Biotechnology-derived protein drugs, usually referred as biologics, represent a significant part of the whole pharmaceutical market. Typically, biologics are produced in genetically modified animal cells, which are regarded by many engineers and practitioners in the pharmaceutical industry as extremely sensitive to hydrodynamic forces. Since during research or normal operation in the biopharmaceutical industry cells are exposed to a range of hydrodynamic forces, this "shear sensitivity" idea often leads to very mild, sub-optimal designing and operating conditions.
To determine the actual levels of hydrodynamic stress capable of affecting the metabolism or viability of a cell line in bioprocessing or analytical devices, a microfluidic contracting-expanding device was developed in our group that exposes cells to controlled, well-defined hydrodynamic forces by means of keeping the flow in laminar conditions. Using this device, changes in cell behavior can be determined as a function of the local energy dissipation rate (EDR). EDR is a scalar value that is intrinsic to any moving fluid, is independent of the flow regime (turbulent/laminar) and accounts for both shear and extensional components of three-dimensional flow. It represents the rate at which work is done on a fluid element or a cell. If laminar flow is maintained, EDR can be reliably calculated using well-established equations for simple geometries or computational fluid dynamics (CFD) software for more complex problems.
The microfluidic device, consisting of a micro-channel bored in a stainless steel sheet in sandwich between two polycarbonate plates, was used in different setups to imitate the environment cells will experience in both bioprocessing and analytical equipment. As a model for analytical devices, it was selected a Fluorescent Activated Cell Sorter (FACS), where cells are forced through a nozzle and interrogated by a laser beam. This instrument was mimicked by passing the cells once through the microfluidic device; on the other hand, as a model for bioprocessing equipment, the hydrodynamic forces a cell experiences in a bioreactor were simulated by recirculating the cells through the microchannel, in an intent to reproduce the cyclic passage of the cells through the high EDR zone around the impeller to zones of relative low EDR intensity away from it. Several cell lines of industrial, research and medical interest were tested using the just described methodology. For single passage, in every case the elicited response was mainly an increase of cell necrosis with larger EDR while only a small fraction of cells became apoptotic when exposed to the highest levels of EDR tested. Changes in medium composition or genetic modifications did not affect this behavior.
The response to repeated, chronic exposure to moderate levels of EDR was case-specific. A research CHO cell line (CHO6E6) stopped growing at the lowest levels of chronic EDR evaluated (2.9x105 W·m-3) and started dying when the EDR intensity was increased. On the other hand, the growth curve of a GS-CHO industrial cell line that produces a fully human antibody was not affected at all even at the highest EDR tested (6.5x106 W·m-3), although the glycosylation pattern of the antibody suffered modifications. Single passage results for both cell lines showed a very similar behavior to previously tested cell lines.
Interestingly, results showed that at least some medical cell lines (THP1) might have an EDR threshold lower than industrial or research cell lines (i.e., more "shear" sensitive), which suggest a possible selection of the tougher individuals after continuous manipulation. This conclusion seems to be also supported by the chronic exposure of the industrial GS-CHO cell line at the highest chronic EDR tested. If this is the case, the microfluidic device could even become a tool for selection of stronger clones in the pharmaceutical industry.