INVESTIGADORES
BOLONTRADE Marcela Fabiana
congresos y reuniones científicas
Título:
Dissecting the crosstalk between mesenchymal stromal cells and hepatocellular carcinoma: clues to improve the use of cells as carriers for therapeutic genes
Autor/es:
BAYO FINA, JUAN; REAL, ALEJANDRINA; FIORE, ESTEBAN; MALVICINI, MARIANA; SGANGA, LEONARDO; BOLONTRADE, MARCELA F.; ANDRIANI, OSCAR; BIZAMA, CAROLINA; FRESNO, CRISTÓBAL; PODHAJCER, OSVALDO; FERNANDEZ, ELMER; GIDEKEL, MANUEL; GARCIA, MARIANA; MAZZOLINI, GUILLERMO
Lugar:
Vancouver
Reunión:
Simposio; International Liver Cancer Association (ILCA); 2016
Institución organizadora:
ILCA
Resumen:
Introduction: The incidence and mortality of hepatocellular carcinoma (HCC) are increasing steadily worldwide; therefore, new therapies are urgently needed1. The carcinogenesis process involves an interchange of secreted factors between HCC cells and their microenvironment, including hepatic stellate cells (HSC)2. It is known that mesenchymal stromal cells (MSCs) migrate in response to most of these factors and were used as carriers of antitumoral genes2. However, the mechanisms involved in the migration of MSC to HCC are not fully elucidated. Our aim was to study the crosstalk between the MSC and HCC microenvironment.Methods: Conditioned medium (CM) were derived from 5 human HCC (tumoral tissue, TT) and adjacent tissue (AT) samples, HCC xenografts (HuH7 and HC-PT-5), and monolayer cultures of HSC (LX-2) or HuH7 cells. The presence of soluble factors was analyzed by antibody arrays. Chemotaxis was studied using modified Boyden chambers. Specific antibodies or siRNA were used for Blocking or knockdown experiments. For MSC stimulation, 18h starved cells were incubated with HCC-CM for 24h. Gene expression profile was analyzed using cDNA microarrays. Cell proliferation was evaluated by [3H]-thymidine incorporation. In vivo HCC growth was analyzed in HuH7 subcutaneous tumor bearing mice after peritumoral administration of MSCs. ANOVA, Kruskal-Wallis test or t-test were used for the statistic analysis.Results: All the HCC CM shared the presence of GRO, MCP-1 and IL-8, being the latter with the highest concentration. Interestingly, AT-CM showed lower MCP-1 and IL-8 levels than TT-CM. Similarly, LX-2 CM showed the same cytokines although IL-6 was the most expressed one. Chemotaxis assays showed that MSC migration was higher to TT-CM than to AT-CM. Blocking and knockdown experiments of MCP-1, IL-8, IL8RA and IL8RB reduced >20 % MSC migration. Simultaneous blockage of AMF (autocrine motility factor), a cytokine involved in the migration of MSCs3, IL8RA and IL8RB resulted in >60% inhibition of MSC migration to HCC. Changes on chemotaxis potential, genes expression pattern and chemokines secretion of MSCs stimulated with HCC-CM (sMSC) were studied. Chemotaxis of sMSC (HuH7, HC-PT-5) showed a 2-fold increase of migration to HCC-CM. In addition, stimulation of MSC (PT-7, PT-12, HuH7 or HC-PT-5)modulate the expression of ~500 genes, being 46 genes related with cell migration and invasion. Furthermore, antibody arrays showed that sMSC (HuH7 or HC-PT-5) increase the secretion of several cytokines including IL-8, GCP-2 and GRO; and decrease of IP-10 and NAP-2. Then we assessed if these changes on sMSC could affect HCC behavior. HuH7 or HC-PT-5 exposed to CM derived from sMSC (HuH7 or HC-PT-5) did not show changes in their proliferation status. Chemotaxis assay showed that, sMSC recruit PBMNCs, fibroblasts (WI-38) and endothelial cells (HMEC-1) through IL-8, GRO and GCP-2. Finally, sMSC (HuH7) inoculated on HCC xenografts did not modify tumor growth.Conclusion: In this work we identified factors responsible for MSC migration to HCC that work in a cooperative way. In addition, HCC soluble factors increase MSC chemotactic potential, and modulate their gene and protein secretion profile. Moreover, HCC-stimulated MSCs could enhance the recruitment of other cells to the tumor microenvironment. MSC therapy did not affect HCC growth after in vivo transplantation. MSC as carriers of therapeutic genes for HCC is a challenging strategy and warrants further investigations.References: 1. El-Serag, H.B. N Engl J Med, 2011. 365(12); 2. Bayo, J., et al. Liver Int, 2014. 34(3); 3. Bayo, J., et al.PLoS One, 2014. 9(4)