Microfluidic device to study cell transmigration under physiological shear stress conditions

The development of new drug therapies relies on studies of cell transmigration in in vitro systems. Migration has traditionally been studied using two methods, the Boyden chamber and a shear flow chamber assay. Though, commonly applied in cell transmigration studies, they are far from imitating a natural migration process. Here we describe a novel in vitro cell transmigration microfluidic assay, which mimicks physiological shear flow conditions in blood vessels. The device was designed to incorporate the principles of both the Boyden chamber and the shear flow chamber assay, i.e. migration through the membrane under flow conditions. The 3D environment of migrating cells is imitated by injecting cell adhesion proteins to coat the membrane in the device. We tested the developed device with Jurkat cells migration towards medium supplemented with serum, and with chemokine induced lymphocytes migration. The applied continuous flow of cell suspension and chemoattractant ensures that the concentration gradient is maintained in time and space. The cell adhesion proteins used to enhance cell migration in the device were fibronectin and VCAM-1. We successfully observed a multistep transmigration process by means of the developed microfluidic migration assay. The presented device is inexpensive, easy to fabricate and disposable, having a potential to be applied in basic research as well as in the drug development process.     

Breast cancer cells can switch between estrogen receptor α and ErbB signaling and combined treatment against both signaling pathways postpones development of resistance

The majority of breast cancers are estrogen responsive, but upon progression of disease other growth promoting pathways are activated, e.g., the ErbB receptor system. The present study focuses on resistance to the pure estrogen antagonist fulvestrant and strategies to treat resistant cells or even circumvent development of resistance. Limited effects were observed when targeting EGFR and ErbB2 with the monoclonal antibodies cetuximab, trastuzumab, and pertuzumab, whereas the pan-ErbB inhibitor CI-1033 selectively inhibited growth of fulvestrant resistant cell lines. CI-1033 inhibited Erk but not Akt signaling, which as well as Erk is important for antiestrogen resistant cell growth. Accordingly, combination therapy with CI-1033 and the Akt inhibitor SH-6 or the Protein Kinase C inhibitor RO-32-0432 was applied and found superior to single agent treatment. Further, the resistant cell lines were more sensitive to CI-1033 treatment when grown in the presence of fulvestrant, as withdrawal of fulvestrant restored signaling through the estrogen receptor α (ERα), partly overcoming the growth inhibitory effects of CI-1033. Thus, the resistant cells could switch between ERα and ErbB signaling for growth promotion. Although parental MCF-7 cell growth primarily depends on ERα signaling, a heregulin-1β induced switch to ErbB signaling rescued MCF-7 cells from the growth inhibition exerted by fulvestrant-mediated blockade of ERα signaling. This interplay between ERα and ErbB signaling could be abrogated by combined therapy targeting both receptor systems. Thus, the present study indicates that upon development of antiestrogen resistance, antiestrogen treatment should be continued in combination with signal transduction inhibitors. Further, upfront combination of endocrine therapy with pan-ErbB inhibition may postpone or even prevent development of treatment resistance.