Generation and characterization of telomerase-transfected human lymphatic endothelial cells with an extended life span.
The study of lymphatic endothelial cells and lymphangiogenesis has, in the past, been hampered by the lack of lymphatic endothelial-specific markers. The recent discovery of several such markers has permitted the isolation of lymphatic endothelial cells (LECs) from human skin. However, cell numbers are limited and purity is variable with the different isolation procedures. To overcome these problems, we have transfected human dermal microvascular endothelial cells (HDMVECs) with a retrovirus containing the coding region of human telomerase reverse transcriptase (hTERT), and have produced a cell line, hTERT-HDLEC, with an extended lifespan. hTERT-HDLEC exhibit a typical cobblestone morphology when grown in culture, are contact-inhibited, and express endothelial cell-specific markers. hTERT-HDLEC also express the recognized lymphatic markers, Prox-1, LYVE-1 and podoplanin, as well as integrin alpha9, but do not express CD34. They also form tube-like structures in three-dimensional collagen gels when stimulated with vascular endothelial growth factors -A and -C. Based on these currently recognized criteria, these cells are LEC. Surprisingly, we also found that the widely studied HMEC-1 cell line expresses recognized lymphatic markers; however, these cells are also CD34-positive. In summary, the ectopic expression of hTERT increases the life span of LECs and does not affect their capacity to form tube-like structures in a collagen matrix. The production and characterization of hTERT-HDLEC will facilitate the study of the properties of lymphatic endothelium in vitro.
Nisato
Riccardo E
RE
Department of Morphology, University Medical Center, 1 rue Michel Servet, 1211 Geneva 4, Switzerland.
Harrison
Jillian A
JA
Buser
Raphaele
R
Orci
Lelio
L
Rinsch
Chris
C
Montesano
Roberto
R
Dupraz
Philippe
P
Pepper
Michael S
MS
D015415:Biomarkers; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D016922:Cellular Senescence; D018920:Coculture Techniques; D024042:Collagen Type I / Q000378:metabolism; D004268:DNA-Binding Proteins; D004729:Endothelium, Lymphatic / Q000166:cytology* / Q000201:enzymology / Q000276:immunology / Q000378:metabolism / Q000648:ultrastructure; D004730:Endothelium, Vascular / Q000166:cytology* / Q000201:enzymology / Q000276:immunology / Q000378:metabolism / Q000648:ultrastructure; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D005782:Gels; D006801:Humans; D007150:Immunohistochemistry; D042601:Lymphatic Vessels / Q000166:cytology; D020782:Matrix Metalloproteinases / Q000032:analysis / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D012190:Retroviridae / Q000235:genetics; D012867:Skin / Q000166:cytology; D019098:Telomerase / Q000235:genetics / Q000378:metabolism*; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
D015415:Biomarkers; D024042:Collagen Type I; D004268:DNA-Binding Proteins; D005782:Gels; D011994:Recombinant Proteins; D042461:Vascular Endothelial Growth Factor A; D016222:Fibroblast Growth Factor 2; D019098:Telomerase; D020782:Matrix Metalloproteinases