Muscle Derived Cells for the Treatment of Gastro-Esophageal Pathologies and Methods of Making and Using the Same

Abstract

The present invention provides muscle-derived progenitor cells that show long-term survival following transplantation into body tissues and which can augment soft tissue following introduction (e.g. via injection, transplantation, or implantation) into a site of soft tissue. Also provided are methods of isolating muscle-derived progenitor cells, and methods of genetically modifying the cells for gene transfer therapy. The invention further provides methods of using compositions comprising muscle-derived progenitor cells for the augmentation and bulking of mammalian, including human, soft tissues in the treatment of various cosmetic or functional conditions, including malformation, injury, weakness, disease, or dysfunction. In particular, the present invention provides treatments and amelioration of symptoms for gastro-esophageal pathologies like gastro-esophageal reflux.

What is claimed is: 
     
         1 . A method of increasing lower esophageal sphincter pressure in a mammalian subject comprising:
 (a) isolating skeletal muscle cells from the mammalian subject;   (b) cooling the cells to a temperature lower than 10° C. and storing the cells for 1-7 days;   (c) suspending the mammalian skeletal muscle cells in a first cell culture container between 30 and 120 minutes;   (d) decanting the media from the first cell culture container to a second cell culture container;   (e) allowing the remaining cells in the media to attach to the walls of the second cell culture container;   (f) isolating the cells from the walls of the second cell culture container, wherein the isolated cells are MDCs;   (g) culturing the cells to expand their number;   (h) freezing the MDCs to a temperature below −30° C.; and   (i) thawing the MDCs and administering the MDCs to the esophagus of the mammalian subject;   
       thereby, increasing lower esophageal sphincter pressure by at least about 50% in a mammalian subject. 
     
     
         2 . The method of  claim 1 , wherein the skeletal muscle cells are isolated from the mammalian subject before the gastro-esophageal reflux disease begins in the mammalian subject. 
     
     
         3 . The method of  claim 1 , wherein increase in lower esophageal sphincter pressure is increased by at least about 100%. 
     
     
         4 . The method of  claim 1 , wherein the MDCs are administered by injecting them into the esophagus. 
     
     
         5 . The method of  claim 1 , wherein the MDCs are injected into the lower esophageal sphincter. 
     
     
         6 . The method of  claim 1 , wherein the mammal is a human. 
     
     
         7 . A method of increasing lower esophageal sphincter pressure in a mammalian subject comprising:
 (a) isolating skeletal muscle cells from the mammalian subject,   (b) suspending mammalian skeletal muscle cells in a first cell culture container for between 30 and 120 minutes;   (c) decanting the media from the first cell culture container to a second cell culture container;   (d) allowing the remaining cells in the media to attach to the walls of the second cell culture container;   (e) isolating the cells from the walls of the second cell culture container, wherein the isolated cells are MDCs; and   (f) administering the MDCs to the esophagus of the mammalian subject; thereby, increasing lower esophageal sphincter pressure in a in a mammalian subject.   
     
     
         8 . The method of  claim 7 , wherein increase in lower esophageal sphincter pressure is increased by at least about 100%. 
     
     
         9 . The method of  claim 7 , wherein the MDCs are administered by injecting them into the esophagus. 
     
     
         10 . The method of  claim 9 , wherein the MDCs are injected into the lower esophageal sphincter. 
     
     
         11 . The method of  claim 7 , wherein the mammal is a human. 
     
     
         12 . The method of  claim 7 , wherein the MDCs are cultured to expand their number before being administered to the esophagus of the mammalian subject. 
     
     
         13 . A method of increasing lower esophageal sphincter pressure in a mammalian subject comprising:
 (a) plating a suspension of skeletal muscle cells from skeletal muscle tissue in a first container to which fibroblast cells of the skeletal muscle cell suspension adhere,   (b) re-plating non-adherent cells from step (a) in a second container, wherein the step of re-plating is after about 15 to about 20% of cells have adhered to the first container;   (c) repeating step (b) at least once;   (d) isolating the skeletal muscle-derived MDCs and administering the MDCs to the esophagus of the mammalian subject;   
       thereby increasing lower esophageal sphincter pressure in a mammalian subject. 
     
     
         14 . The method of  claim 13 , wherein increase in lower esophageal sphincter pressure is increased by at least about 100%. 
     
     
         15 . The method of  claim 13 , wherein the MDCs are administered by injecting them into the esophagus. 
     
     
         16 . The method of  claim 13 , wherein the MDCs are injected into the lower esophageal sphincter. 
     
     
         17 . The method of  claim 13 , wherein the mammal is a human.