Antioxidant Efficacy of Hwangryunhaedok-tang through Nrf2 and AMPK Signaling Pathway against Neurological Disorders In Vivo and In Vitro.

Alzheimer's disease (AD) is a representative cause of dementia and is caused by neuronal loss, leading to the accumulation of aberrant neuritic plaques and the formation of neurofibrillary tangles. Oxidative stress is involved in the impaired clearance of amyloid beta (Aß), and Aß-induced oxidative stress causes AD by inducing the formation of neurofibrillary tangles. Hwangryunhaedok-tang (HHT, Kracie K-09®), a traditional herbal medicine prescription, has shown therapeutic effects on various diseases. However, the studies of HHT as a potential treatment for AD are insufficient. Therefore, our study identified the neurological effects and mechanisms of HHT and its key bioactive compounds against Alzheimer's disease in vivo and in vitro. In a 5xFAD mouse model, our study confirmed that HHT attenuated cognitive impairments in the Morris water maze (MWM) test and passive avoidance (PA) test. In addition, the prevention of neuron impairment, reduction in the protein levels of Aß, and inhibition of cell apoptosis were confirmed with brain tissue staining. In HT-22 cells, HHT attenuates tBHP-induced cytotoxicity, ROS generation, and mitochondrial dysfunction. It was verified that HHT exerts a neuroprotective effect by activating signaling pathways interacting with Nrf2, such as MAPK/ERK, PI3K/Akt, and LKB1/AMPK. Among the components, baicalein, a bioavailable compound of HHT, exhibited neuroprotective properties and activated the Akt, AMPK, and Nrf2/HO-1 pathways. Our findings indicate a mechanism for HHT and its major bioavailable compounds to treat and prevent AD and suggest its potential.

Protective Effects of Red Ginseng Against Tacrine-Induced Hepatotoxicity: An Integrated Approach with Network Pharmacology and Experimental Validation.

Introduction: Tacrine, an FDA-approved acetylcholinesterase inhibitor, has shown efficacy in treating Alzheimer's disease, but its clinical use is limited by hepatotoxicity. This study investigates the protective effects of red ginseng against tacrine-induced hepatotoxicity, focusing on oxidative stress. Methods: A network depicting the interaction between compounds and targets was constructed for RG. Effect of RG was determined by MTT and FACS analysis with cells stained by rhodamine 123. Proteins were extracted and subjected to immunoblotting for apoptosis-related proteins. Results: The outcomes of the network analysis revealed a significant association, with 20 out of 82 identified primary RG targets aligning with those involved in oxidative liver damage including notable interactions within the AMPK pathway. in vitro experiments showed that RG, particularly at 1000µg/mL, mitigated tacrine-induced apoptosis and mitochondrial damage, while activating the LKB1-mediated AMPK pathway and Hippo-Yap signaling. In mice, RG also protected the liver injury induced by tacrine, as similar protective effects to silymarin, a well-known drug for liver toxicity protection. Discussion: Our study reveals the potential of RG in mitigating tacrine-induced hepatotoxicity, suggesting the administration of natural products like RG to reduce toxicity in Alzheimer's disease treatment.

Forsythiaside A Activates AMP-Activated Protein Kinase and Regulates Oxidative Stress via Nrf2 Signaling.

Forsythiaside A (FA) is an active constituent isolated from Forsythia suspensa, a beneficial herb used in traditional medicine known for its antioxidant and anti-inflammatory properties. Although various studies have suggested that FA has the protective effects, its impacts on arachidonic acid (AA) plus iron in vitro models and carbon tetrachloride (CCl4)-induced mouse liver damage in vivo have not been explored. In this study, HepG2 cells were subjected to AA + iron treatment to induce apoptosis and mitochondrial impairment and determine the molecular mechanisms. FA exhibited protective effects by inhibiting cell damage and reactive oxygen species (ROS) production induced by AA + iron, as assessed via immunoblot and flow cytometry analyses. Further molecular investigations revealed that FA resulted in the activation of extracellular-signal-related protein kinase (ERK), which subsequently triggered the activation of AMP-activated protein kinase (AMPK), a critical regulator of cellular oxidative stress. Additionally, FA modulated the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway, which is a significant antioxidant transcription factor regulated by the AMPK pathway. For in vivo studies, mice were orally administered FA and then subjected to induction of CCl4-based hepatotoxicity. The protective effect of FA was confirmed via blood biochemistry and immunohistochemical analyses. In conclusion, our findings demonstrated the protective effects of FA against oxidative stress both in vitro and in vivo, thus indicating that FA is a potential candidate for liver protection. Our study sheds light on the mechanistic pathways involved in the antioxidant effects of FA, highlighting the hepatoprotective potential of naturally occurring compounds in traditional herbs, such as FA.

Kaempferol stimulation of autophagy regulates the ferroptosis under the oxidative stress as mediated with AMP-activated protein kinase.

Recent studies have highlighted the positive effects of Kaempferol (KP), including its anti-inflammatory and antioxidant properties. However, its impact on oxidative damage induced by heavy metals and pro-inflammatory mediators, such as arachidonic acid (AA), has not yet been identified. Our objective was to specifically evaluate liver damage due to AA + iron-induced oxidative stress, both in vitro and in vivo. In HepG2 cells, KP activated the AMP-activated protein kinase (AMPK), suggesting a hepatoprotective effect through AMPK inhibition, as assessed by immunoblot and FACS analysis (EC50 = 10 µM). KP also stimulated autophagy, a degradation process that eliminates aged, damaged, and unnecessary components, via mTOR inhibition and ULK1 phosphorylation. This activation was further validated by the upregulation of autophagy-related genes (ATG5) and Beclin-1, along with the conversion of LC3BI to LC3BII. Ferroptosis, a non-apoptotic type of cell death characterized by oxidative stress from the production of reactive oxygen species (ROS) and excessive iron accumulation, was linked to the activation of autophagy, as confirmed through the protein expression of deferoxamine (DFO) and ferrostatin-1 (Fer-1), the representative ferroptosis inhibitors (positive controls). In mice, oral administration of KP demonstrated protective effects against CCl4-induced hepatotoxicity. In conclusion, KP provides hepatoprotective effects against oxidative stress induced by AA + iron treatment in vitro and CCl4 treatment in vivo.

Systemic and molecular analysis dissect the red ginseng induction of apoptosis and autophagy in HCC as mediated with AMPK.

Background: Hepatocellular carcinoma (HCC) has a high incidence and is one of the highest mortality cancers when advanced stage is proceeded. However, Anti-cancer drugs available for treatment are limited and new anti-cancer drugs and new ways to treat them are minimal. We examined that the effects and possibility of Red Ginseng (RG, Panax ginseng Meyer) as new anti-cancer drug on HCC by combining network pharmacology and molecular biology. Materials and Methods: Network pharmacological analysis was employed to investigate the systems-level mechanism of RG focusing on HCC. Cytotoxicity of RG was determined by MTT analysis, which were also stained by annexin V/PI staining for apoptosis and acridine orange for autophagy. For the analyze mechanism of RG, we extracted protein and subjected to immunoblotting for apoptosis or autophagy related proteins. Results: We constructed compound-target network of RG and identified potential pathways related to HCC. RG inhibited growth of HCC through acceleration of cytotoxicity and reduction of wound healing ability of HCC. RG also increased apoptosis and autophagy through AMPK induction. In addition, its ingredients, 20S-PPD (protopanaxadiol) and 20S-PPT (protopanaxatriol), also induced AMPK mediated apoptosis and autophagy. Conclusion: RG effectively inhibited growth of HCC cells inducing apoptosis and autophagy via ATG/AMPK in HCC cells. Overall, our study suggests possibility as new anti-cancer drug on HCC by proof for the mechanism of the anti-cancer action of RG.

Unraveling the Antioxidant Capacity of Spatholobi caulis in Nonalcoholic Fatty Liver Disease: A Multiscale Network Approach Integrated with Experimental Validation.

Nonalcoholic fatty liver disease (NAFLD) is a global health problem that is closely associated with obesity and metabolic syndrome. Spatholobi caulis (SC) is a herbal medicine with potential hepatoprotective effects; however, its active compounds and underlying mechanisms have not been fully explored. In this study, we combined a multiscale network-level approach with experimental validation to investigate SC's antioxidant properties and their impact on NAFLD. Data collection and network construction were performed, and active compounds and key mechanisms were identified through multi-scale network analysis. Validation was conducted using in vitro steatotic hepatocyte models and in vivo high-fat diet-induced NAFLD models. Our findings revealed that SC treatment improved NAFLD by modulating multiple proteins and signaling pathways, including AMPK signaling pathways. Subsequent experiments showed that SC treatment reduced lipid accumulation and oxidative stress. We also validated SC's effects on AMPK and its crosstalk pathways, emphasizing their role in hepatoprotection. We predicted procyanidin B2 to be an active compound of SC and validated it using a lipogenesis in vitro model. Histological and biochemical analyses confirmed that SC ameliorated liver steatosis and inflammation in mice. This study presents SC's potential use in NAFLD treatment and introduces a novel approach for identifying and validating active compounds in herbal medicine.

Integrative approach to uncover antioxidant properties of Bupleuri Radix and its active compounds: Multiscale interactome-level analysis with experimental validation.

Acute and chronic liver disease are global problems with high morbidity and mortality. Bupleuri Radix (BR) is an herbal medicine that has been prescribed empirically in traditional Asian medicine to modulate liver metabolism. However, its active compounds and therapeutic mechanisms remain unclear. Here, we integrated a network-based approach and experimental validation to elucidate BR's therapeutic potential in treating oxidative liver injury. Our approach incorporated data collection and network construction utilizing bioinformatics tools, and identified active compounds and key mechanisms based on the multiscale interactome. The proposed mechanisms were validated using an in vitro oxidative stress model and an in vivo carbon tetrachloride-induced model. We found that BR ameliorated the oxidative hepatic damage by acting on multiple proteins (STAT3, TNF, and BCL2) and signaling pathways (AMPK and Hippo signaling pathways). Subsequent in vitro experiments confirmed that BR significantly inhibited oxidative stress and mitochondrial damage. We further validated the effect of BR on the AMPK and Hippo-YAP pathways; a key mechanism for the antioxidant properties of BR. We prioritized the active compounds in BR based on a multiscale interactome-based approach, and further experiments revealed that saikosaponin A was a key active compound involved in hepatocyte protection (EC50 = 50 µM), similar to the result using metformin and 5-aminoimidazole-4-carboxamide ribonucleotide. Histochemistry and blood biochemistry established that BR significantly inhibited carbon tetrachloride-induced oxidative tissue damage in mice. Thus, BR can be used to develop novel therapeutics for oxidative liver injury. Moreover, we suggest a novel strategy to prioritize and validate the active compounds and key mechanisms of herbal medicine based on the multiscale interactome.

Coordination of AMPK and YAP by Spatholobi Caulis and Procyanidin B2 Provides Antioxidant Effects In Vitro and In Vivo.

The liver is vulnerable to oxidative attacks from heavy metals, such as iron, as well as some drugs, including acetaminophen. It has been shown that enhanced oxidative stress in the liver leads to excessive ROS production and mitochondrial dysfunction, resulting in organ injury. The beneficial effects of Spatholobi Caulis (SC), a natural herbal medicine, include treating ischemic stroke, inhibiting tumor cell invasion, pro-angiogenic activities, and anti-inflammatory properties. Scientific studies on its effects against hepatotoxic reagents (e.g., iron and acetaminophen), as well as their underlying mechanisms, are insufficient. This study examined the antioxidant effects and mechanisms of SC in vitro and in vivo. In cells, the proinflammatory mediator, arachidonic acid (AA), plus iron, significantly induced an increase in ROS generation, the damage in mitochondrial membrane potential, and the resulting apoptosis, which were markedly blocked by SC. More importantly, SC affected the activation of AMP-activated protein kinase (AMPK)-related proteins, which were vital to regulating oxidative stress in cells. In addition, SC mediated the expression of Yes-associated protein (YAP)-related proteins. Among the active compounds in SC, the procyanidin B2, but not liquiritigenin, daidzein, and genistein, significantly inhibited the cytotoxicity induced by AA + iron, and activated the LKB1-AMPK pathway. In mice, the oral administration of SC alleviated the elevations of ALT and histological changes by the acetaminophen-induced liver injury. These results reveal the potential of SC and a key bioactive component, procyanidin B2, as antioxidant candidates for hepatoprotection.

Angelica gigas NAKAI and Its Active Compound, Decursin, Inhibit Cellular Injury as an Antioxidant by the Regulation of AMP-Activated Protein Kinase and YAP Signaling.

Natural products and medicinal herbs have been used to treat various human diseases by regulating cellular functions and metabolic pathways. Angelica gigas NAKAI (AG) helps regulate pathological processes in some medical fields, including gastroenterology, gynecology, and neuropsychiatry. Although some papers have reported its diverse indications, the effects of AG against arachidonic acid (AA)+ iron and carbon tetrachloride (CCl4) have not been reported. In HepG2 cells, AA+ iron induced cellular apoptosis and mitochondrial damage, as assessed by mitochondrial membrane permeability (MMP) and the expression of apoptosis-related proteins. On the other hand, AG markedly inhibited these detrimental phenomena and reactive oxygen species (ROS) production induced by AA+ iron. AG activated the liver kinase B1 (LKB1)-dependent AMP-activated protein kinase (AMPK), which affected oxidative stress in the cells. Moreover, AG also regulated the expression of yes-associated protein (YAP) signaling as mediated by the AMPK pathways. In mice, an oral treatment of AG protected against liver toxicity induced by CCl4, as indicated by the plasma and histochemical parameters. Among the compounds in AG, decursin had antioxidant activity and affected the AMPK pathway. In conclusion, AG has antioxidant effects in vivo and in vitro, indicating that natural products such as AG could be potential candidate for the nutraceuticals to treat various disorders by regulating mitochondrial dysfunction and cellular metabolic pathways.

A Critical YAP in Malignancy of HCC Is Regulated by Evodiamine.

Liver cancer has relatively few early symptoms and is usually diagnosed in the advanced stage. Sorafenib is the only first-line anticancer drug approved by the Food and Drug Administration (FDA) for advanced HCC; however, its use is limited due to resistance. Therefore, the development of new drugs is essential to achieving customized treatment. Many studies have suggested that Yes-associated protein (YAP)/transcriptional co-activator with PDZ-binding motif (TAZ) is associated with metastasis and cancer formation and progression in various cancers. In the present study, YAP was overexpressed in various patient-derived hepatocarcinoma (HCC) tissues. In addition, this study examined whether evodiamine (which has anticancer effects) can inhibit YAP and, if so, modulate HCC. Evodiamine significantly reduced both the YAP level and cell growth of HCC in a dose-dependent manner. Biochemical analysis indicated mitochondria dysfunction-mediated apoptosis to be the cause of the reduction in HCC cell growth by evodiamine. YAP was overexpressed in metastatic HCC tissues as well when compared to primary HCC tissues. Migration and invasion analysis showed that evodiamine has anti-metastatic ability on Hep3B and Huh-7 cells and reduces the level of vimentin, an EMT marker. In conclusion, YAP is a critical target in HCC therapy, and evodiamine can be an effective HCC anticancer drug by reducing the YAP level.

Improved STR analysis of degraded DNA from human skeletal remains through in-house MPS-STR panel.

DNA analysis of degraded samples and low-copy number DNA derived from skeletal remains, one of the most challenging forensic tasks, is common in disaster victim identification and genetic analysis of historical materials. Massively parallel sequencing (MPS) is a useful technique for STR analysis that enables the sequencing of smaller amplicons compared with conventional capillary electrophoresis (CE), which is valuable for the analysis of degraded DNA. In this study, 92 samples of human skeletal remains (70+ years postmortem) were tested using an in-house MPS-STR system designed for the analysis of degraded DNA. Multiple intrinsic factors of DNA from skeletal remains that affect STR typing were assessed. The recovery of STR alleles was influenced more by DNA input amount for amplification rather than DNA degradation, which may be attributed from the high quantity and quality of libraries prepared for MPS run. In addition, the higher success rate of STR typing was achieved using the MPS-STR system compared with a commercial CE-STR system by providing smaller sized fragments for amplification. The results can provide constructive information for the analysis of degraded sample, and this MPS-STR system will contribute in forensic application with regard to skeletal remain sample investigation.

Genomic profiling of the residual disease of advanced high-grade serous ovarian cancer after neoadjuvant chemotherapy.

The goal of our study was to demonstrate the spectrum of genomic alterations present in the residual disease of patients with advanced high-grade serous ovarian cancer (HGSOC) after neoadjuvant chemotherapy (NAC), including matched pretreatment biopsies. During the study period between 2006 and 2017, we collected pre-NAC and post-NAC tumor tissue samples from patients with advanced HGSOC. We performed combined next-generation sequencing and immunohistochemistry to identify actionable targets and pathway activation in post-NAC residual tumors. We also examined whether post-NAC profiling of residual HGSOC identified targetable molecular lesions in the chemotherapy-resistant component of tumors. Among 102 post-NAC samples, 41 (40%) of patients had mutations in homologous recombination repair (HRR) genes (HRR deficiency). Patients with HRR mutations had higher tumor mutation burdens (p < 0.001) and higher alterations in the PI3K-AKT-mTOR pathway (p = 0.004) than patients without these HRR mutations. Nevertheless, we found no significant differences in progression-free survival (p = 0.662) and overall survival (OS; p = 0.828) between the two groups. Most patients (91%) had alterations in at least one of the targetable pathways, and those patients with cell cycle (p = 0.004) and PI3K-AKT-mTOR signaling (p = 0.005) pathway alterations had poorer OS (Bonferroni-corrected threshold = 0.0083, 0.05/6). We showed the genomic landscape of tumor cells remaining in advanced HGSOC after NAC. Once validated, these data can help inform biomarker-driven adjuvant studies in targeting residual tumors to improve the outcomes of patients with advanced HGSOC after NAC.

Protective effects of a traditional herbal extract from Stellaria dichotoma var. lanceolata against Mycobacterium abscessus infections.

Stellaria dichotoma var. lanceolata (SdLv), a member of the Caryophyllaceae, is a traditional herbal medicine that has been used to treat fever, night sweats, and malaria in East Asia. Inflammation plays an essential role in both host defense and pathogenesis during infection by diverse intracellular pathogens. Herein, we showed that an herbal extract from SdLv effectively attenuated inflammatory responses from infection of Mycobacterium abscessus (Mab), but not Toxoplasma gondii (T. gondii). In primary murine macrophages, Mab infection resulted in the rapid activation of nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK), as well as in the generation of proinflammatory cytokines, such as tumor necrosis factor α and interleukin-6, which were all significantly inhibited by pretreatment with SdLv. However, herbal extracts from Bupleurum chinense DC. (Buch) or Bupleurum falcatum L. (Bufa) did not affect M. abs-induced activation of proinflammatory responses. Importantly, we demonstrated that generation of intracellular reactive oxygen species, which are important signaling intermediaries in the activation of NF-κB and the MAPK signaling pathway, was rapidly increased in Mab-infected macrophages, and this was effectively suppressed by pretreatment with SdLv, but not Buch and Bufa. We further found that the treatment of Buch and Bufa, but not SdLv, led to the activation of NF-κB and the MAPK signaling pathway and the generation of intracellular reactive oxygen species. Moreover, oral administration of SdLv significantly reduced lethality in Mab-infected mice. Collectively, these results suggest the possible use of SdLv as an effective treatment for Mab infection.

NADPH oxidase 4 is required for the generation of macrophage migration inhibitory factor and host defense against Toxoplasma gondii infection.

Nicotinamide adenine dinucleotide phosphate (NADPH) oxidases (Nox) are an important family of catalytic enzymes that generate reactive oxygen species (ROS), which mediate the regulation of diverse cellular functions. Although phagocyte Nox2/gp91phox is closely associated with the activation of host innate immune responses, the roles of Nox family protein during Toxoplasma gondii (T. gondii) infection have not been fully investigated. Here, we found that T. gondii-mediated ROS production was required for the upregulation of macrophage migration inhibitory factor (MIF) mRNA and protein levels via activation of mitogen-activated protein kinase and nuclear factor-κB signaling in macrophages. Interestingly, MIF knockdown led to a significant increase in the survival of intracellular T. gondii in bone marrow-derived macrophages (BMDMs). Moreover, Nox4 deficiency, but not Nox2/gp91phox and the cytosolic subunit p47phox, resulted in enhanced survival of the intracellular T. gondii RH strain and impaired expression of T. gondii-mediated MIF in BMDMs. Additionally, Nox4-deficient mice showed increased susceptibility to virulent RH strain infection and increased cyst burden in brain tissues and low levels of MIF expression following infection with the avirulent ME49 strain. Collectively, our findings indicate that Nox4-mediated ROS generation plays a central role in MIF production and resistance to T. gondii infection.

Effects of MBL2 polymorphisms in patients with diisocyanate-induced occupational asthma.

Diisocyanate (DI) is the most common cause of occupational asthma (OA) in Korea. Mannose-binding lectin (MBL) initiates the lectin complement activation pathway following oxidative stress and plays an important role in the regulation of inflammatory processes. To determine whether there is a genetic association between MBL2 polymorphisms and DI-OA, 99 patients with DI-OA, 99 asymptomatic exposed controls (AECs) and 144 unexposed normal controls were enrolled in this study. Three polymorphisms (-554 G>C, -431A>C and -225 G>C) in the MBL2 promoter were genotyped, and serum MBL levels were determined by enzyme-linked immunosorbent assay. Functional variabilities in the promoter polymorphisms were analyzed by a luciferase reporter assay and electrophoretic mobility shift assay (EMSA). A significantly higher frequency of haplotype (ht) 2 [CAG] was noted in the DI-OA group compared with the AEC group (P=0.044). The patients with DI-OA carrying ht2 [CAG] had significantly lower PC20 methacholine levels (P<0.001) than the non-carriers. The serum MBL levels were significantly higher in the DI-exposed subjects (both the DI-OA patients and AECs) carrying ht1 [GAG] (P=0.028). Luciferase activity was significantly enhanced in ht1 [GAG] compared with ht2 [CAG] in human hepatocarcinoma cells (Hep3B) (P=0.002). The EMSA showed that a -554G probe produced a specific shifted band compared with the -554C probe. These findings suggest that decreased serum MBL levels due to polymorphisms of the MBL2 gene may increase susceptibility to the development of DI-OA in DI-exposed individuals.

Effect of material properties on stability of silver nanoparticles in water.

Increased production and use of silver nanoparticles (AgNPs) could potentially lead to their release into the environment. Estimating the exposure to engineered nanomaterials in the environment is essential for assessing their risk. This study examined the aggregation and sedimentation kinetics behaviors of citrate- (Cit-AgNPs) and polyvinylpyrrolidone-coated silver nanoparticles (PVP-AgNPs) with three different average sizes in calcium chloride (CaCl2) solutions, emphasizing the effects of particle size and type of coating material on both behaviors. As the ionic strength increased, Cit-AgNPs (stabilized by charge repulsion) aggregated rapidly and settled down, while PVP-AgNPs (stabilized by steric repulsion) did not aggregate, even at an ionic strength of 10 mM CaCl2, due to likely steric hiderance effects of PVP coating. Interestingly, however, PVP-AgNPs sedimented without aggregating within 7 days and this tendency seems to having relevance to the particle size. These results suggest that the particle size and type of coating material play important roles in determining nanoparticle fate and transport.

Diagnostic patterns in the evaluation of patients presenting with syncope at the emergency or outpatient department.

PURPOSE: Patterns of syncope evaluation vary widely among physicians and hospitals. The aim of this study was to assess current diagnostic patterns and medical costs in the evaluation of patients presenting with syncope at the emergency department (ED) or the outpatient department (OPD) of a referral hospital. MATERIALS AND METHODS: This study included 171 consecutive patients with syncope, who visited the ED or OPD between January 2009 and July 2009. RESULTS: The ED group had fewer episodes of syncope [2 (1-2) vs. 2 (1-5), p=0.014] and fewer prodromal symptoms (81.5% vs. 93.3%, p=0.018) than the OPD group. Diagnostic tests were more frequently performed in the ED group than in the OPD group (6.2±1.7 vs. 5.3±2.0; p=0.012). In addition, tests with low diagnostic yields were more frequently used in the ED group than in the OPD group. The total cost of syncope evaluation per patient was higher in the ED group than in the OPD group [823,000 (440,000-1,408,000) won vs. 420,000 (186,000-766,000) won, p<0.001]. CONCLUSION: There were some differences in the clinical characteristics of patients and diagnostic patterns in the evaluation of syncope between the ED and the OPD groups. Therefore, a selective diagnostic approach according to the presentation site is needed to improve diagnostic yields and to reduce the time and costs of evaluation of syncope.

Endothelial nitric oxide synthase gene polymorphisms and the risk of silent brain infarction.

Silent brain infarction (SBI), a unique cerebrovascular disorder, is frequently detected on magnetic resonance imaging (MRI) of apparently healthy elderly persons, and it is known to increase the risk of stroke and other related diseases. Although detailed mechanisms of SBI pathogenesis have yet to be determined, recent studies suggest that SBI is significantly influenced by genetic factors. In this study, we investigated polymorphisms in the endothelial nitric oxide synthase (eNOS) gene (i.e., -786T>C, 4a4b and 894G>T) as possible risk factors for SBI. We enrolled 269 patients with SBI and 234 control subjects and examined their fasting plasma homocysteine and folate levels, and analyzed for eNOS polymorphisms and haplotypes. The prevalence of SBI was shown to be significantly higher in patients with the eNOS 894GT genotype (OR, 2.00; 95% CI, 1.30-3.08) and 894GT+TT genotype (OR, 2.05; 95% CI, 1.34-3.16), compared with the 894GG genotype. However, in the case of -786T>C and 4a4b polymorphisms, no significant difference was observed between SBI patients and normal subjects. Interestingly, we found that the prevalence of SBI can increase twice as high when either -786T>C or 4a4b polymorphism was combined with 894G>T polymorphism, -786TC+CC/894GT+TT (OR, 3.83; 95% CI, 1.24-11.80) and 4a4b+4a4a/894GT+TT (OR, 4.08; 95% CI, 1.34-12.40), respectively. According to haplotype analysis, we found that three haplotypes (-786T-4b-894G, -786T-4b-894T and -786C-4a-894T) were shown to be significantly different between SBI patients and control subjects. These results indicate that eNOS polymorphisms and haplotypes serve as risk factors for SBI, and three different polymorphic loci in the eNOS gene play interactively, thereby leading to synergistic effects for the generation of SBI.

Gender-specific association between polymorphism of vascular endothelial growth factor (VEGF 936C>T) gene and patients with stomach cancer.

PURPOSE: Angiogenesis plays an important role in the growth, progression, and metastasis of tumors. Vascular endothelial growth factor (VEGF) overexpression has been associated with advanced stage and poor survival in several cancers. We investigated the present case-control study to determine whether there is an association between the VEGF 936C>T polymorphism and stomach cancer. PATIENTS AND METHODS: The association of functional single nucleotide polymorphisms (SNPs) of the VEGF gene with stomach cancer development was evaluated in a case-control study of 154 Korean stomach cancer patients. Genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. RESULTS: Our results revealed significant association of T allele-bearing genotypes with increased risk for stomach cancer development. Genotype frequencies of the VEGF 936C>T polymorphisms were significantly different between patient and control groups (CT, AOR: 2.007, 95% CI: 1.277-3.156, TT, AOR: 4.790, 95% CI: 1.174-19.539, CT+TT, AOR: 2.147, 95% CI: 1.382-3.337). When stratified by gender and age, genotype frequencies were significantly different for stomach cancer in women and in patients younger than 55 years (in women, CT, OR: 3.049, 95% CI: 1.568-5.930, CT+TT, OR: 3.132, 95% CI: 1.638-5.990; in < 55 years, CT, OR: 3.306, 95% CI: 1.413-7.732, CT+TT, OR: 3.967, 95% CI: 1.729-9.104). In addition, this association partially remained in cases with intestinal and diffuse-type stomach cancer. CONCLUSION: Our present study suggests that the VEGF 936C>T polymorphism is a susceptibility factor for stomach cancer, at least in Korean. These observations, however, require further confirmation by a larger multi-ethnic study.