锦灯笼酸浆苦素B对活化多形核中性粒细胞化学发光及H_2O_2产生的影响

锦灯笼酸浆苦素Ｂ对活化多形核中性粒细胞化学发光及Ｈ＿２Ｏ＿２产生的影响北京中医研究所，山西运城地区高等专科学校李萍，盛巡，王晓中，韩秋萍本实验通过观察中性粒细胞（ｐｏｌｙｍｏｒｐｈｏｎｕｃｌｅａｒｌｅｕｋｏｃｙｔｅ，ＰＭＮ）化学发光及检测Ｈ２Ｏ２产生...     

大鼠肾小球巨噬细胞产生白细胞介素－1及氧自由基在肾小球损伤发生中的作用

本文用大鼠加速肾毒血清肾炎模型，于注射肾毒血清后７２ｈ，用电子自旋共振（ＥＳＫ）和胸腺细胞增殖检测肾炎鼠的肾小球巨噬细胞（ＧＭφ），自身腹腔Ｍφ（ＰＭφ）及正常鼠腹腔Ｍφ（Ｎ－ＰＭφ）所产生羟自由基（ＯＨ）和ＩＬ－１活性。同时观察ＩＬ－１对ＧＭφ、ＰＭφ和Ｎ－ＰＭφ产生ＯＨ的影响。结果显示：肾炎鼠ＧＭφＩＬ－１活性和ｂＨ明显高于ＰＭφ和Ｎ－ＰＭφ；ＩＬ－１能刺激Ｍφ产生ＯＨ，且炎症ＧＭφ的ＯＨ明显高于ＰＭφ和Ｎ－ＰＭφ，揭示：肾小球中浸润的Ｍφ过度活化并产生大量ＩＬ－１及ＯＨ，ＩＬ－１又能进一步刺激Ｍφ产生更多的ＯＨ，这一肾小球内的局部恶性循环，在加速肾毒血清肾损伤的发病机制中可能起着重要作用。     

P物质对人多形核白细胞功能的影响

目的研究Ｐ物质（ＳＰ）对人多形核白细胞（ＰＭＮ）有关方面作用及可能的意义。方法应用硝基四氮唑蓝（ＮＢＴ）还原法，荧光法和Ｇｒｉｅｓｓ反应等测定不同浓度ＳＰ单独或在细菌衍生肽类似物ＦＭＬＰ甲酯存在下对ＰＭＮ产生超氧阴离子（Ｏ－２，）过氧化氢（Ｈ２Ｏ２），一氧化氮（ＮＯ）和ＰＭＮ膜上一功能酶中性内肽酶（ＮＥＰ）活性及膜流动性的影响。结果ＳＰ（≥１０－５ｍｏｌ／Ｌ）可剌激ＰＭＮ显著产生Ｏ－２，Ｈ２Ｏ２和ＮＯ，后者被Ｌ－单甲基精氨酸（ＮＭＭＡ）所抑制；ＳＰ（１０－８～１０－４ｍｏｌ／Ｌ）能显著增加ＦＭＬＰ甲酯剌激ＰＭＮ产生Ｈ２Ｏ２，并显浓度递增依赖性；ＳＰ与ＦＭＬＰ甲酯协同可下调ＮＥＰ活性；ＳＰ能提高ＰＭＮ膜流动性。结论ＳＰ可通过ＰＭＮ介导调节炎症反应，对ＰＭＮ的杀菌功能有增强作用，ＳＰ对ＰＭＮ的这些影响可能是神经系统参与炎症和免疫调节的途径之一。     

ELISA法定量检测兔中性粒细胞CD18的表达

应用其自建的定量检测细胞粘附分子（ｃｅｌｌａｄｈｅｓｉｏｎｍｏｌｅｃｕｌｅｓ，ＣＡＭｓ）表达的细胞酶联免疫吸附测定法（ｃｅｌｌ－ＥＬＩＳＡ），将兔中性粒细胞（ＰＭＮ）用０．５％甲醛－ＨＢＳＳ固定于９６孔板，通过检测ＰＭＮ上结合物的酶活性和ＰＭＮ的蛋白量，以每分钟内每微克ＰＭＮ上的酶活性（ＯＤ／ｍｉｎ／μｇｐｒｏｔｅｉｎ）相对表示细胞粘附分子的表达，并用此法分别检测了静息状态和ＴＮＦ激活的兔ＰＭＮＣＤ１８的表达。     

多形核白细胞在内毒素休克时肝组织损伤中的作用

家兔输注内毒素０．３ｍｇ／ｋｇ复制内毒素休克模型。输注４ｈｒ以后ＰＭＮ吞噬发光和Ｏ＾－２生成呈持续升高（Ｐ＜０．０５～０．０１），同时激活的ＰＭＮ释放氧自由基损伤肝细胞，引起肝细胞内ＭＤＡ含量和培养上清ＬＤＨ活性升高（Ｐ＞０．０５～０．０１）。体外内毒素与ＰＭＮ共孵，ＰＭＮ吞噬发光和Ｏ＾－２生成呈先升高后回降的变化，经内毒素在体外激活的ＰＭＮ也能明显引起肝细胞内ＭＤＡ含量和上清ＬＤＨ活性升高（Ｐ＜     

嗜中性粒细胞致急性组织破坏的机理研究近况

宿主的嗜中性粒细胞（ＰＭＮ）致急性组织破坏的机理首先是ＰＭＮ与内皮粘附并进入组织，然后ＰＭＮ产生并释放毒性介质，在氧化环境和粘附微环境的保护下，毒性介质致组织破坏，本文还介绍了ＮＡＤＰ－１氧化酶产生机理、金属蛋白酶的激活机理、ＰＭＮ能否产生ＯＨ·以及氧代谢产物在体内的确切作用。     

内源性NO在内毒素引起的PAH和肺损伤中的作用

内源性ＮＯ在内毒素引起的ＰＡＨ和肺损伤中的作用河北医科大学病理生理教研室（石家庄０５００１７）万梅凌亦凌谷振勇实验观察了静脉预注入ＮＯ合成酶（ＮＯｓｙｎｔｈａｓｅ，ＮＯＳ）抑制剂Ｎω硝基左旋精氨酸（Ｎω－ｎｉｔｒｏ－Ｌ－ａｒｇｉｎｉｎｅ，Ｌ－ＮＮＡ...     

应用ONE-TOUCHⅡ型血糖监测仪测定毛细血管全血糖

应用ＯＮＥ－ＴＯＵＣＨⅡ型血糖监测仪测定毛细血管全血糖上海市内分泌研究所应用ＯＮＥ－ＴＯＵＣＨⅡ型血糖仪测定毛细血管全血糖（ＣＢＧ）１２６４例次，并与ＢｅｃｋｍａｎＣＸ４自动生化分析仪测定静脉血浆糖（ＶＰＧ）比较。结果表明，ＶＰＧ较ＣＢＧ高１０．２±...     

尼古丁对炎性细胞活化及细胞间粘附分子基因表达的影响

本文观察了尼古丁对炎性细胞活化、炎性细胞与内皮细胞粘附及内皮细胞粘附分子表达的作用，同时观察７６４－３（从丹参中提取的单体）对上述部分作用的影响，将有助于阐明尼古丁在ＣＯＰＤ等慢性炎症疾病发病中的作用并为防治提供实验资料。方法：按以往报道的方法，收集大鼠腹腔ＰＭＮ，观察尼古丁作用下ＰＭＮ释放的β－ｇ及溶菌酶活性，以反映ＰＭＮ的活化；培养脐静脉内皮细胞，加入ＰＭＮ，以观察尼古丁对ＰＭＮ－内皮细胞粘附的影响；转化及扩增含ＩＣＡＭ－ＩｃＤＮＡ的质粒，酶切、ＰＥＧ法纯化及琼脂糖凝胶电泳鉴定ＩＣＡＭ－Ｉｃ…     

低浓度H2O2对肺微血管内皮细胞的迟发效应--凋亡

目的和方法：对大鼠肺微血管内皮细胞（ＰＭＶＥＣ）培养、鉴定。观察了低浓度Ｈ２Ｏ２（０￣１００μＭ）对体外培养的ＰＭＶＥＣ的迟发性损伤效应。结果：ＭＩＴ法发现Ｈ２Ｏ２对ＰＭＶＥＣ的增殖代谢活性具有剂量依赖性抑制作用,而且当Ｈ２Ｏ２去除后,此作用仍可进一步发展。形态观察、流式细胞仪ＤＮＡ测定、３’末端标记ＤＮＡ降解片断原位检测（ＴＵＮＥＬ）均表明,细胞凋亡是Ｈ２Ｏ２对ＰＭＶＥＣ迟发性损伤效应的一种重要     

Stimulated platelets release factor(s) affecting the in vitro response of human polymorphonuclear cells

The metabolic and functional responses of human polymorphonuclear cells (PMNs) to thrombin-activated platelet supernatants were studied. The incubation of PMNs with supernatants from stimulated platelets (SPS) caused a 50% decrease in both killing of Staphylococcus aureus and luminol-enhanced chemiluminescence (CL) by PMNs stimulated by opsonized-zymosan (OZ), Concanavalin A (Con A), or calcium ionophore A23187. The levels of PMN intracellular fluorescence measured by flow cytometry, using the fluorochrome dichlorofluorescein diacetate (DCF-DA), were considerably less in the presence of SPS than in resting platelet supernatants (RPS). No influence of platelet supernatant on O2 consumption and O2- generation by OZ-activated PMNs was observed. The incubation of PMNs with SPS caused a significant increase in the rate of chemotaxis and aggregation elicited by Con A, OZ, and phorbol myristate acetate (PMA). The supernatant from resting platelets did not show any of the above-reported effects. Platelets previously degranulated by thrombin were unable to inhibit CL when activated with agonists. Studies on the differential release of the granules by platelets showed that the CL-quenching activity paralleled the discharge of lysosomal content. The release of myeloperoxidase (MPO) from PMNs elicited by OZ was reduced in the presence of SPS. The platelet supernatant did not affect the MPO activity if PMNs were lysed with Triton X-100. The leakage of lactate dehydrogenase (LDH) from platelets was less than 3%, and no catalase or superoxide dismutase was released. This activity withstood lyophilization, but was destroyed by 10 min heating at 100 degrees C or by treatment with proteolytic enzymes.     

TNF-α priming effect on polymorphonuclear leukocyte reactive oxygen species generation and adhesion molecule expression in hemodialyzed patients

Introduction The study aimed to assess reactive oxygen species generation and the expressions of some surface antigens on polymorphonuclear leukocytes (PMNs) in patients on regular hemodialysis (HD) treatment. Materials and Methods The respiratory burst of PMNs was determined with luminol-dependent chemiluminescence (CL) in resting cells and following N-formyl-methionyl-leucyl-phenylalanine (fMLP), phorbol 12-myristate 13-acetate (PMA), or opsonized zymosan (OZ) stimulation and expressed in arbitrary CL units times assay-time (aU × min). The expressions of CD11b/CD18, CD10, and CD13 receptors were determined with flow cytometry. Results Basal PMN CL was increased in HD patients to up to 1285 ± 129 aU × min compared with 895 ± 88 aU × min in healthy controls (p < 0.05). The CL of unprimed PMNs increased after fMLP stimulation from 3085 ± 746 to 4529 ± 808 aU × min, and after OZ stimulation from 12945 ± 1296 to 14678 ± 1355 aU × min. PMA-stimulated CL of PMNs was similar to control values. The oxidative burst in PMNs from HD patients and healthy controls was similar in response to TNF-α alone. The CL of TNF-α-primed PMNs in HD patients was significantly lower than CL measured in healthy controls (p < 0.05). The expressions of CD10 and CD13 metalloproteinase receptors were also increased (p < 0.05). Although CD11b expression was significantly increased at rest and after fMLP stimulation, the expression of another β-integrin heterodimer compound, CD18, was not increased. Conclusions These results provide evidence that TNF-α priming of PMNs is down-regulated in HD patients despite constitutive up-regulation of resting cytotoxicity and enhanced expression of adhesion and metalloproteinase receptors.     

Antioxidant Properties of Lunularia Cruciata (Bryophyta) Extract

The effects of Lunularia cruciata (L.) Dum (Bryophyta) acetonic extract was studied in vitro by means of luminol-dependent chemiluminescence (CL) emission from human peripheral whole blood phagocytes and isolated polymorphonuclear leukocytes (PMNs). L. crudata adult thalli underwent extraction with acetone. CL emission was evaluated in an automated luminometer, measuring the oxygen free-radical production by phagocytes incubated with opsonized zymosan (OZ) or phorbol myristate acetate (PMA), in absence or in presence of various concentrations of L. crudata extract. The CL results indicated that L. crudata induced significant changes in light emission from whole blood phagocytes, as well as isolated PMNs. Its inhibitory activity was more evident when resting isolated PMNs were studied. When the cells were activated, the greatest inhibitory effect was observed with PMA. The L, crudata activity could be caused by several compounds, such as flavonoids and or sesquiterpenes, present in the acetonic extract.     

The stimulation of superoxide anion production in guinea-pig peritoneal macrophages and neutrophils by phorbol myristate acetate, opsonized zymosan and IgG2-containing soluble immune complexes

The kinetics of superoxide anion production in guinea-pig peritoneal macrophages and neutrophils were determined following in vitro stimulation with phorbol myristate acetate (PMA), opsonized zymosan (OZ) and soluble immune complexes of guinea-pig IgG2 (SIC). Superoxide production was recorded as chemiluminescence (CL) arising from the reductive cleavage of lucigenin. With PMA, both macrophages and neutrophils displayed a two-phase response consisting of a rapid initial burst of CL, which preceded ligand ingestion, followed by a plateau in the CL response which persisted for more than 30 min. By contrast, OZ induced a slow progressive increase in CL in both phagocytes which was consistent with the development of an oxidative burst concomitant with ingestion. The phagocytes differed in their responses to SIC, the macrophages displaying CL kinetics similar to those observed with PMA, whereas the neutrophils responded in the manner observed with OZ. The relationship between disparity in the patterns of macrophage and neutrophil CL responses to SIC and differences in their expression of Fc receptors for IgG2 (Coupland & Leslie, 1983) is discussed.     

Saliva inhibits the chemiluminescence response, phagocytosis, and killing of Staphylococcus epidermidis by polymorphonuclear leukocytes.

Saliva inhibited several functional properties of polymorphonuclear leukocytes (PMNs) from murine peritoneal exudate, namely, luminol-mediated chemiluminescence (CL) induced by either Staphylococcus epidermidis or formylmethionyl-leucyl-phenylalanine (FMLP), phagocytosis, and killing of bacteria in vitro. The concentration of saliva in the reaction mixture that caused a complete inhibition of the CL response of PMNs to both S. epidermidis and FMLP was 25%. However, there was no catalase or superoxide dismutase activity in saliva that could influence the CL response of PMNs. The production of superoxide by PMNs stimulated with S. epidermidis was assayed in the presence or absence of saliva by inhibition of the reduction of cytochrome c by superoxide dismutase. In the presence of 50% saliva, O2- generation by PMNs was only 7.3% of that observed in the absence of saliva. After gel filtration of salivary material through Sephadex G-25 or Sephacryl S-200, several fractions were obtained that inhibited the CL response of PMNs to either FMLP or S. epidermidis or to both. Two inhibitory fractions were analyzed. One contained immunoglobulin A, and the other contained a peptide which was composed of 14 different amino acids. The two fractions of high molecular weight included in the first protein peak of Sephacryl S-200 gel filtration were able to inhibit the CL response to S. epidermidis and to inhibit phagocytic activity, while fractions of low molecular weight (under 12,500 Mr) inhibited the CL response to FMLP and to S. epidermidis but did not inhibit phagocytic activity.     

Ability of polymorphonuclear leukocytes to generate active oxygen species in children with bronchial asthma. Use of chemiluminescence probes with a Cypridina luciferin analog and luminol.

Airway inflammation with polymorphonuclear leukocytes (PMN) may play an important role in bronchial hyperresponsiveness (BHR). PMN generate superoxide anion (O2-) and other oxygen radicals that can damage lung tissue. We investigated the ability of peripheral PMN of children with bronchial asthma and control subjects to generate O2- and other active oxygen species using a 2-methyl-6-(p-methoxyphenyl)-3,7-dihydroimidazo[1,2-a]pyrazin++ +-3-one, a highly sensitive and specific chemiluminescence (CL) probe for O2-, and luminol-dependent CL. The ability of PMN of subjects with asthma to generate O2- and other active oxygen species was significantly greater than that of PMN of control subjects when stimulated with opsonized zymosan (OZ), phorbol myristate acetate or N-formyl-methionyl-leucyl-phenylalanine. Furthermore, in the same asthmatic children, the generation of O2- and other active oxygen species was significantly higher with attacks than without attacks when PMN were stimulated with OZ. We also demonstrated that O2- generation correlated with the degree of BHR to inhaled histamine. These results suggest that PMN of asthmatic children, especially those with attacks, generate more active oxygen species than that of control subjects and that airway inflammation caused by O2- may be closely related to BHR in subjects with bronchial asthma.     

Role of the capsular polysaccharide-like serotype-specific antigen in resistance of Actinobacillus actinomycetemcomitans to phagocytosis by human polymorphonuclear leukocytes.

Serotype b-specific polysaccharide antigen (SPA) of Actinobacillus actinomycetemcomitans Y4 consists of D-fucose and L-rhamnose. To clarify the role of SPA in phagocytosis of the organism by human polymorphonuclear leukocytes (PMNs), monoclonal antibodies (MAbs) against SPA and SPA-defective mutants, which were constructed by inserting the transposon Tn916 into strain Y4, were used in a chemiluminescence (CL) assay and a phagocytic killing assay. The CL responses of human PMNs to strain Y4 were very low, and the organism was not killed by PMNs. In contrast, SPA-defective mutants induced strong CL responses. The addition of immunoglobulin G MAbs against Y4 SPA enhanced significantly both the CL responses to strain Y4 and the killing of the organism in the presence of complement. The CL responses to SPA-defective mutants were little affected by the addition of these MAbs. We conclude that SPA of A. actinomycetemcomitans plays an important role in the resistance to host defenses by PMNs.     

Effect of piliation on interactions of Haemophilus influenzae type b with human polymorphonuclear leukocytes.

Piliated, adherent (P+) and nonpiliated, nonadherent (P-) strains of Haemophilus influenzae type b (Hib) were compared with respect to their ability to induce polymorphonuclear leukocyte (PMN) chemiluminescence (CL) and superoxide (O2-) generation and their susceptibility to phagocytosis by PMNs. P+ strains opsonized in normal human serum (NHS) induced significantly greater CL than did P- strains (500 X 10(5) +/- 112 X 10(5) versus 242 X 10(5) +/- 65 X 10(5) total counts per 60 min; P less than 0.001) when reacted with normal PMNs. Contributions of immunoglobulin and complement to CL activity in these mixtures were shown by findings of lower overall levels of CL when hypogammaglobulinemic serum or heat-inactivated NHS was used to opsonize either P+ or P- organisms. Results obtained with mixtures of hypogammaglobulinemic plus adsorbed heat-inactivated NHS (with P+ or P- organisms) suggested a role for an antipilus antibody in the enhancement of CL by these strains. NHS-opsonized P+ strains also induced significantly greater (P less than 0.002) O2- generation than did P- strains (2.83 +/- 0.08 versus 1.94 +/- 0.14 nmol of ferricytochrome c reduced per 10 min/10(6) PMN). Comparable ingestion of P+ or P- strains opsonized in NHS by PMNs was demonstrated by a radiolabeled uptake technique and transmission electron microscopy, and primary granule release (beta-glucuronidase) was comparable during ingestion of P+ or P- strains. The basis for the observed enhanced capacity of P+ Hib to stimulate PMN oxidative metabolism as compared with P- organisms is uncertain. Possible clinical implications of these findings deserve further study.     

1-naphthyl N-methyl carbamate effect on intra- and extracellular concentrations of arachidonic acid metabolites, and on the chemiluminescence generation by mouse peritoneal macrophages

1-Naphthyl N-methyl carbamate (carbaryl), potent carbamate insecticide with anticholinesterase activity, was tested for its ability to affect mouse peritoneal macrophages in particular arachidonic acid (AA) metabolism and oxidative burst. Carbaryl inhibited in a dose-related manner the reactive oxygen intermediate dependent chemiluminescence (CL) induced by opsonized zymosan (OZ), 12-O-tetradecanoyl phorbol-13-acetate (TPA) and calcium ionophore (A23187); this carbamate did not affect CL-mediated by AA. The intracellular and extracellular concentrations of prostaglandins (PGs) and 5-hydroxyeicosatetraenoic (5-HETE) generated in macrophages stimulated with OZ has been investigated for various periods. Carbaryl effect displayed two successive phases on AA metabolism stimulation. In a first phase (up to 2-15 min), carbaryl did not alter the rapid AA metabolite synthesis (total amount of intra- and extracellular metabolites) but it increased intracellular concentration of PGE2, PGA2, PGF2 alpha and decreased 5-HETE intracellular concentration. In a second phase (after 2-15 min), carbaryl inhibited AA metabolite synthesis. The release of cyclooxygenase (CO) and lipoxygenase (LO) metabolites decreased, in particular PGF2 alpha and PGD2 which in addition seemed to be submit to a cellular retention; the inhibition of other metabolite release appeared essentially related to the inhibition of their synthesis since the intracellular amount did not augment. The inhibition by carbaryl of the NADPH-oxidase dependent CL induced by OZ may be related to the alteration of the intra- and extracellular concentrations of AA metabolites.     

尼古丁对中性粒细胞活化及细胞间粘附分子基因表达的影响

目的:观察尼古丁对中性粒细胞(PMNs)的活化,PMNs与内皮细胞的粘附及内皮细胞表达ICAM-1mRNA,有助于阐明尼古丁在慢性阻塞性肺疾患(COPD)炎症发病中的作用。方法:测定β-葡萄糖醛酸苷酶及溶菌酶活性,以反映PMNs的活化;培养人脐静脉内皮细胞,观察PMNs与内皮细胞的粘附;制备探针,提取总RNA,Northern杂交测细胞间粘附分子-1(ICAM-1)mRNA。结果:尼古丁可活化PMNs,增加PMNs－内皮细胞粘附;增强ICAM-1mRNA表达,764-3可明显抑制尼古丁的上述作用。结论:尼古丁通过活化PMNs,促进PMNs－内皮细胞粘附,在COPD慢性炎症发病中起重要作用。而这种粘附作用的增加与粘附分子表达增强有关;抑制尼古丁的上述作用可能是764-3抗炎作用的部分机理。