RNAlater® Tissue Collection

To test the hypothesis that central and peripheral neural transcriptomes show fundamental quantitative differences, Dr. Mark LeDoux and colleagues (University of Tennessee Health Science Center, Memphis, TN) used Affymetrix GeneChip® expression arrays to compare murine lumbar spinal cord (SC) and dorsal root ganglion (DRG) gene expression [1]. They used the novel technique of perfusion-fixing mice with RNAlater® Solution to preserve RNA integrity before the SC and DRG were harvested. The vascular tree was flushed with DEPC-treated saline, tissues were fixed, and RNA was stabilized by perfusion with RNAlater Solution. The vertebral column was placed in RNAlater Solution and the SC and DRG were dissected out. RNAlater Solution was continually dripped on the samples during dissection. Samples were then placed in TRI Reagent® solution (Applied Biosystems) for RNA extraction.

Placental Samples

Aspartyl-(asparaginyl) β-hydroxylase (AAH) is a transmembrane protein that hydroxylates epidermal growth factor-like domains of proteins that have a functional role in cell motility and invasion. Dr. Fusun Gundogan and colleagues (Brown Medical School, Providence, RI) characterized the potential role of AAH in implantation using human placentas from terminated pregnancies, normal term deliveries, spontaneous abortuses, and small-for-gestational-age term deliveries [2]. Snap-frozen or RNAlater solution-preserved specimens were used for RNA analysis of AAH expression by RT-PCR and protein analysis by Western blotting. Immunohistochemical staining and RNA analysis suggested that AAH may serve as a biomarker of impaired implantation.

Embryonic Samples

The process of testicular descent depends on the gubernaculum, which enlarges and migrates into the scrotum, creating a space into which the testis descends. Development of the fetal gubernaculum requires the expression of leucine rich G protein coupled receptor and its ligand, insulin-like 3. Dr. Julia Barthold and colleagues (A I DuPont Hospital for Children, Wilmington, DE) compared expression levels of these two genes in Long-Evans orl and wild type rats during perinatal development—failure of testicular descent occurs in ~65% of spontaneously cryptorchid Long-Evans orl rats [3]. Males from both strains obtained at gestational days 18–21 and day of birth were preserved in RNAlater Solution for at least 24 hours. The size and position of the testes, kidneys, and gubernacula were determined by microdissection and image analysis. Leucine-rich G protein coupled receptor and insulin-like 3 mRNA expression were analyzed by RT-PCR. Data suggest that the impaired shortening of the Long-Evans orl gubernaculum may interfere with the timing and quality of its inversion at birth, leading to failure of testicular descent in some newborn males.