Isolation and immunocharacterization of lactobacillus salivarius from the intestine of wakame-fed pigs to develop novel "Immunosynbiotics"

Emerging threats of antimicrobial resistance necessitate the exploration of effective alternatives for healthy livestock growth strategies. ?Immunosynbiotics?, a combination of immunoregulatory probiotics and prebiotics with synergistic effects when used together in feed, would be one of the most promising candidates. Lactobacilli are normal residents of the gastrointestinal tract of pigs, and many of them are able to exert beneficial immunoregulatory properties. On the other hand, wakame (Undaria pinnafida), an edible seaweed, has the potential to be used as an immunoregulatory prebiotic when added to livestock feed. Therefore, in order to develop a novel immunosynbiotic, we isolated and characterized immunoregulatory lactobacilli with the ability to utilize wakame. Following a month-long in vivo wakame feeding trial in 8-week-old Landrace pigs (n = 6), sections of intestinal mucous membrane were processed for bacteriological culture and followed by identification of pure colonies by 16S rRNA sequence. Each isolate was characterized in vitro in terms of their ability to assimilate to the wakame and to differentially modulate the expression of interleukin-6 (IL-6) and interferon beta (IFN-β) in the porcine intestinal epithelial (PIE) cells triggered by Toll-like receptor (TLR)-4 and TLR-3 activation, respectively. We demonstrated that feeding wakame to pigs significantly increased the lactobacilli population in the small intestine. We established a wakame-component adjusted culture media that allowed the isolation and characterization of a total of 128 Lactobacilli salivarius colonies from the gut of wakame-fed pigs. Interestingly, several L. salivarius isolates showed both high wakame assimilation ability and immunomodulatory capacities. Among the wakame assimilating isolates, L. salivarius FFIG71 showed a significantly higher capacity to upregulate the IL-6 expression, and L. salivarius FFIG131 showed significantly higher capacity to upregulate the IFN-β expression; these could be used as immunobiotic strains in combination with wakame for the development of novel immunologically active feeds for pigs.Fil: Masumizu, Yuki. Tohoku University; JapónFil: Zhou, Binghui. Tohoku University; JapónFil: Humayun Kober, AKM. Tohoku University; Japón. Chittagong Veterinary and Animal Sciences University; BangladeshFil: Islam, M. Aminul. Agricultural University; Bangladesh. Tohoku University; JapónFil: Iida, Hikaru. Tohoku University; JapónFil: Ikeda-Ohtsubo, Wakako. Tohoku University; JapónFil: Suda, Yoshihito. Department Of Food Agriculture, Miyagi University; JapónFil: Albarracín, Leonardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; Japón. Universidad Nacional de Tucumán; ArgentinaFil: Nochi, Tomonori. Tohoku University; JapónFil: Aso, Hisashi. Tohoku University; JapónFil: Suzuki, Keiichi. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Kitazawa, Haruki. Tohoku University; Japó

Siglec-F is a novel intestinal M cell marker

Intestinal microfold (M) cells are epithelial cells primarily present on Peyer's patches (PPs) in the small intestine. The ability of M cells to shuttle antigens into the PP for appropriate immune responses makes M cells a target for next-generation oral vaccine delivery. In this regard, discovery of M cell specific receptors are of great interest, which could act as molecular tags for targeted delivery of cargo to M cells. Here, using a monoclonal antibody we generated to the Sialic acid-binding immunoglobulin-like lectin F (Siglec-F), we show that Siglec-F is expressed on mouse M cells in the small intestine. Immunohistochemical analysis of the PP tissue sections shows that Siglec-F is expressed on the surface of the M cell membrane exposed to the intestinal lumen. Anti-Siglec-F antibody injected into the mouse small intestine bound to M-cells, demonstrating the potential to target M cells via Siglec-F

Relevant Assay to Study the Adhesion of Plasmodium falciparum-Infected Erythrocytes to the Placental Epithelium

In placental malaria, Plasmodium falciparum-infected erythrocytes adhere to the apical plasma membrane of the placental epithelium, triggering an impairment of placental function detrimental to the fetus. The design of anti-adhesion intervention strategies requires a detailed understanding of the mechanisms involved. However, most adhesion assays lack in vivo relevance and are hardly quantitative. Here, we describe a flow cytometry-based adhesion assay that is fully relevant by using apical epithelial plasma membrane vesicles as the adhesion matrix, and being applicable to infected erythrocytes directly isolated from patients. Adhesion is measured both as the percentage of pathogens bound to epithelial membrane vesicles as well as the mean number of vesicles bound per infected erythrocytes. We show that adhesins alternative to those currently identified could be involved. This demonstrates the power of this assay to advance our understanding of epithelial adhesion of infected erythrocytes and in the design of intervention strategies

ART influences HIV persistence in the female reproductive tract and cervicovaginal secretions

The recently completed HIV prevention trials network study 052 is a landmark collaboration demonstrating that HIV transmission in discordant couples can be dramatically reduced by treating the infected individual with antiretroviral therapy (ART). However, the cellular and virological events that occur in the female reproductive tract (FRT) during ART that result in such a drastic decrease in transmission were not studied and remain unknown. Here, we implemented an in vivo model of ART in BM/liver/thymus (BLT) humanized mice in order to better understand the ability of ART to prevent secondary HIV transmission. We demonstrated that the entire FRT of BLT mice is reconstituted with human CD4+ cells that are shed into cervicovaginal secretions (CVS). A high percentage of the CD4+ T cells in the FRT and CVS expressed CCR5 and therefore are potential HIV target cells. Infection with HIV increased the numbers of CD4+ and CD8+ T cells in CVS of BLT mice. Furthermore, HIV was present in CVS during infection. Finally, we evaluated the effect of ART on HIV levels in the FRT and CVS and demonstrated that ART can efficiently suppress cell-free HIV-RNA in CVS, despite residual levels of HIV-RNA+ cells in both the FRT and CVS

Id2-, RORγt-, and LTβR-independent initiation of lymphoid organogenesis in ocular immunity

The eye is protected by the ocular immunosurveillance system. We show that tear duct–associated lymphoid tissue (TALT) is located in the mouse lacrimal sac and shares immunological characteristics with mucosa-associated lymphoid tissues (MALTs), including the presence of M cells and immunocompetent cells for antigen uptake and subsequent generation of mucosal immune responses against ocularly encountered antigens and bacteria such as Pseudomonas aeruginosa. Initiation of TALT genesis began postnatally; it occurred even in germ-free conditions and was independent of signaling through organogenesis regulators, including inhibitor of DNA binding/differentiation 2, retinoic acid–related orphan receptor γt, lymphotoxin (LT) α1β2–LTβR, and lymphoid chemokines (CCL19, CCL21, and CXCL13). Thus, TALT shares immunological features with MALT but has a distinct tissue genesis mechanism and plays a key role in ocular immunity

M cell-depletion blocks oral prion disease pathogenesis

Many prion diseases are orally acquired. Our data show that after oral exposure, early prion replication upon follicular dendritic cells (FDC) in Peyer's patches is obligatory for the efficient spread of disease to the brain (termed neuroinvasion). For prions to replicate on FDC within Peyer's patches after ingestion of a contaminated meal, they must first cross the gut epithelium. However, the mechanism through which prions are conveyed into Peyer's patches is uncertain. Within the follicle-associated epithelium overlying Peyer's patches are microfold cells (M cells), unique epithelial cells specialized for the transcytosis of particles. We show that following M cell-depletion, early prion accumulation upon FDC in Peyer's patches is blocked. Furthermore, in the absence of M cells at the time of oral exposure, neuroinvasion and disease development are likewise blocked. These data suggest M cells are important sites of prion uptake from the gut lumen into Peyer's patches

DOGS: Reaction-Driven de novo Design of Bioactive Compounds

We present a computational method for the reaction-based de novo design of drug-like molecules. The software DOGS (Design of Genuine Structures) features a ligand-based strategy for automated ‘in silico’ assembly of potentially novel bioactive compounds. The quality of the designed compounds is assessed by a graph kernel method measuring their similarity to known bioactive reference ligands in terms of structural and pharmacophoric features. We implemented a deterministic compound construction procedure that explicitly considers compound synthesizability, based on a compilation of 25'144 readily available synthetic building blocks and 58 established reaction principles. This enables the software to suggest a synthesis route for each designed compound. Two prospective case studies are presented together with details on the algorithm and its implementation. De novo designed ligand candidates for the human histamine H4 receptor and γ-secretase were synthesized as suggested by the software. The computational approach proved to be suitable for scaffold-hopping from known ligands to novel chemotypes, and for generating bioactive molecules with drug-like properties

IL-2 receptor γ-chain molecule is critical for intestinal T-cell reconstitution in humanized mice

Intestinal immune cells are important in host defense, yet the determinants for human lymphoid homeostasis in the intestines are poorly understood. In contrast, lymphoid homeostasis has been studied extensively in mice, where the requirement for a functional common γ-chain molecule has been established. We hypothesized that humanized mice could offer insights into human intestinal lymphoid homeostasis if generated in a strain with an intact mouse common γ-chain molecule. To address this hypothesis, we used three mouse strains (non-obese diabetic (NOD)/severe-combined immunodeficient (SCID) (N/S); NOD/SCID γ-chain−/− (NSG); and Rag2−/− γ-chain−/− (DKO)) and two humanization techniques (bone marrow liver thymus (BLT) and human CD34+ cell bone marrow transplant of newborn mice (hu)) to generate four common types of humanized mice: N/S-BLT, NSG-BLT, NSG-hu, and DKO-hu mice. The highest levels of intestinal human T cells throughout the small and large intestines were observed in N/S-BLT mice, which have an intact common γ-chain molecule. Furthermore, the small intestine lamina propria T-cell populations of N/S-BLT mice exhibit a human intestine-specific surface phenotype. Thus, the extensive intestinal immune reconstitution of N/S-BLT mice was both quantitatively and qualitatively better when compared with the other models tested such that N/S-BLT mice are well suited for the analysis of human intestinal lymphocyte trafficking and human-specific diseases affecting the intestines

"They brought you back to the fact you're not the same": Sense of self after traumatic brain injury

This paper considers contexts following traumatic brain injury, exploring what may be at stake when dominant expectations predict a ‘lost’ or ‘broken’ self. I explore stories co-constructed with one young man and his mother to illustrate their personal and intersubjective understandings of identity, at times conflicting, within family interactions and when encountering normative practices of neurorehabilitation clinicians. The ower relations portrayed confront this man’s narrative attempts to align his present and pre-injury self, including standard assessments delineating change, administered by healthcare professionals. I consider a need for greater attention to interaction-generated disruption to sense of self, wthin contemporary conceptualisations of ‘person-centred care’

The functional maturation of M cells is dramatically reduced in the Peyer's patches of aged mice

The transcytosis of antigens across the follicle-associated epithelium (FAE) of Peyer's patches by microfold cells (M cells) is important for the induction of efficient immune responses to mucosal antigens. The mucosal immune response is compromised by ageing, but effects on M cells were unknown. We show that M-cell density in the FAE of aged mice was dramatically reduced. As a consequence, aged Peyer's patches were significantly deficient in their ability to transcytose particulate lumenal antigen across the FAE. Ageing specifically impaired the expression of Spi-B and the downstream functional maturation of M cells. Ageing also dramatically impaired C-C motif chemokine ligand 20 expression by the FAE. As a consequence, fewer B cells were attracted towards the FAE, potentially reducing their ability to promote M-cell maturation. Our study demonstrates that ageing dramatically impedes the functional maturation of M cells, revealing an important ageing-related defect in the mucosal immune system's ability to sample lumenal antigens