Sam Hou

Cross-priming of CD8+ T cells stimulated by virus-induced type I interferon.

CD8+ T cell responses can be generated against antigens that are not expressed directly within antigen-presenting cells (APCs), through a process known as cross-priming. To initiate cross-priming, APCs must both capture extracellular antigen and receive specific activation signals. We have investigated the nature of APC activation signals associated with virus infection that stimulate cross-priming. We show that infection with lymphocytic choriomeningitis virus induces cross-priming by a mechanism dependent on type I interferon (IFN-α/β). Activation of cross-priming by IFN-α/β was independent of CD4+ T cell help or interaction of CD40 and CD40 ligand, and involved direct stimulation of dendritic cells. These data identify expression of IFN-α/β as a mechanism for the induction of cross-priming during virus infections.

CD8+ T-cell memory to viruses.

Recent experiments show that laboratory mice infected once with an influenza A virus or with the murine parainfluenza type 1 virus, called the Sendai virus, have enhanced numbers of cytotoxic T-lymphocyte precursors ( > 20x background) for life. Neither virus persists at the genome level, and the mice are maintained under conditions where there is no possibility of re-infection. These observations are highly relevant to any understanding of CD8+ cell memory and suggest that the clonal burst size associated with the primary challenge is a key determining factor.

Nasal-Associated Lymphoid Tissue Is a Site of Long-Term Virus-Specific Antibody Production following Respiratory Virus Infection of Mice

ABSTRACT Nasal immunoglobulin A provides an initial defense against inhaled respiratory pathogens. However, it is not known whether the nasal-associated lymphoid tissues (NALT) are able to mount an effective long-lasting pathogen-specific immune response, nor is it known whether functional differences exist between the organized NALT (O-NALT) and the diffuse NALT lining the nasal passages (D-NALT). Here we show that although both the O-NALT and the D-NALT are capable of producing virus-specific antibody in response to influenza virus infection, the frequency of specific antibody-forming cells in the D-NALT is much greater than the frequency observed in the O-NALT. Furthermore, we show that the D-NALT but not the O-NALT is the site of long-term virus-specific humoral immunity which lasts for the life of the animal. These results indicate that the D-NALT is not only the major effector site of the NALT but also the site of local long-term specific antibody production.

Inability To Evoke a Long-Lasting Protective Immune Response to Respiratory Syncytial Virus Infection in Mice Correlates with Ineffective Nasal Antibody Responses

ABSTRACT Long-lasting protective antibody is not normally generated in children following primary respiratory syncytial virus (RSV) infection, frequently leading to reinfection. We used the BALB/c mouse model to examine the role of the nasal-associated lymphoid tissue and the bone marrow in the generation of RSV-specific long-lasting plasma cells, with a view to further understanding the mechanisms responsible for the poorly sustained RSV antibody levels following primary infection. We show here that substantial numbers of RSV-specific plasma cells were generated in the bone marrow following challenge, which were maintained thereafter. In contrast, in the nasal-associated lymphoid tissue, RSV-specific plasma cell numbers waned quickly both after primary infection and after challenge and were not maintained at a higher level after boosting. These data indicate that the inability to generate a robust local mucosal response in the nasal tissues may contribute substantially to the likelihood of subsequent reinfection and that the presence of serum anti-RSV antibody without local protection is not enough to protect against reinfection.

Lymphocytic choriomeningitis virus induces a chronic wasting disease in mice lacking class I major histocompatibility complex glycoproteins

Lymphocytic choriomeningitis virus (LCMV) induces a chronic, wasting syndrome when injected intracerebrally into H-2b mice homozygous for a beta 2-microglobulin (beta 2-m (-/-)) gene disruption. These mice have very few CD8+ T cells and express little class I MHC glycoprotein, though minimal levels of the H-2Db molecule have been detected on in vitro cultured beta 2-m (-/-) cells. The underlying immunopathological process in these beta 2-m (-/-) mice is mediated by virus immune CD4+ effectors. However, adoptively transferred CD8+ T cells from normal, LCMV-infected H-2Db compatible donors induce significant (but low level) meningitis in beta 2-m (-/-) recipients. Such mice develop neither the neurological disease characteristic of LCM nor the persistent, though generally non-fatal, debility that occurs when only the CD4+ T cell subset is involved.

Extent of γδ T cell involvement in the pneumonia caused by sendai virus

Abstract The prevalence of γδ T cells in bronchoalveolar lavage (BAL) populations recovered from the respiratory tract of young, adult C57BL/6J mice infected intranasally (i.n.) with Sendai virus has been assessed by FACS-phenotyping, and by probing cytocentrifuge preparations for expression of TCRγ mRNA. The surface γδ TCR + set comprised from 5 to 20% of the inflammatory lymphocytes in sequential samples taken throughout the course of this nonfatal viral pneumonia. The BAL population also contained numerous cells expressing mRNA for Cγ 1/2 and Cγ4; the C-regions were utilized for productive TCR gene rearrangement. Sorting the lymphocytes from the BAL established that >90% of both the TCRγ and TCRβ mRNA partitioned to cells with the appropriate surface TCR phenotype, while + cells in the total inflammatory exudate were phagocytes that engulfed latex particles. Both the frequency and the total numbers of the γδ TCR + and TCRγ mRNA + cells were increased in mice depleted of αβ T cells by in vivo treatment with mAbs to CD4 and CD8. indicating that the CD4 + and CD8 + αβ and CD4 − 8 − γδ T cell subsets may operate independently in this virus disease. The Cγl/2 mRNA phenotype predominated throughout the course of the active infection, with a transition to maximal prevalence of the Cγ4 mRNA + set occurring very late (Day 20) in the resolving inflammatory process. Large numbers of macrophages expressing mRNA (>50%) for a mammalian 65-kDa heat shock protein (hsp65), a possible target for some of the γδ T cells, were present early (Days 5–7) and remained at lower levels ( + macrophages were much less apparent in BAL populations from mice depleted concurrently of the CD4 + and CD8 + T cell subsets, indicating that exposure to Sendai virus alone is not the major factor inducing the transcription of this endogenous gene. These experiments thus establish that γδ T cells are a minority of the infiltrating lymphocytes in Sendai virus pneumonia and provide new insights into the spectrum of hsp65 mRNA and TCRγ mRNA expression during an inflammatory process.

Intranasal immunisation with inactivated RSV and bacterial adjuvants induces mucosal protection and abrogates eosinophilia upon challenge.

We have previously shown that following intranasal exposure to influenza virus, specific plasma cells are generated in the nasal‐associated lymphoid tissue (NALT) and maintained for the life of the animal. However, we also showed that following infection with respiratory syncytial virus (RSV), specific plasma cells are generated in the NALT but wane quickly and are not maintained even after challenge, even though RSV‐specific serum antibody responses remain robust. Only infection with influenza virus generated sterilising immunity, implying a role for these long‐lived plasma cells in protection. We show here that the RSV‐specific IgA NALT plasma cell population and lung antibody levels can be substantially boosted, both at acute and memory time points, by intranasal immunisation with inactivated RSV (iRSV) in combination with bacterial outer membrane vesicles (OMV) compared to live RSV alone. Finally, challenge with live RSV showed that immunisation with iRSV and OMV protect against both virus replication in the lung and the eosinophil infiltrate generated by either live RSV or iRSV alone. These data show that immunisation with iRSV and OMV maintains a NALT RSV‐specific plasma cell population and generates an efficient protective immune response following RSV infection.

Local and Systemic Antibody Responses in Mice Immunized Intranasally with Native and Detergent-Extracted Outer Membrane Vesicles from Neisseria meningitidis

ABSTRACT The mouse humoral immune response toward native or detergent-extracted outer membrane vesicles (NOMVs and DOMVs, respectively) from Neisseria meningitidis was determined after intranasal immunization. Both preparations elicited high frequencies of NOMV-specific antibody-forming cells (AFCs) locally in the nasal associated lymphoid tissue (NALT) after three or four weekly doses. The diffuse NALT (D-NALT) contained ca. 10-fold more NOMV-specific AFCs than those observed in the mediastinal lymph node, spleen, and bone marrow. AFCs observed in the D-NALT were primarily immunoglobulin A positive (IgA+) and were maintained for at least 1 month. In contrast, the organized NALT (O-NALT) contained low numbers of AFCs, and the response was relatively short-lived. In other lymphoid tissues, AFCs producing various IgG subclasses and IgM were present with IgG2b-producing AFCs being dominant or codominant with IgA or IgG2a. In serum and in all of the tissues examined, with the exception of the NALT, NOMVs clearly induced a stronger antibody response and a broader range of antibody isotypes than DOMVs. The development of NOMV-specific AFCs in spleen and bone marrow after intranasal immunization was slow compared to intravenous immunization but, once established, the intranasally elicited responses increased steadily for at least 75 days. NOMV-specific antibodies induced via several routes of immunization had high bactericidal activities in serum. Our results indicated that intranasally administered OMVs induced strong local and systemic antibody responses in mice that were relatively long-lived.

Infection of mice with respiratory syncytial virus during neonatal life primes for enhanced antibody and T cell responses on secondary challenge

Primary neonatal immune responses to infection or vaccines are weak when compared with those of adults. In addition, memory responses of neonatally primed animals may be absent, weak or T helper type 2 (Th2)‐biased. Respiratory syncytial virus (RSV) is an important pathogen of human infants and infection during the neonatal period has been linked to the development of asthma in later life. Here we report that acute intranasal infection of neonatal mice with RSV induces significant RSV‐specific antibody and CD8 T cell responses. These responses were boosted after RSV rechallenge during adulthood, demonstrating the establishment of memory after neonatal priming. Primary infection during neonatal life was associated, following rechallenge, with limited viral replication in the lung. Recall responses of both spleen and lymph node cells from neonatally primed and adult‐primed mice were associated with interferon‐γ secretion, indicative of a Th1‐type response. However, interleukin (IL)‐4 and IL‐5 secretion were enhanced only in spleen and lymph node cells from neonatally primed mice. Rechallenge of neonatally primed mice was also associated with increased concentrations of chemokines monocyte chemoattractant protein‐1, macrophage inflammatory protein‐1α and regulated upon activation normal T cell expressed and secreted in the lung. These may play a role in the enhanced inflammatory cell recruitment and immunopathology induced following RSV reinfection. Our results demonstrate therefore that immunity to RSV can be established during neonatal life and, importantly, that the quality of the subsequent response is dependent upon the age of first infection.

Nasal mucosal administration of chitin microparticles boosts innate immunity against influenza A virus in the local pulmonary tissue

Influenza virus infection remains a major health concern due to morbidity and mortality associated with epidemics and occasional pandemics. The absence of acquired immunity to antigenically distinct, emerging virus strains stresses the need for a generic drug that protects independent of vaccination. Here, we demonstrate that prophylactic administration of chitin microparticles (CMP) via the intranasal route significantly reduced lung viral titres and clinical signs. Pre-treatment boosted the innate immune response to subsequent infection by recruiting innate cells, such as neutrophils, and increasing inflammatory cytokines. Although an increase in virus-specific T cells was observed, the memory phase was diminished. Our data demonstrate that in the absence of prior exposure to influenza virus, CMP reduce clinical signs by boosting innate immunity.