Serum cholesterol levels define an inverted u-shaped relationship with PBSC mobilization and immune balance in healthy donors

Abstract Background: Peripheral blood stem cell (PBSC) mobilization with granulocyte colony-stimulating factor (G-CSF) is the standard approach for hematopoietic stem cell transplantation. However, considerable inter-individual variability exists in mobilization efficiency, and the underlying metabolic determinants remain incompletely understood. Recent evidence suggests a potential link between lipid metabolism and hematopoietic regulation, but the role of serum cholesterol in modulating mobilization outcomes remains controversial. This study aimed to investigate the impact of cholesterol levels on PBSC mobilization efficacy and immune cell dynamics in healthy donors. Methods:A retrospective analysis was conducted on adult donors who underwent PBSC mobilization and collection at our center. Univariate analyses were performed using the Mann–Whitney U test and Spearman rank correlation, followed by simple and multiple linear regression to identify independent predictors. Single-cell RNA sequencing(scRNA-seq) was employed to explore pathway differences among three donor groups stratified by cholesterol levels, and findings were validated by quantitative PCR (qPCR) and Western blotting (WB). Results: A total of 643 healthy donors were enrolled. Both univariate and multivariate analyses identified cholesterol as an independent predictor of stem cell mobilization (β = −566.77, P = 0.017). Donors were stratified into three groups based on cholesterol levels, the moderate-cholesterol group (TC: 4.20–5.20 mmol/L) demonstrated:optimal CD34+ cell mobilization (P = 0.020 vs. low group); significantly higher CD34+CD38- primitive stem cells in apheresis products (vs. low group, P < 0.05). And donors with moderate cholesterol levels exhibit more balanced immune cell regulation and a superior response to G-CSF during hematopoietic stem cell mobilization. The scRNA-seq sequencing revealed upregulated expression of CXCR4 in the intermediate group, which was further confirmed by qPCR and WB at transcriptomic and protein levels. Conclusions: Cholesterol level is a essential factor influencing PBSC mobilization in healthy donors. An appropriate cholesterol level to enhance CXCR4 expression, thereby improving mobilization efficiency. Keywords: Peripheral blood stem cells; Stem cell mobilization; Cholesterol; CXCR4; Single-cell RNA sequencing.