RESUMO
BACKGROUND: Nonlive vaccine approaches that are simple to deliver and stable at room temperature or 2-8°C could be advantageous in controlling future Ebola virus (EBOV) outbreaks. Using an immunopotent DNA vaccine that generates protection from lethal EBOV challenge in small animals and nonhuman primates, we performed a clinical study to evaluate both intramuscular (IM) and novel intradermal (ID) DNA delivery. METHODS: Two DNA vaccine candidates (INO-4201 and INO-4202) targeting the EBOV glycoprotein (GP) were evaluated for safety, tolerability, and immunogenicity in a phase 1 clinical trial. The candidates were evaluated alone, together, or in combination with plasmid-encoded human cytokine interleukin-12 followed by in vivo electroporation using either the CELLECTRA® IM or ID delivery devices. RESULTS: The safety profile of all 5 regimens was shown to be benign, with the ID route being better tolerated. Antibodies to EBOV GP were generated by all 5 regimens with the fastest and steepest rise observed in the ID group. Cellular immune responses were generated with every regimen. CONCLUSIONS: ID delivery of INO-4201 was well tolerated and resulted in 100% seroreactivity after 2 doses and elicited interferon-γ T-cell responses in over 70% of subjects, providing a new approach for EBOV prevention in diverse populations. Clinical Trials Registration. NCT02464670.
Assuntos
Vacinas contra Ebola/efeitos adversos , Vacinas contra Ebola/imunologia , Imunidade Celular/imunologia , Imunidade Humoral/imunologia , Vacinas de DNA/efeitos adversos , Vacinas de DNA/imunologia , Adolescente , Adulto , Anticorpos Antivirais/imunologia , Ebolavirus/imunologia , Eletroporação/métodos , Feminino , Glicoproteínas/imunologia , Voluntários Saudáveis , Doença pelo Vírus Ebola/imunologia , Humanos , Injeções Intradérmicas/métodos , Interleucina-12/imunologia , Masculino , Pessoa de Meia-Idade , Temperatura , Vacinação/métodos , Adulto JovemRESUMO
The major ion chemistry of the ocean has been assumed to be controlled by river input, hydrothermal circulation at mid-ocean ridges, carbonate production, and low-temperature alteration of seafloor basalt, but marine chemical budgets remain difficult to balance. Here we propose that large-scale groundwater flow and diagenetic reactions in continental shelf sediments have been overlooked as an important contributor to major ion budgets in the ocean. Based on data synthesized from 17 passive margin basins, continental shelves contribute fluid exchanges comparable to hydrothermal circulation at mid-ocean ridges. Chemical exchange is similarly significant, indicating removal of Mg2+ from the oceans at rates similar to mid-ocean ridge convection. Continental shelves likely contribute Ca2+ and K+ to the oceans at rates that, in combination with low-temperature basalt alteration, can close current budget deficits. Flow and reaction in continental shelf sediments should be included in a new generation of studies addressing marine isotope budgets.
RESUMO
OBJECTIVES: To determine whether granulocyte colony-stimulating factor receptor (G-CSFR) autocrine signaling promotes endothelial cell adhesion and invasion of bladder cancer cells through a beta1-integrin-mediated pathway. A significant fraction of invasive bladder carcinomas express both G-CSF and G-CSFR. Bladder carcinoma cell line 5637 constitutively secretes G-CSF but lacks G-CSFR expression. Thus, we studied the effects of G-CSFR expression on cell adhesion and invasion in this unique model system. METHODS: Flow cytometry and adhesion assay were performed to detect expression of beta1-integrin in G-CSFR-expressing 5637 cells and adhesion of these cells to human umbilical vein endothelial cell, respectively. Furthermore, an invasion chamber assay was done with the 5637 cells. Next, we used the G-CSF-specific antibody, siRNA, and a truncated version of G-CSFR (GR19) to block G-CSFR autocrine loop in these cells. We also used a beta1-integrin-specific neutralizing antibody in the adhesion and invasion assays with the 5637 cells. RESULTS: G-CSFR-mediated increased expression (approximately threefold) of beta1-integrin is significantly abrogated by G-CSF specific antibody or siRNA in 5637 cells. GR19 also completely blocked beta1-integrin expression. G-CSFR signaling increased adhesion (approximately 2.5-fold) of 5637 cells to human umbilical vein endothelial cells, which are potently blocked by beta1-integrin-specific antibody. G-CSF/G-CSFR autocrine signaling significantly increased the invasiveness of 5637 cells (approximately 10-fold), which was reduced by either attenuating G-CSF production (G-CSF-specific antibody and siRNA) or interfering with G-CSFR signaling (GR19). Furthermore, beta1-integrin-specific antibody completely blocked G-CSFR-mediated invasion of 5637 cells. CONCLUSIONS: Autocrine G-CSF/G-CSFR signaling in bladder cancer can significantly contribute to cancer cell adhesion and invasion in a beta1-integrin-dependent manner.
Assuntos
Integrina beta1/fisiologia , Receptores de Fator Estimulador de Colônias de Granulócitos/fisiologia , Neoplasias da Bexiga Urinária/fisiopatologia , Adesividade , Comunicação Autócrina , Linhagem Celular Tumoral , Endotélio Vascular/patologia , Citometria de Fluxo , Fator Estimulador de Colônias de Granulócitos/farmacologia , Humanos , Integrina beta1/metabolismo , Invasividade Neoplásica , Transdução de Sinais , Neoplasias da Bexiga Urinária/patologiaRESUMO
OBJECTIVES: To examine the association of granulocyte colony-stimulating factor (G-CSF) and G-CSF receptor (G-CSFR) expression with increased beta1-integrin expression and determine the ability of autocrine G-CSFR signaling to promote bladder cancer cell adhesion by way of beta1-integrin. beta1-integrin is expressed at higher levels in more invasive bladder carcinoma cells and participates in the process of tissue invasion. Cancer cell invasion and metastasis in some ways mimic normal neutrophil behavior. In neutrophils, G-CSF acts through its receptor to enhance adhesion and migration. A significant fraction of bladder carcinoma has been reported to express both G-CSF and G-CSFR. METHODS: We examined bladder carcinoma tissue samples obtained from segmental or radical cystectomy specimens for expression of G-CSF, G-CSFR, and beta1-integrin using reverse transcriptase-polymerase chain reaction, Western blot, and immunohistochemical methods. We determined the G-CSFR-mediated beta1-integrin adhesion using a static adhesion assay and the bladder cancer cell line 5637. RESULTS: Eleven of 14 bladder cancer samples expressed G-CSFR. All 11 G-CSFR positive tumors also expressed G-CSF, and the G-CSF/G-CSFR positive tumors had elevated beta1-integrin protein levels. All but one G-CSFR negative tumor demonstrated low beta1-integrin protein levels. In four G-CSF/G-CSFR positive tumors for which distant urothelium was available for examination, G-CSF, G-CSFR, and beta1-integrin expression was also increased. In the 5637 cell line, we demonstrated G-CSFR-mediated upregulation of beta1-integrin-dependent adhesion to fibronectin and laminin. CONCLUSIONS: G-CSF/G-CSFR expression in some bladder cancers appears to be an early event during malignant transformation that increases beta1-integrin expression and adhesion and thereby may promote tissue invasion.