Analysis of ultrastructural defects in sperm by transmission electron microscopy in asthenozoospermia patients: a study from multiple centers across China

Abstract

This study aimed to identify the ultrastructural features of sperm in Chinese asthenozoospermia patients and to evaluate their clinical implications. A total of 139 individuals, including 106 asthenozoospermia patients and 33 fertile controls, were recruited from multiple centers across China. Sperm ultrastructural defects were examined using transmission electron microscopy (TEM), while conventional sperm quality was assessed by light microscopy. Compared with the control group, the asthenozoospermia group showed significant ultrastructural defects, particularly in the sperm tail, including mitochondrial and axonemal defects. Based on tail ultrastructure, participants were further categorized into four groups (normal ultrastructure, simple abnormal axonemes, simple abnormal mitochondria, and both abnormality in axonemes and mitochondria). All three abnormal groups indicated significantly lower value in sperm (motile/kinematic) quality compared to the normal group. However, no statistically significant differences in sperm quality were observed among the three abnormal groups. These nationwide findings indicated that TEM could remedy the limitations of conventional light microscopy, which is difficult to localize organelle-specific defects. This capability may ultimately inform individualized diagnostic and therapeutic strategies.

Keywords: Asthenozoospermia patient; Sperm ultrastructure; Transmission electron microscopy.

Fig. 1

Electron micrographs of typical morphological features of human sperm. (A) Electron micrographs of normal morphological features in human sperm: (a) mode diagram of the typical morphological features of human sperm; (b) longitudinal section of human sperm: the normal sperm had a symmetrical mid-piece with smooth axoneme surrounded by regularly arranged mitochondria; (c) sperm head (longitudinal section): normal chromatin condensation (Ch), no vacuoles (simple membrane vacuoles or vacuoles with membrane whorls) or small membrane vacuoles occupying a little part of nucleus, the acrosome covers approximately two-thirds of the head without swelling, and the acrosome membrane is intact (Ac); (d) neck of sperm tail (longitudinal section): the neck region appears conserved, with the well-defined centriole (Ce); (e) midpiece of sperm tail (cross-section), the typical axoneme and peri-axoneme structure: the axoneme mainly comprises a “9 + 2” structure, including nine pairs of peripheral doublet microtubules (AX, upper arrow) and a central pair of microtubules (AX, lower arrow). The peri-axoneme structure includes the nine dense fibers (Df), and helical mitochondria (M); (f) principal piece of sperm tail (cross-section), the typical axoneme and peri-axoneme structure: the axoneme mainly comprises a “9 + 2” structure, including nine pairs of peripheral doublet microtubules (AX, upper arrow) and a central pair of microtubules (AX, lower arrow). The peri-axoneme structure includes the nine dense fibers (Df), and the fiber sheath (Fs); (g) endpiece of sperm tail (cross-section), the typical axoneme structure: the axoneme mainly comprises a “9 + 2” structure, including nine pairs of peripheral doublet microtubules and a central pair of microtubules (AX, lower arrow). (B) Electron micrographs of sperm head defects: (a) longitudinal section: trinucleated head (*); (b) longitudinal section: altered shape of the head, severe vacuolar defect of the chromatin (*), granular chromatin; (c) longitudinal section: severe vacuolar defect of the chromatin (*); (d) longitudinal section: partly granular chromatin (triangle); (e) longitudinal section: a displaced acrosomal vesicle (arrow). (C) Electron micrographs of sperm tail defects: (a) longitudinal section: cytoplasmic residues (arrow), the midpiece is enlarged with aggregates of misaligned pale mitochondria; (b) cross-section: supernumerary axonemes (arrow) in midpiece; (c) longitudinal section: number of mitochondria is increasing, swollen, irregular in shape (arrow) in midpiece; (d) longitudinal section: mitochondria are absent or mitochondria are scarce (arrow) in midpiece; (e) cross-section: a supernumerary dense fiber (arrow) in midpiece; (f) cross-section: translocation of axonemes and dense fibers, absence of fibrous sheath in principal piece; (g) cross-section: misassembled axonemal structures (arrow) in endpiece. Ac: Acrosomes, Ax: Axonemes (typical structure of “9 + 2” core axoneme), Ce: Centriole, Ch: Chromatin, Df: Dense fibers, Fs: Fibrous sheath, M: Mitochondria

Fig. 2

Differences of baseline characteristics and sperm parameters among normal ultrastructure group, simple abnormal axonemes group, simple abnormal mitochondria group, and both abnormality group. ALH, amplitude of lateral head displacement; BAAM, both abnormity in axonemes and mitochondria; BCF, beat cross frequency; BMI, body mass index; LIN, linearity; MAD, mean angular displacement; NSM, normal sperm morphology; NU, normal ultrastructure; PSM, progressive sperm motility; SAA, simple abnormal axonemes; SAM, simple abnormal mitochondria; SC, sperm concentration; STR, straightness; SV, semen volume; TSC, total sperm count; TSM, total sperm motility; VAP, velocity average path; VCL, velocity curvilinear; VSL, velocity straight line; WOB, wobble. *: P < 0.05, **: P < 0.01, ***: P < 0.005.