CXC chemokine receptor-2 ligands are required for neutrophil-mediated host defense in experimental brain abscesses

We have developed a mouse brain abscess model by using Staphylococcus aureus, one of the main etiologic agents of brain abscesses in humans. Direct damage to the blood-brain barrier was observed from 24 h to 7 days after S. aureus exposure as demonstrated by the accumulation of serum IgG in the brain parenchyma. Evaluation of brain abscesses by immunohistochemistry and flow cytometry revealed a prominent neutrophil infiltrate. To address the importance of neutrophils in the early containment of S. aureus infection in the brain, mice were transiently depleted of neutrophils before implantation of bacteria-laden beads. Neutrophil-depleted animals consistently demonstrated more severe brain abscesses and higher CNS bacterial burdens compared with control animals. S. aureus led to the induction of numerous chemokines in the brain, including macrophage-inflammatory protein (MIP)-1alpha/CCL3, MIP-1beta/CCL4, MIP-2/CXCL1, monocyte chemoattractant protein-1/CCL2, and TCA-3/CCL1, within 6 h after bacterial exposure. These chemokines also were expressed by both primary cultures of neonatal mouse microglia and astrocytes exposed to heat-inactivated S. aureus in vitro. Because neutrophils constitute the majority of the cellular infiltrate in early brain abscess development, subsequent analysis focused on MIP-2 and KC/CXCL1, two neutrophil-attracting CXC chemokines. Both MIP-2 and KC protein levels were significantly elevated in the brain after S. aureus exposure. Neutrophil extravasation into the brain parenchyma was impaired in CXCR2 knockout mice and was associated with increased bacterial burdens. These studies demonstrate the importance of the CXCR2 ligands MIP-2 and KC and neutrophils in the acute host response to S. aureus in the brain.     

Bacterial clearance and survival are dependent on CXC chemokine receptor-2 ligands in a murine model of pulmonary Nocardia asteroides infection

Survival from murine pulmonary nocardiosis is highly dependent on CXC chemokine receptor-2 (CXCR2) ligand-mediated neutrophil chemotaxis and subsequent clearance of the infectious agent Nocardia asteroides. Intratracheal inoculation of N. asteroides rapidly up-regulated the CXC chemokines macrophage inflammatory protein-2 (MIP-2) and KC within 24 h, with levels remaining elevated through day 3 before returning to near baseline levels by day 7. Coinciding with elevated MIP-2 and KC were the rapid recruitment of neutrophils and clearance of the organism. Anti-Ly-6G Ab-mediated neutrophil depletion before bacterial challenge resulted in strikingly increased mortality to N. asteroides infection. The relative contribution of MIP-2 in neutrophil recruitment was examined by anti-MIP-2 Ab treatment before nocardial infection. MIP-2 neutralization had no detrimental effects on survival, neutrophil recruitment, or bacterial clearance, suggesting the usage of additional or alternative CXCR2-binding ligands. The importance of the CXC family of chemokines was determined by the administration of an anti-CXCR2 Ab capable of blocking ligand binding in vivo. Anti-CXCR2 treatment greatly increased mortality by preventing neutrophil migration into the lung. Paralleling this impaired neutrophil recruitment was a 100-fold increase in lung bacterial burden. Combined, these observations indicate a critical role for neutrophils and CXC chemokines during nocardial pneumonia. These data directly link CXCR2 ligands and neutrophil recruitment and lend further support to the concept of CXC chemokine redundancy. For infections highly dependent on neutrophils, such as nocardial pneumonia, this is of critical importance.     

Impaired pulmonary host defense in mice lacking expression of the CXC chemokine lungkine

Lungkine (CXCL15) is a novel CXC chemokine that is highly expressed in the adult mouse lung. To determine the biologic function of Lungkine, we generated Lungkine null mice by targeted gene disruption. These mice did not differ from wild-type mice in their hematocrits or in the relative number of cells in leukocyte populations of peripheral blood or other tissues, including lung and bone marrow. However, Lungkine null mice were more susceptible to Klebsiella pneumonia infection, with a decreased survival and increased lung bacterial burden compared with infected wild-type mice. Histologic analysis of the lung and assessment of leukocytes in the bronchioalveolar lavage revealed that neutrophil numbers were normal in the lung parenchyma, but reduced in the airspace. The production of other neutrophil chemoattractants in the Lungkine null mice did not differ from that in wild-type mice, and neutrophil migration into other tissues was normal. Taken together, these findings demonstrate that Lungkine is an important mediator of neutrophil migration from the lung parenchyma into the airspace.     

Macrophage inflammatory protein-1 alpha is a critical mediator of host defense against invasive pulmonary aspergillosis in neutropenic hosts

Invasive pulmonary aspergillosis is a devastating complication of immunosuppression that usually occurs in neutropenic patients. In this setting, augmentation of the antifungal activity of available immune cells may improve the outcome of the infection. Macrophage inflammatory protein-1 alpha (MIP-1 alpha) is a CC chemokine with potent chemotactic activity for various subsets of mononuclear leukocytes. We therefore tested the hypothesis that the influx of mononuclear cells into the lung in invasive pulmonary aspergillosis is in part mediated by MIP-1 alpha, and the manipulation of this ligand alters the outcome of the infection. We found that in both immunocompetent and neutropenic mice, MIP-1 alpha was induced in the lungs in response to intratracheal administration of Aspergillus fumigatus conidia. In neutrophil-depleted mice challenged with intratracheal conidia, there was evidence of invasive fungal pneumonia associated with a predominantly mononuclear leukocyte infiltrate. Ab-mediated depletion of MIP-1 alpha resulted in a 6-fold increase in mortality in neutropenic mice, which was associated with a 12-fold increase in lung fungal burden. Studies of single-cell suspensions of whole lungs revealed a 36% decrease in total lung leukocyte infiltration as a result of MIP-1 alpha neutralization. Flow cytometry on whole lung suspensions showed a 41% reduction in lung monocyte/macrophages as a result of MIP-1 alpha neutralization, but no difference in other lung leukocyte subsets. These studies indicate that MIP-1 alpha is a critical mediator of host defense against A. fumigatus in the setting of neutropenia and may be an important target in devising future therapeutic strategies against invasive aspergillosis.     

Dynamic optimization reveals alveolar epithelial cells as key mediators of host defense in invasive aspergillosis

Aspergillus fumigatus is an important human fungal pathogen and its conidia are constantly inhaled by humans. In immunocompromised individuals, conidia can grow out as hyphae that damage lung epithelium. The resulting invasive aspergillosis is associated with devastating mortality rates. Since infection is a race between the innate immune system and the outgrowth of A. fumigatus conidia, we use dynamic optimization to obtain insight into the recruitment and depletion of alveolar macrophages and neutrophils. Using this model, we obtain key insights into major determinants of infection outcome on host and pathogen side. On the pathogen side, we predict in silico and confirm in vitro that germination speed is an important virulence trait of fungal pathogens due to the vulnerability of conidia against host defense. On the host side, we found that epithelial cells, which have been underappreciated, play a role in fungal clearance and are potent mediators of cytokine release. Both predictions were confirmed by in vitro experiments on established cell lines as well as primary lung cells. Further, our model affirms the importance of neutrophils in invasive aspergillosis and underlines that the role of macrophages remains elusive. We expect that our model will contribute to improvement of treatment protocols by focusing on the critical components of immune response to fungi but also fungal virulence traits.     

Neutralization of the CXC chemokine, macrophage inflammatory protein-2, attenuates bleomycin-induced pulmonary fibrosis

Few studies have addressed the importance of vascular remodeling in the lung during the development of bleomycin-induced pulmonary fibrosis. For fibroplasia and deposition of extracellular matrix to occur, there must be a geometric increase in neovascularization. We hypothesized that net angiogenesis during the pathogenesis of fibroplasia and deposition of extracellular matrix during bleomycin-induced pulmonary fibrosis are dependent in part upon an overexpression of the angiogenic CXC chemokine, macrophage inflammatory protein-2 (MIP-2). To test this hypothesis, we measured MIP-2 by specific ELISA in whole lung homogenates in either bleomycin-treated or control CBA/J mice and correlated these levels with lung hydroxyproline. We found that lung tissue from mice treated with bleomycin, compared with that from saline-treated controls, demonstrated a significant increase in the presence of MIP-2 that was correlated to a greater angiogenic response and total lung hydroxyproline content. Neutralizing anti-MIP-2 Abs inhibited the angiogenic activity of day 16 bleomycin-treated lung specimens using an in vivo angiogenesis bioassay. Furthermore, when MIP-2 was depleted in vivo by passive immunization, bleomycin-induced pulmonary fibrosis was significantly reduced without a change in the presence of pulmonary neutrophils, fibroblast proliferation, or collagen gene expression. This was also paralleled by a reduction in angiogenesis. These results demonstrate that the angiogenic CXC chemokine, MIP-2, is an important factor that regulates angiogenesis/fibrosis in pulmonary fibrosis.     

A combination of in vitro techniques for efficient discovery of functional monoclonal antibodies against human CXC chemokine receptor-2 (CXCR2)

Background: Development of functional monoclonal antibodies against intractable GPCR targets.Results: Identification of structured peptides mimicking the ligand binding site, their use in panning to enrich for a population of binders, and the subsequent challenge of this population with receptor overexpressing cells leads to functional monoclonal antibodies.Conclusion: The combination of techniques provides a successful strategic approach for the development of functional monoclonal antibodies against CXCR2 in a relatively small campaign.Significance: The presented combination of techniques might be applicable for other, notoriously difficult, GPCR targets.Summary: The CXC chemokine receptor-2 (CXCR2) is a member of the large ‘family A’ of G-protein-coupled-receptors and is overexpressed in various types of cancer cells. CXCR2 is activated by binding of a number of ligands, including interleukin 8 (IL-8) and growth-related protein α (Gro-α). Monoclonal antibodies capable of blocking the ligand-receptor interaction are therefore of therapeutic interest; however, the development of biological active antibodies against highly structured GPCR proteins is challenging. Here we present a combination of techniques that improve the discovery of functional monoclonal antibodies against the native CXCR2 receptor.The IL-8 binding site of CXCR2 was identified by screening peptide libraries with the IL-8 ligand, and then reconstructed as soluble synthetic peptides. These peptides were used as antigens to probe an antibody fragment phage display library to obtain subpopulations binding to the IL-8 binding site of CXCR2. Further enrichment of the phage population was achieved by an additional selection round with CXCR2 overexpressing cells as a different antigen source. The scFvs from the CXCR2 specific phage clones were sequenced and converted into monoclonal antibodies. The obtained antibodies bound specifically to CXCR2 expressing cells and inhibited the IL-8 and Gro-α induced ß-arrestin recruitment with IC50 values of 0.3 and 0.2 nM, respectively, and were significantly more potent than the murine monoclonal antibodies (18 and 19 nM, respectively) obtained by the classical hybridoma technique, elicited with the same peptide antigen. According to epitope mapping studies, the antibody efficacy is largely defined by N-terminal epitopes comprising the IL-8 and Gro-α binding sites. The presented strategic combination of in vitro techniques, including the use of different antigen sources, is a powerful alternative for the development of functional monoclonal antibodies by the classical hybridoma technique, and might be applicable to other GPCR targets.     

A combination of in vitro techniques for efficient discovery of functional monoclonal antibodies against human CXC chemokine receptor-2 (CXCR2)

Background: Development of functional monoclonal antibodies against intractable GPCR targets.

Results: Identification of structured peptides mimicking the ligand binding site, their use in panning to enrich for a population of binders, and the subsequent challenge of this population with receptor overexpressing cells leads to functional monoclonal antibodies.

Conclusion: The combination of techniques provides a successful strategic approach for the development of functional monoclonal antibodies against CXCR2 in a relatively small campaign.

Significance: The presented combination of techniques might be applicable for other, notoriously difficult, GPCR targets.

Summary: The CXC chemokine receptor-2 (CXCR2) is a member of the large ‘family A’ of G-protein-coupled-receptors and is overexpressed in various types of cancer cells. CXCR2 is activated by binding of a number of ligands, including interleukin 8 (IL-8) and growth-related protein α (Gro-α). Monoclonal antibodies capable of blocking the ligand-receptor interaction are therefore of therapeutic interest; however, the development of biological active antibodies against highly structured GPCR proteins is challenging. Here we present a combination of techniques that improve the discovery of functional monoclonal antibodies against the native CXCR2 receptor.

The IL-8 binding site of CXCR2 was identified by screening peptide libraries with the IL-8 ligand, and then reconstructed as soluble synthetic peptides. These peptides were used as antigens to probe an antibody fragment phage display library to obtain subpopulations binding to the IL-8 binding site of CXCR2. Further enrichment of the phage population was achieved by an additional selection round with CXCR2 overexpressing cells as a different antigen source. The scFvs from the CXCR2 specific phage clones were sequenced and converted into monoclonal antibodies. The obtained antibodies bound specifically to CXCR2 expressing cells and inhibited the IL-8 and Gro-α induced ß-arrestin recruitment with IC50 values of 0.3 and 0.2 nM, respectively, and were significantly more potent than the murine monoclonal antibodies (18 and 19 nM, respectively) obtained by the classical hybridoma technique, elicited with the same peptide antigen. According to epitope mapping studies, the antibody efficacy is largely defined by N-terminal epitopes comprising the IL-8 and Gro-α binding sites. The presented strategic combination of in vitro techniques, including the use of different antigen sources, is a powerful alternative for the development of functional monoclonal antibodies by the classical hybridoma technique, and might be applicable to other GPCR targets.     

肺部侵袭性曲霉菌感染的诊断思路

侵袭性真菌感染(invasive fungal infection,IFI)是免疫低下患者常见的并发症之一,尤其在血液肿瘤患者和造血干细胞移植后患者中,IFI更是影响患者生存和生活质量的重要因素。随着氟康唑在高危人群中的广泛预防性应用,白念珠菌的感染较前已有所减少,而曲霉菌的感染比例则有所增[1]     

T cell vaccination in mice with invasive pulmonary aspergillosis

Aspergillus fumigatus, an opportunistic fungal pathogen, is responsible for multiple airway diseases of an allergic and a nonallergic nature. In a murine model of invasive pulmonary aspergillosis, resistance is associated with a decreased lung inflammatory pathology and the occurrence of an IL-12-dependent Th1-type reactivity that are both impaired by IL-4. In the present study we assess the ability of Aspergillus crude culture filtrate Ags and the recombinant allergen Asp f 2 to induce protective antifungal responses in mice with invasive pulmonary aspergillosis. Similar to what occurred upon nasal exposure to viable A. fumigatus conidia, treatment of immunocompetent mice with Aspergillus crude culture filtrate Ags resulted in the development of local and peripheral protective Th1 memory responses, mediated by Ag-specific CD4+ T cells producing IFN-gamma and IL-2 capable of conferring protection upon adoptive transfer to naive recipients. Protective Th1 responses could not be observed in mice deficient of IFN-gamma or IL-12 and did not occur in response to Asp f 2, which, on the contrary, elicited high level production of inhibitory IL-4. The results show that Ags of Aspergillus exist with the ability to induce both Th1- and Th2-type reactivity during infection, a finding that suggests a possible mechanism through which potentially protective immune responses are inhibited in mice with the infection. However, the occurrence of Th1-mediated resistance upon vaccination with Aspergillus crude culture filtrate Ags, suggests the existence of fungal Ags useful as a candidate vaccine against invasive pulmonary aspergillosis.     

Toll-like receptors are temporally involved in host defense

Toll-like receptors (TLRs) are evolutionarily conserved proteins that recognize microbial molecules and initiate host defense. To investigate how TLRs work together to fight infections, we tested the role of TLRs in host defense against the Gram-negative bacterial pathogen, Salmonella. We show that TLR4 is critical for early cytokine production and killing of bacteria by murine macrophages. Interestingly, later on, TLR2, but not TLR4, is required for macrophage responses. Myeloid differentiation factor 88, an adaptor protein directly downstream of TLRs, is required for both early and late responses. TLR4, TLR2, and myeloid differentiation factor 88 are involved in murine host defense against Salmonella in vivo, which correlates with the defects in host defense observed in vitro. We propose a model where the sequential activation of TLRs tailors the immune response to different microbes.     

The role of CXC chemokine ligand 4/CXC chemokine receptor 3-B in breast cancer progression

Chemokines and their receptors participate in the development of cancers by enhancing tumor cell proliferation, angiogenesis, invasion, metastasis and penetration of tumor immune cells. It remains unclear whether CXC chemokine ligand 4 (CXCL4)/CXC chemokine receptor 3-B (CXCR3-B) can be used as an independent molecular marker for establishing prognosis for breast cancer patients. We evaluated CXCL4 and CXCR3-B expression in 114 breast cancer tissues and 30 matched noncancerous tissues using immunohistochemistry and western blot, and determined the correlation between their expression and clinicopathologic findings. We observed that breast cancer tissues express CXCL4 strongly and CXCR3-B weakly compared to noncancerous tissues. Strong CXCL4 expression was detected in 94.7% and weak CXCR3-B expression was detected in 78.9% of the tissues. Therefore, CXCL4/CXCR3-B might play a crucial role in breast cancer progression. We found no significant correlation between CXCL4 and age, tumor stage, tumor grade or TNM stage. CXCR3-B was associated significantly with tumor grade. Moreover, the Chi-square test of association showed that the expression of CXCL4/CXCR3-B might be an independent prognostic marker for breast cancer. Therefore, we suggest that CXCR3-B is an indicator of poor prognosis and may also be a chemotherapeutic target.     

128例侵袭性肺曲霉病临床疗效分析

目的 通过回顾性病例分析,加强对侵袭性肺曲霉病的认识,重视对侵袭性肺曲霉病的分级诊断,从而制定合理治疗方案.方法 对四川大学华西医院201 1年1 ～12月128例诊断为侵袭性肺曲霉病患者的临床资料进行回顾性分析,重点分析基础疾病、临床表现、影像学表现、诊断及治疗方法.结果 确诊17例;依据宿主危险因素、临床表现及反复痰培养阳性拟诊111例.128例患者均伴有基础疾病.治疗方案包括伏立康唑（86例）,两性霉素B脂质体（1例）,伊曲康唑（12例）,米卡芬净（3例）,卡泊芬净（3例）,两性霉素B去氧胆酸盐（10例）,伏立康唑联合米卡芬净（1例）.治愈或好转81例（63.28％）,自动出院24例（18.75％）,死亡23例（17.97％）.结论 侵袭性肺曲霉病患者多数伴有多种感染危险因素,应当重视侵袭性肺曲霉病的分级诊断,据不同情况给予适当抗真菌治疗,以降低患者病死率.     

The ligands of CXC chemokine receptor 3, I-TAC, Mig, and IP10, are natural antagonists for CCR3

Th1 and Th2 lymphocytes express a different repertoire of chemokine receptors (CCRs). CXCR3, the receptor for I-TAC (interferon-inducible T cell alpha-chemoattractant), Mig (monokine induced by gamma-interferon), and IP10 (interferon-inducible protein 10), is expressed preferentially on Th1 cells, whereas CCR3, the receptor for eotaxin and several other CC chemokines, is characteristic of Th2 cells. While studying responses that are mediated by these two receptors, we found that the agonists for CXCR3 act as antagonists for CCR3. I-TAC, Mig, and IP10 compete for the binding of eotaxin to CCR3-bearing cells and inhibit migration and Ca(2+) changes induced in such cells by stimulation with eotaxin, eotaxin-2, MCP-2 (monocyte chemottractant protein-2), MCP-3, MCP-4, and RANTES (regulated on activation normal T cell expressed and secreted). A hybrid chemokine generated by substituting the first eight NH(2)-terminal residues of eotaxin with those of I-TAC bound CCR3 with higher affinity than eotaxin or I-TAC (3- and 10-fold, respectively). The hybrid was 5-fold more potent than I-TAC as an inhibitor of eotaxin activity and was effective at concentrations as low as 5 nm. None of the antagonists described induced the internalization of CCR3, indicating that they lack agonistic effects and thus qualify as pure antagonists. These results suggest that chemokines that attract Th1 cells via CXCR3 can concomitantly block the migration of Th2 cells in response to CCR3 ligands, thus enhancing the polarization of T cell recruitment.     

Increase of CXC chemokine CXCL10 and CC chemokine CCL2 serum levels in normal ageing

No study has evaluated contemporaneously serum CXC and CC chemokines changes in normal ageing. Serum levels of CXCL10 (sCXCL10) (CXC) and CCL2 (sCCL2) (CC) prototype chemokines have been measured in 164 healthy subjects, from 10 to 79 years of age (82 males/82 females). By simple regression analysis, sCXCL10 and sCCL2 were significantly related with increasing age (r=0.32, p<0.001; r=0.31, p<0.0001, respectively), and with each other (r=0.30, p=0.0004). In a multiple linear regression model, only age and sCCL2 were significantly related to sCXCL10 levels (p<0.001); age and sCXCL10 were significantly related to sCCL2 levels (p<0.001). Subjects with high sCXCL10 levels (>150 pg/ml) were not significantly associated with those with high sCCL2 levels (>559 pg/ml). This study, performed in healthy subjects on an age gradient, demonstrates an increase of sCXCL10 and sCCL2 with advancing age; the differential increase of sCXCL10 or sCCL2 may reflect a general shift towards Th1 or Th2 cytokines pattern, respectively.     

Essential role for the p40 subunit of interleukin-12 in neutrophil-mediated early host defense against pulmonary infection with Streptococcus pneumoniae: involvement of interferon-gamma

Interleukin (IL)-12 is a critical cytokine in the T helper (Th)1 response and host defense against intracellular microorganisms, while its role in host resistance to extracellular bacteria remains elusive. In the present study, we elucidated the role of IL-12 in the early-phase host defense against acute pulmonary infection with Streptococcus pneumoniae, a typical extracellular bacterium, using IL-12p40 gene-disrupted (IL-12p40KO) mice. IL-12p40KO mice were highly susceptible to S. pneumoniae infection, as indicated by the shortened survival time, which was completely restored by the replacement therapy with recombinant (r) IL-12, and increased bacterial counts in the lung. In these mice, recruitment of neutrophils in the lung was significantly attenuated when compared to that in wild-type (WT) mice, which correlated well with the reduced production of macrophage inflammatory protein (MIP-2) and tumor necrosis factor (TNF)-alpha in the infected tissues at the early phase of infection. In vitro synthesis of both cytokines by S. pneumoniae-stimulated lung leukocytes was significantly lower in IL-12p40KO mice than in WT mice, and addition of rIL-12 or interferon (IFN)-gamma restored the reduced production of MIP-2 and TNF-alpha in IL-12p40KO mice. Neutralizing anti-IFN-gamma monoclonal antibody (mAb) significantly decreased the effect of rIL-12. Anti-IFN-gamma mAb shortened the survival time of infected mice and reduced the recruitment of neutrophils and production of MIP-2 and TNF-alpha in the lungs. Our results indicated that IL-12p40 plays a critical role in the early-phase host defense against S. pneumoniae infection by promoting the recruitment of neutrophils to the infected tissues.     

Bronchoalveolar lavage in an immunosuppressed rabbit model of invasive pulmonary aspergillosis

A rabbit model of invasive aspergillosis has been used to investigate the pathogenesis of Aspergillus infection in the immunosuppressed host. The animals received hydrocortisone daily and a single dose of cyclophosphamide 2 days prior to intratracheal instillation of conidia from Aspergillus fumigatus. Bronchoalveolar lavage (BAL) was performed in 3 infected and 2 control saline treated animals sacrificed on days 1, 2, 4, 7 and 10 following inoculation. Infective load within the lung was quantified using an assay for chitin which is an important component of fungal cell walls (in particular the hyphal cell wall) and is not present in vertebrate tissue. The total BAL white cell count did not discriminate between infected and saline treated animals and Aspergillus was cultured from one lavage specimen only. Infected animals developed a marked neutrophil alveolitis by day 2 in contrast to a near total absence of neutrophils in the lavages of the control animals. Phagocytosis of conidia by alveolar macrophages was prominent but did not prevent progressive infection as confirmed by measurement of lung chitin. This pattern of cellular response within the alveolar airspace reflects the complex nature of the response to Aspergillus infection in the immunosuppressed host.     

Early diagnosis of invasive pulmonary aspergillosis in a young immunocompetent patient

A 22-year-old insulin-dependent diabetic male was admitted for diabetic ketoacidosis. He developed hospital-acquired pneumonia (HAP) for which empirical antibiotic and antifungal therapy was started on the ward. On day 6, clinical and laboratory findings worsened, and bronchoalveolar lavage (BAL) was performed. Serum real time-polymerase chain reaction (RT-PCR) indicated invasive pulmonary aspergillosis (IPA) and led to antifungal therapy being initiated 48 hours before the results of the BAL culture were available. Despite early appropriate antifungal therapy, however, the patient died on day 22 while being supported by venovenous extracorporeal membrane oxygenation.     

The chemokine decoy receptor M3 blocks CC chemokine ligand 2 and CXC chemokine ligand 13 function in vivo

Chemokines and their receptors play a key role in immune homeostasis regulating leukocyte migration, differentiation, and function. Viruses have acquired and optimized molecules that interact with the chemokine system. These virus-encoded molecules promote cell entry, facilitate dissemination of infected cells, and enable the virus to evade the immune response. One such molecule in the murine gammaherpesvirus 68 genome is the M3 gene, which encodes a secreted 44-kDa protein that binds with high affinity to certain murine and human chemokines and blocks chemokine signaling in vitro. To test the hypothesis that M3 directly interferes with diverse chemokines in vivo, we examined the interaction of M3 with CCL2 and CXCL13 expressed in the pancreas of transgenic mice. CCL2 expression in the pancreas promoted recruitment of monocytes and dendritic cells; CXCL13 promoted recruitment of B and T lymphocytes. Coexpression of M3 in the pancreas blocked cellular recruitment induced by both CCL2 and CXCL13. These results define M3 as multichemokine blocker and demonstrate its use as a powerful tool to analyze chemokine biology.     

胞浆识别受体NODs及其信号通路在侵袭性肺曲霉病中的作用

【目的】研究天然免疫系统中胞浆识别受体NODs及其信号通路在小鼠侵袭性肺曲霉病(IPA)中的作用。【方法】小鼠随机分为正常对照组、正常+接种烟曲霉菌组(正常感染组)和免疫抑制+接种烟曲霉菌组(IPA组),经鼻吸入烟曲霉孢子后在不同时相点处死小鼠,无菌取肺组织分别进行病理切片,烟曲霉菌落计数,RT-PCR法、Western blot法动态检测小鼠感染烟曲霉菌过程中肺组织NOD1、NOD2、RIP2 mRNA表达,促炎细胞因子TNF-α含量的变化规律。【结果】鼻吸入烟曲霉菌后72 h时,IPA组肺组织出现严重炎症反应,并有大量的菌丝生成,同时各时相点的烟曲霉菌负荷均高于正常感染组;与正常感染组比较,IPA组NOD1、RIP2 mRNA持续低表达,而NOD2 mRNA则在感染最早期(24 h)异常高表达,而在随后的感染过程中一直处于低表达状态;正常小鼠感染烟曲霉菌后,肺组织中促炎细胞因子TNF-α在感染前期皆呈高表达,且最高表达量均出现在48 h或72 h,之后下降并恢复至正常水平。而IPA小鼠促炎症细胞因子TNF-α缓慢且低水平释放。【结论】NOD1、RIP2的表达受到长期抑制,NOD2在感染最早期的过度激活以及随后的抑制表达,引起促炎细胞因子低表达,可能导致了侵袭性肺曲霉的发生发展。