Nuclear factor-�� B inducing kinase is required for graft-versus-host disease

Background

Donor T lymphocytes are directly responsible for graft-versus-host disease. Molecules important in T-cell function may, therefore, be appropriate targets for graft-versus-host disease therapy and/or prophylaxis. Here we analyzed whether nuclear factor-κ B inducing kinase might have a role in graft-versus-host disease.

Design and Methods

We studied the expression of nuclear factor-κ B inducing kinase in human samples from patients with graft-versus-host disease. We also explored the effect of nuclear factor-κ B inducing kinase in a murine model of graft-versus-host disease using donor cells from aly/aly mice (deficient in nuclear factor-κ B inducing kinase) and C57BL/6 mice (control).

Results

We detected expression of nuclear factor-κ B inducing kinase in T-lymphocytes in the pathological lesions of patients with acute graft-versus-host disease. Mice transplanted with aly/aly T lymphocytes did not develop graft-versus-host disease at all, while mice receiving C57BL/6 cells died of a lethal form of the disease. Deficiency of nuclear factor-κ B inducing kinase did not affect the engrafting ability of donor T cells, but severely impaired their expansion capacity early after transplantation, and aly/aly T cells showed a higher proportion of apoptosis than did C57BL/6 T cells. Effector T lymphocytes were the T-cell subset most affected by nuclear factor-κ B inducing kinase deficiency. We also detected lower amounts of inflammatory cytokines in the serum of mice receiving aly/aly T cells than in the serum of mice receiving C57BL/6 T cells.

Conclusions

Our results show that nuclear factor-κ B inducing kinase has a role in graft-versus-host disease by maintaining the viability of activated alloreactive T lymphocytes.     

Hepatic preconditioning of doxorubicin in stop-flow chemotherapy：NF-kB／IkB-α pathway and expression of HSP72

AIM: To provide hepatic protection through administration of doxorubicin before stop-flow chemotherapy (SFC) and to investigate the expression of heat shock protein 72(HSP72) and role of nuclear factor kappa B (NF-kB) in this effect.METHODS: The hepatic preconditioning of doxorubicin was established in a porcine model by injection of doxorubidn(1 mg/kg) before SFC. The experimental animals were randomized into two groups: groups receiving doxorubicin(DOX) and normal saline (NS). Serial serum and tissue samples were taken from both groups to evaluate the protection of doxorubicin. Western blot and immunoprecipitation were applied to detect the expression of HSP72, NF-kB p65 protein, inhibitor kB-α (IkB-α) and phosphorylated IkB-α as well. The expression of tumor necrosis factor α (TNF-α) was estimated by semiquantitative RT-PCR. And the extent of the hepatic injury was estimated with the level of serum aminotransferases.RESULTS: An abundance production of HSP72 in porcine liver was observed after 24 h of intravenous administration of doxorubicin, but without any change in the expression of NF-kB p65 subunit in cytoplasm. NF-kB p65 subunit accumulated in nuclei at the end of SFC and reached its highest level at 30 rain after the restoration of the abdominal circulation and decreased gradually during the 6 h after SFC in NS group, while there was little change in DOX group. There was also a slight decrease of IkB-α at 30 min after the restoration of the abdominal circulation in NS group accompanying with the appearance of phosphorylated IkB-α. The expression of TNF-α was significantly higher in NS group than that in DOX group (average 1.40±0.17 vs 0.62±0.22, P<0.01) at serial time points after SFC. Serum ALT and AST levels of NS group were higher after 24 h than those of DOX group (93.2±7.8 IU/L vs 53.34±13.9 IU/L,217.0±29.4 IU/L vs 155.0±15.6 IU/L for ALT and AST respectively, P<0.05) and alter 48 h than those of DOX group (66.6±18.1 IU/L vs 43.3±16.7 IU/L, 174.4±21.3 IU/Lvs 125.7±10.5 IU/L for ALT and AST respectively, P<0.05).CONCLUSION: Doxorubicin renders the liver to be tolerant to the hepatic influence in SFC in a porcine model through the NF-kB/IkB-α pathway with the expression of HSP72.     

Outcomes in COPD patients receiving tiotropium or salmeterol plus treatment with inhaled corticosteroids

Patients with COPD are frequently prescribed inhaled corticosteroids (ICS); however, it is unclear whether the treatment with ICS might modify responses to inhaled bronchodilators. Two 6-month, randomized, placebo-controlled, double-blind, double-dummy, parallel-group studies of tiotropium 18 μg once daily, compared with salmeterol, 50 μg bid, had been conducted in patients with moderate-to-severe COPD. Efficacy was assessed by spirometry, transition dyspnea index (TDI), St. George’s Respiratory Questionnaire (SGRQ), and exacerbations. Data from both studies were combined to form subgroups with regard to concurrent use of ICS. 796 patients receiving ICS were separately analyzed from 390 patients not receiving ICS. Mean age was 64 years, and pre-bronchodilator FEV₁ was 1.06 L (ICS group) and 1.13 L (non-ICS group). Both bronchodilators increased morning mean ± SE pre-dose FEV₁ compared with placebo (ICS groups: tiotropium 110 ± 20 mL, salmeterol 80 ± 20 mL; non-ICS groups: tiotropium 150 ± 30 mL, salmeterol 110 ± 30 mL; p > 0.05 for tiotropium vs salmeterol). Improvements in TDI and SGRQ and frequency of exacerbations also tended to be more profound for tiotropium. Treatment with tiotropium in patients with moderate-to-severe COPD was superior to salmeterol in lung function, irrespective of concurrent use of ICS.     

Gene therapy that inhibits NF-κB results in apoptosis of human hepatocarcinoma by recombinant adenovirus

AIM: To investigate whether the recombinant adenovirus induces the TNF-alpha-mediated apoptosis in vivo. METHODS: Human hepatocarcinoma cell line (HepG(2)) cells were transfected into BALB/c nude mice, and the tumor growth curve was drawn. We analyzed apoptosis in HepG(2) cells by TUNEL, HE staining and electron microscopy. RESULTS: AdIkappaBalphaM was expressed stably and efficiently in HepG(2) and could not be degraded by induction of TNF-alpha. Tumor growth in mice could be reduced remarkably if treated by AdIkappaBalphaM plus TNF-alpha. There was apoptosis of > 70% of cells treated with AdIkappaBalphaM plus TNF-alpha and about 50% of cells treated with AdIkappaBalphaM. In contrast, there was few cell apoptosis in HepG(2) cells treated with phosphate buffered saline and AdIkappaBalpha. HepG(2) cells in mice also exhibited a high level of apoptosis after in vivo injection with AdIkappaBalphaM. The tumor growth curve indicated the tumor transfected with AdIkappaBalphaM could be restrained. CONCLUSION: AdIkappaBalphaM gene therapy greatly enhances apoptosis due to inhibition of an NF-kappaB-mediated antiapoptosis signaling pathway.     

Oxygen therapy for COPD

Chronic obstructive pulmonary disease (COPD) is a leading cause of disability and death globally, characterised by progressive breathlessness, loss of function and, in its later stages, chronic hypoxaemia. Long-term continuous oxygen therapy increases life expectancy in patients with severe resting hypoxaemia. However, there are few data to support the use of oxygen in patients with only mild hypoxaemia and more research is required to determine any benefits of oxygen supplementation in COPD in such individuals.     

C反应蛋白水平在COPD病人、IHD病人，吸烟者和非吸烟者中的比较研究

目的评价在COPD病人中IHD症状和吸烟对CRP升高的潜在影响，并检验糖皮质激素（ICS）药物治疗与CRP血清水平之间的联系。方法对46名COPD患者与对照组的18名吸烟者（Smoking）和20名不吸烟者（N．Smoking）进行比较。结果COPD病人的CRP血清水平[4．83±1．23mg／L]显著高于两个对照组，具有显著性差异（P〈0．05），但是CRP血清水平在两个对照组中却无统计学意义。ICS治疗后，COPD病人的CRP水平降低。结论对COPD病人使用ICS治疗能够降低CRP水平。CRP可以作为COPD患者体内组织炎症的标志物。     

Expression and significance of nuclear factor κB p65 in colon tissues of rats with TNBS-induced colitis

AIM: To investigate the role of NF-κB in the pathogenesis of TNBS-induced colitis in rats. METHODS: Thirty-two healthy adult Sprague-Dawley (SD) rats were randomly divided into four groups of eight each: normal, NS, model Ⅰ, model Ⅱ groups in our study. Rat colitis model was established through 2-,4-,6-trinitrobenzene sulfonic acid (TNBS) enema. At the end of four weeks, the macroscopical and histological changes of the colon were examined and mucosa myeloperoxidase (MPO)activities assayed. NF-~B p65 expression was determined by Western blot assessment in cytoplasmic and nuclear extracts of colon tissue, and the expressions of TNF-α (and ICAM-1 protein in colon tissue were examined by immunohistochemistry. The relativities between expression of NF-κB p65 and other parameters were analyzed. RESULTS: TNBS enema resulted in pronounced pathological changes of colonic mucosa in model Ⅱ group (macroscopic and histological injury indices 6.25±1.39 and 6.24±1.04, respectively), which were in accordance with the significantly elevated MPO activity (1.69+0.11). And the nuclear level of NF-κB and expression of TNF-α, ICAM-1 in rats of model Ⅱ group were higher than that of normal control(9.7±1.96 vs 1.7±0.15, 84.09±14.52 vs 16.03±6.21,77.69±8.09 vs 13.41±4.91 P<0.01), Linear correlation analysis revealed that there were strong correlations between the nuclear level of NF-κB and the tissue positive expression of TNF-α and ICAM-1, MPO activities, macroscopical and histological indices in TNBS-induced colitis, respectively (r = 0.8235, 0.8780, 0.8572, 0.9152,0.8247; P<0.05). CONCLUSION: NF-κB plays a pivotal role in the pathogenesis of ulcerative colitis, which might account for the up-regulation the expression of TNF-α and ICAM-1.     

Pulmonary haemodynamic changes in patients with severe COPD

Worsening gas exchange during exercise and during exacerbations of COPD contributes to systemic hypoxaemia and reduces quality of life. However, pulmonary haemodynamic changes under such conditions are not well understood. Right heart catheterization was performed in six patients with severe COPD (%FEV(1) < 50%) during rest, exercise and during an exacerbation. Pulmonary artery pressure (Ppa) was slightly elevated at rest. The Ppa, as well as pulmonary artery wedge pressure (Pawp) and cardiac index were significantly increased during bicycle ergometer exercise. In contrast, pulmonary vascular resistance increased significantly during an exacerbation accompanied by a slightly increased Ppa. Supplemental oxygen resulted in significant decreases in Ppa and Pawp during exercise and Ppa during exacerbations. In patients with COPD, haemodynamic changes in the pulmonary circulation may differ during exercise and with exacerbations. Supplementary oxygen is beneficial and associated with reductions in pulmonary arterial pressures.     

Role of N-acetylcysteine in the management of COPD

The importance of the underlying local and systemic oxidative stress and inflammation in chronic obstructive pulmonary disease (COPD) has long been established. In view of the lack of therapy that might inhibit the progress of the disease, there is an urgent need for a successful therapeutic approach that, through affecting the pathological processes, will influence the subsequent issues in COPD management such as lung function, airway clearance, dyspnoea, exacerbation, and quality of life. N-acetylcysteine (NAC) is a mucolytic and antioxidant drug that may also influence several inflammatory pathways. It provides the sulfhydryl groups and acts both as a precursor of reduced glutathione and as a direct reactive oxygen species (ROS) scavenger, hence regulating the redox status in the cells. The changed redox status may, in turn, influence the inflammation-controlling pathways. Moreover, as a mucolytic drug, it may, by means of decreasing viscosity of the sputum, clean the bronchi leading to a decrease in dyspnoea and improved lung function. Nevertheless, as successful as it is in the in vitro studies and in vivo studies with high dosage, its actions at the dosages used in COPD management are debatable. It seems to influence exacerbation rate and limit the number of hospitalization days, however, with little or no influence on the lung function parameters. Despite these considerations and in view of the present lack of effective therapies to inhibit disease progression in COPD, NAC and its derivatives with their multiple molecular modes of action remain promising medication once doses and route of administration are optimized.     

Airway inflammation and anti-protease defences rapidly improve during treatment of an acute exacerbation of COPD

Background and objective:   There are few data on the short-term natural history of airway inflammation during severe episodes of acute exacerbation of COPD (AECOPD). An observational study was performed to determine how rapidly conventional treatment improves airway inflammation in patients admitted to hospital with AECOPD.
Methods:   Twenty-four consecutive patients with AECOPD were recruited and changes in sputum inflammatory indices were assessed after 2 and 4 days of treatment. The primary end-points included presence of bacteria and viruses, changes in sputum total cell counts (TCC) and polymorphonuclear leukocyte (PMN) differential counts, and levels of secretory leukocyte protease inhibitor (SLPI), IL-8 and TNF-α.
Results:   All patients received oral corticosteroids and 17 (71%) were also treated with oral antibiotics. A bacterial or viral pathogen was isolated from 12 patients (50%), and Aspergillus fumigatus was isolated from one. A positive bacterial culture was associated with increased sputum TCC and PMN count ( P  < 0.05), as well as higher levels of IL-8 and TNF-α ( P  < 0.05), and a trend towards lower sputum SLPI levels ( P  = 0.06). Sputum PMN numbers fell by 70% within the first 48 h of admission ( P  < 0.05), accompanied by an increase in sputum SLPI ( P  < 0.001) and reductions in the levels of TNF-α ( P  < 0.005) and IL-8 ( P  = 0.06), with no further significant change at 4 days.
Conclusions:   Conventional treatment for severe AECOPD is associated with rapid reduction of neutrophilic inflammation and improvement in anti-protease defences.     

Exertional hypoxemia is more severe in fibrotic interstitial lung disease than in COPD

Background and objective

Despite its clinical and prognostic significance, few studies have evaluated the severity of exertional oxygen desaturation in fibrotic interstitial lung disease (ILD). Our objectives were to identify clinical and physiological variables that predict the extent of exertional oxygen desaturation in fibrotic ILD and to quantify the severity of desaturation compared to chronic obstructive pulmonary disease (COPD).

Methods

This retrospective study compared the results of 6‐min walk test (6MWT) performed while breathing room air in fibrotic ILD patients and COPD patients eligible for pulmonary rehabilitation. Outcomes included the oxygen saturation (SpO₂) nadir and the change in SpO₂ from rest during a 6MWT. Predictor variables were identified on unadjusted analysis, followed by multivariate analysis to identify independent predictors of desaturation.

Results

The study included 134 patients with fibrotic ILD and 274 patients with COPD. The ILD and COPD cohorts had similar age, sex, frequency of major comorbidities, walk distance, baseline SpO₂ and baseline Borg dyspnoea scores. DL_CO was the strongest predictor of desaturation in both cohorts. Compared to patients with COPD, ILD patients had significantly lower SpO₂ nadir values (88.1 ± 6.4 vs 91.0 ± 4.6) and greater decrease in SpO₂ from baseline (7.4 ± 5.2 vs 4.5 ± 3.7) after adjusting for demographic features and pulmonary physiology (P < 0.0005), with greater between‐group differences at lower DL_CO values.

Conclusion

Patients with fibrotic ILD have greater oxygen desaturation during 6MWT compared to patients with COPD when adjusting for demographic features and pulmonary physiology. These findings suggest the need for disease‐specific studies to evaluate the potential utility of ambulatory oxygen in fibrotic ILD.

     

NIK overexpression amplifies, whereas ablation of its TRAF3-binding domain replaces BAFF:BAFF-R-mediated survival signals in B cells

BAFF-R-dependent activation of the alternative NF-κB pathway plays an essential role in mature B cell survival. Mutations leading to overexpression of NIK and deletion of the TRAF3 gene are implicated in human multiple myeloma. We show that overexpression of NIK in mouse B lymphocytes amplifies alternative NF-κB activation and peripheral B cell numbers in a BAFF-R-dependent manner, whereas uncoupling NIK from TRAF3-mediated control causes maximal p100 processing and dramatic hyperplasia of BAFF-R-independent B cells. NIK controls alternative NF-κB signaling by increasing the protein levels of its negative regulator TRAF3 in a dose-dependent fashion. This mechanism keeps NIK protein levels below detection even when they cause B cell hyperplasia, so that contributions of NIK to B cell pathologies can easily be overlooked.     

IkappaB kinase beta phosphorylates Dok1 serines in response to TNF, IL-1, or gamma radiation

Dok1 is an abundant Ras-GTPase-activating protein-associated tyrosine kinase substrate that negatively regulates cell growth and promotes migration. We now find that IkappaB kinase beta (IKKbeta) associated with and phosphorylated Dok1 in human epithelial cells and B lymphocytes. IKKbeta phosphorylation of Dok1 depended on Dok1 S(439), S(443), S(446), and S(450). Recombinant IKKbeta also phosphorylated Dok1 or Dok1 amino acids 430-481 in vitro. TNF-alpha, IL-1, gamma radiation, or IKKbeta overexpression phosphorylated Dok1 S(443), S(446), and S(450) in vivo, as detected with Dok1 phospho-S site-specific antisera. Moreover, Dok1 with S(439), S(443), S(446), and S(450) mutated to A was not phosphorylated by IKKbeta in vivo. Surprisingly, mutant Dok1 A(439), A(443), A(446), and A(450) differed from wild-type Dok1 in not inhibiting platelet-derived growth factor-induced extracellular signal-regulated kinase 1/2 phosphorylation or cell growth. Mutant Dok1 A(439), A(443), A(446), and A(450) also did not promote cell motility, whereas wild-type Dok1 promoted cell motility, and Dok1 E(439), E(443), E(446), and E(450) further enhanced cell motility. These data indicate that IKKbeta phosphorylates Dok1 S(439)S(443) and S(446)S(450) after TNF-alpha, IL-1, or gamma-radiation and implicate the critical Dok1 serines in Dok1 effects after tyrosine kinase activation.     

Extracellular superoxide dismutase protects against pulmonary emphysema by attenuating oxidative fragmentation of ECM

Extracellular superoxide dismutase (ECSOD or SOD3) is highly expressed in lungs and functions as a scavenger of O₂^{• ─}. ECM fragmentation, which can be triggered by oxidative stress, participates in the pathogenesis of chronic obstructive pulmonary disease (COPD) through attracting inflammatory cells into the lungs. The level of SOD3 is significantly decreased in lungs of patients with COPD. However, the role of endogenous SOD3 in the development/progression of emphysema is unknown. We hypothesized that SOD3 protects against emphysema by attenuating oxidative fragmentation of ECM in mice. To test this hypothesis, SOD3-deficient, SOD3-transgenic, and WT C57BL/6J mice were exposed to cigarette smoke (CS) for 3 d (300 mg total particulate matter/m³) to 6 mo (100 mg/m³ total particulate matter) or by intratracheal elastase injection. Airspace enlargement, lung inflammation, lung mechanical properties, and exercise tolerance were determined at different time points during CS exposure or after elastase administration. CS exposure and elastase administration caused airspace enlargement as well as impaired lung function and exercise capacity in SOD3-null mice, which were improved in mice overexpressing SOD3 and by pharmacological SOD mimetic. These phenomena were associated with SOD3-mediated protection against oxidative fragmentation of ECM, such as heparin sulfate and elastin, thereby attenuating lung inflammatory response. In conclusion, SOD3 attenuates emphysema and reduces oxidative fragmentation of ECM in mouse lung. Thus, pharmacological augmentation of SOD3 in the lung may have a therapeutic potential in the intervention of COPD/emphysema.     

重组人血管内皮抑制素对COPD大鼠肺血流动力学指标的影响

目的观察重组人血管内皮抑制素对慢性阻塞性肺疾病（COPD）病程的影响及机制。方法将64只大鼠随机分为8组各8只,其中A组和B组分别于第1、14天气道内注入生理盐水及脂多糖（LPS）溶液;C组同上注入LPS溶液,同时采用香烟暴露法制备COPD模型,共1个月;D组同上注入LPS溶液并行香烟烟雾暴露及18％氧气吸人制备COPD并肺动脉高压（PAH）模型;B1组、C1组、D1组分别按B、C、D组造模,并腹腔注射重组人血管内皮抑制素;Al组处理方式同A组,并腹腔注射生理盐水。采用肺动脉插管法测定各组肺血流动力学指标,采用ELISA法检测肺泡灌洗液（BALF）中血管内皮生长因子（VEGF）蛋白表达情况,采用RT-PCR法、Western-blot法检测肺组织匀浆中VEGF mRNA及蛋白表达,分析VEGF表达与肺血流动力学指标的相关性。结果C组和D组右心室收缩压（RVSP）、平均肺动脉压（mPAP）、右心室肥厚指数（RVHI）均明显高于A组,Cl、D1组此三项指标均较对应C、D组下降（P均〈0．05）;C组和D组BALF中VEGF蛋白表达水平显著高于A组,Cl、D1组VEGF蛋白表达较对应C、D组显著下降（P均〈0．05）;B组、C组和D组肺组织中VEGF mRNA及蛋白表达逐渐增强,B1、C1、D1组VEGF mRNA表达显著低于B组、C组和D组,C1、Dl组VEGF蛋白表达显著低于C组和D组（P均〈0．05）;肺组织中VEGF mRNA及BALF中VEGF蛋白水平均与RVSP、mPAP呈正相关（P均〈0．05）。结论重组人血管内皮抑制素可延缓COPD病情进展,可能机制为下调VEGF表达;此为COPD药物治疗提供了新的思路。     

Sleep hypoventilation due to increased nocturnal oxygen flow in hypercapnic COPD patients

Background and objective: Several COPD treatment guidelines recommend increasing oxygen flow during sleep to avoid nocturnal desaturation. However, such an increase could have deleterious clinical and gas exchange effects. The objective of this study was to evaluate short‐term gas exchange alterations produced by increasing the nocturnal oxygen flow rate. Methods: Thirty‐eight COPD patients with chronic hypercapnic respiratory failure were evaluated. In a cross‐over study, patients were randomly assigned to receive the daytime oxygen flow rate on one night and an additional litre on the alternate night. Nocturnal pulse oximetry and arterial blood gases at awakening were measured, in each patient, on two consecutive days. Results: The administration of 1 L more oxygen during the night resulted in improved parameters of nocturnal oxygenation (oxygen pulse oximetry saturation—SpO₂; percentage of sleep time spent at SpO₂ < 90%—CT90; PaO₂ at awakening). Nevertheless, such an increase in oxygen flow during the night was also associated with greater hypercapnia and acidosis (p < 0.05) the next morning. Conclusions: The increase of oxygen flow in severe COPD patients with established daytime hypercapnia improved nocturnal oxygenation but it also led to greater hypercapnia and respiratory acidosis at awakening in a considerable proportion of these patients.     

C反应蛋白在AECOPD的意义

目的探讨C反应蛋白（CRP）在慢性阻塞性肺疾病急性加重期（AECOPD）变化的意义及临床应用价值。方法检测152例AECOPD患者及88名COPD缓解期患者CRP、白细胞计数（WBC）、中性粒细胞计数百分比（N）、血沉（ESR）,疾培养及X线胸片,同时对152例AECOPD患者给予抗感染治疗,并比较这些指标治疗前后的变化。结果AECOPD患者血清CRP水平显著高于COPD稳定期患者（P〈0．01）,而在AECOPD患者死亡组血清CRP水平又显著高于存活组（P〈0．001）。结论CRP升高不仅可以提示COPD的急性加重,极度升高又提示COPD患者病情严重并且可能预后不良。CRP在AECOPD是一个良好的观察指标,可反映抗生素的即时疗效,比WBC,ESR更迅速且敏感。