Extraction of RNA from archival tissues and measurement of thrombospondin-1 mRNA in normal, dysplastic, and malignant oral tissues

• Published in: British journal of oral & maxillofacial surgery Vol. 44; no. 2; pp. 116 - 123
• Main Authors: Macluskey, M., Baillie, R., Morrow, H., Schor, S.L., Schor, A.M.
• Format: Journal Article
• Language: English
• Published: Londonc Elsevier Ltd 01.04.2006; Elsevier
• Subjects: Biological and medical sciences; Carcinoma, Squamous Cell - blood supply; Carcinoma, Squamous Cell - chemistry; Dentistry; Female; Gene Expression Regulation, Neoplastic; Histological Techniques; Humans; Male; Medical sciences; Messenger RNA; Middle Aged; Mouth mucosa; Mouth Mucosa - chemistry; Mouth Mucosa - pathology; Mouth neoplasm; Mouth Neoplasms - blood supply; Mouth Neoplasms - chemistry; Neovascularization, Pathologic - genetics; Otorhinolaryngology. Stomatology; Polymerase Chain Reaction - methods; RNA, Messenger - isolation & purification; Thrombospondin 1 - analysis; Thrombospondin 1 - genetics; Thrombospondin-1; Tissue Preservation; Thrombospondin-1; Mouth neoplasm; Messenger RNA; Mouth mucosa; Human; Oral cavity; Tissue; Treatment; Stomatology; Surgery; Malignant tumor; Thrombospondin 1
• ISSN: 0266-4356; 1532-1940
• DOI: 10.1016/j.bjoms.2005.03.001

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 μg/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.