Bone Marrow Deficiency of MCPIP1 Results in Severe Multi-Organ Inflammation but Diminishes Atherogenesis in Hyperlipidemic Mice

Objective

MCPIP1 is a newly identified protein that profoundly impacts immunity and inflammation. We aim to test if MCPIP1 deficiency in hematopoietic cells results in systemic inflammation and accelerates atherogenesis in mice.

Approach and Results

After lethally irradiated, LDLR^−/− mice were transplanted with bone marrow cells from either wild-type or MCPIP1^−/− mice. These chimeric mice were fed a western-type diet for 7 weeks. We found that bone marrow MCPIP1^−/− mice displayed a phenotype similar to that of whole body MCPIP1^−/− mice, with severe systemic and multi-organ inflammation. However, MCPIP1^−/− bone marrow recipients developed >10-fold less atherosclerotic lesions in the proximal aorta than WT bone marrow recipients, and essentially no lesions in en face aorta. The diminishment in atherosclerosis in bone marrow MCPIP1^−/− mice may be partially attributed to the slight decrease in their plasma lipids. Flow cytometric analysis of splenocytes showed that bone marrow MCPIP1^−/− mice contained reduced numbers of T cells and B cells, but increased numbers of regulatory T cells, Th17 cells, CD11b+/Gr1+ cells and CD11b+/Ly6C^low cells. This overall anti-atherogenic leukocyte profile may also contribute to the reduced atherogenesis. We also examined the cholesterol efflux capability of MCPIP1 deficient macrophages, and found that MCPIP1deficiency increased cholesterol efflux to apoAI and HDL, due to increased protein levels of ABCA1 and ABCG1.

Conclusions

Hematopoietic deficiency of MCPIP1 resulted in severe systemic and multi-organ inflammation but paradoxically diminished atherogenesis in mice. The reduced atheroegensis may be explained by the decreased plasma cholesterol levels, the anti-atherogenic leukocyte profile, as well as enhanced cholesterol efflux capability. This study suggests that, while atherosclerosis is a chronic inflammatory disease, the mechanisms underlying atherogenesis-associated inflammation in arterial wall versus the inflammation in solid organs may be substantially different.     

Monitoring of Mass Distribution Interventions for Trachoma in Plateau State,Nigeria

Mass drug administration (MDA) with antibiotics is a key component of the SAFE strategy for trachoma control. Guidelines recommend that where MDA is warranted the whole population be targeted with 80% considered the minimum acceptable coverage. In other countries, MDA is usually conducted by salaried Ministry of Health personnel (MOH). In Plateau State, Nigeria, the existing network of volunteer Community Directed Distributors (CDD) was used for the first trachoma MDA. We conducted a population-based cluster random survey (CRS) of MDA participation to determine the true coverage and compared this to coverage reported from CDD registers. We surveyed 1,791 people from 352 randomly selected households in 24 clusters in three districts in Plateau State in January 2011, following the implementation of MDA. Households were enumerated and all individuals present were asked about MDA participation. Household heads were questioned about household-level characteristics and predictors of participation. Individual responses were compared with the CDD registers. MDA coverage was estimated as 60.3% (95% CI 47.9–73.8%) by the survey compared with 75.8% from administrative program reports. CDD registration books for comparison with responses were available in 19 of the 24 clusters; there was a match for 658/682 (96%) of verifiable responses. CDD registers did not list 481 (41.3%) of the individuals surveyed. Gender and age were not associated with individual participation. Overall MDA coverage was lower than the minimum 80% target. The observed discrepancy between the administrative coverage estimate from program reports and the CRS was largely due to identification of communities missed by the MDA and not reported in the registers. CRS for evaluation of MDA provides a useful additional monitoring tool to CDD registers. These data support modification of distributor training and MDA delivery to increase coverage in subsequent rounds of MDA.     

A species in decline: genetic diversity and conservation of the Victorian eastern barred bandicoot, Perameles gunnii

The eastern barred bandicoot, Perameles gunnii, has undergone a dramatic decline in distribution and abundance on the mainland of Australia during the twentieth century. In 1988 a captive breeding program was initiated to reduce the chance of extinction. With the extinction of the last wild mainland population in the early 1990s, reintroductions from captive-bred P. gunnii have met limited success, and currently only two extant populations persist in predator proof enclosures in the State of Victoria. With ~20 years of breeding, there are concerns that the genetic diversity within the breeding program has declined and may inhibit current and future success of the program. We have used ten nuclear microsatellite loci and sequencing of two partial mitochondrial genes (cytochrome oxidase I and ATPase 6) to determine genetic diversity within current Victorian P. gunnii. These diversity estimates are compared with historic samples from the captive breeding program dating back to 1995, historic samples from the last wild mainland population found at Hamilton in 1992 and contemporary Tasmanian wild populations. Results indicate that the captive P. gunnii population in the State of Victoria has lost significant genetic diversity through time. Genetic diversity is also reduced in populations at Hamilton Community Parklands and Mount Rothwell. Samples from the last wild population at Hamilton collected in 1992, along with samples from Tasmanian P. gunnii, had significantly greater genetic diversity than contemporary mainland populations. The results are discussed with reference to management options for maintaining genetic diversity within Victorian P. gunnii, including crossing Victorian and Tasmanian P. gunnii to increase genetic diversity, adaptability and evolutionary potential.     

The Oxytricha trifallax Macronuclear Genome: A Complex Eukaryotic Genome with 16,000 Tiny Chromosomes

The macronuclear genome of the ciliate Oxytricha trifallax displays an extreme and unique eukaryotic genome architecture with extensive genomic variation. During sexual genome development, the expressed, somatic macronuclear genome is whittled down to the genic portion of a small fraction (∼5%) of its precursor “silent” germline micronuclear genome by a process of “unscrambling” and fragmentation. The tiny macronuclear “nanochromosomes” typically encode single, protein-coding genes (a small portion, 10%, encode 2–8 genes), have minimal noncoding regions, and are differentially amplified to an average of ∼2,000 copies. We report the high-quality genome assembly of ∼16,000 complete nanochromosomes (∼50 Mb haploid genome size) that vary from 469 bp to 66 kb long (mean ∼3.2 kb) and encode ∼18,500 genes. Alternative DNA fragmentation processes ∼10% of the nanochromosomes into multiple isoforms that usually encode complete genes. Nucleotide diversity in the macronucleus is very high (SNP heterozygosity is ∼4.0%), suggesting that Oxytricha trifallax may have one of the largest known effective population sizes of eukaryotes. Comparison to other ciliates with nonscrambled genomes and long macronuclear chromosomes (on the order of 100 kb) suggests several candidate proteins that could be involved in genome rearrangement, including domesticated MULE and IS1595-like DDE transposases. The assembly of the highly fragmented Oxytricha macronuclear genome is the first completed genome with such an unusual architecture. This genome sequence provides tantalizing glimpses into novel molecular biology and evolution. For example, Oxytricha maintains tens of millions of telomeres per cell and has also evolved an intriguing expansion of telomere end-binding proteins. In conjunction with the micronuclear genome in progress, the O. trifallax macronuclear genome will provide an invaluable resource for investigating programmed genome rearrangements, complementing studies of rearrangements arising during evolution and disease.     

Functional Genomic Analysis of the let-7 Regulatory Network in Caenorhabditis elegans

The let-7 microRNA (miRNA) regulates cellular differentiation across many animal species. Loss of let-7 activity causes abnormal development in Caenorhabditis elegans and unchecked cellular proliferation in human cells, which contributes to tumorigenesis. These defects are due to improper expression of protein-coding genes normally under let-7 regulation. While some direct targets of let-7 have been identified, the genome-wide effect of let-7 insufficiency in a developing animal has not been fully investigated. Here we report the results of molecular and genetic assays aimed at determining the global network of genes regulated by let-7 in C. elegans. By screening for mis-regulated genes that also contribute to let-7 mutant phenotypes, we derived a list of physiologically relevant potential targets of let-7 regulation. Twenty new suppressors of the rupturing vulva or extra seam cell division phenotypes characteristic of let-7 mutants emerged. Three of these genes, opt-2, prmt-1, and T27D12.1, were found to associate with Argonaute in a let-7–dependent manner and are likely novel direct targets of this miRNA. Overall, a complex network of genes with various activities is subject to let-7 regulation to coordinate developmental timing across tissues during worm development.     

Human Spermatogenic Failure Purges Deleterious Mutation Load from the Autosomes and Both Sex Chromosomes,including the Gene DMRT1

Gonadal failure, along with early pregnancy loss and perinatal death, may be an important filter that limits the propagation of harmful mutations in the human population. We hypothesized that men with spermatogenic impairment, a disease with unknown genetic architecture and a common cause of male infertility, are enriched for rare deleterious mutations compared to men with normal spermatogenesis. After assaying genomewide SNPs and CNVs in 323 Caucasian men with idiopathic spermatogenic impairment and more than 1,100 controls, we estimate that each rare autosomal deletion detected in our study multiplicatively changes a man''s risk of disease by 10% (OR 1.10 [1.04–1.16], p<2×10⁻³), rare X-linked CNVs by 29%, (OR 1.29 [1.11–1.50], p<1×10⁻³), and rare Y-linked duplications by 88% (OR 1.88 [1.13–3.13], p<0.03). By contrasting the properties of our case-specific CNVs with those of CNV callsets from cases of autism, schizophrenia, bipolar disorder, and intellectual disability, we propose that the CNV burden in spermatogenic impairment is distinct from the burden of large, dominant mutations described for neurodevelopmental disorders. We identified two patients with deletions of DMRT1, a gene on chromosome 9p24.3 orthologous to the putative sex determination locus of the avian ZW chromosome system. In an independent sample of Han Chinese men, we identified 3 more DMRT1 deletions in 979 cases of idiopathic azoospermia and none in 1,734 controls, and found none in an additional 4,519 controls from public databases. The combined results indicate that DMRT1 loss-of-function mutations are a risk factor and potential genetic cause of human spermatogenic failure (frequency of 0.38% in 1306 cases and 0% in 7,754 controls, p = 6.2×10⁻⁵). Our study identifies other recurrent CNVs as potential causes of idiopathic azoospermia and generates hypotheses for directing future studies on the genetic basis of male infertility and IVF outcomes.     

Rational Design of a Fibroblast Growth Factor 21-Based Clinical Candidate,LY2405319

Fibroblast growth factor 21 is a novel hormonal regulator with the potential to treat a broad variety of metabolic abnormalities, such as type 2 diabetes, obesity, hepatic steatosis, and cardiovascular disease. Human recombinant wild type FGF21 (FGF21) has been shown to ameliorate metabolic disorders in rodents and non-human primates. However, development of FGF21 as a drug is challenging and requires re-engineering of its amino acid sequence to improve protein expression and formulation stability. Here we report the design and characterization of a novel FGF21 variant, LY2405319. To enable the development of a potential drug product with a once-daily dosing profile, in a preserved, multi-use formulation, an additional disulfide bond was introduced in FGF21 through Leu118Cys and Ala134Cys mutations. FGF21 was further optimized by deleting the four N-terminal amino acids, His-Pro-Ile-Pro (HPIP), which was subject to proteolytic cleavage. In addition, to eliminate an O-linked glycosylation site in yeast a Ser167Ala mutation was introduced, thus allowing large-scale, homogenous protein production in Pichia pastoris. Altogether re-engineering of FGF21 led to significant improvements in its biopharmaceutical properties. The impact of these changes was assessed in a panel of in vitro and in vivo assays, which confirmed that biological properties of LY2405319 were essentially identical to FGF21. Specifically, subcutaneous administration of LY2405319 in ob/ob and diet-induced obese (DIO) mice over 7–14 days resulted in a 25–50% lowering of plasma glucose coupled with a 10–30% reduction in body weight. Thus, LY2405319 exhibited all the biopharmaceutical and biological properties required for initiation of a clinical program designed to test the hypothesis that administration of exogenous FGF21 would result in effects on disease-related metabolic parameters in humans.     

Vitamin D (25OHD) Serum Seasonality in the United States

Background

Vitamin D is an important micronutrient for health. Hypovitaminosis D is thought to play a role in the seasonality of a number of diseases and adverse health conditions. To refine hypotheses about the links between vitamin D and seasonal diseases, good estimates of the cyclicality of serum vitamin D are necessary.

Objectives

The objective of this study is to describe quantitatively the cyclicality of 25-hydroxyvitamin D (25OHD) in the United States. We provide a statistical analysis with weekly time resolution, in comparison to the quarterly (winter/spring/summer/fall) estimates already in the literature.

Methods

We analyzed time series data on 25OHD, spanning 287 consecutive weeks. The pooled data set comes from 3.44 million serum samples from the United States. We statistically analyzed the proportion of sera that were vitamin D sufficient, defined as 25OHD ng/mL, as a function of date.

Results

In the United States, serum 25OHD follows a lagged pattern relative to the astronomical seasons, peaking in late summer (August) and troughing in late winter (February). Airmass, which is a function of solar altitude, fits the 25OHD data very well when lagged by 8 weeks.

Conclusions

Serum vitamin D levels can be modeled as a function of date, working through a double-log transformation of minimal solar airmass (easily calculated from solar altitude, retrievable from an online solar altitude/azimuth table).     

Genetic Structure of Wild Bonobo Populations: Diversity of Mitochondrial DNA and Geographical Distribution

Bonobos (Pan paniscus) inhabit regions south of the Congo River including all areas between its southerly tributaries. To investigate the genetic diversity and evolutionary relationship among bonobo populations, we sequenced mitochondrial DNA from 376 fecal samples collected in seven study populations located within the eastern and western limits of the species’ range. In 136 effective samples from different individuals (range: 7–37 per population), we distinguished 54 haplotypes in six clades (A1, A2, B1, B2, C, D), which included a newly identified clade (D). MtDNA haplotypes were regionally clustered; 83 percent of haplotypes were locality-specific. The distribution of haplotypes across populations and the genetic diversity within populations thus showed highly geographical patterns. Using population distance measures, seven populations were categorized in three clusters: the east, central, and west cohorts. Although further elucidation of historical changes in the geological setting is required, the geographical patterns of genetic diversity seem to be shaped by paleoenvironmental changes during the Pleistocene. The present day riverine barriers appeared to have a weak effect on gene flow among populations, except for the Lomami River, which separates the TL2 population from the others. The central cohort preserves a high genetic diversity, and two unique clades of haplotypes were found in the Wamba/Iyondji populations in the central cohort and in the TL2 population in the eastern cohort respectively. This knowledge may contribute to the planning of bonobo conservation.     

FireStem2D – A Two-Dimensional Heat Transfer Model for Simulating Tree Stem Injury in Fires

FireStem2D, a software tool for predicting tree stem heating and injury in forest fires, is a physically-based, two-dimensional model of stem thermodynamics that results from heating at the bark surface. It builds on an earlier one-dimensional model (FireStem) and provides improved capabilities for predicting fire-induced mortality and injury before a fire occurs by resolving stem moisture loss, temperatures through the stem, degree of bark charring, and necrotic depth around the stem. We present the results of numerical parameterization and model evaluation experiments for FireStem2D that simulate laboratory stem-heating experiments of 52 tree sections from 25 trees. We also conducted a set of virtual sensitivity analysis experiments to test the effects of unevenness of heating around the stem and with aboveground height using data from two studies: a low-intensity surface fire and a more intense crown fire. The model allows for improved understanding and prediction of the effects of wildland fire on injury and mortality of trees of different species and sizes.