The Study About Established Human Rituximab-Resistant Cell Lines. And Preliminary Exploring The Mechanisms Of Rituximab Resistance By CDNA Microarray

1、THE STUDY ABOUT ESTABLISHED HUMAN RITUXIMAB-RESISTANT CELL LINES.Objective:In order to exploring the mechanisms of rituximab resistance in vitro, we designed this study to establish human rituximab-resistant B-cell lymphoma cell line Jeko-1/R and Raji/R.Methods:The rituximab-resistant human lymphoma cell subline were established in vitro using gradually increased concentration (1μg/ml、5μg/ml、 10μg/ml、20μg/ml、50μg/ml) of rituximab (R) in culture. Flow cytometry (FCM) was performed to measure CD20 expression quantity on the surface of cells then to determine Jeko-1/R and Raji/R cells were rituximab-resistant cells. The Jeko-1/R and Raji/R cells were cultured in 50ug/mL rituximab to maintain the resistance phenotype.Results:In comparison to the parental Jeko-1 and Raji cells, the Jeko-1/R and Raji/R cells exhibited significantly lower CD20 expression. Specifically, the percentages of Jeko-1 and Raji cells expressing CD20 were 99.9% and 96.6%, respectively, whereas the percentages of Jeko-1/R cells and Raji/R cells expressing CD20 were 0.3% and 0.7%.Conclusion:The Jeko-1/R and Raji/R cells were rituximab-resistant lymphoma cell lines, making them ideal cellular modelswith which to explore the mechanism of rituximab resistance.2、PRELIMINARY EXPLORING THE MECHANISMS OF RITUXIMAB RESISTANCE BY cDNA MICROARRAY.Objective:In this study, we aimed to Preliminary Explore the Mechanisms of Rituximab Resistance by cDNA Microarray.Methods:After determining Jeko-1/R and Raji/R cells were rituximab-resistant cells by Flow cytometry (FCM),we Extracted the total RNA of parental cell lines and RRCLs, and then cDNA microarray analysis of parental cell lines and RRCLs was performed. KEGG database and DAVID software were used to subsequent bioinformatics analysis.Results:(1) We identified 201 differentially expressed genes in Jeko-1/R and Raji/R cells versus the parental cell lines,70 genes were upregulated and 42 genes were downregulated. And only 8 genes were more or less than 5 times (fold change≥5 or fold change≤0.2), they were DNAJB1、EPB49、HSPA2A. HSPA3A.HPSA6.PLXNA4、RAPH1 and BRWD1.(2)KEGG pathway analysis found that MAPK signaling pathway was significantly enriched in the Jeko-1/R and Raji/R cells. (3) GO term analysis of differentially expressed genes showed that the biological characteristics of rituximab-resistant cells including "anti-apoptosis" "promoting proliferation" and "blood vessel development".Conclusion:(1) Our results suggest that most closely involved with rituximab resistant genes are DNAJB1, EPB49, HSPA2A, HSPA3A, HPSA6, PLXNA4, RAPH1, BRWD1. (2) The MAPK signaling pathway is most closely associated with rituximab resistance, perhaps by inhibiting apoptosis and promoting proliferation and vascular development. These findings provide a theoretical basis for predicting and overcoming rituximab resistance in the clinical setting.