Innate immunity defines the capacity of antiviral T cells to limit persistent infection

Effective immunity requires the coordinated activation of innate and adaptive immune responses. Natural killer (NK) cells are central innate immune effectors, but can also affect the generation of acquired immune responses to viruses and malignancies. How NK cells influence the efficacy of adaptive immunity, however, is poorly understood. Here, we show that NK cells negatively regulate the duration and effectiveness of virus-specific CD4+ and CD8+ T cell responses by limiting exposure of T cells to infected antigen-presenting cells. This impacts the quality of T cell responses and the ability to limit viral persistence. Our studies provide unexpected insights into novel interplays between innate and adaptive immune effectors, and define the critical requirements for efficient control of viral persistence

Ballet injuries: injury incidence and severity over 1 year.

STUDY DESIGN: Prospective, descriptive single-cohort study. OBJECTIVE: To assess the incidence and severity of injuries to a professional ballet company over 1 year. METHODS: Data for an elite-level ballet company of 52 professional dancers were collected by an in-house medical team using a time-loss injury definition. RESULTS: A total of 355 injuries were recorded, with an overall injury incidence of 4.4 injuries per 1000 hours (female, 4.1; male, 4.8; P>.05) and a mean of 6.8 injuries per dancer (female, 6.3; male, 7.3; P>.05). Mean injury severity was 7 days (female, 4; male, 9; P.05); mean severity of injury was 3 days for females and 9 days for males (P<.05). The percentage of traumatic injuries was 32% for females and 40% for males (P<.05); the corresponding severity was 6 and 10 days, respectively (P<.05). CONCLUSION: The relatively high number of injuries reported and the resulting loss of dance time support the need to introduce interventions to reduce the risk of injury in professional dancers

Spatiotemporally separated antigen uptake by alveolar dendritic cells and airway presentation to T cells in the lung

Asthma pathogenesis is focused around conducting airways. The reasons for this focus have been unclear because it has not been possible to track the sites and timing of antigen uptake or subsequent antigen presentation to effector T cells. In this study, we use two-photon microscopy of the lung parenchyma and note accumulation of CD11b(+) dendritic cells (DCs) around the airway after allergen challenge but very limited access of these airway-adjacent DCs to the contents of the airspace. In contrast, we observed prevalent transepithelial uptake of particulate antigens by alveolar DCs. These distinct sites are temporally linked, as early antigen uptake in alveoli gives rise to DC and antigen retention in the airway-adjacent region. Antigen-specific T cells also accumulate in the airway-adjacent region after allergen challenge and are activated by the accumulated DCs. Thus, we propose that later airway hyperreactivity results from selective retention of allergen-presenting DCs and antigen-specific T cells in airway-adjacent interaction zones, not from variation in the abilities of individual DCs to survey the lung

The regulation of peripheral T cell responses in TCR transgenic mice

Transgenic mice expressing a T cell receptor specific for cytochrome C in association with I-Ek were employed to study the mechanisms regulating the decision between tolerance and immunity. Tolerance and immunity to the same peptide antigen were . induced by using different routes of administration. Thus tolerance was induced by the intravenous route and immunity by the subcutaneous route. The results presented in this thesis provide a detailed map of the cellular events that occur during these two distinct responses. Intravenous immunisation induced activation of CD4+ cells expressing the transgenic TCR (CD4+Tg0L+ cells), resulting in CD69 expression within two hours, priming of T cell proliferation and Th1 cytokine production by 24 hours. Clonal expansion peaked 3- 4 days after immunisation. The number of CD4+Tg0t+ cells decreased rapidly after day four, such that only 50% of initial cell numbers remained 7—10 days after immunisation. Intravenous peptide also upregulated CD44 expression, increasing the proportion and number of CD4+Tg0t+CD44hi cells at the peak of the response, but having no net effect at the resolution of the response. When labelled CD4+Tg0t+ cells were adoptively transferred to syngeneic non-transgenic hosts, deletional tolerance to intravenous peptide was still seen, demonstrating that it was not an artefact of the high precursor frequency in TCR transgenic mice. Antigen re-challenge experiments demonstrated that CD4+Tg0t+ cells remaining at the resolution of the primary response to intravenous peptide could not respond as vigorously as naive cells either in vitro and in vivo. Interestingly, intravenous administration of intact cytochrome C also stimulated T cell activation, but failed to induce peripheral deletion, and resulted instead in a minor increase in the final proportion of CD4+Tg0t+CD44hi cells

Visualizing T Cell Competition for Peptide/MHC Complexes A Specific Mechanism to Minimize the Effect of Precursor Frequency

AbstractIn vivo antigenic competition of naive CD4+ TCR transgenic T cells was visualized by tracking cell division. Competition reduced both recruitment into cell division and burst size per recruited precursor cell, minimizing the effect of differences in precursor frequency while maintaining the dose-response relationship with antigen. Competition was restricted to T cells of the same specificity, indicating that cells were competing for access to Ag-MHC complexes rather than for Ag nonspecific factors. Moreover, the qualitative distinction between the responses to i.v. peptide and s.c. peptide/CFA was unaffected by precursor frequency. These data explain the paradoxical ability of the immune system to tailor responses to the type and dose of Ag even in individuals with large differences in initial precursor frequency

FLT3 Inhibitor AC220 Is a Potent Therapy for Myeloproliferative Disease in c-Cbl RING Finger Mutant Mice

Abstract Abstract 642 Introduction: c-Cbl is a RING finger based E3 ubiquitin ligase that is highly expressed in hematopoietic cells where it directs the polyubiquitylation and degradation of protein tyrosine kinases. c-Cbl also functions as a multi-domain adaptor that recruits and activates proteins such as PI 3-kinase. Oncogenic forms c-Cbl that have lost E3 ligase show enhanced activity of these adaptor functions in addition to the inability to downregulate activated tyrosine kinases. c-Cbl mutations are found in patients with a range of myeloid malignancies that include myelodysplastic syndromes, chronic myelomonocytic leukaemia, juvenile myelomonocytic leukaemia, atypical chronic myeloid leukaemia and acute myeloid leukaemia. Sequencing has shown that the mutations are located in either the linker or RING finger domains and that these mutations abolish E3 ligase activity. Thus the development of leukemia is caused, at least in part, through the loss of this activity. To investigate c-Cbl associated malignancies we generated a mouse with a knockin mutation in the RING finger domain that develops a myeloproliferative disease (MPD) progressing to lethal leukemia. The mutation is a substitution of a cysteine for alanine in the RING finger at 379 (i.e. C379A) that disrupts c-Cbl's ability to function as an E3 ubiquitin ligase. The majority of the c-Cbl C379A mutant mice succumb within a year with markedly elevated white blood cell (WBC) counts, splenomegaly and extensive infiltrations of myeloid cells into peripheral organs. These mice therefore provide a pre-clinical model for studying myeloid malignancies associated with oncogenic forms of c-Cbl. We have found that hematopoietic progenitors from c-Cbl(C379A) mice have enhanced FLT3 signaling and an expanded population of FLT3high cells compared to WT and c-Cbl KO mice. When c-Cbl(C379A) mice are mated to FLT3 ligand KO mice the doubly mutant mice do not develop a severe MPD or leukemia and the FLT3high population is markedly suppressed. Thus targeting FLT3 may offer an effective therapy. AC220 is a FLT3 inhibitor that shows excellent potency, selectivity and pharmacokinetic properties and is currently being investigated in clinical trials treating patients with activating FLT3 mutations. In this study we examined the effects of treating c-Cbl(C379A) mice with AC220 to determine whether AC220 can prevent the development of myeloid malignancies associated with aberrant wild-type FLT3 signaling. Results: To determine the effectiveness of AC220 for treating c-Cbl-associated MPD we established two cohorts, one comprising 16 C57BL/6.CD45.1 mice repopulated with bone marrow (BM) from an 8 week-old c-Cbl(C379A) mouse and one comprising 18 c-Cbl(C379A) mice aged 8–9 months. Mice were bled 5 days before dosing to determine pre-treatment WBC counts and both cohorts were then divided into two groups for daily dosing by oral gavage with either AC220 (10mg/kg) or vehicle (5% cyclodextrin) and bled at 3 and 6 weeks. Dosing with AC220 resulted in a marked decrease in WBC counts in all lineages. The mean WBC counts for the AC220-treated mice transplanted with c-Cbl(C379A) marrow at 3 and 6 weeks were 14.0 and 14.5×109/L respectively, compared to a mean WBC count of 33.5×109/L before treatment commenced. For the 8–9 month old c-Cbl(C379A) mice the 3 and 6 wk counts were 28.5 and 20.0 ×109/L respectively, compared to pre-treatment levels of 64.5×109/L. In contrast the WBC counts remained constantly high in the vehicle treated groups. After 6 weeks of dosing the mice were sacrificed and the BM analyzed by flow cytometry to determine the effect on FLT3high progenitors. The AC220 treatment reduced the FLT3high population of LSK cells by more than two fold in both cohorts. Similarly AC220 treatment had a significant impact on spleen weights with a greater than two-fold reduction in the AC220-treated groups. In addition AC220 dosing resulted in a marked decrease in the myeloid invasiveness into peripheral organs, an outcome that suggests AC220 treatment would enhance survival. Conclusions: These studies are the first to demonstrate in vivo that AC220 is a very effective treatment for diseases or conditions that are driven by WT FLT3. Future studies investigating aberrant FLT3 expression on progenitors from a range of MDS/MPN patients may therefore reveal therapeutic options that would not have otherwise been appreciated without the preclinical studies of the c-Cbl RING finger mutant mouse. Disclosures: No relevant conflicts of interest to declare. </jats:sec

Flt3 inhibitor AC220 is a potent therapy in a mouse model of myeloproliferative disease driven by enhanced wild-type Flt3 signaling

Abstract High levels of expression of wild-type Flt3 characterize many hematopoietic proliferative diseases and neoplasms, providing a potential therapeutic target. Using the c-Cbl RING finger mutant mouse as a model of a myeloproliferative disease (MPD) driven by wild-type Flt3, in the present study, we show that treatment with the Flt3 kinase inhibitor AC220 blocks MPD development by targeting Flt3+ multipotent progenitors (MPPs). We found that daily administration of AC220 caused a marked reduction in Flt3 expression, induction of quiescence, and a significant loss of MPPs within 4 days. Unexpectedly, a robust Flt3 ligand–associated proliferative recovery response soon followed, preventing further loss of MPPs. However, continued AC220 treatment limited MPP recovery and maintained reduced, steady-state levels of cycling MPPs that express low levels of Flt3. Therefore, a finely tuned balance between the opposing forces of AC220 and Flt3 ligand production was established; whereas the Flt3 ligand blunted the inhibitory effects of AC220, the disease was held in remission for as long as therapy was continued. The net effect is a potent therapy indicating that patients with c-Cbl mutations, or those with similarly enhanced Flt3 signaling, may respond well to AC220 even after the induction of high levels of Flt3 ligand.</jats:p