Feasibility of Using a Novel 2.45 GHz Double Short Distance Slot Coaxial Antenna for Minimally Invasive Cancer Breast Microwave Ablation Therapy: Computational Model, Phantom, and In Vivo Swine Experimentation.

Microwave ablation (MWA) by using coaxial antennas is a promising alternative for breast cancer treatment. A double short distance slot coaxial antenna as a newly optimized applicator for minimally invasive treatment of breast cancer is proposed. To validate and to analyze the feasibility of using this method in clinical treatment, a computational model, phantom, and breast swine in vivo experimentation were carried out, by using four microwave powers (50 W, 30 W, 20 W, and 10 W). The finite element method (FEM) was used to develop the computational model. Phantom experimentation was carried out in breast phantom. The in vivo experimentation was carried out in a 90 kg swine sow. Tissue damage was estimated by comparing control and treated micrographs of the porcine mammary gland samples. The coaxial slot antenna was inserted in swine breast glands by using image-guided ultrasound. In all cases, modeling, in vivo and phantom experimentation, and ablation temperatures (above 60°C) were reached. The in vivo experiments suggest that this new MWA applicator could be successfully used to eliminate precise and small areas of tissue (around 20-30 mm2). By modulating the power and time applied, it may be possible to increase/decrease the ablation area.

Tracheal and bronchial polymeric immunoglobulin secretory immune system (PISIS) development in a porcine model.

Polymeric immunoglobulins (pIgs) mucosal secretion is mediated by the pIg secretory immune system (PISIS), which is composed of J-chain (JC) and antibody (IgM/IgA) producing cells (JC-AbPC), pIg receptor (pIgR) epithelial cell expression and the efficient release of secretory Igs (SIgs) to the mucosal lumen. A poor development or disturbances in this system may cause higher infection susceptibility, as observed in young and elderly people. In spite of this system's importance, few detailed studies regarding its development have been described in the lower respiratory tract of humans. Because the porcine model has been reported as an option for translational medicine to humans, we studied the tracheal and bronchial PISIS development in healthy, non-vaccinated, SPF, miniature Vietnamese pigs from birth to adulthood using immunohistochemistry and ELISAs. Our results demonstrated that pIgR was present at birth, and its expression increased with age. In contrast, JC-AbPC were low in neonatal pigs; however, colostrum was a source of IgM, SIgA, total IgA and IgG in respiratory secretions (trachea and bronchoalveolar lavages, nasal secretion and saliva) in piglets. JC-AbPC steadily increased in post-weaned, young and adult pigs, correlating with considerable increases in secretory and total Igs in the trachea and bronchi. These data suggest a compensatory role of maternal Igs at the respiratory mucosa in the absence of a structured PISIS before weaning. Furthermore, monomeric Igs (IgG and IgA) may also play an important role in respiratory protection and deserves a more thorough study.

Partial protection and abomasal cytokine expression in sheep experimentally infected with Haemonchus contortus and pre-treated with Taenia hydatigena vesicular concentrate.

The abomasal expression of IL-2, IL-4, IL-6, IL-10 and IFNγ in lambs experimentally infected with Haemonchus contortus and its relationship to protection induced by a Taenia hydatigena larvae vesicular concentrate (ThLVC) were evaluated. The lambs that were only infected with H. contortus larvae showed a worm burden greater (p<0.05) than the lambs that received ThLVC prior to infection. Moreover, the lambs that received ThLVC showed a greater (p<0.05) number of blood eosinophils than the lambs that did not receive the ThLVC. In general, the lambs that received ThLVC prior to infection had a greater amount of eosinophils and mast cells and higher in situ expression of IFNγ, IL-2, IL-4, IL-6 and IL-10 in the abomasal wall than the lambs that were infected with H. contortus only or that received ThLVC (p<0.05) only. A higher expression of IL-2 and IFNγ in the submucosa compared to the abomasal mucosa and a higher expression of IL-4 in the abomasal mucosa compared to the submucosa was observed (p<0.05). These results suggest that there is a Th1 type response in the abomasal submucosa and a Th2 type response in in the abomasal mucosa. The amount of eosinophils and mast cells and the in situ expression of IFNγ, IL-2, IL-4 and IL-6 in the abomasal walls were negatively correlated with the worm burden (p<0.05). These results suggest that ThLVC is a non-specific immune stimulator for the abomasal immune response, and it is likely that the protection observed is the result of this effect.

Effects of Vitamin D Supplementation during the Induction and Progression of Osteoarthritis in a Rat Model.

Epidemiological studies correlate low levels of vitamin D with the osteoarthritis (OA) progression. Cytokines and metalloproteases play a major role in OA promoting the inflammation and degradation of the cartilage and can be induced through the Toll-like receptor (TLR) pathway. The aim of this study was to evaluate the protective effect of vitamin D supplementation on the development of osteoarthritis (OA) through examining the genetic regulation of TLRs, cytokines, and metalloproteases in chondrocytes as well as the wideness of cartilage in rats with OA. Our results demonstrate that the signaling through TLR-4 is a proinflammatory mechanism in osteoarthritis that drives the upregulation of MMP-3, IL-1ß, and TNF-α gene expression, leading to cartilage degradation and inflammation. Vitamin D supplementation had a protective effect during the onset but not during the chronic stage of OA in the rat model.

Cell death of chondrocytes is a combination between apoptosis and autophagy during the pathogenesis of Osteoarthritis within an experimental model.

The death of chondrocytes and the loss of extracellular matrix are the central features in cartilage degeneration during Osteoarthritis (OA) pathogenesis. The mechanism by which chondrocytes are removed in OA cartilage are still not totally defined, although previous reports support the presence of apoptotic as well as non apoptotic signals. In addition, in 2004 Roach and co-workers suggested the term "Chondroptosis" to design the type of cell death present in articular cartilage, which include the presence of some apoptotic and autophagic processes. To identify the mechanisms, as well as the chronology by which chondrocytes are eliminated during OA pathogenesis, we decided to evaluate apoptosis (by active caspase 3 and TUNEL signal) and autophagy (by LC3II molecule and cytoplasmic vacuolization) using Immunohistochemistry and Western blot techniques in an animal OA model. During OA pathogenesis, chondrocytes exhibit modifications in their death process in each zone of the cartilage. At early stages of OA, the death of chondrocytes starts with apoptosis in the superficial and part of the middle zones of the cartilage, probably as a consequence of a constant mechanical damage in the joint. As the degenerative process progresses, high incidence of active caspase 3 as well as LC3II expression are observed in the same cell, which indicate a combination of both death processes. In contrast, in the deep zone, due the abnormal subchondral bone ossification during the OA pathogenesis, apoptosis is the only mechanism observed.

Intranasal immunization using biphasic lipid vesicles as delivery systems for OmlA bacterial protein antigen and CpG oligonucleotides adjuvant in a mouse model.

The nasal mucosa is an important arm of the mucosal system since it is often the first point of contact for inhaled antigens. The ineffectiveness of the simple delivery of soluble antigens to mucosal membranes for immunization has stimulated extensive studies in appropriate delivery systems and adjuvants. We have evaluated biphasic lipid vesicles as a novel intranasal (i.n.) delivery system (designated as vaccine targeting adjuvant, VTA) containing bacterial antigens and CpG oligodeoxynucleotides (ODNs). Results show that administration of antigen and CpG ODNs in biphasic lipid vesicles resulted in greater induction of IgA levels in serum (P< 0.05) and mucosal antibody responses such as IgA in nasal secretions and lung (P< 0.01) after immunization with a combined subcutaneous (s.c.)/i.n. as compared to s.c./s.c. approach. Based on antibody responses, VTA formulations were found to be suitable as delivery systems for antigens and CpG ODNs by the intranasal route, resulting in a Th2-type of immune response, characterized by IgG1 and IL-4 production at the systemic level.

Development of intraepithelial cells in the porcine small intestine.

The number, phenotype, localisation and development of intraepithelial lymphocytes (IEL) from duodenum (Du) and ileum (Il) were studied by immunohistochemistry (IHC) and light and electron microscopy in unweaned (0-7 weeks old) and six months-old pigs. Developmental changes at birth showed that 38% of the total lymphocytes in the villi were IEL, mainly of the CD2+CD4-CD8- double negative (DN) phenotype. That proportion rose to over 50% at week 5 after birth, resembling adult proportion, although still with fewer cells than in adult pigs. CD4+ cells appeared relatively early in life although they were confined to the lamina propria (LP) and CD8+ cells were found only in low numbers. In the villi of adult animals, almost half of the total number of lymphocytes were IEL (49% Du, 52% Il). Over half of these IEL (52% Du, 53% Il) showed the CD2+CD4-CD8+ phenotype and were localized at the epithelium's basement membrane. Numerous (43% Du, 42% Il) DN IEL were found grouped at the enterocyte nucleus level and relatively few (5% in Du and Il) granular IEL were found apically in the epithelium. These proportions were homogeneously maintained along the villi's tip, middle and bottom, suggesting that the IEL may have their origin in the LP. Therefore, the IEL compartment in the porcine intestine develops slowly with age and is actually composed by a heterogeneous population of cells (null, DN and CD8+). These results may explain the increased susceptibility of young animals to disease during the lactation period and should be taken into account when functional studies are carried out with IEL. The quantitative results of this paper established a model for studies on the effect of age, diet, normal flora, infection and oral immunization on the IEL of the gut.

Regulation of mucosal immune responses in effector sites.

In human disease and rodent models, immune responses in the intestinal mucosa can be damaging. Damage is characterised by villus atrophy, crypt hyperplasia and reduced ability to digest and absorb nutrients. In normal individuals active responses to harmless environmental antigens associated with food and commensal bacteria are controlled by the development of immunological tolerance. Similar pathological changes occur in piglets weaned early from their mothers. Active immune responses to food antigens are observed in these piglets, and we and others have hypothesised that the changes occur as a result of transient allergic immune responses to novel food or bacteria antigens. The normal mechanism for producing tolerance to food antigens may operate at induction (Peyer's patches and mesenteric lymph nodes) or at the effector stage (intestinal lamina propria). In our piglet studies immunological tolerance occurs despite the initial active response. Together with evidence from rodents, this observation suggests that active responses are likely to be controlled at the effector stage, within the intestinal lamina propria. Support for this mechanism comes from the observation that human and pig intestinal T-cells are susceptible to apoptosis, and that this process is accelerated by antigen. We suggest that the role of the normal mature intestinal lamina propria is a balance between immunological effector and regulatory function. In neonatal animals this balance develops slowly and is dependant on contact with antigen. Immunological insults such as weaning may tip the balance of the developing mucosal immune system into excessive effector or regulatory function resulting in transient or chronic allergy or disease susceptibility.

Intragastric immunization of rats with Entamoeba histolytica trophozoites induces cecal mucosal IgE, eosinophilic infiltration, and type I hypersensitivity.

We have investigated the role of IgE in the local immunity of intestinal amebiasis, a parasitic infection known to induce specific antibody-forming cells (AFC) and IgA antibodies in rodents and humans. We found that intragastric immunization of rats with glutaraldehyde-fixed Entamoeba histolytica trophozoites significantly increased antiameba AFC in the Peyer's patches and spleen and that the lamina propria of the cecum from immunized animals was infiltrated by eosinophils armed with IgE antibodies. Morphometric analysis showed that IgE-containing cells and eosinophils were nearly three times more abundant in the cecum of immunized rats. Antigenic challenge with amebal lysates provoked an increase in the short-circuit current and in the transepithelial potential difference in Ussing-chambered cecum preparations from immunized rats. Although eosinophilia and the increase of IgE are common consequences of infection by parasitic worms, our results indicate that local immunity in intestinal amebiasis also involves IgE deposition, eosinophil infiltration, and type I hypersensitivity, which may explain some symptoms of amebic dysentery such as colic, abdominal tension, tenesmus, and bloody stools.

Effect of early weaning on the development of immune cells in the pig small intestine.

The controlled effects of age and weaning on the numbers of CD2+ T cells, subsets (CD4+, CD8+), accessory cells (macrophage/granulocyte) and cells expressing MHC class II (DQw) and IL-2R in the piglet intestine was investigated. At birth low numbers of CD2+CD4-CD8- cells were the only demonstrable T cells in the intestine. Monocyte/granulocyte and MHC class II+ cells were also detected in low numbers and IL-2R+ cells were proportionally quite numerous. All those cell populations, except the IL-2R+ cells, increased thereafter and peaked at Week 7 when the numbers of cells were comparable with those of adult animals. CD4+ cells increased dramatically after Week 1. In contrast, CD8+ remained scarce until after 5-7 weeks of age in unweaned animals. Four days after weaning at 3 weeks old, there were increases in CD2+ (P < 0.001) and macrophage/granulocyte (P < 0.01) cells in proximal small intestinal villi and in CD2+ cells only (P < 0.01) in crypts. No significant changes in cell numbers were demonstrated in the distal small intestine.

Immune cell distribution in the small intestine of the pig: immunohistological evidence for an organized compartmentalization in the lamina propria.

Using monoclonal antibodies in immunohistochemistry, the distribution of the cells with the following surface antigens was studied in samples of proximal and distal small intestine of five 6-month-old pigs: CD2, CD4 (helper/inducer T-cells), CD8 (suppressor/cytotoxic T cells), accessory cell marker (monocyte/granulocyte), MHC Class II (DRw), and interleukin 2 (IL-2) receptor. CD2+ cells were found in high numbers in both the epithelium and the lamina propria. More cells were demonstrated in villis than in crypts (proportion approximately 4:1). At least two subpopulations of intraepithelial lymphocytes were identified: apically in the epithelium there were CD2+CD4-CD8- (double negative) cells, whereas cells expressing CD8 marker were concentrated around the basement membrane. CD4+ cells were localized in the lamina propria towards the villus core. Accessory cells were distributed in crypts and the villus base and more cells were found in ileum than in duodenum. In contrast, MHC Class II+ cells were located predominantly in villi, just underneath the basement membrane, forming a sheath of cells between the CD8+ and the CD4+ cells. Cells expressing IL-2 receptor were sparse but widely distributed in both the lamina propria and the epithelium. This organized cell distribution may be related to the physiology of the mucosal immune system in the gut.