BACKGROUND:There are differentially expressed genes in acute intracerebral hemorrhage,which are related to the occurrence and development of intracerebral hemorrhage. OBJECTIVE:To screen differentially expressed genes and key genes in brain tissue of a rat model with acute intracerebral hemorrhage,to validate them through qPCR,and to analyze the relationships between key genes and the neurological function and brain tissue water content after intracerebral hemorrhage. METHODS:Seventy-eight Sprague-Dawley rats were randomly divided into two groups:in intracerebral hemorrhage group,a rat model of acute intracerebral hemorrhage was made using collagenase injection at the right caudate nucleus;and in sham-operated group,rats were injected with equal amount of saline at the same site.RNA was extracted from rat brain tissues of both groups using the TRIzol method and transcriptome sequencing technology was used to identify differentially expressed genes in brain tissues of acute intracerebral hemorrhage,which were then verified by qPCR and analyzed for the relationships between the genes and neurological function and brain tissue water content after intracerebral hemorrhage.And the key genes were analyzed by GO and KEGG functional enrichment analysis in combination with bioinformatics. RESULTS AND CONCLUSION:Ten key genes were identified,including CXCL8,SERPINE1,TFPI2,CXCR4,GDA,KCNQ5,ERICH3,SCN3B,CACNA1E,and CCL20.The contents of GDA,KCNQ5,ERICH3,SCN3B,and CACNA1E in the intracerebral hemorrhage group were lower than those in the sham-operated group(P＜0.05).The contents of CXCL8,SERPINE1,TFPI2,CXCR4 and CCL20 in the intracerebral hemorrhage group were higher than those in the sham-operated group(P＜0.05).The contents of GDA,KCNQ5,ERICH3,SCN3B,and CACNA1E were positively correlated with brain tissue water content and neurologic deficit score(P＜0.05),while the contents of CXCL8,SERPINE1,TFPI2,CXCR4 and CCL20 were negatively correlated with brain tissue water content and neurologic deficit score(P＜0.05).GO analysis indicated that differentially expressed genes were mainly enriched in two biological processes(leukocyte chemotaxis and chemokine-mediated signaling pathways),two cell components(cation channel complexes and ion channel complexes),and two molecular functions(gated channel activity and ion channel activity).KEGG analysis indicated that differentially expressed genes were concentrated in tumor necrosis factor signaling pathway,glutamatergic synapses and GABAergic synapses.To conclude,the differentially expressed genes in intracerebral hemorrhage include CXCL8,SERPINE1,TFPI2,CXCR4,GDA,KCNQ5,ERICH3,SCN3B,CACNA1E,and CCL20,and these genes are related to brain tissue water content and neurological function after intracerebral hemorrhage.These genes are mainly enriched in cell components,binding functions,cellular protrusions,and other related biological functions.

Liquid chromatography tandem mass spectrometry (LC-MS/MS) has gradually become a promising alternative to ligand binding assay for the bioanalysis of biotherapeutic molecules,due to its rapid method development and high accuracy.In this study,we established a new LC-MS/MS method for the determination of the anti-sclerostin monoclonal antibody (SHR-1222) in cynomolgus monkey serum,and compared it to a previous electrochemiluminescence method.The antibody was quantified by detecting the surrogate peptide obtained by trypsin digestion.The surrogate peptide was carefully selected by investigating its uniqueness,stability and MS response.The quantitative range of the pro-posed method was 2.00-500 μg/mL,and this verified method was successfully applied to the tox-icokinetic assessment of SHR-1222 in cynomolgus monkey serum.It was found that the concentrations of SHR-1222 in cynomolgus monkeys displayed an excellent agreement between the LC-MS/MS and electrochemiluminescence methods (ratios of drug exposure,0.8-1.0).Notably,two monkeys in the 60 mg/kg dose group had abnormal profiles with a low detection value of SHR-1222 in their individual sample.Combining the high-level anti-drug antibodies (ADAs) in these samples and the consistent quantitative results of the two methods,we found that the decreased concentration of SHR-1222 was due to the accelerated clearance mediated by ADAs rather than the interference of ADAs to the detection platform.Taken together,we successfully developed an accurate,efficient and cost-effective LC-MS/MS method for the quantification of SHR-1222 in serum samples,which could serve as a powerful tool to improve the preclinical development of antibody drugs.