Fibrin deposition and exudation of plasma fibrinogen (Fg) have long been recognized as hallmarks of inflammation, cardiovascular disease and neoplasia. The Fg-b 15-42 domain binds to the endothelial cell adhesion molecule, VE-cadherin, promoting endothelial cell proliferation, angiogenesis and leukocyte diapedesis. Furthermore, spontaneous blood-borne and lymphatic metastasis of some types of tumor emboli requires plasma fibrin(ogen); however, the molecular mechanisms by which this occurs are poorly understood. We sought to determine whether Fg-b 15-42 and VEcadherin binding interactions promote endothelial barrier permeability and breast cancer cell transendothelial migration (TEM) using transwell insert culture systems. Synthetic peptides containing/missing residues b 15-17 critical for In 1865, Armand Trousseau reported the observation that patients with an increased incidence of coagulopathies would later manifest visceral malignancies, which became known as Trousseau's syndrome. Ironically, Trousseau diagnosed himself with his own syndrome in 1866 and died of gastric cancer the following year. Causal factors linked to the prothrombotic state of Trousseau's syndrome also facilitate cancer metastasis, including thrombin, tissue factor, selectins, platelets, endothelial cells (EC) and fibrin. 1 Deposition of fibrinogen (Fg) 5 and fibrin commonly occurs within the stroma of most solid tumors, 2 and elevated levels of plasma Fg and fibrin degradation products (FDPs) correlate positively with lymph node involvement and metastasis of colon, ovarian, lung and breast cancers. 1 Studies by Degen and co-workers 3-5 have firmly established that plasma fibrin(ogen) and platelets play critical roles in spontaneous cancer metastasis through both the circulation and lymphatic systems, in part by protecting tumor cells from natural killer cell-mediated lysis.Expression of interleukin (IL)-6 during systemic inflammation upregulates expression of specific plasma proteins in the liver, including Fg. 6 Excessive fibrin deposition is accompanied by local expression of proinflammatory mediators, vascular leakage, and inflammatory cell recruitment and activation leading to amplification of the inflammatory response. 2,7,8 Residues 15-42 on the b chain of Fg have been implicated in functional attributes ascribed to fibrin(ogen). Although the primary structure of fibrinopeptide B (FPB) is poorly conserved across species, the fibrin b 15-42 domain is highly conserved, implying evolutionary conservation of function. 9 The b 15-42 region constitutes a cryptic domain in soluble Fg that is exposed in fibrin after thrombin cleavage. 10 Newly exposed residues, b15-GHRP-18, promote lateral aggregation of fibrin monomers during polymerization and insoluble clot formation in secondary hemostasis. 10 Furthermore, exposure of the b 15-42 domain mediates fibrin binding to EC surfaces, 11 promotes EC adhesion and spreading, 12 and stimulates proliferation of EC, fibroblasts and cancer cells. 13,14 Cadherins mediate homophilic cell-cell adhesion...

Vaccine responses vary by geographic location. We have previously described how HIV-associated inflammation leads to fibrosis of secondary lymph nodes (LNs) and T cell depletion. We hypothesized that other infections may cause LN inflammation and fibrosis, in a process similar to that seen in HIV infection, which may lead to T cell depletion and affect vaccine responses. We studied LNs of individuals from Kampala, Uganda, before and after yellow fever vaccination (YFV) and found fibrosis in LNs that was similar to that seen in HIV infection. We found blunted antibody responses to YFV that correlated to the amount of LN fibrosis and loss of T cells, including T follicular helper cells. These data suggest that LN fibrosis is not limited to HIV infection and may be associated with impaired immunologic responses to vaccines. This may have an impact on vaccine development, especially for infectious diseases prevalent in the developing world.

Summary Background Antigenic drift and shift of influenza viruses require frequent reformulation of influenza vaccines. In addition, seasonal influenza vaccines are often mismatched to the epidemic influenza strains. This stresses the need for a universal influenza vaccine. Methods BALB/c mice were vaccinated with the trivalent live attenuated (LAIV; FluMist) or inactivated (TIV; FluZone) influenza vaccines and challenged with PR8 (H1N1), FM/47 (H1N1), or HK/68 (H3N2) influenza virus. Cytokines and antibody responses were tested by ELISA. Furthermore, different LAIV dosages were applied in BALB/c mice. LAIV vaccinated mice were also depleted of T-cells and challenged with PR8 virus. Results LAIV induced significant protection against challenge with the non-vaccine strain PR8 influenza virus. Furthermore, protective immunity against PR8 was dose-dependent. Of note, interleukin 2 and interferon gamma cytokine secretion in the lung alveolar fluid were significantly elevated in mice vaccinated with LAIV. Moreover, T-cell depletion of LAIV vaccinated mice compromised protection, indicating that T-cell-mediated immunity is required. In contrast, passive transfer of sera from mice vaccinated with LAIV into naïve mice failed to protect against PR8 challenge. Neutralization assays in vitro confirmed that LAIV did not induce cross-strain neutralizing antibodies against PR8 virus. Finally, we showed that three doses of LAIV also provided protection against challenge with two additional heterologous viruses, FM/47 and HK/68. Conclusions These results support the potential use of the LAIV as a universal influenza vaccine under a prime–boost vaccination regimen.

The influenza virus is a respiratory pathogen with a negative-sense, segmented RNA genome. Construction of recombinant influenza viruses in the laboratory was reported starting in the 1980s. Within a short period of time, pioneer researchers had devised methods that made it possible to construct influenza viral vectors from cDNA plasmid systems. Herein, we discuss the evolution of influenza virus reverse genetics, from helper virus-dependent systems, to helper virus-independent 17-plasmid systems, and all the way to 3- and 1- plasmid systems. Successes in the modification of different gene segments for various applications, including vaccine and gene therapies are highlighted.

Microvascular plasma leakage is the hallmark of dengue hemorrhagic fever and dengue shock syndrome. The precise molecular mechanisms leading to microvascular leakage are yet to be determined, but dengue virus (DENV) infection and consequent endothelial cell death has been suggested as its major cause. However, the extent of endothelial cell permissiveness to DENV infection and the magnitude of cell death following DENV infection are controversial. To clarify this issue, we analyzed the kinetics and consequences of DENV infection of human umbilical vein endothelial cells (HUVEC) using a novel molecularly cloned DENV2-16681 virus. Viral replication was detected as early as 24 hr post-infection by RT-PCR and plaque assays. However, merely 2% of HUVEC were DENV antigen-positive even after 96 hr of infection as measured by the FACS indirect immunofluorescence assays. Unlike monocytes/macrophages, HUVEC did not support antibody dependent enhancement of dengue viral infection due to a lack of FcgammaRI and FcgammaRII. Furthermore, DENV infection did not increase HUVEC apoptosis as compared to mock-infected cells. Because in vitro only a small percentage of endothelial cells were productively infected in vitro with no significant apoptosis occurring in either infected or bystander cells, it would be important to re-examine whether direct dengue viral infection of endothelium is the major cause of the extensive vascular leakage observed in patients with dengue hemorrhagic fever and dengue shock syndrome.

Introduction Chikungunya virus (CHIKV) is a re-emerging pathogen responsible for causing outbreaks of febrile disease accompanied with debilitating joint pain. Symptoms typically persist for two weeks, but more severe and chronic chikungunya illnesses have been reported, especially in the elderly. Currently, there are no licensed vaccines or antivirals against CHIKV available. In this study, we combined a CHIK virus-like particle (VLP) vaccine with different adjuvants to enhance immunogenicity and protection in both, adult and aged mice. Methods CHIK VLP-based vaccines were tested in 6-8-week-old (adult) and 18-24-month-old (aged) female C57BL/6J mice. Formulations contained CHIK VLP alone or adjuvants: QuilA, R848, or Imject Alum. Mice were vaccinated three times via intramuscular injections. CHIKV-specific antibody responses were characterized by IgG subclass using ELISA, and by microneutralization assays. In addition, CHIKV infections were characterized in vaccinated and non-vaccinated adult mice and compared to aged mice. Results In adult mice, CHIKV infection of the right hind foot induced significant swelling, which peaked by day 7 post-infection at approximately 170% of initial size. Viral titers peaked at 2.53 × 10 10 CCID 50 /ml on day 2 post-infection. Mice vaccinated with CHIK VLP-based vaccines developed robust anti-CHIKV-specific IgG antibody responses that were capable of neutralizing CHIKV in vitro . CHIK VLP alone or CHIK plus QuilA administered by IM injections protected 100% of mice against CHIKV. In contrast, the antibody responses elicited by the VLP-based vaccines were attenuated in aged mice, with negligible neutralizing antibody titers detected. Unvaccinated, aged mice were resistant to CHIKV infection, while vaccination with CHIKV VLPs exacerbated disease. Conclusions Unadjuvanted CHIK VLP vaccination elicits immune responses that protect 100% of adult mice against CHIKV infection. However, an improved vaccine/adjuvant combination is still necessary to enhance the protective immunity against CHIKV in the aged.

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Streptococcus pneumoniae is a major bacterial respiratory pathogen. Current licensed pneumococcal polysaccharide and polysaccharide-protein conjugate vaccines are administered by an intramuscular injection. In order to develop a new-generation vaccine that can be administered in a needle-free mucosal manner, we have constructed early 1 and 3 gene regions (E1/E3) deleted, replicationdefective adenoviral vectors encoding pneumococcal surface antigen A (PsaA), the N-fragment of pneumococcal surface protein A (N-PspA), and the detoxified mutant pneumolysin (PdB) from S. pneumoniae strain D39. Intranasal vaccination with the three adenoviral vectors (Ad/PsaA, Ad/N-PspA, and Ad/PdB) in mice resulted in robust antigen-specific serum immunoglobulin G responses, as demonstrated by an enzyme-linked immunosorbent assay. In addition, nasal mucosal vaccination with the combination of the three adenoviral vectors conferred protection against S. pneumoniae strain D39 colonization in mouse lungs. Taken together, these data demonstrate the feasibility of developing a mucosal vaccine against S. pneumoniae using recombinant adenoviruses for antigen delivery.