Maintaining mRNA integrity during decalcification of mineralized tissues

Biomineralization of the extracellular matrix occurs inappropriately in numerous pathological conditions such as cancer and vascular disease, but during normal mammalian development calcification is restricted to the formation of the skeleton and dentition. The comprehensive study of gene expression...

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Bibliographic Details
Published in:PloS one 2013-03, Vol.8 (3), p.e58154-e58154
Main Authors: Belluoccio, Daniele, Rowley, Lynn, Little, Christopher B, Bateman, John F
Format: Article
Language:English
Subjects:
Analysis
Animals
Arthritis
Biology
Calcification
Calcification, Physiologic
Cancer
Cartilage - metabolism
Collagen
Decalcification
Dentition
Disease
Dissection
Epiphysis
Ethylenediaminetetraacetic acids
Extracellular matrix
Gene Expression
Genes
Histology
Hybridization
Immunohistochemistry
Integrity
Laboratories
Localization
Male
Messenger RNA
Methods
Mice
Mineralization
Morphology
Pathology
Proteins
RNA polymerase
RNA Stability
RNA, Messenger - metabolism
Sectioning
Skeleton
Studies
Teeth
Tissues
Vascular diseases
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Summary:Biomineralization of the extracellular matrix occurs inappropriately in numerous pathological conditions such as cancer and vascular disease, but during normal mammalian development calcification is restricted to the formation of the skeleton and dentition. The comprehensive study of gene expression in mineralized skeletal tissues has been compromized by the traditional decalcification/fixation methods that result in significant mRNA degradation. In this study we developed a novel RNAlater/EDTA decalcification method that protects the integrity of the mRNA in mature mouse tibial epiphyses. Furthermore, this method preserves the tissue structure to allow histological sectioning and microdissection to determine region-specific gene expression, in addition to immuno- and in situ histology. This method will be widely applicable to the molecular analysis of calcified tissues in various pathological conditions, and will be of particular importance in dissection of the gene expression in mouse bone and joint tissues during development and in important clinical conditions such as arthritis.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0058154