Suppression of phosphorylation of extracellular-signal-regulated kinase and p38 mitogen-activated protein kinase in polymorphonuclear leukocytes by the proton pump inhibitor lansoprazole

Lansoprazole (LPZ) is a proton pump inhibitor that suppresses gastric secretion and exerts anti-inflammatory effects on immune cells. Recently, LPZ has been used for the treatment of peptic ulcer and gastritis, which can be caused by Helicobacter pylori, due to its potent acid-suppressive effects. We focused the aim to the anti-inflammatory effects on the over-activation of neutrophils, and investigated the effects of LPZ on the signal transduction of the mitogen-activated protein kinase (MAPK) family. LPZ slightly phosphorylated p38 MAPK of neutrophils at a concentration of 10 μg/ml, but did not phosphorylate extracellular-signal regulated kinase (ERK) 1/2. Pretreatment of neutrophils with (1–5 μg/ml) LPZ strongly attenuated the phorbol-12-myristate-13-acetate-stimulated phosphorylation of ERK1/2, and LPZ slightly suppressed the lipopolysaccharide (LPS)- and N-formylmethionylleucylphenylalanine-stimulated phosphorylation of p38. ERK1/2 produces the mitochondrial anti-apoptotic proteins, and the signaling pathway from LPS and N-formylmethionylleucylphenylalanine to p38 is the main pathway for reactive oxygen species production. The mechanism of anti-inflammatory effect of LPZ on hyper-activated neutrophils is suggested to be the suppression of signal transduction of ERK1/2 and p38 MAPK.     

In vitro demonstration of transport and delivery of antibiotics by polymorphonuclear leukocytes.

Several antibiotics are concentrated inside polymorphonuclear leukocytes (PMN). To investigate whether PMN could act as vehicles for delivery of antibiotics, we combined an assay measuring PMN chemotaxis under agarose with a bioassay measuring levels of antibiotic in agar. Double-layer plates were made by pouring a layer of chemotaxis agarose into tissue culture plates and then adding a thin layer of Trypticase soy agar. Neutrophils were incubated with antibiotic for 1 h and then were washed and placed in wells made in the plates. After allowing PMN to migrate under the agar toward a chemoattractant well containing formyl-methionine-leucine-phenylalanine for 3 h, Streptococcus pyogenes was streaked on top of the agar and grown overnight. PMN migration and zones of inhibition of bacterial growth were measured. Neutrophils migrated 2.51 +/- 0.16 mm toward the chemoattractant well and 1.48 +/- 0.12 mm toward the medium well; migration was not significantly affected by any of the antibiotics used. Plates with PMN incubated without antibiotic showed insignificant inhibition of bacterial growth toward chemoattractant and medium wells (0.38 +/- 0.18 and 0.14 +/- 0.12 mm, respectively; for both, P > 0.05 for difference from 0). PMN incubated with oxacillin (3 micrograms/ml), a drug not concentrated in PMN, caused a similar lack of inhibition (0.28 +/- 0.09 mm toward chemoattractant; 0.14 +/- 0.03 mm toward medium). Incubation with 30 microns of ciprofloxacin per ml resulted in inhibition that was similar in both directions (1.40 +/- 0.16 versus 1.18 +/- 0.13 mm). However, for PMN incubated with azithromycin (3 micrograms/ml), an agent highly concentrated inside phagocytes, there was a large degree of inhibition which was significantly greater in the direction of chemoattractant than in the direction of medium (3.47 +/- 0.30 versus 1.89 +/- 0.25 mm; P < 0.001), indicating that release of bioactive azithromycin by neutrophils occurred after migration. Likewise, after incubation with rifampin (10 micrograms/ml), which is also concentrated by PMN, inhibition was significantly greater in the direction of chemoattractant than in the direction of medium (1.54 +/- 0.24 versus 0.81 +/- 0.28 mm; P = 0.001). We conclude that for certain antibiotics, PMN may act as vehicles for transport and delivery of active drug to sites of infection.     

RNA and protein synthesis in human peripheral blood polymorphonuclear leukocytes

Polymorphonuclear leukocytes purified from human peripheral blood synthesized RNA and proteins when placed in cell culture. Autoradiography of the cultured cells revealed that a majority of mature PMNs were engaged in macromolecule synthesis, and an analysis of newly synthesized proteins by SDS-polyacrylamide gel electrophoresis showed that many different polypeptide chains were synthesized by these cells. The rate of [3H]uridine incorporation and the pattern of newly synthesized proteins were modulated by Con A and glucocorticoids. These results suggest that in spite of their short lifetime and a large performed enzymatic apparatus, mature PMNs retain a substantial capacity for RNA and protein synthesis; and, further, that modulation of macromolecule synthesis forms part of the mechanism by which PMNs respond to inflammatory and anti-flammatory stimuli.     

Membrane distribution and adsorptive endocytosis by C3b receptors on human polymorphonuclear leukocytes

C3b receptors on human polymorphonuclear leukocytes (PMN) were nonrandomly distributed in small clusters on the plasma membranes of these cells when assessed by indirect immunofluorescence at 0 degree C using monospecific rabbit Fab' or F(ab')2 anti-C3b receptor and tetramethylrhodamine isothiocyanate (TRITC)-conjugated goat IgG anti- F(ab')2. When PMN were incubated with the bivalent anti-C3b receptor at 37 rather than at 0 degree C, almost no immunofluorescence was observed, which indicates that the C3b receptor-F(ab')2 complexes had been rendered inaccessible to TRITC-IgG anti-F(ab')2. Endocytosis of the anti-C3b receptor ligand was quantitated by measuring the binding 131I-IgG anti-F(ab')2 by PMN that had previously taken up 125I-F(ab')2 anti-C3b receptor at 0 and at 37 degree C, respectively. There was a constant 2: 1 molar ratio of anti-F(ab')2 to anti-C3b receptor with PMN that had been incubated with the first antibody at 0 degree C. In contrast, when increments of F(ab')2 anti-C3b receptor were taken up by the cells at 37 degree C, there was a dose-related decline in this molar ratio to a minimum of 0.2 molecules of anti-F(ab')2 anti-F(ab')2 bound per molecule of PMN-associated anti-C3b receptor. 125I-F(ab')2 anti-C3b receptor taken up by PMN at 37 degree C was also inaccessible to release by proteolytic treatment of the cells with pronase. The rate of endocytosis of 125I-F(ab')2 anti-C3b receptor was rapid as the PMN- bound antibody fragment became inaccessible to 131I-IgG anti-F(ab')2 within 10 min during incubation of the cells at 37 degree C. In contrast to these findings, 125I-Fab' anti-C3b receptor that was taken up by PMN at 37 degree C remained accessible to both 131I-IgG anti- F(ab')2 and to proteolytic release by pronase, which suggests that monovalent interaction of ligand with C3b receptors was not sufficient for induction of endocytosis. The requirement for multivalency was also demonstrated using the C3b-OR, the normal ligand for the C3b receptor. 125I-C3b-OR was specifically bound by PMN but remained on the cell receptor. 125I-C3b-OR was specifically bound by PMN but remained on the cell surface, as determined by its accessibility to pronase, unless it was cross-linked with F(ab')2 anti-C3. Although C3b receptors on PMN do not mediate internalization of adsorptive pinocytosis of soluble ligand indicates their potential for the clearance of C3b-bearing immune complexes without recruitment of other cell surface receptors.     

蛋白激酶C在嗜酸细胞凋亡中的调控作用

目的本试验检测蛋白激酶C(PKC)在嗜酸细胞凋亡中的调控作用。方法用免疫组化和原为杂交的方法,选择嗜酸性粒细胞增多为特点的鼻息肉组织26例,检测PKC与抑凋亡基因bcl-2mRNA及其蛋白的相关性;用细胞培养的方法,用PKC抑制剂检测嗜酸性粒细胞凋亡指数。结果嗜酸性粒细胞中PKC与Bcl-2mRNA及其蛋白的表达分别呈明显的阳性正相关(r1=0.0875,r2=0.0823,P<0.01),PKC抑制剂可促进嗜酸性粒细胞凋亡。结论PKC在嗜酸细胞凋亡中可以抑制嗜酸细胞凋亡,为治疗因嗜酸性粒细胞增多性疾病的治疗提供新的途径。     

Hormonal regulation of glycogen synthase and phosphorylase activities in human polymorphonuclear leukocytes

Hormonal regulation of glycogen synthase and phosphorylase activities were studied in human polymorphonuclear leukocytes. Polymorphonuclear leukocytes from normal subjects were incubated with glucose, insulin, D,L-isoproterenol and L-thyroxine, either independently or in different combinations, and changes of the enzyme activity ratios of glycogen synthase (active form (I)/total activity (T)) and glycogen phosphorylase (active form (a)/total activity (T)) were assessed. Neither glucose nor insulin changed the glycogen synthase activity ratio. However, the proportion of the active form (I) of glycogen synthase was increased by the simultaneous addition of glucose and insulin to the incubation mixture, but D,L-isoproterenol or L-thyroxine diminished this effect and caused a decrease in the proportion of the active form of glycogen synthase. Insulin had no effect on the glycogen phosphorylase activity ratio. Glucose decreased the proportion of phosphorylase in the a form. The simultaneous addition of glucose and insulin caused no further changes, whereas in the presence of D,L-isoproterenol or L-thyroxine, this glucose effect was abolished and the proportion of phosphorylase a increased. These results show that both thyroid hormone and a beta-agonist alter glycogen metabolism to reduce glycogen storage in polymorphonuclear leukocytes.     

Ethanol withdrawal-associated allodynia and hyperalgesia: age-dependent regulation by protein kinase C epsilon and gamma isoenzymes.

Ethanol (EtOH) withdrawal increases sensitivity to painful stimuli in adult rats. In this study, withdrawal from a single, acute administration of EtOH dose-dependently produced mechanical allodynia and thermal hyperalgesia in postnatal day 7 (P7) rats. In contrast, P21 rats exhibited earlier and more prolonged mechanical allodynia but not thermal hyperalgesia. For both P7 and P21 rats, blood and spinal cord EtOH levels peaked at 30 minutes after administration, with P7 rats achieving overall higher spinal cord concentrations. Protein kinase C (PKC) has been implicated in mediating pain responses. Inhibitory PKC- and gamma-specific peptides attenuated mechanical allodynia and thermal hyperalgesia in P7 rats, whereas only the PKCgamma inhibitor prevented mechanical allodynia in P21 rats. Immunoreactive PKC in dorsal root ganglion and PKCgamma in lumbar spinal cord increased at 6 hours after EtOH administration in P7 rats. In P21 rats, the density of PKC immunoreactivity remained unchanged, whereas the density of PKCgamma immunoreactivity increased and translocation occurred. These studies demonstrate developmental differences in neonatal nociceptive responses after withdrawal from acute EtOH and implicate a role for specific PKC isozymes in EtOH withdrawal-associated allodynia and hyperalgesia. PERSPECTIVE: This study examines age-specific nociceptive responses after ethanol exposure by using 2 different ages of rats. The results suggest that ethanol age-dependently alters sensitivity to mechanical and thermal stimuli via specific protein kinase C isozymes. These results begin to ascertain the mechanisms that produce abnormal pain after alcohol exposure.     

Inhibition of platelet function by polymorphonuclear leukocytes

Human polymorphonuclear leukocytes (PMNs) were evaluated for their ability to modulate platelet response induced by collagen, thrombin, platelet-activating factor and the stable analog of cyclic endoperoxides U46619. Platelet aggregation was first evaluated in whole blood and in leukocyte-depleted whole blood by the impedance method. This novel approach highlighted the inhibitory role of leukocytes on platelet aggregation in whole blood. The inhibitory role of PMNs on platelet function was subsequently evaluated on washed cells. PMN inhibition of platelet aggregation and beta-thromboglobulin release was more evident with threshold concentrations of stimuli. The inhibition also depended on the number of PMNs incubated in mixed cellular suspensions. Higher concentrations of stimuli may overcome the PMN-dependent inhibition. Under this condition, preincubation of cells with N-formyl-methionyl-leucyl-phenylalanine (a specific PMN agonist) restored the inhibitory effect of PMNs on platelet aggregation in whole blood and in mixed cellular suspensions. Not only PMNs, but also PMN-derived supernatants, dose-dependently inhibited U46619-induced platelet aggregation, suggesting that the inhibition observed may be exerted by chemically stable compound(s). Cytoplasmic Ca2+ movement was measured in aequorin-loaded platelets exposed to thrombin or U46619 to see whether cytoplasmic Ca2+ levels were affected by PMN. Ca2+ levels were similar in the presence or absence of PMNs, suggesting that inhibition may be related to a subsequent platelet response step. A series of bioassay experiments showed that PMNs were able to remove and/or convert adenosine diphosphate available for platelet aggregation but not to reduce U46619 availability. Our findings suggest that (1) unstimulated PMNs may release factor(s) that inhibit platelet aggregation and beta-thromboglobulin release; (2) this in itself is sufficient to block the platelet response to a threshold concentration of stimuli; (3) release of the same or other inhibitory mediators from stimulated PMNs may have to be greater to inhibit platelet response to higher concentrations of stimuli. Data presented here suggest that adenosine diphosphatase activity and chemically stable, as yet unidentified, compounds besides previously well-characterized labile compounds such as nitric oxide and arachidonic acid metabolites are responsible for the PMN-dependent mechanism of inhibition of platelet response that could be relevant in physiopathologic conditions.     

Inhibition of nucleoside transport by protein kinase inhibitors

Recently we reported that the pyridinylimidazole class of p38 mitogen-activated protein (MAP) kinase inhibitors potently inhibited the facilitated transport of nucleosides and nucleoside analogs in K562 cells. These compounds competed with the binding of nitrobenzylthioinosine (NBMPR) to K562 cells, consistent with inhibition of the NBMPR-sensitive equilibrative transporter (ENT1). In this study we examined a large number of additional protein kinase inhibitors for their effects on nucleoside transport. We find that incubation of K562 cells with tyrosine kinase inhibitors (AG825, AG1517, AG1478, STI-571), protein kinase C (PKC) inhibitors (staurosporine, GF 109203X, R0 31-8220, arcyriarubin A), cyclin-dependent kinase inhibitors (roscovitine, olomoucine, indirubin-3'-monoxime), or rapamycin resulted in a dose-dependent reduction of intracellular uptake of [3H]uridine. In contrast, neither the MAP kinase kinase inhibitors (U0126, PD 98059) nor the phosphatidyl inositol-3 kinase inhibitors (wortmannin, LY 294002) affected this process. Furthermore, both transient uptake and prolonged [3H]thymidine incorporation in K562 cells were inhibited by protein kinase inhibitors, inactive analogs of kinase inhibitors (R0 31-6045, SB202474), and NBMPR, independently of effects on cell proliferation as determined by MTT assay. These studies demonstrate that a wide variety of protein kinase inhibitors affect nucleoside uptake through selective inhibition of nucleoside transporters, independently of kinase inhibition.     

Thromboxane generation by human peripheral blood polymorphonuclear leukocytes

Human peripheral blood polymorphonuclear leukocytes were stimulated to generate thromboxane B2 in a time- and concentration-dependent fashion upon exposure to serum-treated zymosan particles. Conversion by stimulated PMN of [14C] arachidonic acid to [14C]thromboxane B2 was confirmed by thin-layer radiochromatography, radio-gas chromatography, and mass spectrometry. Generation of thromboxane B2 was independent of platelet contamination and could be inhibited by the cyclooxygenase inhibitor, indomethacin. Cells rendered incapable of ingesting particles by treatment with cytochalasin B generated comparable amounts of thromboxane B2. These results suggest that human peripheral blood polymorphonuclear leukocytes synthesize thromboxanes in response to surface stimulation independently of phagocytosis.     

Aggregated C-reactive protein binds to human polymorphonuclear leukocytes and potentiates Fc receptor-mediated chemiluminescence

Aggregated C-reactive protein (CRP), an acute phase reactant, binds to and influences the functional activities of human mononuclear leukocytes. Our purpose was to examine the potential interactions between CRP and human polymorphonuclear leukocytes (PMNL). Binding of CRP to PMNL was examined by flow cytometry after cell incubation with fluorescein-labeled heat-aggregated CRP (Agg-CRP) or Agg-CRP and fluorescein-conjugated F(ab')2 anti-CRP antibodies. It was determined that, at an optimal dose of Agg-CRP (100 micrograms), approximately 36% of PMNL were fluorescence positive. This was in contrast to the 70% of monocytes and 8% of lymphocytes that expressed Agg-CRP binding sites. Although less than half of resting PMNL bound Agg-CRP, up to 93% of PMNL activated with 1.0 micrograms/ml of phorbol myristate acetate (PMA) expressed binding sites for Agg-CRP. Exposure to PMA similarly enhanced the amount of Agg-CRP bound per PMNL as determined by mean channel fluorescence. To evaluate whether Agg-CRP binding to PMNL could induce or modify a biologic response, respiratory burst activity was assessed by measuring luminol-enhanced chemiluminescence. Whereas Agg-CRP alone did not elicit a chemiluminescence response, Agg-CRP synergistically enhanced the chemiluminescence response induced by heat-aggregated IgG (Agg-IgG). Although this enhancing effect of Agg-CRP could be observed at both optimal and suboptimal concentrations of Agg-IgG, no enhancement of PMA or serum-opsonized zymosan-induced CL was detected. These data demonstrate that aggregated CRP binds to and selectively modulates the response of PMNL to Fc receptor-mediated activation.     

Ethylene formation by polymorphonuclear leukocytes. Role of myeloperoxidase

Ethylene formation from the thioethers, beta-methylthiopropionaldehyde (methional) and 2-keto-4-thiomethylbutyric acid by phagocytosing polymorphonuclear leukocytes (PMNs) was found to be largely dependent on myeloperoxidase (MPO). Conversion was less than 10% of normal when MPO-deficient PMNs were employed; formation by normal PMNs was inhibited by the peroxidase inhibitors, azide, and cyanide, and a model system consisting of MPO, H2O2, chloride (or bromide) and EDTA was found which shared many of the properties of the predominant PMN system. MPO-independent mechanisms of ethylene formation were also identified. Ethylene formation from methional by phagocytosing eosinophils and by H2O2 in the presence or absence of catalase was stimulated by azide. The presence of MPO-independent, azide-stimulable systems in the PMN preparations was suggested by the azide stimulation of ethylene formation from methional when MPO-deficient leukocytes were employed. Ethylene formation by dye-sensitized photooxidation was also demonstrated and evidence obtained for the involvement of singlet oxygen (1O2). These findings are discussed in relation to the participation of H2O2, hydroxyl radicals, the superoxide anion and 1O2 in the formation of ethylene by PMNs and by the MPO model system.     

Tumor necrosis factor-alpha-stimulated polymorphonuclear leukocytes suppress migration and bactericidal activity of polymorphonuclear leukocytes in a paracrine manner

OBJECTIVE: Polymorphonuclear leukocytes (PMN) and tumor necrosis factor-alpha (TNF-alpha) play prominent roles in acute respiratory distress syndrome, ischemia-reperfusion injury, trauma, and sepsis. Whereas direct effects of TNF-alpha on PMN function and viability are well documented, little data are available addressing the ability of PMN to communicate with each other in response to cytokine stimulation. Therefore, the aim of this study was to determine whether TNF-alpha can modulate PMN function by inducing PMN to secrete products upon stimulation, which would affect other PMN in vitro in a manner independent of cell contact. METHODS: PMN were purified daily from blood obtained from a pool of 22 healthy volunteers. Conditioned media (CM-TNF) was prepared by incubating PMN in Hanks' balanced salt solution plus TNF-alpha for 1-4 hrs. Freshly isolated PMN were resuspended in CM-TNF and analyzed for 1) phagocytosis of opsonized Escherichia coli, 2) oxidative metabolism as measured as an index of DCF-DA activation, and 3) migration to chemoattractants through Transwell inserts. RESULTS: CM-TNF decreased PMN phagocytotic activity by 8% to 15% and completely suppressed oxidative metabolism but did not modulate the expression of receptors associated with these functions. CM-TNF suppressed the migration of PMN to two biologically relevant agents, N-formyl-methionyl-leucyl-phenylalanine and leukotriene B4, by approximately 65%, but had no effect on PMN migration to interleukin-8. This suppression was observed for migration across plastic filters as well as extracellular matrix proteins. CONCLUSION: Our data demonstrate that PMN stimulated with TNF-alpha suppress the immunologic function and migration of other PMN independent of cell-cell contact and suggest that TNF-alpha may participate in a negative feedback loop by inducing a PMN-derived factor that counteracts its activity.     

Regulation of protein kinase C by sphingosine and lysosphingolipids

Protein kinase C (PKC), a calcium and phospholipid dependent protein kinase C, has emerged as a key element in signal transduction and cell regulation. It is activated by sn-1,2-diacylglycerol (DAG) second messengers and it serves as the receptor for phorbol esters, potent tumor promoters. PKC is now known to occur as a family of isoenzymes sharing similar structural features that allow regulation of activity by calcium, phospholipid, and DAG. In vitro mechanisms of PKC regulation by phospholipid, DAG, and phorbol esters have been studied using mixed micelles of Triton X-100/lipids. PKC activation occurs at physiologic mole fractions of phospholipid and DAG, does not require a bilayer, and appears to occur by a two-step mechanism whereby PKC initially interacts with a phospholipid surface and is then activated by the addition of DAG. Similar methodology has been used to explore the inhibition of PKC by different inhibitors that interact with its regulatory domain. Sphingosine and lysosphingolipids are potent inhibitors of PKC that prevent its interaction with DAG/phorbol esters. These naturally occurring metabolites have been shown to affect PKC activity in different cell systems. Disturbances in sphingolipid metabolism may lead to accumulation of lysosphingolipids with consequent inhibition of PKC. Additionally, these naturally occurring metabolites may have physiologic functions in regulating PKC activity by counteracting the action of DAG. The mechanism of action of sphingosine/lysosphingolipids and their possible physiologic function will be discussed.     

Fluorometric measurement of ofloxacin uptake by human polymorphonuclear leukocytes.

A fluorometric assay, based on the natural fluorescence of the quinolone nucleus, was used to determine the uptake of ofloxacin by human polymorphonuclear leukocytes. The ratio of cellular concentration to extracellular concentration (C/E) at 20 min and 37 degrees C was 7.2, using an extracellular concentration of 5 micrograms/ml. Uptake was rapid and was not affected by pH (5 to 9), but required elevated environmental temperature and cell viability. The metabolic inhibitors sodium fluoride and sodium cyanide significantly decreased the uptake of ofloxacin. The penetration of ofloxacin was not affected by the presence of glucose or adenosine, but was decreased by L-amino acids (lysine, leucine, and glycine). These results suggest that ofloxacin could be transported via an amino acid transport system and that the fluorometric assay is a useful method for determining the intracellular penetration of fluoroquinolones, avoiding the use of radiolabeled antimicrobial agents.     

Oxidative cross-linking of immune complexes by human polymorphonuclear leukocytes.

Incubation of human serum albumin-anti-human serum albumin immune complexes bound to a plastic surface, with human polymorphonuclear leukocytes for 1 h at 37 degrees C resulted in covalent cross-linking of 8.5% +/- 0.5 of the complexes, corresponding to a minimum rate of 700 antibody molecules per cell per minute. Similar results were obtained with IgG-anti-IgG and type II collagen-anticollagen II human antibodies. Cross-linking was defined as the antibody remaining attached to plastic-bound antigen after extraction with 3 M MgCl2 and 0.1 N HCl solutions. The effects of addition of oxygen radical scavengers, heme-enzyme inhibitors, and omission of Cl- indicated that the cross-linking process was mediated by the myeloperoxidase-H2O2-Cl- system. Cross-linking was also obtained with cell lysates, polymorphonuclear granules, and purified human myeloperoxidase in the presence of a steady flux of H2O2 provided by glucose oxidase-glucose. Cross-linking by the cell-free systems was also abolished by sodium azide or omission of chloride ions. Cross-linked immune complexes were also generated by incubation with 20 to 50 microM solutions of freshly distilled hypochlorous acid. Addition of 10 mM hypochlorous acid to soluble IgG resulted in the formation of protein precipitates insoluble in 5 M guanidine, 0.1 N HCl, or boiling 2.3 M sodium dodecyl sulfate-1.4 M 2-mercaptoethanol. The remaining soluble IgG contained fluorescent high molecular aggregates (ex: 360 nm; em: 454 nm). Oxidative cross-linking of antigen-antibody molecules, and of immune complexes to connective tissue macromolecules may play a pathogenic role in acute and chronic inflammatory processes.     

人类粒细胞杀菌蛋白治疗脓毒症小鼠

目的通过对脓毒症模型的治疗过程,观察人类杀菌蛋白（ＢＰ）的治疗作用,并与抗ＬＰＳ单克隆抗体１Ａ比较。方法脓毒症模型采用ＮＨ１雄性小鼠腹腔注射Ｏ１１１Ｂ４大肠杆菌悬液。给药分三组,ＢＰ０１ｍｇ,１Ａ１０ｍｇ,生理盐水（ＮＳ）０５ｍｌ。结果各组生存率无显著性差异（Ｐ＞００５）。第二次攻击后３小时外周血白细胞计数,ＮＳ组稍高于其他两组,分别为１９６±５７×１０６／ｍｌ、１５５±３８×１０６／ｍｌ和１５２±３０×１０６／ｍｌ,差异未达到显著性界值。血浆ＬＰＳ水平：ＢＰ和ｌＡ组明显低于ＮＳ组（Ｐ＜００５）。血浆细胞因子活性：ＢＰ组对细胞因子的抑制作用较强,分别在不同时间点对ＩＬ－１、ＩＬ－６和ＴＮＦ的数据低于ＮＳ组,有显著性差异（Ｐ＜００５）;ｌＡ只在一个时间对ＩＬ－６有抑制作用（Ｐ＜００５）。结论抗ＬＰＳ单抗和ＢＰ对ＬＰＳ均具有阻断作用,但是,ＢＰ的作用更强。两者均不能明显改善动物的生存率,应作为辅助治疗。     

Function of irradiated polymorphonuclear leukocytes obtained by buffy- coat centrifugation

Several studies suggest that transfusion of polymorphonuclear leukocytes (PMNs) may be beneficial in the treatment of septic neonatal patients. Because of expense, donor availability, and the technical effort involved in obtaining PMNs by intermittent or continuous flow leukapheresis, buffy coat centrifugation of whole blood has been suggested as an alternative source. An in vitro study was performed to determine whether PMNs collected by this method have adequate oxidative and migratory function measured by chemiluminescence (CL) and chemotaxis under agarose (CT), respectively. Whole blood samples from six adult volunteers were drawn into citrate-phosphate-dextrose-adenine-one and stored at 4 degrees C for 0 to 48 hours. One-half of each sample was irradiated with 1500 rads. PMNs isolated from the buffy coat of these samples had greater than 80 percent normal CT and CL following 0 to 28 hours of storage in whole blood. Irradiation caused no depression in function. Units of whole blood yielded 1.11 +/- 0.40 X 10(9) PMNs per unit. This study indicates that transfusion of radiated PMNs obtained from stored whole blood that is less than 28 hours old is reasonable to use in studies involving PMN transfusions.     

The effect of propranolol on protein synthesis and phagocytosis by polymorphonuclear leukocytes from newborn, children and elderly individuals

The effect of propranolol on protein synthesis by polymorphonuclear (PMN) leukocytes was assessed in subjects from four age categories: elderly individuals, grown-up children, small children and newborn babies. This function was inhibited in PMN cells from individuals of all age groups, but inversely proportional to the age of the individual, i.e. the younger the subject, the more inhibited was the protein synthesis by his PMN leukocytes. The phagocytic capacity of the PMN cells of elderly subjects was decreased in comparison with that of individuals from the younger age groups. Propranolol inhibited this cell function in cells from subjects of all categories, although the inhibition was more pronounced in the cells of the younger groups. The difference in the degree of inhibition of the cell function observed in the various age groups may indicate different functional properties of the cell membrane.