Fischell Department of Bioengineering Research Works

Permanent URI for this collectionhttp://hdl.handle.net/1903/6627

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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.
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.
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, Dirk
The 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.
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.
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.
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-Chiao
Various 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.
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-Qun
Understanding 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.
Chitosan to Connect Biology to Electronics: Fabricating the Bio-Device Interface and Communicating Across This Interface
(MDPI, 2014-12-24) Kim, Eunkyoung; Xiong, Yuan; Cheng, Yi; Wu, Hsuan-Chen; Liu, Yi; Morrow, Brian H.; Ben-Yoav, Hadar; Ghodssi, Reza; Rubloff, Gary W.; Shen, Jana; Bentley, William E.; Shi, Xiaowen; Payne, Gregory F.
Individually, advances in microelectronics and biology transformed the way we live our lives. However, there remain few examples in which biology and electronics have been interfaced to create synergistic capabilities. We believe there are two major challenges to the integration of biological components into microelectronic systems: (i) assembly of the biological components at an electrode address, and (ii) communication between the assembled biological components and the underlying electrode. Chitosan possesses a unique combination of properties to meet these challenges and serve as an effective bio-device interface material. For assembly, chitosan’s pH-responsive film-forming properties allow it to “recognize” electrode-imposed signals and respond by self-assembling as a stable hydrogel film through a cathodic electrodeposition mechanism. A separate anodic electrodeposition mechanism was recently reported and this also allows chitosan hydrogel films to be assembled at an electrode address. Protein-based biofunctionality can be conferred to electrodeposited films through a variety of physical, chemical and biological methods. For communication, we are investigating redox-active catechol-modified chitosan films as an interface to bridge redox-based communication between biology and an electrode. Despite significant progress over the last decade, many questions still remain which warrants even deeper study of chitosan’s structure, properties, and functions.
Engineering Cell Surfaces with Polyelectrolyte Materials for Translational Applications
(MDPI, 2017-01-28) Zhang, Peipei; Bookstaver, Michelle L.; Jewell, Christopher M.
Engineering cell surfaces with natural or synthetic materials is a unique and powerful strategy for biomedical applications. Cells exhibit more sophisticated migration, control, and functional capabilities compared to nanoparticles, scaffolds, viruses, and other engineered materials or agents commonly used in the biomedical field. Over the past decade, modification of cell surfaces with natural or synthetic materials has been studied to exploit this complexity for both fundamental and translational goals. In this review we present the existing biomedical technologies for engineering cell surfaces with one important class of materials, polyelectrolytes. We begin by introducing the challenges facing the cell surface engineering field. We then discuss the features of polyelectrolytes and how these properties can be harnessed to solve challenges in cell therapy, tissue engineering, cell-based drug delivery, sensing and tracking, and immune modulation. Throughout the review, we highlight opportunities to drive the field forward by bridging new knowledge of polyelectrolytes with existing translational challenges.
Catechol-Based Hydrogel for Chemical Information Processing
(MDPI, 2017-07-03) Kim, Eunkyoung; Liu, Zhengchun; Liu, Yi; Bentley, William E.; Payne, Gregory F.
Catechols offer diverse properties and are used in biology to perform various functions that range from adhesion (e.g., mussel proteins) to neurotransmission (e.g., dopamine), and mimicking the capabilities of biological catechols have yielded important new materials (e.g., polydopamine). It is well known that catechols are also redox-active and we have observed that biomimetic catechol-modified chitosan films are redox-active and possess interesting molecular electronic properties. In particular, these films can accept, store and donate electrons, and thus offer redox-capacitor capabilities. We are enlisting these capabilities to bridge communication between biology and electronics. Specifically, we are investigating an interactive redox-probing approach to access redox-based chemical information and convert this information into an electrical modality that facilitates analysis by methods from signal processing. In this review, we describe the broad vision and then cite recent examples in which the catechol–chitosan redox-capacitor can assist in accessing and understanding chemical information. Further, this redox-capacitor can be coupled with synthetic biology to enhance the power of chemical information processing. Potentially, the progress with this biomimetic catechol–chitosan film may even help in understanding how biology uses the redox properties of catechols for redox signaling.
The Binding Effect of Proteins on Medications and Its Impact on Electrochemical Sensing: Antipsychotic Clozapine as a Case Study
(MDPI, 2017-08-01) Banis, George E.; Winkler, Thomas; Barton, Patricia; Chocron, Sheryl E.; Kim, Eunkyoung; Kelly, Deanna L.; Payne, Gregory F.; Ben-Yoav, Hadar; Ghodssi, Reza
Clozapine (CLZ), a dibenzodiazepine, is demonstrated as the optimal antipsychotic for patients suffering from treatment-resistant schizophrenia. Like many other drugs, understanding the concentration of CLZ in a patient’s blood is critical for managing the patients’ symptoms, side effects, and overall treatment efficacy. To that end, various electrochemical techniques have been adapted due to their capabilities in concentration-dependent sensing. An open question associated with electrochemical CLZ monitoring is whether drug–protein complexes (i.e., CLZ bound to native blood proteins, such as serum albumin (SA) or alpha-1 acid-glycoprotein (AAG)) contribute to electrochemical redox signals. Here, we investigate CLZ-sensing performance using fundamental electrochemical methods with respect to the impact of protein binding. Specifically, we test the activity of bound and free fractions of a mixture of CLZ and either bovine SA or human AAG. Results suggest that bound complexes do not significantly contribute to the electrochemical signal for mixtures of CLZ with AAG or SA. Moreover, the fraction of CLZ bound to protein is relatively constant at 31% (AAG) and 73% (SA) in isolation with varying concentrations of CLZ. Thus, electrochemical sensing can enable direct monitoring of only the unbound CLZ, previously only accessible via equilibrium dialysis. The methods utilized in this work offer potential as a blueprint in developing electrochemical sensors for application to other redox-active medications with high protein binding more generally. This demonstrates that electrochemical sensing can be a new tool in accessing information not easily available previously, useful toward optimizing treatment regimens.
Modification and Assembly of a Versatile Lactonase for Bacterial Quorum Quenching
(MDPI, 2018-02-06) Rhoads, Melissa K.; Hauk, Pricila; Gupta, Valerie; Bookstaver, Michelle L.; Stephens, Kristina; Payne, Gregory F.; Bentley, William E.
This work sets out to provide a self-assembled biopolymer capsule activated with a multi-functional enzyme for localized delivery. This enzyme, SsoPox, which is a lactonase and phosphotriesterase, provides a means of interrupting bacterial communication pathways that have been shown to mediate pathogenicity. Here we demonstrate the capability to express, purify and attach SsoPox to the natural biopolymer chitosan, preserving its activity to “neutralize” long-chain autoinducer-1 (AI-1) communication molecules. Attachment is shown via non-specific binding and by engineering tyrosine and glutamine affinity ‘tags’ at the C-terminus for covalent linkage. Subsequent degradation of AI-1, in this case N-(3-oxododecanoyl)-l-homoserine lactone (OdDHL), serves to “quench” bacterial quorum sensing (QS), silencing intraspecies communication. By attaching enzymes to pH-responsive chitosan that, in turn, can be assembled into various forms, we demonstrate device-based flexibility for enzyme delivery. Specifically, we have assembled quorum-quenching capsules consisting of an alginate inner core and an enzyme “decorated” chitosan shell that are shown to preclude bacterial QS crosstalk, minimizing QS mediated behaviors.
Gelatin-Enabled Microsensor for Pancreatic Trypsin Sensing
(MDPI, 2018-01-31) Banis, George; Beardslee, Luke A.; Ghodssi, Reza
Digestive health is critically dependent on the secretion of enzymes from the exocrine pancreas to the duodenum via the pancreatic duct. Specifically, pancreatic trypsin is a major protease responsible for breaking down proteins for absorption in the small intestine. Gelatin-based hydrogels, deposited in the form of thin films, have been studied as potential sensor substrates that hydrolyze in the presence of trypsin. In this work, we (1) investigate gelatin as a sensing material; (2) develop a fabrication strategy for coating sensor surfaces; and (3) implement a miniaturized impedance platform for measuring activity levels of pancreatic trypsin. Using impedance spectroscopy, we evaluate gelatin’s specificity and rate of degradation when exposed to a combination of pancreatic enzymes in neutral solution representative of the macromolecular heterogeneity present in the duodenal environment. Our findings suggest gelatin’s preferential degradation to trypsin compared to enzymes such as lipase and amylase. We further observe their interference with trypsin behavior in equivalent concentrations, reducing film digestion by as much as 83% and 77%, respectively. We achieve film patterns in thicknesses ranging from 300–700 nm, which we coat over interdigitated finger electrode sensors. Finally, we test our sensors over several concentrations to emulate the range of pancreatic secretions. Ultimately, our microsensor will serve as the foundation for developing in situ sensors toward diagnosing pancreatic pathologies.
Nuclear Deformation in Response to Mechanical Confinement is Cell Type Dependent
(MDPI, 2019-05-08) Doolin, Mary T.; Ornstein, Thea S.; Stroka, Kimberly M.
Mechanosensing of the mechanical microenvironment by cells regulates cell phenotype and function. The nucleus is critical in mechanosensing, as it transmits external forces from the cellular microenvironment to the nuclear envelope housing chromatin. This study aims to elucidate how mechanical confinement affects nuclear deformation within several cell types, and to determine the role of cytoskeletal elements in controlling nuclear deformation. Human cancer cells (MDA-MB-231), human mesenchymal stem cells (MSCs), and mouse fibroblasts (L929) were seeded within polydimethylsiloxane (PDMS) microfluidic devices containing microchannels of varying cross-sectional areas, and nuclear morphology and volume were quantified via image processing of fluorescent cell nuclei. We found that the nuclear major axis length remained fairly constant with increasing confinement in MSCs and MDA-MB-231 cells, but increased with increasing confinement in L929 cells. Nuclear volume of L929 cells and MSCs decreased in the most confining channels. However, L929 nuclei were much more isotropic in unconfined channels than MSC nuclei. When microtubule polymerization or myosin II contractility was inhibited, nuclear deformation was altered only in MSCs in wide channels. This work informs our understanding of nuclear mechanics in physiologically relevant spaces, and suggests diverging roles of the cytoskeleton in regulating nuclear deformation in different cell types.
Systematic Evaluation of Light-Activatable Biohybrids for Anti-Glioma Photodynamic Therapy
(MDPI, 2019-08-21) Inglut, Collin T.; Baglo, Yan; Liang, Barry J.; Cheema, Yahya; Stabile, Jillian; Woodworth, Graeme F.; Huang, Huang-Chiao
Photosensitizing biomolecules (PSBM) represent a new generation of light-absorbing compounds with improved optical and physicochemical properties for biomedical applications. Despite numerous advances in lipid-, polymer-, and protein-based PSBMs, their effective use requires a fundamental understanding of how macromolecular structure influences the physicochemical and biological properties of the photosensitizer. Here, we prepared and characterized three well-defined PSBMs based on a clinically used photosensitizer, benzoporphyrin derivative (BPD). The PSBMs include 16:0 lysophosphocholine-BPD (16:0 Lyso PC-BPD), distearoyl-phosphoethanolamine-polyethylene-glycol-BPD (DSPE-PEG-BPD), and anti-EGFR cetuximab-BPD (Cet-BPD). In two glioma cell lines, DSPE-PEG-BPD exhibited the highest singlet oxygen yield but was the least phototoxic due to low cellular uptake. The 16:0 Lyso PC-BPD was most efficient in promoting cellular uptake but redirected BPD’s subcellular localization from mitochondria to lysosomes. At 24 h after incubation, proteolyzed Cet-BPD was localized to mitochondria and effectively disrupted the mitochondrial membrane potential upon light activation. Our results revealed the variable trafficking and end effects of PSBMs, providing valuable insights into methods of PSBM evaluation, as well as strategies to select PSBMs based on subcellular targets and cytotoxic mechanisms. We demonstrated that biologically informed combinations of PSBMs to target lysosomes and mitochondria, concurrently, may lead to enhanced therapeutic effects against gliomas.
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-Chiao
Liposomes 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.
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, Imran
A 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.
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 Morss
Disrupted 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.
Photodynamic Priming Improves the Anti-Migratory Activity of Prostaglandin E Receptor 4 Antagonist in Cancer Cells In Vitro
(MDPI, 2021-10-20) Sorrin, Aaron J.; Liu, Cindy; Cicalo, Julia; Reader, Jocelyn; Najafali, Daniel; Zhang, Yuji; Roque, Dana M.; Huang, Huang-Chiao
The combination of photodynamic agents and biological inhibitors is rapidly gaining attention for its promise and approval in treating advanced cancer. The activity of photodynamic treatment is mainly governed by the formation of reactive oxygen species upon light activation of photosensitizers. Exposure to reactive oxygen species above a threshold dose can induce cellular damage and cancer cell death, while the surviving cancer cells are “photodynamically primed”, or sensitized, to respond better to other drugs and biological treatments. Here, we report a new combination regimen of photodynamic priming (PDP) and prostaglandin E2 receptor 4 (EP4) inhibition that reduces the migration and invasion of two human ovarian cancer cell lines (OVCAR-5 and CAOV3) in vitro. PDP is achieved by red light activation of the FDA-approved photosensitizer, benzoporphyrin derivative (BPD), or a chemical conjugate composed of the BPD linked to cetuximab, an anti-epithelial growth factor receptor (EGFR) antibody. Immunoblotting data identify co-inhibition of EGFR, cAMP-response element binding protein (CREB), and extracellular signal-regulated kinase 1/2 (ERK1/2) as key in the signaling cascades modulated by the combination of EGFR-targeted PDP and EP4 inhibition. This study provides valuable insights into the development of a molecular-targeted photochemical strategy to improve the anti-metastatic effects of EP4 receptor antagonists.
Targeting the Gut Mucosal Immune System Using Nanomaterials
(MDPI, 2021-10-21) McCright, Jacob; Ramirez, Ann; Amosu, Mayowa; Sinha, Arnav; Bogseth, Amanda; Maisel, Katharina
The gastrointestinal (GI) tract is one the biggest mucosal surface in the body and one of the primary targets for the delivery of therapeutics, including immunotherapies. GI diseases, including, e.g., inflammatory bowel disease and intestinal infections such as cholera, pose a significant public health burden and are on the rise. Many of these diseases involve inflammatory processes that can be targeted by immune modulatory therapeutics. However, nonspecific targeting of inflammation systemically can lead to significant side effects. This can be avoided by locally targeting therapeutics to the GI tract and its mucosal immune system. In this review, we discuss nanomaterial-based strategies targeting the GI mucosal immune system, including gut-associated lymphoid tissues, tissue resident immune cells, as well as GI lymph nodes, to modulate GI inflammation and disease outcomes, as well as take advantage of some of the primary mechanisms of GI immunity such as oral tolerance.

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