Extraction of RNA from archival tissues and measurement of thrombospondin-1 mRNA in normal, dysplastic, and malignant oral tissues.
Thrombospondin-1 (TSP-1) is an extracellular matrix glycoprotein implicated in the regulation of angiogenesis and tumour development. Our objectives were to ascertain the quantity and quality of RNA extracted from archival, formalin-fixed, paraffin embedded, oral tissues and their application in measuring the concentrations of TSP-1 mRNA in these tissues. We compared three techniques of isolation of RNA as well as related experimental variables. TSP-1 mRNA was measured in specimens of normal, dysplastic, and malignant oral tissues by real-time reverse transcriptase polymerase chain reaction (RT-PCR). RNA suitable for analysis by real-time RT-PCR was obtained by the three techniques tested, although the yield varied depending on the protocol used (range 0.2-3.6 microg/mm(3)). The mean (S.D.) concentrations of TSP-1 mRNA relative to 18S were 21.1 (7.2) in normal oral tissues (n=9), 11.0 (8.2) in dysplastic tissue (n=8) and 7.3 (5.3) in carcinomatous tissue (n=17). The difference between normal and carcinomatous specimens was significant (p=0.01). This reduction in expression of TSP-1 mRNA from normal to dysplasia to carcinoma may favour the angiogenic drive that accompanies the development of oral tumours.
Macluskey
M
M
Unit of Cell and Molecular Biology, Dental School, University of Dundee, Park Place, Dundee DD1 4HR, UK.
Baillie
R
R
Morrow
H
H
Schor
S L
SL
Schor
A M
AM
D002294:Carcinoma, Squamous Cell / Q000098:blood supply / Q000737:chemistry*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006652:Histological Techniques; D006801:Humans; D008297:Male; D008875:Middle Aged; D009061:Mouth Mucosa / Q000737:chemistry* / Q000473:pathology; D009062:Mouth Neoplasms / Q000098:blood supply / Q000737:chemistry*; D009389:Neovascularization, Pathologic / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000302:isolation & purification; D019700:Thrombospondin 1 / Q000032:analysis* / Q000235:genetics; D014021:Tissue Preservation
D012333:RNA, Messenger; D019700:Thrombospondin 1