Microsatellite genetic distances between oceanic populations of the humpback whale (Megaptera novaeangliae)

Mitochondrial DNA haplotypes of humpback whales show strong segregation between oceanic populations and between feeding grounds within oceans, but this highly structured pattern does not exclude the possibility of extensive nuclear gene flow. Here we present allele frequency data for four microsatellite loci typed across samples from four major oceanic regions: the North Atlantic (two mitochondrially distinct populations), the North Pacific, and two widely separated Antarctic regions, East Australia and the Antarctic Peninsula. Allelic diversity is a little greater in the two Antarctic samples, probably indicating historically greater population sizes. Population subdivision was examined using a wide range of measures, including Fst, various alternative forms of Slatkin's Rst, Goldstein and colleagues' delta mu, and a Monte Carlo approximation to Fisher's exact test. The exact test revealed significant heterogeneity in all but one of the pairwise comparisons between geographically adjacent populations, including the comparison between the two North Atlantic populations, suggesting that gene flow between oceans is minimal and that dispersal patterns may sometimes be restricted even in the absence of obvious barriers, such as land masses, warm water belts, and antitropical migration behavior. The only comparison where heterogeneity was not detected was the one between the two Antarctic population samples. It is unclear whether failure to find a difference here reflects gene flow between the regions or merely lack of statistical power arising from the small size of the Antarctic Peninsula sample. Our comparison between measures of population subdivision revealed major discrepancies between methods, with little agreement about which populations were most and least separated. We suggest that unbiased Rst (URst, see Goodman 1995) is currently the most reliable statistic, probably because, unlike the other methods, it allows for unequal sample sizes. However, in view of the fact that these alternative measures often contradict one another, we urge caution in the use of microsatellite data to quantify genetic distance.      

Evidence of Selection against Complex Mitotic-Origin Aneuploidy during Preimplantation Development

Whole-chromosome imbalances affect over half of early human embryos and are the leading cause of pregnancy loss. While these errors frequently arise in oocyte meiosis, many such whole-chromosome abnormalities affecting cleavage-stage embryos are the result of chromosome missegregation occurring during the initial mitotic cell divisions. The first wave of zygotic genome activation at the 4–8 cell stage results in the arrest of a large proportion of embryos, the vast majority of which contain whole-chromosome abnormalities. Thus, the full spectrum of meiotic and mitotic errors can only be detected by sampling after the initial cell divisions, but prior to this selective filter. Here, we apply 24-chromosome preimplantation genetic screening (PGS) to 28,052 single-cell day-3 blastomere biopsies and 18,387 multi-cell day-5 trophectoderm biopsies from 6,366 in vitro fertilization (IVF) cycles. We precisely characterize the rates and patterns of whole-chromosome abnormalities at each developmental stage and distinguish errors of meiotic and mitotic origin without embryo disaggregation, based on informative chromosomal signatures. We show that mitotic errors frequently involve multiple chromosome losses that are not biased toward maternal or paternal homologs. This outcome is characteristic of spindle abnormalities and chaotic cell division detected in previous studies. In contrast to meiotic errors, our data also show that mitotic errors are not significantly associated with maternal age. PGS patients referred due to previous IVF failure had elevated rates of mitotic error, while patients referred due to recurrent pregnancy loss had elevated rates of meiotic error, controlling for maternal age. These results support the conclusion that mitotic error is the predominant mechanism contributing to pregnancy losses occurring prior to blastocyst formation. This high-resolution view of the full spectrum of whole-chromosome abnormalities affecting early embryos provides insight into the cytogenetic mechanisms underlying their formation and the consequences for human fertility.     

Non-Invasive Prenatal Detection of Trisomy 13 Using a Single Nucleotide Polymorphism- and Informatics-Based Approach

Purpose

To determine how a single nucleotide polymorphism (SNP)- and informatics-based non-invasive prenatal aneuploidy test performs in detecting trisomy 13.

Methods

Seventeen trisomy 13 and 51 age-matched euploid samples, randomly selected from a larger cohort, were analyzed. Cell-free DNA was isolated from maternal plasma, amplified in a single multiplex polymerase chain reaction assay that interrogated 19,488 SNPs covering chromosomes 13, 18, 21, X, and Y, and sequenced. Analysis and copy number identification involved a Bayesian-based maximum likelihood statistical method that generated chromosome- and sample-specific calculated accuracies.

Results

Of the samples that passed a stringent DNA quality threshold (94.1%), the algorithm correctly identified 15/15 trisomy 13 and 49/49 euploid samples, for 320/320 correct copy number calls.

Conclusions

This informatics- and SNP-based method accurately detects trisomy 13-affected fetuses non-invasively and with high calculated accuracy.     

Platelet Lysates Produced from Expired Platelet Concentrates Support Growth and Osteogenic Differentiation of Mesenchymal Stem Cells

Background

Mesenchymal stem cells are promising candidates in regenerative cell therapy. Conventional culture methods involve the use of animal substances, specifically fetal bovine serum as growth supplement. Since the use of animal-derived products is undesirable for human applications, platelet lysates produced from human platelets are an attractive alternative. This is especially true if platelet lysates from already approved transfusion units at blood banks can be utilized. The purpose of this study was to produce human platelet lysates from expired, blood bank-approved platelet concentrates and evaluate their use as growth supplement in the culture of mesenchymal stem cells.

Methodology/Principal Findings

In this study, bone marrow-derived mesenchymal stem cells were cultured with one of three culture supplements; fetal bovine serum, lysates from freshly prepared human platelet concentrates, or lysates from expired human platelet concentrates. The effects of these platelet-derived culture supplements on basic mesenchymal stem cell characteristics were evaluated. All cultures maintained the typical mesenchymal stem cell surface marker expression, trilineage differentiation potential, and the ability to suppress in vitro immune responses. However, mesenchymal stem cells supplemented with platelet lysates proliferated faster than traditionally cultured cells and increased the expression of the osteogenic marker gene RUNX-2; yet no difference between the use of fresh and expired platelet concentrates was observed.

Conclusion/Significance

Our findings suggest that human platelet lysates produced from expired platelet concentrates can be used as an alternative to fetal bovine serum for mesenchymal stem cell culture to the same extent as lysates from fresh platelets.     

Detection of Clonal and Subclonal Copy-Number Variants in Cell-Free DNA from Patients with Breast Cancer Using a Massively Multiplexed PCR Methodology

We demonstrate proof-of-concept for the use of massively multiplexed PCR and next-generation sequencing (mmPCR-NGS) to identify both clonal and subclonal copy-number variants (CNVs) in circulating tumor DNA. This is the first report of a targeted methodology for detection of CNVs in plasma.Using an in vitro model of cell-free DNA, we show that mmPCR-NGS can accurately detect CNVs with average allelic imbalances as low as 0.5%, an improvement over previously reported whole-genome sequencing approaches. Our method revealed differences in the spectrum of CNVs detected in tumor tissue subsections and matching plasma samples from 11 patients with stage II breast cancer. Moreover, we showed that liquid biopsies are able to detect subclonal mutations that may be missed in tumor tissue biopsies. We anticipate that this mmPCR-NGS methodology will have broad applicability for the characterization, diagnosis, and therapeutic monitoring of CNV-enriched cancers, such as breast, ovarian, and lung cancer.     

Species hybridization between a female blue whale (Balaenoptera musculus) and a male fin whale (B. physalus): molecular and morphological documentation.

In 1986 a large, pregnant, female balaenopterid whale was caught in Icelandic waters. The animal had morphological characteristics of both the blue and the fin whale. Molecular analyses of the whale showed that it was a hybrid between a female blue whale and a male fin whale. The descent of the species hybrid was established without access to either parental specimen. Analysis of the fetus showed that it had a blue whale father. The present report of species hybridization between the two largest cetacean species, the blue and the fin whale, documents the occurrence of cetacean species hybridization in the wild. It is also the first example of any cetacean hybridization giving rise to a fertile offspring.     

Chitosan and Chitin Hexamers affect expansion and differentiation of mesenchymal stem cells differently

Chitooligosaccharides are of interest as potential drugs due to their bioactivity and water solubility. We compared the effect of acetylated and deacetylated chitooligomers (Hexamers) on short-term expansion (7 days) and osteogenic differentiation of bone-marrow derived, human mesenchymal stem cells in terms of gene expression, cytokine secretion and quality of osteogenic differentiation. We show that chitooligomers affect hMSC gene expression and cytokine secretion, but not mineralization. The effect of chitooligomers was shown to be dependent on the acetylation degree, with significantly stronger effects when cells are stimulated with chitin-derived Hexamers (N-Acetyl Chitohexaose) than with Chitosan Hexamers (Chitohexaose).