LINC00853 Restrains T Cell Acute Lymphoblastic Leukemia Invasion And Infiltration By Regulating CCR9/CCL25

Background and purpose:T-lineage acute lymphocytic leukemia(T-ALL)is a malignant hematological tumor characterized by abnormal aggregation and infiltration of hematopoietic stem cells or early progenitor cells.In children and adults under the age of 35,the mortality rate of leukemia ranks the first among malignant tumors,and the incidence of T-ALL in children and adults accounts for 15% and 25% of acute lymphoblastic leukemia,posing a serious threat to human health.The invasion and infiltration of leukemia cells into specific organs leads to disease recurrence and poor prognosis.Therefore,researching the mechanism of T-ALL invasion and infiltration is crucial to find new therapeutic targets.Previous studies in our laboratory have shown that CC chemokine receptor 9(CCR9)is highly expressed in T-ALL cells and can bind to its unique receptor,CC chemokine ligand 25(CCL25),specifically,so as to participate in the T-ALL invasion and infiltration.However,the expression regulation mechanism of CCR9 remains unclear.At present,more and more studies have found that long non-coding RNA(lnc RNA)plays an important role in tumor migration,invasion and infiltration by regulating gene expression.In this study,lnc RNA negatively correlating with CCR9 expression was screened out from20 T-ALL datasets in GEO database by bioinformatics method,aims to explore the regulation of CCR9 expression by lnc RNA and the effect of this lnc RNA in T-ALL invasion and infiltration.Methods:Bioinformatics was used to find lnc RNAs negatively correlated with CCR9 expression in T-ALL datasets of GEO database,then ranked from large to small correlation,so LINC00853 was screened.The expression levels of LINC00853 and CCR9 in T-ALL were measured in the GEO dataset.Real-time PCR was further used to detect the expression levels of LINC00853 and CCR9 in T-ALL clinical samples and T-ALL cell lines.Lentivirus transfecting in jurkat cells were used to construct a LINC00853 stable knockdown cell line and adenovirus were used to construct a LINC00853 rescue cell line.Realtime PCR,Western Blot and flow cytometry were used to detected the expression of LINC00853 and CCR9 in the knockdown group,the rescue group and the control group.The migration and invasion ability of jurkat cells in LINC00853 knockdown group,rescue group and control group were decteced by Transwell and Matrigel-Transwell assays.In animal experiments,we built mice model by tail-vein injection of three groups of T-ALL cell line.We compared the body weight gain of three mice groups,calculated the ratio of organ weight to body weight in each group.Flow cytometry was used to detected the proportion of EGFP(a surface molecule sign of jurkat cell after transfection)positive cells in liver,spleen,brain,lung,kidney and ovary tissues.The proportion of Jurkat cells in tissue section were observed by fluorescence microscope.These in vitro and in vivo experiments confirmed that LINC00853 has effects on TALL cells(jurkat)invasion and infiltration.Results:Bioinformatics screened out ten lnc RNAs negatively correlated with CCR9 expression,ranked them from large to small correlation,excluded the ranked first and second lnc RNAs with too many homologous sequences,LINC00853 was selected for deep research.GEO data sets showed low expression of LINC00853 and high expression of CCR9 in T-ALL.Meanwhile,realtime PCR detected the down-regulation of LINC00853 and the up-regulation of CCR9 in T-ALL clinical samples and T-ALL cell lines,which were consistent with the database results.Real-time PCR,Western Blot and flow cytometry detected the expression level of CCR9 was up-regulated after knocking down LINC00853,and after rescuing LINC00853,the expression level of CCR9 was recovered.Transwell and Matrigel-Transwell assays detected that after LINC00853 was knocked down,the migration and invasion abilities of jurkat cells treated with CCL25 were significantly enhanced,while rescuing LINC00853 significantly decreased these abilities.In vivo experiments showed that the weight gain rate of knockdown group mice was the lowest,in rescue group,the level basically recovered to the control group.Liver,spleen,brain,lung,kidney and ovary in knockdown group all showed obvious swelling symptoms,and the degree of inflammation in rescue group was similar to control group.The proportion of EGFP(a surface molecule sign of jurkat cell after transfection)positive cells in liver,spleen,brain,lung,kidney and ovary tissues of knockdown mice group was the largest among three groups,and the proportion in rescue mice group was basically the same as control group.It was suggested that LINC00853 restrained the T-ALL infiltration into organs of mice by regulating the expression of CCR9.Conclusions:LINC00853 regulates CCR9 negatively.LINC00853 restrains the T-ALL invasion and infiltration by regulating the CCR9/CCL25.