Influence of IGF-I and cell density on MDR1 expression in the T-lymphoblastoid cell line CCRF-CEM.
The debate about a direct or indirect effect of GH and IGF-I on the recurrence of malignancy, especially in the case of rhGH therapy in patients with leukemia, is still going on. Recent studies suggested that IGF-I plays a role in drug resistance during anticancer therapy. This resistance to diverse cytotoxic drugs, named multidrug-resistance (MDR), is mainly due to high levels of P-glycoprotein (P-gp). The gene encoding this membrane-associated transporter protein was named MDR1, and increased levels of P-gp are linked to enhanced MDR1 mRNA expression. Our aim was to investigate a possible effect of rhIGF-I on MDR1 gene expression in vitro. We cultured the T-lymphoblastoid cell line CCRF-CEM with different rhIGF-I concentrations (0, 5, 20 and 50 ng/ml) in serum-free medium for 3 days. CCRF-CEM cells are drug-sensitive and express MDR1 at low levels. MDR1 mRNA expression was measured by semiquantitative RT-PCR using a competitive assay with a heterologous DNA construct. In addition, GAPDH mRNA was amplified as an internal control for RNA integrity. P-gp activity was determined by a flow cytometric assay measuring rhodamine 123 accumulation. Furthermore, cell proliferation was monitored in all experiments. Our data do not support an effect of rhIGF-I on MDR1 mRNA expression, P-gp activity or cell proliferation in the CCRF-CEM cell line. MDR1 mRNA levels were inversely correlated to cell density with high significance (p < 0.0001). In conclusion, multidrug resistance linked to P-gp is not induced by IGF-I in CCRF-CEM cells. At high density, CCRF-CEM cells downregulate MDR1 gene expression. Our experimental model provides a very useful tool for monitoring the influence of growth factors on multidrug resistance in vitro.
Schwarze
C P
CP
University Children's Hospital Tübingen, Germany.
Neu
S
S
Beck
J
J
Mavridou
K
K
Ranke
M B
MB
Binder
G
G
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics*; D002452:Cell Count; D015870:Gene Expression / Q000187:drug effects*; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D015459:Leukemia-Lymphoma, Adult T-Cell / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D017526:Receptor, IGF Type 1 / Q000235:genetics; D011994:Recombinant Proteins / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D012333:RNA, Messenger; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D017526:Receptor, IGF Type 1