MAPK-Activated Protein Kinase 2 Contributes to Clostridium difficile-Associated Inflammation

Clostridium difficile infection (CDI) results in toxin-induced epithelial injury and marked intestinal inflammation. Fecal markers of intestinal inflammation correlate with CDI disease severity, but regulation of the inflammatory response is poorly understood. Previous studies demonstrated that C. difficile toxin TcdA activates p38 kinase in tissue culture cells and mouse ilium, resulting in interleukin-8 (IL-8) release. Here, we investigated the role of phosphorylated mitogen-activated protein kinase (MAPK)-activated protein kinase (MK2 kinase, pMK2), a key mediator of p38-dependent inflammation, in CDI. Exposure of cultured intestinal epithelial cells to the C. difficile toxins TcdA and TcdB resulted in p38-dependent MK2 activation. Toxin-induced IL-8 and GROα release required MK2 activity. We found that p38 and MK2 are activated in response to other actin-disrupting agents, suggesting that toxin-induced cytoskeleton disruption is the trigger for kinase-dependent cytokine response. Phosphorylated MK2 was detected in the intestines of C. difficile-infected hamsters and mice, demonstrating for the first time that the pathway is activated in infected animals. Furthermore, we found that elevated pMK2 correlated with the presence of toxigenic C. difficile among 100 patient stool samples submitted for C. difficile testing. In conclusion, we find that MK2 kinase is activated by TcdA and TcdB and regulates the expression of proinflammatory cytokines. Activation of p38-MK2 in infected animals and humans suggests that this pathway is a key driver of intestinal inflammation in patients with CDI.     

Synoviocyte-Derived Angiopoietin-Like Protein 2 Contributes to Synovial Chronic Inflammation in Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by symmetrical polyarticular synovitis of the diarthrodial joints. Several proinflammatory cytokines derived from both infiltrating inflammatory cells and activated resident cells within the RA joint play a fundamental role in the processes that cause inflammation. However, anticytokine treatment is beneficial but not curative, the effects are only partial, and nonresponses are common. Therefore, an effort has been made to identify other key regulators of inflammation in articular structures to develop new therapies to suppress synovial inflammation and joint destruction in RA. Adipose tissue-derived angiopoietin-like protein 2 (Angptl2) activates an inflammatory cascade in endothelial cells and induces chemotaxis of monocytes/macrophages in obesity, resulting in initiation and propagation of inflammation within adipose tissues and obesity-related metabolic diseases. Angptl2 mRNA and protein are abundantly expressed in hyperplastic rheumatoid synovium of RA patients, especially in fibroblast-like and macrophage-like synoviocytes, but not in B and T lymphocytes. Angptl2 concentration in joints of RA patients was also significantly increased in comparison with patients with osteoarthritis, which in comparison with RA represents a significantly lower inflammatory grade form of arthritis. Notably, Angptl2 promoted increased chemotactic activities of CD14⁺CD16⁻ monocytes from synovial fluid of RA patients. Therefore, Angptl2 acts as an important rheumatoid synovium-derived inflammatory mediator in RA pathogenesis.Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by symmetrical polyarticular synovitis of the diarthrodial joints. The RA synovial membrane contains activated B and T lymphocytes, plasma cells, mast cells, and significantly activated monocytes/macrophages. The synovium is normally a relatively acellular structure with a delicate intimal lining, thus these cells are recruited via an intense neovascularization process with associated lymphangiogenesis. Hyperplasia of the intimal lining results from macrophage-like and fibroblast-like synoviocytes. These resident and infiltrating cells within the RA joint could be a source of proinflammatory cytokines that activate inflammatory pathways in a paracrine or autocrine fashion and play a fundamental role in processes underlying inflammation, articular destruction, and comorbidities associated with RA.^1,2,3,4,5,6 In fact, numerous cytokines, such as interleukin (IL)-1, IL-6, IL-15, IL-18, and tumor-necrosis factor (TNF)-α, as well as various chemokines, are active within the synovium and synovial fluid in joints of RA patients.^2,7,8 Continuous anticytokine treatment, such as through use of TNF-α and IL-1β inhibitors, is required for long-term control, and discontinuation of treatment leads to disease flare-up, indicating the importance of cytokine-related inflammation in pathogenesis of RA.² Furthermore, although such anticytokine treatment is beneficial,^{9,10,11,12,13,14} it is not curative, its effects are partial, and nonresponses are common.^2,15 These findings indicate that the mechanism of inflammation in the RA joint is more complex than previously thought, thus suggesting that new factors and mechanisms are operating that could serve as novel therapeutic targets for RA.The Angptl (Angiopoietin-like protein) family was identified as a group of proteins possessing structural similarity to angiopoietin, which contains an N-terminal coiled-coil domain functioning in oligomerization and a C-terminal fibrinogen-like domain serving as a receptor binding site.^16,17 Although Angptls were predicted to function as ligands for the angiopoietin-receptor; Tie-2 or its family member; Tie-1, they do not bind to either, strongly suggesting biological functions different from those of angiopoietins. More recently, we reported that Angptl2, a member of the Angptl family, is expressed in a variety of tissues, especially in obese adipose tissues.¹⁸ Angptl2 expression has been shown to increase by hypoxia and endoplasmic reticulum stress, both of which are commonly observed in pathological conditions. We also showed that Angptl2 signaling via integrins activated an inflammatory cascade in endothelial cells and induced chemotaxis of monocytes/macrophages. Constitutive Angptl2 activation in vivo induced inflammation of the vasculature characterized by abundant attachment of leukocytes to vessel walls and increased permeability. These findings suggest that adipocyte-derived Angptl2 acts as a key chronic inflammatory mediator in obesity, resulting in obesity-related metabolic diseases. These findings, plus the fact that its expression is not restricted to adipose tissues, suggest a possible role of Angptl2 in other chronic inflammatory diseases.The current study showed that Angptl2 mRNA and protein are abundantly expressed in the hyperplastic synoviocytes from RA patients. An in vitro culture analysis revealed that the synoviocytes from RA patients secrete Angptl2. Indeed, the concentration of Angptl2 in RA synovial fluid is significantly higher than that seen in osteoarthritis (OA), which is a lower inflammatory grade arthritis than RA. Angptl2 increases the chemotactic activities of monocytes from RA synovial fluid. Taken together, these findings establish Angptl2 as a hyperplastic rheumatoid synovium-derived inflammatory mediator in RA joints.     

Trillin Reduces Liver Chronic Inflammation and Fibrosis in Carbon Tetrachloride (CCl4) Induced Liver Injury in Mice

Trillin is an active ingredient isolated from Dioscorea nipponica Makino. This study investigated the anti-inflammatory and anti-fibrosis effects of trillin on CCl₄-induced hepatotoxicity in C57BL/6 mice. Chronic inflammation and fibrosis were induced by intraperitoneal administration of CCl₄ 0.5 μL/g of body weight twice a week for 6 weeks. Trillin (50 mg/kg, 100 mg/kg) was administered by gavage for 12 days before finishing the CCl₄ induction. Aspartate amino-transferase (AST) and glutamic-pyruvic transaminase (ALT) in serum were determined by AST and ALT kits. Superoxidase dismutase (SOD) activity and malondialdehyde (MDA) levels in serum were assayed by SOD and MDA kits. Meanwhile, the levels of inflammatory mediators including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β) in serum were detected by enzyme-linked immunosorbent assay (ELISA) method. Pathological changes were observed by hematoxylin-eosin (HE) staining. The proteins of the NF-κB pathway and the TGF-β/Smad pathway were measured by western blot. The trillin-treated group exhibited reduced AST, ALT, MDA, IL-6, TNF-α, and IL-1β, and increased SOD. Histological analyses of the trillin-treated group exhibited reduced inflammatory process and prevented liver fibrosis. Western blot analyses of the trillin-treated group showed reduced NF-κB pathway and TGF-β/Smad pathway. Significance: Based on the results of the present study, trillin can be used as a potential anti-inflammatory drug for chronic hepatic inflammation.     

Mixed Lineage Kinase 3 deficiency delays viral clearance in the lung and is associated with diminished influenza-induced cytopathic effect in infected cells

Influenza virus leads to acute respiratory disease resulting in seasonal epidemics and periodic pandemics. Little is known about the signaling events that regulate host defense to influenza. One particular pathway, the c-Jun amino-terminal kinase (JNK) cascade is activated following influenza infection and blocking JNK leads to enhanced viral replication. We hypothesize that Mixed Lineage Kinase 3 (MLK3), an upstream regulator of JNK, is involved in the host response to influenza. To test this, wild-type and MLK3−/− mice were infected with pathogenic strain of influenza A virus, A/PR/8/34 (PR8). Although, cellular and humoral immune responses were similar between wild-type and MLK3−/− hosts, the viral load in the lungs was comparatively higher in MLK3−/− mice at day 8 post-infection. Consistent with this, MLK3−/− murine lung fibroblast and epithelial cells had prolonged survival and increased virion production following infection compared to wild-type. These findings support a role for MLK3 in viral production during influenza infection.     

Angiogenesis in Mice with Chronic Airway Inflammation : Strain-Dependent Differences

Chronic inflammation is associated with blood vessel proliferation and enlargement and changes in vessel phenotype. We sought to determine whether these changes represent different types of angiogenesis and whether they are stimulus dependent. Chronic airway inflammation, produced by infection with Mycoplasma pulmonis, was compared in strains of mice known to be resistant (C57BL/6) or susceptible (C3H). Tracheal vascularity, assessed in whole mounts after Lycopersicon esculentum lectin staining, increased in both strains at 1, 2, 4, and 8 weeks after infection, but the type of vascular remodeling was different. The number of vessels doubled in tracheas of C57BL/6 mice, with corresponding increases of capillaries and venules. In contrast, neither the number nor the length of vessels changed in C3H mice. Instead, vessel diameter and endothelial cell number doubled, and the proportion of venules doubled with a corresponding decrease of capillaries. Although the infection had no effect on baseline plasma leakage, in both strains it potentiated the leakage produced by substance P. We conclude that the same stimulus can result in blood vessel proliferation or enlargement, depending on the host response. Endothelial cells proliferate in both cases, but in one case new capillaries form whereas in the other capillaries convert to venules.     

Second-Generation Tyrosine Kinase Inhibitors in First-Line Treatment of Chronic Myeloid Leukaemia (CML)

Tyrosine kinase inhibitors (TKIs) have contributed to marked improvements in survival in patients with chronic myeloid leukaemia (CML). This article discusses the place of the second-generation TKIs dasatinib and nilotinib in the first-line treatment of CML and is based on published literature. The new agents are more potent and effective than imatinib. Data from pivotal clinical trials indicate that response to dasatinib and nilotinib is greater and more rapid than that to imatinib, resulting in a higher probability of patients achieving an optimal response to treatment. Differences between the newer agents with respect to patient groups for whom caution is advised, drug interaction potential, haematological toxicity, pulmonary toxicity, changes in the immune system and effects on laboratory parameters are discussed. With similar levels of efficacy, the choice of second-generation agents should be guided by the characteristics of the individual patient and the most suitable dosing regimen.     

前列腺素E1对肾缺血再灌注损伤大鼠内皮的Rho/Rho激酶信号通路的影响

探讨前列腺素E1对肾缺血再灌注损伤大鼠内皮细胞Rho/Rho激酶信号通路的影响.建立大鼠IRI模型,将健康Wistar大鼠75只随机平均分为假手术组、模型组、治疗组,每组各25只;并按术后观察时间分为6、12、24、48、72 h5个不同时段亚组,每组各5只.检测各组血清肌酐(Scr)、血管内皮生长因子(Vascula...     

FMS-Like Tyrosine Kinase 3 Ligand Treatment of Mice Aggravates Acute Lung Injury in Response to Streptococcus pneumoniae: Role of Pneumolysin

FMS-like tyrosine kinase-3 ligand (Flt3L) is a dendritic cell (DC) growth and differentiation factor with potential in antitumor therapies and antibacterial immunization strategies. However, the effect of systemic Flt3L treatment on lung-protective immunity against bacterial infection is incompletely defined. Here, we examined the impact of deficient (in Flt3L knockout [KO] mice), normal (in wild-type [WT] mice), or increased Flt3L availability (in WT mice pretreated with Flt3L for 3, 5, or 7 days) on lung DC subset profiles and lung-protective immunity against the major lung-tropic pathogen, Streptococcus pneumoniae. Although in Flt3L-deficient mice the numbers of DCs positive for CD11b (CD11b^pos DCs) and for CD103 (CD103^pos DCs) were diminished, lung permeability, a marker of injury, was unaltered in response to S. pneumoniae. In contrast, WT mice pretreated with Flt3L particularly responded with increased numbers of CD11b^pos DCs and with less pronounced numbers of CD103^pos DCs and impaired bacterial clearance and with increased lung permeability following S. pneumoniae challenge. Notably, infection of Flt3L-pretreated mice with S. pneumoniae lacking the pore-forming toxin, pneumolysin (PLY), resulted in substantially less lung CD11b^pos DCs activation and reduced lung permeability. Collectively, this study establishes that Flt3L treatment enhances the accumulation of proinflammatory activated lung CD11b^pos DCs which contribute to acute lung injury in response to PLY released by S. pneumoniae.     

吗啡长时程作用对小鼠脑组织蛋白激酶A及C活性的影响

本文通过复制小鼠吗啡耐受依赖模型，观察了吗啡长时程作用对小鼠脑组织ＰＫＡ及ＰＫＣ活性的影响。结果发现：（１）吗啡耐受依赖小鼠纹状体、海马、大脑皮层神经细胞胞浆ＰＫＡ活性显著升高，而小脑胞浆及以上各部位膜相ＰＫＡ活性变化不明显；（２）不同脑组织中ＰＫＣ活性变化不同，吗啡耐受依赖小鼠大脑皮层及小脑胞浆ＰＫＣ活性明显升高，在纹状体胞浆则显著下降，但纹状体膜相ＰＫＣ活性却显著增加，海马及小脑膜相ＰＫＣ活性则明显降低；（３）纳洛酮可拮抗吗啡引起的上述变化。结果提示：一些脑组织胞浆ＰＫＡ活性的升高、ＰＫＣ活性的变化以及可能存在的ＰＫＣ于胞浆和膜相之间的移位可能是吗啡耐受依赖的重要生化基础，且此变化可能由阿片受体所介导。     

Multipotent Mesenchymal Stromal Cells in Patients with Chronic Myeloid Leukemia before Discontinuation of Tyrosine Kinase Inhibitors

Bulletin of Experimental Biology and Medicine - We studied the influence of vascular prostheses made of polytetrafluoroethylene and polyethylene terephthalate on the proliferation, migration, and...     

Plasmacytoid Dendritic Cells Control Lung Inflammation and Monocyte Recruitment in Indirect Acute Lung Injury in Mice

Indirect acute lung injury (ALI, not caused by a direct insult to the lung) represents the first organ dysfunction in trauma patients, with nonpulmonary sepsis being the most common cause of indirect ALI. Dendritic cells (DCs) are thought to participate in a number of inflammatory lung diseases; however, their role in indirect ALI is currently not established. Using a clinically relevant model of indirect ALI induced in mice by hemorrhagic shock followed 24 hours later by polymicrobial septic challenge, we report that mature DC numbers were markedly increased in the lung during indirect ALI. DC depletion induced a significant increase in indirect ALI severity, which was associated with enhanced lung and plasma proinflammatory cytokine concentration and recruitment of proinflammatory CD115⁺ monocytes in response to increased lung monocyte chemotactic protein-1 production. Among the different DC subpopulations, plasmacytoid DCs, which were induced and activated in the lung during indirect ALI, were responsible for this effect because their specific depletion reproduced the observations made in DC-depleted mice. As the recruitment of monocytes to the lung plays a central deleterious role in the pathophysiology of indirect ALI, our data therefore position plasmacytoid DCs as important regulators of acute lung inflammation.Acute lung injury (ALI) and its most severe form, the acute respiratory distress syndrome (ARDS), are the two parts of a clinical syndrome defined by acute hypoxemic respiratory failure, bilateral pulmonary infiltrate attributable to edema, and normal cardiac filling pressures.¹Of those seriously injured trauma victims who survive the first hours immediately after injury, upwards of 50% develop some forms of multiple organ failure. In this respect, ALI is reported to be one of the most common forms of organ dysfunction in these individuals.^1,2 Every year, ALI and ARDS are thus the cause of more than 74,000 deaths in the United States.²This syndrome can be categorized into direct (pulmonary) and indirect (nonpulmonary) ALI. Epidemiologically, direct ALI accounts for 57% of all cases and is caused mainly by pneumonia, aspiration, and lung trauma. Indirect ALI accounts for the residual 43% with nonpulmonary sepsis being the most frequent underlying disease.³ Among all causes, sepsis is associated with the highest risk of progression to ALI (approximately 40%) and sepsis-associated ARDS carries the highest mortality rates from ARDS.^1,2,4 Importantly, patients developing ALI after nonpulmonary sepsis (indirect ALI) present with a higher mortality rate than patients with pulmonary sepsis (direct ALI).⁵Despite almost 35 years of intense investigation, the fundamental mechanisms that initiate and propagate lung injury have not yet been defined completely.¹ In particular, mechanisms leading to direct versus indirect ALI may be different, with pulmonary infections causing lung injury directly via the pathogen and host response versus nonpulmonary infections causing lung injury indirectly via systemic inflammation.¹ Moreover, findings from a number of studies suggest that “priming” of different cell types occurs and appears to play significant roles in mediating the increased inflammation associated with this injury.^1,6 However, despite a good understanding of the process that initiates and promotes host inflammation, little is known about the host immune cells that are responsible for the inhibition of the inflammatory response.Dendritic cells (DCs), both myeloid (mDCs) and plasmacytoid (pDCs), exist in the lung in relatively small numbers.^7,8 In this location, they are ideally positioned to play a central role in the immune response during infection/inflammation.^7,8 Indeed, a role for DCs has been shown in a number of lung inflammatory diseases in human (asthma, chronic obstructive pulmonary disease, lung cancer, or transplant rejection).⁷ Moreover, during ongoing inflammation, DCs migrate to the lung where they not only maintain and enhance local immune response, but also regulate this response.^7,9 Therefore, we hypothesized that DCs have a role in the pathophysiology of indirect ALI. We investigated this using a clinically relevant model of indirect ALI induced in mice by a hemorrhagic shock followed 24 hours later by a polymicrobial septic challenge.