Noteworthy

Bovine spongiform encephalopathy (BSE) is caused by propagation of misfolded normal cellular prion protein PrP^C. Mice do not naturally develop prion diseases, and disruption of PrP^C expression does not result in developmental abnormalities. To assess the effects of PrP^C ablation in a species in which prion disease naturally occurs, investigators produced Holstein bulls with homozygous loss of PrP^C (PRNP^−/−) protein. No developmental, biochemical or neurological deficits were noted from birth to age 20 months. Histological examination of tissues from the central nervous systems was unremarkable, as were cytokine response and T-cell proliferation following mitogen stimulation. Brain homogenates from PRNP^−/− cattle and wild-type cattle were used in a protein misfolding cyclic amplification assay with brain homogenates from a cow with BSE used as the inoculum. In this assay, PrP^BSE was readily detected in the wild-type cattle while the PRNP^−/− cattle were negative. The authors conclude that their novel cattle are a more relevant model for investigating prion diseases and they speculate that these cattle could be used as a source of prion-free bovine-derived products and tissues.

Richt JA, Kasinathan P, Hamir AN, et al. Production of cattle lacking prion protein. Nat Biotechnol 25(1): 132–138, 2007

Cancer genomes are known to harbor multiple somatic mutations. Mutations which undergo positive selection and confer a growth advantage to the transformed cell are known as “driver” mutations while those mutations which are not subject to selection are called “passenger” mutations. In an effort to distinguish driver mutations from passenger mutations and identify new cancer related genes, this group sequenced the coding regions of 518 protein kinase genes in 210 human cancers of various types. The protein kinase family was chosen based on its known involvement in carcinogenesis. By focusing on a specific family of genes, the authors felt that they would be more likely to identify cancer related genes than whole-genome sequencing of cancer cells that other groups have undertaken. A total of 274 megabases of DNA were sequenced and over 1,000 somatic mutations were found. Most of these consisted of single base substitutions. The number of mutations varied widely among the cancer types, which may reflect different exposures to carcinogens and other harmful compounds. The majority of the mutations were determined to be passenger mutations; however, 120 of the kinase genes harbored what is believed to be a driver mutation–a much higher prevalence than was anticipated. This study shows that the cancer genome has more mutations than previously realized, and the investigators were successful in identifying new genes that may play a role in cancer development.

Greenman C, Stephens P, Smith R, et al. Patterns of somatic mutation in human cancer genomes. Nature 446(7132): 153–158, 2007

Many different software packages for image analysis are available. Some are open-source while others are proprietary and can be quite expensive. Additionally, many of these programs are designed for a limited number of applications and making modifications or customizing the programs may not be possible. In this report, the authors describe a new, free, open-source software program that is capable of a wide variety of image analysis applications. CellProfiler™ is compatible with most operating systems and is available online at www.cellprofiler.org. This paper describes various uses of CellProlifer™ to count colonies, analyze colony morphology, perform cell microarray annotation, murine tumor quantification, wound healing assays, and tissue topography measurements. Size measurements include basic parameters such as area, perimeter, and length, but there are 35 other measurements included in these calculations. Tissue topography analysis includes determination of the number of neighbors each call has, which may be useful in studying tumor-stromal interactions. All data captured by the program is easily exported into spreadsheets and databases for further analysis. The website offers the viewer multiple examples of experimental images and analysis applications.

Lamprecht MR, Sabatini DM, Carpenter AE. CellProfiler™: Free, versatile software for automated biological image analysis. Biotechniques 42(1): 71–75, 2007

In pulmonary alveolar proteinosis (PAP) there is accumulation of surfactant in the alveoli with subsequent respiratory insufficiency and sometimes death. PAP is associated with neutralizing autoantibodies to granulocyte-macrophage colony stimulating factor (GM-CSF). Humans with PAP as well as GM-CSF^−/− mice have an increased susceptibility to opportunistic infections, prompting the authors of this paper to hypothesize that the GM-CSF autoantibodies lead to impaired neutrophil function in humans and mice. GM-CSF primes neutrophils by increasing their levels of CD11b which promotes adhesion to the vascular endothelium in response to infection. Neutrophils from PAP patients and GM-CSF^−/− mice had impaired basal functions, including reduced phagocytic and bactericidal activity, cellular adhesion, and oxidative burst. Priming with GM-CSF restored these functions in neutrophils from the knock-out mice but not in neutrophils from PAP patients. Using neutrophils from healthy humans and wild-type mice, the pattern of neutrophil dysfunction could be reproduced by incubation with purified GM-CSF antibodies. These effects appear to be independent and distinct from effects of granulocyte-colony stimulating factor on neutrophil function. These findings shed light on mechanisms of neutrophil function and may be relevant to further investigate the use of GM-CSF therapeutically to augment innate immunity in the setting of severe infection.

Uchida K, Beck DC, Yamamoto T. GM-CSF autoantibodies and neutrophil dysfunction in pulmonary alveolar proteinosis. N Engl J Med 356(6): 567–579, 2007

Identification of canine hematopoietic progenitor cells (HPCs) would be useful for further understanding normal and abnormal hematopoieses, diagnosing disease conditions of the bone marrow, and for therapy involving bone marrow transplantation. This report describes the isolation and phenotypic characterization of multipotent HPCs from canine bone marrow. Equilibrium discontinued density centrifugation followed by in vitro colony formation assays revealed that HPCs were enriched in the relatively low density fraction. Although this fraction contained CD34+ and MHC class II expressing cells, these were not significantly enriched. Further testing showed that multipotent HPCs and cells committed to the myeloid lineage had high affinity for wheat germ agglutination. The WGA^high population demonstrated dull (low) staining with Rhodamin 123. These cells were round and greater than 10 microns in diameter, with a blast-like morphology including a large nucleus and large nucleoli. The ability to isolate this population should prove useful in a number of aspects of canine hematology.

Wijewardana V, Sugiura K, Shigeyama N. Isolation and characterization of hematopoietic progenitor cells in canine bone marrow. Vet Immunol Immunopathol 115(3–4): 230–238, 2007