Impact of erythrocyte species on assays for influenza serology.

The influenza viruses have the ability to agglutinate erythrocytes by binding to sialic acid receptors on the host cell. Human influenza viruses preferentially bind to sialic acid linked to galactose by α 2.6 linkage, while avian influenza viruses preferentially bind to sialic acid linked to Gal by α 2.3 linkage. There is a close correlation between the ability of influenza A viruses to agglutinate erythrocytes from different animal species and their receptor specificity. The haemagglutination and haemagglutination inhibition assays are influenced by the species of erythrocytes. To provide an overview of the expression of sialic acid receptors on different erythrocytes, avian (turkey, chicken, pigeon) and mammalian (sheep, horse, human) species have been analysed by flow cytometry. Chicken, turkey and human erythrocytes display both types of linkages. Horse and sheep erythrocytes show almost exclusively α 2.3 Gal linkages, while pigeon erythrocytes express almost exclusively α 2.6 Gal linkages. The erythrocytes from the same avian and mammalian species have been evaluated by haemagglutination and haemagglutination inhibition assays with seasonal and avian strains. Chicken and turkey erythrocytes seem to be the most appropriate for both assays with seasonal influenza strains, in addition to pigeon erythrocytes, particularly for the B strains. In the case of the avian strain, chicken erythrocytes are suitable for haemagglutination assay and horse erythrocytes for haemagglutination inhibition assay. The choice of erythrocytes has a significant impact on the titres measured by both assays.

Safety and immunogenicity of a recombinant Plasmodium falciparum AMA1-DiCo malaria vaccine adjuvanted with GLA-SE or Alhydrogel® in European and African adults: A phase 1a/1b, randomized, double-blind multi-centre trial.

BACKGROUND: Plasmodium falciparum Apical Membrane Antigen 1 Diversity Covering (PfAMA1-DiCo) candidate vaccine is a formulation of three recombinant variants of AMA1 designed to provide broader protection against parasites with varying AMA1 sequences. METHODS: In this staggered phase Ia/Ib randomized, double blind trial, healthy French adults received AMA1-DiCo with either Alhydrogel® (n=15) or GLA-SE (n=15). Following a safety assessment in French volunteers, GLA-SE was chosen for the phase Ib trial where healthy Burkinabe adults received either AMA1-DiCo/GLA-SE (n=18) or placebo (n=18). AMA1-DiCo (50µg) was administered intramuscularly at baseline, Week 4 and 26. RESULTS: AMAI-DiCo was safe, well tolerated either with Alhydrogel® or GLA-SE. In European volunteers, the ratios of IgG increase from baseline were about 100 fold in Alhydrogel® group and 200-300 fold in GLA-SE group for the three antigens. In African volunteers, immunization resulted in IgG levels exceeding those observed for the European volunteers with a 4-fold increase. DiCo-specific IgG remained higher 26weeks after the third immunization than at baseline in both European and African volunteers. Induced antibodies were reactive against whole parasite derived from different strains. CONCLUSION: AMA1-DiCo vaccine was safe and immunogenic whatever the adjuvant although GLA-SE appeared more potent than Alhydrogel® at inducing IgG responses. CLINICAL TRIALS REGISTRATION: ClinicalTrials.gov NCT02014727; PACTR201402000719423.

New candidate vaccines against blood-stage Plasmodium falciparum malaria: prime-boost immunization regimens incorporating human and simian adenoviral vectors and poxviral vectors expressing an optimized antigen based on merozoite surface protein 1.

Although merozoite surface protein 1 (MSP-1) is a leading candidate vaccine antigen for blood-stage malaria, its efficacy in clinical trials has been limited in part by antigenic polymorphism and potentially by the inability of protein-in-adjuvant vaccines to induce strong cellular immunity. Here we report the design of novel vectored Plasmodium falciparum vaccines capable of overcoming such limitations. We optimized an antigenic insert comprising the four conserved blocks of MSP-1 fused to tandemly arranged sequences that represent both allelic forms of the dimorphic 42-kDa C-terminal region. Inserts were expressed by adenoviral and poxviral vectors and employed in heterologous prime-boost regimens. Simian adenoviral vectors were used in an effort to circumvent preexisting immunity to human adenoviruses. In preclinical studies these vaccines induced potent cellular immune responses and high-titer antibodies directed against MSP-1. The antibodies induced were found to have growth-inhibitory activity against dimorphic allelic families of P. falciparum. These vectored vaccines should allow assessment in humans of the safety and efficacy of inducing strong cellular as well as cross-strain humoral immunity to P. falciparum MSP-1.

Heritability estimates of innate immunity: an extended twin study.

Cytokines are key players in numerous inflammatory processes. Demonstration of a heritable component in the variation of cytokine production would indicate that simultaneous occurrence of conditions might be caused by a heritable inflammatory characteristic. We applied an extended twin study approach to assess heritability estimates of interleukin (IL)-1beta, IL-1ra, IL-10, IL-6, and TNF-alpha production capacity after ex vivo stimulation with lipopolysaccharide. Cytokine production capacity was assessed in 42 monozygotic pairs, 52 dizygotic pairs, one trizygotic triplet, 33 single twins, and 83 additional siblings. Heritability estimates were derived from variance decomposition models using maximum likelihood estimation. For all cytokines, over 50% of the variance was genetically determined. IL-1ra and TNF-alpha had the lowest heritability estimate of 53%. Estimates for IL-6 and IL-10 were 57 and 62%, respectively. IL-1beta had the highest estimate of 86%. We conclude that the production of cytokines is under tight genetic control.

Interaction of atherosclerosis and inflammation in elderly subjects with poor cognitive function.

OBJECTIVE: To test the hypothesis that a pro-inflammatory response is associated with cognitive impairment among individuals with cardiovascular disease. METHOD: All 85-year-old inhabitants of Leiden (n = 599) were visited at their place of residence. A history of cardiovascular disease and an EKG were used as indicators of atherosclerosis. Production of the pro-inflammatory cytokine tumor necrosis factor-alpha and the anti-inflammatory cytokine interleukin-10 was assessed in a whole-blood assay using lipopolysaccharide as a stimulus. Global cognitive functioning was determined with the Mini-Mental State Examination (MMSE); attention, cognitive speed, and memory were determined with four neuropsychological tests; and a history of dementia was obtained. RESULTS: In subjects with cardiovascular disease, median MMSE scores were lower in those with a pro-inflammatory response when compared with those with an anti-inflammatory response (p = 0.02). Similar associations were found for the Stroop Test, measuring attention (p < 0.01), the Coding Test measuring cognitive speed (p = 0.02), the Word Learning Test measuring memory (p < 0.01), and the presence of dementia (p = 0.04). The associations remained unaltered after adjustments for possible confounders such as gender, level of education, use of nonsteroidal anti-inflammatory drugs, use of cardiovascular drugs, and cardiovascular risk factors. In contrast, outcomes of the cognitive tests and presence of dementia were not dependent on the inflammatory response when cardiovascular disease was absent. CONCLUSION: The combination of cardiovascular disease and a pro-inflammatory cytokine response may be associated with cognitive impairment and dementia.

Innate cytokine profile in patients with complex regional pain syndrome is normal.

Complex regional pain syndrome (CRPS) is a disabling disease characterized by the classic symptoms and signs of inflammation. In this study we investigated the innate cytokine profile in patients with CRPS to determine a possible role of the immune system in the pathophysiology of CRPS. The cytokine profile before and after lipopolysaccharide and thrombin stimulation was determined in 26 severely affected CRPS patients and 20 healthy controls. No difference in the production of pro- and anti- inflammatory cytokines between patients and controls was found. Hence, our results do not support a role of genetic factors responsible for the cytokine profile in the pathophysiology of CRPS. These findings encourage further investigations of mechanisms responsible for neurogenic-induced inflammation.

Randomized controlled trial of seroresponses to double dose and booster influenza vaccination in frail elderly subjects.

Responses to influenza vaccination are poor in frail elderly subjects who suffer the greatest morbidity and mortality due to infection. Therefore, a randomized clinical trial was performed to determine the effect of a double dose and booster vaccination on antibody responses after influenza vaccination. A total of 815 patients (median age 83 years, median disability score 8, median disease categories 2 and median number of medications 4) residing in 14 nursing homes in the Netherlands were vaccinated during the influenza season 1997-98. The first vaccine dose (15 or 30 microg) was given on Day 0 followed by a booster dose (placebo or 15 microg) on Day 84. Blood samples were taken before and 25 days after vaccination. There were four treatment groups: (i) 15 microg and placebo, (ii) 15 microg and 15 microg booster, (iii) 30 microg and placebo and (iv) 30 microg and 15 microg booster. Geometric mean antibody titers of those receiving the double vaccine dose was 15% (95% CI, 6% to 24%, P = 0.001) higher as compared to the standard 15 microg dose. A booster dose, given 84 days after the first vaccination, yielded postvaccination titters that were 14% (95% CI, 9% to 19%, P = 0.001) higher as compared to placebo. Subgroup analysis did not reveal patient groups that had a proportionally greater benefit from adapted vaccination strategies. It is concluded that higher antibody responses can be achieved in frail elderly people by a double vaccine dose or a booster vaccination.

Patients with Alzheimer's disease display a pro-inflammatory phenotype.

BACKGROUND: Inflammation plays a pivotal role in amyloid plaque progression thereby contributing to Alzheimer's disease-related neurodegeneration. We hypothesized that patients with Alzheimer's disease have an innate pro-inflammatory phenotype, as compared to control subjects without dementia. METHODS: Patients with a diagnosis of probable Alzheimer's disease (n=12) and control subjects without signs of dementia (n=18) were enrolled. Whole blood samples were stimulated ex vivo with endotoxin under standard conditions. Cytokine levels were assessed by ELISA and compared by Mann-Whitneyll-test after log transformation. RESULTS: Patients with Alzheimer's disease had seven- to ten-fold higher IL-1beta production relative to the amount of IL-10 both at the low (p=0.006) and high concentration of endotoxin (p=0.007). Subjects who display a pro-inflammatory phenotype as defined by a high IL-1beta/IL-10 ratio had 13.0-fold higher odds (95% CI: 2.1-82) to have dementia. CONCLUSION: The data support the hypothesis that a pro-inflammatory phenotype contributes to the development of Alzheimer's disease.

Interaction of indomethacin with cytokine production in whole blood. Potential mechanism for a brain-protective effect.

Both Alzheimer's disease and vascular dementia are featured by inflammatory responses and it is known that non-steroidal anti-inflammatory drugs (NSAIDs) decrease the risk and severity of these diseases. To study the effect of NSAIDs on PGE2 levels and pro- and anti-inflammatory cytokine levels in the whole blood assay, blood samples from 23 elderly persons aged 85 years were stimulated with thrombin or LPS as primary stimulus. Indomethacin was added in concentrations ranging from 0.4 to 16 microg/ml and acetylsalicylic acid was added to in concentrations ranging from 0.5 to 8.0 microg/ml. Indomethacin abrogated thrombin- and LPS-induced PGE2 production at all concentrations tested. In addition, indomethacin reduced the production of thrombin-induced IL-6 and IL-10 (p<0.05) at physiological concentrations. Indomethacin reduced the production of LPS-induced IL-6, IL-1 beta and IL-10 (p<0.05) at the highest indomethacin concentration tested. Similar results were obtained upon incubation with acetylsalicylic acid. It is concluded that indomethacin may reduce the thrombin-induced inflammatory reaction by decreasing IL-6 through inhibition of PGE2 synthesis. This IL-6 reduction may be relevant for the ability of indomethacin to reduce the risk of Alzheimer's disease. However, the decrease in IL-10 production due to indomethacin suggests a more inflammatory state.

Idiopathic senile gait disorders are signs of subclinical disease.

OBJECTIVES: To evaluate survival and causes of death in subjects with idiopathic senile gait disorders. DESIGN: A population-based longitudinal study. SETTING: Survival analysis of the oldest old within the Leiden 85-plus Study. PARTICIPANTS: We distinguished three different groups according to their gait: subjects with a normal gait (n = 25), subjects with senile gait disorders (n = 14), and subjects with gait disorders due to known disease (n = 87). The mean age was 90 years in all groups (range 87 to 97 years). MEASUREMENTS: The risk of all cause mortality and cardiovascular mortality was estimated over 5 years of follow-up in a Cox-proportional hazards model, adjusted for age and sex. RESULTS: Eighty-nine of 126 subjects died during follow-up. Mean survival differed among the three groups (P log-rank = .01). All cause mortality risk was increased in subjects with senile gait disorders compared with subjects with a normal gait (RR = 2.8; 95% CI, 1.1-7.3, P = .03) and was similar to subjects with gait disorders caused by known disease (RR = 1.2; 95% CI: .6-2.5, P = .6). Mortality caused by cardiovascular disease also differed among the three groups (P log-rank = .03). The risk of cardiovascular death in subjects with senile gait disorders was twofold greater than in subjects with a normal gait (RR = 2.1; 95% CI, 0.4-10.3). CONCLUSIONS: Senile gait disorders are related to subclinical, perhaps cardiovascular, disease. Senile gait disorders should not be accepted as an inevitable, benign concomitant of the normal aging process.

The effect of geographic origin on the frequency of HLA antigens and their association with ageing.

The association between HLA antigens and ageing is not clear. Ageing in women was associated with B40 and DR5 in a recent study, but other studies yielded conflicting results. In none of the studies, however, did the young and elderly samples originate from the same homogeneous population. Homogeneity is dependent on geographic origin. The aim of this study was to investigate whether differences in geographic origin between age groups could explain the age-associated differences in the frequencies of B40 and DR5. The authors used the new design of a 'birth-place-restricted comparison' in which the origin of all subjects was ascertained. The total study population comprised 1010 young women aged 25-40 years and 660 elderly women aged 85 years and older. The 'birth-place-restricted comparison' included 66 young and 285 elderly women from one geographic area (Leiden, the Netherlands). Men were not included because ageing in men was not associated with HLA antigens in a recent study. In the total population, the frequency of B40 in young women of different origin varied between 16 and 28%, and the frequency of DR5 between 11 and 23%. Similar differences were observed in the elderly women. In the 'birth-place-restricted comparison', the frequency of B40 was 15% in the young women and 11% in the elderly women (difference 4%, 95% confidence interval, -5 to 13%). The frequency of DR5 was 20% in the young women, and 28% in the elderly women (difference 8%, 95% confidence interval, -4 to 19%). Thus, marked differences in HLA antigen frequency were found between populations of various geographic origins. Definition and ascertainment of the target population are therefore necessary in genetic studies of ageing. In such a 'birth-place-restricted comparison', the authors confirmed that ageing in women was negatively associated with HLA-B40 and positively associated with HLA-DR5.

Reduced IFN-gamma production in elderly people following in vitro stimulation with influenza vaccine and endotoxin.

Cytokine interferon gamma (IFN-gamma) is pivotal in the defence against viruses and intracellular pathogens and an age-related decreased IFN-gamma production may explain the increased infectious disease morbidity and mortality in the elderly. Therefore, we performed a series of clinical experiments evaluating the influence of age and health status on IFN-gamma production following in vitro stimulation with influenza vaccine or endotoxin. Both healthy and frail elderly people produced significantly lower amounts of IFN-gamma following ex vivo stimulation with influenza vaccine or endotoxin. We conclude that ageing is accompanied by a decreased capacity to produce IFN-gamma. This may explain the increased incidence and case-fatality caused by viruses and intracellular pathogens in the elderly.

Influenza vaccination in elderly people.

Influenza is an important cause of morbidity and mortality in the elderly. Influenza vaccine has been shown to successfully reduce influenza- and pneumonia-associated hospitalizations and deaths, but the antibody induction by influenza vaccines is not always optimal in the elderly. The lower serological efficacy of influenza vaccines that is often observed in the elderly may be due to a multitude of factors. Here we will discuss some of these factors. These include health status and previous exposures to influenza viruses. In addition, we will discuss possibilities to improve antibody responses to influenza vaccination.

Dose-dependent antibody response to influenza H1N1 vaccine component in elderly nursing home patients.

The effects of an increased antigen dose on HI, IgG, IgA, and IgM antibody responses to influenza A/Taiwan/1/86 (H1N1) were investigated in 92 elderly nursing-home residents (mean age 81 years) and 104 young subjects (mean age 20 years). At a standard 10-microg dose, HI and IgG titer rises were lower in the elderly. HI titers did not improve at higher vaccine dosages. By contrast, influenza-specific IgG and IgA antibody responses were dose dependent in elderly subjects, but not in young. In the young subjects, IgM antibody responses were dose dependent. The improved antibody responses in the elderly as observed in IgG and IgA were not reflected in the HI response. Therefore, the evaluation of antibody production by HI only may lead to an underestimate of the immune response in elderly people.

T cells and aging (update february 1999).

Deterioration of the immune system with aging ("immunosenescence") is believed to contribute to morbidity and mortality in man due to the greater incidence of infection, as well as possibly autoimmune phenomena and cancer in the aged. Dysregulation of T cell function is thought to play a critical part in these processes. Factors contributing to T cell immunosenescence may include a) stem cell defects, b) thymus involution, c) defects in antigen presenting cells (APC), d) aging of resting immune cells, e) disrupted activation pathways in immune cells, f) replicative senescence of clonally expanding cells. This review aims to consider the current state of knowledge on the scientific basis for and potential clinical relevance of those factors in immunosenescence.

Quality and quantity of the humoral immune response in healthy elderly and young subjects after annually repeated influenza vaccination.

Doubt about the serologic efficacy of annually repeated influenza vaccination prompted investigations into the course of hemagglutination-inhibiting (HI), IgG, and IgA antibody titers and the IgG and IgA avidity index to influenza A/Taiwan/1/86 and A/Beijing/353/89 after annual vaccination. Fifty-four healthy elderly persons >70 years of age and 24 healthy young adults <30 years of age received standard influenza vaccine during 3 consecutive years. On average, prevaccination HI, IgG, and IgA titers to both influenza virus strains increased >=4 fold between the first and the third vaccination (analysis of variance, P<.001). The postvaccination HI and IgG titers remained unchanged after annual vaccination. The avidity index of IgG and IgA antibodies increased somewhat after annual vaccination, although the increase was statistically significant only in the young subjects. These data indicate that annual influenza vaccination of healthy elderly and young subjects results in an overall increase in protective antibodies.

Impact of aging on innate immunity.

Immune responses in higher organisms are triggered by the recognition of a limited diversity of microbiological products by cells of the innate or "natural" immune system. As a result, in addition to the direct protective effect of natural immunity, antigen-presenting cells, particularly dendritic cells, are activated to process and present an enormous number of peptide antigens to the T lymphocytes of the adaptive immune system. These, together with the B lymphocytes, then mediate specific immune responses and maintain acquired immunological memory. The aging immune system is less well able to cope with infectious disease than the youthful immune system; this review will briefly consider what is known of the age-associated alterations in innate immunity, and how these may also impact on adaptive immunity.

Altered antibody response to influenza H1N1 vaccine in healthy elderly people as determined by HI, ELISA, and neutralization assay.

To determine the influence of ageing per se as well as of priming histories on the antibody response to influenza vaccination, haemagglutination inhibition (HI), ELISA IgG, IgA, IgM and neutralizing antibody titres were studied in 43 healthy young subjects (mean age 23 years) and 55 healthy elderly people (mean age 79 years). The HI and ELISA lgG responses to the A/Guizhou/54/89 strain (H3N2) for which both the young and the elderly had similar priming histories were equal. By contrast, the HI and IgG responses to A/Taiwan/1/86 (H1N1), where the priming histories were different, were lower in the elderly (P < 0.05). Influenza-specific IgA responses in the elderly tended to be higher for all vaccine strains. Influenza-specific postvaccination IgM titres were similar or tended to be higher in the elderly. A subgroup of elderly subjects (18%) who did not express HI activity to the A/Taiwan/1/86 (H1N1) vaccine strain, reacted in the HI assay with the closely related A/Singapore/6/86 (H1N1) strain. These elderly people, however, produced lgG antibodies which neutralized A/Taiwan/1/86 virus in vitro. It is concluded that the elderly are capable of mounting antibody responses similar to those observed in the young. Moreover, the observed age-related differences in antibody responses to H1N1 strains are probably not due to ageing of the immune system itself, but are determined by differences in priming histories.