Environmental Engineering Commons^™

High-Throughput Screening Of Clinically Approved Drugs That Prime Nonviral Gene Delivery To Human Mesenchymal Stem Cells, Tyler Kozisek, Andrew Hamann, Albert Nguyen, Michael Miller, Sarah Plautz, Angela K. Pannier

Biological Systems Engineering: Papers and Publications

Background: Human mesenchymal stem cells (hMSCs) are intensely researched for applications in cell therapeutics due to their unique properties, however, intrinsic therapeutic properties of hMSCs could be enhanced by genetic modification. Viral transduction is efficient, but suffers from safety issues. Conversely, nonviral gene delivery, while safer compared to viral, suffers from inefficiency and cytotoxicity, especially in hMSCs. To address the shortcomings of nonviral gene delivery to hMSCs, our lab has previously demonstrated that pharmacological ‘priming’ of hMSCs with the glucocorticoid dexamethasone can significantly increase transfection in hMSCs by modulating transfection-induced cytotoxicity. This work seeks to establish a library of transfection …

Nucleic Acid Delivery To Mesenchymal Stem Cells: A Review Of Nonviral Methods And Applications, Andrew Hamann, Albert Nguyen, Angela K. Pannier

Biological Systems Engineering: Papers and Publications

Background: Mesenchymal stem cells (MSCs) are multipotent stem cells that can be isolated and expanded from many tissues, and are being investigated for use in cell therapies. Though MSC therapies have demonstrated some success, none have been FDA approved for clinical use. MSCs lose stemness ex vivo, decreasing therapeutic potential, and face additional barriers in vivo, decreasing therapeutic efficacy. Culture optimization and genetic modification of MSCs can overcome these barriers. Viral transduction is efficient, but limited by safety concerns related to mutagenicity of integrating viral vectors and potential immunogenicity of viral antigens. Nonviral delivery methods are safer, though limited …

Free Polyethylenimine Enhances Substrate-Mediated Gene Delivery On Titanium Substrates Modified With Rgd-Functionalized Poly(Acrylic Acid) Brushes, Amy Mantz, Alice Rosenthal, Eric Farris, Tyler Kozisek, Eva Bittrich, Saghar Nazari, Eva Schubert, Mathias Schubert, Manfred Stamm, Petra Uhlmann, Angela K. Pannier

Biological Systems Engineering: Papers and Publications

Substrate mediated gene delivery (SMD) is a method of immobilizing DNA complexes to a substrate via covalent attachment or nonspecific adsorption, which allows for increased transgene expression with less DNA compared to traditional bolus delivery. It may also increase cells receptivity to transfection via cell-material interactions. Substrate modifications with poly(acrylic) acid (PAA) brushes may improve SMD by enhancing substrate interactions with DNA complexes via tailored surface chemistry and increasing cellular adhesion via moieties covalently bound to the brushes. Previously, we described a simple method to graft PAA brushes to Ti and further demonstrated conjugation of cell adhesion peptides (i.e., RGD) …

High-Throughput Screening Of Clinically Approved Drugs That Prime Polyethylenimine Transfection Reveals Modulation Of Mitochondria Dysfunction Response Improves Gene Transfer Efficiencies, Albert Nguyen, Jared P. Beyersdorf, Jean-Jack Riethoven, Angela K. Pannier

Biological Systems Engineering: Papers and Publications

Nonviral gene delivery methods are advantageous over viral vectors in terms of safety, cost, and flexibility in design and application, but suffer from lower gene transfer efficiency. In addition to modifications to nucleic acid design and nonviral carriers, new tools are sought to enhance transfection. Priming is the pharmacological modulation of transfection efficiency and transgene expression, and has demonstrated transfection increase in several compounds, for example, chloroquine and glucocorticoids. To develop a library of transfection priming compounds, a highthroughput screen was performed of the NIH Clinical Collection (NCC) to identify clinical compounds that prime polyethylenimine (PEI) transfection. HEK293T cells were …

Temporal Endogenous Gene Expression Profiles In Response To Polymer-Mediated Transfection And Profile Comparison To Lipid-Mediated Transfection, Timothy M. Martin, Sarah A. Plautz, Angela K. Pannier

Biological Systems Engineering: Papers and Publications

Background Design of efficient nonviral gene delivery systems is limited by the rudimentary understanding of specific molecules that facilitate transfection.

Methods Polyplexes using 25-kDa polyethylenimine (PEI) and plasmid encoding green fluorescent protein (GFP) were delivered to HEK 293T cells. After treating cells with polyplexes, microarrays were used to identify endogenous genes differentially expressed between treated and untreated cells (2 h of exposure) or between flow-separated transfected cells (GFP+) and treated, untransfected cells (GFP–) at 8, 16 and 24 h after lipoplex treatment. Cell priming studies were conducted using pharmacologic agents to alter endogenous levels of the identified differentially expressed genes …

Identifying Intracellular Pdna Losses From A Model Of Nonviral Gene Delivery, Timothy Michael Martin, Beata Joanna Wysocki, Tadeusz Antoni Wysocki, Angela K. Pannier

Biological Systems Engineering: Papers and Publications

Nonviral gene delivery systems are a type of nanocommunication system that transmit plasmid packets (i.e., pDNA packets) that are programmed at the nanoscale to biological systems at the microscopic cellular level. This engineered nanocommunication system suffers large pDNA losses during transmission of the genetically encoded information, preventing its use in biotechnological and medical applications. The pDNA losses largely remain uncharacterized, and the ramifications of reducing pDNA loss from newly designed gene delivery systems remain difficult to predict. Here, the pDNA losses during primary and secondary transmission chains were identified utilizing a MATLAB model employing queuing theory simulating delivery of pEGFPLuc …

Temporal Endogenous Gene Expression Profiles In Response To Lipid-Mediated Transfection, Timothy M. Martin, Sarah A. Plautz, Angela K. Pannier

Biological Systems Engineering: Papers and Publications

Background — Design of efficient nonviral gene delivery systems is limited as a result of the rudimentary understanding of the specific molecules and processes that facilitate DNA transfer.

Methods — Lipoplexes formed with Lipofectamine 2000 (LF2000) and plasmid-encoding green fluorescent protein (GFP) were delivered to the HEK 293T cell line. After treating cells with lipoplexes, HG-U133 Affymetrix microarrays were used to identify endogenous genes differentially expressed between treated and untreated cells (2 h exposure) or between flow-separated transfected cells (GFP+) and treated, untransfected cells (GFP–) at 8, 16 and 24 h after lipoplex treatment. Cell priming studies were conducted using …

Parallel Synthesis Of Poly(Amino Ether)-Templated Plasmonic Nanoparticles For Transgene Delivery, James Ramos, Thrimoorthy Potta, Olivia Scheideler, Kaushal Rege

Biological Systems Engineering: Papers and Publications

Plasmonic nanoparticles have been increasingly investigated for numerous applications in medicine, sensing, and catalysis. In particular, gold nanoparticles have been investigated for separations, sensing, drug/nucleic acid delivery, and bioimaging. In addition, silver nanoparticles demonstrate antibacterial activity, resulting in potential application in treatments against microbial infections, burns, diabetic skin ulcers, and medical devices. Here, we describe the facile, parallel synthesis of both gold and silver nanoparticles using a small set of poly(amino ethers), or PAEs, derived from linear polyamines, under ambient conditions and in absence of additional reagents. The kinetics of nanoparticle formation were dependent on PAE concentration and chemical composition. …

Dynamic Analysis Of Dna Nanoparticle Immobilization To Model Biomaterial Substrates Using Combinatorial Spectroscopic Ellipsometry And Quartz Crystal Microbalance With Dissipation, Tadas Kasputis, Alex Pieper, Mathias Schubert, Angela K. Pannier

Biological Systems Engineering: Papers and Publications

Gene expression within cells can be altered through gene delivery approaches, which have tremendous potential for gene therapy, tissue engineering, and diagnostics. Substrate-mediated gene delivery describes the delivery of plasmid DNA or DNA complexed with nonviral vectors to cells from a surface, with the DNA immobilized to a substrate through specific or nonspecific interactions. In this work, DNA-nanoparticle (DNA–NP) adsorption to substrates is evaluated using combinatorial, in situ spectroscopic ellipsometry and quartz crystal microbalance with dissipation (SE/QCM-D), to evaluate the basic dynamic processes involved in the adsorption and immobilization of DNA–NP complexes to substrates. The concentration of DNA–NP solutions influences …