Inflammation has a central role in cancer progression. Metastatic tumors arise at sites of chronic inflammation, and tumors or tumor-infiltrating immune cells produce inflammatory mediators. By contrast, natural killer (NK) cells and cytotoxic T cells (CTLs) help eliminate premalignant lesions and limit the rate of tumor metastasis. <em>Interleukin</em> (IL)-<em>27</em> is an IL-12 family cytokine chiefly produced by antigen-presenting cells (APCs) such as dendritic cells (DCs) and macrophages, and alone or in combination with other cytokines, IL-<em>27</em> boosts antitumor immunity by contributing to the development of NK cells and CTLs - a central immnunomodulatory effect - and by exerting potent antiangiogenic and antimetastatic activities, a local antitumor effect. In this review, we argue that by virtue of its rate-limiting functions in innate and adaptive immune responses, modulating IL-<em>27</em> holds considerable promise for future cancer immunotherapy.

Autism spectrum disorder (ASD) is a complex neurodevelopmental condition that can be reliably diagnosed at age 24 months. Immunological phenomena, including skewed cytokine production, have been observed among children with ASD. Little is known about whether immune dysregulation is present before diagnosis of ASD.

We examined neonatal blood spots from 214 children with ASD (141 severe, 73 mild/moderate), 62 children with typical development, and <em>27</em> children with developmental delay as control subjects who participated in the Childhood Autism Risks from Genetics and the Environment study, a population-based case-control study. Levels of 17 cytokines and chemokines were compared across groups and in relation to developmental and behavioral domains.

Interleukin (IL)-1β and IL-4 were independently associated with ASD compared with typical development, although these relationships varied by ASD symptom intensity. Elevated IL-4 was associated with increased odds of severe ASD (odds ratio [OR] = 1.40, 95% confidence interval [CI], 1.03, 1.91), whereas IL-1β was associated with increased odds of mild/moderate ASD (OR = 3.02, 95% CI, 1.43, 6.38). Additionally, IL-4 was associated with a higher likelihood of severe ASD versus mild/moderate ASD (OR = 1.35, 95% CI, 1.04, 1.75). In male subjects with ASD, IL-4 was negatively associated with nonverbal cognitive ability (β = -3.63, SE = 1.33, p = .04).

This study is part of a growing effort to identify early biological markers for ASD. We demonstrate that peripheral cytokine profiles at birth are associated with ASD later in childhood and that cytokine profiles vary depending on ASD severity. Cytokines have complex roles in neurodevelopment, and dysregulated levels may be indicative of genetic differences and environmental exposures or their interactions that relate to ASD.

Coronaviruses (CoVs) possess an enveloped, single, positive-stranded RNA genome which encodes for four membrane proteins, namely spike (S), envelope (E), membrane (M) and nucleocapsid (N) proteins 3-5 [1]. With regard to pathogenicity, S proteins are essential for viral entry into host cells [2, 3]. SARS-CoV binds to the angiotensin-converting enzyme (ACE)2 which is present on nonimmune cells, such as respiratory and intestinal epithelial cells, endothelial cells, kidney cells (renal tubules) and cerebral neurons and immune cells, such as alveolar monocytes/macrophages [4-6]. Of note, CD209L or liver/lymph node special intercellular adhesion molecule-3-grabbing non-integrin (SIGN) and dendritic cell (DC)-SIGN are alternative receptors for SARS-CoV but with lower affinity [7]. In the case of MERS-CoV, S proteins bind to the host cell receptor dipeptidyl peptidase 4 (DPP4 or CD26) which is broadly expressed on intestinal, alveolar, renal, hepatic and prostate cells as well as on activated leukocytes [8]. Then, viruses replicate in target cells with release of mature virions, which, in turn, invade new target cells [9]. Evidence has been provided that SARSCoV proteins are cleaved into two subunits, S1 and S2, respectively, and the amino acids 318-510 of the S1 represent the receptor-binding domain (RBD) which binds to ACE2 [10, 11]. Quite importantly, in the context of RBD there is the receptor-binding motif (RBM) (amino acids 424- 494), which accounts for complete binding to ACE2 [11]. Moreover, by means of two residues at positions 479 and 487 RBD allows virus progression and tropism [10, 11]. In the case of MERSCoV, its RBM binds to DPP4 with residues 484-567, thus, suggesting that its RBD differs from that of SARS-CoV [12, 13]. In a very recent paper, Wan and associates [14] have investigated the receptor recognition by COVID-19 (a new term to indicate the 2019-nCoV in Wuhan) on the bases of structural studies. In this respect, the sequence of COVID-19 RBM is similar to that of SARSCoV, thus, implicating that ACE2 may represent the binding receptors for COVID-19. Furthermore, gln493 residue of COVID-19 RBM seems to allow interaction with human ACE2, thus, suggesting the ability of this virus to infect human cells. According, to Wan and associates structural analysis [14], COVID-19 binds to human ACE2 with a lesser efficiency than human SARS-CoV (2002) but with higher affinity than human SARS-CoV (2003). Furthermore, same authors predicted that a single mutation at the 501 position may enhance the COVID-19 RBD binding capacity to human ACE2 and this evolution should be monitored in infected patients [14]. These predictive findings by Wan and associates [14] are confirmed by two contemporary studies by Letko and Muster [15] and Peng and associates [16]. In particular, the report by Peng and associates [16], points out the possible origin of COVID-19 from bats [16]. From a pathogenic point of view, evidence has been provided that binding of S2 to ACE2 receptor leads to its down-regulation with subsequent lung damage in the course of SARS-CoV infection [17]. Down-regulation of ACE2 causes excessive production of angiotensin (ANG) II by the related enzyme ACE with stimulation of ANG type 1a receptor (AT1R) and enhanced lung vascular permeability [18]. In particular, same authors have reported that recombinant ACE2 could attenuate severe acute lung injury in mice [18]. Moreover, Battle and associates [19] also proposed to use already available recombinant ACE2 for intercepting COVID-19 and attenuating infection. In the previous paragraphs, the presence of ACE2 on immune cells has been pointed out and, by analogy to epithelial cells, this receptor may also be down-regulated following viral entry. Therefore, in CoV-infected animal models and in infected humans further investigations are required to clarify a possible reduced expression of ACE2 on immune cells. In fact, in the course of SARS-CoV infection, a number of immune disorders have been detected. Three reports have demonstrated the ability of CoV to inhibit interferon (IFN)- production in the course of SARS acting as IFN antagonist [20-22]. In senescent Balb/c mice, depletion of T lymphocytes is associated to more severe interstitial pneumonitis and delayed clearance of SARS-CoV, thus, suggesting a protective role played by these cells [23]. In this connection, both SARS-CoV and MERS-CoV have been shown to induce T cell apoptosis, thus, aggravating the clinical course of disease [24, 25]. Quite interestingly, memory CD8+ T cells specific for SARS-CoV M and N proteins have been detected up to 11 years post-infection [26]. As far as humoral immune responsiveness is concerned, evidence has been provided that S1 subunit from MERS-CoV is highly immunogenic in mice [<em>27</em>]. Moreover, monoclonal antibodies have been shown to be highly neutralizing against MERS-CoV replication and endowed with post exposure effectiveness in susceptible mice [28, 29]. Human neutralizing antibodies have also been isolated from a recovered patient, thus, suggesting the role of humoral immunity in the control of the persistence of CoV in the host [30]. In particular, IgG response occurs early in infection and its prolonged production may serve for virus clearance during recovery also in view of the absence of viremia in convalescent sera from SARS patients [31]. According to current literature, severity of COVID-19 infection correlates with lymphopenia and patients who died from COVID-19 had lower lymphocyte counts when compared to survivors [32, 33]. These data suggest that lymphocyte-mediated anti-viral activity is poorly effective against COVID-19. Despite lymphopenia, evidence for an exaggerate release of proinflammatory cytokines [<em>interleukin</em> (IL)-1 and IL-6] has been reported in the course acute respiratory syndrome in COVID19 infected patients, thus, aggravating the clinical course of disease [34]. As recently reported, during COVID-19 pandemic in both Italy and China higher frequency of fatalities have been observed in the frail elderly population with previous comorbidities [35]. It is well known that decline of immunity occurs in ageing and, therefore, COVID-19 may gain easier access to the respiratory tract in frail elderly patients [36]. There is evidence that ACE2 protects from severe acute lung failure and operates as a negative regulator of the renin-angiotensin system (RAS) [18, 37]. It is well known that ANG II via activation of the AT1R promotes detrimental effects on the host, such as, vasoconstriction, reactive oxygen species generation, inflammation and matrix remodelling [38]. ACE2 counterbalances the noxious effects exhibited by ANG II and AT1R via activation of AT2R which arrests cell growth, inflammation and fibrosis [39]. In this framework, Gurwitz [40] proposed to use AT1R blockers, such as losartan, as a potential treatment of COVID-19 infection. In fact, losartan as well as olmesartan, used for treating hypertension in patients, were able to increase ACE2 expression after 28 days treatment of rats with myocardial infarction [41]. Then, Gurwitz suggests to evaluate severity of symptoms in COVID-19 infected patients under previous chronic treatment with AT1R blockers in comparison to COVID-19 infected patients who did not take AT1R blockers [40]. Quite interestingly, 75% of aged COVID-19 infected patients admitted to Italian hospitals had hypertension [unpublished data]. However, the putative effects of ACE-2 down-regulation on the cardiovascular system in the course of COVID-19 pandemic need more intensive studies. Taken together, these evidences suggest that CoV-induced down-regulation of ACE2 activates RAS with collateral damage to organs, such as lungs, in the course of SARS-related pneumonia. Then, putative therapeutic measures aimed at increasing ACE2 levels on respiratory epithelial cells should be taken into serious consideration. Quite interestingly, over the past few years, three key papers have demonstrated the ability of a polyphenol, resveratrol (RES), to experimentally deactivate the RAS system in maternal and post-weaning high fat diet, arterial ageing and high fat diet, respectively [42-44]. In all these experimental models, RES led to an increase of ACE2 with reduction of organ damage, such as liver steatosis and aorta media thickness and decrease of adipose tissue mass, respectively. As far as the mechanism of action of RES is concerned, this polyphenol is able to activate sirtuin (Sirt)1 [45-47]. In turn, Sirt1 down-regulates AT1R expression via ACE2 up-regulation [43, 48]. Of importance, Lin and associates [48] have demonstrated the ability of RES to in vitro inhibit MERS-CoV infection of Vero E6 cells, thus, prolonging cell survival in virtue of an anti-apoptotic mechanism. These findings suggest a direct antiviral effect exerted by RES. It would be very interestingly to evaluate the direct effects of RES on COVID-19, in vitro. The data above discussed strongly suggest, that RES, as an activators of ACE2, should be investigated in animal models of CoV-induced severe pneumonia, also taking into account the antioxidant, anti-inflammatory and immunomodulating effects exerted by polyphenols [49]. Then, successful animal studies may pave the way for RES-based human trials in COVID-infected patients. Note added in proof During the reviewing process of this perspective other related papers have been published. Hanff and associates [50] have discussed the possible association between COVID-19-associated cardiovascular mortality and dysregulation of the Renin Angiotensin System (RAS). From a pharmacologic point of view, RAS inhibition leads to upregulation of ACE2, thus, attenuating acute respiratory syndrome and myocarditis in COVID-19-infected patients. Conversely, increase in ACE2 expression may facilitate the access into the host of COVID-19, thus, aggravating the clinical picture. Such a dilemma would be solved by clinical trials based on RAS blockade or initiation and monitoring related effects. Contemporarily, Danser and associates [51] claim that there is no evidence to stop RAS blockers in the course of COVID-19 infection. In fact, there are no available data which support that ACE inhibitors or ANG II type I receptor blockers increase COVID-19 infection via its binding to ACE2. Finally, Kuster and associates [52] write that there are no data on the strict relationship between ACE2 activity and SARS-CoV2 mortality. Moreover, in the SARSCoV2, cells expressing ACE2 were not attacked by the virus, while cells lacking ACE2 were bound by the SARS-CoV2 virus [53]. These findings suggest that also in the case of RES effects on COVID-19 infection, the dual role of ACE2 should be taken into serious consideration.

Congenital infection with cytomegalovirus (CMV) can induce immune responses and placental damage. By use of immunoassay panels, <em>27</em> cytokines were assessed in midtrimester amniotic fluid from 8 patients with congenital CMV, in midtrimester sera from 12 pregnant women with primary CMV infection, and in amniotic fluid and serum from uninfected maternal controls. Levels of the cytokines tumor necrosis factor α, <em>interleukin</em> 1β, <em>interleukin</em> 12, and <em>interleukin</em> 17; the chemokines CCL2, CCL4, and CXCL10; and the growth factors granulocyte-macrophage colony-stimulating factor and platelet-derived growth factor bb were significantly elevated in amniotic fluid from congenital CMV patients (P < .01). Only CXCL10 was significantly higher in sera from CMV-infected pregnant women. CMV infection during pregnancy is associated with a shift in cytokine expression toward a proinflammatory state.

Systemic effects on T-cell-mediated antitumor immunity, on expression of T-cell adhesion/homing receptors, and on the promotion of T-cell infiltration of neoplastic tissue may represent key steps for the efficacy of immunological therapies of cancer. In this study, we investigated whether these processes can be promoted by s.c. administration of low-dose (0.5 microg/kg) recombinant human <em>interleukin</em>-12 (rHuIL-12) to metastatic melanoma patients. A striking burst of HLA-restricted CTL precursors (CTLp) directed to autologous tumor was documented in peripheral blood by a high-efficiency limiting dilution analysis technique within a few days after rHuIL-12 injection. A similar burst in peripheral CTLp frequency was observed even when looking at response to a single tumor-derived peptide, as documented by an increase in Melan-A/Mart-1(<em>27</em>-35)-specific CTLp in two HLA-A*0201+ patients by limiting dilution analysis and by staining peripheral blood lymphocytes (PBLs) with HLA-A*0201-melanoma antigen-A/melanoma antigen recognized by T cells (Melan-A/Mart)-1 tetrameric complexes. The CTLp burst was associated, in PBLs, with enhanced expression of T-cell adhesion/homing receptors CD11a/CD18, CD49d, CD44, and with increased proportion of cutaneous lymphocyte antigen (CLA)-positive T cells. This was matched by a marked increase, in serum, of soluble forms of the endothelial cell adhesion molecules E-selectin, vascular cell adhesion molecules (VCAM)-1 and intercellular adhesion molecules (ICAM)-1. Infiltration of neoplastic tissue by CDS+ T cells with a memory and cytolytic phenotype was found by immunohistochemistry in eight of eight posttreatment metastatic lesions but not in five of five pretreatment metastatic lesions from three patients. Increased tumor necrosis and/or fibrosis were also found in several posttherapy lesions of two of three patients in comparison with pretherapy metastases. These results provide the first evidence that rHuIL-12 can boost the frequency of circulating antitumor CTLp in tumor patients, enhances expression of ligand receptor pairs contributing to the lymphocyte function-associated antigen-1/ICAM-1, very late antigen-4/VCAM-1, and CLA/E-selectin adhesion pathways, and promotes infiltration of neoplastic lesions by CD8+ memory T cells in a clinical setting.

We previously reported that IL-<em>27</em>, which belongs to the IL-12 family of cytokines, is elevated in the serum of patients infected with influenza A virus (IAV). Here, we show that the expression of IL-<em>27</em> was significantly up-regulated in A549 human lung epithelial cells and human peripheral blood mononuclear cells infected with IAV. Additionally, IAV triggered IL-<em>27</em> expression through protein kinase A and cAMP-response element-binding protein signaling, which was mediated by cyclooxygenase-2-derived prostaglandin E(2). IL-<em>27</em> inhibited IAV replication by STAT1/2/3 phosphorylation and activated antiviral factor protein kinase R phosphorylation. Clinical analysis showed that IL-<em>27</em> levels were significantly elevated in a cohort of patients infected with IAV compared with healthy individuals and that circulating IL-<em>27</em> levels were tightly and positively correlated with prostaglandin E(2) levels. These results indicate that IL-<em>27</em> expression is one host immune factor produced in response to IAV infection and that elevated IL-<em>27</em> levels inhibit viral replication.

Skeletal muscle is the source of pro- and anti-inflammatory cytokines, and recently, it has been recognized as an important source of <em>interleukin</em>-6 (IL-6). Acute physical exercise is known to induce a pro-inflammatory cytokine profile in the plasma. However, the effect of chronic physical exercise in the production of pro- and anti-inflammatory cytokines by the skeletal muscle has never been examined. We assessed IL-6, TNF-alpha, IL-1beta and IL-10 levels in the skeletal muscle of rats submitted to endurance training. Animals were randomly assigned to either a sedentary group (S, n = 7) or an endurance exercise trained group (T, n = 8). Trained rats ran on a treadmill for 5 days week(-1) for 8 weeks (60% VO(2max)). Detection of IL-6, TNF-alpha, IL-1beta and IL-10 protein expression was carried out by ELISA. We found decreased expression of IL-1beta, IL-6, TNF-alpha and IL-10 (28%, <em>27</em>%, 32% and 37%, respectively, p < 0.05) in the extensor digital longus (EDL) from T, when compared with S. In the soleus, IL-1beta, TNF-alpha and IL-10 protein levels were similarly decreased (34%, 42% and 50%, respectively, p < 0.05) in T in relation to S, while IL-6 expression was not affected by the training protocol. In conclusion, exercise training induced decreased cytokine protein expression in the skeletal muscle. These data show that in healthy rats, 8-week moderate-intensity aerobic training down regulates skeletal muscle production of cytokines involved in the onset, maintenance and regulation of inflammation, and that the response is heterogeneous according to fibre composition.

Histone modifications are critical in regulating gene expression, cell cycle, cell proliferation, and development. Relatively few studies have investigated whether Helicobacter pylori, the major cause of human gastric diseases, affects histone modification. We therefore investigated the effects of H. pylori infection on histone modifications in a global and promoter-specific manner in gastric epithelial cells. Infection of gastric epithelial cells by wild-type H. pylori induced time- and dose-dependent dephosphorylation of histone H3 at serine 10 (H3 Ser10) and decreased acetylation of H3 lysine 23, but had no effects on seven other specific modifications. Different cag pathogenicity island (PAI)-containing-clinical isolates showed similar abilities to induce H3 Ser10 dephosphorylation. Mutation of cagA, vacA, nonphosphorylateable CagA mutant cagA(EPISA), or disruption of the flagella showed no effects, while deletion of the entire cagPAI restored the H3 Ser10 phosphorylation to control levels. Analysis of <em>27</em> cagPAI mutants indicated that the genes that caused H3 Ser10 dephosphorylation were similar to those that were previously found to induce <em>interleukin</em>-8, irrespective of CagA translocation. This effect was independent of ERK or p38 pathways and type I interferon signaling. Additionally, c-Jun and hsp70 gene expression was associated with this histone modification. These results demonstrate that H. pylori alters histone modification and host response via a cagA-, vacA-independent, but cagPAI-dependent mechanisms, which contribute to its persistent infection and pathogenesis.

OBJECTIVE

Exposure to bacterial endotoxin, perhaps due to bowel congestion or ischaemia and altered gut permeability, may result in immune activation that is characteristic for patients with severe heart failure. It is known that blood procalcitonin rises in response to bacterial endotoxin exposure.

METHODS

We measured procalcitonin in a group of 29 patients with acute cardiogenic shock and no sign of infection (all without bacteraemia) and 26 with septic shock. Blood was analysed for procalcitonin, interleukin-6, tumour necrosis factor-alpha (TNF-alpha), c-reactive protein (CRP) and neopterin. Patients were managed conventionally in an intensive care unit with no further experimental procedures.

RESULTS

Three cardiogenic (10%) and seven septic shock patients (27%) survived. Most patients with acute heart failure surviving 12 h or more (18 of 20) developed a pyrexia (738.0 degrees C) of unknown origin in the absence of positive cultures, with a rise in procalcitonin (1.4+/-0.8 to 48.0+/-16.2 ng/ml, P<0.001), CRP (76.5+/-16.4 to 154.7+/-22.9 mg/l, P<0.001) and neopterin (20.7+/-3.5 to 41.2+/-6.7 nmol/l, P<0.001). Patients with septic shock had higher initial levels of cytokines, and higher peak levels. Those with heart failure surviving (n=3) and those dying in the first 12 h (n=9) had no rise in cytokine levels. The patients with high procalcitonin had a higher temperature (38.9+/-0.3 vs. 37.3+/-0.23 degrees C, P<0.05), TNF-alpha (43.95+/-9.64 vs. 16.43+/-4.33 pg/ml; P<0.005) and CRP (146.1+/-18.4 vs. 68.2+/-39.6 mg/ml, P<0.005). Peak procalcitonin levels correlated with peak temperature (r=0.74, P<0.001).

CONCLUSIONS

Cardiogenic shock causes a pyrexia of unknown origin in patients surviving for 12 h and that is associated with a rise in procalcitonin levels. This lends support to the hypothesis that patients with cardiogenic shock may be being exposed to bacterial endotoxin at a time when bowel wall congestion and or ischaemia is likely to be present.

BACKGROUND

In models of COPD, environmental stressors induce innate immune responses, inflammasome activation and inflammation. However, the interaction between these responses and their role in driving pulmonary inflammation in stable COPD is unknown.

OBJECTIVE

To investigate the activation of innate immunity and inflammasome pathways in the bronchial mucosa and bronchoalveolar lavage (BAL) of patients with stable COPD of different severity and control healthy smokers and non-smokers.

METHODS

Innate immune mediators (<em>interleukin</em> (IL)-6, IL-7, IL-10, IL-<em>27</em>, IL-37, thymic stromal lymphopoietin (TSLP), interferon γ and their receptors, STAT1 and pSTAT1) and inflammasome components (NLRP3, NALP7, caspase 1, IL-1β and its receptors, IL-18, IL-33, ST2) were measured in the bronchial mucosa using immunohistochemistry. IL-6, soluble IL-6R, sgp130, IL-7, IL-<em>27</em>, HMGB1, IL-33, IL-37 and soluble ST2 were measured in BAL using ELISA.

RESULTS

In bronchial biopsies IL-<em>27</em>+ and pSTAT1+ cells are increased in patients with severe COPD compared with control healthy smokers. IL-7+ cells are increased in patients with COPD and control smokers compared with control non-smokers. In severe stable COPD IL-7R+, IL-<em>27</em>R+ and TSLPR+ cells are increased in comparison with both control groups. The NALP3 inflammasome is not activated in patients with stable COPD compared with control subjects. The inflammasome inhibitory molecules NALP7 and IL-37 are increased in patients with COPD compared with control smokers. IL-6 levels are increased in BAL from patients with stable COPD compared with control smokers with normal lung function whereas IL-1β and IL-18 were similar across all groups.

CONCLUSIONS

Increased expression of IL-<em>27</em>, IL-37 and NALP7 in the bronchial mucosa may be involved in progression of stable COPD.

Ectopic lymphoid-like structures (ELSs) reminiscent of secondary lymphoid organs often develop at sites of chronic inflammation where they contribute to immune-mediated pathology. Through evaluation of synovial tissues from rheumatoid arthritis (RA) patients, we now show that low <em>interleukin</em>-<em>27</em> (IL-<em>27</em>) expression corresponds with an increased incidence of ELS and gene signatures associated with their development and activity. The presence of synovial ELS was also noted in mice deficient in the IL-<em>27</em> receptor (IL-<em>27</em>R) after the onset of inflammatory arthritis. Here, pathology was associated with increased synovial expression of pro-inflammatory cytokines, homeostatic chemokines, and transcriptional regulators linked with lymphoid neogenesis. In both clinical and experimental RA, synovial ELS coincided with the heightened local expression of cytokines and transcription factors of the Th17 and T follicular helper (Tfh) cell lineages, and included podoplanin-expressing T cells within lymphoid aggregates. IL-<em>27</em> inhibited the differentiation of podoplanin-expressing Th17 cells, and an increased number of these cells were observed in IL-<em>27</em>R-deficient mice with inflammatory arthritis. Thus, IL-<em>27</em> appears to negatively regulate ELS development in RA through control of effector T cells. These studies open new opportunities for patient stratification and treatment.

<em>Interleukin</em> 6 (IL-6) is a pleiotropic inflammatory cytokine and its role in cancer is not yet clear. The effects of IL-6 on four breast cancer cell lines and normal mammary epithelium, cultured from milk were tested. Four different patterns of response to IL-6 were found depending on the differentiation status of the cells. In normal mammary epithelial cultures, the effects of IL-6 were mainly growth inhibitory, whereas in MCF-7, IL-6 had growth inhibitory and anti-adhesive effects. In T-47D and ZR-75-1 the anti-adhesive effects were prominent although the growth inhibitory effects were not. These anti-adhesive effects were associated with epithelioid to fibroblastoid morphological changes and a local decrease in E-cadherin expression. In the highly invasive cell line MDA-MB-231, which does not express E-cadherin, no effects of IL-6 were seen. IL-6 levels in the serum of 60 breast cancer patients were found to be increased in <em>27</em>% (16/60) compared to 2% (1/50) in a control group. Furthermore, it was found that altered E-cadherin expression was seen in 69% of the primary tumours, although no significant association was found between raised serum IL-6 levels and altered E-cadherin expression. Finally IL-6 serum levels did not effect the survival of breast cancer patients. The authors therefore implicate IL-6 as a possible factor important in breast cancer progression and metastasis formation, although the clinical significance of this cytokine in breast cancer patients could not be established.

<em>Interleukin</em> 6 (IL-6) is a pleiotropic cytokine produced by a wide variety of lymphoid and non-lymphoid tissues. We studied the relationship between IL-6 and the liver in an attempt to elucidate this cytokine's role in hepatitis C-induced liver inflammation. We investigated the behavior of serum IL-6 in 25 patients with chronic hepatitis C (divided into three groups depending on severity) and in <em>27</em> healthy controls. Our results showed a significant elevation (P < 0.0001) in serum IL-6 levels in the patients with chronic hepatitis C, correlated with the histological activity index (HAI) and their HCV-RNA serum levels. This rise may represent the expression of the hepatitis C virus-induced inflammatory state.

OBJECTIVE

To identify differentially expressed genes in healing mouse corneas by using cDNA microarrays.

METHODS

Transepithelial excimer laser ablations were performed on mouse corneas, and the wounds were allowed to heal partially in vivo for 18 to 22 hours. Total RNA was isolated from both normal and healing corneas and was used for synthesis of cDNA probes. 33P-labeled exponential cDNA probes were hybridized to mouse cDNA nylon arrays.

RESULTS

Of the 1176 genes on the nylon arrays, the expression of 37 was upregulated and that of <em>27</em> was downregulated more than fivefold in the healing corneas compared with the normal, uninjured corneas. <em>Interleukin</em> (IL)-1beta, laminin-5, and thrombospondin-1, which have been shown to be upregulated in healing corneas, were all found to be induced in the corneas in response to excimer laser treatment. Many genes were identified for the first time to be differentially regulated during corneal wound healing. Among the upregulated genes were intercellular adhesion molecule (ICAM)-1, macrophage inflammatory proteins, suppressors of cytokine signaling proteins (SOCS), IL-10 receptor, and galectin-7. Among the downregulated genes were connexin-31, a gap junction protein; ZO1 and occludin, tight junction proteins; and Smad2, a key component in the TGFbeta signaling pathway. Microarray data were validated on a limited number of genes by semiquantitative RT-PCR and Western blot analyses.

CONCLUSIONS

Gene array technology was used to identify for the first time many genes that are differentially regulated during corneal wound healing. These differentially expressed genes have not previously been investigated in the context of wound healing and represent novel factors for further study of the mechanism of wound healing.

OBJECTIVE

The purpose of this study was to determine if a bedside test, the MMP-8 PTD Check, can be of value in the antenatal identification of funisitis. This test can be performed in 15 min without any laboratory equipment.

METHODS

The relationship between the presence or absence of funisitis and the results of an MMP-8 PTD Check was examined in 139 patients who delivered preterm singleton neonates (gestational age <35 weeks) within 72 h of amniocentesis. Amniotic fluid (AF) was cultured for aerobic and anaerobic bacteria and for genital mycoplasmas. AF was analyzed for white blood cell (WBC) count, interleukin-6 (IL-6) and an MMP-8 PTD Check. The IL-6 concentration was also determined in umbilical cord plasma collected at birth. Funisitis was diagnosed in the presence of neutrophil infiltration into the umbilical vessel walls or Wharton's jelly.

RESULTS

1) Funisitis was present in 27% (38/139) of cases; 2) A positive MMP-8 PTD Check had a sensitivity of 97% (37/38), a specificity of 63% (64/101), a positive predictive value of 50% (37/74) and a negative predictive value of 99% (64/65) in the identification of funisitis; 3) Among cases without funisitis, patients with a positive MMP-8 PTD Check had a significantly higher median AF IL-6 concentration, AF WBC count, and umbilical cord plasma IL-6 concentration at birth than those with a negative MMP-8 PTD Check (P<0.05 for each).

CONCLUSIONS

The MMP-8 PTD Check is a rapid, simple and sensitive bedside test which allows assessment of the risk of funisitis.

<em>Interleukin</em> 1 alpha and tumour necrosis factor-alpha stimulated phosphorylation of three <em>27</em> kDa phosphoproteins in MRC-5 fibroblasts which was sustained for up to 2 h after adding the cytokines. All three phosphoproteins were immunoprecipitated by a specific antiserum to the small mammalian heat shock protein, hsp <em>27</em>. The three phosphoproteins from stimulated or control cells contained phosphoserine but not phosphothreonine or phosphotyrosine. Similar increases in phosphorylation of immunoprecipitable <em>27</em> kDa proteins were seen in U937 cells stimulated by TNF alpha and Hep G2 cells stimulated by IL1 alpha.

Hypertriglyceridemia may contribute to the development of atherosclerosis by increasing expression of cell adhesion molecules (CAMs). Although the cellular expression of CAMs is difficult to assess clinically, soluble forms of CAMs (sCAMs) are present in the circulation and may serve as markers for CAMs. In this study, we examined the association between sCAMs and other risk factors occurring with hypertriglyceridemia, the effect of triglyceride reduction on sCAM levels, and the role of soluble vascular cell adhesion molecule-1 (sVCAM-1) in monocyte adhesion in vitro. Compared with normal control subjects (n=20), patients with hypertriglyceridemia and low HDL (n=39) had significantly increased levels of soluble intercellular adhesion molecule-1 (sICAM-1) (316+/-28.8 versus 225+/-16.6 ng/mL), sVCAM-1 (743+/-52.2 versus 522+/-43.6 ng/mL), and soluble E-selectin (83+/-5.9 versus 49+/-3.6 ng/mL). ANCOVA showed that the higher sCAM levels in patients occurred independently of diabetes mellitus and other risk factors. In <em>27</em> patients who received purified n-3 fatty acid (Omacor) 4 g/d for>> or =7 months, triglyceride level was reduced by 47+/-4.6%, sICAM-1 level was reduced by 9+/-3.4% (P=.02), and soluble E-selectin level was reduced by 16+/-3.2% (P<.0001), with the greatest reduction in diabetic patients. These results support previous in vitro data showing that disorders in triglyceride and HDL metabolism influence CAM expression and treatment with fish oils may alter vascular cell activation. In a parallel-plate flow chamber, recombinant sVCAM-1 at the concentration seen in patients significantly inhibited adhesion of monocytes to <em>interleukin</em>-1-stimulated cultured endothelial cells under conditions of flow by <em>27</em>.5+/-7.2%. Thus, elevated sCAMs may negatively regulate monocyte adhesion.

<em>Interleukin</em> 13 (IL13) binds a receptor that is highly overexpressed in malignant gliomas, IL13Ralpha2. IL13 protein is composed of four helices: alpha-helix A, B, C, and D, and we found a new "hot spot" in alpha-helix D that is crucial for the binding of IL13 to IL13Ralpha2. Lys-105 plus Lys-106 and Arg-109 represent this hot spot. In the current study, we have made substitutions at these three positions in IL13. We examined both neutralization of an IL13-based cytotoxin's glioma cell killing and direct receptor binding of the new IL13 mutants. We observed that Lys-105 and Arg-109 are critical for IL13 binding to IL13Ralpha2, indeed. However, new mutants of important properties were identified with regard to tumor targeting. IL13.K105R mutant, in which lysine was substituted by arginine, neutralized the killing of IL13Ralpha2-positive cells by IL13-based cytotoxin more efficiently than wild-type IL13. However, IL13.K105L or IL13.K105A was deprived of any such activity. Furthermore, IL13.K105R and IL13.R109K competed 77- and <em>27</em>-fold better, respectively, with the binding of [(125)I]IL13 to the IL13Ralpha2 binding sites when compared with wild-type IL13. Thus, we have uncovered the first forms of IL13 of higher avidity toward IL13Ralpha2. These mutants should prove useful in the further design of anticancer diagnostics/therapeutics.

The purpose of this study was to evaluate the safety, tolerability, pharmacokinetics, and possible antitumor activity of a ligand fusion-protein, DAB389IL-2, in a phase I trial. This was a multicenter, open-label, dose-escalation trial. Patients with preserved organ function and histologically confirmed relapsed cutaneous T-cell lymphoma (CTCL), other non-Hodgkin's lymphomas (NHL), or Hodgkin's disease (HD) were eligible if their cancer was shown to express the <em>interleukin</em> (IL)-2 receptor by an immunohistochemical assay for the p55 or the p75 subunit. Patients received up to eight courses of DAB389IL-2 given as a short intravenous infusion daily for 5 days with subsequent courses every 21 days. The maximum tolerated dose (MTD) and tumor response was determined according to standard criteria. Seventy-three patients (44 men/29 women), aged 16 to 81 years (mean, 50.7) with CTCL (n = 35), NHL (n = 17), and HD (n = 21) were enrolled. The patients were extensively treated, failing 0 to 15 previous therapies (median, 4). Patients received one to six courses (mean, 3.3) of DAB389IL-2 over a range of 3 to 31 micrograms/kg/day. The dose-limiting toxicity was asthenia, establishing the maximum tolerated dose of <em>27</em> micrograms/kg/day. Approximately half of all patients had significant titers of antibody to diphtheria toxin or to DAB389IL-2 at the time of enrollment compared with 92% with titers at the end of treatment. The presence of antibody did not preclude clinical response. There were five complete (CR) and eight partial (PR) remissions in patients with CTCL with one CR and two PR occurring in NHL. The median time to response was 2 months and the duration of response was 2 to 39+ months. No responses were documented in patients with HD. DAB389IL-2 is well tolerated with an MTD of <em>27</em> micrograms/kg/day. This ligand fusion-protein showed antitumor effects in patients with IL-2 receptor expressing CTCL and NHL. Additional trials in these diseases are warranted.

Previous research has shown that parent mental health is associated with asthma morbidity in children. However, the biological pathways explaining these relationships are not known. The present study tested whether parent psychological characteristics could predict longitudinal changes in inflammatory markers in children with asthma and a comparison group of healthy children. For this, 33 healthy children (17 m/16 f; 13.5 years) and 50 children with asthma (37 m/13 f; 13.3 years) were assessed at two time points on average 208 days apart. Parent depression (CES-D) and perceived stress (PSS) were assessed at baseline, child depression (CDI) and anxiety (RCMAS) at follow-up. Asthma-relevant inflammatory markers eosinophil cationic protein (ECP) and stimulated <em>interleukin</em>-4 (IL-4) production were measured at baseline and at follow-up. Hierarchical regression analyses controlling for asthma severity and medication use revealed that higher levels of parental perceived stress at baseline were associated with greater increases over time in children's IL-4 production (beta=.29, p=.019) as well as ECP release (beta=.<em>27</em>, p=.004). Additionally, higher levels of parental depression at baseline were associated with increases in ECP over time (beta=.19, p=.046). There was no evidence that these associations were mediated by child depression or anxiety. These results demonstrate that parental stress and depression at baseline predict increases in children's inflammatory profiles over a six month period. This pattern appeared in both children with asthma and healthy children, and was not due to effects on child psychological states. These changes in inflammatory makers may represent one biological mechanism underlying the association between parental distress and child asthma morbidity.

MyD88 couples the activation of the Toll-like receptors and <em>interleukin</em>-1 receptor superfamily with intracellular signaling pathways. Upon ligand binding, activated receptors recruit MyD88 via its Toll-<em>interleukin</em>-1 receptor domain. MyD88 then allows the recruitment of the <em>interleukin</em>-1 receptor-associated kinases (IRAKs). We performed a site-directed mutagenesis of MyD88 residues, conserved in death domains of the homologous FADD and Pelle proteins, and analyzed the effect of the mutations on MyD88 signaling. Our studies revealed that mutation of residues 52 (MyD88(E52A)) and 58 (MyD88(Y58A)) impaired recruitment of both IRAK1 and IRAK4, whereas mutation of residue 95 (MyD88(K95A)) only affected IRAK4 recruitment. Since all MyD88 mutants were defective in signaling, recruitment of both IRAKs appeared necessary for activation of the pathway. Moreover, overexpression of a green fluorescent protein (GFP)-tagged mini-MyD88 protein (GFP-MyD88-(<em>27</em>-72)), comprising the Glu(52) and Tyr(58) residues, interfered with recruitment of both IRAK1 and IRAK4 by MyD88 and suppressed NF-kappaB activation by the <em>interleukin</em>-1 receptor but not by the MyD88-independent TLR3. GFP-MyD88-(<em>27</em>-72) exerted its effect by titrating IRAK1 and suppressing IRAK1-dependent NF-kappaB activation. These experiments identify novel residues of MyD88 that are crucially involved in the recruitment of IRAK1 and IRAK4 and in downstream propagation of MyD88 signaling.

BACKGROUND

Many intervertebral disc diseases cause low back pain (LBP). Proinflammatory cytokines and matrix metalloproteinases (MMPs) participate in disc pathology. In this study, we examined levels of serum cytokines and MMPs in human subjects with diagnoses of disc herniation (DH), spinal stenosis (SS), or degenerative disc disease (DDD) relative to levels in control subjects. Comparison between subjects with DH and those with other diagnoses (Other Dx, grouped from SS and DDD) was performed to elaborate a pathological mechanism based on circulating cytokine levels.

METHODS

Study participants were recruited from a spine neurosurgery practice (n = 80), a back pain management practice (n = <em>27</em>), or a control cohort (n = 26). Serum samples were collected before treatment and were assayed by multiplex assays for levels of <em>interleukin</em> (IL)-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, interferon-γ, tumor necrosis factor-α, MMP-1, MMP-3, and MMP-9. Inflammatory and degradative mediator levels were compared for subjects with LBP and control subjects, by diagnosis and by treatment groups, controlling for effects of sex, age, and reported history of osteoarthritis. Spearman's correlation coefficient was used to examine relationships with age, body mass index (BMI), symptom duration, and smoking history.

RESULTS

Serum levels of IL-6 were significantly higher in subjects with LBP compared with control subjects. Participants with LBP due to Other Dx had significantly higher levels of IL-6 than DH and controls. Serum levels of MMP-1 were significantly lower in LBP subjects, specifically those with DH, than in control subjects. Positive correlations were found between IL-6 levels and BMI, symptom duration, and age. MMP-1 levels were positively correlated with age.

CONCLUSIONS

The findings of the present clinical study are the results of the first examination of circulating cytokine levels in DDD and SS and provide evidence for a more extensive role of IL-6 in disc diseases, where patients with DDD or SS have higher serum cytokine levels than those with DH or control subjects. These findings suggest that LBP subjects have low-grade systemic inflammation, and biochemical profiling of circulating cytokines may assist in refining personalized diagnoses of disc diseases.

Patients with chronic obstructive pulmonary disease (COPD) are prone to acute respiratory exacerbations, which can develop suddenly or subacutely over the course of several days. Exacerbations have a detrimental effect on patients' health status and increase the burden on the healthcare system. Initial treatment is unsuccessful in 24-<em>27</em>% of patients, who have a relapse or a second exacerbation within 30 days of the initial event. No obvious benefit has been seen in recent clinical trials of anti-tumour necrosis factor therapy, anti-leukotriene therapy, intensive chest physiotherapy, or early inpatient pulmonary rehabilitation for treatment of exacerbations. By contrast, clinical trials of prevention rather than acute treatment have shown promising results. Long acting β agonist (LABA) or long acting anti-muscarinic (LAMA) bronchodilators and inhaled corticosteroid-LABA combinations prevent exacerbations in patients at risk, with relative risk reductions averaging 14-<em>27</em>% for each of these drugs relative to placebo. Triple therapy with inhaled corticosteroid-LABA plus LAMA may provide additional benefit, although study results to date are heterogeneous and more studies are needed. Pneumonia is an important complication of treatment with inhaled corticosteroid-LABA products, and the risk of pneumonia seems to be doubled in patients with COPD who use fluticasone. The addition of azithromycin to usual COPD therapy prevents exacerbations, although it may prolong the Q-T interval and increase the risk of death from cardiovascular disease in patients prone to arrhythmia. New potential drugs--including mitogen activated protein kinase inhibitors, phosphodiesterase 3 inhibitors, and monoclonal antibodies to the <em>interleukin</em> 1 receptor--offer additional hope for treatments that may prevent exacerbations in the future.

Induction of the <em>interleukin</em>-12 (IL-12) cytokine family comprising IL-12, IL-23, IL-<em>27</em>, and IL-12p40 by intracellular pathogens is required for orchestration of cell-mediated immune responses. Macrophages (MPhi) have been shown to be a source of IL-12 following TLR4-dependent activation by Salmonella (S.). In this study another antigen-presenting cell type, the conventional dendritic cell (cDC), was analyzed and its cytokine responses compared with those of MPhi. We generated bone marrow-derived conventional dendritic cells (BMDC) and macrophages (BMMPhi) by incubating murine bone marrow cells with supernatants containing granulocyte/macrophage colony-stimulating factor (GM-CSF) or macrophage colony-stimulating factor (M-CSF), respectively. Stimulation of BMDC and BMMPhi with S. enterica serovar Enteritidis (SE) or LPS resulted in the release of IL-12 and IL-23 by BMDC but not by BMMPhi. Furthermore, BMDC secreted approx. 20-fold more IL-12p40 and IL-<em>27</em>p28 than BMMPhi. However, BMDC and BMMPhi produced similar levels of IL-10. Using BMDC originating from wild-type (wt), TLR2(def) and TLR4(def) mice, we show that in BMDC the induction of IL-12, IL-23, and IL-<em>27</em>p28 by SE is dependent on TLR4, whereas low-level production of p40 is also mediated by pattern recognition receptors (PRR) other than TLR4. Interestingly, LPS- and SE-provoked responses of BMDC were remarkably similar indicating that LPS is the primary danger molecule of SE. Taken together, our results point to cDC rather than MPhi as the major producers of the IL-12 family members during in vitro infection with SE. The mechanisms of recognition of SE, however, appear to be the same for cDC and MPhi.