Play in rats: association across contexts and types, and analysis of structure

Play has been proposed as a promising indicator of positive animal welfare. We aimed to study play in rats across contexts (conspecific/heterospecific) and types (social: pinning, being pinned; solitary: scampering), and we investigated its structure using behavioral sequence analysis. Group-housed (three per cage) adolescent male Lister Hooded rats (n = 21) were subjected to a Play-In-Pairs test: after a 3 hour isolation period, a pair of cage-mates was returned to the home cage and both social and solitary play were scored for 20 min. This procedure was repeated for each pair combination across three consecutive days, and individual play scores were calculated. Heterospecific play was measured using a Tickling test: rats were individually tickled by the experimenter through bouts of gentle, rapid finger movements on their underside, and the number of positive 50 kHz frequency modulated vocalizations and experimenter-directed approach behaviors were recorded. Both of the above tests were compared with social play in the home cage. While conspecific play in both the Play-In-Pairs test and home cage were correlated, both seemed to be unrelated to heterospecific play in the Tickling test. During the Play-In-Pairs test, although both solitary and social play types occurred, they were unrelated, and solitary locomotor play of one rat did not predict the subsequent play behavior of its cage mate. Analysis of play structure revealed that social play occurred more often in bouts of repeated behaviors while solitary play sequences did not follow a specific pattern. If play is to be used as an indicator of positive welfare in rats, context, type and structure differences should be taken into account

Chemokine Receptor CCR9 Contributes to the Localization of Plasma Cells to the Small Intestine

Humoral immunity in the gut-associated lymphoid tissue is characterized by the production of immunoglobulin A (IgA) by antibody-secreting plasma cells (PCs) in the lamina propria. The chemokine CCL25 is expressed by intestinal epithelial cells and is capable of inducing chemotaxis of IgA+ PCs in vitro. Using a newly generated monoclonal antibody against murine CCR9, we show that IgA+ PCs express high levels of CCR9 in the mesenteric lymph node (MLN) and Peyer's patches (PPs), but down-regulate CCR9 once they are located in the small intestine. In CCR9-deficient mice, IgA+ PCs are substantially reduced in number in the lamina propria of the small intestine. In adoptive transfer experiments, CCR9-deficient IgA+ PCs show reduced migration into the small intestine compared with wild-type controls. Furthermore, CCR9 mutants fail to mount a regular IgA response to an orally administered antigen, although the architecture and cell type composition of PPs and MLN are unaffected and are functional for the generation of IgA PCs. These findings provide profound in vivo evidence that CCL25/CCR9 guides PCs into the small intestine

Selective Generation of Gut Tropic T Cells in Gut-associated Lymphoid Tissue (GALT): Requirement for GALT Dendritic Cells and Adjuvant

In the current study, we address the underlying mechanism for the selective generation of gut-homing T cells in the gut-associated lymphoid tissues (GALT). We demonstrate that DCs in the GALT are unique in their capacity to establish T cell gut tropism but in vivo only confer this property to T cells in the presence of DC maturational stimuli, including toll-like receptor-dependent and -independent adjuvants. Thus, DCs from mesenteric LNs (MLNs), but not from spleen, supported expression of the chemokine receptor CCR9 and integrin α4β7 by activated CD8+ T cells. While DCs were also required for an efficient down-regulation of CD62L, this function was not restricted to MLN DCs. In an adoptive CD8+ T cell transfer model, antigen-specific T cells entering the small intestinal epithelium were homogeneously CCR9+α4β7+CD62Llow, and this phenotype was only generated in GALT and in the presence of adjuvant. Consistent with the CCR9+ phenotype of the gut-homing T cells, CCR9 was found to play a critical role in the localization of T cells to the small intestinal epithelium. Together, these results demonstrate that GALT DCs and T cell expression of CCR9 play critical and integrated roles during T cell homing to the gut

A Systemically-Administered Small Molecule Antagonist of CCR9 Acts as a Tissue-Selective Inhibitor of Lymphocyte Trafficking

A goal for developers of immunomodulatory drugs has long been a systemically administered small molecule that can selectively inhibit inflammation in specific tissues. The chemokine receptor CCR9 is an attractive target for this approach, as entry of T cells into the small intestine from blood requires interaction between CCR9 and its ligand CCL25. We have tested the ability of a small molecule CCR9 antagonist, CCX8037, to inhibit antigen-mediated T cell accumulation in the intestine. This compound prevented accumulation of gut-imprinted antigen-specific CD8 T cells within epithelium of the small intestine. Interestingly, the antagonist did not affect the robust generation of gut-imprinted CD8 T cells within mesenteric lymph nodes. To distinguish “gut-selective” from “general” T cell inhibition, we tested the drug’s ability to influence accumulation of T cells within skin, a tissue in which CCR9 plays no known role, and we found no appreciable effect. This study demonstrates the feasibility of creating systemically-administered pharmaceuticals capable of tissue-selective immune modulation. This proof of concept is of utmost importance for designing effective treatments against various autoimmune disorders localized to a specific tissue

‘Puppy Dog Eyes’ Are Associated With Eye Movements, Not Communication

The inner brow raiser is a muscle movement that increases the size of the orbital cavity, leading to the appearance of so-called ‘puppy dog eyes’. In domestic dogs, this expression was suggested to be enhanced by artificial selection and to play an important role in the dog-human relationship. Production of the inner brow raiser has been shown to be sensitive to the attentive stance of a human, suggesting a possible communicative function. However, it has not yet been examined whether it is sensitive to human presence. In the current study, we aimed to test whether the inner brow raiser differs depending on the presence or absence of an observer. We used two versions of a paradigm in an equivalent experimental setting in which dogs were trained to expect a reward; however, the presence/absence of a person in the test apparatus was varied. In the social context, a human facing the dog delivered the reward; in the non-social context, reward delivery was automatized. If the inner brow raiser has a communicative function and dogs adjust its expression to an audience, we expect it to be shown more frequently in the social context (when facing a person in the apparatus) than in the non-social context (when facing the apparatus without a person inside). The frequency of the inner brow raiser differed between the two contexts, but contrary to the prediction, it was shown more frequently in the non-social context. We further demonstrate that the inner brow raiser is strongly associated with eye movements and occurs independently in only 6% of cases. This result challenges the hypothesis that the inner brow raiser has a communicative function in dog-human interactions and suggests a lower-level explanation for its production, namely an association with eye movements

Play like me: Similarity in playfulness promotes social play

Social play is associated with the experience of positive emotions in higher vertebrates and may be used as a measure of animal welfare. Altering motivation to play (e.g., through short-term social isolation) can temporarily affect play levels between familiar individuals, a process which may involve emotional contagion. This study investigated how forming groups based on known differences in the personality trait “playfulness” (i.e., the longer-term propensity of an individual to actively play from adolescence to early adulthood) affects social play. Seventy-six adolescent male Lister Hooded rats underwent a Play-in-Pairs test assessing their playfulness, ranked as high (H), intermediate (I) or low (L). At seven weeks of age, rats were resorted into homogenous groups of similar (LLL, III, HHH), or heterogeneous groups of dissimilar (HII, LII) playfulness. Social play was scored in the home cage at Weeks 8, 10, 12 of age. A second Play-in-Pairs test was performed (Week 11) to assess consistency of playfulness. A Social Preference test investigated whether I rats in heterogeneous groups preferred proximity with I, H or L cage mates. It was found that heterogeneous groups played less than homogeneous ones at adolescence (8 weeks of age), while play levels at early adulthood (Weeks 10 and 12) did not differ between groups. Play in the homogeneous groups decreased with age as expected, while it did not change over time in the heterogeneous groups, which did not compensate for the lower play levels shown at adolescence. Play-in-Pairs scores before and after resorting were mildly correlated, indicating some level of consistency over time despite the resorting procedure. In the Social Preference test, subjects did not prefer one playfulness level over another. We conclude that a mismatch in playfulness may negatively affect social play development, and thus the welfare, of rats. Groups made of animals with similar playfulness, even those initially scoring relatively low in this trait, seemed to be more successful in establishing play relationships during adolescence

Settling the thymus: immigration requirements

Thymus settling by precursor cells is essential for the production of T cells, yet the immigration requirements are poorly defined. P-selectin and CC chemokine receptor-9 (CCR9) are involved, and settling is favored when existing residents have moved on. A new study strengthens the correlation between niche emptying and the induction of thymic P-selectin and CCR9 ligand, and provides evidence for feedback from the periphery to thymic P-selectin expression via sphingosine-1-phosphate

CCL25/CCR9 Interactions Regulate Large Intestinal Inflammation in a Murine Model of Acute Colitis

CCL25/CCR9 is a non-promiscuous chemokine/receptor pair and a key regulator of leukocyte migration to the small intestine. We investigated here whether CCL25/CCR9 interactions also play a role in the regulation of inflammatory responses in the large intestine.Acute inflammation and recovery in wild-type (WT) and CCR9(-/-) mice was studied in a model of dextran sulfate sodium (DSS)-induced colitis. Distribution studies and phenotypic characterization of dendritic cell subsets and macrophage were performed by flow cytometry. Inflammatory bowel disease (IBD) scores were assessed and expression of inflammatory cytokines was studied at the mRNA and the protein level.CCL25 and CCR9 are both expressed in the large intestine and are upregulated during DSS colitis. CCR9(-/-) mice are more susceptible to DSS colitis than WT littermate controls as shown by higher mortality, increased IBD score and delayed recovery. During recovery, the CCR9(-/-) colonic mucosa is characterized by the accumulation of activated macrophages and elevated levels of Th1/Th17 inflammatory cytokines. Activated plasmacytoid dendritic cells (DCs) accumulate in mesenteric lymph nodes (MLNs) of CCR9(-/-) animals, altering the local ratio of DC subsets. Upon re-stimulation, T cells isolated from these MLNs secrete significantly higher levels of TNFα, IFNγ, IL2, IL-6 and IL-17A while down modulating IL-10 production.Our results demonstrate that CCL25/CCR9 interactions regulate inflammatory immune responses in the large intestinal mucosa by balancing different subsets of dendritic cells. These findings have important implications for the use of CCR9-inhibitors in therapy of human IBD as they indicate a potential risk for patients with large intestinal inflammation

CCR9-CCL25 interactions promote cisplatin resistance in breast cancer cell through Akt activation in a PI3K-dependent and FAK-independent fashion

<p>Abstract</p> <p>Background</p> <p>Chemotherapy heavily relies on apoptosis to kill breast cancer (BrCa) cells. Many breast tumors respond to chemotherapy, but cells that survive this initial response gain resistance to subsequent treatments. This leads to aggressive cell variants with an enhanced ability to migrate, invade and survive at secondary sites. Metastasis and chemoresistance are responsible for most cancer-related deaths; hence, therapies designed to minimize both are greatly needed. We have recently shown that CCR9-CCL25 interactions promote BrCa cell migration and invasion, while others have shown that this axis play important role in T cell survival. In this study we have shown potential role of CCR9-CCL25 axis in breast cancer cell survival and therapeutic efficacy of cisplatin.</p> <p>Methods</p> <p>Bromodeoxyuridine (BrdU) incorporation, Vybrant apoptosis and TUNEL assays were performed to ascertain the role of CCR9-CCL25 axis in cisplatin-induced apoptosis of BrCa cells. Fast Activated Cell-based ELISA (FACE) assay was used to quantify <it>In situ </it>activation of PI3K^p85, Akt^Ser473, GSK-3β^Ser9and FKHR^Thr24in breast cancer cells with or without cisplatin treatment in presence or absence of CCL25.</p> <p>Results</p> <p>CCR9-CCL25 axis provides survival advantage to BrCa cells and inhibits cisplatin-induced apoptosis in a PI3K-dependent and focal adhesion kinase (FAK)-independent fashion. Furthermore, CCR9-CCL25 axis activates cell-survival signals through Akt and subsequent glycogen synthase kinase-3 beta (GSK-3β) and forkhead in human rhabdomyosarcoma (FKHR) inactivation. These results show that CCR9-CCL25 axis play important role in BrCa cell survival and low chemotherapeutic efficacy of cisplatin primarily through PI3K/Akt dependent fashion.</p