May Argyrophilic Nucleolar Organizer Regions Be Used as a Biomarker for the Detection of the Degree of Ischemic Damage Instead of Tunel in Testicular Torsion?

Abstract

Background and Objectives: It is of great importance to obtain information about the severity of ischemic damage and duration of testicular torsion for an effective treatment strategy. Nucleolar-organizing regions (NORs) are sites of the ribosomal genes composed of ribosomal DNA and proteins. Post-silver staining NORs are termed "AgNOR". Since AgNORs clearly reveals the self-renewal potential of cells damaged in ischemic events, we performed the current study. Materials and Methods: The study was carried out in four groups as control, sham, early, and late T/D. In the surgical groups, testes were corrected after a 4-h ischemia period. Testicular tissue samples were taken on the third day after detorsion in group 1, 2, 3, and on the tenth day after detorsion in group 4. TUNEL and silver stainings were applied to all samples. Results: The differences were significant among the groups for both mean AgNOR number and total AgNOR area/total nuclear area (TAA/TNA). Moreover, the differences between control and early torsion-detorsion (T/D), between control and late T/D, between sham and early T/D, between sham and late T/D, and between early T/D and late were statistically significant for AgNOR amount. Furthermore, statistically significant differences among the groups for an average number of apoptotic cells per tubule and the percentage of apoptotic tubule values were detected. Discussion: The apoptotic index gives the ratio of cells that are damaged and will die in a programmed way and cells that remain intact, rather than show the viability of the returning testicle. However, by measuring cells that regenerate with AgNOR, we can show not only those that survive but also cells that can repair themselves. Conclusion: AgNOR proteins are usable for the early observation of ischemic injury levels. The amount of AgNOR protein can enlighten us about the extent of testicular damage after T/D treatment. It may also help the physician in the development of effective treatment strategies for cases.

Keywords: AgNOR; ischemia; nucleolar organizer regions; rDNA; testis.

Figure 1

Demonstrative examples of TUNEL staining in Seminiferous tubules of each group. (a): control; (b): sham; (c): ET/D (early torsion/detorsion group); (d): LT/D (late torsion/detorsion group).

Figure 2

Comparison of mean AgNOR number (a) and TAA/TNA ratio (b) of groups.

Figure 3

Relation between mean AgNOR numbers of testes cells and ANPCT (a), between mean AgNOR numbers of testes cells and PAT (b). Relation between mean TAA/TNA ratio of testes cells and ANPCT (c) and between TAA/TNA ratio of testes cells and PAT (d).

Figure 4

Representative images of silver stained NOR (a) Control, (b) Sham, (c) Early T/D, (d) Late T/D for testicular cells. The arrows showed the AgNOR proteins.