The Effect Of RhG-CSF On Spleen Transcriptome In Mouse Leukopenia Model Induced By Cyclophosphamide

Myelosuppression is a common adverse effect of chemotherapy. Leukopenia,which tends to be the primary symptom in bone marrow suppression, frequentlydemonstrates a drastic decrease in white blood cell counts and leads to the defect ofthe immune system, which can cause serious infection and threaten a patient`s life.Recombinant human granulocyte colony stimulating factor (rhG-CSF) is the mostefficient, safe and widely used therapy currently to reverse the suppression effects.Previous researches in rhG-CSF were mainly focused on its effect in hematopoieticsystem. However, according to several recent studies, rhG-CSF is able to raise theproportion of the marrow-derived suppression cells (MDSC) in the graft donors,which can lower the incidence of graft-versus-host disease (GVHD), and thusimplicate its potential roles in regulating the immune responses. Hence, it is of greatimportance to take a further step to explore the effects of rhG-CSF on immune systemin vivo.Transcriptomics is an approach to investigate the transcription status and relevantregulatory conditions within the cell as a whole, and thus reveals the potentialmolecular mechanisms during the process of a certain disease. In this study, we adopta Genechip assay to explore the expression conditions in the spleen of leukopeniamice with or without the rhG-CSF therapy, trying to reveal the underlyingmechanisms, through which the rhG-CSF can influence the immune system.To achieve our project, we carry out our research as follows. The mice used inthis study had a C57BL/6background. Mice were divided into three groups: blankcontrol group, cyclophosphamide therapy group and rhG-CSF therapy group. Firstly,mice in the therapy group were intraperitoneally injected with cyclophosphamide(200mg/kg) for a continuous three days to generate the leukopenia animal model.Afterwards, rhG-CSF (20ug/kg) was adopted in the rhG-CSF group for a successivesix days, during which period of time the status of each kind of blood cells in theperipheral blood were monitored. On the third day of rhG-CSF therapy, part of the mice were executed and the spleen was dissected for Genechip assay. All the data wasnormalized and calculated with the RNA algorithm. F-test of random varianceanalysis was adopted to evaluate the expression differences. The resources from GeneOntology and KEGG database were used to annotate the functions of related genes.By resorting to the gene regulatory network information from the KEGG database andthe pertinent resources from the Pathway database, we were able to delineate theunderlying gene network of interaction.In our research, we attain the following results. During the monitoring of theperipheral blood cells, we observed a pancytopenia effect from cyclophosphamide.However, the rhG-CSF can reverse this effect primarily and distinctly in neutrophilsand monocytes, yet with a limitation in its efficiency in lymphocytes. The Genechipassay revealed the difference in expression in3552genes among the three groups.After statistical analysis,74.9%of these genes are fasten to three dominant expressionpatterns due to the use of cyclophosphamide and rhG-CSF, which illustrates that therhG-CSF possibly reverses the cytotoxity of cyclophosphamide and exerts its immunesuppression functions via the regulation of amino acid and nucleic acid metabolism,DNA repair system as well as the gene expression regulatory system. In details, wespeculate that four pathways are involved in the process:1) rhG-CSF lowers theexpression of several adhesion molecules, interferes the connection between immunecells and endothelial cells and diminishes the migration of immune cells;2) rhG-CSFdecreases the expression of CXCR4, which is a lymphocyte chemotactic factor, andthus influences the chemotactic behavior of lymphocyte;3) rhG-CSF down-regulatesthe expression of DNM2and other phagocytosis related genes, restrains thephagocytosis behavior of the scavenger cells;4) rhG-CSF down-regulates theexpression of the core factors in the P38-MAPK pathway, which raises the proportionthe immune suppression cells.In conclusion, we investigated the influence of rhG-CSF on the immune systemby adopting the Genechip assay in the spleen of the cyclophosphamide inducedleukopenia animal model, and delineated the possible regulatory networks involved inthis process. All the work we have completed will facilitate us a better understanding of the rhG-CSF exerting its effects on the immune system.