OBJECTIVETo compare apoptosis gene expression profiling of U937 cells in suspension culture with that cultivated with mesenchymal stem cells (MSCs), and find out the relationship between drug resistance of leukemia cells and hemopoietic microenvironment.

METHODSU937 cells were cultivated in adhesion culture with MSCs and in suspension culture for 48 hours. Cell cycle was determined by flow cytometry and gene expression profiling by cDNA microarray.

RESULTSCompared with that in suspension, G(0)/G(1) fraction of U937 cells increased in adhesion culture (45.3 +/- 3.1)% vs (32.6 +/- 2.1)%, respectively (P < 0.05), whereas G(2)/M fraction and apoptosis rate were decreased. After 48 h twenty-eight differential expression genes were screened out in 487 apoptosis-related genes, among which 27 were up-regulated and were mainly apoptosis-suppressor genes, apoptosis-promoter genes, cell cycle positive control genes and cell cycle negative control genes. But Bcl-XL was up-regulated most obviously. The only one gene down-regulated was an apoptosis promoter gene.

CONCLUSIONAdhesion culture with MSCs can lead to growth suppression and decrease natural apoptosis of U937 cells. The mechanism was multiple gene effects, but Bcl-XL may be of the most importance.

Apoptosis ; genetics ; Cell Adhesion ; Cell Cycle ; genetics ; Cells, Cultured ; Coculture Techniques ; Flow Cytometry ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; Humans ; Lymphoma, Large B-Cell, Diffuse ; genetics ; pathology ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Oligonucleotide Array Sequence Analysis ; U937 Cells

BACKGROUNDThe hematopoietic microenvironment (HM) plays a critical role in malignant cell growth, patient survival, and response to chemotherapy in hematologic malignancies. However, mechanisms associated with this environmental influence remain unclear. In this study, we investigated the role of bone marrow derived mesenchymal stem cells (MSCs) in U937 cell line, to find out the relations between leukemia drug resistance and the MSCs.

METHODSU937 cells were cultured in suspension or grew adherently with MSCs. The cell growth curve was drawn and the cell cycle was measured by flow cytometry. Apoptosis and sensitivity of U937 to daunoblastina (DNR) were quantified by DNA ladder detection and trypan blue exclusion assays, respectively. The gene expression profile chip technology was used to determine and analyze the changes in apoptosis-related gene expression after adherent culture and the expression of MDR1 mRNA was assessed by reverse transcriptional polymerase chain reaction (RT-PCR) at the same time.

RESULTSIn the adherent culture, the proliferation of the U937 cells was inhibited, the G0/G1 phase cells increased (F = 64.9726, P < 0.0001), G2/M phase cells were decreased (F = 98.1361, P < 0.0001) and the natural apoptosis rate was decreased (F = 24.0866, P < 0.0001) compared with those in the suspended culture. U937 cell viability was enhanced and cell apoptosis was blocked during DNR treatment in adherent culture with MSCs. Thirty-nine differently expressed genes were screened from the 487 apoptosis related genes in the adherent culture U937 cells. Among the 37 upregulated genes, Bcl-XL was upregulated most significantly. Two genes were downregulated. Adherent culture did not induce MDR1 mRNA expression in U937 cells.

CONCLUSIONSMSCs play a role in modulating the proliferation of U937 cells and response of U937 cells to DNR, and Bcl-XL apoptosis-inhibiting gene may be most important in determining the sensitivity of leukemic cells to treatment, which is not related to MDR1.

Apoptosis ; drug effects ; Bone Marrow Cells ; physiology ; Cell Proliferation ; Daunorubicin ; pharmacology ; Drug Resistance, Neoplasm ; Genes, MDR ; Humans ; Immunophenotyping ; Mesenchymal Stromal Cells ; physiology ; U937 Cells ; drug effects