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Adipose derived adult stromal cells exhibiting characteristics of endothelial cells

A technology of endothelial cells, cells, applied in the field of composition of adult stromal cells

Inactive Publication Date: 2008-04-02
COGNATE THERAPEUTICS INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Current methods for culturing and obtaining large numbers of EPCs have not been successful

Method used

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  • Adipose derived adult stromal cells exhibiting characteristics of endothelial cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0172] Example 1: Establishment of primary ADAS cultures

[0173] Stromal vascular fraction (SVF) from white adipose tissue obtained by liposuction was digested with Krebs-Ringer bicarbonate buffer containing 0.5% BSA and 125 g / mL type I collagenase (final concentration) at 37 °C 80 minutes, shaking vigorously in 10-minute intervals. After digestion, the suspension was centrifuged at 1,200 rpm for 5 minutes at room temperature, vigorously shaken and then centrifuged again at 1,200 rpm for 5 minutes at room temperature. Aspirate the lipid / adipocyte layer and discard without disturbing the SVF pellet. The pellet was resuspended / washed with stromal cell medium (DMEM / F12 1:1, 10% FBS, IX antibiotic / antimycotic) and resuspended in a total volume of 40 mL of stromal cell medium. 10 mL of this suspension was added to a T-225 flask containing 40 mL of stromal medium. Non-adherent cells were washed out 1 to 3 days after seeding, and the medium was replaced with stromal cell medium...

Embodiment 2

[0174] Example 2: Treatment of ADAS cells

[0175] In two separate experiments, ADAS (P0) cells from a single donor were split at approximately 6 × 10 3 cells / cm 2 The density was inoculated in T-83 flasks (1 generation). Control ADAS cells were cultured in expansion medium containing DMEM / F12 (1:1), 10% FBS, 5 ng / mL hEGF and 1 ng / mL hFGF, and the medium was changed every 2 to 4 days. ADAS cells in the treatment group were subjected to a step-wise treatment protocol, which started 6 days in MII medium (DMEM / F12, N2 supplement, B27 supplement, 2.3 mM glutamine, 10 ng / mL hFGF), after inoculation The medium was changed on days 1, 3 and 5. On day 7, the culture was rinsed twice with D-PBS, and then the ADAS cells were incubated in MIII medium (DMEM / F12, N2 supplement (Invitrogen, Carlsbad, CA), B27 supplement (Invitrogen, Carlsbad, CA) , 2.3mM glutamine, 10mM nicotinamide, 2% FBS) for 4 days. MIII was replaced on day 10 and control and treated ADAS cells were harvested by t...

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Abstract

The present invention encompasses an adipose-derived adult stromal (ADAS) cell exhibiting at least one characteristic of a pre-endothelial cell and / or an endothelial cell. The present invention also encompasses compositions and methods for generating an adipose-derived adult stromal to exhibit at least one characteristic of a pre-endothelial cell and / or an endothelial cell. Methods for using the cells in vascular transplantation, tissue engineering, regulation of angiogenesis, vasculogenesis, and the treatment of numerous disorders including heart disease are also included.

Description

Background of the invention [0001] The developing neonatal period in humans is characterized by the presence of "stem" cells that have the potential to develop along multiple differentiation pathways. The final differentiation of these cells is determined by cytokine and hormonal signals that coordinate organogenesis and tissue architecture. Embryonic stem (ES) cells from mice have been isolated and extensively studied in vitro and in vivo. Using exogenous in vitro stimuli, researchers have induced ES cells to differentiate along multiple lineage pathways. These pathways include neuronal, B-lineage lymphoid, and adipocyte pathways (Dani, et al., 1997, J. Cell Sci. 110:1279; Remoncourt, et al., 1998, Mech.Dev.79:185; O 'Shea, 1999, Anat. Rec. 257:32). [0002] Pluripotent stem cells are also present in the tissues of adult organisms. The most well-characterized examples of adult stem cells are hematopoietic progenitor cells isolated from bone marrow and peripheral blood. L...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/02C12N5/08C12N5/00A61K35/12C12N5/071
CPCC12N5/0667C12N2501/115C12N2501/11C12N5/069C12N2506/1384C12N5/0652
Inventor 詹姆斯·K·亨德里克斯詹姆斯·B·米切尔二世
Owner COGNATE THERAPEUTICS INC
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