Isolamento, coltura e caratterizzazione di cellule staminali mesenchimali dall'apparato genitale della cagna

Abstract

Mesenchymal stem cells (MSC) can be isolated from a variety of adult animal tissues, replicated in vitro and induced to differentiate into different cell lineages. In the dog many districts have been investigated for MSC but never the female genital tract. In the last decade, progenitor cells isolated from dissociated endometrial tissue, have been the subject of many studies in several animal species. Recently, endometrial cells showing characteristics of mesenchymal stem cells (MSC) have been demonstrated in human, pig and cow uterine tissue samples. The aim of the present study was simultaneously studied the tissue collected from three different sites: ovarian bursa adipose tissue, endometrium and abdominal adipose tissue, used as control. All isolated cells were able to adhere to plastic support, showed similar fibroblastic-like morphology and ability of subculture; MSC isolated from abdominal adipose tissue and endometrium showed ability to generate colony-forming units, plasticity and expressed specific surface antigens. Conversely cells isolated from ovarian bursa adipose tissue did not express specific surface antigens and did not show ability to generate colony-forming units and plasticity. Multipotent stromal cells could be isolated from all bitches anrolled in the study (n=7). The multipotecncy of cells was demonstrated by their capacity to differentiate into adypocytic, osteocytic and chondrocytic lineages. Clonogenicity and cell proliferation ability were also tested. Furthermore, gene expression analysis by RT-PCR was used to compare the expression of a set of genes (CD44, CD29, CD34, CD45, CD90, CD13, CD133, CD73, CD31 CD105, Oct4) with adipose tissue derived MSC. Stromal cells isolated from uterine endometrium showed similar morphology, ability of subculture and plasticity, and also expressed a panel of genes comparable with adipose tissue 4 derived MSC. These data suggest that endometrial stromal cells fulfill the basic criteria proposed by the “Mesenchymal and Tissue Stem Cell Committee of the International Society for Cellular Therapy” for the identification of mesenchymal stem cells. Although endometrial mesenchymal stem cells (EnMSC) showed a lower replicative ability in comparison to adipose tissue derived MSC, they could be considered a cell therapeutic agent alternative to adipose tissue or bone marrow derived MSC in dog.