Pre-Existing Cross-Reactive Antibodies to Avian Influenza H5N1 and 2009 Pandemic H1N1 in US Military Personnel

We studied cross-reactive antibodies against avian influenza H5N1 and 2009 pandemic (p) H1N1 in 200 serum samples from US military personnel collected before the H1N1 pandemic. Assays used to measure antibodies against viral proteins involved in protection included a hemagglutination inhibition (HI) assay and a neuraminidase inhibition (NI) assay. Viral neutralization by antibodies against avian influenza H5N1 and 2009 pH1N1 was assessed by influenza (H5) pseudotyped lentiviral particle-based and H1N1 microneutralization assays. Some US military personnel had cross-neutralizing antibodies against H5N1 (14%) and 2009 pH1N1 (16.5%). The odds of having cross-neutralizing antibodies against 2009 pH1N1 were 4.4 times higher in subjects receiving more than five inactivated whole influenza virus vaccinations than those subjects with no record of vaccination. Although unclear if the result of prior vaccination or disease exposure, these pre-existing antibodies may prevent or reduce disease severity.Outbreaks of 1997 avian influenza H5N1 and 2009 pandemic (p) H1N1 in humans have provided an opportunity to gain insight into cross-reactive immunity. The US military periodically collects and stores serum samples from service members linked to medical records.¹ We measured cross-reactive antibodies in stored serum to avian influenza H5N1 and 2009 pH1N1 from US military personnel and identified factors associated with presence of neutralizing antibodies.Two hundred archived serum samples were obtained from the US Department of Defense Serum Repository. They were representative of a wide cross-section of active military personnel at the times of collection, whereas specific geographic information was not available on the individual selected; the cohort represents the general US military population, which is deployed throughout the United States and globally. Fifty samples each were selected from four birth cohorts: (1) < 1949, (2) 1960–1965, (3) 1966–1971, and (4) 1972–1977. Within each cohort, 25 samples were collected in the year 2000 (before the introduction of intranasal live attenuated influenza vaccine [LAIV]), and 25 samples were collected in 2008 (where 51% of donors had received LAIV). It has been suggested that LAIV elicits cross-reactive immunity.^2,3 The samples were all collected before the outbreak of 2009 pH1N1, and there have not been any reported outbreaks of H5N1 in US military personnel.Assays used to measure antibodies included a hemagglutination inhibition (HI) assay and a neuraminidase inhibition (NI) assay.⁴ Viral neutralization by antibodies against H5N1 and 2009 pH1N1 was assessed by influenza (H5) pseudotyped lentiviral particle-based (H5pp)⁵ and microneutralization assays, respectively. Electronic medical and vaccination records from the Defense Medical Surveillance System (DMSS), which captured records before the serum sample date, were linked to samples and compared with the in vitro results.¹The odds ratios (ORs) and 95% confidence intervals (95% CIs) of univariate and multivariate binary logistic regression analyses were used to determine the association between donor characteristics and positive antibody responses. A multiple logistic regression model was constructed, and it included independent variables with a P value of < 0.05 in univariate logistic regression. A P value of < 0.05 was considered to indicate statistical significance. SPSS 12.0 for Windows (SPSS Inc., Chicago, IL) was used to perform all statistical analysis.Cross-reactivity is summarized in 5 and 22.5% for the NI assay. H5pp and NI antibody titers to H5N1 were evenly distributed among birth cohorts and did not differ substantially based on history of vaccination or prior respiratory infections. Of those individuals with neutralizing antibodies to H5N1 (N = 28), 32.1% also had neutralizing antibodies to pH1N1, whereas 19.3% of those individuals with any H5N1-specific antibody response also had neutralizing antibodies to pH1N1 (

The role of intracellular calcium phosphate in osteoblast-mediated bone apatite formation

Mineralization is a ubiquitous process in the animal kingdom and is fundamental to human development and health. Dysfunctional or aberrant mineralization leads to a variety of medical problems, and so an understanding of these processes is essential to their mitigation. Osteoblasts create the nano-composite structure of bone by secreting a collagenous extracellular matrix (ECM) on which apatite crystals subsequently form. However, despite their requisite function in building bone and decades of observations describing intracellular calcium phosphate, the precise role osteoblasts play in mediating bone apatite formation remains largely unknown. To better understand the relationship between intracellular and extracellular mineralization, we combined a sample-preparation method that simultaneously preserved mineral, ions, and ECM with nano-analytical electron microscopy techniques to examine osteoblasts in an in vitro model of bone formation. We identified calcium phosphate both within osteoblast mitochondrial granules and intracellular vesicles that transported material to the ECM. Moreover, we observed calcium-containing vesicles conjoining mitochondria, which also contained calcium, suggesting a storage and transport mechanism. Our observations further highlight the important relationship between intracellular calcium phosphate in osteoblasts and their role in mineralizing the ECM. These observations may have important implications in deciphering both how normal bone forms and in understanding pathological mineralization.     

Acemannan Stimulates Gingival Fibroblast Proliferation; Expressions of Keratinocyte Growth Factor-1, Vascular Endothelial Growth Factor,and Type I Collagen; and Wound Healing

Aloe vera has long been used as a traditional medicine for inducing wound healing. Gingival fibroblasts (GFs) play an important role in oral wound healing. In this study, we investigated the effects of acemannan, a polysaccharide extracted from Aloe vera gel, on GF proliferation; keratinocyte growth factor-1 (KGF-1), vascular endothelial growth factor (VEGF), and type I collagen production; and oral wound healing in rats. [³H]-Thymidine incorporation assay and ELISA were used. Punch biopsy wounds were created at the hard palate of male Sprague Dawley rats. All treatments (normal saline; 0.1% triamcinolone acetonide; plain 1% Carbopol^®; and Carbopol^® containing 0.5%, 1%, and 2% acemannan (w/w)) were applied daily. Wounded areas and histological features were observed at day 7 after treatment. From our studies, acemannan at concentrations of 2, 4, 8, and 16 mg/ml significantly induced cell proliferation (P<0.05). Acemannan concentrations between 2 – 16 mg/ml significantly stimulated KGF-1, VEGF, and type I collagen expressions (P<0.05). Wound healing of animals receiving Carbopol^® containing 0.5% acemannan (w/w) was significantly better than that of the other groups (P<0.05). These findings suggest that acemannan plays a significant role in the oral wound healing process via the induction of fibroblast proliferation and stimulation of KGF-1, VEGF, and type I collagen expressions.     

Apoptosis and pathogenesis of avian influenza A (H5N1) virus in humans

The pathogenesis of avian influenza A (H5N1) virus in humans has not been clearly elucidated. Apoptosis may also play an important role. We studied autopsy specimens from 2 patients who died of infection with this virus. Apoptosis was observed in alveolar epithelial cells, which is the major target cell type for the viral replication. Numerous apoptotic leukocytes were observed in the lung of a patient who died on day 6 of illness. Our data suggest that apoptosis may play a major role in the pathogenesis of influenza (H5N1) virus in humans by destroying alveolar epithelial cells. This pathogenesis causes pneumonia and destroys leukocytes, leading to leukopenia, which is a prominent clinical feature of influenza (H5N1) virus in humans. Whether observed apoptotic cells were a direct result of the viral replication or a consequence of an overactivation of the immune system requires further studies.     

Prevalence of human cytomegalovirus (HCMV) gB genotypes in Thai patients

Human cytomegalovirus (HCMV) infection can cause asymptomatic to symptomic diseases leading to morbidity and mortality especially in immunocompromized patients. One factor of the difference in clinical outcome is the distinction of HCMV strain. As HCMV glycoprotein (g)B plays an important role in viral entry and neutralizing antibody induction, HCMV gB genotypes were determined in 161 clinical specimens containing HCMV-DNA obtained from patients at King Chulalongkorn Memorial Hospital, Bangkok, Thailand during the year 2000 and 2004. Of the 113 (70%) samples that were able to be genotyped, mixed gB genotype was demonstrated in 35%, followed by gB1 (33%), gB3 (15%), gB2 (11%), and untyped (7%); gB4 was not detected. The distribution of HCMV gB genotypes between genders was not significantly different. Mixed gB genotype (35%) was found in HIV- infected patients.