- 2014-12-01 (x)
- 1922-12-30 (x)
- Haynesworth, Stephen E. (x)
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Show moreThe present invention provides methods for directing mesenchymal stem cells cultivated in vitro to differentiate into specific cell lineage pathways prior to, or at the time of, their implantation for the therapeutic treatment of pathologic conditions in humans and other species. Mesenchymal stem cells (MSCs) are the formative pluripotent blast or embryonic-like cells found in bone marrow, blood, dermis, and periosteum that are capable of differentiating into specific types of mesenchymal or connective tissues including adipose, osseous, cartilaginous, elastic, muscular, and fibrous connective tissues. The specific differentiation pathway which these cells enter depends upon various influences from mechanical influences and/or endogenous bioactive factors, such as growth factors, cytokines, and/or local microenvironmental conditions established by host tissues. Although these cells are normally present at very low frequencies in bone marrow, a process for isolating, purifying, and mitotically expanding the population of these cells in tissue culture is reported in Caplan et al. U.S. Pat. Nos. 5,197,985 and 5,226,914. In prenatal organisms, the differentiation of MSCs into specialized connective tissue cells is well established; for example embryonic chick, mouse or human limb bud mesenchymal cells differentiate into cartilage, bone and other connective tissues (1-5). In addition, a clonal rat fetus calvarial cell line has also been shown to differentiate into muscle, fat, cartilage, and bone (6). The existence of MSCs in post-natal organisms has not been widely studied with the objective of showing the differentiation of post-embryonic cells into several mesodermal phenotypes. The few studies which have been done involve the formation of bone and cartilage by bone marrow cells following their encasement in diffusion chambers and in vivo transplantation (7, 8). Recently, bone marrow-derived cells from young rabbits (800-1,000 g) have been shown to form adipocytic and osteogenic cells in vivo (9) and cloned bone marrow stromal cells of post-natal mice were shown to form adipocytes and osteogenic cells (10). Likewise, cells from chick periosteum have been isolated, expanded in culture, and, under high density conditions in vitro, shown to differentiate into cartilage and bone (11). Rat bone marrow-derived mesenchymal cells have been shown to have the capacity to differentiate into osteoblasts and chondrocytes when implanted in vivo (12, 6). Cells from various marrow sources of post-natal organisms have never been observed to exhibit myogenic properties, with multinuclear appearance being the most easily recognized characteristic in culture.
http://www.google.com/patents?vid=USPAT5942225
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Show moreThe present invention provides methods for directing mesenchymal stem cells cultivated in vitro to differentiate into specific cell lineage pathways prior to, or at the time of, their implantation for the therapeutic treatment of pathologic conditions in humans and other species. Mesenchymal stem cells (MSCs) are the formative pluripotent blast or embryonic-like cells found in bone marrow, blood, dermis, and periosteum that are capable of differentiating into specific types of mesenchymal or connective tissues including adipose, osseous, cartilaginous, elastic, muscular, and fibrous connective tissues. The specific differentiation pathway which these cells enter depends upon various influences from mechanical influences and/or endogenous bioactive factors, such as growth factors, cytokines, and/or local microenvironmental conditions established by host tissues. Although these cells are normally present at very low frequencies in bone marrow, a process for isolating, purifying, and mitotically expanding the population of these cells in tissue culture is reported in Caplan et al. U.S. Pat. Nos. 5,197,985 and 5,226,914. In prenatal organisms, the differentiation of MSCs into specialized connective tissue cells is well established; for example embryonic chick, mouse or human limb bud mesenchymal cells differentiate into cartilage, bone and other connective tissues (1-5). In addition, a clonal rat fetus calvarial cell line has also been shown to differentiate into muscle, fat, cartilage, and bone (6). The existence of MSCs in post-natal organisms has not been widely studied with the objective of showing the differentiation of post-embryonic cells into several mesodermal phenotypes. The few studies which have been done involve the formation of bone and cartilage by bone marrow cells following their encasement in diffusion chambers and in vivo transplantation (7, 8). Recently, bone marrow-derived cells from young rabbits (800-1,000 g) have been shown to form adipocytic and osteogenic cells in vivo (9) and cloned bone marrow stromal cells of post-natal mice were shown to form adipocytes and osteogenic cells (10). Likewise, cells from chick periosteum have been isolated, expanded in culture, and, under high density conditions in vitro, shown to differentiate into cartilage and bone (11). Rat bone marrow-derived mesenchymal cells have been shown to have the capacity to differentiate into osteoblasts and chondrocytes when implanted in vivo (12, 6). Cells from various marrow sources of post-natal organisms have never been observed to exhibit myogenic properties, with multinuclear appearance being the most easily recognized characteristic in culture.
http://www.google.com/patents?vid=USPAT5736396
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Show moreThe present invention is directed to isolated and purified human mesenchymal stem cells, to a method for isolating, purifying, and culturally expanding human mesenchymal stem cells (i.e. mesenchymal stem cells or "MSCs"), and to characterization of and uses for such cells. Further, the present invention is directed to monoclonal antibodies that are specific for human mesenchymal stem cells (i.e. mesenchymal stem cells). In addition, the invention is directed to the monoclonal hybridoma cell lines that produce such antibodies, and the use of the monoclonal antibodies for diagnostic and/or therapeutic purposes. The present invention is also directed to various methods and devices for enhancing the implantation and differentiation of mesenchymal stem cells. The present invention is also directed to various methods and devices for treating skeletal and other connective tissue disorders. The methods and devices of the invention utilize isolated mesenchymal progenitor cells which, under certain conditions, can be induced to differentiate into different types of desired connective tissue, such as into bone or cartilage forming cells. Mesenchymal stem cells are the formative pluripotential blast cells found inter alia in bone marrow, blood, dermis and periosteum that are capable of differentiating into any of the specific types of mesenchymal or connective tissues (i.e. the tissues of the body that support the specialized elements; particularly adipose, osseous., cartilaginous, elastic, and fibrous connective tissues) depending upon various influences from bioactive factors, such as cytokines. Although these cells are normally present at very low frequencies in bone marrow, the inventors of the present invention have discovered a process for isolating, purifying, and greatly replicating these cells in culture, i.e. in vitro. In one aspect, the present invention is directed to human mesenchymal stem cells isolated from a tissue specimen and the method of their isolation. Homogeneous human mesenchymal stem cell compositions are provided which serve as the progenitors for all mesenchymal cell lineages. MSCs are identified by specific cell surface markers which are identified with unique monoclonal antibodies. The homogeneous MSC compositions are obtained by positive selection of adherent marrow or periosteal cells which are free of markers associated with either hematopoietic cell or differentiated mesenchymal cells. These isolated mesenchymal cell populations display epitopic characteristics associated with only mesenchymal stem cells, have the ability to regenerate in culture without differentiating, and have the ability to differentiate into specific mesenchymal lineages when either induced in vitro or placed in vivo at the site of damaged tissue. In order to obtain subject human mesenchymal stem cells, it is necessary to isolate rare pluripotent mesenchymal stem cells from other cells in the bone marrow or other MSC source.
http://www.google.com/patents?vid=USPAT6087113
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Show moreThe present invention relates to the field of monoclonal antibodies, particularly monoclonal antibodies that are specific for subsets of human osteogenic cells, hybridoma cell lines that synthesize and secrete these antibodies, and the uses of the monoclonal antibodies. Osteogenic cells are responsible for synthesizing and maintaining the extracellular matrix of both embryonic and adult bone. This family, or lineage, of cells arises from undifferentiated mesenchymal stem cells (MSCs) and progresses through a series of distinct maturational stages defined, in part, by the sequential acquisition and/or loss of specific cell surface antigens (Bruder, S. P. and Caplan, A. I., Dev. Biol., 141:319-329,1990). Although monoclonal antibodies against cell surface antigens on normal cells of the osteogenic lineage have been reported for avian and rodent species (Bruder, S. P. and Caplan, A. I., Bone, 10:359-375, 1989; Bruder, S. P. and Caplan, A. I., Bone, 11:189-198, 1990; Turksen, K. et al., J Histochem Cytochem, 40:1339-1352, 1992), none have been reported for human cells. Some of these antibodies reported also react with cells other than those found in bone. While monoclonal antibodies have been raised against intracellular antigens in normal human osteoblasts and against the surface of transformed human osteogenic cell lines, none have recognized the surface of normal human osteoblasts (Embleton, M. J. et al., Br J Cancer, 43:582-587, 1981; Hosoi, S. et al., Cancer Res, 42:654-661, 1982; Heiner, J. et al., Cancer Res, 47:5377-5384, 1987; Bruland, O. et al., Cancer Res, 48:5302-5308, 1988; Tsai, C. et al., Cancer Res, 50:152-161, 1990; Walsh, S. et al., J Bone Miner Res, 9:1687-1696, 1994). At present, the only monoclonal antibody against a cell surface epitope capable of identifying human osteogenic cells is one directed against alkaline phosphatase (Lawson, G. et al., Clin Chem, 31:381-385, 1985). This well-characterized cell surface enzyme has served as the historical standard for identifying a large family of osteogenic cells, and is readily demonstrated by a simple histochemical stain. A common thread highlighted in these studies is the fact that a monoclonal antibody may also react with non-osteogenic cells while remaining a useful marker of osteogenic differentiation. So long as the monoclonal antibody is selective for cells at discrete stages of differentiation, the probe is both novel and useful. In an experimental sense, osteogenic cells have been shown to arise from purified human mesenchymal stem cells in an in vivo setting (Haynesworth, S. E. et al., Bone, 13:81-88, 1992). Mesenchymal stem cells are the formative multipotential blast cells found inter alia in bone marrow, blood, dermis and periosteum that are capable of differentiating into any of the specific types of mesenchymal or connective tissues (i.e. the tissues of the body that support the specialized elements.
http://www.google.com/patents?vid=USPAT5643736
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Show moreThe present invention is directed to a totally unexplored application of human stem cells, i.e. that of genetically engineered cell that carry within them genes of interest particularly for the expression of physiologically or pharmacologically active proteins or for use in gene therapy. In accordance with the present invention it has been discovered that human mesenchymal stem cells (MSCs) or human mesenchymal progenitor cells can be used as host cells for the expression of exogenous gene products. One aspect of the invention relates to the discovery and development of the technology to isolate these cells, mitotically proliferate them in cell culture and introduce them back in vivo into the same recipient. These culture-expanded cells home back to the marrow and enhance hematopoietic recovery in the marrow transplant setting. Furthermore, these cells can be manipulated for cellular therapy, e.g. expanded, purified, selected and maintained for clinical use while still mantaining their precursor phenotype. Part of this manipulation is the characterization of such cells and their cryopreservation for future use. Mesenchymal stem cells (MSCs) can be derived from marrow, periosteum, dermis and other tissues of mesodermal origin. They are the formative pluripotential blast cells that differentiate into the specific types of connective tissues (i.e. the tissues of the body that support the specialized elements; particularly adipose, areolar, osseous, cartilaginous, elastic, marrow stroma, muscle, and fibrous connective tissues) depending upon various in vivo or in vitro environmental influences. Although these cells are normally present at very low frequencies in bone marrow, the inventors of the present invention have discovered a process for isolating, purifying, and greatly replicating the marrow-derived mesenchymal stems cells in culture, i.e. in vitro. This discovery is the subject of U.S. patents and co-pending applications, for example, Caplan and Haynesworth, U.S. Pat. Nos. 5,197,985 and 5,226,914 and PCT Publication No. WO 92/22584 (published 23 Dec. 1992) as well as numerous literature references by Caplan and Haynesworth. Isolated human hematopoietic stem cells have also been described, for example, in Tsuksamoto et al., U.S. Pat. No. 5,061,620 (October 1991) and reviewed in Edgington, Biotechnology, 10:1099-1106 (1992) and the references cited therein. These are distinguished from MSCs by their ability to differentiate into myeloid and lymphoid blood cells. In its principal embodiment the invention relates to isolated human mesenchymal stem cells capable of expressing incorporated genetic material of interest. Human stem cells are obtained from the individual donor and rendered substantially isolated from other cells and constituitive donor proteins and other components.
http://www.google.com/patents?vid=USPAT5591625
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