Quantification of PERF 15 mRNA in Tissue Sections from Rat Testes

Abstract

We previously conducted basic research to quantify in situ hybridization (ISH) signals in rat testes. In this experimental model, we selected ribosomal RNA (rRNA) as the hybridizable RNA in paraffin sections, since it allowed us to easily analyze ISH signals expressed with digoxygenin (DIG)-labeled probes quantitatively through “posterization” of the images. We applied this method to analyze the quantification of transcript, PERF 15 mRNA. PERF 15 is expressed specifically in the testes and localized in the rigid cytoskeletal structure of the sperm head, and has been considered to be involved in the apoptotic process of spermatogenic cells. Quantification of the signals may help to clarify the detailed function of PERF 15. We further analyzed the signals concomitant with a confocal laser scanning microscope. The peak of PERF 15 mRNA expression was found in diplotene spermatocytes, and the amount of PERF 15 mRNA was greatest in late pachytene and diplotene spermatocytes and early spermatids, followed by early pachytene spermatocytes, and then late spermatids. PERF 15 may be involved in the events leading to meiotic division, in which apoptosis is also involved. The present study may help to determine the concentration of mRNA in tissue sections.

Identification, localization, and regulation of passerine GnRH-I messenger RNA

“…The GnRH-I mRNA density value of each bird was obtained by calculating the mean density of ten randomly chosen cells in the brain section which had the largest numbers of GnRH-I neurons. Quantification of non-radioactive in situ hybridi-zation signal was validated as in previous studies (Ubuka et al 2005, Kogami et al 2006. GnRH-I mRNA density levels were compared between mature and immature female birds using Student's t-test.…”

Maybe repressed mRNAs are not stored in the chromatoid body in mammalian spermatids

“…However, useful information is limited to a relatively small group of studies that utilize non-isotopic hybridization and good fixation and present photographs in which the location of the hybridization signal can be visualized. All of these studies show no localization (Weitzel et al 2003, Iida et al 2004, Ellis et al 2005, Kogami et al 2006. In contrast, Fukuda et al (2004) reported that the MOR23 mRNA is localized close to the nucleus (chromatoid body?…”

MeCP2 Expression and Promoter Methylation of Cyclin D1 Gene Are Associated with Cyclin D1 Expression in Developing Rat Epididymal Duct

“…The intensities of the signal of MeCP2 in epididymal duct, stromal cells and capillary endothelium of normal developing testis of (0- and 7-day-old) rats were analyzed by the one way ANOVA as described [ 10 ] to examine whether there were any statistical differences between the two groups. Twenty-five high-power fields for each of the three independent samples were evaluated.…”