Local Immune Changes in Early Stages of Inflammation and Carcinogenesis Correlate with the Collagen Scaffold Changes of the Colon Mucosa.

Continuous activation of the immune system inside a tissue can lead to remodelling of the tissue structure and creation of a specific microenvironment, such as during the tumour development. Chronic inflammation is a central player in stimulating changes that alter the tissue stroma and can lead to fibrotic evolution. In the colon mucosa, regulatory mechanisms, including TGF-ß1, avoid damaging inflammation in front of the continuous challenge by the intestinal microbiome. Inducing either DSS colitis or AOM colorectal carcinogenesis in AVN-Wistar rats, we evaluated at one month after the end of each treatment whether immunological changes and remodelling of the collagen scaffold were already in development. At this time point, we found in both models a general downregulation of pro-inflammatory cytokines and even of TGF-ß1, but not of IL-6. Moreover, we demonstrated by multi-photon microscopy the simultaneously presence of pro-fibrotic remodelling of the collagen scaffold, with measurable changes in comparison to the control mucosa. The scaffold was significantly modified depending on the type of induced stimulation. These results suggest that at one month after the end of the DSS or AOM inductions, a smouldering inflammation is present in both induced conditions, since the pro-inflammatory cytokines still exceed, in proportion, the local homeostatic regulation of which TGF-ß1 is a part (inflammatory threshold). Such an inflammation appears sufficient to sustain remodelling of the collagen scaffold that may be taken as a possible pathological marker for revealing pre-neoplastic inflammation.

Immune activation by microbiome shapes the colon mucosa: Comparison between healthy rat mucosa under conventional and germ-free conditions.

Germ-free animals (GF) are those without a microbiome since birth. This particular biological model has become one of special interest with the growing evidence of importance of the microbiome in the life, development, adaptation, and immunity of humans and animals in the environments in which they live. Anatomical differences observed in GF compared with conventionally-reared animals (CV) has given rise to the question of the influence of commensal microflora on the development of structure and function (even immunological) of the bowel. Only recently, thanks to achievements in microscopy and associated methods, structural differences can be better evaluated and put in perspective with the immunological characteristics of GF vs. CV animals. This study, using a GF rat model, describes for the first time the possible influence that the presence of commensal microflora, continuously stimulating mucosal immunity, has on the collagen scaffold organization of the colon mucosa. Significant differences were found between CV and GF mucosa structure with higher complexity in the CV rats associated to a more activated immune environment. The immunological data suggest that, in response to the presence of a microbiome, an effective homeostatic regulation in developed by the CV rats in healthy conditions to avoid inflammation and maintain cytokine levels near the spontaneous production found in the GF animals. The results indicated that collagen scaffold adapted to the immune microenvironment; therefore, it is apparent that the microbiome was able to condition the structure of the colon mucosa.

(Anti)mutagenic and immunomodulatory properties of quercetin glycosides.

BACKGROUND: Quercetin-3-O-ß-D-glucopyranoside (isoquercitrin) and quercetin-3-O-rutinoside (rutin) are common components of a normal human diet and are increasingly used in food supplements. Here their effect on mutagenesis and immunity is shown. RESULTS: The in vitro (anti)mutagenic potential was compared with that of quercetin using the Ames test in Salmonella typhimurium His(-) strains TA100, TA98 and TA102. Isoquercitrin only slightly increased the number of revertants, while rutin was totally non-mutagenic. On the other hand, all compounds displayed dose-dependent protective activity against H2O2 - and tert-butyl hydroperoxide-induced oxidative damage to the TA102 strain and at 75 µmol L(-1) inhibited H2O2/Fe(2+)-induced formation of the open circular and linear forms of the DNA plasmid pBSIISK(-). In mice, none of the flavonols (0.86 µmol day(-1), 34 days) induced harmful effects. In immunized animals, all compounds enhanced ex vivo B cell proliferation; quercetin stimulated lymphocyte basal proliferation and increased the number of IgM-producing lymphocytes. Rutin promoted NK cytotoxic activity, supported T cells and enhanced gut epithelium renewal. No effect on IgG-forming cells was found. CONCLUSION: Isoquercitrin displayed negligible and rutin no mutagenicity, but both showed significant antimutagenic and DNA-protective effects against oxidative damage. In vivo, they supported the readiness of the immune system for specific humoral immune response.

In Vivo Targeting of Cutaneous Melanoma Using an Melanoma Stimulating Hormone-Engineered Human Protein Cage with Fluorophore and Magnetic Resonance Imaging Tracers.

Nanoparticle (NP)-based materials are promising agents for enhancing cancer diagnosis and treatment. Once functionalized for selective targeting of tumor-expressed molecules, they can specifically deliver drugs and diagnostic molecules inside tumor cells. In the present work, we evaluated the in vivo melanoma-targeting ability of a nanovector (HFt-MSH-PEG) based on human protein ferritin (HFt), functionalized with both melanoma-targeting melanoma stimulating hormone (α-MSH) and stabilizing poly(ethylene glycol) (PEG) molecules. Independent and complementary techniques, such as whole-specimen confocal microscopy and magnetic resonance imaging, were used to detect in vivo localization of NP constructs with suitable tracers (i.e., fluorophores or magnetic metals). Targeted HFt-MSH-PEG NPs accumulated persistently at the level of primary melanoma and with high selectivity with respect to other organs. Melanoma localization of untargeted HFt-PEG NPs, which lack the α-MSH moiety, was less pronounced. Furthermore, HFt-MSH-PEG NPs accumulated to a significantly lower extent and with a different distribution in a diverse type of tumor (TS/A adenocarcinoma), which does not express α-MSH receptors. Finally, in a spontaneous lung metastasis model, HFt-MSH-PEG NPs localized at the metastasis level as well. These results suggest that HFt-MSH-PEG NPs are suitable carriers for selective in vivo delivery of diagnostic or therapeutic agents to cutaneous melanoma.

Microbiome and colorectal carcinoma: insights from germ-free and conventional animal models.

The mammalian microbiota plays a crucial role in the pathogenesis of many diseases. Thanks to recent advances in metagenomics, proteomics, and metabolomics, microbiome composition and metabolic activity can now be studied in detail. Results obtained by such fascinating and provocative studies would be meaningless without considering the perspective of the whole organism. Our work using gnotobiology as the major tool to unravel the mechanisms of host-microbe interaction has demonstrated the crucial role of microbiota in the initiation and progression of inflammation-associated colorectal neoplasia. Carcinogenesis in the gut is driven by the presence of potentially harmful microbes or by lack of protective ones, by the production of carcinogens generated by microbes, and by the induction of inflammation and modulation of the immune system. Here, we review these mechanisms with special emphasis on those where gnotobiology has yielded important insights.

Immunostimulatory properties and antitumor activities of glucans (Review).

New foods and natural biological modulators have recently become of scientific interest in the investigation of the value of traditional medical therapeutics. Glucans have an important part in this renewed interest. These fungal wall components are claimed to be useful for various medical purposes and they are obtained from medicinal mushrooms commonly used in traditional Oriental medicine. The immunotherapeutic properties of fungi extracts have been reported, including the enhancement of anticancer immunity responses. These properties are principally related to the stimulation of cells of the innate immune system. The discovery of specific receptors for glucans on dendritic cells (dectin-1), as well as interactions with other receptors, mainly expressed by innate immune cells (e.g., Toll-like receptors, complement receptor-3), have raised new attention toward these products as suitable therapeutic agents. We briefly review the characteristics of the glucans from mycelial walls as modulators of the immunity and their possible use as antitumor treatments.

Selective targeting of melanoma by PEG-masked protein-based multifunctional nanoparticles.

BACKGROUND: Nanoparticle-based systems are promising for the development of imaging and therapeutic agents. The main advantage of nanoparticles over traditional systems lies in the possibility of loading multiple functionalities onto a single molecule, which are useful for therapeutic and/or diagnostic purposes. These functionalities include targeting moieties which are able to recognize receptors overexpressed by specific cells and tissues. However, targeted delivery of nanoparticles requires an accurate system design. We present here a rationally designed, genetically engineered, and chemically modified protein-based nanoplatform for cell/tissue-specific targeting. METHODS: Our nanoparticle constructs were based on the heavy chain of the human protein ferritin (HFt), a highly symmetrical assembly of 24 subunits enclosing a hollow cavity. HFt-based nanoparticles were produced using both genetic engineering and chemical functionalization methods to impart several functionalities, ie, the α-melanocyte-stimulating hormone peptide as a melanoma-targeting moiety, stabilizing and HFt-masking polyethylene glycol molecules, rhodamine fluorophores, and magnetic resonance imaging agents. The constructs produced were extensively characterized by a number of physicochemical techniques, and assayed for selective melanoma-targeting in vitro and in vivo. RESULTS: Our HFt-based nanoparticle constructs functionalized with the α-melanocyte-stimulating hormone peptide moiety and polyethylene glycol molecules were specifically taken up by melanoma cells but not by other cancer cell types in vitro. Moreover, experiments in melanoma-bearing mice indicate that these constructs have an excellent tumor-targeting profile and a long circulation time in vivo. CONCLUSION: By masking human HFt with polyethylene glycol and targeting it with an α-melanocyte-stimulating hormone peptide, we developed an HFt-based melanoma-targeting nanoplatform for application in melanoma diagnosis and treatment. These results could be of general interest, because the same strategy can be exploited to develop ad hoc nanoplatforms for specific delivery towards any cell/tissue type for which a suitable targeting moiety is available.

Orally administered marine (1-->3)-beta-D-glucan Phycarine stimulates both humoral and cellular immunity.

(1-->3)-beta-D-Glucans represent highly conserved structural components of cell walls in yeast, fungi, or seaweed. However, it is still unknown how they mediate their effects. The aim of this study was to evaluate both intraperitoneal and oral application of seaweed-derived (1-->3)-beta-D-glucan Phycarine. Phycarine showed significant stimulation of phagocytosis by peripheral blood cells. In addition, the efficiency of chemotherapy of Lewis lung carcinoma with cyclophosphamide was potentiated by Phycarine administration. Phycarine also strongly shortened the recovery of leucopenia caused either by chemotherapy or irradiation. Besides the role in stimulation of cellular immunity, we also found a significant increase of antibody formation. Using a suckling rat model for evaluation of the absorption and tissues distribution of enterally administered (125)I-Phycarine, we found that the majority of Phycarine was detected in the stomach and duodenum 5 min after the administration. This amount sharply decreased during first 30 min. A significant amount of Phycarine entered proximal intestine in a shortly after the gavage. Its transit through proximal intestine was decreasing with time and simultaneously increasing in the ileum. Systemic blood levels were very low (less than 0.5%). Taken together, these observations suggest that Phycarine is similarly effective both after i.p. and oral application, has very strong stimulating effects on three types of experimentally induced leucopenia and stimulates both humoral and cellular branch of immune reactions. The majority of Phycarine can be detected throughout the gastrointestinal tract, supporting the feasibility of enteral administration of Phycarine in the treatment of gastrointestinal diseases.