P-104 production and glucose-dependent secretion of insulin by human spermatozoa
Authors: Carrageta D , Oliveira P.F , Barros A , Monteiro M.P , Alves M.G 
Published: June 2022

Abstract

Study question
Do human spermatozoa express insulin and the enzymes responsible for the cleavage of proinsulin? Is the secretion of insulin by spermatozoa responsive to glucose stimuli?

Summary answer
Human spermatozoa express insulin and PC1/3 and PC2, enzymes responsible for the cleavage of proinsulin. Spermatozoa produce and release insulin in a glucose concentration-dependent manner.

What is known already
Spermatogenesis is dependent on a finely regulated hormonal and metabolic control. Insulin is considered one of the most important regulators in the metabolic regulation of spermatogenesis, acting directly on the differentiation of spermatogonia into primary spermatocytes or indirectly through the metabolic modulation of both Sertoli and Leydig cells. In addition, insulin is hypothesized to play a major role in human spermatozoa capacitation, although the mechanisms that regulate its production and secretion remain to be disclosed.

Study design, size, duration
The main objective of this study was to estimate the capacity of insulin production by human spermatozoa and if they express PC1/3 and PC2, which are enzymes responsible for the cleavage of proinsulin. In addition, we hypothesized that insulin production could respond to glucose stimuli. Seminal samples of normozoospermic men (n = 15) seeking fertility treatment were collected for these experiments.

Participants/materials, setting, methods
Spermatozoa were submitted to a density gradient centrifugation and two fractions were collected according to their motility (high vs low motility). The expression of insulin, PC1/3 and PC2 mRNA was evaluated by RT-qPCR. The protein expression of insulin, PC1/3 and PC2 was evaluated by immunofluorescence. High motility spermatozoa were incubated in BWW medium with increasing glucose concentrations (0, 5.5, 11, and 22 mM). After 6 h, the concentration of secreted insulin was quantified by ELISA.

Main results and the role of chance
We were able to identify the expression of insulin, PC1/3, and PC2 mRNA, as well as the respective proteins, in human spermatozoa. Through immunofluorescence, we observed that insulin and both enzymes are mainly expressed in the midpiece. Human spermatozoa also express insulin receptor, mainly in the midpiece and tail. The mRNA expression of insulin and PC1/3 was found to be higher in the highly motile spermatozoa. Human spermatozoa produce very small quantities of insulin (pg/mL range), which suggests an autocrine or paracrine action. The release of insulin to the medium was found to occur in a glucose concentration-dependent manner.

Limitations, reasons for caution
This is an in vitro study with relatively small sample size. Future studies are required to further evaluate the role of insulin and its signalling pathway on human spermatozoa capacitation and fertilization capacity.

Wider implications of the findings
This study shows that human spermatozoa express PC1/3 and PC2, enzymes responsible for the cleavage of proinsulin. In addition, human spermatozoa release insulin in a glucose concentration-dependent manner. The study of insulin signalling opens the path for a deeper understanding of human spermatozoa bioenergetics and the development of novel therapies.

Trial registration number
Not applicable