The endogenous inflammatory reflex inhibits the inflammatory response to different immune challenges in mice

The splanchnic anti-inflammatory pathway, the efferent arm of the endogenous inflammatory reflex, has been shown to suppress the acute inflammatory response of rats to systemic lipopolysaccharide (LPS). Here we show for the first time that this applies also to mice, and that the reflex may be engaged by a range of inflammatory stimuli. Experiments were performed on mice under deep anaesthesia. Half the animals were subjected to bilateral section of the splanchnic sympathetic nerves, to disconnect the splanchnic anti-inflammatory pathway, while the remainder underwent a sham operation. Mice were then challenged intravenously with one of three inflammatory stimuli: the toll-like receptor (TLR)-4 agonist, LPS (60 µg/kg), the TLR-3 agonist Polyinosinic:polycytidylic acid (Poly I:C, 1 mg/kg) or the TLR-2 and -6 agonist dipalmitoyl-S-glyceryl cysteine (Pam2cys, 34 µg/kg). Ninety minutes later, blood was sampled by cardiac puncture for serum cytokine analysis. The splanchnic anti-inflammatory reflex action was assessed by comparing cytokine levels between animals with cut versus those with intact splanchnic nerves. A consistent pattern emerged: Tumor necrosis factor (TNF) levels in response to all three challenges were raised by prior splanchnic nerve section, while levels of the anti-inflammatory cytokine interleukin 10 (IL-10) were reduced. The raised TNF:IL-10 ratio after splanchnic nerve section indicates an enhanced inflammatory state when the reflex is disabled. These findings show for the first time that the inflammatory reflex drives a coordinated anti-inflammatory action also in mice, and demonstrate that its anti-inflammatory action is engaged, in similar fashion, by inflammatory stimuli mimicking a range of bacterial and viral infections.     

Redox active or thiol reactive? Optimization of rapid screens to identify less evident nuisance compounds

Compounds that exhibit assay interference or undesirable mechanisms of bioactivity are routinely encountered in assays at various stages of drug discovery. We observed that assays for the investigation of thiol-reactive and redox-active compounds have not been collected in a comprehensive review. Here, we review these assays and subject them to experimental optimization to improve their reliability. We demonstrate the usefulness of our assay cascade by assaying a library of bioactive compounds, chemical probes, and a set of approved drugs. These high-throughput assays should complement the array of wet-lab and in silico assays during the initial stages of hit discovery campaigns to pursue only hit compounds with tractable mechanisms of action.     

穿山龙提取物抗急性痛风性关节炎的尿液代谢组学分析

目的:对急性痛风性关节炎大鼠给予穿山龙提取物后的尿液代谢组学进行研究,寻找相关的潜在生物标志物及相关代谢通路。方法:采用尿酸钠(MSU)诱导的急性痛风性关节炎大鼠模型,将SD大鼠40只随机分为空白组、穿山龙提取物组、模型组、穿山龙提取物干预组,每组10只。给药组灌胃给予穿山龙提取物,给药剂量0. 48 g·kg~(-1),每天1次,连续5 d,于末次给药后,收集大鼠尿液,运用UPLC-Q-TOF/MS结合模式识别方法分析,采用正、负离子扫描模式下电喷雾离子源,数据采集范围m/z 100～1 500,采用全扫描方式。结果:鉴别出了12个共同的潜在生物标志物,分别为肌氨酸,二甲基甘氨酸,脱氧胞苷,尿酸,5-HT,L-胱硫醚,4-吡哆酸,脱氧尿苷,褪黑激素,5-甲氧基色胺,富马酸和胞苷。与空白组比较,穿山龙提取物组中这12个潜在生物标志物均明显下调;与模型组比较,在穿山龙提取物干预组的潜在生物标志物中,有10个上调,2个下调,穿山龙提取物对肌氨酸,尿酸,L-胱硫醚,4-吡哆酸,脱氧尿苷,5-甲氧基色胺,胞苷,二甲基甘氨酸,褪黑激素,富马酸这10个标志物均表现出了纠正异常表达的趋势;与急性痛风性关节炎相关性最强的代谢通路为半胱氨酸和甲硫氨酸代谢、色氨酸代谢。结论:穿山龙提取物可能是通过促进半胱氨酸和甲硫氨酸代谢中胱硫醚向半胱氨酸的转化水平,上调色氨酸代谢中褪黑激素,实现对痛风性关节炎的防治作用。     

防己茯苓汤对急性肾损伤患者肾组织蛋白表达的影响

目的:观察防己茯苓汤对急性肾损伤患者肾组织蛋白表达的影响。方法:将150例急性肾损伤患者随机分为治疗组和对照组,每组75例,对照组患者均予基础治疗,包括:饮食控制、营养支持,血压及血糖管理,维持水、电解质及酸碱平衡,祛除诱因、对原发病给予治疗,停用可疑致肾损伤药物。治疗组在对照组治疗基础上加服防己茯苓汤治疗,方药组成:防己9 g,黄芪9 g,茯苓18 g,桂枝9 g,甘草6 g,每日1剂。治疗28 d后采集血液及尿液标本检测血利钠肽(natriuretic peptide,ANP)和尿β2-微球蛋白,用生化仪测定血肌酐(serum creatinine,Scr)、血清半胱氨酸蛋白酶抑制剂C(serum cystatin C,Cys C)的变化。结果:两组患者ANP、尿β2-微球蛋白、Cys C比较,差异均有统计学意义(P0.05);两组患者Scr比较,差异无统计学意义(P0.05)。结论:防己茯苓汤对急性肾损伤疗效显著,且能改善患者临床检验指标。     

N-乙酰半胱氨酸治疗COPD合并肺结核临床观察

目的：评价N-乙酰半胱氨酸（NAC）对慢性阻塞性肺疾病（COPD）合并肺结核患者肺功能、细胞免疫功能和生活质量的影响。方法：93例患者随机分为对照组47例和观察组46例，对照组予常规治疗，观察组加服NAC，6个月为一个疗程。比较抗结核作用、肺功能、细胞免疫功能和生活质量，记录不良反应。结果：治疗后，与对照组相比，观察组肺功能指标FEV₁/FVC和PEF，CD4+T、CD4+Foxp3+调节性T细胞亚群比例明显升高，CD4+/CD8+和SGRQ问卷中症状、活动、影响维度评分明显降低，组间均有显著性差异（P<0.05）。其余指标组间无差异。结论：NAC可明显改善COPD合并肺结核患者的肺功能、细胞免疫功能和生活质量，值得临床推广。     

药食心葆汤对心血瘀阻证冠心病患者血清同型半胱氨酸及凝血功能的影响

目的:观察药食心葆汤联合西药治疗冠心病的临床疗效及其对血清同型半胱氨酸(homocysteine,HCY)水平凝血功能的影响,并探讨其作用机制。方法:选取2015年1月至2016年1月于郑州市中医院心内科入院治疗并确诊为心血瘀阻证的冠心病患者200例,按照随机数字表法分为对照组和观察组,对照组102例,观察组98例,对照组患者采用常规西药治疗,观察组在此基础上联合药食心葆汤治疗,药物干预治疗8周,比较两组患者血清同型半胱氨酸(HCY),凝血功能[凝血酶原时间(prothrombin time,PT),部分凝血活酶时间(activated partial thromboplastin time,APTT),纤维蛋白原(fibrinogen,FIB)]等指标。结果:治疗后观察组患者凝血功能、血清HCY水平改善程度优于对照组(P0.05);观察组可明显降低CHD患者血清HCY水平(P0.05),调节患者的凝血功能(P0.05),组间比较显示两组有明显差别,特别是在改善凝血功能方面,观察组明显优于对照组(P0.05),对照组治疗前后各项指标没有统计学差异。结论:药食心葆汤联合西药治疗冠心病的临床疗效确切,能有效降低CHD患者HCY水平、调节凝血功能,值得临床推广使用。     

Cysteine protease inhibitory activity and levels of salivary cystatins in whole saliva of periodontally diseased patients

The 3 human salivary cystatins S, SA and SN are multifunctional proteins that possess a cysteine protease inhibitory property, but their ability to act as such is very different (SN > SA > S). One form, S, also appears to possess antibacterial properties towards the bacterium Porphyromonas gingivalis, often associated with periodontal diseases. In this study we measured the total cystatin inhibitory activity and the levels of each salivary cystatin in the whole saliva of 8 periodontally diseased patients and 2 groups of control subjects (n = 6 and n = 10). The total cystatin inhibitory activity and the total salivary cystatin concentration in the periodontally diseased patients were found to be lower than the controls (p < or = 0.005). The concentration of S was depleted to levels that would not allow it to be an effective antibacterial agent, and the concentration of SA, although depleted in some cases, was still present at sufficient levels to allow it to act as an effective physiological inhibitor of cathepsin L. The concentration of cystatin SN was also depleted in the periodontally diseased patients, but was still present in sufficient quantities to act as an effective physiological cysteine protease inhibitor of cathepsins H and L. In comparison, the concentration of all 3 salivary cystatins in the control subjects were sufficient to enable these proteins to be both effective physiological cysteine protease inhibitors and antibacterial agents.     

Immunization of Macaca fascicularis against experimental periodontitis using a vaccine containing cysteine proteases purified from Porphyromonas gingivalis

INTRODUCTION: Periodontitis is a common infectious disease to which Porphyromonas gingivalis has been closely linked, in which the attachment tissues of the teeth and their alveolar bone housing are destroyed. We conducted a study to determine if immunization using a purified antigen could alter the onset and progression of the disease. METHODS: Using the ligature-induced model of periodontitis in Macaca fascicularis, we immunized five animals with cysteine protease purified from P. gingivalis and used an additional five animals as controls. Alveolar bone loss was measured by digital subtraction radiography. RESULTS: Immunization induced high titers of specific immunoglobuin G serum antibodies that were opsonic. Total bacterial load, levels of P. gingivalis in subgingival plaque and levels of prostaglandin E(2) in gingival crevicular fluid were significantly reduced. Onset and progression of alveolar bone loss was inhibited by approximately 50%. No manifestations of toxicity were observed. CONCLUSIONS: Immunization using a purified protein antigen from P. gingivalis inhibits alveolar bone destruction in a ligature-induced periodontitis model in M. fascicularis.     

鼠半胱氨酸蛋白酶抑制物cystatin S 对人口腔牙龈卟啉菌生长的抑制

目的：观察鼠半胱氨酸蛋白酶抑制物ｃｙｓｔａｔｉｎＳ对人口腔牙龈卟啉菌（Ｐｏｒｐｈｙｒｏｍｏｎａｓｇｉｎｇｉｖａｌｉｓ）生长的影响。方法：用琼脂扩散法检查了ｃｙｓｔａｔｉｎＳ对人口腔数种牙龈卟啉菌生长的抑制情况。结果：虽然对不同菌株其影响程度略有差异，对实验用牙龈卟啉菌３８１，３８１Ｐ，ＡＴＣＣＷ５０，Ｗ８３，３３２７７，ＮＣＴＣ１１８３４，ＶＰＩ１４０１８生长均有抑制作用，并且具有药物浓度依赖性。结论：鼠半胱氨酸蛋白酶抑制物ｃｙｓｔａｔｉｎＳ对人口腔牙龈卟啉菌生长有抑制作用。     

Identification and characterization of the cysteine protease inhibitor gene MdCPI from Musca domestica

Cysteine proteinase inhibitors (CPIs) are involved in many vital cellular processes such as signalling pathways, apoptosis, immune response and development; however, no CPIs have yet been reported from the housefly Musca domestica. Here we report the isolation and characterization of a housefly CPI gene designated MdCPI. The gene contains an open reading frame of 357 bp encoding a protein of 118 amino acid residues with a putative signal peptide of 17 amino acid residues. Protein alignment demonstrated a high homology to that of Sarcophaga crassipalpis (identity = 51%). Phylogenetic analysis suggested that all CPIs from dipterans, including the housefly, belong to the I25A family and may be descended from a single common ancestor. The gene was expressed in and purified from Escherichia coli. Biochemical studies showed that MdCPI exerts an inhibiting function on papain, which is a classical assay to confirm CPIs. Real-time quantitative PCR and immunolocalization analysis revealed that MdCPI is specifically expressed in haemocytes and fat bodies. It is highly down-regulated in larvae and markedly up-regulated in the pupal stage, suggesting that it may be related to development.