MicroRNA expression profiles in testicular biopsies of patients with impaired spermatogenesis

Abstract

Spermatogenesis is a complex process that involves thousands of genes whose expression during different stages is strictly regulated. Small non-coding microRNAs play an important role in the posttranscriptional regulation of mRNA processing during spermatogenesis. Using Agilent SurePrint v16 microRNA 8 × 60 K microarray kit, we investigated the microRNA expression profiles of 24 formalin-fixed paraffin-embedded testicular biopsies from patients with hypospermatogenesis (n = 10), hypospermatogenesis and azoospermia factor c region on the Y chromosome (AZFc) deletion (n = 3), Sertoli cell-only syndrome (n = 3) and maturation arrest (n = 2), in comparison with subjects with normal spermatogenesis (n = 6). After adjusting for multiple testing, six deregulated miRNAs were detected in the patients with AZFc deletion, 30 in maturation arrest group, 52 in Sertoli cell-only syndrome group of patients, and none in the group of patients with hypospermatogenesis. Some of the deregulated microRNAs were shared between groups, resulting in 58 unique differentially expressed microRNAs. The expression of five microRNAs (hsa-miR-34b, hsa-miR-449b, hsa-miR-517c, hsa-miR-181c, and hsa-miR-605) was validated by qRT-PCR in a total of 74 samples. Using mRNA expression profiles of subjects with matching histopathological patterns of impaired spermatogenesis from publically available Gene Expression Omnibus data sets, we have performed integrated mRNA-microRNA regulatory network analysis. Pathway analysis revealed significantly enriched set of genes for tumor necrosis factor-related apoptosis-inducing ligand signaling pathway, previously shown to be involved in regulation of apoptosis in normal functioning testis. Our results should be considered as preliminary as we have analyzed only a small number of patients in each studied group. Further studies with larger number of patients with impaired spermatogenesis as well as more targeted approaches with parallel microRNA and mRNA expression profiling in isolated subpopulations of somatic or germ cells from different stages of spermatogenesis are needed to clarify the role of the microRNAs in the process of spermatogenesis.

Seminal human papillomavirus originates from the body surface and is not a frequent aetiological factor in azoospermia

“…Since it was not possible to detect HPV DNA in the testicular biopsies from unselected azoospermic men in the present study, HPV is probably not a frequent cause of azoospermia. To highlight the aetiologies of azoospermia, and particularly NOA not explained by other probable causes, it might be relevant to evaluate other factors than HPV, for example possible genetic factors (Stouffs et al, ) including the expression of microRNA (Noveski et al, ).…”

Analysis of mRNA Expression Levels in FFPE Testicular Biopsies Using Real-time PCR Arrays

“…We have previously compared this kit with the RNA isolation method using the Trizol reagent and with two other commercial kits for RNA isolation only; we had found that the AllPrep DNA/RNA FFPE Kit was best suited for our needs because the obtained RNA samples were of consistent quality and that the RNA samples contained small RNAs of sufficient quality and quantity that enabled us to perform miRNA microarray expression study. 8 RNA quantity and purity were determined using the NanoDrop ND-2000 (NanoDrop Technologies, Wilmington, DE, USA), and RNA integrity (as RNA integrity number, RIN) was assessed using the RNA 6000 Nano Chip on a Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA, USA). The isolated RNA was analyzed using the Human Male Infertility RT 2 Profiler PCR Array (Cat# PAHS-165ZC; Qiagen), which profiles the expression of 84 key gene transcripts detected in the male germline that are known to be involved in different fertility-and sperm-related processes in the cell, including spermatogenesis, fertilization, male sex differentiation, cell motility, cell cycle and response to stress.…”

Blood plasma miR‐20a‐5p expression as a potential non‐invasive diagnostic biomarker of male infertility: A pilot study

“…In addition to the classical post‐transcriptional control by RNA‐binding proteins, some studies have demonstrated that small non‐coding micro‐RNAs (miRNAs) may have a crucial role in regulating different stages of spermatogenesis 9‐11 …”

“…Most studies on miRNAs involve animal models and only a few investigate miRNAs expression in humans with different histopathological patterns of testicular dysfunction 11,13 …”