ABSTRACT
BACKGROUND: Several studies have shown that hair follicle bugle cells can differentiate into hair follicles and contribute to the formation of the epidermis and sebaceous gland. Although many lines of evidence have suggested that the renewal and maintenance of the sebaceous gland depends on hair follicle bulge cells, direct evidence supporting the in vitro differentiation of follicle bulge cells into sebaceous gland cells has not been found. METHODS: Rat vibrissa follicle bulge cells were isolated, cultured, and transfected with green fluorescent protein (GFPC1) plasmids carrying the peroxisome proliferator-activated receptor gamma2 (PPARgamma2 ) gene. The transfected cells were cultured in modified medium, and the morphologic changes of the cells were observed. Moreover, the expression of epithelial membrane antigens (EMAs) by the cells was detected by immunocytochemistry, and adipogenesis of the cells was evaluated. RESULTS: After induction culture, the cell body enlarged gradually and contained abundant cytoplasm; lipid droplets appeared in the cytoplasm of some cells, and the cells resembled sebocytes of the sebaceous gland. The cells were positive on oil red O and EMA staining. The expression of PPARgamma2 mRNA and protein was significantly upregulated in PPARgamma2-transfected cells. The rate of oil red O-stained and EMA-positive cells was higher in PPARgamma2-transfected cells after induction than in bulge-PPARgamma2 cells and non-transfected bulge cells. CONCLUSIONS: Rat vibrissa hair follicle bulge cells may differentiate into sebocytes in vitro, and the PPARgamma2 gene plays a crucial role.
Subject(s)
Cell Differentiation/genetics , Hair Follicle/cytology , PPAR gamma/metabolism , Sebaceous Glands/cytology , Animals , Cell Culture Techniques , Cells, Cultured , Hair Follicle/pathology , Immunohistochemistry , Male , Models, Animal , RNA, Messenger/metabolism , Rats , Rats, Wistar , Reverse Transcriptase Polymerase Chain Reaction , Sebaceous Glands/pathology , Transfection/methods , Up-RegulationABSTRACT
BACKGROUND: We constructed a cDNA subtractive library of dermal papilla cells (DPCs) in anagen with suppression subtractive hybridization (SSH) technique and clone differentially expressed genes related to DPCs in anagen. METHODS: Total mRNA was isolated from DPCs of anagen and telogen follicles. Moreover, single-strand (ss) and double-strand (ds) cDNAs were synthesized in turn using SMART PCR cDNA synthesis technology. ds cDNAs then were digested with Rsa I and divided into two groups, and ligated to the specific adaptor 1 and adaptor 2R, respectively. After cDNAs were hybridized with each other twice and underwent two rounds of nested PCR. PCR products were ligated with arms of T/A plasmid vectors to set up the subtractive library. Selected clones were demonstrated by reverse Northern blot and sequenced. The acquired sequence data were aligned against the Genbank nucleotide database. RESULTS: cDNA subtractive library of DPCs in anagen follicles was set up successfully with high subtractive efficiency. Thirty-five genes were identified in this study with 22 known functional genes and 13 unknown functional genes. CONCLUSIONS: All results confirm the effectiveness and sensitivity of SSH in detecting differentially expressed genes from a small amount of clinical samples. Information about such alterations in gene expression could be useful for elucidating the genetic events in hair follicle growth regulation.
