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The poly(A)-binding protein genes, EPAB, PABPC1, and PABPC3 are differentially expressed in infertile men with non-obstructive azoospermia

  • Gamete Biology
  • Published:
Journal of Assisted Reproduction and Genetics Aims and scope Submit manuscript

Abstract

Purpose

Azoospermia is one of the major causes of male infertility and is basically classified into obstructive (OA) and non-obstructive azoospermia (NOA). The molecular background of NOA still largely remains elusive. It has been shown that the poly(A)-binding proteins (PABPs) essentially play critical roles in stabilization and translational control of the mRNAs during spermatogenesis.

Methods

In the present study, we aim to evaluate expression levels of the PABP genes, EPAB, PABPC1, and PABPC3, in the testicular biopsy samples and in the isolated spermatocyte (SC) and round spermatid (RS) fractions obtained from men with various types of NOA including hypospermatogenesis (hyposperm), RS arrest, SC arrest, and Sertoli cell-only syndrome (SCO).

Results

In the testicular biopsy samples, both PABPC1 and PABPC3 mRNA expressions were gradually decreased from hyposperm to SCO groups (P < 0.05), whereas there was no remarkable difference for the EPAB expression among groups. The expression levels of cytoplasmically localized PABPC1 and PABPC3 proteins dramatically reduced from hyposperm to SCO groups (P < 0.05). In the isolated SC and RS fractions, the EPAB, PABPC1, and PABPC3 mRNA expressions were gradually decreased from hyposperm to SC arrest groups (P < 0.05). Similarly, both PABPC1 and PABPC3 proteins were expressed at higher levels in the SC and RS fractions from hyposperm group when compared to the SC and RS fractions from either RS arrest or SC arrest group (P < 0.05).

Conclusion

Our findings suggest that observed significant alterations in the PABPs expression may have an implication for development of different NOA forms.

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Acknowledgments

The authors thank Deniz Ozel-Erkan, M.Sc. for her contribution to statistical analysis of data, and thank Asli Uyar, Ph.D. for critically reading the article.

Authors’ roles

S.O. and N.D. designed the study. S.O., B.S., and F.U. created all data, and S.O. wrote the article. I.C.B. performed pathological analysis of the testicular biopsies. M.F.U. provided the testicular biopsy samples. N.D. and G.A. critically read the article.

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Authors and Affiliations

  1. Department of Histology and Embryology, Akdeniz University, School of Medicine, Campus, 07070, Antalya, Turkey

    Saffet Ozturk, Berna Sozen, Fatma Uysal, Gokhan Akkoyunlu & Necdet Demir

  2. Department of Pathology, Akdeniz University, School of Medicine, Campus, 07070, Antalya, Turkey

    Ibrahim C. Bassorgun

  3. Department of Urology, Akdeniz University, School of Medicine, Campus, 07070, Antalya, Turkey

    Mustafa F. Usta

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  1. Saffet Ozturk
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Correspondence to Necdet Demir.

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Funding

This study was supported by TUBITAK (Grant No. 112S169).

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The authors declare that there is no conflict of interest.

Additional information

Capsule The EPAB, PABPC1 and PABPC3 proteins seem to be important RNA-binding proteins in regulating translation control of the mRNAs in spermatogenic cells during spermatogenesis.

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Supplementary Fig. 1

Representative image of hematoxylin and eosin (HE)-stained sections of the NOA groups. The HE-stained sections of each group were analyzed under a bright-field microscopy, and thereby their histopathological characteristics and Johnsen scores were determined. Classification of the testicular biopsy samples into different non-obstructive azoospermia (NOA) groups were performed based on their spermatogenic activity and germinal epithelial cell content, and they were accordingly separated into groups as follows: a hypospermatogenesis, b round spermatid arrest, c spermatocyte arrest, and d Sertoli cell-only syndrome. SG spermatogonia, SC spermatocyte, RS round spermatid, ES elongated spermatid, SE Sertoli cell, IT intertubular area, L lumen. Scale bar: 50 μm. (JPG 1.39 MB)

Supplementary Fig. 2

Localization of the EPAB transcript in the human testis tissue. We have detected EPAB mRNA localization in the testicular biopsy samples obtained from men with hyposperm. For this purpose, EPAB probe specific to exons 1 and 2 was produced as the following processes: RNA isolation, cDNA synthesis, and PCR amplification. a A representative photomicrograph of the RNA in situ hybridization on the testis section from hyposperm group. We observed that EPAB mRNA was cytoplasmically expressed in the spermatogenic cells as well as in the intertubular somatic cells. b A representative photomicrograph of the negative control section. As expected, there was no reaction for the EPAB unlabeled probe in the negative control section. SG spermatogonia, SC spermatocyte, RS round spermatid, ES elongated spermatid, SE Sertoli cell, IT intertubular area, L lumen. Scale bar: 50 μm. (JPG 1.49 MB)

Supplementary Fig. 3

Particular features of the isolated SC and RS fractions from hyposperm, RS arrest, and SC arrest groups using 2–4 % bovine serum albumin (BSA) gradient. Representative photomicrographs of the SC (a) and RS (b) fractions were captured by an inverted microscopy (Olymus) at original magnification of ×400. Scale bar: 50 μm. To determine purity percent of the isolated spermatocyte and round spermatid fractions, their smear slides were air-dried, and then stained with hematoxylin-eosin. Representative photomicrographs of hematoxylin-eosin-stained SC (c) and RS (d) fractions were taken at original magnification of ×200; inserts were at ×1000 magnification. Scale bar: 50 μm. e The HE-stained slides were examined with the bright-field microscope (Zeiss) to determine purity percent of the isolated SC and RS fractions, which were identified based on their morphological characteristics. In general, the purity rates of isolated spermatocyte and round spermatid fractions were in usable levels (>77 %); however, the RS fraction obtained from RS arrest group was found to be in low levels (∼24 %). SG spermatogonia, SC spermatocyte, RS round spermatid, SE Sertoli cell, OC other cell, Hyposperm hypospermatogenesis, RS arrest round spermatid arrest, SC arrest spermatocyte arrest. Data are presented as mean ± standard deviation (SD). (JPG 1.76 MB)

Supplementary Table 1

Sequences and localizations of primers for EPAB, PABPC1, and PABPC3 genes used in the current study. F, Forward; R, Reverse; bp, base pair. (JPG 690 KB)

Supplementary Table 2

Histopathologic features of the testicular biopsy samples obtained from various types of NOA groups. J.S., Johnsen score. (JPG 2.30 MB)

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Ozturk, S., Sozen, B., Uysal, F. et al. The poly(A)-binding protein genes, EPAB, PABPC1, and PABPC3 are differentially expressed in infertile men with non-obstructive azoospermia. J Assist Reprod Genet 33, 335–348 (2016). https://doi.org/10.1007/s10815-016-0654-z

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