Method of progenitor cell isolation from different organs by natural destruction of extracellular matrix
The present invention is directed to novel methods comprising the isolation of viable fetal progenitor cells from organs of aborted fetus. The invention comprises the use of natural proteolytic, collagenolytic and fibrinolytic activity of autologous abortive placenta tissue extract.
1. A method of isolating progenitor cells from different organs of an aborted fetus by using natural collagenolytic, fibrinolytic and proteolytic activity of an autologous abortive placenta, the method comprising:
preparing a collagenolytic, fibrinolytic, and proteolytic fetal placenta extract by selecting chorionic villi tissue from the abortive placenta, incubating the tissue with PGF2α for activation of proteolytic activity, homogenizing the tissue, incubating with trypsin for activation of collagenase activity followed by addition of trypsin inhibitor; and
isolating fetal progenitor cells from target organs of the aborted fetus by:
performing an in situ three open-loop perfusion process of a fetus blood vascular system of the aborted fetus, wherein the perfusion process comprises removing blood from the fetus vascular system;
lysing remaining blood cells in the fetus blood vascular system by perfusion with NH 4 Cl, KHCO 3 , and EDTA;
perfusing the target organs with solution containing the collagenolytic, fibrinolytic, and proteolytic fetal placenta extract; and
removing target organs and isolating cells by rupturing the organs, filtering, and pelleting cells to generate isolated fetal progenitor cells.
2. The method of claim 1 , wherein the aborted fetus is aborted at 18-20 weeks of gestation.
3. The method of claim 1 , wherein creating the solution comprises activating a proteolysis in placenta tissue by a trypsin short-term reaction which is stopped by a soybean trypsin inhibitor.
4. The method of claim 1 , wherein a first loop of perfusion in the in situ three open-loop perfusion process comprises providing a maternal-fetal blood supply via an umbilical vein.
5. The method of claim 1 , wherein a second loop of perfusion in the in situ three open-loop perfusion process comprises providing a blood supply route through a left ventricle of a heart.
6. The method of claim 1 , wherein a third loop of perfusion in the in situ three open-loop perfusion process comprises providing a blood supply route through an abdominal part of an aorta.
7. The method of claim 1 , wherein the three-open loop perfusion process begins in a femoral vein.
8. The method of claim 1 , wherein the target organs comprise any of a liver, intestines, lungs, a kidney, suprarenal glands, a heart, a thymus, and a spleen.