OBJECTIVE

To assess and correlate the levels of inflammatory mediators in the eyes from non-diabetic and diabetic subjects without retinopathy (NDR), with non-proliferative diabetic retinopathy (NPDR) or with proliferative diabetic retinopathy (PDR) to corresponding erum levels.

METHODS

The levels of <em>interleukin</em> 1β, <em>interleukin</em>-6 (IL-6) and tumour necrosis factor-α (TNF-α) were analysed by an ELISA-mimicking technique in the vitreous from 26 diabetic subjects with active PDR and <em>27</em> non-diabetic subjects, or by a multiplex bead assay in the aqueous humour from 35 diabetic subjects with NDR/NPDR and 40 non-diabetic subjects. Intraocular protein production was estimated in vitreous specimens by calculating a vitreous/serum ratio.

RESULTS

In the vitreous, IL-6 was higher in diabetic [157.5 (25.0-1401.0) pg/ml; median (min-max)] than in non-diabetic subjects [44.0 (5.0-4425) pg/ml; p = 0.021]. The vitreous/serum ratio was high (55.5:1 and 16:1, respectively), suggesting intraocular production. TNF-α was lower in diabetic [18.0 (8.0-46.0) pg/ml] than in non-diabetic subjects [22.0 (13.0-47.0) pg/ml; p = 0.034], but the vitreous/serum ratio was elevated in both groups (2:1 and 3.4:1, respectively). TNF-α levels were higher in serum from diabetic subjects [9.0 (5.0-53.0) pg/ml versus 6.7 (3.0-11.0) pg/ml; p < 0.001]. Aqueous levels of inflammatory mediators did not differ between diabetic subjects with NDR/NPDR and non-diabetic subjects despite elevated TNF-α in serum [<em>27</em>.8 (6.8-153.7) pg/ml versus 16.4 (4.1-42.4) pg/ml; p = 0.021].

CONCLUSIONS

Intraocular inflammation seems to be involved in PDR but does not seem to be prominent in early retinopathy stages, i.e. NDR or NPDR. Diabetic subjects have an overall increased inflammatory activity compared to non-diabetic subjects, as demonstrated by increased serum levels of TNF-α.

OBJECTIVE

To investigate whether marker(s) of systemic inflammation detect, at an early stage of acute pancreatitis, patients who may ultimately develop severe disease.

METHODS

Prospective study.

METHODS

University hospital emergency unit.

METHODS

Thirty patients with mild acute pancreatitis (SEV0 group) and <em>27</em> with severe acute pancreatitis. Of the latter, 11 did not develop organ failure (SEV1 group), whereas the other 16 patients developed acute respiratory failure and 9 of them also developed renal failure (SEV2 group).

METHODS

Blood samples were collected at admission to the hospital (T0), and at 12 hrs (T12) and 24 hrs (T24 after admission.

RESULTS

The plasma concentrations of procalcitonin (PCT), soluble E-selectin (sE-selectin), soluble interleukin-2 receptor (sIL-2R), and the serum concentration of C-reactive protein (CRP) were monitored. PCT levels at T0 were significantly higher in the SEV1 group (median 0.4 ng/mL, range 0.2-2.3) and the SEV2 group (0.8 ng/mL, 0.2-73.5) than in the SEV0 group (0.3 ng/mL, 0.1-3, p < .05 and p < .001, respectively). At T12, PCT level in the SEV2 group was significantly higher than that in the SEV1 group (2.2 ng/mL, 0.2-86.6 vs. 0.4 ng/mL, 0.3-2.8, p = .05), as it also was at T24 (2.2 ng/mL, 0.4-73.3 vs. 0.5 ng/mL, 0.3-4, p < .01). Among SEV2 patients, PCT concentration correlated negatively with the time elapsed between admission and the diagnosis of organ failure. At T12, sIL-2R levels of the SEV1 group (1,011 U/mL, range 334-2,211) and the SEV2 group (1,495 U/ml, range 514-4,526) both differed significantly from the SEV0 group (636 U/ml, range 356-1,678, p < .05 and p < .001, respectively) as they also did at T24. Although CRP level in the SEV1 group at T12 did not differ from the SEV0 group, the difference between SEV2 (<em>27</em>2 microg/mL, range 46-462) and SEV0 was significant (53 microg/mL, range 5-243, p < 0.01). sE-selectin levels did not differ between groups.

CONCLUSIONS

At admission to hospital, concentrations of PCT, but not those of CRP, sE-selectin, or sIL-2R, are higher in patients with severe acute pancreatitis than in patients with mild pancreatitis. PCT test had sensitivity of 94% and specificity of 73% for development of organ failure. PCT may be useful to identify the patients who benefit from novel therapies aimed at modifying the course of systemic inflammation.

Important constituents of extracellular matrix are collagen, fibronectin, hyaluronan, and various types of proteoglycans, such as versican, perlecan, decorin, and biglycan. Remodeling of extracellular matrix occurs continuously and is affected by various cytokines. The aim of this study was to investigate how <em>interleukin</em>-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha), separately and in combination, alter fibroblast proliferation, as well as the production of extracellular matrix molecules produced by human fibroblasts from lung. Fibroblast proliferation was inhibited significantly by all treatments, by 12% with IL-1beta and by 16% with TNF-alpha. The combination of IL-1beta and TNF-alpha increased the inhibition further, by <em>27</em>%. Hyaluronan production was increased by all treatments: 1.7-fold by IL-1beta and 1.5-fold by TNF-alpha. The combination of the two gave a further increase (2.5-fold). Similarly, the production of total proteoglycans was increased. The small proteoglycans, decorin, and biglycan, were regulated differently. Decorin production was inhibited by about 34% by all treatments, while biglycan was upregulated 1.3-fold by TNF-alpha. Versican was upregulated by IL-1beta (1.7-fold), whereas TNF-alpha was without effect. Perlecan was mostly unaffected. The changes in protein production of the various proteoglycans were due to increased transcription, as mRNA levels were also changed to the same extent. Synthesis of mRNA for collagen type I was inhibited by up to 75% with the IL-1beta/TNF-alpha combination. The separate cytokines also decreased the level of collagen type I mRNA, but to a lesser extent: 60% with IL-1beta and 40% with TNF-alpha. In summary, our study indicates that these proinflammatory cytokines affect the regulation of extracellular matrix production, which is of importance for the inflammatory process.

The human interferon-beta 2 gene (IFNB2) product is identical to that for the B-cell stimulation factor-2 (BSF-2), the hybridoma growth factor (HGF) ("<em>interleukin</em>-6"), and the hepatocyte stimulating factor (HSF). Proteins derived from this gene mediate the plasma protein response to tissue injury (acute-phase response) and regulate the growth and differentiation of both B and T cells. By using the enzymes MspI, BstNI, and BglI, three polymorphic systems were detected with probes for the IFNB2 gene. The MspI and BglI polymorphisms are likely to be due to base pair substitutions; the BstNI polymorphism was revealed by nine other enzymes and is likely to be due to DNA insertions within 1 kb of the 3' flanking region of the gene. This region is rich in AT dinucleotides, and slippage at DNA replication may generate the insertions of between 0.07 and 0.23 kb that were observed. The polymorphic MspI site also lies within the vicinity of the fifth exon. The BglI polymorphic site is likely to lie in 5' flanking DNA. The three polymorphisms are separate, and a variety of haplotypes was observed. A low level of linkage disequilibrium exists between the MspI and the BglI alleles. MspI and BstNI polymorphisms were observed in Caucasoids, CAR Pygmies, Zaire Pygmies, Melanesians, and Chinese but at differing frequencies, and not all alleles were present in all populations. The BglI polymorphism was observed in Caucasoids and Africans only. Linkage studies involving the IFNB2 gene and <em>27</em> other chromosome 7 markers have localized it to between D7S135 and D7S370 at 7p22-p21.(ABSTRACT TRUNCATED AT 250 WORDS)

<em>Interleukin</em> (IL)-23 and IL-<em>27</em> are IL-6/IL-12 family members that play a role in the regulation of T helper 1 cell differentiation. Cytokines are known to be involved in the bone remodeling process, although the effects of IL-23 and IL-<em>27</em> have not been clarified. In this study, we examined the possible roles of these cytokines on osteoblast phenotypes and osteoclastogenesis. We found that IL-<em>27</em> induced signal transducers and activators of transcription 3 activation in osteoblasts. However, neither IL-23 nor IL-<em>27</em> showed any significant effects on alkaline phosphatase activity, receptor activator of nuclear factor kappaB ligand (RANKL) expression, mRNA expression such as alkaline phosphatase type I procollagen, or the proliferation of osteoblasts. Osteoclastogenesis from bone marrow cells induced by soluble RANKL was partially inhibited by IL-23 and IL-<em>27</em> with reduced multinucleated cell numbers, but these <em>interleukins</em> did not affect the proliferation of osteoclast progenitor cells. These results indicate that IL-23 and IL-<em>27</em> could partly modify cell fusion or the survival of multinucleated osteoclasts. On the other hand, partially purified T cells, which are activated by 2 microg/ml anti-CD3 antibody, completely inhibited osteoclastogenesis by M-CSF/RANKL. On using T cells activated with 0.2 microg/ml anti-CD3 antibody, in which osteoclastogenesis was partially inhibited, the <em>interleukins</em> had additive effects for inhibiting osteoclastogenesis. Although the consequences of phosphorylated signals in osteoblasts have not been identified, IL-23 and IL-<em>27</em>, partly and indirectly through activated T cells, inhibited osteoclastogenesis, indicating that these <em>interleukins</em> may protect against bone destructive autoimmune disorders.

Ruxolitinib, a selective Janus kinase (JAK) 1&2 inhibitor in development for the treatment of myeloproliferative neoplasms, is primarily metabolized by CYP3A4. The effects of inhibition or induction of CYP3A4 on single oral dose ruxolitinib pharmacokinetics (PK) and pharmacodynamics (PD) were evaluated in healthy volunteers. Coadministration of ketoconazole (a potent CYP3A4 inhibitor) and erythromycin (a moderate CYP3A4 inhibitor) increased total ruxolitinib plasma exposure (AUC(0-∞)) by 91% and <em>27</em>%, respectively, and ruxolitinib PD, as measured by the inhibition of <em>interleukin</em> (IL)-6-stimulated STAT3 phosphorylation in whole blood, was generally consistent with the PK observed. Pretreatment with rifampin, a potent CYP3A4 inducer, decreased ruxolitinib AUC(0-∞) by 71% while resulting in only a 10% decrease in the overall PD activity. This apparent PK/PD discrepancy may be explained, in part, by an increase in the relative abundance of ruxolitinib active metabolites with the rifampin coadministration. The collective PK/PD data suggest that starting doses of ruxolitinib should be reduced by 50% if coadministered with a potent CYP3A4 inhibitor, whereas adjustments in ruxolitinib starting doses may not be needed when coadministered with inducers or mild/moderate inhibitors of CYP3A4. All study doses of ruxolitinib were generally safe and well tolerated when given alone and in combination with ketoconazole, erythromycin, or rifampin.

Foxp3-expressing regulatory T cells (Tregs) are central regulators of immune homeostasis and tolerance. As it has been suggested that proper Treg function is compromised under inflammatory conditions, seeking for a pathway that enhances or stabilizes Treg function is a subject of considerable interest. We report that <em>interleukin</em> (IL)-<em>27</em>, an IL-12 family cytokine known to have both pro- and anti-inflammatory roles in T cells, plays a pivotal role in enhancing Treg function to control T cell-induced colitis, a model for inflammatory bowel disease (IBD) in humans. Unlike wild-type (WT) Tregs capable of inhibiting colitogenic T-cell expansion and inflammatory cytokine expression, IL-<em>27</em>R-deficient Tregs were unable to downregulate inflammatory T-cell responses. Tregs stimulated with IL-<em>27</em> expressed substantially improved suppressive function in vitro and in vivo. IL-<em>27</em> stimulation of Tregs induced expression of Lag3, a surface molecule implicated in negatively regulating immune responses. Lag3 expression in Tregs was critical to mediate Treg function in suppressing colitogenic responses. Human Tregs also displayed enhanced suppressive function and Lag3 expression following IL-<em>27</em> stimulation. Collectively, these results highlight a novel function for the IL-<em>27</em>/Lag3 axis in modulating Treg regulation of inflammatory responses in the intestine.

It is well known that cytokines play an important role in oral diseases. Furthermore, increased levels of <em>interleukin</em> 6 (IL-6) and tumor necrosis factor alpha (TNF-alpha) have been reported in patients with cancer and premalignant lesions such as oral lichen planus and oral submucous fibrosis. The aim of this study was to assess salivary IL-6 and TNF-alpha levels in 30 patients with histopathologically confirmed leukoplakia (age range 24-78, mean 52.3 years) in comparison to 34 controls (age range <em>27</em>-79, mean 52 years). Salivary IL-6 and TNF-alpha were determined by enzyme linked immunoadsorbent assay. Statistical analysis was performed by use of Mann-Whitney test for independent samples and values lower than 0.05 were considered as significant (p<0.05). Significantly higher levels of salivary IL-6 and TNF-alpha in patients with oral leukoplakia when compared to healthy controls were found. The levels of salivary IL-6 and TNF-alpha did not correlate with the size of leukoplakia (lesions) nor with its localization regarding high and low risk sites for malignant transformation. Levels of salivary IL-6 and TNF-alpha were not influenced by smoking habits. We can conclude that increased salivary IL-6 and TNF-alpha might play a certain role in oral leukoplakia.

Natural killer (NK) cells are innate lymphoid cells that are important effectors in the first line of defense toward transformed cells. This is mediated both by direct cytotoxic mechanisms and by production of immunoregulatory cytokines. Recent evidence has shown that NK cells also display memory, similar to the cells of the adaptive immune system. Cytokines are pivotal for the maturation, activation and survival of NK cells. <em>Interleukins</em> (IL)-2, IL-12, IL-15, IL-18, IL-21 and type I interferons positively regulate NK cell function, either independently or in cooperation, whereas other cytokines, such as IL-23 and IL-<em>27</em>, may enhance or suppress NK cell function depending on the context. In the tumor microenvironment, TGFβ, IL-10 and IL-6 suppress NK cell activity not only directly, but also indirectly, by affecting immunosuppressive cells and by antagonizing the effect of stimulatory cytokines, thereby dampening the antitumor response of NK cells and promoting subsequent tumor evasion and progression. Increased understanding of the NK cell response to cytokines has provided a better understanding of their impaired function in tumors which may aid in the development of novel immunotherapeutic strategies to enhance NK cell responses in cancer patients.

Heteronuclear 13C and 15N three-dimensional nuclear magnetic resonance (n.m.r.) techniques have been used to determine the solution structure of human <em>interleukin</em> 4, a four-helix bundle protein. A dynamical simulated annealing protocol was used to calculate an ensemble of structures from an n.m.r. data set of 1735 distance restraints, 101 phi angle restraints and <em>27</em> pairs of hydrogen bond restraints. The protein structure has a left-handed up-up-down-down topology for the four helices with the two long overhand loops in the structure being connected by a short section of irregular antiparallel beta-sheet. Analysis of the side-chains in the protein shows a clustering of hydrophobic residues, particularly leucines, in the core of the bundle with the side-chains of charged residues being located on the protein surface. The solution structure has been compared with a recent structure prediction for human <em>interleukin</em> 4 and with crystal structures of other helix bundle proteins.

The immune response to hepatitis B surface antigen (HBsAg) is mostly genetically determined. <em>Interleukin</em> 10 (IL-10) is a central immunoregulatory cytokine with important effects on B-cells. We have studied the influence of IL-10 promoter polymorphisms on the immune response to HBsAg and hepatitis A vaccination. We vaccinated 202 twin pairs in an open prospective study with a combined recombinant HBsAg/inactivated hepatitis A vaccine. IL-10 promoter polymorphisms were investigated in all individuals and their influence on anti-HBs, and anti-HAV responsiveness was studied. In the multiple regression analysis accounting for smoking, gender, body mass index and age, the ACC haplotype (-1082, -819 and -592) had a strong influence on anti-HBs production. Individuals carrying the ACC haplotype had anti-HBs titres almost twice as high as individuals without this haplotype. In contrast, anti-HAV production was suppressed by the presence of the -1082A allele in comparison with individuals homozygous for the -1082G allele. The contribution of the shared IL-10 promoter haplotype accounted for <em>27</em>% of the genetic influence on anti-HBs antibody response. In conclusion, genetic variability in the IL-10 promoter is an important modulator of the immune response against hepatitis viral antigens.

The recognition that CD4+ T-cell responses could be divided into at least two functional subsets either dominated by production of interferon (IFN)-gamma and associated with cell-mediated immunity (Th1) or characterized by production of <em>interleukin</em> (IL)-4 and IL-5 and associated with humoral immunity (Th2) provided a basis to understand the role of T cells in resistance or susceptibility to different types of pathogens. As a consequence, many studies have focused on the identification of cytokines that influence these events. For example, the development of Th1-type responses is largely dependent on IL-12. However, other cytokines also affect this process, and initial studies revealed that IL-<em>27</em>, a cytokine with close structural and functional similarity to IL-12, can promote Th1 responses required for immunity to Leishmania major. Subsequent work with IL-<em>27</em>R (WSX-1)-deficient mice infected with Toxoplasma gondii or Trypanosoma cruzi revealed that the IL-<em>27</em>/IL-<em>27</em>R system can act as a negative regulator of inflammatory T-cell responses. The aim of this review is to discuss recent studies from these laboratories on the role of IL-<em>27</em> in immunity to parasitic infections in the context of previous work and to highlight the pleiotropic effects of the IL-<em>27</em>/IL-<em>27</em>R system in the development and regulation of Th1 and Th2 responses.

The current study was designed to investigate the impact of genetic heterogeneity on host immune responses to pulmonary intracellular infection by using two mouse strains of distinct genetic background, C57BL/6 and BALB/c mice, and a model intracellular pathogen, Mycobacterium bovis BCG. Upon infection, compared to C57BL/6 mice, BALB/c mice developed an earlier response of <em>interleukin</em> 12 (IL-12), gamma interferon (IFN-gamma), tumor necrosis factor alpha, and macrophage chemoattractive protein 1, and greater neutrophilic influx to the lung by days 7 and 14. However, the level of these cytokines at days <em>27</em>, 43, and 71 was much lower in BALB/c mice than in C57BL/6 mice. The magnitude of cellular responses was also much lower in the lung of BALB/c mice around day <em>27</em>. Histologically, while C57BL/6 mice developed lymphocytic granulomas, BALB/c mice displayed atypical granulomas in the lung. Of importance, the level of type 2 cytokines IL-4 and IL-10 remained low and similar in the lung of both C57BL/6 and BALB/c mice throughout. Furthermore, lymphocytes isolated from systemic and local lymphoid tissues of infected BALB/c mice demonstrated a markedly lower antigen-specific IFN-gamma recall response. While the number of mycobacterial bacilli recovered from both the lung and spleen of BALB/c mice was similar to that in C57BL/6 mice at day 14, it was higher than that in C57BL/6 mice at day 43. However, it was eventually leveled off to that in C57BL/6 counterparts later. These results suggest the following: (i) genetic heterogeneity can lead to differential innate and adaptive cell-mediated immune responses to primary pulmonary mycobacterial infection; (ii) it is the level of adaptive, but not innate, immune response that is critical to host resistance; and (iii) a lower type 1 immune response in BALB/c mice is not accompanied by a heightened type 2 response during pulmonary mycobacterial infection.

OBJECTIVE

Acute myeloid leukemia (AML) accounts for more than half of fatal cases in all pediatric leukemia patients; this observation highlights the need of more effective therapies. Thus, we investigated whether <em>interleukin</em> (IL)-<em>27</em>, an immunomodulatory cytokine, functions as an antitumor agent against pediatric AML cells.

METHODS

Expression of WSX-1 and gp130 on AML cells from 16 pediatric patients was studied by flow cytometry. Modulation of leukemia cell proliferation or apoptosis upon IL-<em>27</em> treatment in vitro was tested by bromodeoxyuridine/propidium iodide (PI) and Ki67, or Annexin V/PI staining and flow cytometric analysis. The angiogenic potential of AML cells treated or not with IL-<em>27</em> was studied by chorioallantoic membrane assay and PCR array. In vivo studies were carried out using nonobese diabetic/severe combined immunodeficient (NOD/SCID)/Il2rg(-/-) mice injected intravenously with five pediatric AML cell samples. Leukemic cells engrafted in PBS and IL-<em>27</em>-treated animals were studied by immunohistochemical/morphologic analysis and by PCR array for expression angiogenic/dissemination-related genes.

RESULTS

We provided the first demonstration that (i) AML cells injected into NOD/SCID/Il2rg(-/-) mice gave rise to leukemia dissemination that was severely hampered by IL-<em>27</em>, (ii) compared with controls, leukemia cells harvested from IL-<em>27</em>-treated mice showed significant reduction of their angiogenic and spreading related genes, and (iii) similarly to what was observed in vivo, IL-<em>27</em> reduced in vitro AML cell proliferation and modulated the expression of different genes involved in the angiogenic/spreading process.

CONCLUSIONS

These results provide an experimental rationale for the development of future clinical trials aimed at evaluating the toxicity and efficacy of IL-<em>27</em>.

<em>Interleukin</em> (IL)-10 is an immunoregulatory cytokine that is produced by diverse cell populations. Studies in mice suggest that the cellular source of IL-10 is a key determinant in various disease pathologies, yet little is known regarding the control of tissue-specific human IL-10 expression. To assess cell type-specific human IL-10 regulation, we created a human IL-10 transgenic mouse with a bacterial artificial chromosome (hIL10BAC) in which the IL10 gene is positioned centrally. Since human IL-10 is biologically active in the mouse, we could examine the in vivo capacity of tissue-specific human IL-10 expression to recapitulate IL-10-dependent phenotypes by reconstituting Il10(-/-) mice (Il10(-/-)/hIL10BAC). In response to LPS, Il10(-/-)/hIL10BAC mice proficiently regulate IL-10-target genes and normalize sensitivity to LPS toxicity via faithful human IL-10 expression from macrophages and dendritic cells. However, in the Leishmania donovani model of pathogen persistence, Il10(-/-)/hIL10BAC mice did not develop the characteristic IL-10(+)IFN-gamma(+)CD4 T cell subset thought to mediate persistence and, like Il10(-/-) mice, cleared the parasites. Furthermore, the IL-10-promoting cytokine IL-<em>27</em> failed to regulate transgenic human IL-10 production in CD4(+) T cells in vitro which together suggests that the hIL10BAC encodes for weak T cell-specific IL-10 expression. Thus, the hIL10BAC mouse is a model of human gene structure and function revealing tissue-specific regulatory requirements for IL-10 expression which impacts disease outcomes.

To evaluate the use of HSV-based vectors for arthritis gene therapy we have constructed a first-generation, ICP4 deficient, replication defective herpes simplex virus (HSV) vector (S/0-) and a second-generation HSV vector derivative (T/0-) deficient for the immediate-early genes ICP4, 22 and <em>27</em>, each carrying a soluble TNF receptor or IL-1 receptor antagonist transgene cassette. A rabbit synovial-fibroblast line in culture, infected by either vector enabled high-level expression of the transgene product. However, following a single intra-articular injection of the vectors into rabbit knee joints, only the second-generation, HSV T/0- vector expressed detectable levels of soluble TNFR in synovial fluid. Synovial lavage fluid from inoculated joints con- tained up to 12 ng/ml of soluble receptor that persisted at detectable, but reduced levels for at least 7 days. When tested in an experimental model of arthritis generated by intra-articular overexpression of <em>interleukin</em>-1beta using retrovirus transduced synovial cells, the HSV T/0- vector expressing the <em>interleukin</em>-1 receptor antagonist was found to inhibit leukocytosis and synovitis significantly. The improved levels and duration of intra-articular transgene expression achieved via HSV-mediated gene delivery suggest that an HSV vector system could be used for therapeutic applications in patients with rheumatoid arthritis (RA) and other joint-related inflammatory diseases.

Protection against intracellular pathogens such as Mycobacterium tuberculosis requires the development of Th1-like T-cell responses. This in turn is dependent on the pattern of cytokine produced from dendritic cells (DCs) after infection. Three heterodimeric cytokines, <em>interleukin</em>-12 (IL-12), IL-23, and IL-<em>27</em>, as well as IL-18, contribute to the differentiation and expansion of naive CD4(+) T cells. In this study we compared the effects of plasmids expressing both chains of IL-12, IL-23, or IL-<em>27</em> as adjuvants for DNA immunization against M. tuberculosis infection. The genes encoding p19 and p40 chains of IL-23 or EBI3 and p28 chains of IL-<em>27</em> were cloned on either side of a self-cleaving peptide from the FMDV2A protein. The secretion of functional cytokines from transfected cells was detected with bioassays. Supernatant from p2AIL-23-transfected cells induced the release of IL-17 from activated lymphocytes, confirming the presence of bioactive IL-23. Further, supernatant from p2AIL-<em>27</em>-transfected cells stimulated a significant increase in the proliferation of peptide-stimulated transgenic CD4(+) T cells. In initial experiments, M. tuberculosis infection of DCs was more potent at inducing IL-12 and IL-23 secretion than infection with the vaccine strain Mycobacterium bovis bacille Calmette-Guérin (BCG), and no significant upregulation of IL-<em>27</em> was observed. Coimmunization of C57BL/6 mice with DNA expressing M. tuberculosis antigen 85B (Ag85B; DNA85B) and plasmids expressing IL-23 or IL-12 stimulated stronger Ag85B-specific T-cell proliferative and IFN-gamma responses than DNA85B alone, whereas the addition of p2AIL-<em>27</em> had no effect. Interestingly, DNA85B codelivered with p2AIL-12, but not p2AIL-23, reduced the immunoglobulin G antibody response. Both p2AIL-23 and p2AIL-12, but not p2AIL-<em>27</em>, enhanced the protective efficacy of DNA85B against aerosol M. tuberculosis challenge. Therefore, both p2AIL-23 and p2AIL-12 are valuable as cytokine adjuvants for increasing the protective antituberculosis immunity induced by DNA vaccines.

The pathogenesis of chronic obstructive pulmonary disease (COPD) is not fully understood, and environment and genetic factors have been investigated. Moreover, cytokine genes play an important role in COPD pathogenesis. However, the molecular mechanism of COPD induced by the factors is still unknown. The present study was undertaken to clarify a role of <em>interleukin</em> (IL)-12 16974A/C and IL-<em>27</em> 4730T/C, -964A/G, and 2905T/G polymorphisms in Chinese subjects with COPD. Polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) and sequence analyses were used to type IL-12 and IL-<em>27</em> polymorphisms in 120 patients with COPD and 100 healthy controls. There were significant differences in the genotype and allele distribution of -964A/G and 2905T/G polymorphisms of the IL-<em>27</em> gene among cases and controls in a Chinese population. When compared with the control group, subjects with AG genotype of the IL-<em>27</em> -964A/G had a 2.22-fold decreased risk of COPD (odds ratio [OR] = 0.450, 95% confidence interval [CI]: 0.245-0.826; p = 0.009), and subjects with TG genotype of the IL-<em>27</em> 2905T/G had a 2.85-fold decreased risk of COPD (OR = 0.351, 95% CI: 0.137-0.899; p = 0.024). Compared with the TAT haplotype, the TGG haplotype was associated with a significantly decreased risk of COPD (OR = 0.29, 95% CI: 0.108-0.784; p = 0.010). Even after Bonferroni corrections, significant associations with COPD were observed for the AG genotype of the IL-<em>27</em> -964A/G and the TGG haplotype of the IL-<em>27</em> gene. Our data suggest that polymorphisms in the IL-<em>27</em> gene may play a role in the development of COPD in Chinese population.

BACKGROUND

This study analyzes the impact of Staphylococcus aureus enterotoxins (SAEs) and the inflammatory pattern in polyps from China.

METHODS

Nasal tissue was obtained from <em>27</em> consecutive bilateral nasal polyps and 15 control patients and assayed for eotaxin, <em>interleukin</em>-5, soluble <em>interleukin</em>-2 receptor, transforming growth factor (TGF) beta, myeloperoxidase, eosinophil cationic protein, total IgE, and specific IgE to SAEs. Activated eosinophils were stained using EG2 antibodies in polyps from Chinese and comparative white patients.

RESULTS

The number of EG2+ eosinophils was significantly lower in polyps from Chinese patients versus white patients. Chinese polyps showed significantly increased IgE and soluble <em>interleukin</em>-2 receptor versus control samples, whereas TGF-beta1 was significantly decreased. Ten of <em>27</em> samples in the polyp group versus 0 of 15 controls contained SAE-IgE (p < 0.01). TGF-beta1 was significantly down-regulated in SAE+ samples versus SAE- samples (p = 0.04).

CONCLUSIONS

Nasal polyps from China are characterized by B- and T-cell activation, a minor eosinophilic inflammation compared with polyps from white subjects, and a decrease in TGF-beta1 in comparison with control inferior turbinate tissue. One-third of patients with polyps showed an IgE response to SAEs.

We have established a sensitive ELISPOT assay measuring interferon gamma (IFN gamma) release on a single-cell basis to detect influenza peptide-specific CD8+ T cells in uncultured peripheral blood mononuclear cells (PBMC). Using this method, we studied the T cell response to HLA-A1 and HLA-A2.1 binding peptide epitopes derived from the MAGE-1 and MAGE-3 proteins, from the melanoma-associated antigens tyrosinase, Melan-A/MART-1 and gp100, and from influenza proteins in stage IV melanoma patients and healthy controls. In 18 of 24 HLA-A2-positive donors (75%), but only in 9 of 25 HLA-A2-positive melanoma patients (36%) T cells reactive with the influenza matrix peptide were demonstrated (p = 0.007). T cells responding to one or several of the melanoma-associated peptides were detected in 5 of 25 HLA-A2-positive patients with metastatic melanoma. Four of these 5 patients had been treated with <em>interleukin</em>-2- and IFN alpha-containing therapy. Two of the 24 healthy donors had T cells reactive with the MART-1 <em>27</em>-35 peptide. No reactivity with the HLA-A1-binding peptides from MAGE-1 or MAGE-3 was detected in any of the HLA-A1-positive healthy controls or melanoma patients. These results show that the IFN gamma-ELISPOT assay is suitable to determine quantitatively T cells reactive with melanoma-associated and influenza peptide epitopes in uncultured PBMC. The failure to detect T cells responding to influenza in many melanoma patients with progressive disease may indicate an impairment of their T cell function.

BACKGROUND

Interleukin (IL)-1 has been associated with joint inflammation and damage in rheumatoid arthritis (RA). Endogenous IL-1 receptor antagonist (IL-1Ra) may inhibit IL-1-mediated effects by binding to the IL-1 receptors.

OBJECTIVE

These studies were conducted to determine the efficacy and safety of human recombinant IL-1 receptor antagonist (IL-1ra) in blunting the effects of IL-1 in patients with RA.

METHODS

Three randomized clinical trials have been completed. First, in a 24-week, double-blind, placebo-controlled study of 472 patients with severe RA, patients were randomly placed into 4 groups: placebo or IL-1ra at 30, 75, or 150 mg/d. In a second study, 309 patients from the placebo-controlled trial enrolled in a 24-week extension study. Patients who had been receiving placebo were randomized into 1 of the 3 treatment groups. Those receiving treatment continued to receive their previous dosages. Finally, in a second double-blind, placebo-controlled study, 419 patients with RA receiving methotrexate at 12.5 to 25 mg/wk for at least 6 months were randomly placed into 1 of 6 groups: placebo or IL-1ra at 0.04, 0.1, 0.4, 1.0, or 2.0 mg/kg/d.

RESULTS

In the first study, the primary therapeutic endpoint-an American College of Rheumatology 20% response (ACR20)-was reached by 27% of the placebo group, compared with 43% of the IL-1ra 150-mg/d group. Individual clinical responses, as well as the arrest of joint damage, were also superior in treated patients. In the second trial, 52% of the randomized patients achieved an ACR20. This was maximal (71%) in those receiving the highest treatment dosage. Of patients continuing with the same dosage, 49% maintained an ACR20 at 48 weeks. Radiologic evaluation showed that the reduced rate of cartilage degradation observed in treated patients during the first 24 weeks was maintained, and the rate of joint erosion was slowed even more significantly in the second phase of the study. In the third study, significantly more patients receiving IL-1ra at 1.0 (46%, P =.001) or 2.0 (38%, P =.007) mg/kg/d achieved an ACR20 at week 12 than those receiving placebo (19%). Similar responses were noted at week 24. IL-1ra was generally well tolerated, with injection site reaction the most frequent adverse event. No serious adverse events attributable to IL-1ra treatment were observed.

CONCLUSIONS

These studies suggest an important role for IL-1ra as a novel therapeutic agent in the treatment of RA.

In both Crohn's disease (CD) and ulcerative colitis, the pathologic process is almost certainly driven by an aberrant local immune response directed against normal components of the bacterial microflora. Mucosal immune cells interact with nonimmune cells such as epithelial cells and fibroblasts to promote tissue damage; cytokines are essential mediators of this cross talk. Accumulating evidence now suggests that <em>interleukin</em>-21 (IL-21), the newest member of the common gamma-chain-dependent cytokine family, is a key component of the inflammatory cascade. IL-21 is highly produced by activated CD4+ lymphocytes in the inflamed gut of patients with CD, where it contributes to sustaining the ongoing Th1 inflammation. IL-21 also increases the secretion of extracellular matrix-degrading enzymes by fibroblasts and of MIP-3alpha by epithelial cells. Two other cytokines, IL-<em>27</em> and IL-32, may also be important in the inflammatory pathways that operate in IBD.

Nasopharyngeal carcinoma (NPC) is a common cancer among southern Chinese. The profile of gene expression in NPC cells is largely unknown. In this study, we have examined differential gene expression in non-malignant and malignant nasopharyngeal epithelial (NPE) cells using a cDNA array hybridization method. A total of 42 genes were identified to be expressed in either non-malignant and malignant NPE cells or both. Thirteen of these genes were overexpressed in malignant NPE cells. These includes nuclear factor (NF90), FOS-related antigen 1 (FRA- 1), cytoplasmic dynein light chain (HDLC1), replication factor C (RFC1), nucleoside diphosphate kinase B, UV excision repair protein (RAD23A), insulin-like growth factor receptor II, transcription initiation factor TFIID subunit (TAFII31), growth factor receptor-bound protein 2 (GRB2), UV excision repair protein (RAD23B), glutathione peroxidase, Y box binding protein 1 and heat shock protein 86. In contrast, expression of nine genes was suppressed in malignant NPE cells. These includes calgranulin A, calgranulin B, neutrophil activating protein (ENA-78), heat shock protein <em>27</em>, integrin beta-1, integrin beta-4, cyclin-dependent kinase inhibitor 1A (p21), <em>interleukin</em>-8 and tyrosine protein kinase receptor (RET). Differential expression of calgranulin A, calgraunlin B, ENA-78, FRA-1 and NF90 in non-malignant and malignant nasopharyngeal epithelial cells was confirmed by RT-PCR analysis.

BACKGROUND

Neuropeptide Y (NPY) is emerging as a modulator of communication between the brain and the immune system. However, in spite of increasing evidence that supports a role for NPY in the modulation of microglial cell responses to inflammatory conditions, there is no consistent information regarding the action of NPY on microglial phagocytic activity, a vital component of the inflammatory response in brain injury. Taking this into consideration, we sought to assess a potential new role for NPY as a modulator of phagocytosis by microglial cells.

METHODS

The N9 murine microglial cell line was used to evaluate the role of NPY in phagocytosis. For that purpose, an IgG-opsonized latex bead assay was performed in the presence of lipopolysaccharide (LPS) and an <em>interleukin</em>-1β (IL-1β) challenge, and upon NPY treatment. A pharmacological approach using NPY receptor agonists and antagonists followed to uncover which NPY receptor was involved. Moreover, western blotting and immunocytochemical studies were performed to evaluate expression of p38 mitogen-activated protein kinase (MAPK) and heat shock protein <em>27</em> (HSP<em>27</em>), in an inflammatory context, upon NPY treatment.

RESULTS

Here, we show that NPY inhibits phagocytosis of opsonized latex beads and inhibits actin cytoskeleton reorganization triggered by LPS stimulation. Co-stimulation of microglia with LPS and adenosine triphosphate also resulted in increased phagocytosis, an effect inhibited by an <em>interleukin</em>-1 receptor antagonist, suggesting involvement of IL-1β signaling. Furthermore, direct application of LPS or IL-1β activated downstream signaling molecules, including p38 MAPK and HSP<em>27</em>, and these effects were inhibited by NPY. Moreover, we also observed that the inhibitory effect of NPY on phagocytosis was mediated via Y1 receptor activation.

CONCLUSIONS

Altogether, we have identified a novel role for NPY in the regulation of microglial phagocytic properties, in an inflammatory context.