Phenylephrine Attenuated Sepsis-Induced Cardiac Inflammation and Mitochondrial Injury Through an Effect on the PI3K/Akt Signaling Pathway.

OBJECTIVE: To investigate whether phenylephrine (PE) inhibits sepsis-induced cardiac dysfunction, cardiac inflammation, and mitochondrial injury through the PI3K/Akt signaling pathway. METHODS: A rat model of sepsis was established by cecal ligation and puncture. PE and/or wortmannin (a PI3K inhibitor) were administered to investigate the role of PI3K/Akt signaling in mediating the effects of PE on inhibiting sepsis-induced cardiac dysfunction, cardiac inflammation, and mitochondrial injury. Hematoxylin-eosin staining, echocardiography, and Langendorff system were used to examine the myocardial injury and function. The concentrations of TNF-α and IL-6 were analyzed by enzyme-linked immunosorbent assay. Intercellular cell adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), myeloperoxidase, mitochondria-related fusion/fission proteins, and PI3K/Akt signaling pathway-associated proteins were analyzed by Western blotting. RESULTS: PE improved the cardiac function and survival in septic rats. PE decreased TNF-α, IL-6, ICAM-1, VCAM-1, and myeloperoxidase contents in the myocardium of septic rats. Meanwhile, PE increased the fusion-related proteins and decreased the fission-related proteins in the myocardial mitochondria of septic rats. On the other hand, PE activated the PI3K/Akt signaling pathway in the cecal ligation and puncture-treated rats, and all the protective effects of PE were abolished by wortmannin. CONCLUSIONS: PE attenuated sepsis-induced cardiac dysfunction, cardiac inflammation, and mitochondrial injury through the PI3K/Akt signaling pathway.

Baicalin administration attenuates hyperglycemia-induced malformation of cardiovascular system.

In this study, the effects of Baicalin on the hyperglycemia-induced cardiovascular malformation during embryo development were investigated. Using early chick embryos, an optimal concentration of Baicalin (6 µM) was identified which could prevent hyperglycemia-induced cardiovascular malformation of embryos. Hyperglycemia-enhanced cell apoptosis was reduced in embryos and HUVECs in the presence of Baicalin. Hyperglycemia-induced excessive ROS production was inhibited when Baicalin was administered. Analyses of SOD, GSH-Px, MQAE and GABAA suggested Baicalin plays an antioxidant role in chick embryos possibly through suppression of outwardly rectifying Cl(-) in the high-glucose microenvironment. In addition, hyperglycemia-enhanced autophagy fell in the presence of Baicalin, through affecting the ubiquitin of p62 and accelerating autophagy flux. Both Baicalin and Vitamin C could decrease apoptosis, but CQ did not, suggesting autophagy to be a protective function on the cell survival. In mice, Baicalin reduced the elevated blood glucose level caused by streptozotocin (STZ). Taken together, these data suggest that hyperglycemia-induced embryonic cardiovascular malformation can be attenuated by Baicalin administration through suppressing the excessive production of ROS and autophagy. Baicalin could be a potential candidate drug for women suffering from gestational diabetes mellitus.

Ginsenosides from stems and leaves of ginseng prevent ethanol-induced lipid accumulation in human L02 hepatocytes.

OBJECTIVE: To investigate the effect of ginsenosides from stems and leaves of ginseng on ethanol-induced lipid deposition in human L02 hepatocytes. METHODS: L02 cells were exposed to ethanol for 36 h and treated with or without ginsenosides. The viability of L02 cells was evaluated by methylthiazolyldiphenyl-tetrazolium bromide assay and the triglyceride (TG) content was detected. Lipid droplets were determined by oil red O staining. Intracellular reactive oxygen species (ROS) production and the mitochondrial membrane potential were tested by flow cytometry. The ATP level was measured by reverse phase high performance liquid chromatography. The expression of cytochrome p450 2E1 (CYP2E1) and peroxisome proliferator-activated receptor α (PPARα) was detected by reverse transcriptase-polymerase chain reaction and Western blotting, respectively. RESULTS: Ethanol exposure resulted in the increase of TG level, lipid accumulation and ROS generation, and the decrease of mitochondrial membrane potential and ATP production in the cells. However, ginsenosides significantly reduced TG content (9.69±0.22 µg/mg protein vs. 4.93±0.49 µg/mg protein, P<0.01), and ROS formation (7254.8±385.7 vs. 5825.2±375.9, P<0.01). Meanwhile, improvements in mitochondrial membrane potential (10655.33±331.34 vs. 11129.52±262.35, P<0.05) and ATP level (1.20±0.18 nmol/mg protein vs. 2.53±0.25 nmol/mg protein, P<0.01) were observed by treatment with ginsenosides. Furthermore, ginsenosides could down-regulate CYP2E1 expression (P<0.01) and upregulate PPARα expression (P<0.01) in ethanol-treated cells. CONCLUSIONS: Ginsenosides could prevent ethanol-induced hepatocyte steatosis in vitro related to the inhibition of oxidative stress and the improvement of mitochondrial function. In addition, the modulation of CYP2E1 and PPARα expression may also play an important role in the protective effect of ginsenosides against lipid accumulation.

From the Cover: Exposing Imidacloprid Interferes With Neurogenesis Through Impacting on Chick Neural Tube Cell Survival.

As a neonicotinoid pesticide, imidacloprid is widely used to control insects in agriculture and fleas on domestic animals. However, it is not known whether imidacloprid exposure negatively affects neurogenesis during embryonic development. In this study, using a chick embryo model, we investigated the effects of imidacloprid exposure on neurogenesis at the earliest stage and during late-stage embryo development. Exposing HH0 chick embryos to imidacloprid in EC culture caused neural tube defects (NTDs) and neuronal differentiation dysplasia as determined by NF/Tuj1 labeling. Furthermore, we found that F-actin accumulation on the apical side of the neural tube was suppressed by exposure to imidacloprid, and the expression of BMP4 and Shh on the dorsal and ventral sides of the neural tubes, respectively, were also reduced, which in turn affects the dorsolateral hinge points during bending of the neural plate. In addition, exposure to imidacloprid reduced cell proliferation and increased cell apoptosis, as determined by pHIS3 labeling and TUNEL staining, respectively, also contributing to the malformation. We obtained similar results in late-stage embryos exposed to imidacloprid. Finally, a bioinformatics analysis was employed to determine which genes identified in this study were involved in NTDs. The experimental evidence and bioinformatics analysis suggested that imidacloprid exposure during chick embryo development could increase the risk of NTDs and neural dysplasia.

Imidacloprid Exposure Suppresses Neural Crest Cells Generation during Early Chick Embryo Development.

Imidacloprid is a neonicotinoid pesticide that is widely used in the control pests found on crops and fleas on pets. However, it is still unclear whether imidacloprid exposure could affect early embryo development-despite some studies having been conducted on the gametes. In this study, we demonstrated that imidacloprid exposure could lead to abnormal craniofacial osteogenesis in the developing chick embryo. Cranial neural crest cells (NCCs) are the progenitor cells of the chick cranial skull. We found that the imidacloprid exposure retards the development of gastrulating chick embryos. HNK-1, PAX7, and Ap-2α immunohistological stainings indicated that cranial NCCs generation was inhibited after imidacloprid exposure. Double immunofluorescent staining (Ap-2α and PHIS3 or PAX7 and c-Caspase3) revealed that imidacloprid exposure inhibited both NCC proliferation and apoptosis. In addition, it inhibited NCCs production by repressing Msx1 and BMP4 expression in the developing neural tube and by altering expression of EMT-related adhesion molecules (Cad6B, E-Cadherin, and N-cadherin) in the developing neural crests. We also determined that imidacloprid exposure suppressed cranial NCCs migration and their ability to differentiate. In sum, we have provided experimental evidence that imidacloprid exposure during embryogenesis disrupts NCCs development, which in turn causes defective cranial bone development.

Effects of Senegenin against hypoxia/reoxygenation-induced injury in PC12 cells.

OBJECTIVE: To investigate the effect and the potential mechanism of Senegenin (Sen) against injury induced by hypoxia/reoxygenation (H/R) in highly differentiated PC12 cells. METHODS: The cultured PC12 cells were treated with H/R in the presence or absence of Sen (60 µmol/L). Four groups were included in the experiment: control group, H/R group, H/R+Sen group and Sen group. Cell viability of each group and the level of lactate dehydrogenase (LDH) in culture medium were detected for the pharmacological effect of Sen. Hoechst 33258 staining and annexin V/propidium iodide double staining were used to analyze the apoptosis rate. Moreover, mitochondrial membrane potential (△Ψm), reactive oxygen species (ROS) and intracellular free calcium ([Ca(2+)]i) were measured by fluorescent staining and flow cytometry. Cleaved caspase-3 and activity of NADPH oxidase (NOX) were determined by colorimetric protease assay and enzyme linked immunosorbent assay, respectively. RESULTS: Sen significantly elevated cell viability (P<0.05), decreased the leakage of LDH (P<0.05) and apoptosis rate (P<0.05) in H/R-injured PC12 cells. Sen maintained the value of △Ψm (P<0.05) and suppressed the activity of caspase-3 (P<0.05). Moreover, Sen reduced ROS accumulation P<0.05) and [Ca(2+)]i increment (P<0.05) by inhibiting the activity of NOX (P<0.05). CONCLUSION: Sen may exert cytoprotection against H/R injury by decreasing the levels of intracellular ROS and [Ca(2+)]i, thereby suppressing the mitochondrial pathway of cellular apoptosis.

Angiogenesis is repressed by ethanol exposure during chick embryonic development.

It is now known that excess alcohol consumption during pregnancy can cause fetal alcohol syndrome to develop. However, it is not known whether excess ethanol exposure could directly affect angiogenesis in the embryo or angiogenesis being indirectly affected because of ethanol-induced fetal alcohol syndrome. Using the chick yolk sac membrane (YSM) model, we demonstrated that ethanol exposure dramatically inhibited angiogenesis in the YSM of 9-day-old chick embryos, in a dose-dependent manner. Likewise, the anti-angiogenesis effect of ethanol could be seen in the developing vessel plexus (at the same extra-embryonic regions) during earlier stages of embryo development. The anti-angiogenic effect of ethanol was found associated with excess reactive oxygen species (ROS) production; as glutathione peroxidase activity increased while superoxide dismutase 1 and 2 activities decreased in the YSMs. We further validated this observation by exposing chick embryos to 2,2'-azobis-amidinopropane dihydrochloride (a ROS inducer) and obtained a similar anti-angiogenesis effect as ethanol treatment. Semiquantitative reverse transcription-polymerase chain reaction analysis of the experimental YSMs revealed that expression of angiogenesis-related genes, vascular endothelial growth factor and its receptor, fibroblast growth factor 2 and hypoxia-inducible factor, were all repressed following ethanol and 2,2'-azobis-amidinopropane dihydrochloride treatment. In summary, our results suggest that excess ethanol exposure inhibits embryonic angiogenesis through promoting superfluous ROS production during embryo development.

Ginsenoside Rg1 relieves tert-Butyl hydroperoxide-induced cell impairment in mouse microglial BV2 cells.

Microglial activation plays an important role in neurodegenerative diseases associated with oxidative stress. tert-Butyl hydroperoxide (t-BHP), an analog of hydroperoxide, mimics the oxidative damage to microglial cells. It has been reported that ginsenoside Rg1 (G-Rg1), an active ingredient of Panax ginseng, has anti-stress and anti-inflammatory properties. The present study aims to investigate the ability of G-Rg1 to decrease the t-BHP-mediated cell damage of BV2 microglial cells. We performed flow cytometry assays to facilitate the detection of reactive oxygen species as well as Western blotting analyses and immunofluorescence assays using specific antibodies, such as antibodies against phospho-mitogen-activated protein kinases (p-MAPKs), phospho-nuclear factor-κB (p-NF-κB), B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X (Bax), Caspase-3, autophagy marker light chain 3 (LC3), and Becline-1. We found that treatment with 50 µM G-Rg1 protected microglial cells against oxidative damage induced by 10 µM t-BHP.

Daucosterol protects neurons against oxygen-glucose deprivation/reperfusion-mediated injury by activating IGF1 signaling pathway.

We previously reported that daucosterol (a sterolin) up-regulates the expression of insulin-like growth factor I (IGF1)(1) protein in neural stem cells. In this study, we investigated the effects of daucosterol on the survival of cultured cortical neurons after neurons were subjected to oxygen and glucose deprivation and simulated reperfusion (OGD/R)(2), and determined the corresponding molecular mechanism. The results showed that post-treatment of daucosterol significantly reduced neuronal loss, as well as apoptotic rate and caspase-3 activity, displaying the neuroprotective activity. We also found that daucosterol increased the expression level of IGF1 protein, diminished the down-regulation of p-AKT(3) and p-GSK-3ß(4), thus activating the AKT(5) signal pathway. Additionally, it diminished the down-regulation of the anti-apoptotic proteins Mcl-1(6) and Bcl-2(7), and decreased the expression level of the pro-apoptotic protein Bax(8), thus raising the ratio of Bcl-2/Bax. The neuroprotective effect of daucosterol was inhibited in the presence of picropodophyllin (PPP)(9), the inhibitor of insulin-like growth factor I receptors (IGF1R)(10). Our study provided information about daucosterol as an efficient and inexpensive neuroprotectants, to which the IGF1-like activity of daucosterol contributes. Daucosterol could be potentially developed as a medicine for ischemic stroke treatment.

Autophagy is involved in high glucose-induced heart tube malformation.

Both pre-gestational and gestational diabetes have an adverse impact on heart development, but little is known about the influence on the early stage of heart tube formation. Using early gastrulating chick embryos, we investigated the influence of high glucose on the process of heart tube formation, specifically during the primary heart field phase. We demonstrated that high-glucose exposure resulted in 3 types of heart tube malformation: 1) ventricular hypertrophy, 2) ventricular hypertrophy with dextrocardia and 3) ventricular hypertrophy and dextrocardia with the fusion anomaly of a bilateral primary heart tube. Next, we found that these malformation phenotypes of heart tubes might mainly originate from the migratory anomaly of gastrulating precardiac mesoderm cells rather than cell proliferation in the developmental process of bilateral primary heart field primordia. The treatment of rapamycin (RAPA), an autophagy inducer, led to a similar heart tube malformation phenotype as high glucose. Additionally, high-glucose exposure promoted the expression of the key autophagy protein LC3B in early chick tissue. Atg7 is strongly expressed in the fusion site of bilateral primary heart tubes. All of these data imply that autophagy could be involved in the process of high-glucose-induced malformation of the heart tube.

Daucosterol promotes the proliferation of neural stem cells.

Neural stem cells (NSCs) are self-regenerating cells, but their regenerative capacity is limited. The present study was conducted to investigate the effect of daucosterol (a sterolin) on the promotion of NSC proliferation and determine the corresponding molecular mechanism. Results of cell counting kit-8 (CCK-8) assay showed that daucosterol significantly increased the quantity of viable cells and the effectiveness of daucosterol was similar to that of basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). Flow cytometry detection of CFSE-labeled (CFSE, carboxyfluorescein diacetate succinimidyl ester) NSCs showed that Div Index (or the average number of cell divisions) and % Divided (or the percentage of cells that divided at least once) of the cells were increased, indicating that daucosterol increased the percentage of NSCs re-entering the cell cycle. mRNA microarray analysis showed that 333 genes that are mostly involved in the mitotic cell cycle were up-regulated. By contrast, 627 genes that are mostly involved in differentiation were down-regulated. In particular, insulin-like growth factor I (IGF1) was considered as an important regulatory gene that functionally promoted NSC proliferation, and the increased expression of IGF1 protein was validated by ELISA. In addition, the phosphorylation of AKT was increased, indicating that the proliferation-enhancing activity of daucosterol may be involved in IGF1-AKT pathway. Our study provided information about daucosterol as an efficient and inexpensive growth factor alternative that could be used in clinical medicine and research applications.

[Effects of antagonist and agonist of nicotinic acetylcholine receptors on injury of rat neurons induced by amyloid ß-protein].

OBJECTIVE: To explore the chronic effects of nicotinic antagonist and agonist on rat neurons injury induced by ß-amyloid protein. METHODS: The rat model of neuron injury was established by the exposure to Aß25-35 and the intervention agent was either methyllycaconitine (MLA) or nicotine (Nic). And the experimental groups were control (distilled water), Aß25-35, MLA (MLA and Aß25-35) and Nic (Nic and Aß25-35). Cellular viability was detected by methyl thiazolyl tetrazolium (MTT) chromatometry while apoptosis and necrosis were detected by flow cytometer. RESULTS: Compared with control, cellular viability decreased while the apoptotic and necrotic rates increased in Aß25-35 group(P = 0.00). The values of cellular viability at (0.75 ± 0.02) and (0.75 ± 0.09) in Aß25-35 and MLA groups respectively were significantly lower than that of Nic group (0.81 ± 0.02, P = 0.01) at Day 3 and 7. No significant differences existed in cellular viability between Aß25-35 and MLA groups. At Day 14, the differences of cellular viability were not obvious in all groups. At Day 21, cell viability of MLA group (0.64 ± 0.10) was significantly higher than those of Aß25-35 (0.57 ± 0.04, P = 0.019) and Nic groups (0.56 ± 0.04, P = 0.008). The apoptotic rate was lower than that of Aß25-35 group (3.70% ± 0.20% vs 4.70% ± 0.46%, P = 0.008) while the necrotic rate lower than that of Aß25-35 group (7.73% ± 0.86% vs 16.30% ± 1.05%, P = 0.00) and Nic group (16.03% ± 1.53%, P = 0.00). However, no significant differences existed in cellular viability or apoptotic and necrotic rate between Aß25-35 and Nic groups. CONCLUSION: With chronic treatment, the protective effect of α7 nicotinic antagonist methyllycaconitine increases whereas that of nicotinic agonist nicotine decreases.

Berberine inhibits norepinephrine-induced apoptosis in neonatal rat cardiomyocytes via inhibiting ROS-TNF-α-caspase signaling pathway.

OBJECTIVE: To determine the effect of berberine (Ber) on norepinephrine (NE)-induced apoptosis in neonatal rat cardiomyocytes. METHODS: The cultured neonatal rat cardiomyocytes were treated with NE in the presence or absence of Ber. The activity of lactate dehydrogenase (LDH) in the culture medium was examined, and apoptosis of cardiomyocytes was assessed by Hoechst 33258, isothiocyanate (FITC)-conjugated annexin-V, and propidine iodide (PI) staining. In addition, the activities of caspases-2 and-3 were measured by a fluorescent assay kit. The level of secreted tumor necrosis factor α (TNF-α) and production of intracellular reactive oxygen species (ROS) were also determined. RESULTS: NE at a concentration of 50 µ mol/L induced an obvious increase in the activity of LDH in the culture medium (P<0.05), which was inhibited by coincubation with 0.5, 1.0, or 2.0 µ mol/L Ber (P<0.05). Ber also significantly attenuated NE-induced apoptosis in a dose-dependent manner (P<0.01). Moreover, Ber at a dose of 2 µ mol/L markedly decreased the ROS and TNF-α productions (P <0.05) and inhibited the activation of caspases-2 and -3 in cardiomyocytes exposed to NE (P<0.05)h. CONCLUSION: The present study suggested that Ber could reduce NE-induced apoptosis in neonatal rat cardiomyocytes through inhibiting the ROS-TNF-α-caspase signaling pathway.

Microarray Analysis of mRNA and MicroRNA Expression Profile Reveals the Role of ß -Sitosterol-D-glucoside in the Proliferation of Neural Stem Cell.

Neural stem cells (NSCs) are self-regenerating cells, but their regenerative capacity is limited. The present study was conducted to investigate the effect of ß -sitosterol-D-glucoside (BSSG) on the proliferation of hippocampal NSCs and to determine the corresponding molecular mechanism. Results of CCK-8 assay showed that BSSG significantly increased NSC proliferation and the effectiveness of BSSG was similar to that of basic fibroblast growth factor and epidermal growth factor. mRNA expression profiling showed that 960 genes were differentially expressed after NSCs were treated with BSSG. Among the 960 genes, IGF1 is considered as a key regulatory gene that functionally promotes NSC proliferation. MicroRNA (miRNA) expression profiling indicated that 30 and 84 miRNAs were upregulated and downregulated, respectively. miRNA-mRNA relevance analysis revealed that numerous mRNAs including IGF1 mRNA were negatively regulated by miRNAs with decreased expression, thereby increasing the corresponding mRNA expression. The increased expression of IGF1 protein was validated by ELISA. Picropodophyllin (PPP, an inhibitor of IGF-1R) inhibition test confirmed that the proliferation-enhancing effect depended on IGF1. This study provided information about BSSG as an efficient and inexpensive growth factor alternative, of which the effect is closely involved in IGF1.

Luteolin reduces primary hippocampal neurons death induced by neuroinflammation.

OBJECTIVES: This study examined whether luteolin may exert an anti-inflammatory effect in microglia and may be neuroprotective by regulating microglia activation. METHODS: We treated BV2 microglia with 1.0 µg/ml lipopolysaccharide (LPS) after incubation with luteolin for 1 hour, the nitric oxide (NO) levels were determined by a Griess reaction, the inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor-alpha (TNF-alpha), and interleukin 1beta (IL-1beta) mRNA expression were determined by real-time PCR analysis, the iNOS and COX-2 protein induction were determined by Western blot analysis, and the levels of prostaglandin E(2) (PGE(2)), TNF-alpha, and IL-1beta were determined by enzyme-linked immunosorbent assay (ELISA) kits. Rat primary hippocampal neurons were co-cultured with LPS-activated BV2 microglia with 20 µM luteolin for 24 hours, the hippocampal neurons viability was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and the number of apoptotic hippocampal neurons was determined by immunofluorescence detection. RESULTS: Luteolin significantly inhibited the expression of iNOS and COX-2 in LPS-induced BV2 microglia. Moreover, the compound down-regulated the proinflammatory cytokines (TNF-alpha and IL-1beta) as well as the production of NO and PGE(2) in these cells. When hippocampal neurons were co-cultured with LPS-stimulated BV2 microglia, the administration of 20 µM luteolin increased the neurons viability and reduced the number of apoptotic neurons. CONCLUSION: These data demonstrate that anti-inflammatory activity of luteolin in microglia contributes to its neuroprotective effect and suggest that it may have a potential therapeutic application in the treatment of neurodegenerative diseases.

Berberine protects against lipopolysaccharide-induced intestinal injury in mice via alpha 2 adrenoceptor-independent mechanisms.

AIM: To investigate the mechanisms responsible for the protective action of berberine (Ber) against gut damage in endotoxemic mice. METHODS: Male BALB/c mice were administered intragastrically with distilled water (0.1 mL/10 g), Ber (50 mg/kg) alone, yohimbine (2 mg/kg) alone, or Ber (50 mg/kg) in combination with yohimbine (2 mg/kg) for 3 d. On the third day, lipopolysaccharide (LPS, 18 mg/kg) or normal saline was intraperitoneally injected one hour after the intragastric administration. Following the treatment, intestinal injury in the ileum was histopathologically accessed; enterocyte apoptosis was examined using TUNEL method; Toll-like receptor 4 (TLR4) mRNA expression was measured using RT-PCR assay; inhibitor protein-κBα (I-κBα) phosphorylation and myeloperoxidase content were examined using Western blloting. The macrophage inflammatory protein-2 (MIP-2) production was measured using ELISA assay. RESULTS: Mice challenged with LPS caused extensive ileum injury, including a significantly increased injury score, decreased intestinal villus height, reduced gut mucosal weight and increased intestinal permeability. Furthermore, LPS significantly induced enterocyte apoptosis, increased TLR4 mRNA expression, I-κBα phosphorylation, MIP-2 production and myeloperoxidase content in the ileum. Pretreatment with Ber significantly alleviated all the alterations in the ileum in the endotoxemic mice. Pretreatment with the α2-adrenoceptor antagonist yohimbine did not block the protective action of Ber against LPS-induced intestinal injury. In addition, treatment with yohimbine alone did not prevent LPS-induced intestinal injury. CONCLUSION: Pretreatment with Ber provides significant protection against LPS-induced intestinal injury in mice, via reducing enterocyte apoptosis, inhibiting the TLR4-nuclear factor κB-MIP-2 pathway and decreasing neutrophil infiltration that are independent of α2-adrenoceptors.

Luteolin inhibits microglial inflammation and improves neuron survival against inflammation.

Microglia activation is one of the causative factors for neuroinflammation, which results in brain damage during neurodegenerative disease. Accumulating evidence has shown that the flavonoid luteolin (Lut) possesses potent anti-inflammatory properties; however, its effect on microglia inhibition is currently unknown. Moreover, it is not clear whether Lut also has indirect neuroprotective effects by reducing inflammatory mediators and suppressing microglia activation. In this study, we examined the effects of Lut on lipopolysaccharide (LPS)-induced proinflammatory mediator production and signaling pathways in murine BV2 microglia. In addition, we cocultured microglia and neurons to observe the indirect neuroprotective effects of Lut. Lut inhibited the LPS-stimulated expression of inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor alpha (TNF-α), and interleukin-1ß (IL-1ß) as well as the production of nitric oxide (NO) and prostaglandin E(2) (PGE(2)). Moreover, Lut blocked LPS-induced nuclear factor kappa B (NF-κB) activation. Preincubation of microglia with Lut diminished the neurotoxic effects, owing to the direct anti-inflammatory effects of the compound. Taken together, our findings suggest that Lut may have a potential therapeutic application in the treatment of neuroinflammatory disorders.

Effect of siRNAs on HSV-1 plaque formation and relative expression levels of RR mRNA.

RNA interference (RNAi) is a process by which introduced small interfering RNA (siRNA) can cause the specific degradation of mRNA with identical sequences. The human herpes simplex virus type 1 (HSV-1) RR is composed of two distinct homodimeric subunits encoded by UL39 and UL40, respectively. In this study, we applied siRNAs targeting the UL39 and UL40 genes of HSV-1. We showed that synthetic siRNA silenced effectively and specifically UL39 and UL40 mRNA expression and inhibited HSV-1 replication. Our work offers new possibilities for RNAi as a genetic tool for inhibition of HSV-1 replication.

Pretreatment with berberine and yohimbine protects against LPS-induced myocardial dysfunction via inhibition of cardiac I-[kappa]B[alpha] phosphorylation and apoptosis in mice.

Myocardial dysfunction is a common complication in sepsis and significantly contributes to the mortality of patients with septic shock. Our previous study demonstrated that pretreatment with berberine (Ber) protected against the lethality induced by LPS, which was enhanced by yohimbine, an [alpha]2-adrenergic receptor antagonist, and Ber combined with yohimbine also improved survival in mice subjected to cecal ligation and puncture. However, no studies have examined whether Ber and yohimbine reduce LPS-induced myocardial dysfunction. Here, we report that pretreatment with Ber, Ber combined with yohimbine, or yohimbine significantly reduced LPS-induced cardiac dysfunction in mice. LPS-provoked cardiac apoptosis, I-[kappa]B[alpha] phosphorylation, IL-1[beta], TNF-[alpha], and NO production were attenuated by pretreatment with Ber and/or yohimbine, whereas cardiac Toll-like receptor 4 mRNA expression, malondialdehyde content, and superoxide dismutase activity were not affected. These data demonstrate for the first time that pretreatment with Ber and/or yohimbine prevents LPS-induced myocardial dysfunction in mice through inhibiting myocardial apoptosis, cardiac I-[kappa]B[alpha] phosphorylation, and TNF-[alpha], IL-1[beta], and NO production, suggesting that activation of [alpha]2-adrenergic receptor in vivo may be responsible at least in part for LPS-induced cardiac dysfunction, and Ber in combination with yohimbine may be a potential agent for preventing cardiac dysfunction during sepsis.

In vitro anti-viral activity of the total alkaloids from Tripterygium hypoglaucum against herpes simplex virus type 1.

Herpes simplex virus type 1 (HSV-1) is a commonly occurring human pathogen worldwide. There is an urgent need to discover and develop new alternative agents for the management of HSV-1 infection. Tripterygium hypoglaucum (level) Hutch (Celastraceae) is a traditional Chinese medicine plant with many pharmacological activities such as anti-inflammation, anti-tumor and antifertility. The usual medicinal part is the roots which contain about a 1% yield of alkaloids. A crude total alkaloids extract was prepared from the roots of T. hypoglaucum amd its antiviral activity against HSV-1 in Vero cells was evaluated by cytopathic effect (CPE) assay, plaque reduction assay and by RT-PCR analysis. The alkaloids extract presented low cytotoxicity (CC(50) = 46.6 µg/mL) and potent CPE inhibition activity, the 50% inhibitory concentration (IC(50)) was 6.5 µg/mL, noticeably lower than that of Acyclovir (15.4 µg /mL). Plaque formation was significantly reduced by the alkaloids extract at concentrations of 6.25 µg/mL to 12.5 µg/mL, the plaque reduction ratio reached 55% to 75 which was 35% higher than that of Acyclovir at the same concentration. RT-PCR analysis showed that, the transcription of two important delayed early genes UL30 and UL39, and a late gene US6 of HSV-1 genome all were suppressed by the alkaloids extract, the expression inhibiting efficacy compared to the control was 74.6% (UL30), 70.9% (UL39) and 62.6% (US6) respectively at the working concentration of 12.5 µg/mL. The above results suggest a potent anti-HSV-1 activity of the alkaloids extract in vitro.