烧伤后白细胞与内皮细胞粘附分子介导作用的研究

为研究白细胞粘附依赖于细胞粘附分子的介导作用,作者通过离体实验观察分析了烧伤血清对外周血中性粒细胞（ＰＭＮ）ＣＤ１１ｂ／ＣＤ１８表达的影响及ＣＤ１１ｂ／ＣＤ１８在烧伤后ＰＭＮ粘附中的介导作用。结果表明：（１）烧伤血清使ＰＭＮＣＤ１１ｂ／ＣＤ１８分子表达和ＰＭＮ与肺微血管内皮细胞（ＰＭＥＣ）的粘附率增高。（２）ＣＤ１１ｂ／ＣＤ１８单抗不仅能使正常ＰＭＮ与ＰＭＥＣ的粘附率下降５０％,也使烧伤血清激活的ＰＭＮ与ＰＭＥＣ粘附率下降至烧伤血清激活前水平。我们认为,严重烧伤能使外周血ＰＭＮＣＤ１１ｂ／ＣＤ１８表达明显增高,ＣＤ１１ｂ／ＣＤ１８不但介导基础状态下的ＰＭＮ粘附,更是介导烧伤后引起的大量ＰＭＮ与ＰＭＥＣ粘附的主要分子。     

LPS-stimulated PMN activation and proinflammatory mediator synthesis is downregulated by phosphodiesterase inhibition: role of pentoxifylline

BACKGROUND: Excessive production of reactive oxygen species by PMN is associated with tissue damage during inflammation. LPS interacts with the cell surface receptor CD14, which generates transmembrane signals through Toll-like protein 4 leading to mitogen activated protein kinase (MAPK) p38 activation, cytokine synthesis, PMN beta2-integrin expression and oxidative burst. Phosphodiesterase inhibition decreases proinflammatory cytokine production and tissue injury after LPS challenge. Its effects on PMN function after LPS stimulation, however, have not been fully investigated. We hypothesized that LPS-induced TNF-alpha synthesis and subsequent PMN beta2-integrin expression and oxidative burst are downregulated by concomitant treatment with the non-specific phosphodiesterase inhibitor pentoxifylline (PTX). METHODS: Whole blood was incubated with HBSS (control), LPS (100 microg/mL), fMLP (1 micromol/L), LPS+PTX (2 mmol/L) and fMLP+PTX for different time intervals at 37C. Oxidative burst, CD14, and CD-11b expression were measured by flow cytometry. Serum TNF-alpha levels were measured by ELISA. In an attempt to localize the site of action of PTX (proximal or distal to PKC) cell surface receptors were bypassed by PMA stimulation (1 microg/mL) and oxidative burst was measured with and without PTX. RESULTS: Up-regulation of CD14 expression was similar in LPS and LPS+PTX groups. LPS stimulation caused a significant increase in PMN oxidative burst, CD11b expression, and TNF-alpha serum levels. In addition, PMA and fMLP stimulation also caused significant increase in oxidative burst compared with controls. Concomitant addition of PTX to LPS led to a significant decrease in PMN oxidative burst (65%; p < 0.0001), PMN CD11b expression (20%; p = 0.012), and TNF-alpha levels (93%; p < 0.0001). Also, PMA- and fMLP-induced PMN oxidative burst were significantly decreased by PTX [77.5% (p < 0.0001) and 50% (p < 0.01), respectively]. CONCLUSIONS: These results suggest that PTX-inhibition of oxidative burst occurs distal to PKC and may be either due to direct inhibition of NADPH oxidase or inhibition of MAPK phosphorylation, leading to decreased adhesion molecule expression and TNF-alpha synthesis. Its use in clinical scenarios in which PMN are primed may be of clinical relevance.     

The influence of laparoscopic surgery on postoperative polymorphonuclear leukocyte function

Background: Laparoscopic surgery is thought to result in a better preservation of patients' immunological defenses. Polymorphonuclear leukocytes (PMN) are the most important effector cells in the elimination of pathogenic microorganisms. Because little is known about their function after laparoscopic surgery, we studied PMN phagocytosis, antigen expression, and oxygen radical production. Methods: In this study, 17 patients scheduled for Nissen fundoplication were randomly assigned to undergo either a laparoscopic or conventional procedure. To study phagocytic capacity, PMN were incubated with fluorescein isothiocyanate (FITC)-labeled Staphylococcus aureus. Plasma opsonic capacity was measured by comparing PMN phagocytosis in the presence of patients' own plasma with phagocytosis in the presence of control plasma. Cellular activation was measured by the expression of various cell surface markers and by assessment of PMA-stimulated oxidative burst. Results: Phagocytosis by PMN in the presence of patients' plasma was significantly lower 2 h after the conventional operation. No decrease in phagocytosis was observed when control plasma was used, indicating a decreased opsonic capacity of plasma after conventional surgery. No changes were observed after laparoscopic surgery. Furthermore, CD11b expression was significantly lower after the laparoscopic approach, indicating a blunted cellular activation. A significantly lower PMA-stimulated oxidative burst further confirmed the tempered stimulation after laparoscopic surgery. Conclusions: Laparoscopic surgery results in a preservation of the plasma opsonic capacity, and thereby the ability of PMN to phagocytose bacteria. Moreover, the postoperative cellular activation is reduced. The preserved phagocytosis and the blunted activation may prevent the development of postoperative infectious complications. Received: 12 February 1999/Accepted: 30 September 1999/Online publication: 9 August 2000     

烧伤后粘附分子介导中性粒细胞粘附对肺血管通透性的影响

目的观察比较烧伤血清及烧伤后中性白细胞（ＰＭＮ）对肺血管通透性的影响,分析ＰＭＮ粘附及粘附分子ＣＤ１１ｂ／ＣＤ１８在该影响中的介导作用。方法应用离体肺灌流技术,通过肺重量增加（ＬＷＧ）、液体滤过系数（Ｋｆ）和通透性表面积乘积（ＰＳ）分别观察肺水肿程度、肺血管对小分子物质和大分子物质的通透性。结果烧伤血清能使ＬＷＧ、Ｋｆ和ＰＳ明显增加,以Ｋｆ增加最为明显;烧伤后ＰＭＮ也能使Ｋｆ和ＰＳ增加,以ＰＳ增加为明显;用单抗封闭烧伤后ＰＭＮ膜上ＣＤ１１ｂ／ＣＤ１８后,ＰＭＮ在肺血管内的滞留减少,Ｋｆ和ＰＳ值增加被抑制,并以ＰＳ改变为显著。结论（１）被激活的ＰＭＮ释放的介质类物质如氧自由基、蛋白酶等物质对肺微血管内皮细胞（ＰＭＥＣ）的损伤作用部分依赖于ＰＭＮ与内皮细胞的粘附。（２）烧伤后被激活的ＰＭＮ释放的介质类物质主要介导肺血管对小分子物质通透性的影响。烧伤后ＰＭＮ与肺微血管内皮细胞（ＰＭＥＣ）的粘附除了使介质类物质的作用放大外,还介导肺血管对大分子物质通透作用。（３）ＰＭＮ膜上ＣＤ１１ｂ／ＣＤ１８分子可能通过与ＰＭＥＣ细胞间粘附分子的结合,本身具有对内皮细胞的生物学调控作用     

Whole-Blood Assay to Measure Oxidative Burst and Degranulation of Neutrophils for Monitoring Trauma Patients

AbstractBackground and Purpose: Polymorphonuclear neutrophils (PMNs) protect the host from invading microorganisms, but excessive PMN activation after trauma causes tissue injury. Rapid monitoring of PMN function is critical for the assessment of the inflammatory state of trauma patients. Here, the authors adapted two simple and rapid methods to measure oxidative burst and degranulation of human PMNs in whole blood to avoid potential interference of cell isolation procedures with the assessment of PMN function.Material and Methods: Heparinized blood was drawn from healthy volunteers or trauma patients, preincubated at 37 °C for 5 min, and stimulated with N-formyl-methionyl-leucyl-phenylalanine (fMLP). Four assays for oxidative burst were tested: (1) cytochrome C; (2) homovanillic acid (HVA); (3) Amplex^® Red; and (4) flow cytometry with dihydrorhodamine 123 (DHR). PMN degranulation was assessed with flow cytometry using antibodies to: (1) CD11b/Mac-1 (CD18); (2) CD63; and (3) CD66b (CD67).Results: With the exception of the DHR method, all methods to measure oxidative burst were found to be unsuitable in whole blood due to interference of plasma proteins and hemoglobin with the fluorimetric or photometric readouts. By contrast, all degranulation methods were suitable for whole-blood studies. However, for the assessment of formyl peptide-induced degranulation, anti-antibodies to CD11b/Mac-1 and CD66b were up to five times more sensitive than antibodies to CD63. Thus, the degranulation and DHR methods were optimized for increased sensitivity, speed, and specificity and their usefulness to measure PMN function in trauma patients was tested.Conclusion: The whole-blood methods based on flow cytometry with DHR, anti-CD11b/Mac-1, and anti- CD66b are rapid, simple, and reliable techniques to assess PMN function for trauma research.     

CD11／CD18在烧伤早期中性粒细胞对内皮细胞粘附中的…

为了解烧伤早期病人中性粒细胞（ＰＭＮ）膜表面ＣＤ１１ａ／ＣＤ１８和ＣＤ１１ｂ／Ｃｄ１８变化规律及其在烧伤早期ＰＭＮ对内皮细胞（ＥＣ）粘附及损伤中的作用，为临床抗白细胞粘附治疗提供依据，将烧伤早期ＰＭＮ与体外培养的人脐静脉内皮细胞（ＨＵＶＥＣ）孵育２４小时后，观察烧伤早期ＰＭＮ与ＥＣ粘附百分率和烧伤早期ＰＭＮ引起内皮细胞单层流出液生成速率（Ｊｖ）和滤过系数（ｋｆ）变化，ＣＤ１１／ＣＤ１８单抗（ｍＡｂ     

Endotoxin suppresses matrix protein-induced upregulation of PMN candicidal activity: an effect reversed by low-dose TNF-alpha.

We investigated the relationship of polymorphonuclear leukocyte (PMN) candicidal activity, matrix proteins, and lipopolysaccharide (LPS) to determine how LPS modulates the normal enhancing effect of matrix proteins on PMN candicidal activity. LPS reduced PMN candicidal activity when PMN were adhered in the presence of either fibronectin or laminin. In the presence of fibronectin or laminin, LPS reduced CD11b/CD18 expression (the fibronectin receptor) as assessed using sheep erythrocytes coated with C3bi. Experiments with 125I-fibronectin and 125I-RGDS (Arg-Gly-Asp-Ser) demonstrated that LPS reduced both the binding of fibronectin and the bioavailability of the binding epitope on the PMN surface. Stimulating the PMN oxidative burst with PMA but not FMLP also reduced fibronectin and RGDS binding. Incubation of LPS-treated PMN with staurosporine blocked the decrease in fibronectin and RGDS binding. Exposure of PMN to LPS plus low-dose TNF-alpha restored both fibronectin and RGDS binding with a concomitant increase in CD11b/CD18 surface expression. Low-dose TNF-alpha restored PMN candicidal activity in the presence of LPS and was most effective if PMN were preadhered to fibronectin. These results demonstrate that: (1) matrix proteins enhance normal PMN candicidal activity, (2) LPS reduces PMN candicidal activity in the presence of matrix proteins, (3) stimulation of the PMN oxidative burst in particular via protein kinase c activation reduces the bioavailability of the fibronectin receptor, and (4) low-dose TNF-alpha may restore PMN candicidal activity in part by upregulating the surface receptor for fibronectin binding.     

Influence of colloid fluids on polymorphonuclear granulocyte function in vivo

BACKGROUND: Granulocytes have a role in the immediate immune response. In a previous investigation we could demonstrate in vitro a moderate increase of the complement receptors CR1 (CD35) and CR3 (CD11b/CD18) on the surface of polymorphonuclear neutrophils (PMN) after incubation of whole blood with colloids. To elucidate the clinical significance, we investigated if these changes were also present in vivo. METHODS: The study was performed prior to anaesthesia for orthopaedic surgery. A total of 60 ASA-I patients was evaluated. Patients received in a randomised manner 7 mL/kg of the following solutions: human albumin 5% (HA), gelatine 4% (GEL), hydroxyethylstarch solution 6% with MW 200,000 Da, degree of substitution 0.5 (HES), or Ringer's solution. Prior to the infusion, at the end (30 min) and again 30 min later, blood samples were taken. Blood was incubated with fluorescein-conjugated monoclonal antibodies (CD11b, CD16, CD35, CD62L) and analysed with flow cytometry. RESULTS: HA, GEL, HES, and Ringer's solution failed to induce significant differences in the expression of complement receptors CR1 (CD35) and CR3 (CD11b/CD18), Fc gamma receptor IIIb (CD16), and of L-selectin (CD62L) receptor on the surface of PMN. CONCLUSIONS: Application of colloids like HA, GEL, or HES in moderate amounts shows no short-term effect on adhesion or activation molecules on granulocytes. However, in high doses, infused in situations such as multiple trauma and sepsis, the consequences on the function of PMN may be speculative and require further investigations.     

Improvement of lung preservation -- from experiment to clinical practice

BACKGROUND: Reperfusion injury represents a severe early complication following lung transplantation. Among the pathogenetic factors, the high potassium content of Euro-Collins(reg) solution is discussed. MATERIAL AND METHODS: In a pig model of orthotopic left-sided lung transplantation we investigated the effect of Euro-Collins solution (EC: n = 6) versus low potassium dextran (LPD: Perfadex: n = 6). Sham-operated (n = 6) animals served as control. Transplant function, cellular energy metabolism and endothelial morphology served as parameters. In a clinical investigation, 124 patients were evaluated following single (EC: n = 31; LPD n = 37) or double (EC: n = 17; LPD n = 39) lung transplantation, whose organs where preserved with EC (n = 48) or LPD (n = 76). Duration of ischemia, duration of ventilation and stay on ICU were registered. Primary transplant function was evaluated according to AaDO(2) values. Cause of early death (30 days) was declared. RESULTS: Experimental results: After flush with EC and 18 h ischemia, a reduction of tissue ATP content (p < 0.01 vs inital value and LPD) was noted. Endothelial damage after ischemia was severe (p < 0.05 vs control), paO(2) was significantly decreased. Clinical results: In the LPD group, duration of ischemia was longer for the grafts transplanted first (SLTx and DLTx: p = 0.0009) as well as second (2. organ DLTx: p = 0.045). Primary transplant function was improved (day 0: SLTx: p = 0.0015; DLTx: p = 0.0095, both vs EC). Duration of ventilation and stay on ICU were shorter (n.s.). Reperfusion injury-associated death was reduced from 8% (EC) to 0 (LPD). CONCLUSION: In experimental lung preservation, LPD lead to an improved graft function. These results were confirmed in clinical lung transplantation. Clinical lung preservation, therefore, should be carried out by use of LPD.     

Flush perfusion with low potassium dextran solution improves early graft function in clinical lung transplantation.

OBJECTIVES: We have previously demonstrated experimentally an amelioration of reperfusion injury of the lung after preservation using low potassium dextran (LPD) solution compared to Euro-Collins (EC) solution. Now we report on early graft function in 106 lung transplant recipients of LPD or EC preserved grafts. METHODS: Initial graft function was assessed by measurement of lung compliance and oxygenation index 2 h after transplantation. Length of stay on the intensive care unit and hours of mechanical ventilation were compared. Correlation of donor oxygenation, ischemic time, type of transplant, recipient age and sex as well as initial lung compliance and oxygenation with early postoperative course were calculated. RESULTS: Dynamic lung compliance was significantly (P<0.05) improved in the LPD group. PO(2)/fiO(2) was comparable in both groups (303+/-122 mmHg LPD, 282+/-118 mmHg EC). Mechanical ventilation was used for 321+/-500 h in the EC group and 189+/-365 h in the LPD group (P=0.006). Intensive care therapy was required for 17.2+/-23.7 days in the EC group and 10.4+/-16 days in the LPD group (P=0.012). Significantly higher lung function parameters were obtained in extubated recipients of LPD preserved grafts 2 weeks after TX. Thirty day graft survival was improved in the LPD group (P=0.045). In the EC group, 30 day mortality was 14.2 and 8% in the LPD group. CONCLUSIONS: A reduction of perioperative mortality and morbidity suggests that LPD solution has superior early graft function compared to lung preservation using EC solution.     

Functional and phenotypic changes in polymorphonuclear neutrophils induced by catecholamines

PURPOSE: To elucidate differential functional and phenotypic changes in response to relevant catecholamines, the generation of oxidative free radicals by PMN, and changes in the expression of L-selectin and Mac-1 on the surface of PMN were examined in the presence of epinephrine, norepinephrine and dopamine in physiological and pharmacological concentrations. MATERIALS AND METHODS: Human polymorphonuclear neutrophils were obtained from healthy donors and pretreated with 0.5 nM or 500 nM epinephrine; 1.18 nM or 1 180 nM norepinephrine; or 0.26 nM or 261 nM dopamine, followed by stimulation with FMLP. Stimulated neutrophils were incubated with antibodies against CD 11 b or CD 62 l and assessed by flow cytometry. Additional probes were assessed by flow cytometry for the generation of oxidative free radicals. RESULTS: All catecholamines in high concentration inhibited the suppression of CD 62 l expression and CD 11 b upregulation following stimulation with FMLP. A high concentration of epinephrine suppresses generation of oxidative free radicals. CONCLUSIONS: The effect of catecholamines on the expression of CD 62 l explains the increased expression of L-selection on PMN observed after trauma. The suppression of CD 11 b reduces leukocyte adherence and consecutive abnormalities in microvascular flow. Epinephrine inhibits the generation of oxidative free radicals by PMN with potentially detrimental effects with respect to bacterial clearance.     

The mechanism of action of the two-layer (Euro-Collins' solution/perfluorochemical) cold storage method in canine pancreas preservation

To clarify the mechanism of action of a two-layer [Euro-Collins' solution (EC)/perfluorochemical (PFC) ] cold storage method in the preservation of the pancreas, pancreatic viability and tissue concentrations of adenosine triphosphate (ATP) were examined in the canine model of pancreatic autotransplantation after preservation for 24 and 48 h by simple cold storage in EC (group 1), the two-layer, EC/PFC, method (group 2) and the two-layer, EC + 2, 4 dinitrophenol (DNP)/PFC, method (group 3). DNP is an uncoupler of oxidative phosphorylation. Maintenance of normoglycemia for at least 5 days after transplantation was considered a successful preservation. After preservation for 24 h, the functional success rates of groups 1, 2 and 3 were 100% (4/4), 100% (5/5) and 80% (4/5) respectively. One of five dogs in group 3 died of a cause unrelated to the pancreas. ATP tissue concentrations in group 2 were significantly higher than in group 1 (7.47 ± 0.47 gmol/g dry weight vs 1.41 ± 0.53 gmol/g dry weight, P < 0.01) and ATP tissue concentrations in group 3 were significantly lower than in group 2 (1.25 ± 0.37 gmol/g dry weight vs 7.47 ± 0.47 gmol/g dry weight, P < 0.01). It was apparent that ATP was not an essential factor for successful 24-hour preservation of the canine pancreas in EC because all the pancreatic grafts except one of five grafts in group 3 remained viable after preservation for 24 h, regardless of ATP tissue concentrations. On the other hand, after preservation for 48 h, the functional success rates for groups 1, 2 and 3 were 0% (0/4), 100% (4/4) and 0% (0/3) respectively. ATP tissue concentrations in group 2 were significantly higher than in group 1 (7.91 ± 1.21 gmol/g dry weight vs 1.21 ± 0.314tmol/g dry weight, P < 0.01) and ATP tissue concentrations in group 3 were significantly lower than in group 2 (0.61 ± 0.07 gmol/g dry weight vs 7.91 ± 1.21 µmol/g dry weight, P < 0.01). It was clear that preservation of the pancreas for 48 h was unsuccessful by simple cold storage in EC (group 1) and the two-layer method (group 2) made preservation for 48 h possible by increasing ATP tissue concentrations. However, DNP (group 3) inhibited the synthesis of ATP and the effectiveness of the two-layer method for 48-hour preservation of the pancreas. It was clear that maintenance of high ATP tissue concentrations during preservation was essential for the successful preservation of the canine pancreas in EC by the two-layer method for more than 48 h. We concluded that an adequate supply of oxygen to the pancreas during preservation by the two-layer method led to sufficient production of ATP to maintain cellular integrity and permitted the improvement of pancreatic preservation.     

甘氨酸对急性坏死性胰腺炎大鼠早期外周血中性粒细胞CD11b/CD18表达的影响

目的探讨急性坏死性胰腺炎早期外周血中性粒细胞（PMN）膜上CD11b／CD18及血中IL-6、LPS的动态变化及甘氨酸对急性坏死性胰腺炎（ANP）大鼠的保护作用。方法采用5％牛磺胆酸钠逆行胆胰管注射建立大鼠ANP肝损害模型，90只大鼠，随机分成三组。30只大鼠作为对照组，甘氨酸腹腔注射治疗30只，急性坏死性胰腺炎组30只，于不同时间取血作流式细胞仪分析，测定血IL-6、LPS含量变化。结果建立ANP模型后6、12、24h，外周血中性粒细胞（PMN）膜上CD11b／CD18及血中IL-6、LPS明显升高。甘氨酸治疗后，PMN CD11b／CD18表达及IL-6、LPS水平降低。结论CD11b／CD18表达和IL-6、LPS参与急性坏死性胰腺炎的病理机制。急性坏死性胰腺炎外周血中性粒细胞（PMN）膜上CD11b／CD18的变化可能与炎性介质的变化密切相关。甘氨酸可降低CD11b／CD18表达和IL-6、LPS的水平。     

The impact of liver preservation in HTK and UW solution on microcirculation after liver transplantation

Severe microcirculatory disturbances due to endothelial cell damage and leukocyte adherence during reperfusion of transplanted livers are considered to contribute to early graft failure. Since the degree of reperfusion injury after liver transplantation depends on the length of preservation time and the solution used for preservation, the aim of our study was to assess three solutions with respect to microvascular perfusion and leukocyte adhesion. Therefore, rat livers were stored up to 24 h in Euro-Collins (EC), University of Wisconsin (UW), or histidin-tryphtophan-ketoglutarate (HTK) solutions prior to orthotopic transplantation. The livers were studied in situ 60 min postoperatively using intravital fluorescence video microscopy. Using simple syringe flushing (10 ml), sinusoidal perfusion decreased below 50% in EC preserved livers after 8 h preservation, in HTK preserved livers after 16 h preservation, and remained higher than 70% in livers preserved in UW up to 24 h. Permanent adhesion of leukocytes was increased more rapidly in organs after 1, 8, 16, and 24 h preservation in HTK (16%, 15%, 34%, and 49.7% ± 4.7%) compared to those preserved in UW (15%, 18%, 17%; and 32.7% ± 3.3%; P < 0.05). Using a 10-fold volumn of the organ weight of HTK solution during the harvesting procedure, with an 8 min equilibration period, sinusoidal perfusion (39.6 ± 4.7%) and leukocyte adhesion (42.7 ± 3.1%) were not improved after 24 h. In contrast, equilibration with a volumn of approximately 40-times the liver weight improved sinusoidal perfusion (70.8% ± 2.7%; P < 0.01) and leukocyte adhesion (24.9% ± 3.1%; P < 0.01) significantly. Thus, using HTK solution, simple flushing prior to long-term cold storage resulted in microcirculatory disturbances when compared to UW solution. Larger volumns of HTK solution with an additional equilibration period of 8 min, however, reduced leukocyte adhesion and improved sinusoidal perfusion to a similar degree as UW solution.     

Randomized clinical trial on acute effects of i.v. iron sucrose during haemodialysis

Aim: Haemodialysis induces endothelial dysfunction by oxidation and inflammation. Intravenous iron administration during haemodialysis could worsen endothelial dysfunction. The aim of this study was to ascertain if iron produces endothelial dysfunction and the possible neutralizing effect of N‐acetylcysteine when infused before iron. The oxidative and inflammatory effects of iron during haemodialysis were also assessed. Methods: Forty patients undergoing haemodialysis were studied in a randomized and cross‐over design with and without N‐acetylcysteine infused before iron sucrose (50 or 100 mg). Plasma Von Willebrand factor (vWF), soluble intercellular adhesion molecule‐1 (sICAM‐1) levels, malondialdehyde, total antioxidant capacity, CD11b/CD18 expression in monocytes, interleukin (IL)‐8 in monocytes and plasma IL‐8 were studied at baseline and during haemodialysis. Results: Haemodialysis produced significant (P < 0.001) increase in plasma vWF, sICAM‐1, malondialdehyde, IL‐8 and CD11b/CD18 expression in monocytes, as well as decrease in total antioxidant capacity. Iron induced significant increase in plasma malondialdehyde and IL‐8 in monocytes, but had no effect on total antioxidant capacity, CD11b/CD18 expression, plasma IL‐8, vWF and sICAM‐1. The addition of N‐acetylcysteine to 50 mg of iron produced a significant (P = 0.040) decrease in malondialdehyde. Conclusion: Standard (100 mg) and low (50 mg) doses of iron during haemodialysis had no effects on endothelium. Iron only had minor effects on inflammation and produced an increase in oxidative stress, which was neutralized by N‐acetylcysteine at low iron dose. Haemodialysis caused a significant increase in oxidative stress, inflammation and endothelial dysfunction markers.     

Effects of spermidine and p-cresol on polymorphonuclear cell apoptosis and function

Polymorphonuclear leukocytes (PMNs) from chronic kidney disease (CKD) patients display accelerated apoptosis and dysfunction, which may predispose CKD patients to infections. In this study, we investigated the effect of spermidine and p-cresol on apoptosis and function on PMN from healthy subjects. We measured the effect of spermidine and p-cresol on apoptosis, ROS production unstimulated and stimulated (S. aureus and PMA) and expression of CD95, caspase 3, and CD11b on PMN. After incubation with p-cresol and spermidine, we did not observe any changes in apoptosis, viability or expression of caspase 3 and CD95 in PMN from healthy subjects. PMN incubated for 10 minutes with spermidine demonstrated a significant reduction in spontaneous, S. aureus and PMA-stimulated ROS production. p-cresol induced a decrease in PMA-stimulated ROS production. Spermidine and p-cresol also induced a decrease in the expression of CD11b on PMN. Spermidine and p-cresol decreased the expression of CD11b and oxidative burst of PMN from healthy subjects and had no effect on PMN apoptosis and viability.     

Antagonisation of platelet activating factor - a new therapeutic concept for improvement of organ quality in lung preservation

The release of platelet activating factor (PAF) is thought to be one of the most important pathophysiological pathways in the development of ischemic lung injury. We investigated the use of a PAF antagonist (PAF-a) in a canine model in reducing PAF-mediated pulmonary dysfunction following lung preservation and transplantation. Twelve combined heterotopic heart and orthotopic left lung allotransplantations were performed after 6 h of cold ischemia. Following administration of prostacyclin (PGI₂), Euro-Collins solution (EC) was used for pulmonary artery flush in all donors, while in six animals the PAF-a, WEB 2170 BS, was administered to the donor (0.15 mg/kg for 30 min), to the storage solution (0.3 mg/kg) and to the recipient during reperfusion for a total of 6 h (0.3 mg/kg per h) EC/PAF-a). In all donors myocardial preservation was achieved using St. Thomas Hospital solution. Postoperatively, cardiorespiratory function was evaluated seperately for donor and recipient organs at an FiO₂ of 0.4 for a maximum of 12 h. The quality of lung preservation was assessed by means of postoperative oxygenation (pO₂), pulmonary artery pressure (PAP) and pulmonary vascular resistance index (PVRI). In the EC/PAF-a group, pO₂ of the donor lung was significantly elevated (P < 0.01) and PVRI was significantly lower (P < 0.05) when compared to the EC group, while PAP showed no significant differences between both groups and throughout the entire postoperative course. We concluded that a significant improvement in the current clinical standard for lung preservation could be obtained by the application of WEB 2170 BS in combination with EC flush as demonstrated by improved oxygenation and lower PVRI of the transplantated organs.     

The effect of organ preservation solutions on kidney tubular and endothelial cells

Organ preservation solutions have primarily been tested in whole organ animal models. In the current study, we have examined the effect of commonly used organ preservation solutions on both kidney tubular and endothelial cells. Primary human endothelial and kidney tubular cells were incubated at 4 °C in the following solutions: 0.9 % saline (NS), EuroCollins (EC), University of Wisconsin (UW), or Hank's balanced salts with 5 % polyethylene glycol (PEG). Cell viability was assessed by colorometric measurement of mitochondrial reduction of 3 (4,5-dimethylthiazol-2-yl)-2-,5-diphenyltetrazolium bromide (MTT) to purple 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan. After hypothermic storage, cells were incubated at 37 °C in media with MTT, and the amount of reduced formazan present was quantified. Endothelial cells preserved in PEG displayed the best viability (P < 0.05). UW provided better cellular viability than EC or NS (P < 0.05). Control endothelial cells preserved in culture media at 37 °C displayed the highest absorbance values (P < 0.01).For kidney tubular cells, UW and PEG provided the best cellular protection (P < 0.05). Control kidney tubular cells cultured in complete media at 37 °C displayed the highest absorbance values (P < 0.01). Although the model presented here was not part of a truly morphological study, it may be more reliable for the rapid assessment of preservation-induced cell injury than models presented in previous morphological studies and may help in the development of improved preservation techniques. Received: 11 March 1997 Received after revision: 20 June 1997 Accepted: 1 August 1997     

Impact of lung preservation solutions, Euro-Collins vs. low-potassium dextran, on early graft function: a review of five clinical studies.

Currently, intracellular fluid type solutions that are prepared by modification of Euro-Collins solution (EC) have been the most commonly used by clinical lung transplant programs, which have been associated with a high rate of reperfusion injury. On the basis of the wealth of experimental data demonstrating the benefit of extracellular fluid type solutions over EC, one of the extracellular solutions, low-potassium dextran solution (LPD), became commercially available and has recently been applied to clinical settings. To date there have been 5 clinical studies comparing the effect of EC with LPD on early lung graft function. This article reviewed these 5 reports to determine whether the beneficial effects of LPD, as observed in experimental settings, can also be identified in clinical lung preservation. Four studies were retrospective reviews and one was a prospective non-randomized study. Of these 5 studies, 3 studies demonstrated significantly improved recipient oxygenation in the LPD group compared with the EC group in the early post-operative period. In addition, a significantly shorter duration on mechanical ventilation in the LPD group was shown in 2 studies, and improved reperfusion scores or pulmonary compliance was shown in one study each. Four of 5 studies concluded that graft preservation with LPD leads to superior early graft function after lung transplantation. The impact of LPD on prolonged lung preservation and also on long term results of graft function will be a subject of future investigation.     

Lung procurement by low-potassium dextran and the effect on preservation injury. Munich Lung Transplant Group.

BACKGROUND: This clinical study was performed to evaluate the effect of low-potassium dextran (LPD) solution on organ function in human lung transplantation. METHODS: A total of 80 patients were included in this study. Donor lungs were flushed with Euro-Collins (EC) solution in 48 cases or LPD (Perfadex) in 32 cases. Subsequently, single- (EC: n = 31; LPD: n = 15) or double-lung transplantations (EC: n = 17; LPD: n = 17) were performed. The evaluation parameters of transplant function were the reperfusion injury score (grade I to V); the alveolar/arterial oxygen ratio; the duration of respirator therapy; and the length of intensive care treatment and survival. RESULTS: Incidence and severity of reperfusion injury score were more severe in the EC group (31 of 48: grade I: n = 13; II: n = 8; III: n = 5; IV: n = 2; V: n = 3; LPD group: 17 of 32 patients; grade I: n = 12; II: n = 1; III: n = 3; IV: n = 0 grade V: n = 0), leading to death in three patients. In the LPD group, despite of the use of cardiopulmonary bypass, alveolar/arterial oxygen ratio values were significantly (P = 0.009) better during the early postoperative phase. Thirty-day mortality was 12% in the EC group and 6% in the LPD group. The one-year survival rate was 79% after the use of LPD (vs. EC: 62%). CONCLUSIONS: Graft preservation using LPD leads to better immediate and intermediate graft function after pulmonary transplantation and also results in better long-term survival.