What question did this study set out to answer?

The research investigates how thrombospondin-1 (THBS1) contributes to the dysfunction of granulosa cells in polycystic ovary syndrome (PCOS).

June 4, 2026Open Access

THBS1 Induces Dysfunction of Ovarian Granulosa Cells in Patients with Polycystic Ovary Syndrome by Activating the TGF-β/Smad Pathway

Key Points

The research investigates how thrombospondin-1 (THBS1) contributes to the dysfunction of granulosa cells in polycystic ovary syndrome (PCOS).
21 PCOS patients and 21 controls' granulosa cells and follicular fluids were analyzed for THBS1 expression.
AAV9-mediated THBS1 knockdown was conducted in a DHEA-induced rat model of PCOS to observe changes.
In-vitro studies using KGN cells evaluated the effects of THBS1 on cell proliferation, apoptosis, and steroid hormone production.
THBS1 levels were higher in PCOS subjects and correlated with increased serum AMH/LH (p = 0.006).
THBS1 knockdown in rats improved cystic morphology and restored estrous cyclicity, normalizing hormone levels.
Overexpressed THBS1 in KGN cells reduced proliferation, triggered apoptosis, and disturbed steroidogenic enzyme activity.

Abstract

Objective: This study aims to investigate the role of thrombospondin-1 (THBS1) in polycystic ovary syndrome (PCOS) pathogenesis and its mechanism in regulating granulosa cell (GC) function. Methods: Follicular fluid and granulosa cells from 21 PCOS patients and 21 age-matched non-PCOS controls were analysed for THBS1 expression and clinical correlations. A dehydroepiandrosterone (DHEA) -induced PCOS rat model with adeno-associated virus serotype 9 (AAV9) -mediated THBS1 knockdown was used to assess phenotypic changes. The KGN human granulosa-like cell line was employed to evaluate THBS1 overexpression effects on proliferation, apoptosis, and steroidogenesis. Mechanistic studies included RNA sequencing with Gene Set Enrichment Analysis (GSEA), co-immunoprecipitation, molecular docking against the latent TGF-β1 crystal structure (PDB 9VJJ), molecular dynamics simulation, an active/total TGF-β1 ELISA, and pharmacological TGF-β receptor inhibition. Results: THBS1 was elevated in PCOS follicular fluid and granulosa cells and correlated positively with serum AMH and LH after Benjamini–Hochberg FDR correction. AAV9-mediated ovarian THBS1 knockdown (37. 4% protein reduction, p = 0. 006) ameliorated cystic morphology, restored estrous cyclicity, and normalised serum AMH/LH/T. In KGN cells, THBS1 overexpression suppressed proliferation, induced apoptosis and inflammatory cytokines, and dysregulated steroidogenic enzymes. Transcriptome analysis revealed upregulation of canonical TGF-β/Smad pathway components (SERPINE1, SMAD7, TGFB2, INHBA, CCN2, COL1A1/2). Molecular docking and 100-ns dynamics simulation supported a stable interaction between THBS1 and latent TGF-β1 (ΔGTOTAL ≈ −120 kcal·mol−1). Co-immunoprecipitation confirmed physical association in cells, and ELISA showed elevated TGF-β1 in PCOS follicular fluid and rat serum, both attenuated by THBS1 knockdown. Pharmacological TGF-β receptor inhibition with SB-431542 rescued THBS1-induced cellular dysfunction. Conclusions: THBS1 is associated with PCOS-related granulosa cell dysfunction through the TGF-β/Smad pathway and represents a candidate biomarker and exploratory therapeutic target that warrants validation in independent multicentre cohorts.

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