Fig. 1: Sialokinin modulates monocyte activation via NK receptors and the PI3K/Akt signaling pathway.

a RNA-Seq profiling of primary human monocytes (n = 3) at 24 h post-treatment. Volcano plot indicates differentially expressed genes (DEGs) between sialokinin-treated vs. non-treated monocytes. DEG analysis was performed using edgeR, with multiple testing corrected via the Benjamini–Hochberg FDR method. Significantly upregulated and downregulated genes are shown in red and blue, respectively. b Flow cytometry shows reduced surface expression of Siglec-1/CD169 in sialokinin-treated monocytes compared with untreated monocytes (n = 10 biological replicates) at 24 h post-treatment. Data were analyzed by paired two-tailed t-test. c Pearson correlation matrix for the expressions of 12 significant DEGs from RNA-Seq data (n = 3). d Western blot analysis shows that sialokinin stimulation activated the PI3K-Akt pathway in human monocytes, with increased levels of phosphorylated PI3K (P-PI3K) and Akt (P-Akt) at 20, 40, and 60 min. Pre-treatment with either the NK₁R inhibitor CP-96345 or the NK₂R inhibitor GR-159897 led to inhibition of PI3K and Akt as early as 20 min post-stimulation. Relative expression levels of P-PI3K/PI3K (n = 4 biological replicates) and P-Akt/Akt (n = 5 biological replicates) in monocytes at the 20-min time point were quantified with ImageJ. Levels are expressed as fold change relative to the non-treated controls. Bar graphs represent the mean ± standard deviation (SD). Statistical comparisons between sialokinin-treated sample and untreated control were performed using paired two-tailed t-test. e Schematic representation of CD169 downregulation in response to sialokinin via NK receptor/PI3K/Akt activation. f The monocytes were pre-treated with the NK₁R antagonist (CP-96345; n = 5 biological replicates), NK₂R antagonist (GR-159897; n = 5 biological replicates), both antagonists in combination (n = 4 biological replicates), or the PI3K inhibitor (LY-294002; n = 5 biological replicates) prior to sialokinin stimulation. CD169 levels at 24 h post-treatment were measured by mean fluorescence intensity (MFI) and expressed as fold change relative to non-treated controls. Statistical comparisons were performed using paired two-tailed t-test or Wilcoxon signed-rank test, depending on data distribution.