Gregory D Sempowski

Duke University, Durham, North Carolina, United States

Are you Gregory D Sempowski?

Claim your profile

Publications (90)521.93 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Lymphocytes are sensitive to ionizing radiation and naive lymphocytes are more radiosensitive than their memory counterparts. Less is known about radiosensitivity of memory cell subsets. We examined the radiosensitivity of naive (TN), effector memory (TEM), and central memory (TCM) T cell subsets in C57BL/6 mice and found TEM to be more resistant to radiation-induced apoptosis than either TN or TCM. Surprisingly, we found no correlation between the extent of radiation-induced apoptosis in T cell subsets and 1) levels of pro- and antiapoptotic Bcl-2 family members or 2) the H2AX content and maximal γH2AX fold change. Rather, TEM cell survival correlated with higher levels of immediate γH2AX marking, immediate break binding and genome-wide open chromatin structure. T cells were able to mark DNA damage seemingly instantly (30 s), even if kept on ice. Relaxing chromatin with the histone deacetylase inhibitor valproic acid following radiation or etoposide treatment improved the survival of TCM and TN cells up to levels seen in the resistant TEM cells but did not improve survival from caspase-mediated apoptosis. We conclude that an open genome-wide chromatin state is the key determinant of efficient immediate repair of DNA damage in T cells, explaining the observed T cell subset radiosensitivity differences.
    Journal of immunology (Baltimore, Md. : 1950). 07/2014;
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Using A/J mice, which are susceptible to Staphylococcus aureus, we sought to identify genetic determinants of susceptibility to S. aureus, and evaluate their function with regard to S. aureus infection. One QTL region on chromosome 11 containing 422 genes was found to be significantly associated with susceptibility to S. aureus infection. Of these 422 genes, whole genome transcription profiling identified five genes (Dcaf7, Dusp3, Fam134c, Psme3, and Slc4a1) that were significantly differentially expressed in a) S. aureus -infected susceptible (A/J) vs. resistant (C57BL/6J) mice and b) humans with S. aureus blood stream infection vs. healthy subjects. Three of these genes (Dcaf7, Dusp3, and Psme3) were down-regulated in susceptible vs. resistant mice at both pre- and post-infection time points by qPCR. siRNA-mediated knockdown of Dusp3 and Psme3 induced significant increases of cytokine production in S. aureus-challenged RAW264.7 macrophages and bone marrow derived macrophages (BMDMs) through enhancing NF-κB signaling activity. Similar increases in cytokine production and NF-κB activity were also seen in BMDMs from CSS11 (C57BL/6J background with chromosome 11 from A/J), but not C57BL/6J. These findings suggest that Dusp3 and Psme3 contribute to S. aureus infection susceptibility in A/J mice and play a role in human S. aureus infection.
    PLoS Pathogens 06/2014; 10(6):e1004149. · 8.14 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Adherence of pathogens to cellular targets is required to initiate most infections. Defining strategies that interfere with adhesion is important for the development of preventative measures against infectious diseases. As an adhesin to host extracellular matrix proteins and human keratinocytes, the trimeric autotransporter adhesin DsrA, a proven virulence factor of the Gram-negative bacterium Haemophilus ducreyi, is a potential target for vaccine development. A recombinant form of the N-terminal passenger domain of DsrA from H. ducreyi class I strain 35000HP, termed rNT-DsrAI, was tested as a vaccine immunogen in the experimental swine model of H. ducreyi infection. Viable homologous H. ducreyi was not recovered from any animal receiving four doses of rNT-DsrAI administered with Freund's adjuvant at two-week intervals. Control pigs receiving adjuvant only were all infected. All animals receiving the rNT-DsrAI vaccine developed antibody endpoint titers between 3.5 and 5 logs. All rNT-DsrAI antisera bound the surface of the two H. ducreyi strains used to challenge immunized pigs. Purified anti-rNT-DsrAI IgG partially blocked binding of fibrinogen at the surface of intact H. ducreyi. Overall, immunization with the passenger domain of the trimeric autotransporter adhesin DsrA therefore accelerated clearance of H. ducreyi in experimental lesions, possibly by interfering with fibrinogen binding.
    Vaccine. 05/2014;
  • [Show abstract] [Hide abstract]
    ABSTRACT: IL-17 and IFN-γ production by Th17 and Th1 cells, respectively, is critical for survival during primary respiratory infection with the pathogenic bacterium, Francisella tularensis Live Vaccine Strain (LVS). The importance, however, of these T cell subsets and their soluble mediators is not well understood during a secondary or memory response. We measured the number of CD4(+) T cells producing IFN-γ or IL-17 in the spleen and lungs of vaccinated mice on day four of the secondary response using intracellular cytokine staining in order to identify protective T cell subsets participating in the memory response. Few bacteria were present in spleens of vaccinated mice on day four and a T cell response was not observed. In the lung, where more bacteria were present, there was a robust Th1 response in vaccinated mice but Th17 cells were not present at higher numbers in vaccinated mice compared to unvaccinated mice. These data show that the lung is the dominant site of the secondary immune response and suggest that Th17 cells are not required for survival during secondary challenge. To further investigate the importance of IFN-γ and IL-17 during the secondary response to F. tularensis, we neutralized either IFN-γ or IL-17 in vivo using monoclonal antibody treatment. Vaccinated mice treated with anti-IFN-γ lost more weight and had higher bacterial burdens compared to vaccinated mice treated with isotype control antibody. In contrast, treatment with anti-IL-17A antibody did not alter weight loss profiles or bacterial burdens compared to mice treated with isotype control antibody. Together, these results suggested that IFN-γ is required during both primary and secondary respiratory F. tularensis infection. IL-17, on the other hand, is only critical during the primary response to respiratory F. tularensis but dispensable during the secondary response.
    Vaccine. 05/2014;
  • [Show abstract] [Hide abstract]
    ABSTRACT: Luminex bead array assays are widely used for rapid biomarker quantification due to the ability to measure up to 100 unique analytes in a single well of a 96-well plate. There has been, however, no comprehensive analysis of variables impacting assay performance, nor development of a standardized proficiency testing program for laboratories performing these assays. To meet this need, the NIH/NIAID and the Cancer Immunotherapy Consortium of the Cancer Research Institute collaborated to develop and implement a Luminex assay proficiency testing program as part of the NIH/NIAID-sponsored External Quality Assurance Program Oversight Laboratory (EQAPOL) at Duke University. The program currently monitors 25 domestic and international sites with two external proficiency panels per year. Each panel includes a de-identified commercial Luminex assay kit with standards to quantify human IFNγ, TNFα, IL-6, IL-10 and IL-2, and a series of recombinant cytokine-spiked human serum samples. All aspects of panel development, testing and shipping are performed under GCLP by EQAPOL support teams. Following development testing, a comprehensive site proficiency scoring system comprised of timeliness, protocol adherence, accuracy and precision was implemented. The overall mean proficiency score across three rounds of testing has remained stable (EP3:76%, EP4:75%, EP5:77%); however, a more detailed analysis of site reported results indicates a significant improvement of intra- (within) and inter- (between) site variation, suggesting that training and remediation for poor performing sites may be having a positive impact on proficiency. Through continued proficiency testing, identification of variables affecting Luminex assay outcomes will strengthen efforts to bring standardization to the field.
    Journal of immunological methods 05/2014; · 2.35 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The induction of an intense inflammatory response by Neisseria gonorrhoeae and the persistence of this pathogen in the presence of innate effectors is a fascinating aspect of gonorrhea. Phosphoethanolamine (PEA) decoration of lipid A increases gonococcal resistance to complement-mediated bacteriolysis and cationic antimicrobial peptides (CAMPs), and recently we reported that wild-type N. gonorrhoeae strain FA1090 has a survival advantage relative to a PEA transferase A (lptA) mutant in the human urethral challenge and murine lower genital tract infection models. Here we tested the immunostimulatory role of this lipid A modification. Purified lipooligosaccharide (LOS) containing lipid A devoid of the PEA modification and an lptA mutant of strain FA19 induced significantly lower levels of NFκB in human embryonic kidney Toll-like receptor 4 (TLR4) cells and murine embryonic fibroblasts compared to the wild-type LOS or the parent strain. Moreover, in contrast to mice infected with the wild-type and complemented lptA mutant bacteria, vaginal proinflammatory cytokines and chemokines were not elevated in female mice infected with the isogenic lptA mutant compared to uninfected mice. We also demonstrated that lptA mutant bacteria were more susceptible to the human and murine cathelicidins due to increased binding by these peptides, and that the differential induction of NFκB by wild-type and unmodified lipid A was more pronounced in the presence of CAMPs. This work demonstrates that PEA-decoration of lipid A plays both protective and immunostimulatory roles and that host-derived CAMPs may further reduce the capacity of PEA-deficient lipid A to interact with TLR4 during infection.
    Infection and immunity 03/2014; · 4.21 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The adaptive immune response to Francisella tularensis is dependent on the route of inoculation. Intradermal inoculation with F. tularensis live vaccine strain (LVS) results in a robust Th1 response in the lung whereas intranasal inoculation produces fewer Th1 cells and instead many Th17 cells. Interestingly, bacterial loads in the lung are similar early after inoculation regardless of the inoculation route. We hypothesize that the adaptive immune response is influenced by local events in the lung, such as the type of cells that are first infected with Francisella. Using fluorescent activated cell sorting, we identified alveolar macrophages as the first cell type infected in the lungs of mice intranasally inoculated with F. novicida U112, LVS, or F. tularensis Schu S4. Following bacterial dissemination from the skin to the lung, interstitial macrophages or neutrophils are infected. Overall, we identified the early interactions between Francisella and the host following two different routes of inoculation.
    Infection and immunity 03/2014; · 4.21 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Establishment of humoral immunity against pathogens is dependent on events that occur in the germinal center and the subsequent induction of high-affinity neutralizing antibodies. Quantitative assays that allow monitoring of affinity maturation and duration of antibody responses can provide useful information regarding the efficacy of vaccines and adjuvants. Using an anthrax protective antigen (rPA) and alum model antigen/adjuvant system, we describe a methodology for monitoring antigen-specific serum antibody concentration and avidity by surface plasmon resonance during primary and secondary immune responses. Our analyses showed that following a priming dose in mice, rPA-specific antibody concentration and avidity increases over time and reaches a maximal response in about six weeks, but gradually declines in the absence of antigenic boost. Germinal center reactions were observed early with maximal development achieved during the primary response, which coincided with peak antibody avidity responses to primary immunization. Boosting with antigen resulted in a rapid increase in rPA-specific antibody concentration and five-fold increase in avidity, which was not dependent on sustained GC development. The described methodology couples surface plasmon resonance-based plasma avidity measurements with germinal center analysis and provides a novel way to monitor humoral responses that can play a role in facilitating vaccine and adjuvant development.
    Journal of immunological methods 12/2013; · 2.35 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Cryptococcus neoformans is an opportunistic fungal pathogen that initiates infection following inhalation. As a result, the pulmonary immune response provides a first line of defense against C. neoformans. Surfactant protein D (SP-D) is an important regulator of pulmonary immune responses and is typically host-protective against bacterial and viral respiratory infections. However, SP-D is not protective against C. neoformans. This is evidenced by previous work from our laboratory demonstrating that SP-D deficient mice infected with C. neoformans have a lower fungal burden and live longer, compared to wild-type (WT) control animals. We hypothesized that SP-D alters susceptibility to C. neoformans by dysregulating the innate pulmonary immune response following infection. Thus inflammatory cells and cytokines were compared in the bronchoalveolar lavage fluid from WT and SP-D(-/-) mice after C. neoformans infection. Post-infection, mice lacking SP-D have reduced eosinophil infiltration and IL-5 in lung lavage fluid. To further explore the interplay of SP-D, eosinophils, and IL-5, mice expressing altered levels of eosinophils and/or IL-5 were infected with C. neoformans to assess the role of these innate immune mediators. IL-5 overexpressing mice have increased pulmonary eosinophilia and are more susceptible to C. neoformans infection as compared to WT mice. Furthermore, susceptibility of SP-D(-/-) mice to C. neoformans infection could be restored to that of WT mice by increasing IL-5 and eosinophils, via crossing the IL-5 overexpressing mice with SP-D(-/-) mice. Together, these studies support the conclusion that SP-D increases susceptibility to C. neoformans infection by promoting C. neoformans-driven pulmonary IL-5 and eosinophil infiltration.
    Infection and immunity 11/2013; · 4.21 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Neisseria gonorrhoeae causes gonorrhea, a sexually transmitted infection characterized by inflammation of the cervix or urethra. However, a significant subset of patients with N. gonorrhoeae remains asymptomatic without evidence of localized inflammation. Inflammatory responses to N. gonorrhoeae are generated by host innate immune recognition of N. gonorrhoeae by several innate immune signaling pathways including lipooligosaccharide and other pathogen-derived molecules through activation of innate immune signaling systems including toll-like receptor 4 (TLR4) and the IL-1β processing complex known as the inflammasome. The lipooligosaccharide of N. gonorrhoeae has a hexa-acylated lipid A. N. gonorrhoeae that carry an inactivated msbB (also known as lpxL1) gene produce a penta-acylated lipid A and exhibit reduced biofilm formation, survival in epithelial cells, and induction of epithelial cell inflammatory signaling. We now show that msbB-deficient N. gonorrhoeae induces less inflammatory signaling in human monocytic cell lines and murine macrophages than the parent organism. The penta-acylated LOS exhibits reduced toll-like receptor 4 signaling but does not effect N. gonorrhoeae-mediated activation of the inflammasome. We demonstrate that N. gonorrhoeae msbB is dispensible for initiating and maintaining infection in a murine model of gonorrhea. Interestingly, infection with msbB-deficient N. gonorrhoeae is associated with less localized inflammation. Combined, these data suggest that TLR4-mediated recognition of N. gonorrhoeae LOS plays an important role the pathogenesis of symptomatic gonorrhea infection and that alterations in lipid A biosynthesis may play a role in determining symptomatic and asymptomatic infections.
    Infection and immunity 10/2013; · 4.21 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Macrophages and dendritic cells (DC) are distributed throughout the body and play important roles in pathogen detection and tissue homeostasis. In tissues, resident macrophages exhibit distinct phenotypes and activities, yet the transcriptional pathways that specify tissue-specific macrophages are largely unknown. We investigated the functions and origins of two peritoneal macrophage populations in mice: small and large peritoneal macrophages (SPM and LPM, respectively). SPM and LPM differ in their ability to phagocytose apoptotic cells, as well as in the production of cytokines in response to LPS. In steady-state conditions, SPM are sustained by circulating precursors, whereas LPM are maintained independently of hematopoiesis; however, both populations are replenished by bone marrow precursors following radiation injury. Transcription factor analysis revealed that SPM and LPM express abundant CCAAT/enhancer binding protein (C/EBP)-β. Cebpb(-/-) mice exhibit elevated numbers of SPM-like cells but lack functional LPM. Alveolar macrophages are also missing in Cebpb(-/-) mice, although macrophage populations in the spleen, kidney, skin, mesenteric lymph nodes, and liver are normal. Adoptive transfer of SPM into Cebpb(-/-) mice results in SPM differentiation into LPM, yet donor SPM do not generate LPM after transfer into C/EBPβ-sufficient mice, suggesting that endogenous LPM inhibit differentiation by SPM. We conclude that C/EBPβ plays an intrinsic, tissue-restricted role in the generation of resident macrophages.
    The Journal of Immunology 09/2013; · 5.52 Impact Factor
  • Melissa S Ventevogel, Gregory D Sempowski
    [Show abstract] [Hide abstract]
    ABSTRACT: The thymus is a vital organ for homeostatic maintenance of the peripheral immune system. It is within this mediastinal tissue that T cells develop and are extensively educated and exported to the periphery for establishment of a functional and effective immune system. A striking paradoxical feature of this critical lymphoid tissue is that it undergoes profound age-associated involution. Thymic decline is of minimal consequence to healthy individuals, but the reduced efficacy of the immune system with age has direct etiological linkages with an increase in diseases including opportunistic infections, autoimmunity, and incidence/burden of cancer. Furthermore the inability of adults to restore immune function following insult induced by chemotherapy, ionizing radiation exposure or therapy, and infections (e.g. HIV-1) leads to increased morbidity and often mortality in the elderly. For these reasons, it is important that investigators strive to translate their understanding of mechanisms that drive thymic involution, and develop safe and effective strategies to rejuvenate the thymus in settings of clinical need. In this review, we present a discussion of the current status of thymic rejuvenation efforts associated with: sex steroid ablation, cytokines, growth factors, and hormones.
    Current opinion in immunology 07/2013; · 10.88 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: BACKGROUND & AIMS: The liver's response to injury is fibrosis, and when chronic, cirrhosis. Age is a critical factor impacting many immune-mediated processes, potentially including the liver's wounding response to injury. METHODS: The effects of age on acute and chronic liver injury were evaluated using a carbon tetrachloride model in mice. Lymphocyte and macrophage populations were assessed by flow cytometry and immunohistochemical analysis. RESULTS: Acute liver injury was greater in 18-month-old (old) mice than in 9-month-old (middle-aged) mice as judged by changes in aminotransferases. Similarly, livers of 18-month-old mice had a significantly greater fibrogenic response to injury than did livers of 9-month-old mice after chronic injury (assessed by col1α1 mRNA expression, morphometric analysis and hydroxyproline measurement). Interestingly, livers from young mice (6 weeks old) also exhibited an increase in fibrogenesis compared to 9-month-old mice, albeit not to the same degree as in old mice. Consistent with a role for macrophages in fibrogenesis, the number of liver macrophages in young and 9-month-old mice increased, while in chronically injured livers of 18-month-old mice, the number of macrophages was reduced, and was less than in the livers of young and 9-month-old injured livers. CONCLUSIONS: Our data indicate that the fibrogenic response to injury varies substantially with age, and moreover that macrophage recruitment and dynamics may be an important component in differential age-associated fibrotic disease.
    Liver international: official journal of the International Association for the Study of the Liver 04/2013; · 3.87 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Bacterial attenuation is typically thought of as reduced bacterial growth in the presence of constant immune pressure. Infection with Francisella tularensis elicits innate and adaptive immune responses. Several in vivo screens have identified F. tularensis genes necessary for virulence. Many of these mutations render F. tularensis defective for intracellular growth. However, some mutations have no impact on intracellular growth leading us to hypothesize that these F. tularensis mutants are attenuated because they induce an altered host immune response. We were particularly interested in the F. tularensis LVS mutant clpB (FTL_0094) because this strain was attenuated in pneumonic tularemia yet induced a protective immune response. LVS clpB's attenuation was not due to an intracellular growth defect as LVS clpB grew similarly to LVS in primary bone marrow derived macrophages and a variety of cell lines. We therefore determined whether LVS clpB induced an altered immune response compared to LVS in vivo. We found that LVS clpB induced pro-inflammatory cytokine production in the lung early after infection, a process not observed during LVS infection. LVS clpB provoked a robust adaptive immune response similar in magnitude to LVS, but with increased IFN-γ and IL-17A production as measured by mean fluorescence intensity. Altogether, our results indicate that LVS clpB is attenuated due to altered host immunity and not an intrinsic growth defect. These results also indicate that disruption of non-essential gene(s) that are involved in bacterial immune evasion, like F. tularensis clpB, can serve as a model for the rational design of attenuated vaccines.
    Infection and immunity 03/2013; · 4.21 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Chikungunya virus (CHIKV) is a mosquito-borne alphavirus responsible for recent epidemic outbreaks of debilitating disease in humans. Alphaviruses are known to interact with members of the C-type lectin receptor family of pattern recognition proteins, and given that the dendritic cell inhibitory receptor (DCIR) is known to act as a negative regulator of the host inflammatory response and has previously been associated with rheumatoid arthritis, we evaluated DCIR's role in response to CHIKV infection. Although we observed an increase in the proportion of dendritic cells at the site of CHIKV infection 24-36 hpi, these cells showed decreased cell surface DCIR, suggestive of DCIR triggering and internalization. In vitro, DCIR-/- bone marrow derived dendritic cells exhibited altered cytokine expression following exposure to CHIKV. DCIR-/- mice exhibited more severe disease signs than wild type C57BL6/J mice following CHIKV infection, including more rapid and more severe onset of virus-induced edema and enhanced weight loss. Histological examination revealed that DCIR deficient animals exhibited increased inflammation and damage in both the fascia of the inoculated foot and ankle joint and DCIR deficiency skewed the CHIKV-induced cytokine response at the site of infection at multiple times post infection. Early differences in virus induced disease between C57BL6/J and DCIR-/- mice were independent of viral replication while extended viral replication correlated with enhanced foot swelling and tissue inflammation and damage in DCIR-/- compared to C57BL6/J mice 6-7 dpi. These results suggest that DCIR plays a protective role in limiting the CHIKV-induced inflammatory response and subsequent tissue and joint damage.
    Journal of Virology 03/2013; · 5.08 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: BACKGROUND: Little is known about the immunologic events surrounding pancreatic ischemia-reperfusion injury (IRI) because of a lack of established experimental models. The purpose of this study was to develop a mouse model for pancreatic IRI to serve as a basis for the immunologic characterization of pancreatic organ damage at transplantation. METHODS: Reversible ischemia was surgically induced by vascular isolation of the distal pancreas for 0, 10, 20, or 30 min. Mice receiving laparotomy without clamping served as sham-operated controls. After reperfusion, mice were serially assayed for biochemical and histologic evidence of inflammation, proinflammatory cytokine and chemokine production, and inflammatory gene upregulation. RESULTS: After induction of pancreatic IRI, serum amylase and lactate dehydrogenase peaked at 6 hr and returned to baseline by 120 hr after injury in all groups. Mice undergoing 30 min of IRI demonstrated the greatest biochemical evidence of inflammation. Histologic scoring similarly demonstrated marked inflammation in mice subjected to 30-min IRI compared with controls. Serum cytokine/chemokine analysis demonstrated significant upregulation of granulocyte colony-stimulating factor, interferon γ, tumor necrosis factor α, interleukin (IL)-2, IL-1β, IL-6, chemokine (C-C motif) ligand-2, chemokine (C-C motif) ligand-5, chemokine (C-X-C motif) ligand-1, and macrophage inflammatory protein 2. A similar upregulation of ccl2, il1b, il6, fos, hspa1a, hspd1, and cd14 gene expression was detected by real-time polymerase chain reaction analysis of pancreatic tissue. CONCLUSIONS: This novel model of distal pancreatic IRI in the mouse demonstrates time-limited pancreatic inflammation and injury by histologic and biochemical indices. Inflammation may be, in part, a result of the immunologic effects of IL-1β, IL-6, and CCL-2. This model provides a method by which immunologic mechanisms of pancreatic IRI can be elucidated.
    Transplantation 02/2013; · 3.78 Impact Factor
  • Heather E Lynch, Gregory D Sempowski
    [Show abstract] [Hide abstract]
    ABSTRACT: This chapter provides protocols necessary for quantifying human, mouse, and nonhuman primate signal joint T cell receptor excision circles (sjTRECs) produced during T cell receptor alpha (TCRA) gene rearrangement. These non-replicated episomal circles of DNA are generated by the recombination process used to produce antigen-specific T cell receptors. The number of sjTRECs per mg of thymus tissue or per 100,000 lysed cells has been shown to be a molecular marker of thymopoiesis and naïve T cells. This technology is beneficial to investigators interested in quantitating the level of naïve T cell production occurring under various circumstances in a variety of systems, and complements traditional phenotypic analyses of thymopoiesis. This chapter specifically describes procedures required for rapid detection and quantitation of sjTRECs in thymus tissue or isolated cells using real-time quantitative polymerase chain reaction (PCR). The sjTREC assay system comprises species-specific forward and reverse primers for amplification of a unique site on the T cell receptor δ (TCRD) sjTREC, a fluorescently labeled (FAM/ZEN/IABkFQ) species-specific real-time probe, and a species-specific sjTREC DNA plasmid standard for quantitation.
    Methods in molecular biology (Clifton, N.J.) 01/2013; 979:147-59. · 1.29 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The continued development of nuclear weapons and the potential for thermonuclear injury necessitates the further understanding of the immune consequences after radiation combined with injury (RCI). We hypothesized that sublethal ionization radiation exposure combined with a full-thickness thermal injury would result in the production of immature myeloid cells. Mice underwent either a full-thickness contact burn of 20% total body surface area or sham procedure followed by a single whole-body dose of 5-Gy radiation. Serum, spleen, and peripheral lymph nodes were harvested at 3 and 14 days after injury. Flow cytometry was performed to identify and characterize adaptive and innate cell compartments. Elevated proinflammatory and anti-inflammatory serum cytokines and profound leukopenia were observed after RCI. A population of cells with dual expression of the cell surface markers Gr-1 and CD11b were identified in all experimental groups, but were significantly elevated after burn alone and RCI at 14 days after injury. In contrast to the T-cell-suppressive nature of myeloid-derived suppressor cells found after trauma and sepsis, myeloid cells after RCI augmented T-cell proliferation and were associated with a weak but significant increase in interferon γ and a decrease in interleukin 10. This is consistent with previous work in burn injury indicating that a myeloid-derived suppressor cell-like population increases innate immunity. Radiation combined injury results in the increase in distinct populations of Gr-1CD11b cells within the secondary lymphoid organs, and we propose these immature inflammatory myeloid cells provide innate immunity to the severely injured and immunocompromised host.
    Shock (Augusta, Ga.) 10/2012; 38(5):532-42. · 2.87 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Plague is caused by Yersinia pestis, a bacterium that disseminates inside of the host at remarkably high rates. Plague bacilli disrupt normal immune responses in the host allowing for systematic spread that is fatal if left untreated. How Y. pestis disseminates from the site of infection to deeper tissues is unknown. Dissemination studies for plague are typically performed in mice by determining the bacterial burden in specific organs at various time points. To follow bacterial dissemination during plague infections in mice we tested the possibility of using bioluminescence imaging (BLI), an alternative non-invasive approach. Fully virulent Y. pestis was transformed with a plasmid containing the luxCDABE genes, making it able to produce light; this lux-expressing strain was used to infect mice by subcutaneous, intradermal or intranasal inoculation. We successfully obtained images from infected animals and were able to follow bacterial dissemination over time for each of the three different routes of inoculation. We also compared the radiance signal from animals infected with a wild type strain and a Δcaf1ΔpsaA mutant that we previously showed to be attenuated in colonization of the lymph node and systemic dissemination. Radiance signals from mice infected with the wild type strain were larger than values obtained from mice infected with the mutant strain (linear regression of normalized values, P < 0.05). We demonstrate that BLI is useful for monitoring dissemination from multiple inoculation sites, and for characterization of mutants with defects in colonization or dissemination.
    BMC Microbiology 07/2012; 12:147. · 3.10 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Immunologic reconstitution after allogeneic hematopoietic cell transplantation is a critical component of successful outcome. Umbilical cord blood (UCB) transplantation in adult recipients is associated with slow and often inadequate immune recovery. We characterized the kinetics and extent of immune recovery in 95 adult recipients after a dual UCB (n = 29) and matched sibling donor (n = 33) or matched unrelated donor (n = 33) transplantation. All patients were treated with myeloablative conditioning. There were no differences in the immune recovery profile of matched sibling donor and matched unrelated donor recipients. Significantly lower levels of CD3+, CD4+, and CD8+ T cells were observed in UCB recipients until 6 months after transplantation. Lower levels of regulatory T cells persisted until 1 year after transplantation. Thymopoiesis as measured by TCR rearrangement excision circle was comparable among all recipients by 6 months after transplantation. In a subset of patients 1 year after transplantation with similar levels of circulating T cells and TCR rearrangement excision circle, there was no difference in TCR diversity. Compared to HLA-identical matched sibling donor and matched unrelated donor adult hematopoietic cell transplantation recipients, quantitative lymphoid recovery in UCB transplantation recipients is slower in the first 3 months, but these differences disappeared by 6 to 12 months after transplantation.
    Biology of blood and marrow transplantation: journal of the American Society for Blood and Marrow Transplantation 06/2012; 18(11):1664-1676.e1. · 3.15 Impact Factor

Publication Stats

3k Citations
521.93 Total Impact Points

Institutions

  • 2004–2014
    • Duke University
      Durham, North Carolina, United States
  • 1998–2014
    • Duke University Medical Center
      • • Duke Human Vaccine Institute
      • • Department of Pathology
      • • Department of Medicine
      • • Department of Pediatrics
      Durham, North Carolina, United States
  • 2013
    • University of North Carolina at Chapel Hill
      • Department of Microbiology and Immunology
      Chapel Hill, NC, United States
  • 2002
    • Max Planck Institute of Molecular Cell Biology and Genetics
      Dresden, Saxony, Germany