Physical contact with endothelial cells through β1- and β2- integrins rescues chronic lymphocytic leukemia cells from spontaneous and drug-induced apoptosis and induces a peculiar gene expression profile in leukemic cells.
BACKGROUND
Chronic lymphocytic leukemia B cells display prolonged survival in vivo, but when cultured in vitro rapidly undergo spontaneous apoptosis. We hypothesize that interactions with endothelial cells in infiltrated tissues and during recirculation may have a pathogenic role in chronic lymphocytic leukemia.


METHODS
We evaluated apoptosis of leukemic cells after co-culture on a monolayer of human umbilical vein endothelial cells with addition of fludarabine and antibodies that block adhesion. Then, we compared microarray-based gene expression profiles between leukemic cells at baseline and after co-culture.


RESULTS
We found that the endothelial layer protected leukemic cells from apoptosis inducing a 2-fold mean decrement in apoptotic cells after 2 days of co-culture. Moreover, the endothelial layer decreased the sensitivity of chronic lymphocytic leukemia B cells to fludarabine-induced apoptosis. Physical contact with endothelium mediated by both β(1)- and β(2)- integrins is essential for the survival advantage of leukemic cells. In particular, blocking CD106 on endothelial cells or CD18 on leukemic B cells led to the almost complete abrogation of the survival advantage (>70% inhibition of viability). However, a reduction of apoptosis was also measured in leukemic cells cultured in conditioned medium collected after 2 days of co-culture, implying that survival is partially mediated by soluble factors. Overall, the contact with endothelial cells modulated 1,944 genes in chronic lymphocytic leukemia B cells, establishing a peculiar gene expression profile: up-regulation of angiogenesis-related genes, an increase of genes involved in TGFβ and Wnt signaling pathways, secretion of cytokines recruiting stromal cells and macrophages and up-regulation of anti-apoptotic molecules such as Bcl2 and Survivin.


CONCLUSIONS
Our study supports the notion that endothelial cells are major players in the chronic lymphocytic leukemia microenvironment. Adhesion to endothelium strongly supports survival, protects from drug-induced apoptosis and extensively modifies the gene expression profile of leukemic cells.
Hematology Unit, Department of Oncology, Hematology and Respiratory Diseases, University of Modena and Reggio Emilia, Modena, Italy.
Rossana
R
Maffei
Stefania
S
Fiorcari
Jenny
J
Bulgarelli
Silvia
S
Martinelli
Ilaria
I
Castelli
Silvia
S
Deaglio
Giulia
G
Debbia
Marcella
M
Fontana
Valeria
V
Coluccio
Goretta
G
Bonacorsi
Patrizia
P
Zucchini
Franco
F
Narni
Giuseppe
G
Torelli
Mario
M
Luppi
Roberto
R
Marasca
D057134:Antibodies, Neutralizing / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001402:B-Lymphocytes / Q000378:metabolism* / Q000473:pathology; D018821:CD18 Antigens / Q000235:genetics / Q000378:metabolism*; D002448:Cell Adhesion / Q000187:drug effects / Q000235:genetics; D002450:Cell Communication / Q000187:drug effects / Q000235:genetics; D018920:Coculture Techniques; D017077:Culture Media, Conditioned / Q000494:pharmacology; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D061307:Human Umbilical Vein Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D019012:Integrin beta1 / Q000235:genetics / Q000378:metabolism*; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D061251:Primary Cell Culture; D015398:Signal Transduction / Q000235:genetics; D019010:Vascular Cell Adhesion Molecule-1 / Q000235:genetics / Q000378:metabolism; D014740:Vidarabine / Q000031:analogs & derivatives / Q000494:pharmacology
D057134:Antibodies, Neutralizing; D000970:Antineoplastic Agents; D018821:CD18 Antigens; D017077:Culture Media, Conditioned; D019012:Integrin beta1; D009363:Neoplasm Proteins; D019010:Vascular Cell Adhesion Molecule-1; D014740:Vidarabine; C024352:fludarabine