Methods of evaluating cells and cell cultures

Abstract

Methods of evaluating the composition of a cell culture (e.g., to distinguish chondrocytes from fibroblasts) and methods for evaluating the phenotype of an individual cell (e.g., as a chondrocyte) are disclosed. The methods may be used, for example, for assessing chondrocyte cultures used for treatment of cartilage defects. In some embodiments, the invention involves identifying cell culture composition or the identity of a cell based on expression level of a fibroblast marker. In other embodiments, the invention involves comparing expression levels of at least one chondrocyte marker and at least one fibroblast marker in a cell culture sample or in an individual cell. In illustrative embodiments, the chondrocyte marker is hyaluronan and proteoglycan link protein 1 (HAPLN1), and the fibroblast marker is microfibrillar associated protein 5 (MFAP5).

1 .- 35 . (canceled) 
     
     
         36 . A composition comprising:
 a) at least one polynucleotide chosen from (i) an RNA isolated from a cell culture comprising chondrocytes or (ii) a cDNA prepared from an RNA isolated from a cell culture comprising chondrocytes; and   b) at least one oligonucleotide specific for microfibrillar associated protein 5 (MFAP5).   
     
     
         37 . The composition of  claim 36 , further comprising at least one oligonucleotide specific for a chondrocyte marker. 
     
     
         38 . The composition of  claim 37 , wherein the chondrocyte marker is chosen from hyaluronan and proteoglycan link protein 1 (HAPLN1), matrix Gla protein (MGP), GF-like repeats and discoidin I-like domains 3 (EDIL3), WNT1 inducible signaling pathway protein 3 (WISP3), aggrecan 1 (AGC1), cartilage oligomeric matrix protein (COMP), type II collagen (COL2A1), type IX collagen (COL9A1), type XI collagen (COL11A1), leukocyte cell derived chemotaxin 1 protein (LECT1), S100 calcium binding protein beta (S100B), cartilage acidic protein 1 (CRTAC1), SRY-box9 protein (SOX9), and nebulette (NEBL). 
     
     
         39 . The composition of  claim 37 , wherein the chondrocyte marker is hyaluronan and proteoglycan link protein 1 (HAPLN1). 
     
     
         40 . The composition of  claim 39 , wherein the at least one oligonucleotide specific for the chondrocyte marker comprises at least one of SEQ ID NOs: 16-18. 
     
     
         41 . The composition of  claim 39 , wherein the at least one oligonucleotide specific for the chondrocyte marker comprises at least one of SEQ ID NOs: 22-24. 
     
     
         42 . The composition of  claim 36 , wherein the at least one oligonucleotide specific for MFAP5 comprises at least one of SEQ ID NOs: 19-21. 
     
     
         43 . The composition of  claim 36 , wherein the at least one oligonucleotide specific for MFAP5 comprises at least one of SEQ ID NOs: 25-27. 
     
     
         44 . The composition of  claim 36 , wherein the at least one polynucleotide is an RNA. 
     
     
         45 . The composition of  claim 36 , wherein the at least one oligonucleotide is a probe or a primer. 
     
     
         46 . The composition of  claim 36 , wherein the at least one polynucleotide is chosen from (i) an RNA isolated from a cell culture comprising chondrocytes obtained from a cartilage biopsy or (ii) a cDNA prepared from an RNA isolated from a cell culture comprising chondrocytes obtained from a cartilage biopsy. 
     
     
         47 . The composition of  claim 36 , wherein the cartilage biopsy is taken from a knee joint. 
     
     
         48 . The composition of  claim 36 , wherein the at least one polynucleotide is chosen from (i) an RNA isolated from a primary cell culture comprising chondrocytes or (ii) a cDNA prepared from an RNA isolated from a primary cell culture comprising chondrocytes. 
     
     
         49 . The composition of  claim 36 , wherein the at least one polynucleotide is chosen from (i) an RNA isolated from a cell culture comprising passaged chondrocytes or (ii) a cDNA prepared from an RNA isolated from a cell culture comprising passaged chondrocytes. 
     
     
         50 . The composition of  claim 36 , wherein the at least one polynucleotide is chosen from (i) an RNA isolated from a cell culture comprising passaged, de-differentiated chondrocytes or (ii) a cDNA prepared from an RNA isolated from a cell culture comprising passaged, de-differentiated chondrocytes. 
     
     
         51 . The composition of  claim 36 , wherein the at least one polynucleotide is chosen from (i) an RNA isolated from a cell culture comprising human chondrocytes or (ii) a cDNA prepared from an RNA isolated from a cell culture comprising human chondrocytes. 
     
     
         52 . The composition of  claim 36 , wherein the at least one polynucleotide is chosen from (i) an RNA isolated from a cell culture comprising at least 50% chondrocytes or (ii) a cDNA prepared from an RNA isolated from a cell culture comprising at least 50% chondrocytes. 
     
     
         53 . The composition of  claim 36 , wherein the at least one polynucleotide is chosen from (i) an RNA isolated from a cell culture that further comprises synoviocytes or (ii) a cDNA prepared from an RNA isolated from a cell culture that further comprises synoviocytes. 
     
     
         54 . The composition of  claim 36 , wherein the at least one polynucleotide is chosen from (i) an RNA isolated from a cell culture that further comprises fibroblasts or (ii) a cDNA prepared from an RNA isolated from a cell culture that further comprises fibroblasts. 
     
     
         55 . The composition of  claim 36 , wherein the at least one polynucleotide is chosen from (i) an RNA isolated from a cell culture comprising a collagen matrix loaded with chondrocytes or (ii) a cDNA prepared from an RNA isolated from a cell culture comprising a collagen matrix loaded with chondrocytes.