Procedure of CLI treatment with cell THERAPY (autologenous bone marrow mononuclear cells)
- The place of the surgery around the lumbar bone scapulae must be clean and disinfected, the operating room must be covered in sterile masks in compliance with good clinical praxis procedures. Vital signs monitoring including non-invasive measurement of blood pressure, heart rate and tcpO2 saturation must be taken during the procedure. The bone marrow is collected in intravenous sedation.
- The bone marrow is collected from both lumbar bone scapulae by standard technique of puncture-aspiration. Altogether 240 ml of bone marrow is collected in small aliquots (10-30 ml). Collected bone marrow is transferred into a bone marrow collecting bag containing anticoagulating solution and 2 ml non-fractionated heparin containing 2000 IU/ml.
- A gradient density centrifuge is used in the separation step. Its goal is to separate various parts of the bone marrow including white cells, platelets and red cells.
- In each processing dish allows to process 60 ml of aspirate fluid and to obtain 7-10 ml final concentrate that way. This is how 40 ml of concentrate can be produced for the transplantation.
METHOD DESCRIPTION – CELL THERAPY:
This method uses the differential potential of adult stem cells and their ability to create new vessels with neoangiogenesis process (neovascularization). The goal is to induce angiogenesis and vasculogenesis which would prevent from progression of ischemia and hypoxia, improve perfusion and substance exchange and that way stop the CLI progression towards the limb amputation. This is applied for example in treatments of wounds where the effect of a growth hormone from VGF family caused stimulation of pericytes, smooth muscle cells and endothelial cells with subsequent formation of a new vessel. Applicable stem cells can be obtained from fat tissue, bone marrow, umbilical cord blood, period blood, by tissue engineering, etc.
This method also operates with assumption of arteriogenesis – i.e. growth of collateral arteries – i.e. transformation of pre-existing collateral arteries into functional collateral arteries able to compensate for the loss of large occluded arteries. The original diameter of small, originally non-perfused, arteries can widen up to 20 times during the arteriogenesis process.
Arteriogenesis is mechanically induced by increased shear-pressure following a large artery occlusion. Proliferation in collateral endothelium is started by up-regulation of adhesion molecules and release of cytokines followed by attraction and perivascular migration of bone marrow derived monocytes.
Another proved mechanism is proliferation, migration and remodelation of vessel wall. Bone marrow monocyte cells excrete growth factors, induce proteases matrix and in later stage also factors which stabilize vessels. At sufficient quantity of bone marrow cells participating on arteriogenesis in perivascular space, circa 6-8 weeks in needed to grow collateral artery net. When used in primary transplantation of bone marrow core cells concentrate, the Harvest BMAC technique imitates very accurately physiological repair mechanisms which occur during ischemia.