Anti-thrombotic strategies for microfluidic blood processing

Abstract

The redundant mechanisms involved in blood coagulation are crucial for rapid hemostasis. Yet they also create challenges in blood processing in medical devices and lab-on-a-chip systems. In this work, we investigate the effects of both shear stress and hypothermic blood storage on thrombus formation in microfluidic processing. For fresh blood, thrombosis occurs only at high shear, and the glycoprotein IIb/IIIa inhibitor tirofiban is highly effective in preventing thrombus formation. Blood storage generally activates platelets and primes them towards thrombosis via multiple mechanisms. Thrombus formation of stored blood at low shear can be adequately inhibited by glycoprotein IIb/IIIa inhibitors. At high shear, von Willebrand factor-mediated thrombosis contributes significantly and requires additional treatments with thiol-containing antioxidants-such as N acetylcysteine and reduced glutathione-that interfere with von Willebrand factor polymerization. We further demonstrate the effectiveness of these anti-thrombotic strategies in microfluidic devices made of cyclic olefin copolymer, a popular material used in the healthcare industry. This work identifies effective anti-thrombotic strategies that are applicable in a wide range of blood- and organ-on-a-chip applications.

A microfluidic bleeding model to investigate the effects of blood flow shear on microvascular hemostasis

“…Studies of plasma fibrinogen and platelet integrin α IIb β 3 [32,33] support the above viewpoints, suggesting that under relatively low shear conditions (< 1,000 s -1 ), integrin α IIb β 3 on the surface of free-flowing platelets can engage fibrinogen adsorbed onto the surface of the thrombus; in addition, the subsequent stimulation of platelets by locally generated soluble agonists induces a platelet shape change and an increase in integrin α IIb β 3 affinity, which helps stabilize and sustain integrin α IIb β3-fibrinogen bonds. Studies on plasma vWF [25,27,31] also showed that, at a low shear of < 30 dyn•cm -2 (converted into a shear rate of ~187 s -1 , when considering that the blood viscosity in this model is 1.6 × 10 -2 Pa•s, which is estimated from the empirical relationship for blood flow in tubes with diameters of < 1 mm [34]), vWF remains inactive. When the shear increases beyond 80 dyn•cm -2 (~500 s -1 ), vWF polymerization occurs and begins to induce platelet aggregation; the mechanisms mediated by integrin α IIb β 3 and vWF are not mutually exclusive, and both operate within the intermediate shear range; at an extremely high pathological shear level of > 300 dyn•cm -2 (~1875 s -1 ), the relative contribution of vWF to thrombosis increased significantly and eventually replaced integrin α IIb β 3 as the main mechanism.…”

“…Fluorescently label: platelets (green) and fibrin (ogen) (red), respectively. vWF activity by interfering with disulfide bonding, thereby preventing vWF-mediated thrombosis [31]. As shown in Figs.…”

“…3(a), it can be inferred that vWF dominates the platelet aggregation in a high-shear state, whereas in a low-shear state, it is most likely dominated by another mechanism, integrin α IIb β 3 -fibrinogen interactions. Large number of previously published studies on thrombosis have shown that the platelet aggregation mechanism is regulated by the blood flow shear [25][26][27]31]. Studies on the mechanism of platelet aggregation [26,30] indicate that, under conditions of relatively low shear (0-1,000 s -1 ), platelet aggregation is primarily mediated by soluble fibrinogen, which physically cross-links platelets through the engagement of integrin α IIb β 3 , which is affected by the local concentration of fibrinogen; in addition, at progressively higher wall shear rates (> 1,000 s -1 ), the aggregation becomes more vWF-dependent, and plasma vWF binds to the platelet receptor GP Ib , leading to platelet activation and subsequent shear-induced platelet aggregation.…”

Isolation of circulating tumor cells

“…Though CTC-iChip could process 10 mL of blood per hour, the very narrow microchannels of its 128 parallel DLD arrays limited scalability to larger blood volumes and incurred loss of CTCs as small as the lower half of WBCs. DLDs also suffer from shear-induced clogging ( Loutherback et al., 2012 ; Wong et al., 2018 ) made worse by prolonged contact of blood with tiny posts in narrow channels. Since the original DLD-based CTC-iChip , blood throughput has been raised by replacing the DLD debulking modules with an array-format inertial focusing device, as described in ( Mutlu et al., 2017 ).…”

Microfluidic concentration and separation of circulating tumor cell clusters from large blood volumes

“…More recently, deterministic lateral displacement (DLD) was used to isolate CTCCs 16 . DLDs require channels near in size to sorted cells to ensure bumping across streamlines and thus struggle to process large blood volumes without clogging 21 (platelettargeted additives can aid processing 22 ).…”