Infrared Thermography and Ultrasonography to Indirectly Monitor the Influence of Liner Type and Overmilking on Teat Tissue Recovery

Eight Danish Holstein cows were milked with a 1-mm thick specially designed soft liner on their right rear teat and a standard liner mounted under extra high tension on their left rear teat. Four of the animals were overmilked for 5 min. Rear teats were subjected to ultrasound examination on the first day and to infrared thermography on the second day. Teats were submersed in ethanol 20 min post-milking on the second day. Ultrasonography measurements showed that teat canal length increased by 30–41% during milking. Twenty minutes after milking, teats milked with modified standard liners still had elongated teat canals while teats milked with the soft liner were normalized. Overmilking tended to increase teat wall thickness. Approximately 80% of variability in teat canal length, from before teat preparation to after milking, could be explained by changes during teat preparation. Thermography indicated a general drop in teat temperature during teat preparation. Teat temperature increased during milking and continued to increase until the ethanol challenge induced a significant drop. Temperatures approached pre-challenge rather than pre-milking temperatures within 10 minutes after challenge. Teat temperatures were dependent on type of liner. Mid-teat temperatures post-challenge relative to pre-teat preparation were dependent on overmilking. Thermography and ultrasound were considered useful methods to indirectly and non invasively evaluate teat tissue integrity.     

Germline Mutations in NFKB2 Implicate the Noncanonical NF-κB Pathway in the Pathogenesis of Common Variable Immunodeficiency

Common variable immunodeficiency (CVID) is a heterogeneous disorder characterized by antibody deficiency, poor humoral response to antigens, and recurrent infections. To investigate the molecular cause of CVID, we carried out exome sequence analysis of a family diagnosed with CVID and identified a heterozygous frameshift mutation, c.2564delA (p.Lys855Serfs^∗7), in NFKB2 affecting the C terminus of NF-κB2 (also known as p100/p52 or p100/p49). Subsequent screening of NFKB2 in 33 unrelated CVID-affected individuals uncovered a second heterozygous nonsense mutation, c.2557C>T (p.Arg853^∗), in one simplex case. Affected individuals in both families presented with an unusual combination of childhood-onset hypogammaglobulinemia with recurrent infections, autoimmune features, and adrenal insufficiency. NF-κB2 is the principal protein involved in the noncanonical NF-κB pathway, is evolutionarily conserved, and functions in peripheral lymphoid organ development, B cell development, and antibody production. In addition, Nfkb2 mouse models demonstrate a CVID-like phenotype with hypogammaglobulinemia and poor humoral response to antigens. Immunoblot analysis and immunofluorescence microscopy of transformed B cells from affected individuals show that the NFKB2 mutations affect phosphorylation and proteasomal processing of p100 and, ultimately, p52 nuclear translocation. These findings describe germline mutations in NFKB2 and establish the noncanonical NF-κB signaling pathway as a genetic etiology for this primary immunodeficiency syndrome.     

Shared Segment Analysis and Next-Generation Sequencing Implicates the Retinoic Acid Signaling Pathway in Total Anomalous Pulmonary Venous Return (TAPVR)

Most isolated congenital heart defects are thought to be sporadic and are often ascribed to multifactorial mechanisms with poorly understood genetics. Total Anomalous Pulmonary Venous Return (TAPVR) occurs in 1 in 15,000 live-born infants and occurs either in isolation or as part of a syndrome involving aberrant left-right development. Previously, we reported causative links between TAVPR and the PDGFRA gene. TAPVR has also been linked to the ANKRD1/CARP genes. However, these genes only explain a small fraction of the heritability of the condition. By examination of phased single nucleotide polymorphism genotype data from 5 distantly related TAPVR patients we identified a single 25 cM shared, Identical by Descent genomic segment on the short arm of chromosome 12 shared by 3 of the patients and their obligate-carrier parents. Whole genome sequence (WGS) analysis identified a non-synonymous variant within the shared segment in the retinol binding protein 5 (RBP5) gene. The RBP5 variant is predicted to be deleterious and is overrepresented in the TAPVR population. Gene expression and functional analysis of the zebrafish orthologue, rbp7, supports the notion that RBP5 is a TAPVR susceptibility gene. Additional sequence analysis also uncovered deleterious variants in genes associated with retinoic acid signaling, including NODAL and retinol dehydrogenase 10. These data indicate that genetic variation in the retinoic acid signaling pathway confers, in part, susceptibility to TAPVR.     

Evolution of body condition‐dependent dispersal in metapopulations

Body condition‐dependent dispersal strategies are common in nature. Although it is obvious that environmental constraints may induce a positive relationship between body condition and dispersal, it is not clear whether positive body conditional dispersal strategies may evolve as a strategy in metapopulations. We have developed an individual‐based simulation model to investigate how body condition–dispersal reaction norms evolve in metapopulations that are characterized by different levels of environmental stochasticity and dispersal mortality. In the model, body condition is related to fecundity and determined either by environmental conditions during juvenile development (adult dispersal) or by those experienced by the mother (natal dispersal). Evolutionarily stable reaction norms strongly depend on metapopulation conditions: positive body condition dependency of dispersal evolved in metapopulation conditions with low levels of dispersal mortality and high levels of environmental stochasticity. Negative body condition‐dependent dispersal evolved in metapopulations with high dispersal mortality and low environmental stochasticity. The latter strategy is responsible for higher dispersal rates under kin competition when dispersal decisions are based on body condition reached at the adult life stage. The evolution of both positive and negative body condition‐dependent dispersal strategies is consequently likely in metapopulations and depends on the prevalent environmental conditions.     

重组腺相关病毒2型转导人外周血单核细胞来源树突状细胞的研究

To find out the infection efficiency of recombinant adeno-as sociated virus 2-mediated exogenou s genes in human peripheral blood monocyte-derived dendritic cells(D Cs),the process of transfection wa s investigated by FITC-labeled rAA V2,and observed under confocal mic roscope.Newly separated dendritic cells were tranfected by rAAV2-luc and rAAV2-GFP at different MOI,and transfection efficiency were detec ted by luminometer for rAAV2-luc a nd flow cytometry for rAAV2-GFP.Th e results were elucidated in four different assay systems:(1)60min w as needed for rAAV2 to bind on den dritic cells,and got into cells in the following 10min;(2)the express ion of luc could be detected at th e MOI as low as 1×10~5v.g/cell,and the expression plateau was reached by the MOI of 10~6～10~7v.g/cell,furt her increase of MOI had no functio n on expression level;(3)transgene expression was detected after 48h, and maintained a higher expression level from 96h to 240h after infec tion;(4)7days postinfection of rAA V2-GFP,5%～18% dendritic cells were GFP positive. These data suggest th at rAAV2 vector can efficiently in fect monocyte-derived dendritic ce lls and mediate exogenous gene exp ression,and that the application o f rAAV2 as vector may be useful fo r gene transfer to dendritic cells in ex vivo immunotherapy.     

Clinical and Biochemical Function of Polymorphic NR0B1 GGAA-Microsatellites in Ewing Sarcoma: A Report from the Children's Oncology Group

Background

The genetics involved in Ewing sarcoma susceptibility and prognosis are poorly understood. EWS/FLI and related EWS/ETS chimeras upregulate numerous gene targets via promoter-based GGAA-microsatellite response elements. These microsatellites are highly polymorphic in humans, and preliminary evidence suggests EWS/FLI-mediated gene expression is highly dependent on the number of GGAA motifs within the microsatellite.

Objectives

Here we sought to examine the polymorphic spectrum of a GGAA-microsatellite within the NR0B1 promoter (a critical EWS/FLI target) in primary Ewing sarcoma tumors, and characterize how this polymorphism influences gene expression and clinical outcomes.

Results

A complex, bimodal pattern of EWS/FLI-mediated gene expression was observed across a wide range of GGAA motifs, with maximal expression observed in constructs containing 20–26 GGAA motifs. Relative to white European and African controls, the NR0B1 GGAA-microsatellite in tumor cells demonstrated a strong bias for haplotypes containing 21–25 GGAA motifs suggesting a relationship between microsatellite function and disease susceptibility. This selection bias was not a product of microsatellite instability in tumor samples, nor was there a correlation between NR0B1 GGAA-microsatellite polymorphisms and survival outcomes.

Conclusions

These data suggest that GGAA-microsatellite polymorphisms observed in human populations modulate EWS/FLI-mediated gene expression and may influence disease susceptibility in Ewing sarcoma.     

Genetic evidence for larger African population size during recent human evolution

Genetic evidence suggests that the long-term average effective size of sub-Saharan Africa is larger than other geographic regions. A method is described that allows estimation of relative long-term regional population sizes. This method is applied to 60 microsatellite DNA loci from a sample of 72 sub-Saharan Africans, 63 East Asians, and 120 Europeans. Average heterozygosity is significantly higher in the sub-Saharan African sample. Expected heterozygosity was computed for each region and locus using a population genetic model based on the null hypothesis of equal long-term population sizes. Average residual heterozygosity is significantly higher in the sub-Saharan African sample, indicating that African population size was larger than other regions during recent human evolution. The best fit of the model is with relative population weights of 0.73 for sub-Saharan Africa, 0.09 for East Asia, and 0.18 for Europe. These results are similar to those obtained using craniometric variation for these three geographic regions. These results, combined with inferences from other genetic studies, support a major role of Africa in the origin of modern humans. It is less clear, however, whether complete African replacement is the most appropriate model. An alternative is an African origin with non-African gene flow. While Africa is an important region in recent human evolution, it is not clear whether the gene pool of our species is completely out of Africa or predominately out of Africa.     

Genetic similarities within and between human populations

The proportion of human genetic variation due to differences between populations is modest, and individuals from different populations can be genetically more similar than individuals from the same population. Yet sufficient genetic data can permit accurate classification of individuals into populations. Both findings can be obtained from the same data set, using the same number of polymorphic loci. This article explains why. Our analysis focuses on the frequency, omega, with which a pair of random individuals from two different populations is genetically more similar than a pair of individuals randomly selected from any single population. We compare omega to the error rates of several classification methods, using data sets that vary in number of loci, average allele frequency, populations sampled, and polymorphism ascertainment strategy. We demonstrate that classification methods achieve higher discriminatory power than omega because of their use of aggregate properties of populations. The number of loci analyzed is the most critical variable: with 100 polymorphisms, accurate classification is possible, but omega remains sizable, even when using populations as distinct as sub-Saharan Africans and Europeans. Phenotypes controlled by a dozen or fewer loci can therefore be expected to show substantial overlap between human populations. This provides empirical justification for caution when using population labels in biomedical settings, with broad implications for personalized medicine, pharmacogenetics, and the meaning of race.