Immunization against SIVmne in macaques using multigenic DNA vaccines

All structural and regulatory genes of SIVmne were cloned into mammalian expression vectors to optimize expression in vitro and immunogenicity in mice. Macaca fascicularis were immunized four times with plasmid DNA (n = 4), or two DNA priming inoculations followed by two boosts of recombinant gp160 plus Gag-Pol particles (n = 4). Following intrarectal challenge with SIVmne, all macaques became infected. Three monkeys immunized with DNA alone maintained low plasma virus loads by 1 year post-challenge; the fourth exhibited high virus loads and significant CD4+ cell decline. Two of the DNA plus boost and three control macaques had high virus loads and associated CD4+ cell decline. Both vaccine protocols elicited antibodies and comparable helper T-cell proliferative responses to gp160. Cytokine mRNA levels in activated peripheral blood mononuclear cells (PBMC) taken at time of challenge suggested a dominant T helper (Th) 1 state in three DNA-immunized and one protein-boosted macaque, which correlated with low virus loads and high CD4+ cell counts post-challenge.     

Islet Autoimmunity Identifies a Unique Pattern of Impaired Pancreatic Beta-Cell Function,Markedly Reduced Pancreatic Beta Cell Mass and Insulin Resistance in Clinically Diagnosed Type 2 Diabetes

There is a paucity of literature describing metabolic and histological data in adult-onset autoimmune diabetes. This subgroup of diabetes mellitus affects at least 5% of clinically diagnosed type 2 diabetic patients (T2DM) and it is termed Latent Autoimmune Diabetes in Adults (LADA). We evaluated indexes of insulin secretion, metabolic assessment, and pancreatic pathology in clinically diagnosed T2DM patients with and without the presence of humoral islet autoimmunity (Ab). A total of 18 patients with at least 5-year duration of clinically diagnosed T2DM were evaluated in this study. In those subjects we assessed acute insulin responses to arginine, a glucose clamp study, whole-body fat mass and fat-free mass. We have also analyzed the pancreatic pathology of 15 T2DM and 43 control cadaveric donors, using pancreatic tissue obtained from all the T2DM organ donors available from the nPOD network through December 31, 2013. The presence of islet Ab correlated with severely impaired β-cell function as demonstrated by remarkably low acute insulin response to arginine (AIR) when compared to that of the Ab negative group. Glucose clamp studies indicated that both Ab positive and Ab negative patients exhibited peripheral insulin resistance in a similar fashion. Pathology data from T2DM donors with Ab or the autoimmune diabetes associated DR3/DR4 allelic class II combination showed reduction in beta cell mass as well as presence of autoimmune-associated pattern A pathology in subjects with either islet autoantibodies or the DR3/DR4 genotype. In conclusion, we provide compelling evidence indicating that islet Ab positive long-term T2DM patients exhibit profound impairment of insulin secretion as well as reduced beta cell mass seemingly determined by an immune-mediated injury of pancreatic β-cells. Deciphering the mechanisms underlying beta cell destruction in this subset of diabetic patients may lead to the development of novel immunologic therapies aimed at halting the disease progression in its early stage.     

Deciphering microbial landscapes of fish eggs to mitigate emerging diseases

Animals and plants are increasingly suffering from diseases caused by fungi and oomycetes. These emerging pathogens are now recognized as a global threat to biodiversity and food security. Among oomycetes, Saprolegnia species cause significant declines in fish and amphibian populations. Fish eggs have an immature adaptive immune system and depend on nonspecific innate defences to ward off pathogens. Here, meta-taxonomic analyses revealed that Atlantic salmon eggs are home to diverse fungal, oomycete and bacterial communities. Although virulent Saprolegnia isolates were found in all salmon egg samples, a low incidence of Saprolegniosis was strongly correlated with a high richness and abundance of specific commensal Actinobacteria, with the genus Frondihabitans (Microbacteriaceae) effectively inhibiting attachment of Saprolegniato salmon eggs. These results highlight that fundamental insights into microbial landscapes of fish eggs may provide new sustainable means to mitigate emerging diseases.     

Protection of Macaca nemestrina from disease following pathogenic simian immunodeficiency virus (SIV) challenge: utilization of SIV nucleocapsid mutant DNA vaccines with and without an SIV protein boost

Molecular clones were constructed that express nucleocapsid (NC) deletion mutant simian immunodeficiency viruses (SIVs) that are replication defective but capable of completing virtually all of the steps of a single viral infection cycle. These steps include production of particles that are viral RNA deficient yet contain a full complement of processed viral proteins. The mutant particles are ultrastructurally indistinguishable from wild-type virus. Similar to a live attenuated vaccine, this approach should allow immunological presentation of a full range of viral epitopes, without the safety risks of replicating virus. A total of 11 Macaca nemestrina macaques were inoculated with NC mutant SIV expressing DNA, intramuscularly (i.m.) in one study and i.m. and subcutaneously in another study. Six control animals received vector DNA lacking SIV sequences. Only modest and inconsistent humoral responses and no cellular immune responses were observed prior to challenge. Following intravenous challenge with 20 animal infectious doses of the pathogenic SIV(Mne) in a long-term study, all control animals became infected and three of four animals developed progressive SIV disease leading to death. All 11 NC mutant SIV DNA-immunized animals became infected following challenge but typically showed decreased initial peak plasma SIV RNA levels compared to those of control animals (P = 0.0007). In the long-term study, most of the immunized animals had low or undetectable postacute levels of plasma SIV RNA, and no CD4(+) T-cell depletion or clinical evidence of progressive disease, over more than 2 years of observation. Although a subset of immunized and control animals were boosted with SIV(Mne) proteins, no apparent protective benefit was observed. Immunization of macaques with DNA that codes for replication-defective but structurally complete virions appears to protect from or at least delay the onset of AIDS after infection with a pathogenic immunodeficiency virus. With further optimization, this may be a promising approach for vaccine development.     

Genetic risk factors for BMI and obesity in an ethnically diverse population: Results from the population architecture using genomics and epidemiology (PAGE) study

Objective:

Several genome–wide association studies (GWAS) have demonstrated that common genetic variants contribute to obesity. However, studies of this complex trait have focused on ancestrally European populations, despite the high prevalence of obesity in some minority groups.

Design and Methods:

As part of the “Population Architecture using Genomics and Epidemiology (PAGE)” Consortium, we investigated the association between 13 GWAS‐identified single‐nucleotide polymorphisms (SNPs) and BMI and obesity in 69,775 subjects, including 6,149 American Indians, 15,415 African‐Americans, 2,438 East Asians, 7,346 Hispanics, 604 Pacific Islanders, and 37,823 European Americans. For the BMI‐increasing allele of each SNP, we calculated β coefficients using linear regression (for BMI) and risk estimates using logistic regression (for obesity defined as BMI ≥ 30) followed by fixed‐effects meta‐analysis to combine results across PAGE sites. Analyses stratified by racial/ethnic group assumed an additive genetic model and were adjusted for age, sex, and current smoking. We defined “replicating SNPs” (in European Americans) and “generalizing SNPs” (in other racial/ethnic groups) as those associated with an allele frequency‐specific increase in BMI.

Results:

By this definition, we replicated 9/13 SNP associations (5 out of 8 loci) in European Americans. We also generalized 8/13 SNP associations (5/8 loci) in East Asians, 7/13 (5/8 loci) in African Americans, 6/13 (4/8 loci) in Hispanics, 5/8 in Pacific Islanders (5/8 loci), and 5/9 (4/8 loci) in American Indians.

Conclusion:

Linkage disequilibrium patterns suggest that tagSNPs selected for European Americans may not adequately tag causal variants in other ancestry groups. Accordingly, fine‐mapping in large samples is needed to comprehensively explore these loci in diverse populations.     

Emerging cytopathic and antigenic simian immunodeficiency virus variants influence AIDS progression

Genetic variants of human and simian immunodeficiency virus (HIV and SIV) that evolve during the course of infection and progression to AIDS are phenotypically and antigenically distinct from their progenitor viruses present at early stages of infection. However, it has been unclear how these late variants, which are typically T-cell tropic, cytopathic and resistant to neutralizing antibodies, influence the development of clinical AIDS. To address this, we infected macaques with cloned SIVs representing prototype variants from early-, intermediate- and late-stage infection having biological characteristics typical of viruses found at similar stages of HIV infection in humans. These studies demonstrate that sequential, phenotypic and antigenic variants represent viruses that have become increasingly fit for replication in the host, and our data support the hypothesis that emerging variants have increased pathogenicity and drive disease progression in SIV and HIV infection.     

Intrarectal inoculation of macaques by the simian immunodeficiency virus,SIVmne E11S: CD4 + depletion and AIDS

Macaca nemestrina and Macaca fascicularis were inoculated with various doses of a single-cell clone of SIV_mne-infected HuT 78 cells (E11S) by both the intravenous and intrarectal routes. Animals inoculated intravenously at each dose seroconverted and virus was isolated from peripheral blood mononuclear cells, but only the high-dose intrarectally exposed macaques became viremic and seroconverted. However, some seronegative, virus isolation negative intrarectally inoculated macaques showed evidence of infection and disease.     

Passive neutralizing antibody controls SHIV viremia and enhances B cell responses in infant macaques

Maternal HIV-1-specific antibodies are efficiently transferred to newborns, but their role in disease control is unknown. We administered neutralizing IgG, including the human neutralizing monoclonal IgG1b12, at levels insufficient to block infection, to six newborn macaques before oral challenge with simian-HIV strain SF162P3 (SHIV(SF162P3)). All of the macaques rapidly developed neutralizing antibodies and had significantly reduced plasma viremia for six months. These studies support the use of neutralizing antibodies in enhancing B cell responses and viral control in perinatal settings.     

Association analysis of 33 lipoprotein candidate genes in multi-generational families of African ancestry

African ancestry individuals have a more favorable lipoprotein profile than Caucasians, although the mechanisms for these differences remain unclear. We measured fasting serum lipoproteins and genotyped 768 tagging or potentially functional single nucleotide polymorphisms (SNPs) across 33 candidate gene regions in 401 Afro-Caribbeans older than 18 years belonging to 7 multi-generational pedigrees (mean family size 51, range 21–113, 3,426 relative pairs). All lipoproteins were significantly heritable (P < 0.05). Gender-specific analysis showed that heritability for triglycerides was much higher (P < 0.01) in women than in men (women, 0.62 ± 0.18, P < 0.01; men, 0.13 ± 0.17, P > 0.10), but the heritability for LDL cholesterol (LDL-C) was higher (P < 0.05) in men than in women (men, 0.79 ± 0.21, P < 0.01; women, 0.39 ± 0.12, P < 0.01). The top 14 SNPs that passed the false discovery rate threshold in the families were then tested for replication in an independent population-based sample of 1,750 Afro-Caribbean men aged 40+ years. Our results revealed significant associations for three SNPs in two genes (rs5929 and rs6511720 in LDLR and rs7517090 in PCSK9) and LDL-C in both the family study and in the replication study. Our findings suggest that LDLR and PCSK9 variants may contribute to a variation in LDL-C among African ancestry individuals. Future sequencing and functional studies of these loci may advance our understanding of genetic factors contributing to LDL-C in African ancestry populations.Lipoprotein abnormalities, characterized by elevated levels of LDL cholesterol (LDL-C) and triglycerides (TRIG) and low levels of HDL cholesterol (HDL-C), have a central role in the development of atherosclerotic coronary heart disease (CHD). A recent meta-analysis, including 3,000 individuals with CHD-related deaths, showed that HDL-C and LDL-C are independently associated with CHD risk (1). There is also considerable evidence that high levels of TRIG are an additional, independent risk factor for CHD (2, 3), although this is still controversial (4).Individuals of African ancestry have a more favorable lipoprotein profile than Caucasians, characterized by lower levels of TRIG and higher levels of HDL-C (5–8). The mechanisms responsible for these ethnic differences remain to be defined. In particular, the differences in TRIG levels are independent of the greater degree of obesity among individuals of African ancestry and several other risk factors and appear to be consistent across African populations in different environments (9), indicating a possible role of genetic factors. Although genetic factors are important in determining lipoprotein levels, little data exists regarding the importance of heredity and specific genetic factors in determining lipoprotein levels in populations of African ancestry, especially outside the US, and the findings from previous studies in African-Americans may not necessarily apply to other African ancestry populations. Recently, several genome-wide association studies identified a number of loci contributing to inter-individual variation in lipoprotein levels (10, 11). However, the majority of these studies were restricted to Caucasian populations. Given the ethnic differences in lifestyle and environmental factors, as well as in genetic background, it is important to examine genes related to lipoprotein metabolism in different ethnic groups. Therefore, we examined the heritability of fasting, serum levels of HDL-C, LDL-C, and TRIG and systematically screened for association with 33 positional and biological candidate genes in large, multigenerational families of African ancestry.