Colon cancer cell-derived 12(S)-HETE induces the retraction of cancer-associated fibroblast via MLC2, RHO/ROCK and Ca2+ signalling

• DOI: 10.1007/s00018-016-2441-5
• Publication Date: 2016-12-24T19:36:51Z
• Authors: Serena Stadler; Chi Huu Nguyen; Helga Schachner; Daniela Milovanovic; Silvio Holzner; Stefan Brenner; Julia Eichsteininger; Mira Stadler; Daniel Senfter; Liselotte Krenn; Wolfgang M. Schmidt; Nicole Huttary; Sigurd Krieger; Oskar Koperek; Zsuzsanna Bago-Ho...; Konstantin Alexan...; Brigitte Marian; Oliver de Wever; Robert M. Mader; Benedikt Giessrigl; Walter Jäger; Helmut Dolznig; Georg Krupitza
• Keywords: EXPRESSION; 0301 basic medicine; Myosin Light Chains; Colon; 301209 Pharmazie; INTRAVASATION; CARCINOMA-CELLS; KAPPA-B; Signal transduction; COLORECTAL-CANCER; ACTIVATION; Cellular and Molecular Neuroscience; 03 medical and health sciences; 301209 Pharmacy; Tumour progression; SDG 3 - Good Health and Well-being; Cancer-Associated Fibroblasts; Cell Movement; Cell Line, Tumor; Medicine and Health Sciences; CAMP PRODUCTION; Humans; Neoplasm Invasiveness; 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; 3D invasion model; Molecular Biology; Arachidonic acid metabolite; Pharmacology; rho-Associated Kinases; ECM; RECEPTOR; Rectum; Biology and Life Sciences; ARACHIDONIC-ACID; IN-VITRO; Cell Biology; ENDOTHELIAL-CELLS; 3. Good health; BARRIER FUNCTION; SDG 3 – Gesundheit und Wohlergehen; LIGHT-CHAIN KINASE; Molecular Medicine; Original Article; Calcium; Colorectal Neoplasms; Cardiac Myosins; PHOSPHOLIPASE-C-EPSILON; Signal Transduction

Retraction of mesenchymal stromal cells supports the invasion of colorectal cancer cells (CRC) into the adjacent compartment. CRC-secreted 12(S)-HETE enhances the retraction of cancer-associated fibroblasts (CAFs) and therefore, 12(S)-HETE may enforce invasivity of CRC. Understanding the mechanisms of metastatic CRC is crucial for successful intervention. Therefore, we studied pro-invasive contributions of stromal cells in physiologically relevant three-dimensional in vitro assays consisting of CRC spheroids, CAFs, extracellular matrix and endothelial cells, as well as in reductionist models. In order to elucidate how CAFs support CRC invasion, tumour spheroid-induced CAF retraction and free intracellular Ca2+ levels were measured and pharmacological- or siRNA-based inhibition of selected signalling cascades was performed. CRC spheroids caused the retraction of CAFs, generating entry gates in the adjacent surrogate stroma. The responsible trigger factor 12(S)-HETE provoked a signal, which was transduced by PLC, IP3, free intracellular Ca2+, Ca2+-calmodulin-kinase-II, RHO/ROCK and MYLK which led to the activation of myosin light chain 2, and subsequent CAF mobility. RHO activity was observed downstream as well as upstream of Ca2+ release. Thus, Ca2+ signalling served as central signal amplifier. Treatment with the FDA-approved drugs carbamazepine, cinnarizine, nifedipine and bepridil HCl, which reportedly interfere with cellular calcium availability, inhibited CAF-retraction. The elucidation of signalling pathways and identification of approved inhibitory drugs warrant development of intervention strategies targeting tumour-stroma interaction.