ABSTRACT
Developmental potential was assessed in 8 intra-specific and 20 inter-specific hybrid clones obtained by fusion of embryonic stem (ES) cells with either splenocytes or fetal fibroblasts. Number of chromosomes derived from ES cells in these hybrid clones was stable while contribution of somatic partner varied from single chromosomes to complete complement. This allowed us to compare pluripotency of the hybrid cells with various numbers of somatic chromosomes. Three criteria were used for the assessment: (i) expression of Oct-4 and Nanog genes; (ii) analyses of teratomas generated by subcutaneous injections of the tested cells into immunodeficient mice; (iii) contribution of the hybrid cells in chimeras generated by injection of the tested cells into C57BL blastocysts. All tested hybrid clones showed expression of Oct-4 and Nanog at level comparable to ES cells. Histological and immunofluorescent analyses demonstrated that most teratomas formed from the hybrid cells with different number of somatic chromosomes contained derivatives of three embryonic layers. Tested hybrid clones make similar contribution in various tissues of chimeras in spite of significant differences in the number of somatic chromosomes they contained. The data indicate that pluripotency is manifested as a dominant trait in the ES hybrid cells and does not depend substantially on the number of somatic chromosomes. The latter suggests that the developmental potential derived from ES cells is maintained in ES-somatic cell hybrids by cis-manner and is rather resistant to trans-acting factors emitted from the somatic one.
Subject(s)
Chromosomes, Mammalian , Embryonic Stem Cells/physiology , Hybrid Cells/physiology , Pluripotent Stem Cells/physiology , Animals , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Embryonic Stem Cells/cytology , Homeodomain Proteins/genetics , Homeodomain Proteins/metabolism , Hybrid Cells/cytology , Karyotyping , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Nude , Nanog Homeobox Protein , Octamer Transcription Factor-3/genetics , Octamer Transcription Factor-3/metabolism , Pluripotent Stem Cells/cytology , Teratoma/metabolism , Teratoma/pathology , Tissue DistributionABSTRACT
Long-term effects of in vivo exposures to proestrogen methoxychlor (MXC) or estradiol-17beta (E) were studied during early pregnancy (preimplantation) in ICR mice. Pregnant dams received either subcutaneous injections of 1 microg of E on Day 2 of pregnancy only (vaginal plug = Day 1), or 5.0mg of MXC on Days 2-4 of pregnancy in sesame oil. Pregnant control mice were treated with the vehicle only. Litter size, postnatal survival, sex ratio at birth, and anogenital distance (AGD) in offspring of both sexes were examined, as well as vaginal opening in female offspring. High mortality rate was recorded in MXC-exposed offspring due to infanticide. Exposures to either E or MXC did not change sex ratio at birth, but the litter size was smaller in the former group. On postnatal Day 21, male pups exposed to either E or MXC at preimplantation stage exhibited shorter AGD than the controls, with the change most pronounced after MXC treatments. AGD in female offspring was unaffected after MXC exposures, but E treatments produced longer AGD in the females than that recorded in the controls. Preimplantation exposures to E or MXC also accelerated sexual maturation as significantly more females exhibited precocious vaginal opening at weaning. Our study shows that exposures to MXC or E at preimplantation stages cause long term alteration of sexual development during weaning in offspring of both sexes. Also, MXC treatments retarded both growth and weight of both sexes of offspring, in comparison to controls.
