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1.
Article in English | MEDLINE | ID: mdl-29600197

ABSTRACT

Herpes Simplex Virus type 1 (HSV-1), a neurotropic pathogen widespread in human population, infects the enteric nervous system (ENS) in humans and rodents and causes intestinal neuromuscular dysfunction in rats. Although infiltration of inflammatory cells in the myenteric plexus and neurodegeneration of enteric nerves are common features of patients suffering from functional intestinal disorders, the proof of a pathogenic link with HSV-1 is still unsettled mainly because the underlying mechanisms are largely unknown. In this study we demonstrated that following intragastrical administration HSV-1 infects neurons within the myenteric plexus resulting in functional and structural alterations of the ENS. By infecting mice with HSV-1 replication-defective strain we revealed that gastrointestinal neuromuscular anomalies were however independent of viral replication. Indeed, enteric neurons exposed to UV-inactivated HSV-1 produced monocyte chemoattractant protein-1 (MCP-1/CCL2) to recruit activated macrophages in the longitudinal muscle myenteric plexus. Infiltrating macrophages produced reactive oxygen and nitrogen species and directly harmed enteric neurons resulting in gastrointestinal dysmotility. In HSV-1 infected mice intestinal neuromuscular dysfunctions were ameliorated by in vivo administration of (i) liposomes containing dichloromethylene bisphosphonic acid (clodronate) to deplete tissue macrophages, (ii) CCR2 chemokine receptor antagonist RS504393 to block the CCL2/CCR2 pathway, (iii) Nω-Nitro-L-arginine methyl ester hydrochloride (L-NAME) and AR-C 102222 to quench production of nitrogen reactive species produced via iNOS. Overall these data demonstrate that HSV-1 infection makes enteric neurons recruit macrophages via production of a specific chemoattractant factor. The resulting inflammatory reaction is mandatory for intestinal dysmotility. These findings provide insights into the neuro-immune communication that occurs in the ENS following HSV-1 infection and allow recognition of an original pathophysiologic mechanism underlying gastrointestinal diseases as well as identification of novel therapeutic targets.


Subject(s)
Enteric Nervous System/drug effects , Enteric Nervous System/virology , Gastrointestinal Motility/drug effects , Herpes Simplex/metabolism , Herpesvirus 1, Human/pathogenicity , Macrophages/metabolism , Neurons/drug effects , Adaptive Immunity , Animals , Arginine/analogs & derivatives , Arginine/metabolism , Chemokine CCL2/metabolism , Clodronic Acid , Disease Models, Animal , Enteric Nervous System/metabolism , Enteric Nervous System/pathology , Herpes Simplex/immunology , Herpes Simplex/pathology , Herpes Simplex/virology , Ileum/immunology , Ileum/pathology , Ileum/virology , Inflammation/metabolism , Liposomes/metabolism , Male , Mice , Mice, Inbred C57BL , Myenteric Plexus/drug effects , Myenteric Plexus/metabolism , Myenteric Plexus/pathology , Myenteric Plexus/virology , NG-Nitroarginine Methyl Ester/metabolism , Neurons/virology , Rats , Reactive Nitrogen Species/metabolism , Reactive Nitrogen Species/toxicity , Reactive Oxygen Species/metabolism , Reactive Oxygen Species/toxicity , Receptors, Chemokine , Virus Internalization , Virus Replication
2.
Vaccine ; 34(19): 2216-24, 2016 Apr 27.
Article in English | MEDLINE | ID: mdl-27002499

ABSTRACT

Mucosal HSV infection remains a public health issue in developing and developed world. However, an effective vaccine is still missing, partly because of the incomplete knowledge of correlates of protection. In this study we have investigated the kinetics and quality of immunity elicited by an attenuated HSV1 vector expressing the immunomodulatory Tat protein of HIV-1 (HSV1-Tat). Animals were immunized by intravaginal (IVag) or intradermal (ID) route with HSV1-Tat or with a control HSV1 vector expressing the LacZ gene (HSV1-LacZ) and immune responses were characterized in different anatomical districts. IVag immunization with HSV1-Tat enhanced both expansion and memory phases of HSV-specific immunodominant CD8 responses at systemic, but not local, level and induced short- and long-term protection against mucosal challenge. Conversely, ID immunization with HSV1-Tat favored HSV-subdominant CD8 responses, which protected mice only at early time points after immunization. IVag immunization, in particular with HSV1-Tat, compared to ID immunization, induced the differentiation of CD8(+) T lymphocytes into short-lived effector (SLEC) and effector memory (Tem) cells, generating more robust recall responses associated with increased control of virus replication. Notably, systemic SLEC and Tem contributed to generate protective local secondary responses, demonstrating their importance for mucosal control of HSV. Finally, IgG responses were observed mostly in IVag HSV1-Tat immunized animals, although seemed dispensable for protection, which occurred even in few IgG negative mice. Thus, HSV1 vectors expressing Tat induce protective anti-HSV1 immune responses.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , Herpes Simplex/prevention & control , Herpesvirus Vaccines/immunology , Immunity, Mucosal , tat Gene Products, Human Immunodeficiency Virus/immunology , Administration, Intravaginal , Animals , Antibodies, Viral/blood , Female , Herpesvirus 1, Human , Immunoglobulin G/blood , Immunologic Memory , Mice , Mice, Inbred C57BL
3.
PLoS One ; 9(7): e100844, 2014.
Article in English | MEDLINE | ID: mdl-25033084

ABSTRACT

Herpes simplex virus types 1 and 2 (HSV1 and HSV2) are common infectious agents in both industrialized and developing countries. They cause recurrent asymptomatic and/or symptomatic infections, and life-threatening diseases and death in newborns and immunocompromised patients. Current treatment for HSV relies on antiviral medications, which can halt the symptomatic diseases but cannot prevent the shedding that occurs in asymptomatic patients or, consequently, the spread of the viruses. Therefore, prevention rather than treatment of HSV infections has long been an area of intense research, but thus far effective anti-HSV vaccines still remain elusive. One of the key hurdles to overcome in anti-HSV vaccine development is the identification and effective use of strategies that promote the emergence of Th1-type immune responses against a wide range of epitopes involved in the control of viral replication. Since the HIV1 Tat protein has several immunomodulatory activities and increases CTL recognition of dominant and subdominant epitopes of heterologous antigens, we generated and assayed a recombinant attenuated replication-competent HSV1 vector containing the tat gene (HSV1-Tat). In this proof-of-concept study we show that immunization with this vector conferred protection in 100% of mice challenged intravaginally with a lethal dose of wild-type HSV1. We demonstrate that the presence of Tat within the recombinant virus increased and broadened Th1-like and CTL responses against HSV-derived T-cell epitopes and elicited in most immunized mice detectable IgG responses. In sharp contrast, a similarly attenuated HSV1 recombinant vector without Tat (HSV1-LacZ), induced low and different T cell responses, no measurable antibody responses and did not protect mice against the wild-type HSV1 challenge. These findings strongly suggest that recombinant HSV1 vectors expressing Tat merit further investigation for their potential to prevent and/or contain HSV1 infection and dissemination.


Subject(s)
Herpes Simplex Virus Vaccines/immunology , Herpes Simplex/prevention & control , Herpesvirus 1, Human/genetics , Herpesvirus 1, Human/immunology , tat Gene Products, Human Immunodeficiency Virus/genetics , 3T3 Cells , Animals , Antibodies, Viral/immunology , Cell Line , Chlorocebus aethiops , Epitopes, T-Lymphocyte/immunology , HIV-1/immunology , HeLa Cells , Humans , Immunoglobulin G/immunology , Lac Operon/genetics , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , T-Lymphocytes, Cytotoxic/immunology , Th1 Cells/immunology , Vaccination , Vero Cells , tat Gene Products, Human Immunodeficiency Virus/immunology
4.
Gene Ther ; 11(13): 1087-98, 2004 Jul.
Article in English | MEDLINE | ID: mdl-15057264

ABSTRACT

To improve the utility of herpes simplex virus type 1 (HSV-1) vectors for gene therapy, the viral envelope needs to be manipulated to achieve cell-specific gene delivery. In this report, we have engineered an HSV-1 mutant virus, KgBpK(-) gC(-), deleted for the glycoprotein C (gC) and the heparan sulfate-binding domain (pK) of gB, in order to express gC:preS1 and gC:preS1 active peptide (preS1ap) fusion molecules. PreS1, and a 27 amino acid active peptide inside preS1 (preS1ap), are supposed to be the molecules that the human hepatitis B virus (HBV) needs to bind specifically to hepatocytes. Biochemical analysis demonstrated that the gC:preS1ap fusion molecule was expressed and incorporated into the envelope of the recombinant HSV-1 virus KgBpK(-)gC:preS1ap. Moreover, KgBpK(-)gC:preS1ap recombinant virus gained a specific binding activity to an hepatoblastoma cell line (HepG2) with a consequent productive infection. In addition, anti-preS1-specific antibodies were shown to neutralize recombinant virus infectivity, and a synthetic preS1ap peptide was able to elute KgBpK(-)gC:preS1ap virus bound on HpeG2 cells. These data provide further evidence that HSV-1 can productively infect cells through a specific binding to a non-HSV-1 receptor. Furthermore, these data strongly support the hypothesis that the HBV preS1ap molecule is an HBV ligand to hepatocytes.


Subject(s)
Genetic Vectors/metabolism , Hepatitis B Surface Antigens/metabolism , Hepatocytes/metabolism , Herpesvirus 1, Human/metabolism , Protein Precursors/metabolism , Recombinant Fusion Proteins/metabolism , Animals , Cell Line, Tumor , Chlorocebus aethiops , Gene Deletion , Genetic Engineering , Herpesvirus 1, Human/genetics , Humans , Protein Binding , Transfection , Vero Cells , Viral Envelope Proteins/genetics
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