The Effects of Growth Factor on Testicular Germ Cell Apoptosis in the Stallion

Apoptosis is necessary for both initiation and control of spermatogenesis; however, an increase in apoptosis can lead to subfertility/infertility in stallions, causing substantial financial loss in the equine industry. The ability of stem cell factor (SCF), leukemia-inhibiting factor (LIF), granulocyte–macrophage colony-stimulating factor (GM-CSF), and estradiol (E₂), alone or in combination, to prevent apoptosis of germ cells in short-term equine testicular cultures was examined. Testicular tissue was sectioned into approximately 2-mm cubes and placed in media-filled culture chambers. Concentrations of SCF (100 ng/mL), LIF (10 ng/mL), GM-CSF (5 ng/mL), and E₂ (10⁻⁹ mol/l) were added alone or in combination to each well. After 6 hours in culture, the tissue was fixed and immunohistochemically (terminal deoxynucleotidyl transferase-mediated nick-end labeling; TUNEL) stained for apoptosis detection. Apoptotic cells per 100 Sertoli cell nuclei within seminiferous tubules were counted until the 500^th Sertoli cell nuclei was reached. This counting procedure was used for each slide. An analysis of variance (ANOVA) with a Tukey's test was used to compare apoptotic rates. In comparison with the control, GM-CSF alone lowered apoptosis by 34.77%. GM-CSF–treated tissue combined with SCF and LIF as well as GM-CSF combined with SCF, LIF, and E₂ reduced apoptosis when compared with the control (37.45% and 44.40%, respectively) or other treatment combinations. GM-CSF alone reduced apoptosis; results suggest possible synergy for the combinations of SCF and LIF with GM-CSF and for E₂ with SCF, LIF, and GM-CSF.     

猪繁殖与呼吸综合征病毒感染对猪肺泡巨噬细胞外源性抗原加工递呈相关分子和共刺激分子的影响

采用定量竞争PCR技术对猪繁殖与呼吸综合征病毒（PRRSV）感染后不同时相猪肺泡巨噬细胞（PAM）中外源性抗原加工递呈相关分子SLA-DR、Ii链和SLA-DM分子及共刺激分子CD40、CD80、CD86的mRNA转录动态进行了定量检测。结果表明,PRRSV感染后PAM的SLA-DR mRNA转录水平在3、7和14 d下调,低于对照组;感染后3 d,Ii链、SLA-DM和CD40的mRNA转录水平极显著下调（P〈0.01）;CD80和CD86mRNA转录水平也在感染后3 d明显下调（P〈0.05）。用流式细胞术检测PAM表面的SLA-DR抗原的结果表明,在感染后3 d短暂上升,7 d之后一直低于对照组。由此说明,PRRSV感染早期对猪肺泡巨噬细胞的外源性抗原加工和递呈功能产生显著影响,导致其外源性抗原递呈功能下降,进而影响机体的体液免疫应答。     

Interleukin 1, a mediator of immunoadjuvant peptidoglycans

Bacterial peptidoglycans and the synthetic analog muramyl dipeptide possess various immunomodulating properties (adjuvant effect, increase of resistance to infectious agents and to tumor growth). They are able to induce B cell activation and to stimulate macrophages to produce monokines such as Interleukin 1 (IL 1). IL 1 plays an essential role in immune response. It promotes thymocytes maturation and Interleukin 2 secretion by antigen sensitive T cells, which in turn triggers regulatory T cells. Moreover, it is involved in the proliferation and differentiation of B cells.

There is a correlation between the immunoenhancing effect of PG of a definite structure and their ability to induce IL 1 secretion. Non-adjuvant PG were inactive. This suggests that one of the major mechanisms of action of adjuvant PG could be the stimulation of IL 1 synthesis.     

草鱼巨噬细胞的集结及相关的组织变化

给草鱼腹腔注射细微炭粒（墨汁）后不同时间取样，比较观察了草鱼胸腺、肝、脾、头肾和肾组织中巨噬细胞集结（ｍ ａｃｒｏｐｈａｇｅ ａｇｇｒｅｇａｔｉｏｎ ， Ｍ Ａ）状况及相应组织的变化。结果表明，在头肾和脾组织中， Ｍ Ａ 的数量和体积均较肝和肾中的多而大，胸腺中所产生的 Ｍ Ａ 数量较肝和肾中的少，体积也小；注射墨汁后 １～３ 周内，在胸腺、头肾、肝和脾组织中， Ｍ Ａ 的数量和体积均有不同程度的增加，但在注射墨汁后第 ４ 周的肾组织中， Ｍ Ａ 的体积缩小而且密度降低，同时肾组织结构有逐渐恢复的趋势，可能是由于 Ｍ Ａ 分解后通过肾小球和肾小管排泄到膀胱或体外的缘故。对草鱼 Ｍ Ａ 的组织分布、排除等分析认为， Ｍ Ａ 可作为草鱼非特异细胞防御作用的解剖学和细胞学的一种生物标记。     

STAT6介导的巨噬细胞极化对布鲁氏菌胞内存活的影响

旨在探讨STAT6介导的巨噬细胞极化对布鲁氏菌胞内生存的影响。本研究采用布鲁氏菌光滑株S2308（S2308）和粗糙型疫苗株RB51（RB51）侵染巨噬细胞。利用qRT-PCR检测M1型巨噬细胞标志因子p65、NOS2和IL-1β,M2型巨噬细胞标志因子STAT6、ARG1、IL-10的mRNA表达水平;流式细胞术检测M1型标记分子CD86和M2型标记分子CD206的表达;Western blot检测p-STAT6蛋白及抑制剂AS对蛋白的抑制作用;ELISA检测M1型细胞因子TNF-α、IL-12和M2型细胞因子IL-4、IL-10的表达量;最后对胞内菌落进行CFU计数。qRT-PCR结果显示,在感染8、12 h时可显著诱导M1型因子mRNA转录表达,72 h时低表达,而M2型因子在72 h时高表达;流式细胞术结果显示,S2308感染12 h可显著诱导CD86的表达,感染72 h可显著诱导CD206的表达,但RB51对二者无影响;Western blot结果显示,S2308菌株在感染72 h时激活STAT6信号通路,而RB51几乎不激活该通路,抑制剂AS在2 μmol·L^-1浓度时抑制效果最佳;ELISA结果显示,AS抑制剂可显著抑制IL-4、IL-10的释放,并促进TNF-α、IL-12的释放;CFU计数结果显示,S2308组的胞内菌呈先降低后显著上升趋势,加入AS抑制剂后可显著抑制布鲁氏菌胞内复制。布鲁氏菌S2308在感染后期能够通过STAT6诱导M1型巨噬细胞向M2型转化,并促进Th2型细胞因子的释放,从而有利于布鲁氏菌的胞内生存。而RB51几乎不激活该通路,不影响胞内生存。     

甘草多糖对鼠伤寒沙门菌诱导的巨噬细胞氧化-抗氧化平衡紊乱的调解

为了探讨甘草多糖对鼠伤寒沙门菌诱发巨噬细胞氧化-抗氧化平衡紊乱的调节作用。将0、1、20和100 mg/L甘草多糖4个质量浓度组与小鼠腹腔巨噬细胞共孵育,同时每组按1∶1加入鼠伤寒沙门菌SL1344;利用ELISA试剂盒检测各组丙二醛(malondialdehyde,MDA),活性氧(reactive oxygen species,ROS),一氧化氮(nitricoxide,NO),诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS),谷胱甘肽过氧化物酶(glutathione peroxidase,GSHPX),超氧化物歧化酶(superoxide dismutase,SOD)和总抗氧化能力(total antioxidant capacity,T-AOC)变化情况;利用荧光显微镜和荧光酶标仪观察甘草多糖对鼠伤寒沙门菌引起巨噬细胞产生ROS的影响;利用乳酸脱氢酶(lactate dehydrogenase,LDH)细胞毒性试剂盒检测巨噬细胞上清LDH水平。结果表明,鼠伤寒沙门菌可诱导小鼠腹腔巨噬细胞发生氧化损伤,导致NO(4.65±1.07)μmol/L、iNOS(15.46±0.92)U/mL、MDA(3.46±0.39)nmol/L和ROS(3.69±0.31)U/mL氧化水平以及LDH水平显著升高,而GSH-PX(15.58±1.35)nmol/L、SOD(13.32±0.92)NU/mL和T-AOC(8.36±0.60)U/mL抗氧化水平显著降低。荧光显微镜和荧光酶标仪检测显示,鼠伤寒沙门菌感染小鼠腹腔巨噬细胞ROS水平显著升高。甘草多糖呈剂量依赖明显降低细胞氧化水平和增强抗氧化水平。甘草多糖作为重要的免疫调节剂,能够调节鼠伤寒沙门菌引起的巨噬细胞氧化损伤,维持细胞氧化-抗氧化平衡。     

Characterization of an intravenous lipopolysaccharide inflammation model in calves with respect to the acute-phase response

Our objective was to develop a lipopolysaccharide (LPS) inflammation model in calves to evaluate the acute-phase response with respect to the release of pro-inflammatory cytokines and acute-phase proteins, fever development and sickness behaviour. Fourteen 4-week-old male Holstein Friesian calves were included and randomly assigned to a negative control group (n = 3) and an LPS-challenged group (n = 11). The latter received an intravenous bolus injection of 0.5 μg of LPS/kg body weight. Blood collection and clinical scoring were performed at 0, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 5, 6, 8, 12, 18, 24, 28, 32, 48, 54 and 72 h post LPS administration (p.a.). In the LPS group, the following clinical signs were observed successively: tachypnoea (on average 18 min p.a.), decubitus (29 min p.a.), general depression (1.75 h p.a.), fever (5 h p.a.) and tachycardia (5 h p.a.). Subsequent to the recovery from respiratory distress, general depression was prominent, which deteriorated when fever increased. One animal did not survive LPS administration, whereas the other animals recovered on average within 6.1 h p.a. Moreover, the challenge significantly increased plasma concentrations of tumour necrosis factor-α, interleukin 6, serum amyloid A and haptoglobin, with peaking levels at 1, 3.5, 24 and 18 h p.a., respectively. The present LPS model was practical and reproducible, caused obvious clinical signs related to endotoxemia and a marked change in the studied inflammatory mediators, making it a suitable model to study the immunomodulatory properties of drugs in future research.