CXCL5-secreting pulmonary epithelial cells drive destructive neutrophilic inflammation in tuberculosis

Successful host defense against numerous pulmonary infections depends on bacterial clearance by polymorphonuclear leukocytes (PMNs); however, excessive PMN accumulation can result in life-threatening lung injury. Local expression of CXC chemokines is critical for PMN recruitment. The impact of chemokine-dependent PMN recruitment during pulmonary Mycobacterium tuberculosis infection is not fully understood. Here, we analyzed expression of genes encoding CXC chemokines in M. tuberculosis–infected murine lung tissue and found that M. tuberculosis infection promotes upregulation of Cxcr2 and its ligand Cxcl5. To determine the contribution of CXCL5 in pulmonary PMN recruitment, we generated Cxcl5^–/– mice and analyzed their immune response against M. tuberculosis. Both Cxcr2^–/– mice and Cxcl5^–/– mice, which are deficient for only one of numerous CXCR2 ligands, exhibited enhanced survival compared with that of WT mice following high-dose M. tuberculosis infection. The resistance of Cxcl5^–/– mice to M. tuberculosis infection was not due to heightened M. tuberculosis clearance but was the result of impaired PMN recruitment, which reduced pulmonary inflammation. Lung epithelial cells were the main source of CXCL5 upon M. tuberculosis infection, and secretion of CXCL5 was reduced by blocking TLR2 signaling. Together, our data indicate that TLR2-induced epithelial-derived CXCL5 is critical for PMN-driven destructive inflammation in pulmonary tuberculosis.     

Primary and radiation induced skull base osteosarcoma: a systematic review of clinical features and treatment outcomes

Journal of Neuro-Oncology - We aim to systematically review and summarize the demographics, clinical features, management strategies, and clinical outcomes of primary and radiation-induced...     

Evaluation of the antidiarrheal and antioxidant properties of Justicia hypocrateriformis

Content: Justicia hypocrateriformis Vahl (Acanthaceae) is used as an herbal remedy for diarrhea in Cameroon folk medicine.

Objective: This study evaluates the antidiarrheal and antioxidant properties of the aqueous extract of J. hypocrateriformis (JH).

Materials and methods: Preliminary phytochemical screening and an acute toxicity testing of the extract were carried out. The antidiarrheal activity of JH extract (100, 250, and 500?mg/kg) was assessed at curative and preventive levels in castor oil-induced diarrhea in mice. The antioxidant activity was measured by ferric reducing antioxidant power (FRAP), total phenolic content, and radical scavenging activity.

Results: A high lethal dose (LD₅₀) of 14.35?g/kg obtained in acute toxicity implies the extract is not toxic. Phytochemical screening revealed the presence of phenols, tannins, flavonoids, saponins, anthraquinones, and anthocyanins. JH showed a significant protection against castor oil-induced diarrhea as evidenced by a decrease in the number of defecation and wet stool. JH (100–500?mg/kg, p.o.) produced a non-significant dose-dependent decrease in castor oil-induced intestinal transit in the preventive study. In the curative and in healthy mice study, the decrease was only significant at 500?mg/kg. JH possessed a radical scavenging activity with an IC₅₀ of 9.93?mg/ml compared to 4.90?mg/ml for catechin. JH FRAP of 2703.77?±?0?mg/g (catechin equiv) and phenolic concentration of 14?169.99?±?612.39?mg/g (catechin equiv) were also obtained.

Conclusion: Justicia hypocrateriformis extract possesses antidiarrheal activity supported by its antioxidant potential and phytochemical constituents.     

Priming of protective T cell responses against virus-induced tumors in mice with human immune system components

Many pathogens that cause human disease infect only humans. To identify the mechanisms of immune protection against these pathogens and also to evaluate promising vaccine candidates, a small animal model would be desirable. We demonstrate that primary T cell responses in mice with reconstituted human immune system components control infection with the oncogenic and persistent Epstein-Barr virus (EBV). These cytotoxic and interferon-γ–producing T cell responses were human leukocyte antigen (HLA) restricted and specific for EBV-derived peptides. In HLA-A2 transgenic animals and similar to human EBV carriers, T cell responses against lytic EBV antigens dominated over recognition of latent EBV antigens. T cell depletion resulted in elevated viral loads and emergence of EBV-associated lymphoproliferative disease. Both loss of CD4⁺ and CD8⁺ T cells abolished immune control. Therefore, this mouse model recapitulates features of symptomatic primary EBV infection and generates T cell–mediated immune control that resists oncogenic transformation.Mice are a preferred species for many avenues of immunological research in vivo. Because of the evolutionary divergence of mouse and man 65 million years ago, however, these two species have inhabited different ecological niches and have been challenged with minimally overlapping groups of pathogens. The human and mouse immune systems, evolving to meet these challenges, have therefore accumulated many differences (1), making genes related to immunity, together with genes involved in reproduction and olfaction, the most divergent between the two species (2).Pathogens that drive this divergence are those that exclusively infect either humans or mice with a high frequency of population penetration and life-threatening pathology. One such example is EBV, a human γ-herpesvirus that infects >90% of the human adult population. Humans are the only known natural reservoir for EBV, and this virus is associated with malignancies of lymphocyte and epithelial cell origin (3, 4). EBV is thought to cause or contribute to these life-threatening tumors by its ability to induce proliferation and to protect infected cells from apoptosis (5). All EBV-associated lymphomas and carcinomas express latent EBV antigens, whereas lytic EBV particle production does not cause significant pathology. Latent EBV antigen expression differs between the various EBV-associated malignancies. Burkitt lymphoma expresses only the nuclear antigen 1 of EBV (EBNA1) as the sole latent gene product (latency I). In contrast, Hodgkin lymphoma and nasopharyngeal carcinoma express, in addition, one or both latent membrane proteins (LMPs), LMP1 and 2 (latency II). Only in immune-compromised patients, like HIV-infected individuals or transplant recipients, do B cell lymphomas occur that express the five additional EBNA proteins 2, 3A, B, C, and LP (latency III). In addition, all EBV-infected B cells express small nontranslated virally encoded RNAs, including the EBV-encoded RNAs (EBERs). All three types of EBV latencies can also be found in healthy EBV carriers, and this graded EBV protein expression depends on the B cell differentiation stage of the infected cell (6, 7). Healthy virus carriers therefore have already established tumor-associated latent EBV protein expression. Hence, the increased risk of immune-compromised individuals to develop EBV-associated tumors indicates loss of EBV-specific immune control that normally prevents viral tumorigenesis.T cells constitute the decisive component of EBV-specific immune control against virus-associated malignancies because adoptive transfer of EBV-specific T cell lines can eradicate EBV-associated posttransplant lymphomas (8). During primary infection, CD8⁺ T cells targeting epitopes of EBV lytic antigens make up the majority of EBV-specific T cells, but T cells targeting latent epitopes can also be detected. Different latency patterns confer different degrees of immunogenicity for recognition by cytotoxic CD8⁺ and helper CD4⁺ T cells (9, 10). In healthy EBV carriers, the EBNA3 proteins are the dominant targets of CD8⁺ T cell responses, whereas EBNA1, 2, and 3C are the most consistently recognized CD4⁺ T cell antigens. In contrast, the LMPs are subdominant T cell antigens. At least a subset of investigated CD4⁺ and CD8⁺ T clones, however, recognize both dominant and subdominant specificities directly on EBV-infected cells. Both CD4⁺ and CD8⁺ T cells therefore contribute to immune control against virus-infected cells (11), in addition to maintenance of functional CD8⁺ T cell memory by CD4⁺ T cells (12).The lack of an animal model of EBV infection prevents assignment of a protective value to the known T cell reactivities. Design and evaluation of vaccines against EBV-associated tumors and symptomatic primary infection, however, require knowledge about the degree of protection conferred by different EBV-specific immune responses. Mouse models that partially reconstitute human immune system components after engraftment of hematopoietic progenitor cells (HPCs) are of particular interest to study vaccine candidates and EBV-specific immune responses in vivo. In this respect, three novel models of human immune system reconstitution are being considered. The first one uses BALB/c Rag2^−/− γ_c^−/− mice, which reconstitute macrophages, T cells, B cells, natural killer cells, and dendritic cells after neonatal intrahepatic HPC transfer (13, 14). The second model reconstitutes NOD-scid γ_c^−/− mice by intravenous injection of human HPCs (15, 16), which also leads to significant development of human myeloid and lymphoid cells. Finally, the most labor-intensive model is the BLT mouse, which requires implantation of human fetal liver and thymus pieces under the kidney capsule of NOD-scid mice in addition to intravenous HPC injection (17). Immune compartment reconstitution in peripheral blood of BLT mice is very similar to human. However, although signs of primary immune responses were reported in all three of these current mouse models of human immune system reconstitution, the protective value of this immunocompetence and, thus, the potential of these in vivo systems as challenge models for vaccine development against pathogens with exclusive tropism for humans has not been evaluated. Therefore, we analyzed the ability of mice with reconstituted immune system components to exert antigen-specific T cell–mediated control of EBV infection and EBV-associated lymphomas.     

Targeting the nuclear antigen 1 of Epstein-Barr virus to the human endocytic receptor DEC-205 stimulates protective T-cell responses

Dendritic cells (DCs) express many endocytic receptors that deliver antigens for major histocompatibility class (MHC) I and II presentation to CD8(+) and CD4(+) T cells, respectively. Here, we show that targeting Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA1) to one of them, the human multilectin DEC-205 receptor, in the presence of the DC maturation stimulus poly(I:C), expanded EBNA1-specific CD4(+) and CD8(+) memory T cells, and these lymphocytes could control the outgrowth of autologous EBV-infected B cells in vitro. In addition, using a novel mouse model with reconstituted human immune system components, we demonstrated that vaccination with alphaDEC-205-EBNA1 antibodies primed EBNA1-specific IFN-gamma-secreting T cells and also induced anti-EBNA1 antibodies in a subset of immunized mice. Because EBNA1 is the one EBV antigen that is expressed in all proliferating cells infected with this virus, our data suggest that DEC-205 targeting should be explored as a vaccination approach against symptomatic primary EBV infection and against EBV-associated malignancies.     

NK cell survival mediated through the regulatory synapse with human DCs requires IL-15Ralpha

DCs activate NK cells during innate immune responses to viral infections. However, the composition and kinetics of the immunological synapse mediating this interaction are largely unknown. Here, we report the rapid formation of an immunological synapse between human resting NK cells and mature DCs. Although inhibitory NK cell receptors were polarized to this synapse, where they are known to protect mature DCs from NK cell lysis, the NK cell also received activation signals that induced mobilization of intracellular calcium and CD69 upregulation. The high-affinity component of the receptor for IL-15, IL-15Ralpha, accumulated at the synapse center on NK cells, and blocking of IL-15Ralpha increased NK cell apoptosis and diminished NK cell survival during their interaction with DCs. Furthermore, IL-15Ralpha-deficient NK cells, obtained from donors with a history of infectious mononucleosis, showed diminished survival in culture with DCs. Synapse formation was required for IL-15Ralpha-mediated NK cell survival, because synapse disruption by adhesion molecule blocking decreased DC-induced NK cell survival. These results identify what we believe to be a novel regulatory NK cell synapse with hallmarks of spatially separated inhibitory and activating interactions at its center. We suggest that this synapse formation enables optimal NK cell activation by DCs during innate immune responses.     

Determination of cocaine,cocaine metabolites and cannabinoids in single hairs by MALDI Fourier transform mass spectrometry – preliminary results

In a preliminary test, four single hairs of a drug abuser were analyzed for the presence of drugs by MALDI Fourier transform mass spectrometry (MALDI‐FTMS). Washed hair strains were directly fixed on a sample plate using pressure stable, double‐sided adhesive tape; α‐cyano‐4‐hydroxycinnamic acid matrix was manually spotted onto the hair strains. FTMS full scans were obtained moving from the hair root region towards the hair tip. Cocaine (accurate m/z ratio 304.15433) was identified mostly from the root of the hair and then later again towards the hair tip. This was confirmed by analysis of a second hair. Additionally cocaine metabolites with m/z ratio 290.13868 (benzoylecgonine), and m/z 318.16998 (cocaethylene) were detected for plausibility control. Using the MALDI technique, time‐related information was obtained concerning the behavioural pattern of the consumer with high resolution compared to conventional procedures. However, in two hairs of the same individual which were analyzed under the same conditions, negative results were achieved. These preliminary results confirm the applicability of MALDI‐MS for the determination of drugs and pharmaceuticals in hair samples being useful in forensic toxicology. The high chronological resolution allows an enhanced interpretation concerning the periods of drug administration. However, the negative results with two negative hairs have also demonstrated that hair analysis of single hairs can lead to misinterpretation. Different growth rates have to be considered, and particularly the phenomenon of different growth phases (anagen, catagen, telogen) require attention. Copyright © 2013 John Wiley & Sons, Ltd.