Hypoxic-ischemic brain damage (HIBD) is a common cause of death or mental retardation in newborns. Ferroptosis is a novel form of iron-dependent cell death driven by lipid peroxidation, and recent studies have confirmed that ferroptosis plays an important role in the development of HIBD. However, HIBD ferroptosis-related biomarkers remain to be discovered. An artificial neural network (ANN) was established base on differentially expressed genes (DEGs) related to HIBD and ferroptosis and validated by external dataset. The protein–protein interaction (PPI) network, support vector machine-recursive feature elimination (SVM-RFE) algorithms, and random forest (RF) algorithm were utilized to identify core genes of HIBD. An in vitro model of glutamate-stimulated HT22 cell HIBD was constructed, and glutamate-induced ferroptosis and mitochondrial structure and function in HT22 cells were examined by propidium iodide (PI) staining, flow cytometry, Fe²⁺ assay, Western blot, JC-1 kit, and transmission electron microscopy (TEM). In addition, Western blot and immunofluorescence assays were used to detect the NF-κB/STAT3 pathway. An HIBD classification model was constructed and presented excellent performance. The PPI network and two machine learning algorithms indicated two hub genes in HIBD. Lipocalin 2 (LCN2) was the core gene correlated with the risk of HIBD according to the results of differential expression analysis and logistic regression diagnostics. Subsequently, we verified in an in vitro model that LCN2 is highly expressed in glutamate-induced ferroptosis in HT22 cells. More importantly, LCN2 silencing significantly inhibited glutamate-stimulated ferroptosis in HT22 cells. We also found that glutamate-stimulated HT22 cells produced mitochondrial dysfunction. Furthermore, in vitro experiments confirmed that NF-κB and STAT3 were activated and that silencing LCN2 could have the effect of inhibiting their activation. In short, our findings reveal a molecular mechanism by which LCN2 may promote ferroptosis in HIBD through activation of the NF-κB/STAT3 pathway, providing new and unique insights into LCN2 as a biomarker for HIBD and suggesting new preventive and therapeutic strategies for HIBD.

Fucoidan is a sulfated algal polyanionic polysaccharide that possesses many biological activities. In this paper, a fucoidan (SHF) polysaccharide was extracted from Sargassum hemiphyllum collected in the South China Sea. The SHF, with a molecular weight of 1166.48 kDa (44.06%, w/w), consisted of glucose (32.68%, w/w), galactose (24.81%, w/w), fucose (20.75%, w/w), xylose (6.98%, w/w), mannose (2.76%, w/w), other neutral monosaccharides, and three uronic acids, including glucuronic acid (5.39%, w/w), mannuronic acid (1.76%, w/w), and guronuronic acid (1.76%, w/w). The SHF exhibited excellent immunostimulatory activity. An immunostimulating assay showed that SHF could significantly increase NO secretion in macrophage RAW 264.7 cells via upregulation of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) levels based on both gene expression and protein abundance. These results suggest that SHF isolated from Sargassum hemiphyllum has great potential to act as a health-boosting ingredient in the pharmaceutical and functional-food fields.

• A heteropolysaccharide Sargassum Zhangii fucoidan (SZF) has an Mw of 111.28 kDa. • The main monosaccharides of SZF include galactose, fucose, xylose, and uronic acids. • SZF possesses a main chain of (1,4)-α-d-linked-galactose, (3,4)-α-l-fucose, etc. • SZF has significant immune-stimulating properties compared to commercial fucoidans. • SZF has great potential to act as ingredient of functional food or immunodulator.