DNA Flow cytometric analysis of the human testicular tissues to investigate the status of spermatogenesis in azoospermic patients

Arka Baksi,Rajan R Dighe,S S Vasan

doi:10.1038/s41598-018-29369-8

Arka Baksi, Rajan R Dighe + Show 1 more

Open Access

https://doi.org/10.1038/s41598-018-29369-8

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Journal: Scientific Reports	Publication Date: Jul 24, 2018
Citations: 9	License type: open-access

Affiliation: Indian Institute of Science Bangalore

Abstract

A single, rapid and reproducible diagnostic test to predict the type of azoospermia and outcome of sperm retrieval is not yet available. So the feasibility of employing DNA flow cytometry for rapid investigation of the status of spermatogenesis in the patients with azoospermia was investigated. Testicular biopsies of 44 patients with azoospermia undergoing sperm-retrieval surgery and 4 controls were analyzed by flow cytometry to ascertain their testicular germ-cell patterns. The observed germ-cell pattern was further confirmed by RT-PCR analysis of the cell-specific markers and histology for some patients. The patients with Obstructive Azoospermia (OA) exhibited normal spermatogenesis similar to the control fertile patients showing the presence of diploid, double-diploid and haploid cells. The non-obstructive azoospermia (NOA) patients exhibited disrupted spermatogenesis with arrest at the pre-meiotic (only diploid cells present) or meiotic (diploid and double-diploid cells present) stages. The germ-cell pattern, as ascertained by flow cytometry, provided a clear picture of the intra-testicular spermatogenesis and the presence of spermatozoa in the patients’ testes, which was prognostic of their sperm-retrieval. DNA flow cytometry test to ascertain the testicular germ-cell pattern is simple in execution, analysis and interpretation, requires small amount of tissue and provides quantitative data about the status of spermatogenesis in patients. This test would allow comparable analysis of the status of spermatogenesis in patients across clinics and may form the basis for deciding future treatment and intervention strategies.

Highlights

Azoospermia, a type of male infertility associated with absence of measurable spermatozoa in the semen, accounts for 20–30% of all male infertility cases[1] affecting about 1% of the male population[2,3]
Fourteen patients (Group I), who suffered from Obstructive Azoospermia (OA), showed three distinct germ cell peaks clearly indicating that spermatogenesis in these patients was complete and equivalent to the control group of patients
The results described above clearly show that flow cytometric test can be employed to rapidly establish the status of intra-testicular spermatogenesis in patients with azoospermia, as well as, to predict the exact nature of the arrest in spermatogenesis

Summary

Introduction

Azoospermia, a type of male infertility associated with absence of measurable spermatozoa in the semen, accounts for 20–30% of all male infertility cases[1] affecting about 1% of the male population[2,3]. Analysis of several pre-operative variables including FSH and LH levels, testicular size and volume have been employed to diagnose the type of azoospermia and predict the outcome of sperm retrieval. These factors have low sensitivity and specificity in predicting the success of sperm retrieval[14,15]. We explore the feasibility of employing DNA flow cytometry for rapid and accurate investigation of the germ cell status of the individual patients and show that the germ cell pattern accurately reflects the status of spermatogenesis and is predictive of the success of sperm retrieval post-surgery

Methods

Results

Conclusion