The aim of this study is to explore the effect of tumor necrosis factor‐α (TNF‐α) inhibition in rats with neonatal hypoxic–ischemic encephalopathy (HIE) and ascertain the relevant signaling pathways. The Zea–Longa score was used to evaluate the neurological function of the rats. ImageJ was used for quantification of the brain edema volume. Triphenyl tetrazolium chloride (TTC) staining of brain tissue was performed 24 h after hypoxic–ischemic (HI) to detect right brain infarction. The expression of TNF‐α was detected by quantitative reverse transcription polymerase chain reaction (qRT‐PCR). Immunofluorescence staining was used to identify the localization of TNF‐α; Then, the effective shRNA fragment of TNF‐α was used to validate the role of TNF‐α in HIE rats, and the change of neurotrofin‐3 (NT‐3) and tyrosine kinase receptor‐C (TRKC) was examined after TNF‐α‐shRNA lentivirus transfection to determine downstream signaling associated with TNF‐α. Protein interaction analysis was carried out to predict the links among TNF‐α, NT‐3, and TRKC. Cerebral edema volume and infarction increased in the right brain after the HI operation. The Zea–Longa score significantly increased within 24 h after the HI operation. The relative expression of TNF‐α was upregulated after the HI operation. TNF‐α was highly expressed in the right hippocampus post HI through immunofluorescence staining. Bioinformatics analysis found a direct or an indirect link among TNF‐α, NT‐3, and TRKC. Moreover, the interference of TNF‐α increased the expression of NT‐3 and TRKC. TNF‐α interference might alleviate brain injury in HIE by upregulating NT‐3 and TRKC.

This study aimed to decipher the effect of glycoprotein nonmetastatic melanoma protein B (GPNMB) on neonatal hypoxic-ischemic encephalopathy (NHIE) and its potential molecular mechanism. The hypoxic-ischemic (HI) model was established in 7-day-old rats, and then, Zea-Longa scores and Nissl staining were performed to measure brain damage post-HI. In addition, gene sequencing was used to detect the differential expression genes (DEGs), and then, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes databases were used to determine the function of DEGs. Furthermore, an oxygen-glucose deprivation (OGD) model was developed in SY5Y cells and human fetal neurons, and then, the level of GPNMB was verified by quantitative real-time polymerase chain reaction. In addition, methyl thiazolyl tetrazolium and cell counting kit-8 assays were applied after GPNMB interference. Finally, the alternative splicing of GPNMB expression was analyzed using Splice Grapher software. The results indicated that HI induced marked neurological impairment and neuron injury in rats. Also, GPNMB was the most obviously upregulated gene in DEGs. Additionally, GPNMB was upregulated significantly in SY5Y and fetal neurons after OGD, and GPNMB-si promoted an increase in cell viability and number. Moreover, we found that the GPNMB alternative splicing type was the Alternative 3′ splice site, with the alternative splicing site in 143382985:143404102. Herein, GPNMB promotes a crucial regulatory mechanism with alternative splicing for neuronal survival after NHIE.