Mouse mutants from chemically mutagenized embryonic stem cells.

The drive to characterize functions of human genes on a global scale has stimulated interest in large-scale generation of mouse mutants. Conventional germ-cell mutagenesis with N-ethyl-N-nitrosourea (ENU) is compromised by an inability to monitor mutation efficiency, strain and interlocus variation in mutation induction, and extensive husbandry requirements. To overcome these obstacles and develop new methods for generating mouse mutants, we devised protocols to generate germline chimaeric mice from embryonic stem (ES) cells heavily mutagenized with ethylmethanesulphonate (EMS). Germline chimaeras were derived from cultures that underwent a mutation rate of up to 1 in 1,200 at the Hprt locus (encoding hypoxanthine guanine phosphoribosyl transferase). The spectrum of mutations induced by EMS and the frameshift mutagen ICR191 was consistent with that observed in other mammalian cells. Chimaeras derived from ES cells treated with EMS transmitted mutations affecting several processes, including limb development, hair growth, hearing and gametogenesis. This technology affords several advantages over traditional mutagenesis, including the ability to conduct shortened breeding schemes and to screen for mutant phenotypes directly in ES cells or their differentiated derivatives.

Embryonic stem cells and embryoid bodies express lymphocyte costimulatory molecules.

Despite the importance of the costimulatory proteins B7-1 (CD80), B7-2 (CD86), and their counterreceptors CD28 and CTLA-4 (CD154) in the regulation of T cell proliferation in the adult immunological system, the initial appearance of these proteins during embryonic development has not been investigated. Using in vitro cultures of undifferentiated mouse embryonic stem (ES) cells and differentiating embryoid bodies as a model of very early embryonic development, we examined these cells for the presence of mRNA and protein corresponding to the B7 and CD28 families of costimulatory molecules. By flow cytometry, a stochastically regulated subpopulation of B7-1+ cells comprising 33% of total cells was detected in ES cell cultures, while negligible staining was found for B7-2, CTLA-4, and CD28. When ES cells were differentiated into embryoid bodies for 12 days, a CD45+ subpopulation of embryoid body cells were found to stain positively for B7-1, B7-2, and CD28. RT-PCR confirmed cell staining data by revealing amplification products corresponding to B7-1, B7-2, and CD28 in corresponding samples. Very low levels of CTLA-4 amplification products were found in all samples; however, surface staining of CTLA-4 was never detected. The functional capacity of ES cell B7-1 to bind its ligand was verified by the ability of the soluble fusion protein CTLA-4-Ig to bind ES cells and the ability of this reagent to block anti-B7-1 antibody binding in cell based competition assays. These results demonstrate that expression of costimulatory molecules arises very early during in vitro development and suggests that the early embryonic environment may utilize cellular signaling systems analogous to those seen in the immune system.

Commentary on nature-culture parallelisms.

PIP: The legitimacy of the concept of parallel outcomes in animal and human behavior was assessed through fieldwork conducted in Logoli in Western Kenya. This area was selected because it has the highest population density in East Africa. According to evolutionary ecology, populations in high-density conditions demonstrate heightened competitive abilities. In comparison to other Kenyan populations, the Logoli have developed more innovative land use adaptations, including more efficient use of fertilizers and the introduction of cash cropping and farmers' sales cooperatives. Psychological testing of Logoli high school students has revealed above average achievement concerns, valuation of education, and a below average degree of psychosomatic symptoms. These observations suggest that higher order cognitive processes are unable to obliterate nature-culture parallelisms. Moreover, it is likely that behavioral phenomenon widespread in animals and adopted in human society become further entrenched through social institutionalization. An important implication of this observation is that attempts to modify institutional arrangements that have their basis in parallelism (e.g., male dominance) will meet with resistance.^ieng

Clubfoot analysis with three-dimensional computer modeling.

Which way are the bones rotated in a clubfoot? This question has long been debated by clubfoot surgeons. Opinions have been based on observations from surgery, radiographs, and autopsies. These methods all have pitfalls and are subject to misinterpretation. We used three-dimensional computer modeling to analyze histologic sections of a newborn clubfoot and a newborn normal foot. Relative to the bimalleolar axis in the axial plane, the normal talus demonstrated 5 degrees of internal rotation of its body and 25 degrees internal rotation of its neck. The clubfoot talus showed 14 degrees of external rotation of its body and 45 degrees of internal rotation of its neck. The calcaneus was externally rotated 5 degrees in the normal foot and internally rotated 22 degrees in the clubfoot.