Time-resolved ICP-MS analysis of mineral element contents and distribution patterns in spermatogenic cells of different types

Abstract

Mineral elements play an important role in the spermatogenesis, maturation, and fertilization of sperm. It is of great scientific significance to study the role of mineral elements in spermatogenesis by accurately measuring the content of elements in different spermatogenic cells and analyzing the distribution pattern of elements in spermatogenesis. Here, time-resolved inductively coupled plasma mass spectrometry (ICP-MS) was used to analyze the content and distribution patterns of mineral elements in spermatogenic cells of different types at the single cell level. Firstly, spermatogonia, spermatocytes, round spermatids and elongating spermatids were successfully isolated from testis of mice of different weeks of age by differential adherent method and discontinuous bovine serum albumin (BSA) density gradient method. Then, signal profiles and elemental distributions of ²⁴Mg, ³¹P, ⁵²Cr, ⁵⁵Mn, ⁵⁶Fe and ⁶⁶Zn in spermatogenic cells were obtained with dwell time at 0.1 ms. Based on the results of acid digestion, we derived a formula to calculate element content in single cell from peak area for each element, and the feasibility and universality of the formula in the quantitative detection of single cell elements were verified by sperm samples to a certain extent. The detection results of element content in single cell showed that the content of ³¹P in elongating spermatids was significantly higher than that in spermatogonia, spermatocytes and round spermatids (P < 0.01), and the distribution range was wider. However, the ⁵²Cr and ⁵⁶Fe content of elongating spermatids was lower than that of spermatogonia, spermatocytes and round spermatids (P < 0.05). When spermatogonia developed into round spermatids, the contents of ⁵⁵Mn and ⁶⁶Zn in single cell increased significantly (P < 0.05), then decreased to the lowest in elongating spermatids. In addition, the significant decrease of ⁵²Cr, ⁵⁵Mn, ⁵⁶Fe and ⁶⁶Zn content in elongating spermatids also be visually observed from the center of the fitting curve of the element signal intensity distribution moving to the left. This study provides an elemental view of the changes in elemental content at various stages of spermatogenesis at the single-cell level. Time-resolved ICP-MS is used to detect mineral elements content and distribution patterns in spermatogenic cells of testis, which is helpful to better explore the stages and modes of action of various elements in spermatogenesis, and provide direct evidence for revealing the effects of element content changes on spermatogenesis and semen quality regulation.

Keywords: ICP-MS; Mineral element; Single-cell analysis; Spermatogenic cells.