Fischell Department of Bioengineering Research Works
Permanent URI for this collectionhttp://hdl.handle.net/1903/6627
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Item Increased phagocytosis capacity of circulating neutrophils in patients on continuous flow ventricular assist device support(Wiley, 2023-12-22) Awad, Morcos A.; Sun, Wenji; Han, Dong; Griffith, Bartly P.; Wu, Zhongjun J.Background Neutrophils take part in the innate immune response, phagocytosis, and pro-inflammatory cytokine release. The phagocytic capacity of circulating neutrophils in patients on continuous flow (CF) ventricular assist device (VAD) has not been well studied. Methods Blood samples from 14 patients undergoing CF-VAD implantation were collected and analyzed preoperatively (at baseline) and on postoperative days (POD) 3, 7, 14, and 28. Flow cytometry was used to assess the surface expression levels of CD62L, CD162, and macrophage antigen-1 (MAC-1) and neutrophil phagocytic capacity. Interleukin 1 (IL1), IL6, IL8, TNF-α, neutrophil elastase, and myeloperoxidase in plasma were measured using enzyme-linked immunosorbent assays. Results Among the 14 patients, seven patients had preoperative bridge device support. Relative to baseline, patients with no bridge device had elevated leukocyte count and neutrophil elastase by POD3 which normalized by POD7. Neutrophil activation level, IL6, IL8, and TNF-α increased by POD3 and sustained elevated levels for 7–14 days postoperatively. Elevated neutrophil phagocytic capacity persisted even until POD28. Similar patterns were observed in patients on a preoperative bridge device. Conclusions Neutrophil activation and phagocytic capacity increased in response to VAD support, while inflammatory cytokines remain elevated for up to 2 weeks postoperatively. These findings may indicate that VAD implantation elicits circulating neutrophils to an abnormal preemptive phagocytotic phenotype.Item Investigation of the role of von Willebrand factor in shear-induced platelet activation and functional alteration under high non-physiological shear stress(Wiley, 2023-12-19) Han, Dong; Sun, Wenji; Clark, Kiersten P.; Griffith, Bartley P.; Wu, Zhongjun J.Background von Willebrand factor (vWF) plays a crucial role in physiological hemostasis through platelet and subendothelial collagen adhesion. However, its role in shear-induced platelet activation and functional alteration under non-physiological conditions common to blood-contacting medical devices (BCMDs) is not well investigated. Methods Fresh healthy human blood was treated with an anti-vWF antibody to block vWF–GPIbα interaction. Untreated blood was used as a control. They were exposed to three levels of non-physiological shear stress (NPSS) (75, 125, and 175 Pa) through a shearing device with an exposure time of 0.5 s to mimic typical shear conditions in BCMDs. Flow cytometric assays were used to measure the expression levels of PAC-1 and P-Selectin and platelet aggregates for platelet activation and the expression levels of GPIbα, GPIIb/IIIa, and GPVI for receptor shedding. Collagen/ristocetin-induced platelet aggregation capacity was characterized by aggregometry. Results The levels of platelet activation and aggregates increased with increasing NPSS in the untreated blood. More receptors were lost with increasing NPSS, resulting in a decreased capacity of collagen/ristocetin-induced platelet aggregation. In contrast, the increase in platelet activation and aggregates after exposure to NPSS, even at the highest level of NPSS, was significantly lower in treated blood. Nevertheless, there was no notable difference in receptor shedding, especially for GPIIb/IIIa and GPVI, between the two blood groups at the same level of NPSS. The block of vWF exacerbated the decreased capacity of collagen/ristocetin-induced platelet aggregation. Conclusions High NPSS activates platelets mainly by enhancing the vWF–GPIbα interaction. Platelet activation and receptor shedding induced by high NPSS likely occur through different pathways.Item Investigating the origin of the 13C lactate signal in the anesthetized healthy rat brain in vivo after hyperpolarized [1-13C]pyruvate injection(Wiley, 2023-11-22) Zhu, Minjie; Jhajharia, Aditya; Josan, Sonal; Park, Jae Mo; Yen, Yi-Fen; Pfefferbaum, Adolf; Hurd, Ralph E.; Spielman, Daniel M.; Mayer, DirkThe goal of this study was to investigate the origin of brain lactate (Lac) signal in the healthy anesthetized rat after injection of hyperpolarized (HP) [1-13C]pyruvate (Pyr). Dynamic two-dimensional spiral chemical shift imaging with flow-sensitizing gradients revealed reduction in both vascular and brain Pyr, while no significant dependence on the level of flow suppression was detected for Lac. These results support the hypothesis that the HP metabolites predominantly reside in different compartments in the brain (i.e., Pyr in the blood and Lac in the parenchyma). Data from high-resolution metabolic imaging of [1-13C]Pyr further demonstrated that Lac detected in the brain was not from contributions of vascular signal attributable to partial volume effects. Additionally, metabolite distributions and kinetics measured with dynamic imaging after injection of HP [1-13C]Lac were similar to Pyr data when Pyr was used as the substrate. These data do not support the hypothesis that Lac observed in the brain after Pyr injection was generated in other organs and then transported across the blood–brain barrier (BBB). Together, the presented results provide further evidence that even in healthy anesthetized rats, the transport of HP Pyr across the BBB is sufficiently fast to permit detection of its metabolic conversion to Lac within the brain.Item Computational fluid dynamics-based design and in vitro characterization of a novel pediatric pump-lung(Wiley, 2023-10-05) Han, Dong; Zhang, Jiafeng; He, Ge; Griffith, Bradley P.; Wu, Zhongjun J.Background Although extracorporeal membrane oxygenation (ECMO) has been used to provide temporary support for pediatric patients suffering severe respiratory or cardiac failure since 1970, ECMO systems specifically designed for pediatric patients, particularly for long-term use, remain an unmet clinical need. We sought to develop a new pediatric ECMO system, that is, pediatric pump-lung (PPL), consisting of a unique cylinder oxygenator with an outside-in radial flow path and a centrifugal pump. Methods Computational fluid dynamics was used to analyze the blood fluid field for optimized biocompatible and gas exchange performances in terms of flow characteristics, hemolysis, and gas transfer efficiency. Ovine blood was used for in vitro hemolysis and gas transfer testing. Results Both the computational and experimental data showed that the pressure drop through the PPL's oxygenator is significantly low, even at a flow rate of more than 3.5 L/min. The PPL showed better hemolysis performance than a commercial ECMO circuit consisting of the Quadrox-iD pediatric oxygenator and the Rotaflow pump at a 3.5 L/min flow rate and 250 mm Hg afterload pressure. The oxygen transfer rate of the PPL can reach over 200 mL/min at a flow rate of 3.5 L/min. Conclusions The PPL has the potential to provide adequate blood pumping and excellent respiratory support with minimal risk of hemolysis for a wide range of pediatric patients.Item High performance anion exchange chromatography purification of probiotic bacterial extracellular vesicles enhances purity and anti-inflammatory efficacy(Wiley, 2023-08-09) Pirolli, Nicholas H.; Reus, Laura Samantha C.; Mamczarz, Zuzanna; Lhan, Sulayman; Bentley, William E.; Jay, Steven M.Bacterial extracellular vesicles (BEVs), including outer membrane vesicles, have emerged as a promising new class of vaccines and therapeutics to treat cancer and inflammatory diseases, among other applications. However, clinical translation of BEVs is hindered by a current lack of scalable and efficient purification methods. Here, we address downstream BEV biomanufacturing limitations by developing a method for orthogonal size- and charge-based BEV enrichment using tangential flow filtration (TFF) in tandem with high performance anion exchange chromatography (HPAEC). The data show that size-based separation coisolated protein contaminants, whereas size-based TFF with charged-based HPAEC dramatically improved purity of BEVs produced by probiotic Gram-negative Escherichia coli and Gram-positive lactic acid bacteria (LAB). Escherichia coli BEV purity was quantified using established biochemical markers while improved LAB BEV purity was assessed via observed potentiation of anti-inflammatory bioactivity. Overall, this work establishes orthogonal TFF + HPAEC as a scalable and efficient method for BEV purification that holds promise for future large-scale biomanufacturing of therapeutic BEV products.Item Test method for evaluating the photocytotoxic potential of fluorescence imaging products(Wiley, 2023-07-26) Vig, Shruti; Gaitan, Brandon; Frankle, Lucas; Chen, Yu; Elespuru, Rosalie; Pfefer, T. Joshua; Huang, Huang-ChiaoVarious fluorescence imaging agents are currently under clinical studies. Despite significant benefits, phototoxicity is a barrier to the clinical translation of fluorophores. Current regulatory guidelines on medication-based phototoxicity focus on skin effects during sun exposure. However, with systemic and local administration of fluorophores and targeted illumination, there is now possibility of photochemical damage to deeper tissues during intraoperative imaging procedures. Hence, independent knowledge regarding phototoxicity is required to facilitate the development of fluorescence imaging products. Previously, we studied a cell-free assay for initial screening of reactive molecular species generation from fluorophores. The current work addresses a safety test method based on cell viability as an adjunct and a comparator with the cell-free assay. Our goal is to modify and implement an approach based on the in vitro 3T3 neutral red uptake assay of the Organization for Economic Co-Operation and Development Test Guideline 432 (OECD TG432) to evaluate the photocytotoxicity of clinically relevant fluorophores. These included indocyanine green (ICG), proflavine, methylene blue (MB), and IRDye800, as well as control photosensitizers, benzoporphyrin derivative (BPD) and rose bengal (RB). We performed measurements at agent concentrations and illumination parameters used for clinic imaging. Our results aligned with prior studies, indicating photocytotoxicity in RB and BPD and an absence of reactivity for ICG and IRDye800. DNA interactive agents, proflavine and MB, exhibited drug/light dose–response curves like photosensitizers. This study provides evidence and insights into practices useful for testing the photochemical safety of fluorescence imaging products.Item Abnormal coordination of upper extremity during target reaching in persons post stroke(Springer Nature, 2023-08-08) Koh, Kyung; Oppizzi, Giovanni; Kehs, Glenn; Zhang, Li-QunUnderstanding abnormal synergy of the upper extremity (UE) in stroke survivors is critical for better identification of motor impairment. Here, we investigated to what extent stroke survivors retain the ability to coordinate multiple joints of the arm during a reaching task. Using an exoskeleton robot, 37 stroke survivors’ arm joint angles (θ) and torques (τ) during hand reaching in the horizontal plane was compared to that of 13 healthy controls. Kinematic and kinetic coordination patterns were quantified as variances of the multiple-joint angles and multiple-joint torques across trials, respectively, that were partitioned into task-irrelevant variance (TIVθ and TIVτ) and task-relevant variance (TRVθ and TRVτ). TIVθ and TRVθ (or TIVτ and TRVτ) led to consistent and inconsistent hand position (or force), respectively. The index of synergy (ISθ and ISτ) was determined as ISθ = (TIVθ - TRVθ)/(TIVθ + TRVθ) and ISτ = (TIVτ - TRVτ)/(TIVτ + TRVτ) for kinematic and kinetic coordination patterns, respectively. Both kinematic ISθ and kinetic ISτ in the stroke group were significantly lower than that of the control group, indicating stroke survivors had impaired reaching abilities in utilizing the multiple joints of the UE for successful completion of a reaching task. The reduction of kinematic ISθ in the stroke group was mainly attributed to the lower TIVθ as compared to the control group, while the reduction of kinetic ISτ was mainly due to the higher TRVτ as well as lower TIVτ. Our results also indicated that stroke may lead to motor deficits in formation of abnormal kinetic synergistic movement of UE, especially during outward movement. The findings in abnormal synergy patterns provides a better understanding of motor impairment, suggesting that impairment-specific treatment could be identified to help improve UE synergies, focusing on outward movements.Item Immunological and Toxicological Considerations for the Design of Liposomes(MDPI, 2020-01-22) Inglut, Collin T.; Sorrin, Aaron J.; Kuruppu, Thilinie; Vig, Shruti; Cicalo, Julia; Ahmad, Haroon; Huang, Huang-ChiaoLiposomes hold great potential as gene and drug delivery vehicles due to their biocompatibility and modular properties, coupled with the major advantage of attenuating the risk of systemic toxicity from the encapsulated therapeutic agent. Decades of research have been dedicated to studying and optimizing liposomal formulations for a variety of medical applications, ranging from cancer therapeutics to analgesics. Some effort has also been made to elucidate the toxicities and immune responses that these drug formulations may elicit. Notably, intravenously injected liposomes can interact with plasma proteins, leading to opsonization, thereby altering the healthy cells they come into contact with during circulation and removal. Additionally, due to the pharmacokinetics of liposomes in circulation, drugs can end up sequestered in organs of the mononuclear phagocyte system, affecting liver and spleen function. Importantly, liposomal agents can also stimulate or suppress the immune system depending on their physiochemical properties, such as size, lipid composition, pegylation, and surface charge. Despite the surge in the clinical use of liposomal agents since 1995, there are still several drawbacks that limit their range of applications. This review presents a focused analysis of these limitations, with an emphasis on toxicity to healthy tissues and unfavorable immune responses, to shed light on key considerations that should be factored into the design and clinical use of liposomal formulations.Item Flow-induced Shear Stress Confers Resistance to Carboplatin in an Adherent Three-Dimensional Model for Ovarian Cancer: A Role for EGFR-Targeted Photoimmunotherapy Informed by Physical Stress(MDPI, 2020-03-28) Nath, Shubhankar; Pigula, Michael; Khan, Amjad P.; Hanna, William; Ruhi, Mustafa Kemal; Dehkordy, Farzaneh Mahmoodpoor; Pushpavanam, Karthik; Rege, Kaushal; Moore, Kaitlin; Tsujita, Yujiro; Conrad, Christina; Inci, Faith; del Carmen, Marcela G.; Franco, Walfre; Celli, Jonathan P.; Demirci, Utkan; Hasan, Tayyaba; Huang, Huang-Chiao; Rizvi, ImranA key reason for the persistently grim statistics associated with metastatic ovarian cancer is resistance to conventional agents, including platinum-based chemotherapies. A major source of treatment failure is the high degree of genetic and molecular heterogeneity, which results from significant underlying genomic instability, as well as stromal and physical cues in the microenvironment. Ovarian cancer commonly disseminates via transcoelomic routes to distant sites, which is associated with the frequent production of malignant ascites, as well as the poorest prognosis. In addition to providing a cell and protein-rich environment for cancer growth and progression, ascitic fluid also confers physical stress on tumors. An understudied area in ovarian cancer research is the impact of fluid shear stress on treatment failure. Here, we investigate the effect of fluid shear stress on response to platinum-based chemotherapy and the modulation of molecular pathways associated with aggressive disease in a perfusion model for adherent 3D ovarian cancer nodules. Resistance to carboplatin is observed under flow with a concomitant increase in the expression and activation of the epidermal growth factor receptor (EGFR) as well as downstream signaling members mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK) and extracellular signal-regulated kinase (ERK). The uptake of platinum by the 3D ovarian cancer nodules was significantly higher in flow cultures compared to static cultures. A downregulation of phospho-focal adhesion kinase (p-FAK), vinculin, and phospho-paxillin was observed following carboplatin treatment in both flow and static cultures. Interestingly, low-dose anti-EGFR photoimmunotherapy (PIT), a targeted photochemical modality, was found to be equally effective in ovarian tumors grown under flow and static conditions. These findings highlight the need to further develop PIT-based combinations that target the EGFR, and sensitize ovarian cancers to chemotherapy in the context of flow-induced shear stress.Item 13C Metabolic Flux Analysis Indicates Endothelial Cells Attenuate Metabolic Perturbations by Modulating TCA Activity(MDPI, 2021-04-07) Moiz, Bilal; Garcia, Jonathan; Basehore, Sarah; Sun, Angela; Li, Andrew; Padmanabhan, Surya; Albus, Kaitlyn; Jang, Cholsoon; Sriram, Ganesh; Clyne, Alisa MorssDisrupted endothelial metabolism is linked to endothelial dysfunction and cardiovascular disease. Targeted metabolic inhibitors are potential therapeutics; however, their systemic impact on endothelial metabolism remains unknown. In this study, we combined stable isotope labeling with 13C metabolic flux analysis (13C MFA) to determine how targeted inhibition of the polyol (fidarestat), pentose phosphate (DHEA), and hexosamine biosynthetic (azaserine) pathways alters endothelial metabolism. Glucose, glutamine, and a four-carbon input to the malate shuttle were important carbon sources in the baseline human umbilical vein endothelial cell (HUVEC) 13C MFA model. We observed two to three times higher glutamine uptake in fidarestat and azaserine-treated cells. Fidarestat and DHEA-treated HUVEC showed decreased 13C enrichment of glycolytic and TCA metabolites and amino acids. Azaserine-treated HUVEC primarily showed 13C enrichment differences in UDP-GlcNAc. 13C MFA estimated decreased pentose phosphate pathway flux and increased TCA activity with reversed malate shuttle direction in fidarestat and DHEA-treated HUVEC. In contrast, 13C MFA estimated increases in both pentose phosphate pathway and TCA activity in azaserine-treated cells. These data show the potential importance of endothelial malate shuttle activity and suggest that inhibiting glycolytic side branch pathways can change the metabolic network, highlighting the need to study systemic metabolic therapeutic effects.