Epidemiologic studies demonstrate acute and serious adverse effects of particulate air pollution on respiratory health, especially in people who are susceptible to bacterial infection. However, the underlying mechanism remains to be elucidated. To provide experimental evidence for the epidemiologic data, we determined the effects of diesel exhaust particles (DEP), major participants in particulate pollutants, on lung injury related to bacterial endotoxin in mice. Intratracheal instillation of DEPs synergistically enhanced lung injury related to endotoxin from gram-negative bacteria, which was characterized by neutrophil sequestration, interstitial edema, and alveolar hemorrhage. In the presence of endotoxin, DEPs further activated the nuclear translocation of p65 subunit of nuclear factor-kappaB (NF-kappaB) in the lung and increased the lung expression of intercellular adhesion molecule-1, interleukin-1beta, macrophage chemoattractant protein-1, keratinocyte chemoattractant (KC), macrophage inflammatory protein-1alpha, and Toll-like receptors. DEPs given alone increased the lung expression of Toll-like receptor 4 and the nuclear localization of p50 subunit of NF-kappaB. The combined exposure to DEPs and endotoxin decreased nuclear localization of CCAAT/enhancer binding protein beta. These results provide the first experimental evidence that DEPs enhance neutrophilic lung inflammation related to bacterial endotoxin. The enhancement is mediated by the induction of proinflammatory molecules, likely through the expression of Toll-like receptors and the activation of p65-containing dimer(s) of NF-kappaB, such as p65/p50.

DEP-OC, rather than washed DEP, exaggerated allergic airway inflammation through the enhancement of T-helper type 2 responses. The coexistence of OC with carbonaceous nuclei caused the most remarkable aggravation. DEP components might diversely affect various types of respiratory diseases, while whole DEP might mostly aggravate respiratory diseases.

SUMMARYThe effect of prenatal exposure to bisphenol A (BPA) on the immune system in mice was investigated. Virgin female mice were fed varying doses of BPA, on a daily basis, over a period of 18 days commencing on the day of pairing with stud males (day 0). On day 77, their male offspring of 8 weeks were immunized with hen egg lysozyme (HEL). Three weeks later, anti-HEL immunoglobulin G (IgG) in sera, and proliferative responses of spleen cells to the antigen, were measured. Anti-HEL IgG2a and interferon-c (IFN-c), secreted from splenic lymphocytes, were measured as indicators of T helper 1 (Th1) immune responses, while anti-HEL IgG1 and interleukin-4 (IL-4) were measured as indicators of Th2 responses. The results showed that fetal exposure to BPA was followed by significant increases in anti-HEL IgG as well as antigenspecific cell proliferation. Both Th1 responses (including anti-HEL IgG2a and IFN-c production) and Th2 responses (including anti-HEL IgG1 and IL-4 production) were augmented by prenatal exposure to BPA, although the augmentation of Th1 responses appeared to be greater than that of Th2 responses. Two-colour flow cytometric analysis showed that mice exposed prenatally to BPA had 29% and 100% more splenic CD3 + CD4 + and CD3 + CD8 + cells, respectively, than control animals. Similar results were obtained from females whose mothers had consumed BPA during pregnancy. These results suggest that prenatal exposure to BPA may result in the up-regulation of immune responses, especially Th1 responses, in adulthood.

Neutrophilic airway inflammation is a hallmark of patients with severe asthma. Although we have reported that both IL-33 and IL-17A contributed to IgE-mediated neutrophilic inflammation in mice, the relationship remains unclear. In this article, we examined how IL-17A modifies IL-33–induced neutrophilic inflammation and airway hyperresponsiveness (AHR). IL-33 was intratracheally administered to BALB/c mice on days 0–2; furthermore, on day 7, the effect of the combination of IL-33 and IL-17A was evaluated. Compared with IL-33 or IL-17A alone, the combination exacerbated neutrophilic inflammation and AHR, associated with more increased levels of lung glutamic acid-leucine-arginine+ CXC chemokines, including CXCL1, CXCL2, and CXCL5, and infiltration by alveolar macrophages expressing CXCR2. Treatment with anti-CXCR2 mAb or depletion of alveolar macrophages repressed neutrophilic inflammation and AHR; in addition, depletion of neutrophils suppressed AHR. These findings prompted us to examine the role of CXCR2 in IgE-sensitized mice; a single treatment with anti-CXCR2 mAb in the seventh Ag challenge inhibited late-phase airway obstruction, AHR, and neutrophilic inflammation. In addition to inhibition, multiple treatments during the fourth to seventh challenge attenuated early-phase airway obstruction, eosinophilic inflammation, and goblet cell hyperplasia associated with the reduction of Th2 cytokine production, including IL-4, IL-5, and IL-13. Collectively, IL-33 cooperated with IL-17A to exacerbate AHR by enhancing neutrophilic inflammation via CXCR2 signaling; furthermore, CXCR2 signaling derived Th2 responses. We thus suggest the underlying mechanisms of IL-33 and IL-17A in allergic asthma and CXCR2 as potential therapeutic targets for the disease.

BackgroundWe previously conducted a phase I trial for advanced colorectal cancer (CRC) using five HLA-A*2402-restricted peptides, three derived from oncoantigens and two from vascular endothelial growth factor (VEGF) receptors, and confirmed safety and immunological responses. To evaluate clinical benefits of cancer vaccination treatment, we conducted a phase II trial using the same peptides in combination with oxaliplatin-based chemotherapy as a first-line therapy.MethodsThe primary objective of the study was the response rates (RR). Progression free survival (PFS), overall survival (OS), and immunological parameters were evaluated as secondary objective. The planned sample size was more than 40 patients for both HLA2402-matched and -unmatched groups. All patients received a cocktail of five peptides (3 mg each) mixed with 1.5 ml of IFA which was subcutaneously administered weekly for the first 12 weeks followed by biweekly administration. Presence or absence of the HLA-A*2402 genotype were used for classification of patients into two groups.ResultsBetween February 2009 and November 2012, ninety-six chemotherapy naïve CRC patients were enrolled under the masking of their HLA-A status. Ninety-three patients received mFOLFOX6 and three received XELOX. Bevacizumab was added in five patients. RR was 62.0% and 60.9% in the HLA-A*2402-matched and -unmatched groups, respectively (p = 0.910). The median OS was 20.7 months in the HLA-A*2402-matched group and 24.0 months in the unmatched group (log-rank, p = 0.489). In subgroup with a neutrophil/lymphocyte ratio (NLR) of < 3.0, patients in the HLA-matched group did not survive significantly longer than those in the unmatched group (log-rank, p = 0.289) but showed a delayed response.ConclusionsAlthough no significance was observed for planned statistical efficacy endpoints, a delayed response was observed in subgroup with a NLR of < 3.0. Biomarkers such as NLR might be useful for selecting patients with a better treatment outcome by the vaccination.Trial registrationTrial registration: UMIN000001791.

Allergic asthma is a chronic inflammatory disorder of the airways characterized by biphasic airway obstruction and airway hyperresponsiveness. In this study, we attempted to elucidate the contribution of the complement C3a to these asthmatic symptoms. BALB/c mice sensitized by i.p. injections of OVA plus alum were challenged with OVA intratracheally four times. The fourth challenge caused a biphasic asthmatic response peaking at 10 min and 3–4 h, as well as airway hyperresponsiveness to methacholine. Histological examination revealed increased expression of C3a receptors in the lung on the fourth challenge. Additionally, the C3 level in serum 4 h after the fourth challenge was significantly reduced compared with that before the challenge. When a C3a receptor antagonist, SB290157, was administered i.p. 30 min before the fourth challenge, the late-phase asthmatic response and airway hyperresponsivness induced by the fourth challenge were significantly inhibited, although the early-phase response was not influenced. In bronchoalveolar lavage fluid, neutrophil infiltration 24 h after the fourth challenge was reduced by the treatment. On the other hand, SB290157 suppressed the increased expression of IL-1β in the lung in this model, and the intratracheal administration of IL-1β induced airway obstruction, airway hyperresponsiveness, and neutrophil infiltration in normal mice. These results illustrate that C3a is involved in the development of the late asthmatic response and airway hyperresponsiveness. The mechanism leading to the development of these symptoms may correlate with the recruitment of neutrophils and/or the production of IL-1β induced by C3a.

BackgroundTo evaluate the safety of combination vaccine treatment of multiple peptides, phase I clinical trial was conducted for patients with advanced colorectal cancer using five novel HLA-A*2402-restricted peptides, three peptides derived from oncoantigens, ring finger protein 43 (RNF43), 34 kDa-translocase of the outer mitochondrial membrane (TOMM34), and insulin-like growth factor–II mRNA binding protein 3 (KOC1), and the remaining two from angiogenesis factors, vascular endothelial growth factor receptor 1 (VEGFR1) and VEGFR2.MethodsEighteen HLA- A*2402-positive colorectal cancer patients who had failed to standard therapy were enrolled in this study. 0.5 mg, 1.0 mg or 3.0 mg each of the peptides was mixed with incomplete Freund’s adjuvant and then subcutaneously injected at five separated sites once a week. We also examined possible effect of a single site injection of “the cocktail of 5 peptides” on the immunological responses. ELISPOT assay was performed before and after vaccinations in the schedule of every 4 weeks.ResultsThe vaccine treatment using multiple peptides was well tolerated without any severe treatment-associated systemic adverse events. Dose-dependent induction of peptide-specific cytotoxic T lymphocytes was observed. The single injection of “peptides cocktail” did not diminish the immunological responses. Regarding the clinical outcome, one patient achieved complete response and 6 patients revealed stable disease for 4 to 7 months. The median overall survival time (MST) was 13.5 months. Patients, in which we detected induction of cytotoxic T lymphocytes specific to 3 or more peptides, revealed significantly better prognosis (MST; 27.8 months) than those with poorer immune responses (MST; 3.7 months) (p = 0.032).ConclusionOur cancer vaccine treatment using multiple peptides is a promising approach for advanced colorectal cancer with the minimum risk of systemic adverse reactions.Clinical trial registrationUMIN-CTR number UMIN000004948.

BackgroundCD3 +  and CD8 + T-cell infiltration were reported as positive predictive markers of survival in colorectal cancer (CRC) patients. Here, we demonstrate the prognostic significance of CD4 + and FOXP3 + T-cell densities in CRC.MethodsWe quantified the intratumoural densities of CD3 + , CD8 + , CD4 +  and FOXP3 + T cells by immunohistochemistry and digital pathology in 342 CRC patients who underwent curative resection. Microsatellite instability was also assessed in 322 specimens. Patient demographics, clinicopathological features and survival rates were analysed.ResultsHigh CD3 + , CD4 +  and FOXP3 + T-cell densities were associated with improved relapse-free survival (RFS); high CD8 + , CD4 +  and FOXP3 + T-cell densities were associated with improved disease-specific survival (DSS). Patients with low CD4 + and low FOXP3 + T-cell densities exhibited extremely poor prognoses. T stage, vascular/lymphatic invasion and CD4 + T-cell density were independent prognostic indicators for DSS. The distributions of CD4 + and FOXP3 + T-cell densities were not significantly different between the high microsatellite instability group and other groups, in contrast to those of CD3 + and CD8 + T-cell densities.ConclusionsIntratumoural CD4 + T-cell density and combined CD4 + and FOXP3 + T-cell densities were stronger prognostic indicators than other clinicopathological features. These results may facilitate the establishment of novel prognostic factors and therapeutic strategies for CRC.