Bone Marrow Mesenchymal Stem Cells Are an Attractive Donor Cell Type for Production of Cloned Pigs As Well As Genetically Modified Cloned Pigs by Somatic Cell Nuclear Transfer

Abstract

The somatic cell nuclear transfer (SCNT) technique has been widely applied to clone pigs or to produce genetically modified pigs. Currently, this technique relies mainly on using terminally differentiated fibroblasts as donor cells. To improve cloning efficiency, only partially differentiated multipotent mesenchymal stem cells (MSCs), thought to be more easily reprogrammed to a pluripotent state, have been used as nuclear donors in pig SCNT. Although in vitro-cultured embryos cloned from porcine MSCs (MSCs-embryos) were shown to have higher preimplantation developmental ability than cloned embryos reconstructed from fibroblasts (Fs-embryos), the difference in in vivo full-term developmental rate between porcine MSCs-embryos and Fs-embryos has not been investigated so far. In this study, we demonstrated that blastocyst total cell number and full-term survival abilities of MSCs-embryos were significantly higher than those of Fs-embryos cloned from the same donor pig. The enhanced developmental potential of MSCs-embryos may be associated with their nuclear donors' DNA methylation profile, because we found that the methylation level of imprinting genes and repeat sequences differed between MSCs and fibroblasts. In addition, we showed that use of transgenic porcine MSCs generated from transgene plasmid transfection as donor cells for SCNT can produce live transgenic cloned pigs. These results strongly suggest that porcine bone marrow MSCs are a desirable donor cell type for production of cloned pigs and genetically modified cloned pigs via SCNT.

The effect of amniotic membrane stem cells as donor nucleus on gene expression in reconstructed bovine oocytes

“…The higher appropriate remodeling of stem cell-NT derived embryos has proved in different species (Bosch et al, 2006, Faast et al, 2006, Kumar et al, 2012, Lee et al, 2006, Rideout et al, 2000. Some reports, however, have rejected transcendence of stem cells over somatic cells in normal reprogramming of NT derived embryos (Berg et al, 2007, Li et al, 2013, Sung et al, 2006. In the first report on the use of bovine MSCs for NT, no priority in developmental potential was observed in stem cell derived embryos compared to non-stem cell derived counterparts (Kato et al, 2004).…”

Production of functional human nerve growth factor from the saliva of transgenic mice by using salivary glands as bioreactors

“…Previously, our group and others have described the generation of TG animals expressing microorganism-derived digestive enzymes such as phytase and cellulase specifically in the salivary glands for improvements of feed nutrient utilization11212223. Here we report the production of a therapeutic protein, the human nerve growth factor (hNGF), in the salivary glands of TG mice.…”

Multipotent Cell Types in Primary Fibroblast Cell Lines Used to Clone Pigs using Somatic Cell Nuclear Transfer

“…Instead these workers suggested that the development of cloned embryos should be assessed in terms of the number of blastocyst that develop to term rather than the number of embryos that develop to the blastocyst stage. Interestingly Lee et al [8] and Li et al [9] have reported that porcine MSCs increase blastocyst development and cell number as well as live born when these cells are used for SCNT compared with fibroblasts.…”

“…We have also reported that the use of porcine mesenchymal stem cells (MSCs) isolated from the bone marrow can increase the proportion of SCNT embryos that develop to the blastocyst stage compared with adult fibroblasts obtained from the same animal (Faast et al). Subsequent studies by Lee et al [8] and Li et al [9] have suggested that blastocyst cell number and development to term is also increased when MSCs are used compared with fibroblasts. MSCs are found in the stromal compartment of the bone marrow and can be differentiated into adipocytes, chondrocytes and osteocytes [10].…”