Recovery and expression of messenger RNA from postmortem human brain tissue.
The Bryan Alzheimer's Disease Research Center Rapid Autopsy Program at Duke University Medical Center obtains postmortem human brain tissue for experimental investigations. We evaluated 19 brains for RNA integrity and mRNA gene expression. Nine were from patients diagnosed with Alzheimer's disease, and ten were from nondemented controls. In all cases, the following variables were recorded: postmortem procurement delay (range, 1 hour and 10 minutes to 14 hours), pH of cerebrospinal fluid, premortem fever or sepsis, provision of supplemental oxygen in the agonal period, and temporal relation to time of death (either sudden death or protracted illness). Total RNA was extracted, quantified, and evaluated by agarose gel electrophoresis and quantitative gene expression analysis of 18S rRNA and edg-1 using TaqMan technology. All samples appeared to yield intact RNA without significant degradation, and expression of the edg-1 gene was detected by the real time reverse transcriptase polymerase chain reaction in all cases. We conclude that intact RNA can be obtained from postmortem human brain tissue, even in patients with severe premortem illnesses and delayed postmortem tissue procurement intervals. However, we caution that the successful expression of certain genes from postmortem brain tissue may require enhanced procurement efforts to maximize RNA integrity.
Cummings
T J
TJ
Department of Pathology, Duke University Medical Center, Box 3712, Durham, NC 27710, USA. cummi008@mc.duke.edu
Strum
J C
JC
Yoon
L W
LW
Szymanski
M H
MH
Hulette
C M
CM
D000368:Aged; D000369:Aged, 80 and over; D001921:Brain / Q000378:metabolism*; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics; D008297:Male; D008875:Middle Aged; D011180:Postmortem Changes; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D049312:Receptors, Lysophospholipid; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013048:Specimen Handling / Q000592:standards; D013997:Time Factors
D017874:Immediate-Early Proteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D049312:Receptors, Lysophospholipid