in-vivo-targeting-neural-progenitor-cells-ferret-neocortex-utero

In Vivo Targeting of Neural Progenitor Cells in Ferret Neocortex by In Utero Electroporation

Manipulation of gene expression in vivo during embryonic development is the method of choice when analyzing the role of individual genes during mammalian ...

Max Planck Institute of Molecular Cell Biology and Genetics

Mutations in ASPM (abnormal spindle-like microcephaly associated) cause primary microcephaly in humans, a disorder characterized by a major reduction in brain size in the apparent absence of nonneurological anomalies. The function of the Aspm protein in neural progenitor cell expansion, as well as its localization to the mitotic spindle and midbody, suggest that it regulates brain development by a cell division-related mechanism. Furthermore, evidence that positive selection affected ASPM during primate evolution has led to suggestions that such a function changed during primate evolution. Here, we report that in Aspm mutant mice, truncated Aspm proteins similar to those causing microcephaly in humans fail to localize to the midbody during M-phase and cause mild microcephaly. A human ASPM transgene rescues this phenotype but, interestingly, does not cause a gain of function. Strikingly, truncated Aspm proteins also cause a massive loss of germ cells, resulting in a severe reduction in testis and ovary size accompanied by reduced fertility. These germline effects, too, are fully rescued by the human ASPM transgene, indicating that ASPM is functionally similar in mice and humans. Our findings broaden the spectrum of phenotypic effects of ASPM mutations and raise the possibility that positive selection of ASPM during primate evolution reflects its function in the germline.

Mutations in mouse Aspm (abnormal spindle-like microcephaly associated) cause not only microcephaly but also major defects in the germline.

Soy isoflavones (IF) are in the focus of biomedical research since more than two decades. To assess their bioactivity, IF are investigated in rats and mice as a model. As the biological activity of IF is affected by their biotransformation, our aim was to comprehensively compare the conjugative and microbial metabolism of daidzein and genistein in adult humans, rats and mice of both sexes. One identical soy extract and a validated LC-MS method were used for all studies. We detected considerable differences between the three species. In rats and mice, sex-specific differences were observed in addition. The major plasma phase II metabolites in humans were the 7-sulfo-4'-glucuronides (39-49 %) and, in case of genistein, also the diglucuronide (34 %), whereas in mice monosulfates (33-41 %) and monoglucuronides (30-40 %) predominated. In male rats the disulfates (23-62 %) and 7-sulfo-4'-glucuronides (19-54 %) were predominant, while in female rats the 7-glucuronides (81-93 %) exhibited highest concentrations. The portion of aglycones was low in humans (0.5-1.3 %) and rats (0.5-3.1 %) but comparatively high in mice (3.1-26.0 %), especially in the case of daidzein. Furthermore, substantial differences were observed between daidzein and genistein metabolism. In contrast to humans, all rats and mice were equol producer, independent of their sex. In conclusion, there are marked differences between humans, rats and mice in the profile of major metabolites following IF phase II metabolism. These differences may contribute to resolve inconsistencies in results concerning the bioactivity of IF and should be considered when applying findings of animal studies to humans, e.g., for risk assessment. 

Phase II metabolism of the soy isoflavones genistein and daidzein in humans, rats and mice: a cross-species and sex comparison.

Erratum to: Phase II metabolism of the soy isoflavones genistein and daidzein in humans, rats and mice: a cross-species and sex comparison.

One of the most commonly used protein sources in rodent diets is soy, which is naturally rich in phytoestrogens. Although phytoestrogens have shown potential health benefits in humans, they may also have the ability to disrupt reproduction. Consequently, there has been a tendency to try to exclude them from rodent diets. In the current study, we investigated whether phytoestrogen content in the mouse diet could affect reproduction in mice used as embryo donors. Donor mice (C57BL/6JOlaHsd) were maintained with three different diets: high phytoestrogen (ca. 400 mg/kg genistein), low phytoestrogen (ca. 10 mg/kg genistein) and standard breeding diet (ca. 120 mg/kg genistein). Mice fed a high phytoestrogen diet had a high yield of plugs, embryos, and injectable embryos, as well as producing good quality embryos. Results from donor mice fed a low phytoestrogen diet were consistently but only slightly inferior, whereas mice fed a standard diet performed the poorest. Interestingly, the largest number of born and weaned offspring were observed when recipient females received embryos from the standard diet group. Sperm yield and quality of stud males did not differ between the groups. We surmize that for experimental endpoints requiring fertilized embryos it may be more beneficial to feed mice a diet containing phytoestrogen, but if the goal is to produce transgenic mice, a diet high in phytoestrogen may be inadvisable. In conclusion, care should be taken when selecting a diet for experimental mouse colonies as phytoestrogen could influence the study outcome.

Jussi Helppi

Max Planck Institute of Molecular Cell Biology and Genetics

In Vivo Targeting of Neural Progenitor Cells in Ferret Neocortex by In Utero Electroporation