Method for the in vitro or ex vivo amplification of human adipose tissue stem cells
AssigneesUniversité CôTe D'Azur, Institut National de la Sante et de la Recherche Medicale, Centre National de la Recherche Scientifique (CNRS), Chu De Nice
InventorsChristian Dani, Alain Doglio, Vincent Dani-Davesne, Philippe Letertre
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Abstract
The method for in vitro or ex vivo amplification of human adipose tissue stem cells includes: —extracting a stromal vascular fraction of a human adipose tissue including endothelial cells of the human adipose tissue vascular network and human adipose tissue stem cells, and an extracellular matrix of the human adipose tissue, the extracellular matrix including endothelial cells of the human adipose tissue vascular network, human adipose tissue stem cells and collagen; —mixing the stromal vascular fraction and the extracellular matrix; and —culturing the mixture obtained in the preceding step, in suspension, in a culture medium.
Claims10
A method for in vitro or ex vivo amplification of human adipose tissue stem cells, comprising:
- (1) providing a human adipose tissue comprising:
- an extracellular matrix comprising collagen,
- a vascular network comprising endothelial cells, and
- stem cells;
- (2) performing extraction, wherein the extraction comprises:
- (2a) performing centrifugation of the human adipose tissue to obtain at least two distinct fractions, the at least two distinct fractions including:
- a fraction A comprising a centrifuged extracellular matrix fraction, the centrifuged extracellular matrix fraction comprising the extracellular matrix comprising the collagen, and
- a fraction B comprising a stromal vascular fraction, wherein the stromal vascular fraction comprises cells expressing DPP4, wherein the cells expressing DPP4 in the fraction B constitute a majority of the cells expressing DPP4 initially contained in the provided human adipose tissue,
- wherein the centrifuged extracellular matrix fraction comprises a first portion of the endothelial cells, and the stromal vascular fraction comprises a second portion of the endothelial cells different from the first portion of the endothelial cells,
- wherein the centrifuged extracellular matrix fraction comprises a first portion of the stem cells, and the stromal vascular fraction comprises a second portion of the stem cells different from the first portion of the stem cells;
- and
- (2b) separating the fraction A from the fraction B, and either
- (i) performing mechanical dissociation of the fraction A, so as to obtain a fraction A′ comprising the extracellular matrix comprising the collagen,
- wherein the fraction A′ comprises the first portion of the endothelial cells and the first portion of the stem cells, mechanically dissociated from the extracellular matrix;
- or
- (ii) performing mechanical dissociation of the fraction A, so as to obtain the fraction A′ comprising the extracellular matrix comprising the collagen, wherein the fraction A′ comprises the first portion of the endothelial cells and the first portion of the stem cells, mechanically dissociated from the extracellular matrix, and further performing centrifugation of the fraction A′ to obtain at least two distinct fractions, the at least two distinct fractions including a fraction A″ comprising the extracellular matrix comprising the collagen, and a fraction B′ comprising the first portion of the endothelial cells and the first portion of the stem cells,
- (3) mixing the fraction B with either
- (i) the fraction A′ obtained by the mechanical dissociation of the fraction A;
- or
- (ii) the fraction B′ and the fraction A″ obtained by the mechanical dissociation of the fraction A and further centrifugation of the fraction A′,
- to obtain a mixture;
- and
- (4) culturing the mixture obtained from the mixing, in suspension, in a culture medium.
Dependencies are inferred from the claim text.