Abstract
Circulation is an important delivery method for both natural and synthetic molecules, but microenvironment interactions, regulated by endothelial cells and critical to the molecule's fate, are difficult to interpret using traditional approaches. In this work, we analyzed and predicted growth factor capture under flow using computer modeling and a three-dimensional experimental approach that includes pertinent circulation characteristics such as pulsatile flow, competing binding interactions, and limited bioavailability. An understanding of the controlling features of this process was desired. The experimental module consisted of a bioreactor with synthetic endothelial-lined hollow fibers under flow. The physical design of the system was incorporated into the model parameters. The heparin-binding growth factor fibroblast growth factor-2 (FGF-2) was used for both the experiments and simulations. Our computational model was composed of three parts: (1) media flow equations, (2) mass transport equations and (3) cell surface reaction equations. The model is based on the flow and reactions within a single hollow fiber and was scaled linearly by the total number of fibers for comparison with experimental results. Our model predicted, and experiments confirmed, that removal of heparan sulfate (HS) from the system would result in a dramatic loss of binding by heparin-binding proteins, but not by proteins that do not bind heparin. The model further predicted a significant loss of bound protein at flow rates only slightly higher than average capillary flow rates, corroborated experimentally, suggesting that the probability of capture in a single pass at high flow rates is extremely low. Several other key parameters were investigated with the coupling between receptors and proteoglycans shown to have a critical impact on successful capture. The combined system offers opportunities to examine circulation capture in a straightforward quantitative manner that should prove advantageous for biologicals or drug delivery investigations.
Highlights
The bioavailability of molecules as they circulate through the bloodstream is a crucial factor in their signaling capability
We introduced a computational model based on a flow environment in which the competitive binding of fibroblast growth factor-2 (FGF-2), FGFR, and heparan sulfate proteoglycans (HSPG) in a pulsatile flow environment was addressed to mimic blood vessel-like hollow fibers [16,17]
Looking at cell binding at the entrance of the cell-lined hollow fiber as a function of time after FGF-2 has been introduced with constant FGFR (16104 #/cell) and variable HSPG, we found that there was a significant increase in bound FGF-2 at the higher HSPG (16105 #/cell) when compared to the lower values and that the FGFR binding was essentially all coupled to HSPG (Figure 11B)
Summary
The bioavailability of molecules as they circulate through the bloodstream is a crucial factor in their signaling capability. Halflife in circulation can determine the effectiveness of a drug by regulating the opportunities a molecule has to interact with the vessel wall. While some molecules may have a long circulation life, many may have only a single opportunity to interact with the blood vessel walls before being filtered through the liver or kidneys. Bevacizumab has been shown to increase the circulating concentration of VEGF in cancer patients when compared to patients not undergoing therapy because of the increased half-life of the growth factor-antibody complex; the complex is unable to bind to VEGF receptors [3] making delivery of the VEGF questionable. In order to better understand the vessel microenvironment and to accurately monitor drug interactions in the context of that microenvironment, better tools are needed to provide meaningful measurements that can predict the fate of molecules in circulation
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