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. 2010 Sep;24(5):404-10.
doi: 10.1016/S1674-8301(10)60054-1.

Evaluation of sperm mitochondrial function using rh123/PI dual fluorescent staining in asthenospermia and oligoasthenozoospermia

Tiejun Zou  1 , Xiang Liu, Shangshu Ding, Junping Xing
Affiliations

Evaluation of sperm mitochondrial function using rh123/PI dual fluorescent staining in asthenospermia and oligoasthenozoospermia

Tiejun Zou et al. J Biomed Res. 2010 Sep.

Abstract

Objective: The recent advent of flow cytometry (FCM), coupled with fluorescent dyes, has been successfully applied to assess mitochondrial function. The aim of this study was to investigate the feasibility and clinical significance of detecting sperm mitochondrial function and to evaluate sperm mitochondrial function by using Rhodamine 123/propidium (Rh123/PI) dual fluorescent staining and FCM in asthenospermia and oligoasthenozoospermia.

Methods: Twenty-five fertile men (with normal sperm parameters) and 230 infertile patients were examined. Fifty-five patients of the above 230 patients were selected for idiopathic infertility samples and were divided into two groups: asthenospermia (n = 30) and oligoasthenozoospermia (n = 25). Rh123/PI dual fluorescent staining and FCM were carried out to examine sperm mitochondrial function.

Results: Significant differences were found between the normal and abnormal semen samples (P < 0.05) when Rh123(+)/PI(-), Rh123(-)/PI(+) and Rh123(-)/PI(-) sperm were examined by FCM, but there was no significant difference between the asthenospermia (P = 0.469) and oligoasthenozoospermia group (P = 0.950) when Rh123(+)/PI(-) and Rh123(-)/PI(+) sperm were then examined; however, a significant difference was found between the 2 groups (P = 0.003) when Rh123(-)/PI(-) sperm were examined. There was no correlation between Rh123(-)/PI(-) sperm and semen parameters in the normal group, but there was a significant negative correlation between the sperm concentration and Rh123(-)/PI(-) sperm in asthenospermia and oligoasthenozoospermia patients (r = -0.509, -0.660; P = 0.018, 0.038).

Conclusion: Rh123/PI dual fluorescent staining and FCM can provide reliable information to assess the quality of sperm and reveal differences in mitochondrial membrane potential in asthenospermia and oligoasthenozoospermia.

Keywords: Rhodamine 123/propidium double fluorescent staining; asthenospermia; flow cytometry; mitochondrial membrane potential; oligoasthenozoospermia.

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Figures

Fig. 1
Fig. 1. Sperm fluorescent staining.
A: After staining with Rh123, green fluorescence was centralized on the sperm body under fluorescence microscope, suggesting that the mitochondria functioned well (×400); B: After double fluorescent staining with Rh123/PI, dead sperms from asthenospermia patients were marked by red heads (×400).
Fig. 2
Fig. 2. Results of sperm FCM.
The lower right quadrant of each graph, Rh123+/PI−, shows the sperm with normal mitochondrial function; the lower left quadrant shows Rh123−/PI−, which means the sperm losing mitochondrial function; the upper left quad-rant, Rh123−/PI+, shows necrotic sperm. A: normal seminal sample. B: the asthenospermia group. C: the oligoasthenozoospermia group.
See this image and copyright information in PMC

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