Cigarette smoke induces molecular responses in respiratory tissues of ApoE(-/-) mice that are progressively deactivated upon cessation.

Cigarette smoking is the primary etiology of chronic obstructive pulmonary disease (COPD) and a risk factor for both lung and cardiovascular (CV) diseases, which are rarely investigated concomitantly. Although smoking cessation shows clear CV risk benefit, lung-related disease risk remains higher in former smokers than in never smokers. We sought to determine the differential molecular responses of murine respiratory tissues to better understand the toxicity pathways involved in smoking-related disease risk and those related to the benefits of smoking cessation. ApoE(-/-) mice were exposed to mainstream cigarette smoke (CS) or a smoking cessation-mimicking protocol for up to 6 months and transcriptomics analysis of nasal epithelium and lung parenchyma performed. We supported our gene expression profiling approach with standard lung histopathology and bronchoalveolar lavage fluid (BALF) analysis. Many BALF analytes involved in functions ranging from inflammation to cell proliferation and tissue remodeling were found elevated in BALF. Gene expression levels of these molecules were also increased in lung tissue, suggesting that the inflammatory response was the result of local tissue activation and the contribution of recruited inflammatory cells. Gene set enrichment analysis (GSEA) of expression data from murine lungs and nasal epithelium showed distinct activation patterns of inflammation, complement, and xenobiotic metabolism pathways during CS exposure that were deactivated upon smoking cessation. Pathways involved in cell proliferation and tissue remodeling were activated by CS and progressively deactivated upon smoke exposure cessation. Differential CS-mediated responses of pulmonary and nasal tissues reflect common mechanisms but also the varying degrees of epithelial functional specialization and exposure along the respiratory tract.

Lung inflammatory effects, tumorigenesis, and emphysema development in a long-term inhalation study with cigarette mainstream smoke in mice.

Cigarette smoking is the leading cause of lung cancer and chronic obstructive pulmonary disease, yet there is little mechanistic information available in the literature. To improve this, laboratory models for cigarette mainstream smoke (MS) inhalation-induced chronic disease development are needed. The current study investigated the effects of exposing male A/J mice to MS (6h/day, 5 days/week at 150 and 300 mg total particulate matter per cubic meter) for 2.5, 5, 10, and 18 months in selected combinations with postinhalation periods of 0, 4, 8, and 13 months. Histopathological examination of step-serial sections of the lungs revealed nodular hyperplasia of the alveolar epithelium and bronchioloalveolar adenoma and adenocarcinoma. At 18 months, lung tumors were found to be enhanced concentration dependently (up to threefold beyond sham exposure), irrespective of whether MS inhalation had been performed for the complete study duration or was interrupted after 5 or 10 months and followed by postinhalation periods. Morphometric analysis revealed an increase in the extent of emphysematous changes after 5 months of MS inhalation, which did not significantly change over the following 13 months of study duration, irrespective of whether MS exposure was continued or not. These changes were found to be accompanied by a complex pattern of transient and sustained pulmonary inflammatory changes that may contribute to the observed pathogeneses. Data from this study suggest that the A/J mouse model holds considerable promise as a relevant model for investigating smoking-related emphysema and adenocarcinoma development.

The transcriptome of Nrf2-/- mice provides evidence for impaired cell cycle progression in the development of cigarette smoke-induced emphysematous changes.

Cigarette smoke (CS) imposes a strong oxidative burden on exposed tissues resulting in a severely disturbed oxidant/antioxidant balance, which in the context of chronic exposure is assumed to be a key contributor to CS-related diseases. Because of its emerging central role in orchestrating the general cellular antioxidant response, the pathway leading to the activation of the transcription factor Nrf2 has received mounting attention over the past decade in investigations aimed at elucidating CS-induced pathophysiological mechanisms. To comprehensively characterize the impact of Nrf2 in acute and subchronic smoking scenarios, Nrf2(-/-) mice and their wild-type (wt) ICR littermates were exposed to either ambient air (sham exposure) or one of three doses of CS for up to 5 months, with two postexposure endpoints of 1 and 13 days. The lungs of the mice were monitored for transcriptomic changes on a genome-wide level, which confirmed an impaired expression of antioxidant and phase 2-related genes in CS-exposed Nrf2(-/-) mice. Importantly, in comparison to wt mice, an attenuated cell cycle/mitotic response and intensified stress gene expression pattern were observed in exposed Nrf2(-/-) mice, which was paralleled by clear dose-dependent effects on alveolar destruction and impaired lung function. In contrast, the inflammation-related transcriptional response and scores for various bronchioalveolar inflammation parameters were qualitatively and quantitatively similar in CS-exposed mice of both genotypes. Taken together, these results confirm the protective nature of Nrf2 in oxidative stress scenarios and suggest that the enhanced emphysematous phenotype exhibited by CS-exposed Nrf2(-/-) mice is more likely caused by an imbalance in cell loss and regeneration than by increased inflammation.

Effect of adjuvant on reactogenicity and long-term immunogenicity of the malaria Vaccine ICC-1132 in macaques.

ICC-1132 is a malaria vaccine candidate based on a modified hepatitis B virus core particle (HBc) bearing putative protective epitopes from the circumsporozoite protein (CS) of Plasmodium falciparum. While the epitope carrier itself is immunogenic, its potency can be increased by formulation with adjuvants. As a prelude to Phase I clinical trials, rhesus macaques were immunised twice with GMP grade ICC--1132 in saline or formulated with the adjuvants Alhydrogel (Alhydrogel) or Montanide((R)) ISA 720 (Montanide). Both adjuvant formulations gave significant humoral responses after the first injection, with titres increasing further after the second dose. The Montanide formulation was the most immunogenic, but undesirable reactogenicity in the form of sterile abscesses was associated with higher dosage levels of ICC--1132. These side effects could be avoided with lower antigen load, or by formulation of the second dose in Alhydrogel. Such measures also reduced peak titres and longevity of antibodies against CS, demonstrating the delicate balance between immunogenicity and reactogenicity of new vaccine formulations.

Protection of macaques against Mycobacterium tuberculosis infection by a subunit vaccine based on a fusion protein of antigen 85B and ESAT-6.

Various new tuberculosis (TB) vaccine candidates in combination with new delivery systems, including subunit vaccines, are currently being evaluated by a number of laboratories. One vaccine candidate that has shown promising protective capacity in mice and guinea pigs is a fusion of Ag85B and ESAT-6. In this study, we have investigated the efficacy of this Ag85B-ESAT-6 fusion protein vaccine in a non-human primate model for TB. Vaccination of cynomolgus monkeys with the Ag85B-ESAT-6 fusion protein in two different adjuvant (DDA/MPL, AS02A) resulted in a reduction in bacterial number and/or lung pathology in animals challenged with Mycobacterium tuberculosis. Vaccination prevented an increase in C-reactive protein serum levels, general activation of CD4 and CD8 subsets and boosted development of humoral and cellular immune responses to a spectrum of mycobacterial antigens on exposure to M. tuberculosis infection. We show, in two independent experiments, that vaccination of primates with Ag85B-ESAT-6 induces protective immune responses, suggesting that Ag85B-ESAT-6 is a strong candidate for further clinical evaluation. As far as we are aware this is the first report of protection in primates with a subunit vaccine.

High-resolution X-ray microtomography for the detection of lung tumors in living mice.

In the present study, the feasibility of applying high-resolution microtomography (micro-CT) for the detection of lung tumors was investigated in live mice at an early and more advanced stage of tumor development. The chest area of anesthesized mice was scanned by X-ray micro-CT. In mice with a minor and heavy tumor load, micro-CT proved to be a fast and noninvasive imaging device for the detection of lung tumors. After validation of the CT data by histologic sectioning, it was shown that the majority of tumors could be distinguished in the reconstructed virtual slices obtained by micro-CT. The data from micro-CT were also confirmed by visual inspection of the inflated and excised lungs postmortem. In vivo micro-CT opens broad perspectives for imaging tumor development and its progression in a noninvasive way. Micro-CT also allows for longitudinal evaluation of the treatment of lung cancer by drugs.

Experimental infection of the olive baboon (Paplio anubis) with Plasmodium knowlesi: severe disease accompanied by cerebral involvement.

Experimental systems that model some of the complex interactions between parasite and host can be extremely valuable in identifying and developing new prophylactics and therapeutics against human diseases. Because primates have similar immune systems to humans, we have characterized a baboon model for understanding host response to Plasmodium knowlesi. Ten intact olive baboons (Papio anubis) of either sex were experimentally infected with P. knowlesi H strain erythrocytic parasites. The infection in these baboons was either acute or chronic. Animals with acute infection developed multiple system organ dysfunction and cerebral involvement. In chronically infected animals, only the spleen was moderately enlarged. The P. knowlesi parasitemia profile in baboons and rhesus monkeys was comparable. However, some clinical symptoms of the baboons and P. falciparum-infected humans were similar. These studies demonstrate for the first time that P. anubis is a suitable host for P. knowlesi for studying clinical symptoms and pathology.