TLR4-dependent adjuvant activity of Neisseria meningitidis lipid A

The adjuvant activity of Neisseria meningitidis serogroup B lipopoly(oligo)saccharide (LOS) from wild-type and genetically defined LOS mutants and unglycosylated meningococcal lipid A was assessed in C3H/HeN and C3H/HeJ mice. Meningococcal lipid A, a weak agonist for TLR4/MD-2 in human macrophages, was found to have adjuvant activity similar to that of wild-type and KDO(2)-lipid A LOS in C3H/HeN mice. All meningococcal LOS structures as adjuvants induced high titers of IgG1, IgG2a and IgG2b but very little IgG3 to OMP compared to no adjuvant PBS controls. In addition, induced OMP antibodies were shown to have high bactericidal activity against serogroup B meningococci. Purified LOS and lipid A structures failed to induce any adjuvant activity in C3H/HeJ mice indicating that meningococcal LOS as an adjuvant was TLR4-dependent. Unglycosylated meningococcal lipid A because of its weak agonist activity for human macrophages and retention of adjuvant activity may be a candidate for use in serogroup B meningococcal OMP and OMV vaccines and for use as an adjuvant in other vaccines.     

Phenotypic and functional maturation of dendritic cells induced by polysaccharide isolated from Paecilomyces cicadae

Paecilomyces cicadae Miquel Samson is the anamorph of Cordyceps cicadae Shing and is used in functional foods for the prevention and treatment of various diseases. In the present study, we examined the effects of P. cicadae polysaccharide (PCP) on dendritic cell (DC) maturation. Phenotypic maturation of DCs by PCP was confirmed by the elevated expressions of CD80, CD86, major histocompatibility complex (MHC)-I, and MHC-II molecules and functional maturation by increased expression of interleukin-12, interleukin-1β, and tumor necrosis factor-α, enhanced allogenic T cell stimulation, and decreased endocytosis. PCP induced the maturation of DCs from C3H/HeN and C57BL/6 mice but not from Toll-like receptor (tlr) 4?/? knockout mice and TLR4-mutated C3H/HeJ mice, which suggests that TLR4 is the membrane receptor for PCP. PCP increased the degradation of inhibitor of nuclear factor-κB (NF-κB) α/β, which enhanced the nuclear translocation of NF-κB p50/p65 and induced the phosphorylation of extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 mitogen-activated protein kinases, which are signaling molecules downstream of TLR4. These results indicate that PCP induces DC maturation through TLR4 signaling.     

ω-3多不饱和脂肪酸对脂多糖所致新生大鼠脑损伤Toll-NF-κB信号通路调节作用

目的 探讨ω-3 PUFAs在脂多糖(LPS)诱导的新生大鼠脑损伤中对TLR4、NF-κB信号通路的调节作用。方法 将96只SD新生大鼠按随机数字法分为对照组,ω-3组,ω-6组和LPS组。4组大鼠分别采用生理盐水或LPS腹腔注射后,立即注射生理盐水或不同的脂肪乳剂,使用时间分别为1天或5天,建立感染所致新生大鼠脑损伤模型;1天或5天后处死新生大鼠,检测TLR4 mRNA、NF-κB mRNA和对应蛋白水平的变化。结果 LPS注射后的第1天、第5天,新生大鼠TLR4 mRNA、NF-κB mRNA表达及其对应的蛋白水平均明显高于对照组(P均＜0.05)。1 d 时ω-3组TLR4 mRNA、NF-κB mRNA表达及其对应的蛋白水平均低于ω-6组和LPS组(P均＜0.05)。5 d时ω-3组TLR4 mRNA、NF-κB mRNA表达及其对应的蛋白水平均低于ω-6组和LPS组(P均＜0.05)。不论第1 d和第5 d,ω-3 PUFAs均可明显降低TLR4 mRNA、NF-κB mRNA表达及其对应的蛋白水平(P均＜0.05);而ω-6 PUFAs能提高TLR4 mRNA、NF-κB mRNA表达及对应的蛋白水平(P均＜0.05)。结论 ω-3 PUFAs 能下调 TLR4、NF-κB活性,在LPS所致的脑损伤中具有抗炎作用。     

二硫化碳对实验大鼠神经系统神经营养因子的影响

目的探讨二硫化碳染毒对大鼠神经生长因子(NGF)和脑源性神经营养因子(BDNF)蛋白表达的影响。方法健康雄性Wistar大鼠20只随机分成对照组和二硫化碳组,每组10只。二硫化碳组大鼠腹腔注射二硫化碳玉米油溶液,开始4周染毒剂量为150mg/kg,1次/d,6次/周,后4周染毒剂量为300mg/kg,1次/d,6次/周。对照组腹腔注射等剂量的玉米油溶液。染毒结束后对周围神经损伤体征进行评分,同时检测胫神经运动神经传导速度和腓肠神经感觉神经传导速度;并应用免疫组化方法检测NGF、BDNF在海马及坐骨神经中的蛋白表达情况。结果二硫化碳染毒8周后染毒组大鼠周围神经体征评分明显升高;与对照组比较,神经电生理测定结果显示二硫化碳染毒组胫神经运动神经和腓肠神经感觉神经传导速度明显减慢,潜伏期延长,波幅减低。染毒组大鼠海马及坐骨神经中NGF和BDNF表达较对照组显著降低。结论二硫化碳染毒导致的神经组织NGF、BDNF蛋白表达下降可能参与了其神经毒作用。     

Effects of olive polyphenols administration on nerve growth factor and brain-derived neurotrophic factor in the mouse brain

ObjectivePolyphenols are chemicals derived from plants known to possess antioxidant and anti-inflammatory properties. High intake of fruit and vegetables is believed to be beneficial to human health. Various studies have suggested that dietary polyphenols may protect against cancer and cardiometabolic and neurodegenerative diseases. Nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are neurotrophins that play key roles in brain cell development, growth, and survival. The aim of this study was to investigate whether or not administration of olive (Olea europaea L.) polyphenols could have an effect on NGF and BDNF content and the expression of their receptors, TrkA and TrkB, respectively, in the mouse brain.MethodsNGF and BDNF were measured by enzyme-linked immunosorbent assay. TrkA and TrkB were measured by Western blotting.ResultsWe found NGF and BDNF elevation in the hippocampus and olfactory bulbs and a decrease in the frontal cortex and striatum. These data were associated with potentiated expression of TrkA and TrkB in the hippocampus and olfactory bulbs but no differences between groups in the striatum and frontal cortex. Polyphenols did not affect some behavioral mouse parameters associated with stressing situations.ConclusionsAltogether, this study shows that olive polyphenols in the mouse may increase the levels of NGF and BDNF in crucial areas of the limbic system and olfactory bulbs, which play a key role in learning and memory processes and in the proliferation and migration of endogenous progenitor cells present in the rodent brain.     

Immunization with the Chlamydia trachomatis major outer membrane protein, using adjuvants developed for human vaccines, can induce partial protection in a mouse model against a genital challenge

To test several vaccines for Chlamydia trachomatis we vaccinated BALB/c and C3H/HeN female mice with a purified preparation of the native mouse pneumonitis (MoPn) major outer membrane protein (MOMP). The MOMP was formulated with anyone of three different adjuvants MF59, LT-K63 or LT-R72. As a negative control the animals were immunized with ovalbumin. Positive controls were inoculated intranasally (i.n.) with 10(4) inclusion-forming units (IFU) of C. trachomatis MoPn. High levels of Chlamydia-specific antibodies were detected in the serum and vaginal washes of the mice immunized with MOMP. Using a lymphoproliferative assay (LPA) a significant response was obtained in splenocytes from most of the groups of mice vaccinated with MOMP. Two weeks after the last immunization the mice were challenged in the left ovarian bursa with 10(5) IFU of C. trachomatis MoPn and vaginal cultures were collected for a period of 6 weeks. Overall, BALB/c and C3H/HeN mice immunized with MOMP showed a decrease in the severity and length of the infection but the difference with the controls was not statistically significant. Following mating the percentage of mice with bilateral fertility was not significantly different between mice vaccinated with MOMP and their respective ovalbumin-immunized controls. However, the C3H/HeN mice immunized with MOMP using MF59 or LTR72 as adjuvants had significantly more embryos per mouse than the control groups. In conclusion, mice immunized with native MOMP and adjuvants developed for human vaccines showed significant Chlamydia-specific immune response and a limited protection against infection and long-term sequelae.     

BDNF及高亲和力受体TrKB在胆红素神经损伤豚鼠皮层及海马表达变化的实验研究

目的:探讨新生豚鼠胆红素脑神经损伤元时脑源性神经营养因子(BDNF)及受体酪氨酸激酶B(TrKB)在大脑皮质及海马表达特点。方法:取生后2～5天内的豚鼠60只,随机分为3组,C组、T1组、T2组。C组为对照组;T1组:腹腔注射晶体胆红素100μg/g;T2组腹腔注射胆红素200μg/g,分别在4h、8h处死。做脑组织切片,电镜,光镜观察病理改变,并采用免疫组化和图像分析,观察不同时间点BDNF、TrKB在皮层及海马表达变化。结果:①胆红素脑神经元损伤模型成功建立。②T14h、T24h在早期皮层及海马是降低的。随时间延长,BDNF表达明显上升,T14h、T24h与C4h比较,P<0.05,T14h、T24h与T18h、T28h比较,P<0.05。③TrKB在皮层和海马表达是随时间延长和损伤加重表达增加,T14h、T18h、T24h、T28h与C4h、C8h比较,P<0.05。结论:胆红素脑神经元损伤中,皮层和海马BDNF、TrKB升高可能减轻神经元损伤,在抑制神经元凋亡和神经元修复中发挥重要作用,有利神经元修复和再生,可能是胆红素脑神经元损伤时机体的自我保护机制之一。     

High-dose dietary supplementation of vitamin A induces brain-derived neurotrophic factor and nerve growth factor production in mice with simultaneous deficiency of vitamin A and zinc

Marginal vitamin A and zinc (Zn) deficiency often co-exist in many populations. Vitamin A plays a trophic role in brain and is important for its development. We investigated effects of dietary supplementation of vitamin A on brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) production in mice depleted for vitamin A and Zn. After 3 months' feeding with a low vitamin A and Zn (LVA-LZ) diet, mice were divided into two groups and replenished with either normal or high vitamin A with low Zn diet for an additional 2 months. Levels of BDNF and NGF were measured from extracts of hippocampus, cortex and cerebellum at the end of the third and fifth months. The LVA-LZ group tended to show decreased amounts of the BDNF and NGF, while animals supplemented with high vitamin A along with Zn deficiency had high BDNF and NGF concentrations. From these results, we conclude that vitamin A may increase BDNF and NGF levels.     

邻苯二甲酸二异丁酯对小鼠海马cAMP/PKA-CREB信号转导通路的影响

目的观察邻苯二甲酸二异丁酯(DiBP)对小鼠海马cAMP/PKA-CREB信号通路的影响。方法 30只雄性昆明小鼠适应性喂养3天后,按体重随机分为一个对照组和4个实验组(Ⅰ、Ⅱ、Ⅲ、Ⅳ)。对照组小鼠灌胃给予玉米油溶剂;实验组小鼠灌胃给予含DiBP的玉米油溶液,剂量分别为:50、250、500、1000mg/kg BW。实验周期8周,普通饲料喂饲,自由饮水。实验结束后冰浴条件下取海马,采用酶联免疫法测定环磷酸腺苷(cAMP)含量;蛋白印迹法测定磷酸化蛋白激酶A(P-PKA C)、磷酸化环磷腺苷反应元件结合蛋白(P-CREB)表达水平;实时荧光定量PCR法测定CREB、脑源性神经营养因子(BDNF)、c-fos和c-jun的mRNA表达水平。结果第Ⅳ组海马组织中cAMP含量和P-PKA C蛋白表达水平明显低于对照组,差异具有统计学意义(P<0.05);各剂量组P-CREB蛋白表达量均低于对照组,其中第Ⅱ组与对照组相比差异有统计学意义(P<0.05),同时CREB、BDNF、c-fos和c-jun的mRNA表达水平与对照组相比都呈不同程度的下调。结论小鼠海马cAMP/PKA-CREB信号通路中多种信号分子的表达异常,可能是DiBP导致小鼠认知损伤的重要机制之一。     

Lactobacillus crispatus KT-11 enhances intestinal immune functions in C3H/HeN mice

We investigated the effect of Lactobacillus crispatus KT-11 (KT-11) on intestinal immune systems in C3H/HeN mice. The level of intestinal total immunoglobulin (Ig) A was significantly higher in mice given KT-11 than in mice not given KT-11. Gene expression relating to antibody production and innate immune response increased more than 2-fold in the former compared with the later. Moreover, the number of IL-6(+)CD11b(+) cells was significantly higher in Peyer's patch cells cultured with KT-11 than in those cultured without KT-11, although the number of CD4(+) cells and the cell ratio of CD4(+)/CD8(+) were remarkably lower in the culture with KT-11. These results indicate that KT-11 enhances intestinal IgA production and innate immune response in C3H/HeN mice.     

High-dose dietary supplementation of vitamin A induces brain-derived neurotrophic factor and nerve growth factor production in mice with simultaneous deficiency of vitamin A and zinc

Abstract

Marginal vitamin A and zinc (Zn) deficiency often co-exist in many populations. Vitamin A plays a trophic role in brain and is important for its development. We investigated effects of dietary supplementation of vitamin A on brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) production in mice depleted for vitamin A and Zn. After 3 months' feeding with a low vitamin A and Zn (LVA-LZ) diet, mice were divided into two groups and replenished with either normal or high vitamin A with low Zn diet for an additional 2 months. Levels of BDNF and NGF were measured from extracts of hippocampus, cortex and cerebellum at the end of the third and fifth months. The LVA-LZ group tended to show decreased amounts of the BDNF and NGF, while animals supplemented with high vitamin A along with Zn deficiency had high BDNF and NGF concentrations. From these results, we conclude that vitamin A may increase BDNF and NGF levels.     

Entamoeba histolytica in inbred mice: evidence of liver involvement

Following observations that certain mouse strains appeared to be susceptible to liver invasion by Entamoeba histolytica, mouse strains CBA/HN, CBA/HN, nu/nu, CBA/HN nu/+, CBA/HN Dh/+, CBA/HN +/+, C3H/HeJ Lps/Lps and AB/H, were infected intra-intestinally with strain SAW 408 (zymodeme II) of the amoeba. A number of mice from each group died during the first 14 days with acute lesions. The remainder were killed at 4 and 6 months after infection. CBA/HN, CBA/HN Dh/+, and CBA/HN +/+ mice all had amoeba-infected livers, although only 2 mice had extensive, typical liver lesions.     

Radiation-induced apoptosis in peritoneal resident macrophages of C3H mice

Gamma ray-radiation induced significant apoptosis in peritoneal resident macrophages (PRMs) of C3H/HeJ (C3H) mice, but not in other strains of mice. To investigate the role of DNA damage in the apoptosis, DNA damage was quantified in PRMs by use of the alkaline single-cell gel electrophoresis (Comet) assay. No significant difference was found between C3H and C57Black/6 mice in either radiation-induced DNA damage or repair. Radiation induced apoptosis at the same levels in PRMs of p53 knockout mice and atm knockout mice as those of wild-type C3H mice; however radiation-induced apoptosis was significantly less extensive in the thymocytes of these mutant mice than in those of wild-type mice. Apoptosis was also induced at the same level by an irradiation in PRMs of C3H scid mice as in those of wild-type C3H mice. Therefore it was suggested that radiation-induced DNA damage and TP53, ATM, or DNA-PK-mediated cellular responses occurring downstream thereof were not involved in the radiation-induced apoptotic cell death in C3H mouse PRMs.     

围产期PFOS暴露对幼鼠海马BDNF/TrkB/CREB信号通路影响

目的 探讨观察母孕鼠围产期全氟辛烷磺酸盐(perfluorooctane sulphonate,PFOS)暴露对幼鼠海马组织BDNF/TrkB/CREB信号通路关键基因表达的影响。 方法 20只昆明种雌性小鼠随机分为对照组和低、中、高剂量组,从孕鼠怀孕第2 d开始(gestation day2, GD2)到幼鼠出生后21 d(postnatal day21,PND21)分别给予低、中、高剂量组孕鼠PFOS剂量为 0.1、1.0、5.0 mg/(kg·bw)灌胃染毒,对照组给予等体积的0.05% Tween-20水溶液。灌胃量为0.1 ml/10 (g·bw)。PND 21 d处死幼鼠,收集脑组织,分离海马及皮层。HE染色观察脑组织常规病理改变,实时荧光定量PCR(Quantitative Real-time PCR,QPCR)检测海马组织中BDNF、TrkB、CREB、Syn1及Syp的mRNA表达水平。 结果 与对照组相比较,低、中、高三个剂量组对幼鼠的死亡率和体质量的影响差异无统计学意义(P>0.05)。但是在高剂量组幼鼠海马组织出现空泡,海马BDNF、TrkB、CREB mRNA水平显著降低,分别由对照组的(0.98±0.11)、(1.03±0.09)、(1.08±0.12)下降到(0.22±0.21)、(0.71±0.14)、(0.37±0.26),并在中、高剂量组引起了幼鼠突触相关蛋白Syn1和Spy的mRNA水平显著降低,分别由对照组的(1.10±0.09)、(0.97±0.08)下降到中剂量的(0.41±0.23)、(0.71±0.17)和高剂量的(0.39±0.19)、(0.63±0.19),差异均有统计学意义(P<0.05)。 结论 PFOS损伤海马BDNF/TrkB/CREB信号通路可能是PFOS神经发育毒性之一。     

Adverse cardiovascular effects with acute particulate matter and ozone exposures: interstrain variation in mice

OBJECTIVES: Increased ambient particulate matter (PM) levels are associated with cardiovascular morbidity and mortality, as shown by numerous epidemiology studies. Few studies have investigated the role of copollutants, such as ozone, in this association. Furthermore, the mechanisms by which PM affects cardiac function remain uncertain. We hypothesized that PM and O(3) induce adverse cardiovascular effects in mice and that these effects are strain dependent. STUDY DESIGN: After implanting radiotelemeters to measure heart rate (HR) and HR variability (HRV) parameters, we exposed C57Bl/6J (B6), C3H/HeJ (HeJ), and C3H/HeOuJ (OuJ) inbred mouse strains to three different daily exposures of filtered air (FA), carbon black particles (CB), or O(3) and CB sequentially [O(3)CB; for CB, 536 +/- 24 microg/m(3); for O(3), 584 +/- 35 ppb (mean +/- SE)]. RESULTS: We observed significant changes in HR and HRV in all strains due to O(3)CB exposure, but not due to sequential FA and CB exposure (FACB). The data suggest that primarily acute HR and HRV effects occur during O(3)CB exposure, especially in HeJ and OuJ mice. For example, HeJ and OuJ mice demonstrated dramatic increases in HRV parameters associated with marked brady-cardia during O(3)CB exposure. In contrast, depressed HR responses occurred in B6 mice without detectable changes in HRV parameters. CONCLUSIONS: These findings demonstrate that important interstrain differences exist with respect to PM- and O(3)-induced cardiac effects. This interstrain variation suggests that genetic factors may modulate HR regulation in response to and recuperation from acute copollutant exposures.     

In vivo gene transfer in mouse preimplantation embryos after intraoviductal injection of plasmid DNA and subsequent in vivo electroporation

The aim of this study was to investigate whether direct injection of nonviral DNA into the oviductal lumen and subsequent in vivo electroporation leads to in vivo gene transfer in mouse preimplantation embryos present within an oviduct, as an alternative to the pre-existing pronuclear microinjection-based transgenesis. With this technique, effects of expression of the gene of interest (GOI) on mouse preimplantation development can be monitored with relative ease. Superovulated 4-week-old B6C3F1 female mice (hybrids between C57BL/6N and C3H/HeN) were mated with adult B6C3F1 male mice. Two days later, females that had been identified as pregnant, based on the presence of copulation plugs, were injected with 1?μl of a solution containing an enhanced green fluorescent protein (EGFP) expression plasmid (0.5?μg) and 0.05% trypan blue. The entire oviduct was then electroporated using tweezer-type electrodes with 8 square-wave pulses of 50 V each with 50-ms duration. The next day, the 8-cell stage embryos were collected, and their number, morphology, and EGFP-derived fluorescence were recorded. Of the 12 oviducts (6 females used) examined, 3 contained fluorescent 8-cell stage embryos (33%, 19/58 tested), but the intensity of fluorescence varied among the embryos. In total, 10% (19/192 tested) of the embryos were fluorescent and the fluorescence was maintained in these embryos after 1 day of culture. However, the fluorescence disappeared in the late gestational stage fetuses, and the transgenes could not be detected. Our results indicate that it is possible to transfect in vivo preimplantation embryos, although the success rate appears to be relatively low and gene expression is transient. This technology may provide a new method for manipulating preimplantation embryos in vivo, by using, for example, Cre-mediated conditional DNA recombination.     

Role of acute ethanol exposure and TLR4 in early events of sepsis in a mouse model

Sepsis is a major cause of death worldwide. The associated risks and mortality are known to significantly increase on exposure to alcohol (chronic or acute). The underlying mechanisms of the association of acute ethanol ingestion and poor prognosis of sepsis are largely unknown. The study described here was designed to determine in detail the role of ethanol and TLR4 in the pathogenesis of the sepsis syndrome. The effects of acute ethanol exposure and TLR4 on bacterial clearance, spleen cell numbers, peritoneal macrophage numbers, and cytokine production were evaluated using wild-type and TLR4 hyporesponsive mice treated with ethanol and then challenged with a nonpathogenic strain of Escherichia coli. Ethanol-treated mice exhibited a decreased clearance of bacteria and produced lesser amounts of most pro-inflammatory cytokines in both strains of mice at 2 h after challenge. Neither ethanol treatment nor a hyporesponsive TLR4 had significant effects on the cell numbers in the peritoneal cavity and spleen 2 h postinfection. The suppressive effect of acute ethanol exposure on cytokine and chemokine production was more pronounced in the wild-type mice, but the untreated hyporesponsive mice produced less of most cytokines than untreated wild-type mice. The major conclusion of this study is that acute ethanol exposure suppresses pro-inflammatory cytokine production and that a hyporesponsive TLR4 (in C3H/HeJ mice) decreases pro-inflammatory cytokine levels, but the cytokines and other mediators induced through other receptors are sufficient to ultimately clear the infection but not enough to induce lethal septic shock. In addition, results reported here demonstrate previously unknown effects of acute ethanol exposure on leukemia inhibitory factor and eotaxin, and provide the first evidence that interleukin (IL)-9 is induced through TLR4 in vivo.     

Buckwheat Flour and Its Starch Prevent Age-Related Cognitive Decline by Increasing Hippocampal BDNF Production in Senescence-Accelerated Mouse Prone 8 Mice

Buckwheat is an important pseudo-cereal crop worldwide. This study investigated whether long-term administration of buckwheat can suppress age-related cognitive decline in senescence-accelerated mouse prone 8 (SAMP8) mice. For 26 weeks, 18-week-old male SAMP8 mice were fed a standard diet containing 5% (w/w) buckwheat, Tartary buckwheat, wheat, or rice flour. In the Barnes maze and passive avoidance tests, mice fed buckwheat whole flour (BWF) showed improved cognitive performance compared to those fed a control diet, while no improvement was noticed in case of the other diets. Analysis of the gut microbiota showed that BWF and buckwheat outer flour administration increased the abundance of Lactococcus and Ruminiclostridium, respectively, at the genus level. The expression levels of brain-derived neurotrophic factor (BDNF), postsynaptic Arc and PSD95, and the mature neuronal marker NeuN in the hippocampus were increased after BWF administration, which was induced by the activation of the ERK/CREB signaling pathway and histone H3 acetylation. A similar increase in cognitive performance-related hippocampal BDNF expression in SAMP8 mice was observed after the oral administration of starch prepared from BWF. Therefore, the long-term administration of BWF suppresses cognitive decline by increasing hippocampal BDNF production in SAMP8 mice.