Magnetic Field Changes Macrophage Phenotype.

Article Magnetic Field Changes Macrophage Phenotype Jarek Wosik,1,2,* Wei Chen,3,4 Kuang Qin,1,2 Rafik M. Ghobrial,3,5 Jacek Z. Kubiak,6,7 and Malgorzata Kloc3,5,8,* 1 Electrical and Computer Engineering Department and 2Texas Center for Superconductivity, University of Houston, Houston, Texas; 3The Houston Methodist Research Institute, Houston, Texas; 4Department of Nephrology, Second Xiangya Hospital, Central South University, Changsha, China; 5Department of Surgery, The Houston Methodist Hospital, Houston, Texas; 6Univ Rennes, CNRS, IGDR (Institute of Genetics and Development of Rennes), UMR 6290, Cell Cycle Group, Faculty of Medicine, Rennes, France; 7Department of Regenerative Medicine, Military Institute of Hygiene and Epidemiology (WIHE), Warsaw, Poland; and 8Department of Genetics, The University of Texas, M.D. Anderson Cancer Center, Houston, Texas ABSTRACT Macrophages play a crucial role in homeostasis, regeneration, and innate and adaptive immune responses. Functionally different macrophages have different shapes and molecular phenotypes that depend on the actin cytoskeleton, which is regulated by the small GTPase RhoA. The naive M0 macrophages are slightly elongated, proinflammatory M1 are round, and M2 antiinflammatory macrophages are elongated. We have recently shown in the rodent model system that genetic or pharmacologic interference with the RhoA pathway deregulates the macrophage actin cytoskeleton, causes extreme macrophage elongation, and prevents macrophage migration. Here, we report that an exposure of macrophages to a nonuniform magnetic field causes extreme elongation of macrophages and has a profound effect on their molecular components and organelles. Using immunostaining and Western blotting, we observed that magnetic force rearranges the macrophage actin cytoskeleton, the Golgi complex, and the cation channel receptor TRPM2, and modifies the expression of macrophage molecular markers. We have found that the magnetic-field-induced alterations are very similar to changes caused by RhoA interference. We also analyzed magnetic-field-induced forces acting on macrophages and found that the location and alignment of magnetic-fieldelongated macrophages correlate very well with the simulated distribution and orientation of such magnetic force lines. INTRODUCTION Macrophages are phenotypically and functionally diverse and play a crucial role in homeostasis, regeneration, innate and adaptive immune responses, and rejection of transplanted organs (1–3). Macrophages have several functionally different phenotypes/subtypes. M0 macrophages are naive/unpolarized macrophages. Two of the most common subtypes of activated macrophages are M1 proinflammatory ‘‘killer’’ macrophages, which produce damaging reactiveoxygen species and express the nitric oxide synthase iNOS, and M2 antiinflammatory ‘‘repair’’ macrophages, which produce the enzyme arginase-1 that depletes L-arginine and deprives iNOS of its substrate (1–7). Macrophages, like all eukaryotic cells, contain an actinfilament cytoskeleton. Macrophage migration occurs via dynamic rearrangements of actin filaments. Our recent studies showed that pharmacologic or genetic interference with the small GTPase RhoA pathway, which is the master regulator of actin, causes extreme elongation of macrophages (hummingbird phenotype), disrupts the Golgi/endosomal Submitted November 15, 2017, and accepted for publication March 6, 2018. *Correspondence: or Editor: Alexander Dunn. https://doi.org/10.1016/j.bpj.2018.03.002 pathway, prevents macrophage migration into the graft (through the clustering of the CX3CR1 receptor), and inhibits chronic rejection in the rodent model system (1–7). Here, we were interested in finding out whether an external magnetic field, in conjunction with transduction processes, could induce cytoskeletal rearrangements in macrophages and change their shape and molecular and organellar phenotype. It is already known that external mechanical force applied to the cell has a direct impact and can affect the cell cytoskeleton (8,9). It is also known that nonuniform magnetic fields can create such magnetic-force-driven stimuli (10). The cell responds to the external stimuli by remodeling the cytoskeleton, which is viscoelastic and provides a continuous mechanical coupling throughout the cell as it changes. This, in turn, induces an internal cell stress and changes in certain cellular components and components such as actin-filament polymerization, focal adhesions, etc. Such conversion of mechanical forces to biochemical interactions is referred to as mechanotransduction. There are reports that changes in ion-channel activity at the plasma membrane of cells may convey mechanical stresses from the cell membrane to internal organelles, causing changes in gene transcription and inducing apoptosis (11). Ó 2018 Biophysical Society. Biophysical Journal 114, 2001–2013, April 24, 2018 2001 Wosik et al. Other reports show that pathways of mechanically induced cell damage can include activation of the caspase-3 protease pathway (12) and tumor-necrosis-factor-related apoptosis-inducing ligand (13), and also cleavage of caspases 3 and 9 (14). In spite of outstanding recent progress in research of the influence of electromagnetic fields on the biology of cells and the expanding use of magnetic materials in biomedical applications, surprisingly little is known about the influence of a magnetic field at the cellular level (15,16). The nature and strength of interactions of electromagnetic fields with cell or tissue mainly depend on electric- and magneticfield-produced polarizations. The ability to induce such polarizations is measured by electric- and magnetic-field susceptibilities. There are significant differences between interactions of both fields with cells/tissue because for a typical tissue, the electric susceptibility is 105–106 times larger than the magnetic-field susceptibility and, as a result, the presence of the electric-field can cause significant cell/tissue damage, whereas magnetic-field interactions with cell/tissue are relatively weak (17,18). There are mixed reports about the influence of magnetic fields on cell growth and functions, and most—but not all—studies suggest that there is no obvious observable effect, even at as high as 10 T or higher values of uniform magnetic fields. In addition, although such fields in some studies have been shown to affect cell differentiation and viability, they did not have long-lasting, damaging effects (19). Furthermore, the nonuniform magnetic fields, in contrast to the uniform fields, were proven to generate sufficiently large magnetically induced mechanical forces able to affect cell morphology, differentiation, and functionality (20,21). As a result, a few in vitro studies carried out for spatially modulated magnetic fields showed a clear cellmagnetic field interaction (22–24). Although there are many technical challenges to generate sufficiently hi (...truncated)