Spontaneous Expression of Interleukin-2 In Vivo in Specific Tissues of Young Mice

In situ hybridization and immunohistochemistry were used to determine the spectrum of tissues in which interleukin-2 (IL-2) mRNA and protein are found in healthy, normal young mice. In neonatal animals, IL-2 is expressed specifically by distinct, isolated cells at three major sites: the thymus, skin, and gut. Based on morphology and distribution, the IL-2-expressing cells resemble CD3ε + T cells that are also present in all these locations. Within the thymus of postweanling animals, both TcRαβ and TcRγδ lineage cells secrete "haloes" of the cytokine that diffuse over many cell diameters. Within the skin, isolated cells expressing IL-2 are seen at birth in the mesenchyme, and large numbers of IL-2-expressing cells are localized around hair follicles in the epidermis in 3-week-old animals. At this age, a substantial subset of CD3ε + cells is similarly localized in the skin. Significantly, by 5 weeks of age and later when the CD3ε + cells are evenly distributed throughout the epidermis, IL-2 RNA and protein expression are no longer detectable. Finally, within the intestine, IL-2 protein is first detected in association with a few discrete, isolated cells at day 16 of gestation and the number of IL-2 reactive cells increases in frequency through El9 and remains abundant in adult life. In postnatal animals, the frequency of IL- 2-positive cells in villi exceeds by greater than fivefold that found in mesenteric lymph node or Peyer's patches. Overall, these temporal and spatial patterns of expression provide insight into the regulation of IL-2 in vivo and suggest a role for IL-2 expression distinct from immunological responses to antigen

Developmental and Anatomical Patterns Of IL-2 Gene Expression in Vivo in The Murine Thymus

Interleukin-2 (IL-2) is a potent growth factor that mature T lymphocytes synthesize and use as a proliferation signal. Much controversy has arisen concerning whether it is used to drive the extensive proliferation of immature pre-T cells in the thymus. Immature thymocytes acquire the competence to express IL-2 at an early stage, but it has remained uncertain whether they are activated to exercise this competence in vivo. Therefore, we have used in situ hybridization and immunohistochemistry on serial sections obtained from fetal and adult thymuses of normal C57BL/6 mice and of mice bearing the scid defect to determine where, when, and whether IL-2 is expressed in vivo. Our results show a striking spatial and temporal pattern of IL-2 expression in the normal fetal thymus. We detected a burst of IL-2 mRNA accumulation at day 14.5 of gestation, which rapidly decreased by day 15. At day 15, we observed maximal IL-2 protein production that subsequently decreased by day 16 of gestation. Both in situ hybridization and immunohistochemical staining revealed an unexpectedly strict localization of IL-2 expressing cells to patches around the periphery of the fetal thymus, creating a previously unrecognized compartment of high IL-2 protein content. IL-2 production in the day-15 fetal thymus appeared to be unaffected by the scid mutation, indicating that this response is likely to be T-cell receptor (TcR)-independent. Several features distinguish the IL-2 induction pattern in the adult thymus from that in the fetal thymus. In the normal adult thymus, IL-2-expressing cells are extremely rare (found at a frequency of 10^(-7)), but they are reproducibly detectable as isolated cells in the outer cortex and subcapsular region of the thymus. Unlike the fetal thymic IL-2 producers, the IL-2 producers in the adult thymus are completely eliminated in mice homozygous for the scid mutation. This suggests that the IL-2-expressing cells in the normal adult thymus are of a more mature phenotype than the immature, TcR-negative cells that accumulate in the scid adult thymus. Thus, our work demonstrates that two developmentally distinct types of cell interactions induce IL-2 expression in vivo: one, a broadly localized interaction in day 14-15 fetal thymus that is unaffected by the scid mutation; the other, a rare event that occurs asynchronously from late fetal through adult life, but which is completely eliminated by the scid defect. These results imply that significant differences exist between the physiological processing of thymocytes in the fetal and postnatal thymic microenvironments

Formulating Older Driver Licensing Policy: An Evaluation of Older Driver Crash History and Performance

This research sought to understand the relationship between licensing policy and the opportunity for the development of a scientifically-based approach to identifying high risk older drivers based on prior driving history. This research focused on five tasks: 1) review of the literature, 2) compilation of information on licensing policy for use by decision-makers, 3) assessment of charges and payer source for older driver crashes using linked crash and hospital data , and 4) the development and 5) validation of an older driver crash prediction model. There is relatively little available in the way of information for policymakers regarding licensing, and there is even less information available on evaluation of licensing practice effectiveness. Emergency department charges for older males were lower than females even though males accounted for a larger percentage of the injured population. Older drivers were no more likely to be covered by public insurance than the comparison group. Crash and citation data used to develop a driver history showed no differences between drivers in injury causing crashes and drivers in non-injury crashes. Logistic regression, Poisson regression, and negative binomial regression models were unable to effectively predict crash involvement based on driver history. This is likely due to self-selection bias for older drivers and truncated distribution of count variable (injury causing crashes). Recommendations resulting from this research include Massachusetts and national policy recommendations and additional research. Massachusetts should expand beyond its referral-based system for reviewing older drivers, consider restriction rather than only revocation, review medical advisory board practices, conduct evaluation of any policies it does implement, and conduct a thorough review of alternative transportation options. Nationally, efforts should focus on developing effective cognitive/functional testing by licensing agents, identification of effective second phase of testing, determination of a mechanism for determining when to retest, and assessment of the differences between older males and females for potential use in training, education, and testing. Research recommendations include continued exploration of the potential for systematic identification of high risk drivers using administrative data and in-depth analyses of the differences between males and females in terms of aging and driver safety

Dynamic Transformations of Genome-wide Epigenetic Marking and Transcriptional Control Establish T Cell Identity

T cell development comprises a stepwise process of commitment from a multipotent precursor. To define molecular mechanisms controlling this progression, we probed five stages spanning the commitment process using RNA-seq and ChIP-seq to track genome-wide shifts in transcription, cohorts of active transcription factor genes, histone modifications at diverse classes of cis-regulatory elements, and binding repertoire of GATA-3 and PU.1, transcription factors with complementary roles in T cell development. The results highlight potential promoter-distal cis-regulatory elements in play and reveal both activation sites and diverse mechanisms of repression that silence genes used in alternative lineages. Histone marking is dynamic and reversible, and though permissive marks anticipate, repressive marks often lag behind changes in transcription. In vivo binding of PU.1 and GATA-3 relative to epigenetic marking reveals distinctive factor-specific rules for recruitment of these crucial transcription factors to different subsets of their potential sites, dependent on dose and developmental context

In-packet Bloom filters: Design and networking applications

The Bloom filter (BF) is a well-known space-efficient data structure that answers set membership queries with some probability of false positives. In an attempt to solve many of the limitations of current inter-networking architectures, some recent proposals rely on including small BFs in packet headers for routing, security, accountability or other purposes that move application states into the packets themselves. In this paper, we consider the design of such in-packet Bloom filters (iBF). Our main contributions are exploring the design space and the evaluation of a series of extensions (1) to increase the practicality and performance of iBFs, (2) to enable false-negative-free element deletion, and (3) to provide security enhancements. In addition to the theoretical estimates, extensive simulations of the multiple design parameters and implementation alternatives validate the usefulness of the extensions, providing for enhanced and novel iBF networking applications.Comment: 15 pages, 11 figures, preprint submitted to Elsevier COMNET Journa

Ikaros represses and activates PU.1 cell-type-specifically through the multifunctional Sfpi1 URE and a myeloid specific enhancer

Generation of myeloid and lymphoid cells from progenitors involves dynamic changes in transcription factor expression and use, and disruption of hematopoietic transcription factor function and expression can contribute to leukemic transformation. PU.1 and Ikaros are pivotal factors whose expression and utilization are dynamically altered during hematopoietic development. Here, we demonstrate that expression of PU.1, encoded by the Sfpi1 gene, is divergently regulated by Ikaros in distinct cell type-specific contexts. Chromatin immune precipitation analysis and functional perturbations revealed that Ikaros can directly repress or activate Sfpi1 transcription via different PU.1 cis-elements, with PU.1 and Ikaros collaborating at myeloid-specific elements but not at other elements. Our results thus shed light on how PU.1 and Ikaros can act as lineage competency factors to facilitate both myeloid and lymphoid developmental programs