L-MSCs and A-MSCs exhibited the typical morphology, immunophenotype, and proliferation and differentiation pattern of MSCs. differentiation. Cultures were characterized for morphology, immunophenotype, and by quantitative real-time reverse transcriptionpolymerase chain reaction for expression of pluripotency genes or markers of differentiation. Bone marrowderived MSCs were also analyzed for comparison of these parameters. L-MSCs and A-MSCs exhibited the typical morphology, immunophenotype, and proliferation and differentiation pattern of MSCs. The analysis of gene expression showed a higher potential of adipose tissuederived MSCs toward the osteogenic pathway and of lung-derived MSCs to chondrogenic differentiation, representing an important contribution for the definition of the type of cell to be used in clinical trials of cell therapy and tissue engineering. == Introduction == Mesenchymal stem cells(MSCs) were first described as bone marrow cells capable of originating fibroblast colonies (colony-forming unit-fibroblasts [CFU-Fs], [1]). The enumeration of CFU-Fs from fresh tissue samples has been considered indicative of the frequency of MSCs, but a direct relationship between the 2 has not been clearly established [2]. In 1985, a relationship between these cells and the bone marrow stroma was proposed by Owen [3], who proposed the existence of stromal stem cells that are able to self-renew and generate BIX-01338 hydrate mature conjunctive/stromal cell types. The term MSCs was introduced by Caplan in 1991 [4] and is currently used for stem cells with an intrinsic potential to give rise to different mesenchymal cell types such as osteoblasts, chondrocytes, adipocytes, tenocytes, and others. The Mesenchymal and Tissue Stem Cell Committee of the International Society for Cell Therapy has established the minimal criteria to define human MSCs: the capacity to proliferate as adherent cells, a defined surface phenotype (positive for CD105, CD73, and CD90, and negative for CD45, CD34, CD14 or CD11b, CD79a or CD19, and HLA-DR), and the capacity to differentiate into osteoblasts, adipocytes, and chondroblasts [5]. Although conventionally isolated from the bone marrow, we and others have shown that MSCs are distributed throughout the whole organism, suggesting that they reside in association with blood vessels [6,7]. We have also suggested that the perivascular location of MSCs, associated to their remarkable immunoregulatory capacity, implies a role in the maintenance of tissue homeostasis: in the case of tissue injury, MSCs secrete a panel of cytokines and factors that control the immune response to avoid BIX-01338 hydrate an autoimmune process [8]. These studies have shown that, although similar in general characteristics, MSCs isolated from different tissues, such as brain, spleen, liver, kidney, lung, bone marrow, muscle, thymus, and pancreas, exhibit particular biological features, raising the question on whether they are identical cell populations or have important differences at the molecular level [9]. Cellular and molecular mechanisms underlying one of the fundamental properties of stem cells, self-renewal, have been the subject of many studies (reviewed in ref. [10]). While these studies provide an acceptable framework for defining MSCs at the molecular level, the presence of a large BIX-01338 hydrate number of housekeeping genes prevents proper evaluation of their specific genetic message [11]. Gene expression analyses have shown that the differentiation of MSCs into mature cell types is controlled temporally, and that the regulation of the process involves the activity of transcription factors, growth factors, and signaling pathways (reviewed in ref. [12]). Transcription factors, such as Oct3/4 [13,14], Sox2 [15], and Nanog [16], maintain the pluripotency of embryonic stem cells, and may also be expressed in MSCs [17]. Other genes regulate Mouse monoclonal to ERBB3 the differentiation of stem cells into specific lineages. For example, the lipoprotein lipase [18], enhancer-binding protein (C/EBP) [19,20], and adiponectin receptor 1 (ADIPOR1) [19] genes are expressed in the adipogenic differentiation pathway.COL2A1[21], collagen type X-alpha 1 (COL10A1) [22], and transient receptor potential cation channel, subfamily V, member 4 BIX-01338 hydrate (TRPV4) [23] are highly expressed in the chondrogenic differentiation pathway, while osteomodulin (OMD) [20], alkaline phosphatase (ALP) [24], and osteocalcin [25] regulate the osteogenic differentiation process. To better understand the relationship between MSCs residing in different tissues, we analyzed the expression of genes related to pluripotency and to adipogenic, osteogenic, and chondrogenic differentiation in cultures derived from endodermal (lung) and mesodermal (adipose) tissue maintained in different conditions. These variables were also analyzed in cultures isolated from the bone marrow, which represents the typical source of MSCs. These results are also important in the determination of the best source of stem cells to be used in therapeutic applications. == Materials and Methods == == Reagents, culture media, and solutions == Complete culture medium (CCM) was composed of Dulbecco’s modified Eagle’s medium with 10 mM HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid, free acid, 2.53.7g/L) and 10% fetal bovine serum.