Near-Infrared Light Emitting Diode Based Non-Invasive Glucose Detection System.

Diabetes mellitus is a common disease that has affected people since antiquity. The complexity of diabetes mellitus lies in the fact that it requires continuous management in order to prevent serious complications. Various methods for managing diabetes mellitus through monitoring blood glucose levels have been studied and developed, and the most widely used method includes using an invasive blood glucose meter. Since the invasive blood glucose meter poses a major inconvenience for patients, in this study, we sought to develop a non-invasive blood glucose meter. We therefore proposed the development of a non-invasive blood glucose measurement system that is based on near-infrared spectroscopy. The system developed was composed of a light source for emitting different wavelengths, a light detector unit, and a computing system for recording signals. A prepared glucose solution was injected into a quartz cuvette and light at 780-1650 nm was emitted to pass through the cuvette. The degree of reaction was determined by recording the change in wavelength. The wavelength band used for the experiment was 780-1000 nm and the resolution was 20 nm. The glucose concentration was determined to be between 50 to 400 mg/dl compared to the normal range of 80 to 120 mg/dl. By examining which wavelength bands specifically reacted with glucose, we observed that the wavelength bands that decreased or increased in response to the glucose concentration were at 780 nm and 940 nm. For wavelength bands at 1000 nm or above, light was also absorbed by water and therefore it was difficult to distinguish the results. The most reliable wavelength band was at 940 nm with an R² of 0.9806. In conclusion, the near-infrared light emitting diode based non-invasive glucose detection system performed well and is expected to be a superior method for monitoring blood glucose levels in diabetes mellitus.

Anti-Inflammatory Effects of Step Electrical Stimulation on Complete Freund's Adjuvant (CFA) Induced Rheumatoid Arthritis Rats.

Rheumatoid arthritis is a chronic inflammatory disease that affects joints and induces pain and swelling. We evaluated the anti-inflammatory effects of step electrical stimulation (SES) in this study. SES was carried out by increasing the voltage (3 V/s) from 5 V to 100 V for 60 cycles. The viability of mouse embryonic fibroblasts (NIH-3T3) was evaluated after step-electrical stimulation. After the injection of complete Freund's adjuvant (CFA) on the right hind paw of Sprague Dawley (SD) rats (6 weeks old), the degree of swelling was measured using a digital plethysmometer and Vernier caliper. Histological changes in inflamed tissues were observed with hematoxylin and eosin (H&E) staining, while the degree of inflammation was evaluated from the expression level of inflammatory factors such as cyclooxygenase-2 (COX-2), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6). As a result, we found no difference in cell viability after SES treatment between the control and SES-treated groups. On day 21 after CFA injection, the swelling of right hind paws decreased by 1.09 times in SES-treated group as compared with the untreated group. In addition, the levels of COX-2, TNF-α and IL-6 significantly decreased after SES treatment. Thus, SES treatment decreased paw swelling and alleviated inflammation.

GAREM1 regulates the PR interval on electrocardiograms.

PR interval is the period from the onset of P wave to the start of the QRS complex on electrocardiograms. A recent genomewide association study (GWAS) suggested that GAREM1 was linked to the PR interval on electrocardiograms. This study was designed to validate this correlation using additional subjects and examined the function of Garem1 in a mouse model. We analyzed the association of rs17744182, a variant in the GAREM1 locus, with the PR interval in 5646 subjects who were recruited from 2 Korean replication sets, Yangpyeong (n = 2471) and Yonsei (n = 3175), and noted a significant genomewide association by meta-analysis (P = 2.39 × 10-8). To confirm the function of Garem1 in mice, Garem1 siRNA was injected into mouse tail veins to reduce the expression of Garem1. Garem1 transcript levels declined by 53% in the atrium of the heart (P = 0.029), and Garem1-siRNA injected mice experienced a significant decrease in PR interval (43.27 ms vs. 44.89 ms in control, P = 0.007). We analyzed the expression pattern of Garem1 in the heart by immunohistology and observed specific expression of Garem1 in intracardiac ganglia. Garem1 was expressed in most neurons of the ganglion, including cholinergic and adrenergic cells. We have provided evidence that GAREM1 is involved in the PR interval of ECGs. These findings increase our understanding of the regulatory signals of heart rhythm through intracardiac ganglia of the autonomic nervous system and can be used to guide the development of a therapeutic target for heart conditions, such as atrial fibrillation.

Phosphorylation and inactivation of glycogen synthase kinase 3ß (GSK3ß) by dual-specificity tyrosine phosphorylation-regulated kinase 1A (Dyrk1A).

Glycogen synthase kinase 3ß (GSK3ß) participates in many cellular processes, and its dysregulation has been implicated in a wide range of diseases such as obesity, type 2 diabetes, cancer, and Alzheimer disease. Inactivation of GSK3ß by phosphorylation at specific residues is a primary mechanism by which this constitutively active kinase is controlled. However, the regulatory mechanism of GSK3ß is not fully understood. Dual-specificity tyrosine phosphorylation-regulated kinase 1A (Dyrk1A) has multiple biological functions that occur as the result of phosphorylation of diverse proteins that are involved in metabolism, synaptic function, and neurodegeneration. Here we show that GSK3ß directly interacts with and is phosphorylated by Dyrk1A. Dyrk1A-mediated phosphorylation at the Thr(356) residue inhibits GSK3ß activity. Dyrk1A transgenic (TG) mice are lean and resistant to diet-induced obesity because of reduced fat mass, which shows an inverse correlation with the effect of GSK3ß on obesity. This result suggests a potential in vivo association between GSK3ß and Dyrk1A regarding the mechanism underlying obesity. The level of Thr(P)(356)-GSK3ß was higher in the white adipose tissue of Dyrk1A TG mice compared with control mice. GSK3ß activity was differentially regulated by phosphorylation at different sites in adipose tissue depending on the type of diet the mice were fed. Furthermore, overexpression of Dyrk1A suppressed the expression of adipogenic proteins, including peroxisome proliferator-activated receptor γ, in 3T3-L1 cells and in young Dyrk1A TG mice fed a chow diet. Taken together, these results reveal a novel regulatory mechanism for GSK3ß activity and indicate that overexpression of Dyrk1A may contribute to the obesity-resistant phenotype through phosphorylation and inactivation of GSK3ß.

ATM-deficient human fibroblast cells are resistant to low levels of DNA double-strand break induced apoptosis and subsequently undergo drug-induced premature senescence.

DNA DSBs are induced by IR or radiomimetic drugs such as doxorubicin. It has been indicated that cells from ataxia-telangiectasia patients are highly sensitive to radiation due to defects in DNA repair, but whether they have impairment in apoptosis has not been fully elucidated. A-T cells showed increased sensitivity to high levels of DNA damage, however, they were more resistant to low doses. Normal cells treated with combination of KU55933, a specific ATM kinase inhibitor, and doxorubicin showed increased resistance as they do in a similar manner to A-T cells. A-T cells have higher viability but more DNA breaks, in addition, the activations of p53 and apoptotic proteins (Bax and caspase-3) were deficient, but Akt expression was enhanced. A-T cells subsequently underwent premature senescence after treatment with a low dose of doxorubicin, which was confirmed by G2 accumulation, senescent morphology, and SA-ß-gal positive until 15 days repair incubation. Finally, A-T cells are radio-resistant at low doses due to its defectiveness in detecting DNA damage and apoptosis, but the accumulation of DNA damage leads cells to premature senescence.

Association between TGFBR2 gene polymorphism (rs2228048, Asn389Asn) and intracerebral hemorrhage in Korean population.

Transforming growth factor, beta receptor II (TGFBR2) is mainly expressed by neurons in the central nervous system, and reduced neuronal TGFBR2 signaling results in accelerated age-dependent neurodegeneration. To investigate whether TGFBR2 polymorphisms are associated with ischemic stroke (IS) and intracerebral hemorrhage (ICH), two single nucleotide polymorphisms (SNPs) of TGFBR2 gene (rs764522, -1444C/G; rs2228048, Asn389Asn) were selected and genotyped by direct sequencing in 247 stroke patients (120 IS and 127 ICH) and 655 control subjects (260 for IS and 395 for ICH). SNPStats, SNPAnalyzer, Helixtree, and Haploview version 4.2 were used to analyze genetic data. Multiple logistic regression models (codominant, dominant, recessive, and log-additive) were performed to evaluate odds ratios (ORs), 95% confidence intervals (CIs), and p values. The synonymous SNP rs2228048 was significantly associated with ICH (p = 0.032 in codominant 2 model, p = 0.024 in dominant model, p = 0.020 in recessive model, and p = 0.005 in log-additive model) and Fisher's exact test (p = 0.009). Allele frequencies of rs2228048 were different between ICH and controls (p = 0.006). In Bonferroni correction, these correlations were also significant. These results suggest that the synonymous SNP rs2228048 of TGFBR2 gene may be associated with development of ICH in Korean population.

Prothrombin kringle-2 induces death of mesencephalic dopaminergic neurons in vivo and in vitro via microglial activation.

We have shown that prothrombin kringle-2 (pKr-2), a domain of human prothrombin distinct from thrombin could activate cultured rat brain microglia in vitro. However, little is known whether pKr-2-induced microglial activation could cause neurotoxicity on dopaminergic (DA) neurons in vivo. To address this question, pKr-2 was injected into the rat substantia nigra (SN). Tyrosine hydroxylase (TH) immunohistochemistry experiments demonstrate significant loss of DA neurons seven days after injection of pKr-2. In parallel, pKr-2-activated microglia were detected in the SN with OX-42 and OX-6 immunohistochemistry. Reverse transcription PCR and double-label immunohistochemistry revealed that activated microglia in vivo exhibit early and transient expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and several proinflammatory cytokines. The pKr-2-induced loss of SN DA neurons was partially inhibited by the NOS inhibitor N(G)-nitro-L-arginine methyl ester hydrochloride, and the COX-2 inhibitor DuP-697. Extracellular signal-regulated kinase 1/2, c-Jun N-terminal kinase and p38 mitogen-activated protein kinase were activated in the SN as early as 1 hr after pKr-2 injection, and localized within microglia. Inhibition of these kinases led to attenuation of mRNA expression of iNOS, COX-2 and several proinflammatory cytokines, and rescue of DA neurons in the SN. Intriguingly, following treatment with pKr-2 in vitro, neurotoxicity was detected exclusively in co-cultures of mesencephalic neurons and microglia, but not microglia-free neuron-enriched mesencephalic cultures, indicating that microglia are required for pKr-2 neurotoxicity. Our results strongly suggest that microglia activated by endogenous compound(s), such as pKr-2, are implicated in the DA neuronal cell death in the SN.

Differential effects of 5-fluorouracil on glucose transport and expressions of glucose transporter proteins in gastric cancer cells.

Although 5-fluorouracil (5-FU) is a widely used chemotherapeutic agent in the treatment of gastric cancer, the underlying mechanism for 5-FU resistant phenotype, has yet to be elucidated. We hypothesized that the sensitivity of gastric cancer to 5-FU treatment might be related to the rate of glucose transport (GLUT), and investigated the expressions of GLUT1, 2, 3, and 4 in two different gastric cancer cells (SNU-216, moderately differentiated gastric adenocarcinoma; and SNU-668, signet ring cell gastric carcinoma). Immunohistochemistry of GLUT1 and GLUT4 and immunoblot analysis of glycogen synthase kinase 3 were also performed. Hexokinase activity was measured. We found that 5-FU suppressed glucose uptake in SNU-216, while it stimulated GLUT in SNU-668. Further analysis revealed that 5-FU decreased the expression levels of GLUT1, 2, and 4 in SNU-216 cells and increased the expression levels of GLUT1, 2, and 4 in SNU-668 cells. Consistent with GLUT expression levels, immunohistochemistry analysis showed that 5-FU increased GLUT1 and GLUT4 levels in SNU-216 and decreased GLUT1 and GLUT4 levels in SNU-668. We also observed that glycogen synthase kinase 3 activity was decreased in SNU-216 and increased in SNU-668 with 5-FU treatment. No significant difference in hexokinase activities was observed with 5-FU treatment. Taken together, these results suggest that 5-FU exerts differential effects on GLUT depending on gastric cancer cell types, which may indicate a possible explanation, at least in part, for the differing responses to 5-FU chemotherapy in gastric cancer.

Epidermal growth factor increases prostaglandin E2 production via ERK1/2 MAPK and NF-kappaB pathway in fibroblast like synoviocytes from patients with rheumatoid arthritis.

High concentration of epidermal growth factor (EGF) is found in the synovial fluid of rheumatoid arthritis (RA) that might imply the involvement of EGF in the pathogenesis of arthritic diseases. In order to investigate if EGF is involved in the regulation of cyclooxygenase-2 (COX-2) and the prostaglandin E(2) (PGE(2)) production in fibroblast like synoviocytes (FLS) from patients with RA. The levels of COX-2 and microsomal prostaglandin E synthase-1 (mPGES-1) were evaluated using RT-PCR and Western blot analysis. Electrophoretic mobility shift assay (EMSA) was performed to investigate EGF mediated DNA binding activity of nuclear factor-kappaB (NF-kappaB). PGE(2) levels were analyzed by ELISA. EGF enhanced both COX-2 protein and mRNA expressions. mPGES-1 mRNA level was also increased by EGF treatment. EGF also stimulated ERK1/2 MAPK activity and the inhibition of ERK1/2 by PD098059 (ERK1/2 specific inhibitor) resulted in the suppression of EGF-induced COX-2 expression. The DNA binding activity of NF-kappaB was remarkably increased by EGF treatment and the pretreatment of PD098059 abolished EGF-stimulated NF-kappaB activity. We also observed that the level of PGE(2) was significantly elevated with the treatment of EGF in FLS, and the pretreatment of PD098059 abolished this stimulating effect. These results suggest that EGF is involved in the inflammatory process of RA by stimulating COX-2 expression and PGE(2) production. And EGF enhanced PGE(2) production appears to be mediated via ERK1/2 MAPK and NF-kappaB pathway in FLS.

Melatonin inhibits human fibroblast-like synoviocyte proliferation via extracellular signal-regulated protein kinase/P21(CIP1)/P27(KIP1) pathways.

The excessive proliferation and migration of synoviocytes are well-characterized phenomena that play key roles in the pathophysiology of rheumatoid arthritis (RA). Melatonin has been shown to have potent anti-proliferative effect in various cancer cells such as breast and prostate cancer cells. In this study, we examined the role of melatonin on synoviocyte proliferation in primary cultured human fibroblast-like synoviocytes (FLSs) by analyzing protein expression of P21(CIP1) (P21) and P27(KIP1) (P27), the cyclin-dependent kinase inhibitors that are important in cell cycle control, and the phosphorylation of mitogen-activated protein kinases (MAPKs). RA-FLS proliferation was determined by a [(3)H]-thymidine incorporation assay. Western blot analysis was applied to examine the underlying mechanisms of melatonin's effect. Melatonin inhibited RA-FLS proliferation in a dose-dependent manner. It reduced proliferation of passage 2 FLSs by 25% at 10 microm and by nearly 40% at 100 microm concentrations. The inhibitory effect of melatonin on RA-FLS proliferation was also observed in passages 4 and 6. Melatonin upregulated the expression levels of P21 and P27 dose-dependently (24 hr), induced the phosphorylation of extracellular signal-regulated protein kinase (ERK) time-dependently (10 microm), but did not affect phosphorylation of P38 in RA-FLSs. In addition, the expression of P21 and P27 triggered by melatonin was inhibited by the pretreatment of the ERK inhibitor, PD98059 (10 microm). The anti-proliferative action of melatonin in RA-FLSs was also blocked by PD98059. Taken together, these results suggest that melatonin exerts the inhibitory effect of the proliferation of RA-FLSs through the activation of P21 and P27 mediated by ERK. Hence we suggest that melatonin could be used as a therapeutic agent for the treatment of RA.

The association between genetic polymorphisms in CYP19 and breast cancer risk in Korean women.

Aromatase encoding by the CYP19 gene catalyzes the conversion of androgens to estrogens. In order to determine if polymorphisms of the CYP19 gene are associated with breast cancer risk, we analyzed the frequency of tetranucleotide (TTTA) tandem repeats and a 3-bp insertion (I)/deletion (D) polymorphism in intron 4 of the CYP19 gene in genomic DNA from 70 Korean breast cancer patients and 102 age-matched, healthy women. The 3-bp deletion allele was found more frequently in the breast cancer group than in the control group (p=0.001). Logistic regression analysis of the CYP 19 insertion/deletion (I/D) genotype showed a strong association between ID polymorphisms and breast cancer. The frequency of DD and ID alleles was significantly increased in the breast cancer group (DD genotype p=0.004, OR=12.81; and ID genotype p=0.005, OR=2.62). However, there were no differences in the genotype distributions of the (TTTA)n polymorphism of CYP19 between breast cancer patients and healthy controls. A positive association was noted between TTTA polymorphisms with 10 or more repeats and ER-negative tumors, as well as between lower repeat polymorphisms and ER-positive tumors (p=0.019). With respect to TTTA polymorphisms, we confirmed that the expression of aromatase in ER-positive MCF7 cells with 7-3 and 11 allele heterozygosity was significantly higher than in ER-negative MDA-MB231 cells with 11 allele homozygosity. These results suggest that 3-bp I/D polymorphisms of the CYP19 gene may be associated with breast cancer and that the (TTTA)n repeat genotype would be useful in selecting candidates for tamoxifen therapy, as well as predicting breast cancer risk in Korean women.

Activation of AMP-activated protein kinase by kainic acid mediates brain-derived neurotrophic factor expression through a NF-kappaB dependent mechanism in C6 glioma cells.

AMP-activated protein kinase (AMPK) is a key regulator of energy homeostasis. Kainic acid (KA), a prototype excitotoxin is known to induce brain-derived neurotrophic factor (BDNF) in brain. In this study, we examined the role of AMPK in KA-induced BDNF expression in C6 glioma cells. We showed that KA and KA receptor agonist induced activation of AMPK and KA-induced AMPK activation was blocked by inhibition of Ca(2+)/calmodulin-dependent protein kinase kinase (CaMKK) beta. We then showed that inhibition of AMPK by compound C, a selective inhibitor of AMPK, or small interfering RNA of AMPKalpha1 blocked KA-induced BDNF mRNA and protein expression. Inhibition of AMPK blocked KA-induced phosphorylation of CaMKII and I kappaB kinase (IKK) in C6 cells. Finally, we showed that inhibition of AMPK reduced DNA binding and transcriptional activation of nuclear factor-kappaB (NF-kappaB) in KA-treated cells. These results suggest that AMPK mediates KA-induced BDNF expression by regulating NF-kappaB activation.

Lipopolysaccharide induces hypoxia-inducible factor-1 alpha mRNA expression and activation via NADPH oxidase and Sp1-dependent pathway in BV2 murine microglial cells.

Hypoxia-inducible factor-1 (HIF-1), the key transcription factor of hypoxia-inducible genes, is known to be involved in inflammation and immune response, but little is known about the regulation of HIF-1 during microglial activation. Thus, we examined effect of lipopolysaccharide (LPS) on HIF-1 activation and its signaling mechanism in BV2 microglial cells. LPS induced HIF-1alpha mRNA and protein expression as well as HIF-1 transcriptional activation. Moreover, HIF-1alpha knockdown by small interfering RNA (siRNA) decreased LPS-induced expression of hypoxia responsive genes, VEGF, iNOS, and COX-2. We then showed that LPS-induced HIF-1alpha mRNA expression was blocked by an antioxidant, NADPH oxidase inhibitors, and siRNA of gp91phox, a subunit of NADPH oxidase. In addition, we showed that specific pharmacological inhibitors of PI 3-kinase and protein kinase C decreased LPS-induced HIF-1alpha mRNA expression. Finally, we showed that inhibition of transcription factor Sp1 by mithramycin A or Sp1 siRNA decreased LPS-induced HIF-1alpha mRNA and protein expression. Consistently, LPS increased Sp1 DNA binding and its transcriptional activity. Taken together, these results suggest that LPS induces HIF-1alpha mRNA expression and activation via NADPH oxidase and Sp1 in BV2 microglia.

Modulation of M1/M2 polarization by capsaicin contributes to the survival of dopaminergic neurons in the lipopolysaccharide-lesioned substantia nigra in vivo.

The present study examined the neuroprotective effects of capsaicin (CAP) and explored their underlying mechanisms in a lipopolysaccharide (LPS)-lesioned inflammatory rat model of Parkinson's dieases (PD). LPS was unilaterally injected into the substantia nigra (SN) in the absence or presence of CAP or capsazepine (CZP, a TRPV1 antagonist). The SN tissues were prepared for immunohistochemical staining, reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, western blot analysis, blood-brain barrier (BBB) permeability evaluation, and reactive oxygen species (ROS) detection. We found that CAP prevented the degeneration of nigral dopamine neurons in a dose-dependent manner and inhibited the expression of proinflammatory mediators in the LPS-lesioned SN. CAP shifted the proinflammatory M1 microglia/macrophage population to an anti-inflammatory M2 state as demonstrated by decreased expression of M1 markers (i.e., inducible nitric oxide synthase; iNOS and interleukin-6) and elevated expression of M2 markers (i.e., arginase 1 and CD206) in the SN. RT-PCR, western blotting, and immunohistochemical analysis demonstrated decreased iNOS expression and increased arginase 1 expression in the CAP-treated LPS-lesioned SN. Peroxynitrate production, reactive oxygen species levels and oxidative damage were reduced in the CAP-treated LPS-lesioned SN. The beneficial effects of CAP were blocked by CZP, indicating TRPV1 involvement. The present data indicate that CAP regulated the M1 and M2 activation states of microglia/macrophage in the LPS-lesioned SN, which resulted in the survival of dopamine neurons. It is therefore likely that TRPV1 activation by CAP has therapeutic potential for treating neurodegenerative diseases, that are associated with neuroinflammation and oxidative stress, such as PD.

[Corrigendum] Neurotoxin ß­N­methylamino­L­alanine induces endoplasmic reticulum stress­mediated neuronal apoptosis.

Following the publication of the article, the authors noted an error associated with the presentation of Fig. 4A. Fig 4 showed that overexpression of HSP70 suppresses ER stress-mediated neuronal death induced by ß­N­methylamino­L­alanine (BMAA). An error was made in the compilation of this Figure, and the band images shown in the HA panel for Fig. 1A were selected incorrectly. A corrected version of Fig. 4 is shown below. This change affects neither the interpretation of the data nor conclusions of this work. We regret that this error occurred, and thank the Editor for allowing us the opportunity to publish this Corrigendum. [the original article was published in the Molecular Medicine Reports 14: 4873-4880, 2016; DOI: 10.3892/mmr.2016.5802].

[Corrigendum] Neurotoxin ß­N­methylamino­L­alanine induces endoplasmic reticulum stress­mediated neuronal apoptosis.

Following the publication of the above article and a Corrigendum published in July 2018, the authors have noted an additional error, associated with the presentation of Fig. 1C. Fig 1C showed that ß­N­methylamino­L­alanine induces neuronal apoptotic cell death; however, an error was made in the compilation of this figure and an incorrect band image was selected for α­actinin, the loading control panel for Fig. 1C. A corrected version of Fig. 1 is shown opposite, incorporating the correct α­actinin protein bands in Fig. 1C. This change affects neither the interpretation of the data nor conclusions of this work. We regret that this further error went unnoticed at the time, and thank the Editor for allowing us the opportunity to publish this additional Corrigendum. [the original article was published in the Molecular Medicine Reports 14: 4873­4880, 2016; DOI: 10.3892/mmr.2016.5802].

Cordycepin-enriched WIB-801C from Cordyceps militaris improves functional recovery by attenuating blood-spinal cord barrier disruption after spinal cord injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Cordyceps militaris is an ingredient of traditional Chinese medicine and have been widely used for inflammatory diseases and cancer. Cordycepin is one of the major bioactive components of Cordyceps militaris, and has been known to have anti-inflammatory and anti-oxidant effects. AIM OF THIS STUDY: In the present study, we examined whether WIB-801C, a standardized and cordycepin-enriched extract of caterpillar fungus (Cordyceps militaris), would attenuate blood-spinal cord barrier (BSCB) disruption by inhibiting matrix metalloprotease (MMP)-9 activity, leading to improvement of functional outcomes after spinal cord injury (SCI). MATERIALS AND METHODS: Male Sprague-Dawley rats were subjected to contusive SCI using a New York University (NYU) impactor, and WIB-801C (50mg/kg) was administered at 2h and 8h after injury orally and further treated once a day for indicated time points. BSCB disruption, MMP-9 activity, blood infiltration, inflammation, neuronal apoptosis, axonal loss, demyelination, and neurological deficit were evaluated. RESULTS: We found that WIB-801C significantly attenuated BSCB disruption by inhibiting MMP-9 expression and activation after injury. The infiltration of neutrophils at 1 d and macrophage at 5 d after SCI was also ameliorated by WIB-801C as compared with vehicle control. In addition, the expression of inflammatory cytokines and mediators such as Tnf-α, IL-1ß, IL-6, Cox-2, and inos as well as chemokines such as Gro-α and Mip-2α was significantly inhibited by WIB-801C. Furthermore, WIB-801C inhibits p38MAPK activation and proNGF production in microglia after injury. These events eventually led to the inhibition of apoptotic cell death of neurons and oligodendrocytes, improved functional recovery and attenuated demyelination and axon loss after SCI. CONCLUSION: Our results suggest that WIB-801C can be used as a therapeutic agent after SCI by attenuating BSCB disruption followed inflammation.

Positive association of obesity with single nucleotide polymorphisms of syndecan 3 in the Korean population.

CONTEXT: Very recently the unforeseen role of syndecan 3 (SDC3), a family of membrane-bound heparin sulfate proteoglycans, in the regulation of energy balance has been discovered in the Sdc3 null female mice. OBJECTIVE: The objective of the study was to test the hypothesis that single nucleotide polymorphisms (SNPs) in SDC3 are associated with obesity in the Korean population. DESIGN/SETTING/SUBJECTS: We conducted a population-based cohort study consisting of 229 control and 245 study subjects and a second independent study consisting of 192 control and 115 study subjects. MAIN OUTCOME MEASUREMENT: Body mass index (BMI) was measured. RESULTS: First, Sdc3 mRNA expression in the brain of ob/ob mice was profoundly increased, compared with control mice. Next, all three nonsynonymous SNPs [T271I (rs2282440, C>T), D245N (rs4949184, C>T), and V150I (rs2491132, C>T)] in the SDC3 gene in control female subjects (BMI < 23, n = 229) and obese female subjects (BMI > 30, n = 245) were genotyped. We demonstrated the presence of clear ethnic differences in three nonsynonymous SDC3 SNPs among African-Americans, Chinese, Europeans, and Koreans. Of three SNPs in SDC3, rs4949184 was not associated with obesity and the other two SNPs (rs2282440 and rs2491132) were strongly associated with obesity (P < 0.0001), and the results were confirmed in the second independent study group. Haplotype analysis also revealed strong association with obesity (chi2 = 76.92, P < 0.000001). CONCLUSIONS: There are ethnic differences in the SDC3 polymorphisms, and the polymorphisms are strongly associated with obesity.

Activation of adenosine A3 receptor suppresses lipopolysaccharide-induced TNF-alpha production through inhibition of PI 3-kinase/Akt and NF-kappaB activation in murine BV2 microglial cells.

Adenosine is an endogenous nucleoside that regulates many processes, including inflammatory responses, through activation of its receptors. Adenosine receptors have been reported to be expressed in microglia, which are major immune cells of brain, yet little is known about the role of adenosine receptors in microglial cytokine production. Thus, we investigated the effect of adenosine and adenosine A3 receptor ligands on LPS-induced tumor necrosis factor (TNF-alpha) production and its molecular mechanism in mouse BV2 microglial cells. Adenosine and Cl-IB-MECA, a specific adenosine A3 receptor agonist, suppressed LPS-induced TNF-alpha protein and mRNA levels. Moreover, MRS1523, a selective A3 receptor antagonist, blocked suppressive effects of both adenosine and Cl-IB-MECA on TNF-alpha. We further examined the effect of adenosine on signaling molecules, such as PI 3-kinase, Akt, p38, ERK1/2, and NF-kappaB, which are involved in the regulation of inflammatory responses. Adenosine inhibited LPS-induced phosphatidylinositol (PI) 3-kinase activation and Akt phosphorylation, whereas it had no effect on the phosphorylation of p38 and ERK1/2. We also found that adenosine as well as Cl-IB-MECA inhibited LPS-induced NF-kappaB DNA binding and luciferase reporter activity. Taken together, these results suggest that adenosine A3 receptor activation suppresses TNF-alpha production by inhibiting PI 3-kinase/Akt and NF-kappaB activation in LPS-treated BV2 microglial cells.

Fluoxetine increases the nitric oxide production via nuclear factor kappa B-mediated pathway in BV2 murine microglial cells.

A body of recent evidence implicates that antidepressants affect the inflammatory response and immune system. The present study is focused on the effects of the most widely used antidepressant agent, fluoxetine on the production of nitric oxide (NO) in BV2 microglial cells. In this study, we observed interesting result that NO production was increased by fluoxetine. The mRNA level of nitric oxide synthase (iNos, Nos2) by RT-PCR was also stimulated by fluoxetine. We next conducted electophoretic mobility shift assay (EMSA) to determine the DNA binding activity of nuclear factor kappa B (Nfkappab), an important upstream modulator for Nos2 expression, to find that fluoxetine increased DNA binding activity of Nfkappab. By Western blot analysis, phosphorylation levels of p38 mitogen-activated protein kinase (p38 Mapk, Mapk14) and extracellular signal-related kinase (Erk)1/2 Mapk, upstream signaling mediators of Nfkappab were found to be increased by fluoxetine. In addition, the mRNA expressions of other proinflammatory cytokines, interleukin 6 (Il6) and tumor necrosis factor alpha (Tnfalpha) were examined. The expressions of both Il6 and Tnfalpha by fluoxetine treatment were similar to those of Nos2 and Nfkappab. Taken together, our results show that fluoxetine stimulates NO production via Nfkappab-mediated pathway in BV2 cells.