Elevated levels of thymus and activation-regulated chemokine (TARC) in pleural effusion samples from patients infested with Paragonimus westermani

To investigate the pathogenic mechanisms of eosinophilic pleural effusion in patients with paragonimiasis, we measured the levels of various chemokines including thymus and activation-regulated chemokine (TARC), eotaxin, RANTES and IL-8 in pleural effusion samples. Samples were obtained from 11 patients with Paragonimus westermani infection, six patients with pleural transudate, eight with tuberculous pleurisy and five with empyema. High percentages of eosinophils were detected in pleural fluid (range 9-100%, median 81%) of patients with paragonimiasis. TARC concentrations in pleural effusions of paragonimiasis were markedly higher than those of other groups. Eotaxin levels were also higher in pleural effusions of paragonimiasis patients, although significant difference was noted only against transudate samples. There was a significant correlation between TARC concentrations and percentages of eosinophils, and between TARC and eotaxin concentrations in pleural effusion. There were also significant correlations between TARC concentration and the titre of anti-P. westermani IgG and between eotaxin concentration and the titre of anti-P. westermani IgG. Our findings suggest that TARC contributes to the pathogenesis of eosinophilic pleural effusion in paragonimiasis.     

Granulocyte-macrophage colony-stimulating factor (GM-CSF) but not granulocyte colony-stimulating factor (G-CSF) induces plasma membrane expression of proteinase 3 (PR3) on neutrophils in vitro

The theoretical risk of triggering vasculitis resulting from administration of G-CSF and GM-CSF to patients with anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitides (AAV), such as Wegener's granulomatosis (WG), who develop agranulocytosis due to cytotoxic therapy, is unknown. Since there is strong evidence that activation of polymorphonuclear neutrophils (PMN) induced by binding of ANCA to PR3 or myeloperoxidase (MPO) expressed on their plasma membrane is involved in the pathogenesis of systemic vasculitides (SV), we studied the surface expression of PR3 and MPO on PMN from healthy donors in response to G-CSF and GM-CSF in vitro by flow cytometric analysis. Increasing doses of G-CSF did not alter PR3 expression on either untreated or tumour necrosis factor-alpha (TNF-alpha)-primed donor PMN significantly. In contrast, GM-CSF significantly increased PR3 membrane expression on both intact PMN and neutrophils primed with TNF-alpha. MPO expression was not significantly altered by either G-CSF or GM-CSF. In summary, these data demonstrate that GM-CSF, but not G-CSF, induces plasma membrane expression of PR3 on PMN in vitro. Since in AAV accessibility of the antigen (PR3 or MPO) to the antibody (ANCA) on the plasma membrane of PMN is thought to be essential for neutrophil activation by ANCA, the results of the present study suggest that administration of GM-CSF to patients with WG with neutropenia implies a definite theoretical risk of deterioration of vasculitis via this mechanism.     

IL-32 up-regulation is associated with inflammatory cytokine production in allergic rhinitis

IL-32 is a described pro-inflammatory cytokine produced by T lymphocytes, natural killer cells, monocytes, and epithelial cells. However, the specific mechanism of IL-32 on allergic rhinitis (AR) has not been elucidated. Here, we report a significant increase of IL-32 protein and mRNA in the nasal mucosa of AR patients. In addition, in nasal mucosa tissue from AR patients, the level of IL-32 production correlated with inflammation, IL-1β, IL-18, and granulocyte-macrophage colony-stimulating factor (GM-CSF). In an AR animal model, IL-32 significantly increased IgE and inflammatory cytokine levels. IL-32 expression was induced by recombinant human GM-CSF via activation of caspase-1 in eosinophils. In addition, depletion of IL-32 prevents the production of inflammatory cytokines in eosinophils. In conclusion, IL-32 is an important cytokine involved in the inflammation of AR. The regulation of IL-32 expression may form the basis of a new strategy for the treatment of AR.     

Correlation of serum IL-13 and IL-5 levels with clinical response to Glatiramer acetate in patients with multiple sclerosis

Glatiramer acetate (GA) is effective in the treatment of Multiple Sclerosis (MS) presumably by the induction of an immunoregulatory T-cell response. We have previously shown that GA directly induces the Th2 cytokines IL-13 and IL-5 in T-cells in vitro. In the present study we compared the in vitro response to GA in healthy controls, untreated and GA-treated MS patients and tested whether the induction of IL-13 and IL-5 secretion is also detectable in the serum of 25 MS patients treated with GA. Patients were grouped into clinical responders and nonresponders in order to determine a possible correlation with the immunological response. As a result we found a significant increase of IL-13 in the serum of clinical GA-responders whereas IL-13 was not detectable in controls, untreated MS (P < 0.001) and nonresponders (P = 0.015). Similarly, GA-treatment increased serum levels of IL-5 (P = 0.001). The correlation of serum IL-5 and clinical response was also significant (P = 0.039), however, there was an overlap between the different groups. The selective induction of IL-13 and IL-5 but not IL-4 by GA treatment suggests that the specific biological functions of these cytokines might be important for the therapeutic mechanism of GA. Measurement of serum IL-13 and IL-5 levels is a simple and inexpensive tool for monitoring the response to GA in MS patients.     

Serum interferon-gamma-inducing factor/IL-18 levels in primary biliary cirrhosis

Primary biliary cirrhosis is an autoimmune disease of the liver in which T helper 1 cytokines predominate over those of T helper 2 in the pathogenesis. Interleukin- 18 (IL-18), for which the gene was recently cloned, is a novel T helper 1 cytokine, which augments interferon-gamma production. We designed this study to clarify the role of IL-18 in primary biliary cirrhosis and to examine whether serum IL-18 level can be a prognostic indicator for the disease. Serum IL-18 levels were measured using an enzyme linked immuno sorbent assay with mouse monoclonal antibodies. Twenty-two healthy volunteers, 31 patients with primary biliary cirrhosis (Scheuer's stage I, 13; II, 10; and IV, 8), 20 patients with autoimmune hepatitis, 11 patients with virus-related liver cirrhosis and six patients with obstructive jaundice were enrolled. Significant differences of serum IL-18 levels were observed between patients with Scheuer's stage IV and those with stage I, or II, virus-related liver cirrhosis and obstructive jaundice (P < 0.05). The IL-18 levels in primary biliary cirrhosis increased according to the disease progression, and fell promptly after living-related liver transplantation. Moreover, serum IL-18 levels in primary biliary cirrhosis were correlated with serum bilirubin concentrations and the Risk scores of the Mayo Clinic prognostic model for the disease. The IL-18 levels observed in patients with autoimmune hepatitis were also elevated, and correlated with the activity of the disease. These results indicate that serum interleukin-18 levels reflect the severity of primary biliary cirrhosis, the activity of autoimmune hepatitis, and may be an additive prognostic indicator in primary biliary cirrhosis.     

支气管哮喘患儿治疗前后血清IL-2、IL-6和GM-CSF检测的临床意义

目的：探讨了支气管哮喘患儿治疗前后血清IL-2、IL-6和GM-CSF含量的变化及临床意义。方法：应用放射免疫分析对36例支气管哮喘患儿进行治疗前后血清IL-2、IL-6和GM-CSF测定,并与30名正常健康儿做比较。结果：在治疗前血清IL-6和GM-CSF水平非常显著地高于正常儿组（P〈0.01）,而IL-2水平又非常显著地低于正常儿组（P〈0.01）,经治疗一个月后则与正常儿比较无显著差异（P〉0.05）,直线相关分析显示：血清IL-2、IL-6和GM-CSF水平呈明显负相关（r=-0.5846、-0.6018,P〈0.01）。结论：血清IL-2、IL-6和GM-CSF在支气管哮喘患儿的发生、发展过程中相互作用,观察其浓度的变化对探讨其发病机理及指导用药均有重要的临床价值。     

Distribution of the hematopoietic growth factor G‐CSF and its receptor in the adult human brain with specific reference to Alzheimer's disease

The granulocyte colony‐stimulating factor (G‐CSF), being a member of the hematopoietic growth factor family, is also critically involved in controlling proliferation and differentiation of neural stem cells. Treatment with G‐CSF has been shown to result in substantial neuroprotective and neuroregenerative effects in various experimental models of acute and chronic diseases of the central nervous system. Although G‐CSF has been tested in a clinical study for treatment of acute ischemic stroke, there is only fragmentary data on the distribution of this cytokine and its receptor in the human brain. Therefore, the present study was focused on the immunohistochemical analysis of the protein expression of G‐CSF and its receptor (G‐CSF R) in the adult human brain. Since G‐CSF has been shown not only to exert neuroprotective effects in animal models of Alzheimer's disease (AD) but also to be a candidate for clinical treatment, we have also placed an emphasis on the regulation of these molecules in this neurodegenerative disease. One major finding is that both G‐CSF and G‐CSF R were ubiquitously but not uniformly expressed in neurons throughout the CNS. Protein expression of G‐CSF and G‐CSF R was not restricted to neurons but was also detectable in astrocytes, ependymal cells, and choroid plexus cells. However, the distribution of G‐CSF and G‐CSF R did not substantially differ between AD brains and control, even in the hippocampus, where early neurodegenerative changes typically occur.     

Resistance/susceptibility to Echinococcus multilocularis infection and cytokine profile in humans. I. Comparison of patients with progressive and abortive lesions

To clarify the role of Th1- and Th2-type cytokines in the various outcomes of human alveolar echinococcosis (AE), the cytokine immune response of self-cured patients was studied and compared with those of progressive AE patients and healthy subjects. Self-cured patients were divided into two groups according to the following clinical features: subjects who had positive Echinococcus multilocularis serologies and hepatic calcifications typical of AE were classified as 'abortive AE' patients, and those who had positive E. multilocularis serologies but no hepatic lesions or calcifications detectable by ultrasonography were classified as 'positive serology' subjects. Secretions of IL-5, IL-10 and interferon-gamma, and expression of IL-5 mRNA were evaluated in peripheral blood mononuclear cells (PBMC) stimulated in vitro with the mitogen phytohaemagglutinin-C or specific E. multilocularis antigenic preparations. The cytokine profile of abortive AE patients was the opposite of that observed in progressive AE patients. An intermediate profile was observed in positive serology subjects. PBMC from abortive AE patients, whether non-stimulated or stimulated with PHA and antigenic preparations, secreted significantly lower levels of IL-10 than those isolated from progressive AE patients. Our observations seem to confirm the regulatory role of IL-10 in the immunopathology of human AE.     

Expression of IL-18 by Mycobacterium avium-infected human monocytes; association with M. avium virulence

Disseminated Mycobacterium avium infection is the most frequent bacterial infection in patients with advanced AIDS and also associated with interferon-gamma (IFN-gamma) or IL-12 receptor deficiency. IFN-gamma is a key cytokine in host defence against M. avium infection. Expression of IL-18, a potent IFN-gamma inducer, and IFN-gamma by human monocytes after infection with M. avium was examined. Monocytes were co-cultured with isogenic smooth-transparent (SmT: virulent) or smooth-domed (SmD: avirulent) M. avium strains (10 organisms per monocyte). Infection with the SmD strain induced significantly higher concentration of IL-18 and IFN-gamma in culture supernatants than did the SmT strain. IFN-gamma production in response to M. avium was partially inhibited by anti-human IL-18 MoAb. Both recombinant human IL-12 (77 +/- 42 pg/ml, control versus 1492 +/- 141 pg/ml, cultures with IL-12 1 ng/ml) and IL-18 (126 +/- 37 pg/ml, control versus 2683 +/- 864 pg/ml, cultures with IL-18 10 ng/ml) augmented M. avium-induced IFN-gamma production. Freshly isolated uninfected monocytes expressed constitutive levels of IL-18. Following infection with M. avium, enhancement of IL-18 mRNA expression peaked at 3-6 h. IL-18 protein was detected in monocyte lysates as early as 1 h after infection with both SmT and SmD M. avium strains by Western blotting. Higher IL-18 expression by monocytes infected with the avirulent strain may result in more IFN-gamma production, thus modulating its pathogenicity. Local induction of IL-18 may be important both for M. avium pathogenicity and host defence and become a potential candidate for immunotherapy.     

Divergent regulation of cell surface protease expression in HL-60 cells differentiated into macrophages with granulocyte macrophage colony stimulating factor or neutrophils with retinoic acid

Taking advantage of the recently demonstrated presence of N-aminopeptidasesand the serine protease dipeptidyi aminopeptidase IV (DPP IV)at the surface of human myeloblastic HL-60 cells, the regulationof these protease activities in HL-60 cell differentiation hasbeen assessed using combined spectrophotometric and flow cytometricassays. Addition of human recombinant granulocyte macrophagecolony stimulating factor (rHu-GM-CSF) to HL-60 cells to inducedifferentiation into macrophages led to a time and dose-dependentincrease in both cell surface N-aminopeptidase and DPP IV activities.Protease up-regulation was due to an enhancement in cell surfaceprotease number, associated with a slight rise in apparent affinitiesof the enzymes for their substrates. In contrast, in HL-60 cellsinduced to differentiate into neutrophils in the presenceofretinoic acid, expression of cell surface N-amlnopeptidaseswas almost completely abolished in a time-and dose-dependentfashion, and this down-regulation was accompanied by a weakbut significant decrease in affinity. However, no noticeabledifference was seen in serine DPP IV expression between retinoicacid-treated and untreated HL-60 cells. Retinoic acid treatmentalso reduced soluble protease activity in vitro indicating thatdown-regulation of membrane aminopeptldases was not due to theirproteolytic clip. No modulation in the activity of any of theenzymes tested was seen with human recombinant tumor necrosisfactor- or retinol which do not induce HL-60 cell differentiation.The up-regulation of cell surface protease expression in HL-60cells differentiated into macrophages was similar to that observedin monocytes isolated from peripheral blood: both DPP IV andN-aminopeptidase activities strictly increased on cells thatundergo macrophage maturation (up to 5-fold) and independentlyof the nature of the differentiation inducer. Thus, the distinctivepatterns of N-aminopeptidase and DPP IV expression that areseen in differentiating neutrophils and macrophages appear tobe relatedto differences in stage of myeloid maturation. Becausecell surface proteases are crucially involved in leukocyte functions,the data presented suggest that alterations in cell surfaceprotease expression are associated with events controlling thedifferentiation of immature cells.     

Antitumor‐specific T‐cell responses induced by oncolytic adenovirus ONCOS‐102 (AdV5/3‐D24‐GM‐CSF) in peritoneal mesothelioma mouse model

Oncolytic adenoviral immunotherapy activates the innate immune system with subsequent induction of adaptive tumor‐specific immune responses to fight cancer. Hence, oncolytic viruses do not only eradicate cancer cells by direct lysis, but also generate antitumor immune response, allowing for long‐lasting cancer control and tumor reduction. Their therapeutic effect can be further enhanced by arming the oncolytic adenovirus with costimulatory transgenes and/or coadministration with other antitumor therapies. ONCOS‐102 has already been found to be well tolerated and efficacious against some types of treatment‐refractory tumors, including mesothelin‐positive ovarian cancer (NCT01598129). It induced local and systemic CD8+ T‐cell immunity and upregulated programmed death ligand 1. These results strongly advocate the use of ONCOS‐102 in combination with other therapeutic strategies in advanced and refractory tumors, especially those expressing the mesothelin antigen. The in vivo work presented herein describes the ability of the oncolytic adenovirus ONCOS‐102 to induce mesothelin‐specific T‐cells after the administration of the virus in bagg albino (BALB/c) mice with mesothelin‐positive tumors. We also demonstrate the effectiveness of the interferon‐γ the enzyme‐linked immunospot (ELISPOT) assay to detect the induction of T‐cells recognizing mesothelin, hexon, and E1A antigens in ONCOS‐102‐treated mesothelioma‐bearing BALB/c mice. Thus, the ELISPOT assay could be useful to monitor the progress of therapy with ONCOS‐102.     

Neutrophil apoptosis: impact of granulocyte macrophage colony stimulating factor on cell survival and viability in chronic kidney disease and hemodialysis patients

Introduction

Altered neutrophil apoptosis might be responsible for recurrent bacterial infections encountered in hemodialysis (HD) and chronic kidney disease (CKD) patients. This work was designed to assess the neutrophil apoptotic activity and the impact of implementation of granulocyte macrophage colony stimulating factor (GM-CSF), as a survival factor, on neutrophil apoptosis among these patients.

Material and methods

Twenty-five patients on regular HD along with 34 CKD patients on conservative treatment, as well as 15 healthy controls, were investigated for apoptotic rate via assessment of neutrophil expression of Annexin-V by flow cytometry, before and after 20 h culture in absence and presence of GM-CSF. Neutrophil viability was determined using light microscopy. The preservation of neutrophil activation in these patients was analyzed by flow cytometric CD18 neutrophil expression. Chronic inflammatory state was evaluated by estimating C-reactive protein (CRP) and soluble intercellular adhesion molecule-1 (sICAM-1). Obtained data were statistically analyzed.

Results

Compared to controls, both HD and CKD groups had a significant increase of Annexin-V and CD18 expression and significant decrease in neutrophil viability. Culture of their neutrophils with GM-CSF showed significant decrease of apoptosis accompanied by improvement of neutrophil viability compared to their cultured cells without GM-CSF. These patients also showed significant elevation of CRP and sICAM-1.

Conclusions

Granulocyte macrophage colony stimulating factor demonstrated an evident impact on improving in vitro neutrophil survival and viability in HD and CKD patients. Therefore, this may represent promising preventive and/or therapeutic strategies against infection frequently observed in these patients and causing morbidity and mortality.     

慢性前列腺炎患者治疗前后血清IL-6、IL-8和M-CSF测定的临床价值

目的:探讨慢性前列腺炎患者治疗前后血清IL-6、IL-8和M-CSF检测的临床意义.方法:应用放射免疫分析对33例慢性前列腺炎患者进行了治疗前后血清IL-6、IL-8和M-CSF检测,并与35例正常健康人作比较.结果:慢性前列腺炎患者在治疗前血清IL-6、IL-8和M-CSF水平非常显著地高于正常人组(P<0.01),...     

GM-CSF and IL-12 production by malignant plasma cells promotes cell-mediated immune responses against monoclonal Ig determinants in a light chain myeloma model

Immune responses towards malignant plasma cells have clearly been demonstrated in the course of monoclonal B cell dyscrasias and shown to be mostly specific for idiotypic determinants of the monoclonal immunoglobulin (Ig). These responses are specifically efficient against lymphoma cells expressing a membrane form of the monoclonal Ig. In myeloma, such immune responses are often weak and a number of strategies are currently assayed in order to boost the cell-mediated responses against the secreted monoclonal Ig. The use of cytokines promoting Th1 responses could be helpful for the induction of anti-tumour immunity and the control of residual disease in patients treated with myeloablative therapy, and such strategies need to be evaluated. In a light chain myeloma model where the monoclonal Ig can only be secreted, we tried to induce protective immune responses through immunization of animals with transfected malignant plasma cells. An expression plasmid encoding GM-CSF and IL-12 proved to be highly efficient for the induction of both cytotoxic and proliferative responses after immunization of animals with transfected and irradiated tumour cells. Anti-tumour immunization according to this protocol was successful in protecting 93.4% of the animals against a subsequent tumour challenge.     

Interleukin‐2 reverses CD8+ T cell exhaustion in clinical malignant pleural effusion of lung cancer

Malignant pleural effusion (MPE) is a poor prognostic sign for cancer patients, whereas the functional condition of MPE CD8⁺ T cells is unknown. Intracavitary immunotherapy with interleukin (IL)‐2 has been proven effective in controlling MPE. To elucidate the underlying mechanism, 35 lung cancer (LC) patients with MPE and 12 healthy donors were included in this study. For the IL‐2 therapy experiments, after draining partial MPE, we treated 14 patients by administrating IL‐2 (3 or 5 × 10⁶ U in 50 ml saline) into the thoracic cavity. Before and after IL‐2 treatment (40‐48 h), the MPE and peripheral blood (PB) were obtained from the subjects. PB from healthy volunteers was collected as control. The expression of programmed cell death 1 (PD‐1), granzyme B (GzmB), interferon (IFN)‐γ and the proliferation were analysed in CD8⁺ T cells from MPE and PB. The CD8⁺ T cells in the MPE of LC patients showed lowest GzmB, IFN‐γ and proliferation but highest PD‐1 expression, compared with that in PB of LC patients and healthy donors. IL‐2 treatment reduced the expression of PD‐1, increased the expression of GzmB and IFN‐γ and enhanced the proliferation of CD8⁺ T cells in MPE. In addition, IL‐2 treatment reduced carcino‐embryonic antigen (CEA) level in MPE. These results indicate that MPE CD8⁺ T cells exhibit exhaustion phenotype which can be reversed by IL‐2 therapy.     

IL-2 and IL-10 serum levels in HIV-1-infected patients with or without active antiretroviral therapy

We analyzed IL-2 and IL-10 serum levels in 26 HIV-1-infected patients naive of antiretroviral treatment and in 34 patients receiving highly active antiretroviral therapy (HAART). All patients without treatment were asymptomatic. When they were stratified according to levels of CD4+ T cells, IL-2 levels were significantly increased in patients with > or =200 CD4+/microl and IL-10 levels were significantly increased in patients with <200 CD4+/microl compared to controls. A significant negative correlation was observed between IL10 levels and CD4+ T-cell counts. No correlation was observed between IL-2 and IL-10 levels and viral load due to the wide range of variability in the number of HIV copies/ml present in the different patients. However, IL-2 levels were higher in patients with high viral load than in patients with low viral load. In patients with HAART, IL-2 and IL-10 levels were similar to the control group and no differences were detected respecting CD4+ T cells counts and viral load. Our findings show that the modifications in IL-2 and IL-10 serum levels in HIV-1-infected patients naive of antiretroviral treatment are associated with the progression of immunological damage. Furthermore, they show a dysbalance of type-1/type-2 cytokines with an involvement of type-2 cytokines in later stages of HIV infection. Cytokine dysregulation can be reversed by HAART in the context of immune restoration and viral suppression.     

Resistance/susceptibility to Echinococcus multilocularis infection and cytokine profile in humans. II. Influence of the HLA B8, DR3, DQ2 haplotype

Differences have been shown between HLA characteristics of patients with different courses of alveolar echinococcosis (AE). Notably the HLA B8, DR3, DQ2 haplotype was associated with more severe forms of this granulomatous parasitic disease. We compared IL-10, IL-5, interferon-gamma (IFN-gamma) and tumour necrosis factor (TNF) secretion by peripheral blood mononuclear cells (PBMC) isolated from eight HLA-DR3+, DQ2+, B8+ AE patients and from 10 HLA-DR3-, DQ2-, B8- patients after non-specific mitogenic and specific Echinococcus multilocularis antigenic in vitro stimulation. PBMC from seven HLA-DR3+, DQ2+, B8+ healthy subjects and nine HLA-DR3-, DQ2-, B8- subjects were also studied as controls. PBMC from AE patients with HLA DR3+, DQ2+ haplotype secreted higher levels of IL-10 without any stimulation and after specific antigenic stimulation than did patients without this haplotype. Higher levels of IL-5 and IFN-gamma were also produced by these patients' PBMC after stimulation with non-purified parasitic antigenic preparations; however, the specific alkaline phosphatase antigen extracted from E. multilocularis induced only Th2-type cytokine secretion. A spontaneous secretion of TNF by HLA DR3+, DQ2+ B8+ AE patients was also found. These results suggest that HLA characteristics of the host can influence immune-mediated mechanisms, and thus the course of AE in humans; specific antigenic components of E. multilocularis could contribute to the preferential Th2-type cytokine production favoured by the genetic background of the host.