The anti-inflammatory role of α7 nicotinic acetylcholine receptor in microglial cells and its mechanisms / 药学实践杂志

Objective To investigate the anti-inflammatory role of α7 nicotinic acetylcholine receptor (α7nAChR) under inflammatory stress and its mechanisms. Methods PNU282987 was used for the activation of α7nAChR and LPS was administrated as inflammatory stressor. Realtime PCR was used for the detection of IL-1β, IL-6, TNF-α, M1 macrophage marker CD68, CD86 and M2 macrophage marker CD206, Arg1. Cell immunofluorescence was used for the detection of M1/M2 ratio and Western blot was applied for the detection of autophagy-related proteins. Results Under the stimulation of LPS, the mRNA levels of proinflammatory cytokines IL-1β, IL-6 and TNF-α, the proportion of M1 macrophage and autophagy process were increased in BV2 microglial cells. However, the administration of PNU282987 significantly decreased the mRNA levels of IL-1β, IL-6 and TNF-α and the proportion of M1 macrophage while increased the proportion of M2 macrophage and the level of autophagy process. Conclusion Activating α7nAChR plays an anti-inflammatory role in microglial cells under inflammatory stress due to the regulation of M1/M2 macrophage ratio and increase of autophagy level.

Electroacupuncture at "Jiaji" (EX-B2) induced down-regulation of OX-42 and P2X4 in lumbar spinal cord contributes to its analgesic after-effect in rats with chronic constriction injury / 针刺研究

OBJECTIVE: To investigate the effect of electroacupuncture (EA) at "Jiaji" (EX-B2) at different time points on the expression of OX-42 (a monoclonal antibody with specific expression of complement receptor-3 in spinal microglial cells) and purinergic receptor P2X4 (P2X4) in rats with chronic constriction injury (CCI) of the sciatic nerve, as well as the possible after-effect mechanism of EA analgesia in neuropathic pain. METHODS: Sprague-Dawley rats were randomly divided into blank group, model group, immediately after EA group, 0.5-hour after EA group, 1-hour after EA group, 2-hour after EA group, 4-hour after EA group, 12-hour after EA group, and 24-hour after EA group, with 6 rats in each group. The rats in the model group and the EA groups were used to establish a model of CCI-induced neuropathic pain, and those in the immediately after EA group and the 0.5, 1, 2, 4, 12 and 24 hours after EA groups were treated with EA at bilateral L3 and L5 "Jiaji" points for 20 min after 7 d of modeling. Samples were collected immediately and at 0.5, 1, 2, 4, 12, and 24 hours after EA, and for the rats in the blank group and the model group, samples were collected after fixation the rats for 20 min. Heat pain threshold was observed before and after intervention, and immunohistochemistry was used to measure the protein expression of OX-42 and P2X4 in the spinal cord lumbar enlargement. RESULTS: After 7 days of modeling (before intervention), compared with the blank group, the heat pain threshold had a significant reduction in the model group and the EA groups (P<0.01). Compared with the model group after intervention, the immediately after EA group and the 0.5, 1 and 2 hours after EA groups had a significant increase in heat pain threshold (P<0.05). Compared with the model group, immediately after EA, the 0.5, 1 and 2 hours after EA groups had a significant reduction in the protein expression of OX-42 (P<0.01), and immediately after EA, the 0.5 and 1 hour after EA groups had a significant reduction in the protein expression of P2X4 (P<0.01). CONCLUSION: EA at "Jiaji" points can significantly increase heat pain threshold and down-regulate the protein expression of OX-42 and P2X4 in the spinal cord of CCI rats. The analgesic effect can last for 2 h.

Attenuation of lipopolysaccharide-induced neuroinflammatory events in BV-2 microglial cells by Moringa oleifera leaf extract

Objective: To determine the anti-neuroinflammatory activity of Moringa oleifera leaf extract (MLE) under lipopolysaccharide stimulation of mouse murine microglia BV2 cells in vitro. Methods: The cytotoxicity effect of MLE was investigated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide assay. The inflammatory response of BV-2 cells were induced with lipopolysaccharide. The generation of nitric oxide levels was determined by using Griess assay and the level of pro-inflammatory cytokines (IL-1β, IL-6 and TNF-α) was evaluated by ELISA kit. The expression of iNOS, COX-2 as well as IκB-α was carried out by immunoblot analysis. Results: MLE reduced the nitric oxide production in concentration-dependent manner, and maintained the viability of BV-2 microglial cells which indicated absence of toxicity. In addition, MLE repressed the activation of nuclear factor kappa B by arresting the deterioration of IκB-α, consequently resulted in suppression of cytokines expression such as COX-2 and iNOS. Conclusions: MLE inhibitory activities are associated with the inhibition of nuclear factor kappa B transcriptional activity in BV2 microglial cells. Thus MLE may offer a substantial treatment for neuroinflammatory diseases.

Attenuation of lipopolysaccharide-induced neuroinflammatory events in BV-2 microglial cells by Moringa oleifera leaf extract

Objective: To determine the anti-neuroinflammatory activity of Moringa oleifera leaf extract (MLE) under lipopolysaccharide stimulation of mouse murine microglia BV2 cells in vitro. Methods: The cytotoxicity effect of MLE was investigated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5- diphenyl-tetrazolium bromide assay. The inflammatory response of BV-2 cells were induced with lipopolysaccharide. The generation of nitric oxide levels was determined by using Griess assay and the level of pro-inflammatory cytokines (IL-1β, IL-6 and TNF-α) was evaluated by ELISA kit. The expression of iNOS, COX-2 as well as IκB-α was carried out by immunoblot analysis. Results: MLE reduced the nitric oxide production in concentration-dependent manner, and maintained the viability of BV-2 microglial cells which indicated absence of toxicity. In addition, MLE repressed the activation of nuclear factor kappa B by arresting the deterioration of IκB-α, consequently resulted in suppression of cytokines expression such as COX-2 and iNOS. Conclusions: MLE inhibitory activities are associated with the inhibition of nuclear factor kappa B transcriptional activity in BV2 microglial cells. Thus MLE may offer a substantial treatment for neuroinflammatory diseases.

Anti-neuroinflammatory Effects of 12-Dehydrogingerdione in LPS-Activated Microglia through Inhibiting Akt/IKK/NF-κB Pathway and Activating Nrf-2/HO-1 Pathway

Ginger, one of worldwide consumed dietary spice, is not only famous as food supplements, but also believed to exert a variety of remarkable pharmacological activity as herbal remedies. In this study, a ginger constituent, 12-dehydrogingerdione (DHGD) was proven that has comparable anti-inflammatory activity with positive control 6-shogaol in inhibiting LPS-induced interleukin (IL)-6, tumor necrosis factor (TNF)-α, prostaglandin (PG) E₂, nitric oxide (NO), inducible NO synthase (iNOS) and cyclooxygenase (COX)-2, without interfering with COX-1 in cultured microglial cells. Subsequent mechanistic studies indicate that 12-DHGD may inhibit neuro-inflammation through suppressing the LPS-activated Akt/IKK/NF-κB pathway. Furthermore, 12-DHGD markedly promoted the activation of NF-E2-related factor (Nrf)-2 and heme oxygenase (HO)-1, and we demonstrated that the involvement of HO-1 on the production of pro-inflammatory mediators such as NO and TNF-α by using a HO-1 inhibitor, Zinc protoporphyrin (Znpp). These results indicate that 12-DHGD may protect against neuro-inflammation by inhibiting Akt/IKK/IκB/NF-κB pathway and promoting Nrf-2/HO-1 pathway.

Effects of electroacupuncture on activation of microglia cells in spinal cord in rats with neuropathic pain / 中国针灸

<p><b>OBJECTIVE</b>To observe the effects of electroacupuncture (EA) on the activation of microglia cells in the Lto Lspinal cord in rats with neuropathic pain, so as to investigate whether EA could inhibit the activation of spinal microglial cells and regulate the expression of brain-derived neurotrophic factor (BDNF) to achieve the analgesic effect.</p><p><b>METHODS</b>Forty male Sprague Dawley rats were randomly divided into a normal group, a sham-model group, a model group and an EA group, 10 rats in each one. The rats in the normal group received no treatment; the rats in sham-model group were treated with operation to exposure sciatic nerve for 2 to 3 min (no knot); the rats in the remaining groups were treated with model establishment of chronic constrictive injury (CCI). 7 days after model establishment, the rats in the EA group were treated with EA at "Zusanli" (ST 36) and "Yanglingquan" (GB 34), 30 min per time, once a day for consecutive 7 days. Only immobilization was used in the remaining groups the mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) of affected side feet were respectively measured before model establishment and 3 days, 5 days, 7 days, 10 days, 12 days and 14 days after model establishment; 14 days after model establishment, rats were sacrificed; the immunohistochemical method was used to measure the expression of Iba1 and BDNF in the sample of Lto Lspinal cord; real-time fluorescent quantitative PCR was used to measure the expression BDNF mRNA.</p><p><b>RESULTS</b>Compared with the sham-model group, the pain threshold was decreased significantly in the model group (<0.05), leading to hyperpathia. After EA treatment, compared with the model group, the pain threshold was increased significantly in the EA group (<0.05). 14 days after operation, the microglia cells in the Lto Lspinal cord, expression of BDNF and level of mRNA in the model group were significantly higher than those in the normal group and sham-model group (all<0.01); those in the EA group were significantly lower than those in the model group (all<0.01).</p><p><b>CONCLUSIONS</b>The analgesic effect on neuropathic pain is likely to be achieved by EA through inhibiting the activation of spinal microglia cells and down-regulating the expression of BDNF.</p>

The Anti-Inflammatory Activity of Eucommia ulmoides Oliv. Bark. Involves NF-κB Suppression and Nrf2-Dependent HO-1 Induction in BV-2 Microglial Cells

In the present study, we investigated the anti-inflammatory properties of Eucommia ulmoides Oliv. Bark. (EUE) in lipopolysaccharide (LPS)-stimulated microglial BV-2 cells and found that EUE inhibited LPS-mediated up-regulation of pro-inflammatory response factors. In addition, EUE inhibited the elevated production of pro-inflammatory cytokines, mediators, and reactive oxygen species (ROS) in LPS-stimulated BV-2 microglial cells. Subsequent mechanistic studies revealed that EUE suppressed LPS-induced phosphorylation of mitogen-activated protein kinases (MAPKs), phosphoinositide-3-kinase (PI3K)/Akt, glycogen synthase kinase-3β (GSK-3β), and their downstream transcription factor, nuclear factor-kappa B (NF-κB). EUE also blocked the nuclear translocation of NF-κB and inhibited its binding to DNA. We next demonstrated that EUE induced the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) and upregulated heme oxygenase-1 (HO-1) expression. We determined that the significant up-regulation of HO-1 expression by EUE was a consequence of Nrf2 nuclear translocation; furthermore, EUE increased the DNA binding of Nrf2. In contrast, zinc protoporphyrin (ZnPP), a specific HO-1 inhibitor, blocked the ability of EUE to inhibit NO and PGE2 production, indicating the vital role of HO-1. Overall, our results indicate that EUE inhibits pro-inflammatory responses by modulating MAPKs, PI3K/Akt, and GSK-3β, consequently suppressing NF-κB activation and inducing Nrf2-dependent HO-1 activation.

Vaccinium bracteatum Thunb. Exerts Anti-Inflammatory Activity by Inhibiting NF-κB Activation in BV-2 Microglial Cells

This study was designed to evaluate the pharmacological effects of Vaccinium bracteatum Thunb. methanol extract (VBME) on microglial activation and to identify the underlying mechanisms of action of these effects. The anti-inflammatory properties of VBME were studied using lipopolysaccharide (LPS)-stimulated BV-2 microglial cells. We measured the production of nitric oxide (NO), inducible NO synthase (iNOS), cyclooxygenase (COX)-2, prostaglandin E₂ (PGE₂), tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and interleukin-6 (IL-6) as inflammatory parameters. We also examined the effect of VBME on intracellular reactive oxygen species (ROS) production and the activity of nuclear factor-kappa B p65 (NF-κB p65). VBME significantly inhibited LPS-induced production of NO and PGE₂ and LPS-mediated upregulation of iNOS and COX-2 expression in a dose-dependent manner; importantly, VBME was not cytotoxic. VBME also significantly reduced the generation of the pro-inflammatory cytokines TNF-α, IL-1β, and IL-6. In addition, VBME significantly dampened intracellular ROS production and suppressed NF-κB p65 translocation by blocking IκB-α phosphorylation and degradation in LPS-stimulated BV2 cells. Our findings indicate that VBME inhibits the production of inflammatory mediators in BV-2 microglial cells by suppressing NF-κB signaling. Thus, VBME may be useful in the treatment of neurodegenerative diseases due to its ability to inhibit inflammatory mediator production in activated BV-2 microglial cells.

Inhibitory effects of acteoside on LPS-induced inflammatory response on BV-2 microglial cells / 中国中药杂志

To investigate the inhibitory effects of acteoside (ACT) on BV-2 microglial cells and the potential mechanism,LPS was used to treat BV-2 cells with or without ACT (12.5,25,50 μmol•L ⁻¹). Then, the expressions of inflammatory factors (NO,TNF-α,IL-6) and inflammation related proteins (iNOS,COX-2,p-IKKβ,IKKβ,p-ⅠκB,ⅠκB) were detected. In addition,the nuclear translocation of NF-κB was explored. The results showed that ACT could significantly suppress the inflammatory response against LPS stimulation by decreasing the expressions of NO,IL-6,TNF-α,iNOS,COX-2 and the phosphorylations of IKKβ and IκB. Moreover,the nuclear translocation of NF-κB p65 was inhibited by ACT. Taken together, ACT could significantly inhibit the inflammatory response of BV-2 microglial cells which were induced by LPS via inhibition of NF-κB signaling pathway.

Effect of baicalin on BV2 cell activation damage caused by rotenone / 中国生化药物杂志

Objective To observe Effect of baicalin on BV2 cell activation damage caused by rotenone.Methods Rotenone infected BV2 cell viability were detected by MTT assay, and OX42 expression by ICC method assay, the inner cell Ca2 + content by fluorescence spectrophotometry, intracellular iron content by ICP-AES when low concentrations iron of load without damage in cells and extracellular H2 O2 and intracellular content of MDA, NO, NOS with related kits.ResuIts Rotenone exposure BV2 cell reduced cell viability, the expression of cell marker protein OX42 and intracellular Ca2 +content and intracellular iron content were all significantly increased(P<0.05), resulting in oxidative damage to cells.Baicalin reduced OX42 expression of rotenone exposure BV2 cell, increased cells survival, reduced intracellular iron content and H2 O2 release and oxidative damage of BV2(P<0.05).ConcIusion Baicalin inhibits BV2 activation and damage by rotenone-induced, and it has neuroprotective effects.The protection mechanism may relate to reduce iron into cells .

3,4,5-Trihydroxycinnamic Acid Inhibits LPS-Induced iNOS Expression by Suppressing NF-kappaB Activation in BV2 Microglial Cells

Although various derivatives of caffeic acid have been reported to possess a wide variety of biological activities such as neuronal protection against excitotoxicity and anti-inflammatory property, the biological activity of 3,4,5-trihydroxycinnamic acid (THC), a derivative of hydroxycinnamic acids, has not been clearly examined. The objective of the present study is to evaluate the anti-inflammatory effects of THC on lipopolysaccharide (LPS)-stimulated BV2 microglial cells. THC significantly suppressed LPS-induced excessive production of nitric oxide (NO) and expression of iNOS, which is responsible for the production of iNOS. THC also suppressed LPS-induced overproduction of pro-inflammatory cytokines such as IL-1beta and TNF-alpha in BV2 microgilal cells. Furthermore, THC significantly suppressed LPS-induced degradation of IkappaB, which retains NF-kappaB in the cytoplasm. Therefore, THC attenuated nuclear translocation of NF-kappaB, a major pro-inflammatory transcription factor. Taken together, the present study for the first time demonstrates that THC exhibits anti-inflammatory activity through the suppression of NF-kappaB transcriptional activation in LPS-stimulated BV2 microglial cells.

Microglia and white matter injury in preterm infants / 国际儿科学杂志

Periventricular leukomalacia in premature infants is the most common brain injury.It often results in spastic cerebral palsy,cognitive and behavioral abnormalities and other sequelae.The microglia is the primary immune response cell in brain,and it plays an important role in balancing mechanism for the defense response and brain injury.The suppression of its activated reaction can reduce the cytotoxic effect and pathological damage in the early phase.

Expression of milk fat globule-epidermal growth factor 8 in microglial cells of neural retinal layer in developing rats / 中华实验眼科杂志

Background The retina microglia play a eliminating effect on apoptotie cells in the neural retinal layer of normal rats during postnatal development.Milk fat globule epidermal growth factor 8(MFG.E8)can combine specifically with phosphatidylinositol serine of the surface of apoptotie cells and enhance macrophage phagoeytosis of apoptotic cells.Objective Present study was to evaluate the localization and expression of MFG-E8 and its relevant cytokines in the neural retinal layer of normal rats during postnatal development Methods Normal royal college of surgeon(RCS)rats were divided into P0,P3,P7,Pi4,P30,P45 groups according to their postnatal days,and the 30-day-old RCS rats(2 rats)served as controls.Double stain of M FG.E8 and microglial cells marker(CD11b)was performed by immunofluorescence.Expressions of MFG-E8,integrin β5,CD11b and interleukin-6(IL-6)mRNA in the neural retina were analyzed by real-time quantitative reverse transcription polymerase chain reaction(RT-PCR).The utilization of animals complied with the Regulation for the Administration of Affair Concerning Experimental Animals by State and Science and Technology Commission.Results MFG-E8 and CD11b were positively co-expressed in retinal ganglion cell layer and external plexiform layer with the green fluorescence for FITC-labeled IgG and red fluorescence for cy3-labeled lgG respectively in normal adult rats.RT-PCR showed that the mRNA of MFG-E8,integrin 85,CD11b and IL-6 was detectable at P0 rats.The expression level of these eytokines began to rise fterward and reached peak value at P14 rats and then declined gradually,showing significant differences among different ages groups in various cytokines mRNA expression(all P<0.05).Conclusion MFG-E8 can be specifically expressed in the neural layer of retina microglia in RCS rat.

Anti-inflammatory Activity of 1-docosanoyl Cafferate Isolated from Rhus verniciflua in LPS-stimulated BV2 Microglial Cells

Although various derivatives of caffeic acid have been reported to possess a wide variety of biological activities such as protection of neuronal cells against excitotoxicity, the biological activity of 1-docosanoyl cafferate (DC) has not been examined. The objective of the present study was to evaluate the anti-inflammatory effects of DC, isolated from the stem bark of Rhus verniciflua, on lipopolysaccharide (LPS)-stimulated BV2 microglial cells. Pretreatment of cells with DC significantly attenuated LPS-induced NO production, and mRNA and protein expression of iNOS in a concentration-dependent manner. DC also significantly suppressed LPS-induced release of cytokines such as TNF-alpha and IL-1beta . Consistent with the decrease in cytokine release, DC dose-dependently and significantly attenuated LPS-induced mRNA expression of these cytokines. Furthermore, DC significantly suppressed LPS-induced degradation of IKB, which retains NF-kB in the cytoplasm. Therefore, nuclear translocation of NF-kB induced by LPS stimulation was significantly suppressed with DC pretreatment. Taken together, the present study suggests that DC exerts its anti-inflammatory activity through the suppression of NF-kB translocation to the nucleus.

Contact-Independent Cell Death of Human Microglial Cells due to Pathogenic Naegleria fowleri Trophozoites

Free-living Naegleria fowleri leads to a fatal infection known as primary amebic meningoencephalitis in humans. Previously, the target cell death could be induced by phagocytic activity of N. fowleri as a contact-dependent mechanism. However, in this study we investigated the target cell death under a non-contact system using a tissue-culture insert. The human microglial cells, U87MG cells, co-cultured with N. fowleri trophozoites for 30 min in a non-contact system showed morphological changes such as the cell membrane destruction and a reduction in the number. By fluorescence-activated cell sorter (FACS) analysis, U87MG cells co-cultured with N. fowleri trophozoites in a non-contact system showed a significant increasse of apoptotic cells (16%) in comparison with that of the control or N. fowleri lysate. When U87MG cells were co-cultured with N. fowleri trophozoites in a non-contact system for 30 min, 2 hr, and 4 hr, the cytotoxicity of amebae against target cells was 40.5, 44.2, and 45.6%, respectively. By contrast, the cytotoxicity of non-pathogenic N. gruberi trophozoites was 10.2, 12.4, and 13.2%, respectively. These results suggest that the molecules released from N. fowleri in a contact-independent manner as well as phagocytosis in a contact-dependent manner may induce the host cell death.

Comparison of Inhibitory Potency of Various Antioxidants on the Activation of BV2 Microglial Cell Lines Induced by LPS

Antioxidant properties have been proposed as a mechanism for the putative anti-inflammatory effects of phenolic compounds. To reveal the relationship between antioxidant activity and anti-inflammatory effects of various antioxidants, we measured 1, 1-diphenyl-2-picryhydrazyl (DPPH)-reducing activity and examined the inhibitory effects on LPS-induced inflammation-related gene expression in the BV2 microglial cell line. Lipopolysaccharide (LPS) (0.2microgram/ml) was used with or without antioxidants to treat cells, and the regulation of iNOS and cytokine gene expression was monitored using an RNase protection assay (RPA). Although, all tested antioxidants had similar DPPH-reducing activity and inhibited nitrite production, but the curcuminoid antioxidants (ferulic acid, caffeic acid, and curcumin) inhibited LPS-induced gene expression (iNOS, TNF-alpha, IL-1beta, IL-6, and IL-1 Ra) in a concentration-dependent manner. Other tested antioxidants did not exhibit the same effects; N-acetylcysteine (NAC) only began to suppress IL-1beta gene expression just below the concentration at which cytotoxicity occurred. Moreover, the antioxidant potency of curcuminoids appeared to have no correlation with anti-inflammatory potency. Only curcumin could inhibit LPS-induced microglial activation at a micromolar level. These data suggest that curcumin may be a safe antioxidant possessing anti-inflammatory activity.

Estrés oxidativo, péptido beta-amiloide y enfermedad de Alzheimer / Oxidative stress, beta-amyloide peptide and Alzheimer's disease

La enfermedad de Alzheimer es la causa más común de demencia en la población de edad avanzada. Una de las características histopatológicas de esta enfermedad es la formación de placas seniles, cuyo componente proteínico es el péptido β-amiloide (Aβ) en su forma insoluble. Este péptido se produce normalmente en forma monomérica soluble y circula en concentraciones bajas en el líquido cefalorraquídeo y sangre. En concentraciones fisiológicas actúa como factor neurotrófico y neuroprotector, sin embargo con el envejecimiento y sobre todo en la enfermedad de Alzheimer se acumula, forma fibrillas insolubles y causa neurotoxicidad. La toxicidad del Aβ se ha asociado a la generación de radicales libres que causan peroxidación de lípidos y oxidación de proteínas entre otros daños. Se ha planteado que el Aβ pueda reconocer a receptores específicos que median a su vez neurotoxicidad. Entre estos se encuentra el receptor scavenger o pepenador que se expresa en la microglia y es capaz de internalizar agregados de este péptido. Independientemente de la vía de entrada del péptido a la célula, éste genera un estado de estrés oxidativo que eventualmente desencadena la muerte celular. Estudios recientes desarrollados en nuestro laboratorio muestran que el proceso de traducción de proteínas que intervienen en el proceso de endocitosis mediada por un receptor puede ser afectado por una condición de estrés oxidativo. Este es el caso de la β-adaptina, proteína clave en la formación del pozo cubierto.

Alzheimer's disease, the leading cause of dementia in the elderly is characterized by the presence in the brain of senile plaques formed of insoluble fibrillar deposits of beta-amyloid peptide. This peptide is normally produced in a monomeric soluble form and it is present in low concentrations in the blood and spinal fluid. At physiological concentrations, this peptide is a neurotrophic and neuroprotector factor; nevertheless, with aging and particularly in Alzheimer's disease this peptide accumulates, favors the formation of insoluble fibrils and causes neurotoxicity. beta-Amyloid peptide toxicity has been associated with the generation of free radicals that in turn promote lipid peroxidation and protein oxidation. Through the recognition of specific receptors such as the scavenger receptor, the beta-amyloid peptide becomes internalized in the form of aggregates. Independently of the way the peptide enters the cell, it generates oxidative stress that eventually triggers a state of neurotoxicity and cell death. Recent studies in our laboratory have shown the effect caused by an extracellular oxidative stress upon the internalization of the scavenger receptor. We have also demonstrated that the process of protein translation of molecules implicated in the mechanism of endocytosis through the scavenger receptor, such as the case of beta-adaptin, is arrested in microglial cells treated with beta-amyloid.

Enhancement of Nitric Oxide Production by Corticotropin-releasing Hormone (CRH) in Murine Microglial Cells, BV2

BACKGROUND: Microglial cells, major immune effector cells in the central nervous system, become activated in neurodegenerative disorders. Activated microglial cells produce proinflammatory mediators such as nitric oxide (NO), tumor necrosis factor-alpha and interleukin-1beta(IL-1beta). These proinflammatory mediators have been shown to be significantly increased in the neurodegenerative disorders such as Alzhimer's disease and Pakinson's disease. It was known that one of the neurodegeneration source is stress and it is important to elucidate mechanisms of the stress response for understanding the stress-related disorders and developing improved treatments. Because one of the neuropeptide which plays a main role in regulating the stress response is corticotropin- releasing hormone (CRH), we analyzed the regulation of NO release by CRH in BV2 murine microglial cell as macrophage in the brain. METHODS: First, we tested the CRH receptor expression in the mRNA levels by RT-PCR. To test the regulation of NO release by CRH, cells were treated with CRH and then NO release was measured by Griess reagent assay. RESULTS: Our study demonstrated that CRH receptor 1 was expressed in BV2 murine microglial cells and CRH treatment enhanced NO production. Furthermore, additive effects of lipopolysaccaride (LPS) and CRH were confirmed in NO production time dependantly. CONCLUSION: Taken together, these data indicated that CRH is an important mediator to regulate NO release on microglial cells in the brain during stress.

Nitric Oxide Production in Mouse's Microglial Cells by Human Chorionic Gonadotropin / 대한산부인과학회잡지

Nitric oxide ( NO ) is now recognized as a mediator of several biological and immunological functions, but unlike classical neurotransmitters, NO simply diffuse of the postsynaptic cell and around affecting cells. Human chorionic gonadotropin ( hCG ), produced by placental trophoblasts may act as stimulator on NO synthesis in oocytes of mouse's ovary. How-ever, in the various organs or cells, the action of hCG on NO synthesis is unknown. We have examined that the effect of hCG on NO synthesis in microglial cells of murine's brain, using the Griess method. And this study was evident that hCG did not induce NO produc-tion without recombinant interferon gamma ( rIFN-gamma), whereas hCG ( 10~500 IU/ml ) with rIFN-gamma effectively produced NO in microglial cells of brain. As result, NO production in microglial cells increased most significantly in dose of 100 IU/ml of the hCG and the pro-duction of NO was dependent on the dose of hCG ( Table 1 and Fig. 1 ). And N(G)-monomethyl-L-arginine ( N(G)MMA ), competitive inhibitor of NO synthase, reduced the NO production by hCG stimulation with rIFN-gamma in microglial cells of murine. Conclusively, this study sugge-sted that hCG stimulate NO production at microglial cells in brain, which may be an important factor for mediating immune and neuroendocrinologic regulation in nervous system.

Influence of Mycoplasma Pneumoniae Infection on the Growth, Phagocytie Activities and Induction of Nitric Oxide Production of the Microglial Cells of Mice

In this study, the distribution and reisolation of Mycoplasma pneumoniae(Mp) were observed from the various tissues of BALB/c mice which were intraperitoneally pre-inoculated with Mp. In addition, the effect of Mp on the growth, phagocytic activities and nitric oxide production of microglial cells were also examined. The results were as follows; 1) Mp was reisolated from the various tissues such as lymph node, spleen, liver, kidney, brain and blood from one hour through 48 hours after intra-peritoneal inoculation of Mp in mice by the cultural method. Furthermore, it could also be confirmed from those tissues up to 72 hours by the indirect immunofluorescent antibody method. 2) There was no difference in the phagocytic activities between the control microglial cells and Mp stimulated microglial cells. 3) The growth of microglial cells in the medium was significantly increased by the stimulation with Mp compared with that of the control. 4) Nitric oxide production of mouse microglial cells was increased by the combined treatment if IFN-r and LPS or IFN-r and Mp or IFN-r, LPS and Mp, whereas, no increase was observed by either LPS or Mp alone. 5) Nitric oxide production of microglial cells primed with IFN-r was closely related with the dose of LPS and Mp in the dose dependent manner rather than that of the IFN-r. These results suggest that; i) Mp spreads to the various tissues of mice within one hour after intraperitoneal inoculation, ii) the growth of microglial cells increases by the infection of Mp, iii) microglial cells have phagocytic activities to C.albicans and iv) nitric oxide production of microglial cells was augmented by the infection of Mp. Increased nitric oxide production of microglial cells is regarded as an increase of the intracellular bactericidal activiteis of microglial cells. It is suggested, nonetheless, that the inflammatory response of the Mp infected tissues is augmented by the increase of nitric oxide.