Subsensitivity of T lymphocytes to sympathomimetic and cholinergic stimulation in bronchial asthma.

The effect of adrenergic and cholinergic drugs on short incubation “active” E rosette formation was studied in 19 patients with bronchial asthma and 17 healthy controls. Both groups had an equal absolute number of baseline “active” E rosettes, but the asthmatics demonstrated a higher percent baseline value. The beta adrenergic drug isoproterenol (10^?3 M) inhibited the formation of “active” E rosettes in asthmatics by only 18.0% as compared to a 60.8% inhibition in the control group. Carbamylcholine (10^?5 M) a cholinergic agonist, also showed a lower than normal response in asthmatics, 34.3% enhancement of “active” E rosetting compared to a 52.4% enhancement in the controls. The alpha adrenergic agent phenylephrine (10^?5 M) exhibited equal enhancing effects in both groups, 34.2% in the asthmatics and 36.5% in the controls. Isoproterenol (10^?3 M) had a minimal effect on inhibition of long incubation “total” E rosettes in both groups studied. The beta adrenergic abnormality conforms to the beta blockade theory of asthma of Szentivanyi. The cholinergic abnormality is unexplained in view of the hyperresponsiveness of patients with asthma to cholinergic agents in vivo. Patients with bronchial asthma probably have an autonomic dysfunction that may play a role in the pathogenesis of their disease.     

Differential role of dopamine in drug- and lithium-conditioned saccharin avoidance

Rats learn to avoid palatable saccharin solutions that predict the systemic administration of reinforcing drugs as well as malaise-inducing lithium chloride (conditioned saccharin avoidance, CSA). In the present study the involvement of dopamine (DA) transmission in the acquisition of morphine, nicotine and lithium-conditioned CSA was investigated in a two-bottle choice paradigm. Nicotine tartrate (0.2 and 0.4 mg/kg s.c.) administered 15 min after saccharin presentation induced CSA, with a maximum effect at 0.4 mg/kg. The DA D1 receptor antagonist, SCH 39166 (0.1 mg/kg s.c.) and the DA D2 receptor antagonist raclopride (0.3 mg/kg s.c.), administered immediately after saccharin, prevented CSA induced by the lower but not by the higher dose of nicotine. However, combined administration of the two antagonists prevented CSA induced by the higher dose of nicotine. SCH 39166 prevented CSA induced by all morphine doses while raclopride prevented only CSA induced by the lowest dose of morphine (1.75 mg/kg). CSA induced by different doses of lithium given by the same schedule of drug-CSA (i.e. two pairings, 15 min after saccharin) was not affected by SCH 39166. However SCH 39166 impaired the acquisition of lithium-CSA when lithium was given 60 min after saccharin. In contrast, raclopride failed to affect lithium-CSA independently from the delay between saccharin and lithium. These results suggest that DA can play different roles in drug- and in lithium-CSA and are consistent with a different mechanism of drug- as compared to lithium-CSA.     

Study on nonspecific immunity in pregnant women: II. Effect of hormones on chemiluminescence response of peripheral blood phagocytes.

To analyze the mechanisms of increased nonspecific immunity in pregnant women, the effect of various hormones on the phagocytic activity was estimated by a luminol-dependent chemiluminescence (CL) response during phagocytosing opsonized zymosan. The CL response of whole blood supplemented with exogenous human chorionic gonadotropin (hCG) increased significantly in all the male and female subjects and pregnant women. An approximate two- to fourfold increase was observed in comparison with the unsupplemented control in each subject at concentrations ranging from 1 to 1,000 IU/ml after 48 h of incubation (P less than 0.05). Progesterone slightly stimulated the CL response in female subjects only, but had no effect on male and pregnant women. Estradiol (E2) did not stimulate the CL response in any subject. The expression of Fc and C3b receptors on the surface of polymorphonuclear leucocytes (PMNL) in pregnant women was also investigated by measuring the immunofluorescence stained with monoclonal antibody to Fc and C3b receptors, respectively. The relative numbers of Fc receptors increased significantly in the third trimester compared to those of female control (P less than 0.05). Those of C3b receptor also increased in the second and third trimester (P less than 0.005). These results suggested that the nonspecific immunity represented by phagocytic activity in pregnant women increased with both oxidative metabolic responsiveness and the expression of membrane receptors. Besides, the increased phagocytic activity of the maternal host is probably due to the stimulatory effect of both endogenous and exogenous hCG on their peripheral blood phagocytes.     

微小RNA-23b-3p通过靶向TGFBR3促进人心房肌成纤维细胞纤维化相关基因表达

目的:探究微小RNA-23b-3p(mi R-23b-3p)对人心房肌成纤维细胞中纤维化相关基因表达的作用及其可能作用靶基因。方法:分离并体外培养房颤患者心耳中原代心房肌成纤维细胞,并用细胞免疫荧光染色实验鉴定;双萤光素酶报告基因实验检测mi R-23b-3p与潜在靶基因转化生长因子β受体3(TGFBR3) 3'端非翻译区(3'-UTR)的结合作用; CCK-8、Ed U染色及Transwell实验检测细胞活力、增殖及迁移能力,RT-qPCR和Western blot法检测TGFBR3及纤维化相关基因的m RNA和蛋白表达。结果:在人心房肌成纤维细胞中过表达mi R-23b-3p不影响细胞的活力、增殖及迁移能力,但可显著增强细胞中纤维化相关基因COL1A1、COL3A1和ACTA2的表达(P 0. 05或P 0. 01)。双萤光素酶报告基因实验显示mi R-23b-3p与TGFBR3 3'-UTR有结合作用。RT-qPCR和Western blot结果证实mi R-23b-3p可在转录水平抑制TGFBR3表达。过表达mi R-23b-3p和沉默TGFBR3均能显著促进人心房肌成纤维细胞中Smad3激活和纤维化相关基因表达(P 0. 05或P 0. 01)。结论:TGFBR3是mi R-23b-3p的作用靶基因,并介导mi R-23b-3p促进心房肌成纤维细胞中纤维化相关基因表达。     

血管内皮损伤标志物在肾移植患者中的应用价值

目的 :通过对肾移植患者血浆可溶性血栓调节蛋白 (solublethrombomdulin ,sTM)、可溶性血管内皮细胞蛋白C受体 (endothelialproteinCreceptor ,sEPCR)、血管性血友病因子 (vonWillebrandfactor,vWF)含量水平的动态检测分析 ,探讨其在肾移植及移植排斥反应中的应用价值。方法 :采用双抗体夹心ELISA法检测 4 2例肾移植患者手术前后、排斥反应治疗过程中的sTM、sEPCR、vWF含量。结果 :肾移植术后sTM、sEPCR、vWF含量水平较术前明显增高 (P <0 0 5 )、与正常对照组比较有显著性差异 (P <0 0 1) ;发生移植排斥反应者较无排斥者有明显差异 (P <0 0 5 ) ;肾移植术前、排斥反应有效治疗后与正常对照组比较无显著差异 (P >0 0 5 ) ;肾移植患者sTM、sEPCR、vWF各指标间呈正相关 (r =0 5 95 ,P <0 0 1,r =0 5 81,P <0 0 1,r =0 6 0 6 ,P <0 0 1)。结论 :sTM、sEPCR、vWF均可作为肾移植患者血管内皮的免疫损伤标志物 ,联合动态监测sTM、sEPCR、vWF的含量在观察肾移植手术创伤程度、排斥反应的早期诊断和治疗效果的监测等方面均有重要的临床应用价值。     

Immunogenicity and T cell recognition in swine of foot-and-mouth disease virus polymerase 3D

Immunization of domestic pigs with a vaccinia virus (VV) recombinant expressing foot-and-mouth disease virus (FMDV) 3D protein conferred partial protection against challenge with infectious virus. The severity reduction of the clinical symptoms developed by the challenged animals occurred in the absence of significant levels of anti-3D circulating antibodies. This observation suggested that the partial protection observed was mediated by the induction of a 3D-specific cellular immune response. To gain information on the T cell recognition of FMDV 3D protein, we conducted in vitro proliferative assays using lymphocytes from outbred pigs experimentally infected with FMDV and 90 overlapping peptides spanning the complete 3D sequence. The use of pools of two to three peptides allowed the identification of T cell epitopes that were efficiently recognized by lymphocytes from at least four of the five animals analyzed. This recognition was heterotypic because anti-peptide responses increased upon reinfection of animals with a FMDV isolate from a different serotype. The results obtained with individual peptides confirmed the antigenicity observed with peptide pools. Detection of cytokine mRNAs by RT-PCR in lymphocytes stimulated in vitro by individual 3D peptides revealed that IFN-gamma mRNA was the most consistently induced, suggesting that the activated T cells belong to the Th 1 subset. These results indicate that 3D protein contains epitopes that can be efficiently recognized by porcine T lymphocytes from different infected animals, both upon primary and secondary (heterotypic) FMDV infection. These epitopes can extend the repertoire of viral T cell epitopes to be included in subunit and synthetic FMD vaccines.     

Structural and functional complexity of the humoral response against the Trypanosoma cruzi ribosomal P2 beta protein in patients with chronic Chagas' heart disease

High levels of antibodies against the C-terminus of the Trypanosoma cruzi TcP2 beta ribosomal protein, defined by the peptide EEEDDDMGFGLFD, named R13, have been measured in sera from patients with chronic Chagas' Heart Disease (cChHD). These antibodies also recognize an epitope on the second extracellular loop of the beta 1-adrenergic receptor, inducing a functional response on cardiomyocytes. The aim of this study was to gain novel insights into the structural basis of this cross-reactivity as well as to evaluate the origin of anti-M2- cholinergic receptor antibodies, which are also commonly found in cChHD patients. To address these questions we immunopurified anti-R13 antibodies and studied the structural requirements of epitope recognition. Results showed that the immunopurified antibodies recognized a conformation of R13 in which the third Glu residue was essential for binding, explaining their low affinity for the mammalian homologue (peptide H13: EESDDDMGFGLFD). Alanine mutation scanning showed individual variations in epitope recognition in each of the studied patients. The importance of a negatively charged residue at position 3 for the recognition of anti-R13 antibodies was further confirmed by competition experiments using a Ser3-phosphorylated H13 analogue, which had 10 times more affinity for the anti-R13 antibody than the native H13 peptide. Moreover, anti-R13 antibodies stimulated either the beta 1-adrenergic or the M2-cholinergic receptor, in strict agreement with the functional properties of the IgG fractions from which they derived, demonstrating that the same parasite antigen may generate antibody specificities with different functional properties. This may be a clue to explain the high variability of electrophysiological disturbances found in cChHD.     

凝血酶对大鼠脑微血管内皮细胞的影响

目的 :探讨凝血酶 (Thrombin ,TM )对脑微血管内皮的影响。方法 :将大鼠脑微血管内皮细胞进行培养 ,培养液中加入 10U的TM或10U的TM + 0 .4mU的组织蛋白酶G(CaspethsinG ,CATG ) ,相差显微镜动态观察内皮细胞形态的变化 ,免疫组织化学技术检测基质金属蛋白酶 2 (MatrixMetalloproteinase 2 ,MMP 2 )表达的改变。 结果 :TM使内皮细胞发生收缩 ,细胞收缩程度具有时间依赖性 ,使内皮细胞MMP 2表达水平明显增加。TM +CATG加入培养液后 ,细胞形态、MMP 2表达与对照组比较均无明显统计学差异 (P >0 .0 5 )。结论 :TM通过激活蛋白酶激活受体 1(proteaseactivatedreceptor 1,PAR 1) ,使内皮细胞发生收缩 ,促进MMP 2表达 ,是TM增加血脑屏障 (BloodBrainBarrier,BBB)通透性的可能机制。