Hormonal regulation of leydig cell proliferation and differentiation

Abstract

Regulation of Leydig Cell Proliferation and Function by LH and FSH in Rats

Abstract and Synopsis Introduction Leydig cells are the source of testosterone, essential for male phenotype, reproductive tract development, and spermatogenesis. Located in the interstitial compartment of the testis, Leydig cells are primarily regulated by Luteinizing Hormone (LH), which stimulates steroidogenic enzyme expression and testosterone production. Sertoli cell-derived factors, influenced by Follicle Stimulating Hormone (FSH) and testosterone, also play roles in Leydig cell regulation.

In rats, Leydig cells undergo a three-stage differentiation process:

Progenitor Leydig Cells (PLCs) (day 14 postnatal)

Immature Leydig Cells (ILCs) (day 28)

Adult Leydig Cells (ALCs) (day 60)

LH binding capacity, smooth endoplasmic reticulum, and testosterone production increase with differentiation.

Objectives

Investigate the role of LH in Leydig cell proliferation and early differentiation.

Evaluate the role of FSH in Leydig cell function during different developmental stages.

Use passive neutralization approaches with specific antisera to LH and FSH in immature and EDS-treated rat models.

Methods

Isolation of Leydig cells: Collagenase dispersion and Percoll density fractionation. Purity assessed by histochemical staining for 3 -hydroxysteroid dehydrogenase.

Antiserum production: Ovine LH and FSH purified, immunized in bonnet monkeys, titers assessed by binding assays. Neutralization confirmed by testosterone assays.

Proliferation assays: Cyclin D proteins, PCNA levels (Western blot), and BrdU incorporation (ELISA).

Gene expression: Semi-quantitative RT-PCR for IGF-1, IGF-1 receptor, StAR, aromatase, and steroidogenic enzymes.

Key Findings

LH and Leydig Cell Proliferation

PLCs (day 21) showed maximum DNA synthesis, highest PCNA and Cyclin D3 levels.

LH deprivation abolished Cyclin D3 expression and reduced PCNA, confirming LH’s role in proliferation.

LH stimulates proliferation via IGF-1 signaling, as IGF-1 mRNA and receptor levels decreased with LH deprivation.

FSH and Leydig Cell Function

In neonatal rats, FSH neutralization increased testicular testosterone and steroidogenic enzyme mRNA, suggesting FSH normally inhibits Leydig cell function via estradiol regulation.

In immature/adult rats, FSH neutralization decreased steroidogenic capacity, confirming a positive role for FSH in Leydig cell function.

Recombinant FSH increased testosterone production, while FSH receptor antiserum reduced it.

FSH regulates Leydig cells indirectly via Sertoli cell-derived IGF-1.

EDS-Treated Rat Model

EDS destroyed Leydig cells, followed by regeneration resembling postnatal development.

LH neutralization blocked Leydig cell maturation and testosterone synthesis, but precursor cell repopulation occurred independently of LH.

FSH neutralization reduced testosterone production, steroidogenic enzyme expression, and spermatogenesis, confirming its role in Leydig cell function.

Conclusions LH is indispensable for Leydig cell proliferation, acting via IGF-1 signaling.

FSH plays a dual role: inhibitory in neonatal rats (via estradiol regulation) but stimulatory in immature and adult rats (via Sertoli cell-derived IGF-1).

Leydig cell repopulation after EDS occurs independently of LH, but maturation requires LH.

Both LH and FSH are critical regulators of Leydig cell function and spermatogenesis.