Stem cell conditioned media and methods of producing the same

Abstract

Stem cell conditioned media and methods of producing the same to treat mammalian injuries or insults. In at least one embodiment of a method of producing a stem cell conditioned media of the present disclosure, the method comprises the steps of culturing at least one stem cell in a first cell culture medium, replacing some or all of the first cell culture medium with a second cell culture medium and further culturing the at least one stem cell in the second cell culture medium, and collecting a quantity of the second cell culture medium after a culture duration, wherein the quantity of the second cell culture medium contains a cell culture byproduct effective to treat a mammalian insult or injury. A stem cell culture supernatant containing at least one factor capable of exerting effective neuroprotection to treat a mammalian neural injury or insult.

1 . A method of producing a stem cell conditioned medium, comprising the steps of:
 providing at least one stem cell selected from the group consisting of a human pluripotent stem cell from skeletal muscle, a human brain stem cell, a human common pluripotent stem cell, or a human progenitor cell derived from a mesenchymal stem cell;   providing at least one medium for culturing the at least one stem cell;   culturing the at least one stem cell in the at least one medium;   replacing some or all of the at least one medium with a growth-factor free basal medium and further conditioning the at least one stem cell in either normoxic or hypoxic conditions; and   collecting cell supernatants.   
     
     
         2 . The method of producing a stem cell conditioned medium, as in  claim 1 , wherein the at least one stem cell is conditioned in either normoxic or hypoxic conditions for at least 24 hours. 
     
     
         3 . The method of producing a stem cell conditioned medium, as in  claim 1 , wherein the at least one medium is EGM2MV. 
     
     
         4 . The method of producing a stem cell conditioned medium, as in  claim 1 , wherein the at least one stem cell has a surface marker profile of lin-/CD45−/c-kit-/CD90+. 
     
     
         5 . The method of producing a stem cell conditioned medium, as in  claim 1 , wherein the at least one stem cell display the surface marker profile CD34+ positive. 
     
     
         6 . The method of producing a stem cell conditioned medium, as in  claim 1 , wherein the cell supernatants are effective to treat neural insults. 
     
     
         7 . A method of producing a stem cell conditioned medium, the method comprising the steps of:
 culturing at least one stem cell in a first cell culture medium, wherein the at least one stem cell is selected from the group consisting of a human pluripotent stem cell from skeletal muscle, a human brain stem cell, a human common pluripotent stem cell, or a human progenitor cell derived from a mesenchymal stem cell;   replacing some or all of the first cell culture medium with a second cell culture medium and further culturing the at least one stem cell in the second cell culture medium to generate a plurality of cultured cells;   collecting a quantity of the second cell culture medium after a culture duration, wherein the quantity of the second cell culture medium contains secretions from the plurality of cultured cells;   wherein the step of culturing comprises culturing the at least one stem cell having a surface marker profile selected from the group consisting of lin-/CD45−/c-kit-/CD90+, lin-/CD45−/c-kit-/CD90+/Sca-1+, and lin-/CD45−/c-kit-/CD90+/CD34+.   
     
     
         8 . The method of producing a stem cell conditioned medium, as in  claim 7 , wherein the step of replacing comprises replacing some or all of the first cell culture medium with a basal medium that is at least substantially growth factor-free. 
     
     
         9 . The method of producing a stem cell conditioned medium, as in  claim 7 , wherein the step of collecting comprises collecting the quantity of the second cell culture medium containing at least one factor selected from the group consisting of at least one angiogenic factor and at least one antiapoptotic factor. 
     
     
         10 . The method of producing a stem cell conditioned medium, as in  claim 7 , wherein the step of culturing comprises culturing the at least one stem cell in the first cell culture medium comprising a first amount of growth factor, and wherein the step of replacing comprising replacing some or all of the first cell culture medium with the second cell culture medium having a second amount of growth factor, and wherein the second amount is less than the first amount. 
     
     
         11 . The method of producing a stem cell conditioned medium, as in  claim 7 , wherein the step of culturing comprises culturing the at least one stem cell to confluence. 
     
     
         12 . The method of producing a stem cell conditioned medium, as in  claim 7 , further comprising the step of fractionating the quantity of the second cell culture medium to remove substances less than a specified kDa range selected from the group consisting of less than about 5 kDa, less than about 10 kDa, less than about 20 kDa, less than about 30 kDa, less than about 40 kDa, less than about 50 kDa, and greater than about 50 kDa. 
     
     
         13 . The method of producing a stem cell conditioned medium, as in  claim 7 , wherein the step of replacing comprises replacing some or all of the first cell culture medium with the second cell culture medium for conditioning in either normoxic or hypoxic conditions. 
     
     
         14 . A stem cell conditioned medium, comprising:
 a cell culture supernatant containing at least one factor capable of exerting effective neuroprotection to treat a mammalian neural injury or insult, the cell culture supernatant produced by culturing at least one stem cell to produce the at least one factor;   wherein the at least one stem cell comprises a marker profile of lin-/CD45−/c-kit-/CD90+/CD34+ marker before culturing and loses the CD34+ marker during culturing; and   wherein the at least one stem cell is selected from the group consisting of a human pluripotent stem cell from skeletal muscle, a human brain stem cell, a human common pluripotent stem cell, or a human progenitor cell derived from a mesenchymal stem cell.   
     
     
         15 . The stem cell conditioned medium of  claim 14 , wherein the cell culture supernatant is produced by culturing the at least one stem cell in a first cell culture medium and later removing some or all of the first cell culture medium and further culturing the at least one stem cell in a second cell culture medium. 
     
     
         16 . The stem cell conditioned medium of  claim 14 , wherein the second cell culture medium comprises a growth factor-free basal medium. 
     
     
         17 . The stem cell conditioned medium of  claim 14 , wherein the first cell culture medium comprises a first amount of growth factor, wherein the second cell culture medium comprises a second amount of growth factor, and wherein the second amount is less than the first amount. 
     
     
         18 . The stem cell conditioned medium of  claim 14 , wherein the cell culture supernatant is collected after the at least one stem cell has been cultured in the second cell culture medium. 
     
     
         19 . The stem cell conditioned medium of  claim 14 , wherein the at least one factor is selected from the group consisting of at least one angiogenic factor and at least one antiapoptotic factor. 
     
     
         20 . The stem cell conditioned medium of  claim 2 , wherein the at least one stem cell is cultured in the second cell culture medium under a condition selected from the group consisting of a normoxic condition and a hypoxic condition.