimental animals. Signs of steatosis were observed at 24 h after administration of high dose of MTX. From microarray data analysis, 908 genes were selected as MTX-responsive genes (P<0.05, two-way ANOVA; cutoff > or =1.5-fold). Database for Annotation, Visualization and Integrated Discovery (DAVID) analysis revealed that the predominant biological processes associated with these genes are response to unfolded proteins, phosphate metabolism, and cellular lipid metabolism. Functional categorization of these genes identified 28 genes involved in lipid metabolism that was interconnected with the biological pathways of biosynthesis, catabolism, and transport of lipids and fatty acids. Taken together, these data provide a better understanding of the molecular mechanisms of MTX-induced steatogenic hepatotoxicity, and useful information for predicting hepatotoxicity through pattern recognition.

Inhibition of glycogen synthase kinase-3 reduces L-DOPA-induced neurotoxicity.

The neurotoxicity of l-3,4-dihydroxyphenylalanine (L-DOPA), used for the treatment of Parkinson's disease, remains controversial. Although there are many reports suggesting that long-term treatment of L-DOPA causes neuronal death, an increasing body of recent evidence has proposed that L-DOPA might be neuroprotective rather than neurotoxic. We investigated the effect of L-DOPA on neuronally differentiated PC12 (nPC12) cells by treating cells with various concentrations of L-DOPA for 24h. We also studied whether glycogen synthase kinase (GSK)-3 activation is related to L-DOPA-induced neurotoxicity by simultaneously treating cells with several concentrations of L-DOPA and a GSK-3 inhibitor for 24h. MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, trypan blue staining, cell counting kit-8, and DAPI staining all showed that L-DOPA decreased nPC12 cell viability at high concentrations. In addition, 100 microM L-DOPA treatment significantly increased the activity of GSK-3 and death signals including cytochrome c, activated caspase-3 and cleaved PARP, and decreased survival signals including heat shock transcription factor-1 in a concentration-dependent manner. Treatment with GSK-3 inhibitor VIII or lithium chloride prevented L-DOPA-induced cell death. Together, these results suggest that L-DOPA induces neuronal cell death at high concentrations and that the neurotoxic effect of L-DOPA might be mediated in part by GSK-3 activation.

Gene expression profiling in human lung fibroblast following cadmium exposure.

Cadmium is a naturally occurring metallic element with food and smoking being the main sources of exposure in the non-occupationally exposed population. Chronic exposure to cadmium leads to tumors in a number of tissues including lung. In the present study we investigated genes whose expression is modified by Cd exposure in human lung fibroblast WI38-VA13 cells. We employed a cDNA microarray hybridization method to identify changes in the gene expression profile. Thirty five genes were identified as cadmium-responsive. Their level of expression differed significantly from controls (significance analysis of microarray; SAM, q<5%). The largest groups of gene products affected by cadmium exposure were those involved in cell cycle, immunity and defense, nucleoside metabolism and signal transduction. Repressed expression of E2f1, Tubb and Actg2 following cadmium exposure may contribute to the cell cycle arrest. Down-regulation of Eno1 indicates a potential for causing protooncogene expression and possibly for cadmium-induced carcinogenicity. These results may contribute to better understand the toxic mechanism of cadmium toxicity. Moreover, the gene expression profile of cadmium could provide potential biomarkers for cadmium exposure.

Gene expression profiles of murine fatty liver induced by the administration of valproic acid.

Valproic acid (VPA) has been used as anticonvulsants, however, it induces hepatotoxicity such as microvesicular steatosis and necrosis in the liver. To explore the mechanisms of VPA-induced steatosis, we profiled the gene expression patterns of the mouse liver that were altered by treatment with VPA using microarray analysis. VPA was orally administered as a single dose of 100 mg/kg (low-dose) or 1000 mg/kg (high-dose) to ICR mice and the animals were killed at 6, 24, or 72 h after treatment. Serum alanine aminotransferase and aspartate aminotransferase levels were not significantly altered in the experimental animals. However, symptoms of steatosis were observed at 72 h with low-dose and at 24 h and 72 h with high-dose. After microarray data analysis, 1910 genes were selected by two-way ANOVA (P<0.05) as VPA-responsive genes. Hierarchical clustering revealed that gene expression changes depended on the time rather than the dose of VPA treatment. Gene profiling data showed striking changes in the expression of genes associated with lipid, fatty acid, and steroid metabolism, oncogenesis, signal transduction, and development. Functional categorization of 1156 characteristically up- and down-regulated genes (cutoff >1.5-fold) revealed that 60 genes were involved in lipid metabolism that was interconnected with biological pathways for biosynthesis of triglyceride and cholesterol, catabolism of fatty acid, and lipid transport. This gene expression profile may be associated with the known steatogenic hepatotoxicity of VPA and it may provide useful information for prediction of hepatotoxicity of unknown chemicals or new drug candidates through pattern recognition.

Primary diffuse leptomeningeal gliomatosis: report of a case presenting with chronic meningitis.

Neoplastic meningitis occurs in approximately 5% of patients with cancer. Primary diffuse leptomeningeal gliomatosis is a rare condition whereby a glioma arises from heterotopic cell nests in the leptomeninges. We report here a case presenting with clinical features similar to those of chronic infectious meningitis without positive cerebrospinal fluid cytology. Neurological signs in our patient deteriorated progressively without responding to antitubercular, antiviral, or antibiotic therapy. Leptomeningeal biopsy sampling revealed the condition to be primary diffuse leptomeningeal gliomatosis.

Role of GSK-3beta activity in motor neuronal cell death induced by G93A or A4V mutant hSOD1 gene.

Point mutations such as G93A and A4V in the human Cu/Zn-superoxide dismutase gene (hSOD1) cause familial amyotrophic lateral sclerosis (fALS). In spite of several theories to explain the pathogenic mechanisms, the mechanism remains largely unclear. Increased activity of glycogen synthase kinase-3 (GSK-3) has recently been emphasized as an important pathogenic mechanism of neurodegenerative diseases, including Alzheimer's disease and ALS. To investigate the effects of G93A or A4V mutations on the phosphatidylinositol-3-kinase (PI3-K)/Akt and GSK-3 pathway as well as the caspase-3 pathway, VSC4.1 motoneuron cells were transfected with G93A- or A4V-mutant types of hSOD1 (G93A and A4V cells, respectively) and, 24 h after neuronal differentiation, their viability and intracellular signals, including PI3-K/Akt, GSK-3, heat shock transcription factor-1 (HSTF-1), cytochrome c, caspase-3 and poly(ADP-ribose) polymerase (PARP), were compared with those of wild type (wild cells). Furthermore, to elucidate the role of the GSK-3beta-mediated cell death mechanism, alterations of viability and intracellular signals in those mutant motoneurons were investigated after treating the cells with GSK-3beta inhibitor. Compared with wild cells, viability was greatly reduced in the G93A and A4V cells. However, the treatment of G93A and A4V cells with GSK-3beta inhibitor increased their viability by activating HSTF-1 and by reducing cytochrome c release, caspase-3 activation and PARP cleavage. However, the treatment did not affect the expression of PI3-K/Akt and GSK-3beta. These results suggest that the G93A or A4V mutations inhibit PI3-K/Akt and activate GSK-3beta and caspase-3, thus becoming vulnerable to oxidative stress, and that the GSK-3beta-mediated cell death mechanism is important in G93A and A4V cell death.

Phosphatidylinositol 3-kinase activator reduces motor neuronal cell death induced by G93A or A4V mutant SOD1 gene.

The primary pathogenic mechanism of amyotrophic lateral sclerosis (ALS) remains largely unclear. We recently observed that motoneuron cell death mediated by G93A or A4V mutant SOD1, causing familial ALS, was related with decrease of survival signals, such as phosphatidylinositol 3-kinase (PI3-K) and Akt, which play a pivotal role in neuronal survival. Using a G93A or A4V mutant SOD1 transfected VSC4.1 motoneuron cells (G93A or A4V cells, respectively), we presently investigated whether PI3-K activator could reduce mutant SOD1-mediated motoneuron cell death. To investigate the effect of PI3-K activator on viability of G93A and A4V cells, these cells were treated with 10, 50 or 100ng/ml PI3-K activator for 24h and viability and intracellular signals, including Akt, glycogen synthase kinase-3 (GSK-3), heat shock transcription factor-1 (HSTF-1), cytosolic cytochrome c, caspase-3 and poly(ADP-ribose) polymerase (PARP), were compared with those without treatment (control). Compared with non-treated control G93A or A4V cells, the PI3-K activator treatment increased their viability by enhancing the survival signals, including pAkt, pGSK-3, and by inhibiting the death signals, including caspase-3 activation and PARP cleavage. These results suggest that PI3-K activator protects G93A or A4V cells from mutant SOD1-mediated motoneuron cell death by both activating survival signals and inactivating death signals.

Protective effect of diallyl disulfide on oxidative stress-injured neuronally differentiated PC12 cells.

The effects of diallyl disulfide (DADS), a garlic-derived compound, on the viability of neuronal cells and cell signals, including phosphatidylinositol 3-kinase (PI3K)/Akt, glycogen synthase kinase-3 (GSK-3), cytochrome c, caspase-3, and poly(ADP-ribose) polymerase (PARP), were investigated in PC12 cells neuronally differentiated by nerve growth factor. To evaluate the toxicity of DADS itself, nPC12 cells were treated with several concentrations of DADS, and 3,(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and trypan blue stain revealed that the viability was not affected by low concentration of DADS, up to 20 microM, but it was decreased at higher than this concentration. The levels of free radicals and membrane lipid peroxidation were significantly increased in nPC12 cells when treated with more than 50 microM DADS, and treatment of PC12 cells with 100 microM DADS killed the cells by inhibiting PI3K/Akt and by promoting activation of GSK-3 and caspase-3, release of cytochrome c, and cleavage of PARP. To evaluate the protective effects of low concentration of DADS on oxidative stress-injured nPC12 cells, the viability of the cells (pretreated with DADS for 2 h vs. not pretreated) was evaluated 24 h after exposure to 100 microM H2O2 for 30 min. Compared to the cells treated with 100 microM H2O2 only, pretreatment of the cells with 20 microM DADS before exposure to 100 microM H2O2 increased the viability and induced activation of PI3K and Akt, inactivation of GSK-3, and inhibition of cytochrome c release, caspase-3 activation, and PARP cleavage. These results indicate that low concentration of DADS has neuroprotective effects by activating PI3K/Akt and by inhibiting GSK-3 activation, cytochrome c release, caspase-3 activation, and PARP cleavage, whereas high concentration is rather cytotoxic. Therefore, some specific optimum concentration of DADS may be a new potential therapeutic strategy for oxidative stress injured in vitro model of neurodegenerative diseases.