RESUMEN
UNLABELLED: Microneedle patches (MN) provide a novel method of vaccine delivery to the skin with the objective of targeting the large network of resident antigen-presenting cells to induce an efficient immune response. Our previous reports demonstrated that cutaneous delivery of inactivated influenza virus-coated MN to mice protects against lethal infection. Protection is correlated with sustained levels of anti-influenza virus serum antibodies, hemagglutination inhibition titers, and robust cellular responses that are often stronger than those generated by intramuscular vaccination. Here we dissect the early events occurring in murine skin after microneedle delivery of inactivated influenza virus. We demonstrate correlation of immunization against influenza virus with a local increase of cytokines important for recruitment of neutrophils, monocytes and dendritic cells at the site of immunization. We also observed prolonged antigen deposition, and migration of matured dendritic cells bearing influenza virus antigen from the skin. IMPORTANCE: The immunological mechanisms by which MN vaccination confers protective immunity are not well understood. The present study provides a first analysis of the early immune events after microneedle-based vaccination.
Asunto(s)
Inmunización/métodos , Vacunas contra la Influenza/administración & dosificación , Vacunas contra la Influenza/inmunología , Inyecciones Intradérmicas/métodos , Piel/inmunología , Animales , Citocinas/metabolismo , Células Dendríticas/inmunología , Femenino , Citometría de Flujo , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Monocitos/inmunología , Neutrófilos/inmunología , Piel/patología , Vacunas de Productos Inactivados/administración & dosificación , Vacunas de Productos Inactivados/inmunología , Imagen de Cuerpo EnteroRESUMEN
BACKGROUND: A major goal in influenza vaccine development is induction of serological memory and cellular responses to confer long-term protection and limit virus spread after infection. Here, we investigate induction of long-lived immunity against the 2009 H1N1 virus after skin vaccination. METHODS: BALB/c mice received a single dose of 5 µg inactivated A/California/04/09 virus via coated metal microneedles (MN) applied to skin or via subcutaneous injection. RESULTS: MN or subcutaneous vaccination elicited similar serum IgG and hemagglutination inhibition titers and 100% protection against lethal viral challenge 6 weeks after vaccination. Six months after vaccination, the subcutaneous group exhibited a 60% decrease in functional antibody titers and extensive lung inflammation after challenge with 10 × LD(50) of homologous virus. In contrast, the MN group maintained high functional antibody titers and IFN-γ levels, inhibition of viral replication, and no signs of lung inflammation after challenge. MN vaccination conferred complete protection against lethal challenge, whereas subcutaneous vaccination induced only partial protection. These findings were further supported by high numbers of bone marrow plasma cells and spleen antibody-secreting cells detected in the MN group. CONCLUSIONS: A single skin vaccination with MN induced potent long-lived immunity and improved protection against the 2009 H1N1 influenza virus, compared with subcutaneous injection.
Asunto(s)
Inmunidad Humoral/fisiología , Inmunidad Mucosa/fisiología , Memoria Inmunológica , Subtipo H1N1 del Virus de la Influenza A/inmunología , Vacunas contra la Influenza/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Animales , Anticuerpos Antivirales/fisiología , Células de la Médula Ósea/fisiología , Línea Celular , Perros , Femenino , Vacunas contra la Influenza/administración & dosificación , Inyecciones Intradérmicas , Inyecciones Subcutáneas , Ratones , Ratones Endogámicos BALB C , Bazo/fisiología , Replicación ViralRESUMEN
Influenza prophylaxis would benefit from a vaccination method enabling simplified logistics and improved immunogenicity without the dangers posed by hypodermic needles. Here we introduce dissolving microneedle patches for influenza vaccination using a simple patch-based system that targets delivery to skin's antigen-presenting cells. Microneedles were fabricated using a biocompatible polymer encapsulating inactivated influenza virus vaccine for insertion and dissolution in the skin within minutes. Microneedle vaccination generated robust antibody and cellular immune responses in mice that provided complete protection against lethal challenge. Compared to conventional intramuscular injection, microneedle vaccination resulted in more efficient lung virus clearance and enhanced cellular recall responses after challenge. These results suggest that dissolving microneedle patches can provide a new technology for simpler and safer vaccination with improved immunogenicity that could facilitate increased vaccination coverage.
Asunto(s)
Vacunas contra la Influenza/administración & dosificación , Gripe Humana/prevención & control , Agujas , Polímeros/administración & dosificación , Vacunación/métodos , Administración Cutánea , Adsorción , Animales , Formación de Anticuerpos/fisiología , Formas de Dosificación , Humanos , Inmunización Secundaria/métodos , Vacunas contra la Influenza/farmacocinética , Inyecciones Intradérmicas , Ratones , Agujas/estadística & datos numéricos , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/mortalidad , Infecciones por Orthomyxoviridae/patología , Infecciones por Orthomyxoviridae/prevención & control , Polímeros/farmacocinética , Vacunación/instrumentaciónRESUMEN
BACKGROUND: Influenza is a contagious disease caused by a pathogenic virus, with outbreaks all over the world and thousands of hospitalizations and deaths every year. Due to virus antigenic drift and short-lived immune responses, annual vaccination is required. However, vaccine coverage is incomplete, and improvement in immunization is needed. The objective of this study is to investigate a novel method for transdermal delivery using metal microneedle arrays (MN) coated with inactivated influenza virus to determine whether this route is a simpler and safer approach than the conventional immunization, capable to induce robust immune responses and confer protection against lethal virus challenge. METHODOLOGY/PRINCIPAL FINDINGS: Inactivated A/Aichi/2/68 (H3N2) influenza virus was coated on metal microneedle arrays and applied to mice as a vaccine in the caudal dorsal skin area. Substantial antibody titers with hemagglutination inhibition activity were detected in sera collected two and four weeks after a single vaccine dose. Challenge studies in mice with 5 x LD(50) of mouse adapted Aichi virus demonstrated complete protection. Microneedle vaccination induced a broad spectrum of immune responses including CD4+ and CD8+ responses in the spleen and draining lymph node, a high frequency of antigen-secreting cells in the lung and induction of virus-specific memory B-cells. In addition, the use of MN showed a dose-sparing effect and a strong Th2 bias when compared to an intramuscular (IM) reference immunization. CONCLUSIONS/SIGNIFICANCE: The present results show that delivery of inactivated influenza virus through the skin using metal microneedle arrays induced strong humoral and cellular immune responses capable of conferring protection against virus challenge as efficiently as intramuscular immunization, which is the standard vaccination route. In view of the convenience of delivery and the potential for self-administration, vaccine-coated metal microneedles may provide a novel and highly effective immunization method.
Asunto(s)
Administración Cutánea , Subtipo H3N2 del Virus de la Influenza A/inmunología , Vacunas contra la Influenza/administración & dosificación , Agujas , Vacunación/métodos , Animales , Formación de Anticuerpos/efectos de los fármacos , Diseño de Equipo , Inmunidad Celular/efectos de los fármacos , RatonesRESUMEN
OBJECTIVE: To extend our studies on the prolonged and differential effect of natalizumab on T lymphocyte numbers in the cerebrospinal fluid, we investigated the number and phenotypes of leukocytes and the expression of major histocompatibility complex (MHC) classes I and II in cerebral perivascular spaces (CPVS). We hypothesized that natalizumab reduces the number of antigen presenting cells in CPVS. DESIGN: A case-control study in which inflammatory cell numbers in the CPVS of cerebral tissue were assessed by immunohistochemical staining. SUBJECTS: A patient with multiple sclerosis (MS) who developed progressive multifocal leukoencephalopathy (PML) during natalizumab therapy. Controls included location-matched cerebral autopsy material of patients without disease of the central nervous system, patients with MS not treated with natalizumab, and patients with PML not associated with natalizumab therapy. RESULTS: The absolute number of CPVS in the patient with MS treated with natalizumab was significantly lower than in the control groups owing to extensive destruction of the tissue architecture. The expression of MHC class II molecules and the number of CD209+ dendritic cells were significantly decreased in the CPVS of the patient with MS treated with natalizumab. No CD4+ T cells were detectable. CONCLUSIONS: Our observations may explain the differential and prolonged effects of natalizumab therapy on leukocyte numbers in the cerebrospinal fluid.