Methods and compositions to facilitate repair of avascular tissue
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Publication Number
US-11278573-B2
Publication Date
2022-03-22
Expiration Date
Abstract
Compositions and methods are provided for repairing damaged avascular zones, including intervertebral disc, in a patient in need thereof.
Core Innovation
The invention relates to methods and compositions for treating an avascular zone in a patient in need thereof by isolating and expanding viable mesenchymal stem cells from whole marrow-derived nucleated cells. The isolation and expansion process includes obtaining whole marrow from the patient, centrifuging to separate nucleated cells from red blood cells, washing and counting the nucleated cells, and culturing the nucleated cells under selective pressure of about 1% to about 10% oxygen, or under oxygen conditions decreasing from about 10% oxygen to about 1% oxygen or to about 5% oxygen, for 1-28 days.
The culture medium is maintained at a pH from 6.6 to 6.9 and comprises 10% to 20% human platelet lysate, followed by selecting viable mesenchymal stem cells capable of growth under the selective oxygen pressure. The method further includes providing the selected viable mesenchymal stem cells for implantation in the avascular zone and implanting platelet cell lysate or platelets prior to, during, or after implanting the selected viable mesenchymal stem cells, with the implanting occurring in the avascular zone.
In described embodiments, the implantation target is in a degenerative intervertebral disc and can be narrowed to the posterior disc annulus. The document also describes augmenting the implantation with additional biological supplements administered epidurally and/or in combination with the implanted cells and platelet lysate or platelets, including growth factors, cytokines, integrins, and cadherins.
Claims Coverage
The document includes three independent method claims that share a common core: isolating and expanding whole-marrow-derived nucleated cells into viable mesenchymal stem cells under defined selective oxygen conditions with a specified culture medium composition, and implanting the resulting cells in the avascular zone together with platelet cell lysate or platelets before, during, or after stem cell implantation.
Low-oxygen selection of marrow-derived mesenchymal stem cells with platelet lysate
Culturing nucleated cells in a culture medium under selective pressure of about 1% to about 10% oxygen for 1-28 days, wherein the culture medium is maintained at a pH from 6.6 to 6.9 and comprises 10% to 20% human platelet lysate; selecting viable mesenchymal stem cells capable of growth in the culture medium under the selective pressure of about 1% to about 10% oxygen.
Sequential oxygen reduction for selection of viable mesenchymal stem cells with platelet lysate
Culturing nucleated cells in a culture medium under a selective pressure of about 10% oxygen and then culturing under decreasing levels of oxygen until about 1% oxygen, wherein the culturing from about 10% oxygen to about 1% oxygen is for 1-28 days, wherein the culture medium is maintained at a pH from 6.6 to 6.9 and comprises 10% to 20% human platelet lysate; selecting viable mesenchymal stem cells capable of growth in the culture medium under the selective pressure of about 10% to about 1% oxygen.
Oxygen-controlled expansion to about 5% oxygen for selection with platelet lysate
Culturing nucleated cells in a culture medium under a selective pressure of about 10% oxygen and then culturing under decreasing levels of oxygen until about 5% oxygen, wherein the culturing from about 10% oxygen to about 5% oxygen is for 1-28 days, wherein the culture medium is maintained at a pH from 6.6 to 6.9 and comprises 10% to 20% human platelet lysate; selecting viable mesenchymal stem cells capable of growth in the culture medium under the selective pressure of about 10% to about 5% oxygen.
Implantation of mesenchymal stem cells in an avascular zone with platelet cell lysate or platelets
Providing the selected, viable mesenchymal stem cells for implantation in the avascular zone; implanting platelet cell lysate or platelets prior to, during, or after implanting the selected, viable mesenchymal stem cells; wherein the implanting of the platelet cell lysate or platelets and the selected, viable mesenchymal stem cells is in the avascular zone.
Across the independent claims, the inventive approach is to expand marrow-derived nucleated cells into viable mesenchymal stem cells under selective low-oxygen conditions using a culture medium with pH 6.6 to 6.9 and 10% to 20% human platelet lysate, and then implant the selected viable mesenchymal stem cells into the avascular zone with platelet cell lysate or platelets applied before, during, or after the stem cell implantation.
Stated Advantages
The overall concept links selective low-oxygen expansion with platelet lysate or platelet co-implantation in and around the avascular zone to improve repair outcomes.
Example clinical workflow and outcomes are described for three treated subjects, reporting reduction in lumbar disc protrusion size and improved pain/function in two of three cases.
The described approach uses hostile avascular-like conditions to support viability and growth for subsequent implantation in the damaged avascular region.
Documented Applications
Treatment of an avascular zone in a patient in need thereof.
Implantation in a degenerative intervertebral disc.
Implantation narrowed to the posterior disc annulus.
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