Adipose-derived stem cells and lattices

Abstract

The present invention provides adipose-derived stem cells and lattices. In one aspect, the present invention provides a lipo-derived stem cell substantially free of adipocytes and red blood cells and clonal populations of connective tissue stem cells. The cells can be employed, alone or within biologically-compatible compositions, to generate differentiated tissues and structures, both in vivo and in vitro. Additionally, the cells can be expanded and cultured to produce hormones and to provide conditioned culture media for supporting the growth and expansion of other cell populations. In another aspect, the present invention provides a lipo-derived lattice substantially devoid of cells, which includes extracellular matrix material from adipose tissue. The lattice can be used as a substrate to facilitate the growth and differentiation of cells, whether in vivo or in vitro, into anlagen or even mature tissues or structures.

1 . A mammalian lipo-derived stem cell substantially free of mature adipocytes. 
   
   
       2 . The cell of  claim 1 , which can be cultured in DMEM+about 10% fetal bovine serum for at least 15 passages without differentiating. 
   
   
       3 . The cell of  claim 2 , which has two or more developmental phenotypes selected from the group of developmental phenotypes consisting of adipogenic, chondrogenic, cardiogenic, dermatogenic, hematopoetic, hemangiogenic, myogenic, nephrogenic, neurogenic, neuralgiagenic, urogenitogenic, osteogenic, pericardiogenic, peritoneogenic, pleurogenic, splanchogenic, and stromal developmental phenotypes. 
   
   
       4 . The cell of any of  claims 1 - 3 , which is human. 
   
   
       5 . The cell of any of  claims 1 - 4 , which is genetically modified. 
   
   
       6 . The cell of any of  claims 1 - 5 , which has a cell-surface bound intercellular signaling moiety. 
   
   
       7 . The cell of any of  claims 1 - 5 , which secretes a hormone. 
   
   
       8 . The cell of  claim 7 , wherein the hormone is selected from the group of hormones consisting of cytokines and growth factors. 
   
   
       9 . A defined cell population comprising a cell of any of  claims 1 - 8 . 
   
   
       10 . The defined cell population of  claim 9 , which is heterogeneous. 
   
   
       11 . The defined cell population of  claim 9  or  10 , further compressing a stem cell selected from the group of cells consisting of neural stem cells (NSC), hematopoetic stem cells (HPC), embryonic stem cells (ESC) and mixtures thereof. 
   
   
       12 . The defined cell population of  claim 9 , which consists essentially of cells according to any of  claims 1 - 8 . 
   
   
       13 . The defined cell population of  claim 9  or  12 , which is substantially homogenous. 
   
   
       14 . The defined cell population of  claim 13 , which is clonal. 
   
   
       15 . A defined cell population consisting essentially of mesodermal stem cells (MHC), connective tissue stem cell (CTSC), or mixtures thereof, wherein the population is clonal. 
   
   
       16 . The population of  claim 15 , wherein the stem cells have two or more developmental phenotypes selected from the group of developmental phenotypes consisting of adipogenic, chondrogenic, cardiogenic, dermatogenic, hematopoetic, hemangiogenic, myogenic, nephrogenic, neurogenic, neuralgiagenic, urogenitogenic, osteogenic, pericardiogenic, peritoneogenic, pleurogenic, splanchogenic, and stromal developmental phenotypes. 
   
   
       17 . A lipo-derived lattice comprising adipose tissue extracellular matrix matter substantially devoid of cells. 
   
   
       18 . The lipo-derived lattice of  claim 17 , comprising a human protein, proteoglycan, glycoprotein, hyaluronin, or fibronectin molecule. 
   
   
       19 . The lipo-derived lattice of  claim 17  or  18 , comprising a collagen selected from the group of collagens consisting of type I, type II, type III, type IV, type V, type VI collagen. 
   
   
       20 . The lipo-derived lattice of any of  claims 17 - 19 , comprising a hormone. 
   
   
       21 . The lipo-derived lattice of  claim 20 , wherein the hormone is selected from the group of hormones consisting of cytokines and growth factors. 
   
   
       22 . The lipo-derived lattice of any of  claims 17 - 21 , which is substantially anhydrous. 
   
   
       23 . The lipo-derived lattice of any of  claims 17 - 22 , which is lyophilized. 
   
   
       24 . The lipo-derived lattice of any of  claims 17 - 21 , which is hydrated. 
   
   
       25 . A kit comprising the lipo-derived lattice of any of  claims 17 - 24  and one or more components selected from the group of components consisting of hydrating agents, cell culture substrates, cell culture media, antibiotic compounds, and hormones. 
   
   
       26 . A composition comprising a cell and the lipo-derived lattice of any of  claims 17 - 24 . 
   
   
       27 . A composition comprising the cell of any of  claims 1 - 8  and a biologically compatible lattice. 
   
   
       28 . A composition comprising the population of any of  claims 9 - 16  and a biologically compatible lattice. 
   
   
       29 . The composition of  claim 27  or  28 , wherein the lattice comprises polymeric material. 
   
   
       30 . The composition of  claim 29 , wherein the polymeric material is formed of polymer fibers as a mesh or sponge. 
   
   
       31 . The composition of  claim 29  or  30 , wherein the polymeric material comprises monomers selected from the group of monomers consisting of glycolic acid, lactic acid, propyl fumarate, caprolactone, hyaluronan, hyaluronic acid and combinations thereof. 
   
   
       32 . The composition of any of  claims 29 - 31 , wherein the polymeric material comprises proteins, polysaccharides, polyhydroxy acids, polyorthoesters, polyanhydrides, polyphosphazenes, synthetic polymers or combinations thereof. 
   
   
       33 . The composition of any of  claims 29 - 32 , wherein the polymeric material is a hydrogel formed by crosslinking of a polymer suspension having the cells dispersed therein. 
   
   
       34 . The composition of any of  claims 29 - 33 , wherein the lattice further comprises a hormone selected from the group of hormones consisting of cytokines and growth factors. 
   
   
       35 . The composition of any of  claims 29 - 34 , wherein the lattice is the lipo-derived lattice of any of  claims 17 - 24 . 
   
   
       36 . A method of obtaining a genetically-modified cell comprising exposing the cell of any of  claims 1 - 8  to a gene transfer vector comprising a nucleic acid including a transgene, whereby the nucleic acid is introduced into the cell under conditions whereby the transgene is expressed within the cell. 
   
   
       37 . The method of  claim 36 , wherein the transgene encodes a protein conferring resistance to a toxin. 
   
   
       38 . A method of delivering a transgene to an animal comprising (a) obtaining a genetically-modified cell in accordance with  claim 36  or  37  and (b) introducing the cell into the animal, such that the transgene is expressed in vivo. 
   
   
       39 . A method of differentiating the cell of any of  claims 1 - 8  comprising culturing the cell in a morphogenic medium under conditions sufficient for the cell to differentiate. 
   
   
       40 . The method of  claim 39 , wherein the medium is an adipogenic, chondrogenic, cardiogenic, dermatogenic, embryonic, fetal, hematopoetic, hemangiogenic, myogenic, nephrogenic, neurogenic, neuralgiagenic, urogenitogenic, osteogenic, pericardiogenic, peritoneogenic, pleurogenic, and splanchogenic, or stromogenic media. 
   
   
       41 . The method of  claim 39  or  40 , wherein the morphogenic medium is an adipogenic medium and the cell is monitored to identify adipogenic differentiation. 
   
   
       42 . The method of  claim 39  or  40 , wherein the morphogenic medium is a chondrogenic medium and the cell is monitored to identify chondrogenic differentiation. 
   
   
       43 . The method of  claim 39  or  40 , wherein the morphogenic medium is an embryonic or fetal medium and the cell is monitored to identify embryonic or fetal phenotype. 
   
   
       44 . The method of  claim 39  or  40 , wherein the morphogenic medium is a myogenic medium and the cell is monitored to identify myogenic differentiation. 
   
   
       45 . The method of  claim 39  or  40 , wherein the morphogenic medium is an osteogenic medium and the cell is monitored to identify osteogenic differentiation. 
   
   
       46 . The method of  claim 39  or  40 , wherein the morphogenic medium is a stromal medium and the cell is monitored to identify stromal or hematopoetic differentiation. 
   
   
       47 . The method of any of  claims 39 - 46 , wherein the cell differentiates in vitro. 
   
   
       48 . The method of any of  claims 39 - 46 , wherein the cell differentiates in vivo. 
   
   
       49 . A method of producing hormones, comprising (a) culturing the cell of  claim 7  or  8  within a medium under conditions sufficient for the cell to secrete the hormone into the medium and (b) isolating the hormone from the medium. 
   
   
       50 . A method of promoting the closure of a wound within a patient comprising introducing the cell of  claim 7  or  8  into the vicinity of a wound under conditions sufficient for the cell to produce the hormone, whereby the presence of the hormone promotes closure of the wound. 
   
   
       51 . A method of promoting neovascularization within tissue, comprising introducing the cell of  claim 7  or  8  into the tissue under conditions sufficient for the cell to produce the hormone, whereby the presence of the hormone promotes neovascularization within the tissue. 
   
   
       52 . The method of  claim 51 , wherein the tissue is within an animal. 
   
   
       53 . The method of  claim 51  or  52 , wherein the tissue is a graft. 
   
   
       54 . The method of any of  claims 49 - 53 , wherein the hormone is a growth factor selected from the group of growth factor consisting of human growth factor, nerve growth factor, vascular and endothelial cell growth factor, and members of the TGFβ superfamily. 
   
   
       55 . A method of conditioning culture medium comprising exposing a cell culture medium to the cell of any of  claims 1 - 7  under conditions sufficient for the cell to condition the medium. 
   
   
       56 . The method of  claim 55 , wherein the medium is separated from the cell after it has been conditioned. 
   
   
       57 . The method of any of  claims 36 - 56 , wherein the cell is within a population of any of  claims 9 - 16 . 
   
   
       58 . A conditioned culture medium produced in accordance with the method of  claim 55  or  56 . 
   
   
       59 . The conditioned culture medium of  claim 58 , which is substantially free of a cell of any of  claims 1 - 7 . 
   
   
       60 . A method of culturing a stem cell comprising maintaining a stem cell in the conditioned medium of  claim 58  or  59  under conditions for the stem cell to remain viable. 
   
   
       61 . The method of  claim 60 , which further comprises permitting successive rounds of mitotic division of the stem cell to form an expanded population of stem cells. 
   
   
       62 . The method of  claim 60  or  61 , wherein the medium is substantially free of the lipo-derived cells of any of  claims 1 - 7 . 
   
   
       63 . The method of any of  claims 60 - 62 , wherein the medium contains lipo-derived cells of any of  claims 1 - 7 . 
   
   
       64 . The method of  claim 63 , wherein a stem cell and a lipo-derived cell are in contact. 
   
   
       65 . The method of any of  claims 60 - 64 , wherein a stem cell is a hemopoietic stem cell. 
   
   
       66 . A method of producing animal matter comprising maintaining the composition of any of  claims 18 - 26  under conditions sufficient for the cells within the composition to expand and differentiate to form the matter. 
   
   
       67 . The method of  claim 66 , wherein the matter comprises a tissue type selected from the group of tissues consisting of adipose, cartilage, heart, dermal connective tissue, blood tissue, muscle, kidney, bone, pleural, and splancnic tissues, and combinations thereof. 
   
   
       68 . The method of  claim 66  or  67 , wherein the matter comprises more than one tissue type. 
   
   
       69 . The method of any of  claims 66 - 68 , wherein the matter comprises at least a portion of an animal organ. 
   
   
       70 . The method of  claim 66 - 68 , wherein the matter comprises at least a portion of an animal limb. 
   
   
       71 . The method of any of  claims 66 - 70 , wherein the composition is maintained in vitro. 
   
   
       72 . The method of any of  claims 66 - 70 , wherein the composition is introduced into an animal and maintained in vivo. 
   
   
       73 . An implant comprising the cell of any of  claims 1 - 7 . 
   
   
       74 . An implant comprising the population of any of  8 - 13 . 
   
   
       75 . An implant comprising the lipo-derived lattice of any of  claims 14 - 16 . 
   
   
       76 . An implant comprising the composition of any of  claims 17 - 26 . 
   
   
       77 . A kit for isolating stem cells from adipose tissues comprising a means for isolating adipose tissue from a patient and a means for separating stem cells from the remainder of the adipose tissue. 
   
   
       78 . The kit of  claim 77 , further comprising a medium for differentiating the stem cells. 
   
   
       79 . The kit of  claim 78 , wherein the medium is selected from the group of media consisting of adipogenic, chondrogenic, cardiogenic, dermatogenic, embryonic, fetal, hematopoetic, hemangiogenic, myogenic, nephrogenic, neurogenic, neuralgiagenic, urogenitogenic, osteogenic, pericardiogenic, peritoneogenic, pleurogenic, and splanchogenic, and stromogenic media.