Protection against influenza is mediated by neutralizing antibodies, and their induction at high and sustained titers is key for successful vaccination. Optimal B cells activation requires delivery of help from CD4 + T lymphocytes. In lymph nodes and tonsils, T-follicular helper cells have been identified as the T cells subset specialized in helping B lymphocytes, with interleukin-21 (IL-21) and inducible costimulatory molecule (ICOS1) playing a central role for this function. We followed the expansion of antigen-specific IL-21 + CD4 + T cells upon influenza vaccination in adults. We show that, after an overnight in vitro stimulation, influenza-specific IL-21 + CD4 + T cells can be measured in human blood, accumulate in the CXCR5 − ICOS1 + population, and increase in frequency after vaccination. The expansion of influenza-specific ICOS1 + IL-21 + CD4 + T cells associates with and predicts the rise of functionally active antibodies to avian H5N1. We also show that blood-derived CXCR5 − ICOS1 + CD4 + T cells exert helper function in vitro and support the differentiation of influenza specific B cells in an ICOS1-and IL-21-dependent manner. We propose that the expansion of antigen-specific ICOS1 + IL-21 + CD4 + T cells in blood is an early marker of vaccine immunogenicity and an important immune parameter for the evaluation of novel vaccination strategies.CD4 help | predictivity | humoral response T o confer protection, human vaccines rely on the induction of neutralizing antibodies and on the generation of a pool of memory lymphocytes able to mount an accelerated response upon encounter with the target pathogen. In recent years, novel vaccines, adjuvants, and delivery systems that are able to improve vaccine immunogenicity while reducing their reactogenicity have been developed. As vaccines are given to healthy subjects, their development is a challenging endeavor that requires extensive studies to assess safety, immunogenicity, and clinical efficacy. To accelerate the screening of novel candidates, research has focused on the identification of early biomarkers, molecular and transcriptional signatures predicting vaccine efficacy (1). Predictors should be easy to test in large clinical trials and have a clear mechanistic relationship with the correlates or surrogates of protection taken as the study endpoint. We have previously shown that an early postvaccination increase in the number of vaccine-specific CD4 + T cells is correlated in a predictive manner with the rise and long-term maintenance of protective antibody titers to avian influenza (2). The aim of the present study was to characterize the CD4 + T cells subset responsible for this function.T follicular helper (Tfh) cells have been identified in lymph nodes and tonsils as the CD4 + T cells subpopulation specialized in providing help to B cells (3-11). The recent identification of a circulating counterpart of this T cells subset in blood led us to investigate whether vaccine-specific IL-21 + CD4 + T cells are detectable in human blood, if their frequency is...

؉ cell count were observed at any time point. The results showed good safety and immunogenicity for both vaccines under study for both uninfected and HIV-1-infected adults, confirming current recommendations for immunization of this high-risk category.

Influenza is a major cause of worldwide morbidity and mortality through frequent seasonal epidemics and infrequent pandemics. Morbidity and mortality rates from seasonal influenza are highest in the most frail, such as the elderly, those with underlying chronic conditions and very young children. Antigenic mismatch between strains recommended for vaccine formulation and circulating viruses can further reduce vaccine efficacy in these populations. Seasonal influenza vaccines with enhanced, cross‐reactive immunogenicity are needed to address these problems and can confer a better immune protection, particularly in seasons were antigenic mismatch occurs. A related issue for vaccine development is the growing threat of pandemic influenza caused by H5N1 avian strains. Vaccines against strains with pandemic potential offer the best approach for reducing the potential impact of a pandemic. However, current non‐adjuvanted pre‐pandemic vaccines offer suboptimal immunogenicity against H5N1. For both seasonal and pre‐pandemic vaccines, the addition of adjuvants may be the best approach for providing enhanced cross‐reactive immunogenicity. MF59®, the first oil‐in‐water emulsion licensed as an adjuvant for human use, can enhance vaccine immune responses through multiple mechanisms. A trivalent MF59‐adjuvanted seasonal influenza vaccine (Fluad®) has shown to induce significantly higher immune responses to influenza vaccination in the elderly, compared with non‐adjuvanted vaccines, and to provide cross‐reactive immunity against divergent influenza strains. Similar results have been generated with a MF59‐adjuvanted H5N1 pre‐pandemic vaccine, which showed higher and broader immunogenicity compared with non‐adjuvanted pre‐pandemic vaccines.

Squalene is a naturally occurring oil which has been used in the development of vaccine adjuvants, such as the oil-in-water emulsion MF59. In past years, by use of noncontrolled and nonvalidated assays, a claim was made that antisqualene antibodies were detectable in the sera of individuals with the so-called Gulf War syndrome. Using a validated enzyme-linked immunosorbent assay for the quantitation of immunoglobulin G (IgG) and IgM antibodies against squalene, we demonstrated that antisqualene antibodies are frequently detectable at very low titers in the sera of subjects who were never immunized with vaccines containing squalene. More importantly, vaccination with a subunit influenza vaccine with the MF59 adjuvant neither induced antisqualene antibodies nor enhanced preexisting antisqualene antibody titers. In conclusion, antisqualene antibodies are not increased by immunization with vaccines with the MF59 adjuvant. These data extend the safety profile of the MF59 emulsion adjuvant.Squalene is a triterpenoid hydrocarbon oil (C 30 H 50 ) produced by plants and is present in many foods. Squalene is also produced abundantly by human beings, for whom it serves as a precursor of cholesterol and steroid hormones (8) It is synthesized in the liver and the skin, transported in the blood by very-low-density lipoproteins (VLDL) and low-density lipoproteins (LDL), and secreted in large amounts by sebaceous glands (10,17).Since it is a natural component of the human body and is biodegradable, squalene has been used as a component of vaccine adjuvants. One of these adjuvants is MF59, an oil-inwater emulsion developed by Chiron (14). MF59 has been shown in various preclinical and clinical studies to significantly enhance the immune response to a wide variety of vaccine antigens (15). MF59 is a part of an influenza subunit vaccine which has been licensed in various European countries since 1997. More than 20 million doses of this vaccine have been given, and it has been shown to have an excellent safety profile. The safety of vaccines with the MF59 adjuvant has also been shown by various investigational clinical studies using recombinant antigens from hepatitis B virus, hepatitis C virus, cytomegalovirus, herpes simplex virus, human immunodeficiency virus, uropathogenic Escherichia coli, etc., with various age groups, including 1-to 3-day-old newborns (16).In 2000, antisqualene antibodies were reported to be present in the sera of veterans returning from the first Persian Gulf War with a series of symptoms diagnosed by the authors of the report as representing the so-called Gulf War syndrome (4). The conclusions of this work, based on Western blot assays, were severely criticized on technical grounds (1) and were considered inconclusive by the Institute of Medicine (7).Despite the fact that vaccines given to veterans returning with Gulf War syndrome did not contain squalene (6) and despite that fact that symptoms similar to those of the so-called Gulf War syndrome have been reported after several wars, including the American...