Publications by authors named "Prajnaparamita Dhar"

27 Publications

  • Page 1 of 1

Hyaluronic Acid Hydrogel Microspheres for Slow Release Stem Cell Delivery.

ACS Biomater Sci Eng 2021 08 29;7(8):3754-3763. Epub 2021 Jul 29.

Likarda LLC, 10330 Hickman Mills Drive, Suite B, Kansas City, Missouri 64137, United States.

Cell therapies are hampered by a lack of available delivery systems, resulting in inconsistent outcomes in animal studies and human clinical trials. Hydrogel encapsulants offer a broad range of tunable characteristics in the design of cell delivery vehicles. The focus of the hydrogel field has been on durable encapsulants that provide long-term paracrine function of the cells. However, some cell therapies require cell-to-cell contact in order to elicit their effect. Controlled release microencapsulants would be beneficial in these situations, but appropriate polymers have not been adaptable to microsphere manufacturing because they harden too slowly. We developed and tested a novel microencapsulant formulation (acrylated hyaluronic acid: AHA) with degradation characteristics as a controlled release cell delivery vehicle. The properties of AHA microspheres were evaluated and compared to those of poly(ethylene glycol) diacrylate (PEGDA), a durable hydrogel. AHA microspheres possessed a higher swelling ratio, lower diffusion barrier, faster degradation rate, a lower storage modulus, and a larger average diameter than microspheres composed of PEGDA. Additionally, cell viability and release and short-term biocompatibility in immune competent Sprague-Dawley rats was assessed for each microsphere type. Compared to PEGDA, microspheres composed of AHA resulted in significantly less foreign body response as measured by a lack of cellularity or fibrotic ring in the surrounding tissue and no cellular infiltration into the microsphere. This study illustrates the potential of AHA microspheres as a degradable cell delivery system with superior encapsulated cell viability and biocompatibility with the surrounding tissue.
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http://dx.doi.org/10.1021/acsbiomaterials.1c00658DOI Listing
August 2021

Lung Surfactant Decreases Biochemical Alterations and Oxidative Stress Induced by a Sub-Toxic Concentration of Carbon Nanoparticles in Alveolar Epithelial and Microglial Cells.

Int J Mol Sci 2021 Mar 7;22(5). Epub 2021 Mar 7.

Department of Drug and Health Sciences, University of Catania, 95125 Catania, Italy.

Carbon-based nanomaterials are nowadays attracting lots of attention, in particular in the biomedical field, where they find a wide spectrum of applications, including, just to name a few, the drug delivery to specific tumor cells and the improvement of non-invasive imaging methods. Nanoparticles inhaled during breathing accumulate in the lung alveoli, where they interact and are covered with lung surfactants. We recently demonstrated that an apparently non-toxic concentration of engineered carbon nanodiamonds (ECNs) is able to induce oxidative/nitrosative stress, imbalance of energy metabolism, and mitochondrial dysfunction in microglial and alveolar basal epithelial cells. Therefore, the complete understanding of their "real" biosafety, along with their possible combination with other molecules mimicking the in vivo milieu, possibly allowing the modulation of their side effects becomes of utmost importance. Based on the above, the focus of the present work was to investigate whether the cellular alterations induced by an apparently non-toxic concentration of ECNs could be counteracted by their incorporation into a synthetic lung surfactant (DPPC:POPG in 7:3 molar ratio). By using two different cell lines (alveolar (A549) and microglial (BV-2)), we were able to show that the presence of lung surfactant decreased the production of ECNs-induced nitric oxide, total reactive oxygen species, and malondialdehyde, as well as counteracted reduced glutathione depletion (A549 cells only), ameliorated cell energy status (ATP and total pool of nicotinic coenzymes), and improved mitochondrial phosphorylating capacity. Overall, our results on alveolar basal epithelial and microglial cell lines clearly depict the benefits coming from the incorporation of carbon nanoparticles into a lung surfactant (mimicking its in vivo lipid composition), creating the basis for the investigation of this combination in vivo.
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http://dx.doi.org/10.3390/ijms22052694DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7962095PMC
March 2021

Impact of Polysorbate 80 Grade on the Interfacial Properties and Interfacial Stress Induced Subvisible Particle Formation in Monoclonal Antibodies.

J Pharm Sci 2021 02 26;110(2):746-759. Epub 2020 Sep 26.

Bioengineering Program, School of Engineering, The University of Kansas, 1530 W 15th Street, Lawrence, KS 66045, USA; Department of Chemical and Petroleum Engineering, The University of Kansas, 1530 W 15th Street, Lawrence, KS 66045, USA. Electronic address:

Polysorbate 80 is a nonionic surfactant that is added to therapeutic protein formulations to mitigate protein particle formation when subjected to various mechanical stresses. Variations in the PS80 grade has recently sparked questions surrounding the effect of oleic acid content (OAC) on surfactant's ability to mitigate interface-induced protein particle formation when stressed. In this work, a Langmuir trough was used to apply interfacial dilatational stress to two IgG molecules (mAb1 and mAb2) in formulations containing Chinese pharmacopeia (CP) and multicompendial (MC) grades of PS80. The interfacial properties of these mAb formulations, with and without interfacial dilatational stresses, were correlated with subvisible particle count and particle size/morphology distributions as measured by Micro-flow imaging (MFI). Overall, differences in interfacial properties correlated well with protein particle formation for both molecules in the two PS80 formulations. Further, the impact of grade of PS80 on the interfacial properties and interfacial stress-induced protein particle formation depends on the adsorption kinetics of the IgG molecules as well as the concentration of the surfactant used. This study demonstrates that measuring the interfacial properties of mAb formulations can be a useful tool to predict interfacial stress induced protein particle formation in the presence of different excipients of varying quality.
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http://dx.doi.org/10.1016/j.xphs.2020.09.035DOI Listing
February 2021

Viscoelastic Properties of ECM-Rich Embryonic Microenvironments.

Front Cell Dev Biol 2020 31;8:674. Epub 2020 Aug 31.

Department of Anatomy & Cell Biology, University of Kansas Medical Center, Kansas City, KS, United States.

The material properties of tissues and their mechanical state is an important factor in development, disease, regenerative medicine and tissue engineering. Here we describe a microrheological measurement technique utilizing aggregates of microinjected ferromagnetic nickel particles to probe the viscoelastic properties of embryonic tissues. Quail embryos were cultured in a plastic incubator chamber located at the center of two pairs of crossed electromagnets. We found a pronounced viscoelastic behavior within the ECM-rich region separating the mesoderm and endoderm in Hamburger Hamilton stage 10 quail embryos, consistent with a Zener (standard generalized solid) model. The viscoelastic response is about 45% of the total response, with a characteristic relaxation time of 1.3 s.
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http://dx.doi.org/10.3389/fcell.2020.00674DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7487363PMC
August 2020

Development of lipid membrane based assays to accurately predict the transfection efficiency of cell-penetrating peptide-based gene nanoparticles.

Int J Pharm 2020 Apr 9;580:119221. Epub 2020 Mar 9.

Department of Chemical & Petroleum Engineering, University of Kansas, Lawrence, KS 66047, USA. Electronic address:

Successful gene therapy requires the development of vectors that enable efficient delivery of genetic materials (e.g., pDNA or siRNA) to targeted cells, without degradation of the genetic materials. We have shown that nanoparticles formed by combining cell-penetrating peptide and pDNA (CPP-pDNA) into complexes and condensing them with calcium chloride can provide gene nanoparticles with high transfection efficiency and low cytotoxicity. In this work, we compare in situ measurements of the membrane insertion potential of three arginine-based gene nanoparticles (RW9-NPs, R9-NPs, and RH9-NPs) using four lipid compositions and two types of model membrane (Langmuir monolayers vs. supported bilayers) with their transfection efficiency in two human cancer cell lines. Using a Langmuir trough, we measured the membrane insertion potential of our gene nanoparticles to model membrane monolayers. A Quartz Crystal Microbalance with Dissipation (QCM-D) technique was used to monitor the adsorption of these nanoparticles to lipid bilayers of various compositions. Finally, gene expression using these nanoparticles was measured in breast cancer and cervical cancer cell lines. Our cell culture studies indicate that although R9-NPs and RW9-NPs show a significant increase in transfection efficiency compared to free pDNA, RH9-NPs do not show any significant difference. Both the Langmuir monolayer and QCM-D bilayer studies show that these results are best reflected in the in situ measurement assays when lipid systems containing a mixture of phospholipids, cholesterol, and sphingolipids are used. It is important to note that the mechanism of penetration is expected to differ for RW9 vs. R9; however, gene nanoparticles containing either of these CPPs show similar transfection efficiency. Our results therefore demonstrate that the design of predictive assays for gene therapy using CPPs must involve carefully chosen model lipid membrane systems that accurately represent the varying compositions of cell membranes.
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http://dx.doi.org/10.1016/j.ijpharm.2020.119221DOI Listing
April 2020

Impact of Engineered Carbon Nanodiamonds on the Collapse Mechanism of Model Lung Surfactant Monolayers at the Air-Water Interface.

Molecules 2020 Feb 7;25(3). Epub 2020 Feb 7.

Department of Chemical and Petroleum Engineering, The University of Kansas, Lawrence, KS 66045, USA.

Understanding interactions between inhaled nanoparticles and lung surfactants (LS) present at the air-water interface in the lung, is critical to assessing the toxicity of these nanoparticles. Specifically, in this work, we assess the impact of engineered carbon nanoparticles (ECN) on the ability of healthy LS to undergo reversible collapse, which is essential for proper functioning of LS. Using a Langmuir trough, multiple compression-expansion cycles are performed to assess changes in the surface pressure vs. area isotherms with time and continuous cyclic compression-expansion. Further, theoretical analysis of the isotherms is used to calculate the ability of these lipid systems to retain material during monolayer collapse, due to interactions with ECNs. These results are complemented with fluorescence images of alterations in collapse mechanisms in these monolayer films. Four different model phospholipid systems, that mimic the major compositions of LS, are used in this study. Together, our results show that the ECN does not impact the mechanism of collapse. However, the ability to retain material at the interface during monolayer collapse, as well as re-incorporation of material after a compression-expansion cycle is altered to varying extent by ECNs and depends on the composition of the lipid mixtures.
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http://dx.doi.org/10.3390/molecules25030714DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7037128PMC
February 2020

Novel machine learning application for prediction of membrane insertion potential of cell-penetrating peptides.

Int J Pharm 2019 Aug 21;567:118453. Epub 2019 Jun 21.

Department of Pharmaceutics, Faculty of Pharmacy, King Abdulaziz University, Jeddah, Saudi Arabia. Electronic address:

Cell-penetrating peptides (CPPs) are often used as transporter systems to deliver various therapeutic agents into the cell. We developed a novel machine learning application which can quantitatively screen the insertion/interaction potential of various CPPs into three model phospholipid monolayers. An artificial neural network (ANN) was designed, trained, and ultimately tested on an external dataset using Langmuir experimental data for 13 CPPs (hydrophilic and amphiphilic) together with various features related to the insertion/interaction efficiency of CPPs. The trained ANN provided accurate predictions of the maximum change in surface pressure of CPPs when injected below three membrane models at pH 7.4. The accuracy of predictions was high for the dataset which was used to construct the model (r = 0.986) as well as for the external "prospective" dataset (r = 0.969). In conclusion, this study demonstrates the promising potential of ANNs for screening the insertion potential of CPPs into membrane models for efficient intracellular delivery of therapeutic agents.
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http://dx.doi.org/10.1016/j.ijpharm.2019.118453DOI Listing
August 2019

Self-Assembled Coacervates of Chitosan and an Insect Cuticle Protein Containing a Rebers-Riddiford Motif.

Biomacromolecules 2018 07 9;19(7):2391-2400. Epub 2018 May 9.

Department of Biochemistry and Molecular Biophysics , Kansas State University , 141 Chalmers Hall , Manhattan , Kansas 66506 , United States.

The interactions among biomacromolecules within insect cuticle may offer new motifs for biomimetic material design. CPR27 is an abundant protein in the rigid cuticle of the elytron from Tribolium castaneum. CPR27 contains the Rebers-Riddiford (RR) motif, which is hypothesized to bind chitin. In this study, active magnetic microrheology coupled with microscopy and protein particle analysis techniques were used to correlate alterations in the viscosity of chitosan solutions with changes in solution microstructure. Addition of CPR27 to chitosan solutions led to a 3-fold drop in viscosity. This change was accompanied by the presence of micrometer-sized coacervate particles in solution. Coacervate formation had a strong dependence on chitosan concentration. Analysis showed the existence of a critical CPR27 concentration beyond which a significant increase in particle count was observed. These effects were not observed when a non-RR cuticular protein, CP30, was tested, providing evidence of a structure-function relationship related to the RR motif.
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http://dx.doi.org/10.1021/acs.biomac.7b01637DOI Listing
July 2018

Non-toxic engineered carbon nanodiamond concentrations induce oxidative/nitrosative stress, imbalance of energy metabolism, and mitochondrial dysfunction in microglial and alveolar basal epithelial cells.

Cell Death Dis 2018 02 14;9(2):245. Epub 2018 Feb 14.

Oasi Research Institute - IRCCS, 94018, Troina, Italy.

Engineered nanoparticles are finding a wide spectrum of biomedical applications, including drug delivery and capacity to trigger cytotoxic phenomena, potentially useful against tumor cells. The full understanding of their biosafety and interactions with cell processes is mandatory. Using microglial (BV-2) and alveolar basal epithelial (A549) cells, in this study we determined the effects of engineered carbon nanodiamonds (ECNs) on cell viability, nitric oxide (NO) and reactive oxygen species (ROS) production, as well as on energy metabolism. Particularly, we initially measured decrease in cell viability as a function of increasing ECNs doses, finding similar cytotoxic ECN effects in the two cell lines. Subsequently, using apparently non-cytotoxic ECN concentrations (2 µg/mL causing decrease in cell number < 5%) we determined NO and ROS production, and measured the concentrations of compounds related to energy metabolism, mitochondrial functions, oxido-reductive reactions, and antioxidant defences. We found that in both cell lines non-cytotoxic ECN concentrations increased NO and ROS production with sustained oxidative/nitrosative stress, and caused energy metabolism imbalance (decrease in high energy phosphates and nicotinic coenzymes) and mitochondrial malfunctioning (decrease in ATP/ADP ratio).These results underline the importance to deeply investigate the molecular and biochemical changes occurring upon the interaction of ECNs (and nanoparticles in general) with living cells, even at apparently non-toxic concentration. Since the use of ECNs in biomedical field is attracting increasing attention the complete evaluation of their biosafety, toxicity and/or possible side effects both in vitro and in vivo is mandatory before these highly promising tools might find the correct application.
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http://dx.doi.org/10.1038/s41419-018-0280-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5833425PMC
February 2018

pH-Induced Changes in the Surface Viscosity of Unsaturated Phospholipids Monitored Using Active Interfacial Microrheology.

Langmuir 2018 01 9;34(3):1159-1170. Epub 2017 Nov 9.

Department of Pharmaceutics and Industrial Pharmacy, Faculty of Pharmacy, King Abdulaziz University , Jeddah, KSA.

Lipid membranes, a major component of cells, are subjected to significant changes in pH depending on their location in the cell: the outer leaflet of the cell membrane is exposed to a pH of 7.4 whereas lipid membranes that make up late endosomes and lysosomes are exposed to a pH of as low as 4.4. The purpose of this study is to evaluate how changes in the environmental pH within cells alter the fluidity of phospholipid membranes. Specifically, we studied pH-induced alterations in the surface arrangement of monounsaturated lipids with zwitterionic headgroups (phosphoethanolamine (PE) and phosphocholine (PC)) that are abundant in plasma membranes as well as anionic lipids (phosphatidylserine (PS) and phosphatidylglycerol (PG)) that are abundant in inner membranes using a combination of techniques including surface tension vs area measurements, interfacial microrheology, and fluorescence/atomic force microscopy. Using an active interfacial microrheology technique, we find that phospholipids with zwitterionic headgroups show a significant increase in their surface viscosity at acidic pH. This increase in surface viscosity is also found to depend on the size of the lipid headgroup, with a smaller headgroup showing a greater increase in viscosity. The observed pH-induced increase in viscosity is also accompanied by an increase in the cohesion pressure between zwitterionic molecules at acidic pH and a decrease in the average molecular area of the lipids, as measured by fitting the surface pressure isotherms to well-established equations of state. Because fluorescent images show no change in the phase of the lipids, we attribute this change in surface viscosity to the pH-induced reorientation of the P-N dipoles that form part of the polar lipid headgroup, resulting in increased lipid-lipid interactions. Anionic PG headgroups do not demonstrate this pH-induced change in viscosity, suggesting that the presence of a net negative charge on the headgroup causes electrostatic repulsion between the headgroups. Our results also show that active interfacial microrheology is a sensitive technique for detecting minute changes in the lipid headgroup orientation induced by changes in the local membrane environment, even in unsaturated phospholipids where the surface viscosity is close to the experimental detection limit.
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http://dx.doi.org/10.1021/acs.langmuir.7b02803DOI Listing
January 2018

Evaluating the Role of the Air-Solution Interface on the Mechanism of Subvisible Particle Formation Caused by Mechanical Agitation for an IgG1 mAb.

J Pharm Sci 2016 05 26;105(5):1643-1656. Epub 2016 Mar 26.

Department of Chemical and Petroleum Engineering, University of Kansas, Lawrence, Kansas 66045. Electronic address:

Mechanical agitation of monoclonal antibody (mAb) solutions often leads to protein particle formation. In this study, various formulations of an immunoglobulin G (IgG) 1 mAb were subjected to different controlled interfacial stresses using a Langmuir trough, and protein particles formed at the interface and measured in bulk solution were characterized using atomic force microscopy and flow digital imaging. Results were compared to mAb solutions agitated in glass vials and unstressed controls. At lower pH, mAb solutions exhibited larger hysteresis in their surface pressure versus area isotherms and increased number of particles in bulk solution, when subjected to interfacial stresses. mAb samples subjected to 750-1000 interfacial compression-expansion cycles in 6 h contained high particle numbers in bulk solution, and displayed similar particulation trends when agitated in vials. At compression rates of 50 cycles in 6 h, however, particle levels in mAb solutions were comparable to unstressed controls, despite protein aggregates being present at the air-solution interface. These results suggest that while the air-solution interface serves as a nucleation site for initiating protein aggregation, the number of protein particles measured in bulk mAb solutions depends on the total number of compression cycles that proteins at the air-solution interface are subjected to within a fixed time.
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http://dx.doi.org/10.1016/j.xphs.2016.02.027DOI Listing
May 2016

Physiochemical Properties of Aluminum Adjuvants Elicit Differing Reorganization of Phospholipid Domains in Model Membranes.

Mol Pharm 2016 05 30;13(5):1731-7. Epub 2016 Mar 30.

Department of Chemical and Petroleum Engineering, University of Kansas , Lawrence, Kansas 66045, United States.

Most vaccines contain aluminum adjuvants; however, their exact mechanism of action remains unclear. A novel mechanism by Shi and colleagues proposes aluminum adjuvants may enhance immune activation by binding and reorganizing lipids that are key components of lipid rafts. To better understand the specificity of interaction between aluminum adjuvants and the cell membrane lipids, we present a biophysical study of lipid domain clustering in simple model phospholipid monolayers containing dipalmitoyl-phosphatidylcholine (DPPC) and dioleoyl-phosphatidylcholine (DOPC) exposed to two aluminum adjuvants, Alhydrogel and Adju-Phos. Surface pressure measurements and fluorescence microscopy images verified aluminum adjuvant-induced increase in lipid domain size, even in the key lipid raft components. Additionally, adjuvant induced lipid clustering differed based on the physicochemical properties of the adjuvants. Alhydrogel appeared to reduce monolayer compressibility and insert into the monolayer, while Adju-Phos induced more significant changes in domain size, without compromising the integrity of the monolayer. The Alhydrogel and Adju-Phos-mediated reorganization of phospholipid domains reported here supports the new mechanistic paradigm proposed by Shi and co-workers, and further suggests that lipid clustering is induced even in simple phospholipid membranes. The results present the basis for future exploration into lipid-mediated mechanisms of action for adjuvants.
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http://dx.doi.org/10.1021/acs.molpharmaceut.6b00111DOI Listing
May 2016

Charge Type, Charge Spacing, and Hydrophobicity of Arginine-Rich Cell-Penetrating Peptides Dictate Gene Transfection.

Mol Pharm 2016 Mar 24;13(3):1047-57. Epub 2016 Feb 24.

Department of Pharmaceutical Chemistry, University of Kansas , Lawrence, Kansas 66047, United States.

Noncovalent complexation of plasmid DNA (pDNA) with cell-penetrating peptides (CPPs) forms relatively large complexes with poor gene expression. Yet, condensing these CPP-pDNA complexes via addition of calcium chloride produces small and stable nanoparticles with high levels of gene expression. This simple formulation offered high transfection efficiency and negligible cytotoxicity in HEK-293 (a virus-immortalized kidney cell) and A549 (a human lung cancer cell line). Small changes in CPP charge type, charge spacing, and hydrophobicity were studied by using five arginine-rich CPPs: the well-known hydrophilic polyarginine R9 peptide, a hydrophilic RH9 peptide, and three amphiphilic peptides (RA9, RL9, and RW9) with charge distributions that favor membrane penetration. R9 and RW9 nanoparticles were significantly more effective than the other CPPs under most formulation conditions. However, these CPPs exhibit large differences in membrane penetration potential. Maximum transfection resulted from an appropriate balance of complexing with pDNA, releasing DNA, and membrane penetration potential.
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http://dx.doi.org/10.1021/acs.molpharmaceut.5b00871DOI Listing
March 2016

Combined effect of synthetic protein, Mini-B, and cholesterol on a model lung surfactant mixture at the air-water interface.

Biochim Biophys Acta 2016 Apr 15;1858(4):904-12. Epub 2016 Jan 15.

Department of Chemical Engineering, University of Kansas, KS 66045, United States. Electronic address:

The overall goal of this work is to study the combined effects of Mini-B, a 34 residue synthetic analog of the lung surfactant protein SP-B, and cholesterol, a neutral lipid, on a model binary lipid mixture containing dipalmitolphosphatidylcholine (DPPC) and palmitoyl-oleoyl-phosphatidylglycerol (POPG), that is often used to mimic the primary phospholipid composition of lung surfactants. Using surface pressure vs. mean molecular area isotherms, fluorescence imaging and analysis of lipid domain size distributions; we report on changes in the structure, function and stability of the model lipid-protein films in the presence and absence of varying composition of cholesterol. Our results indicate that at low cholesterol concentrations, Mini-B can prevent cholesterol's tendency to lower the line tension between lipid domain boundaries, while maintaining Mini-B's ability to cause reversible collapse resulting in the formation of surface associated reservoirs. Our results also show that lowering the line tension between domains can adversely impact monolayer folding mechanisms. We propose that small amounts of cholesterol and synthetic protein Mini-B can together achieve the seemingly opposing requirements of efficient LS: fluid enough to flow at the air-water interface, while being rigid enough to oppose irreversible collapse at ultra-low surface tensions.
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http://dx.doi.org/10.1016/j.bbamem.2016.01.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5705005PMC
April 2016

Phospholipid composition modulates carbon nanodiamond-induced alterations in phospholipid domain formation.

Langmuir 2015 May 28;31(18):5093-104. Epub 2015 Apr 28.

†Department of Chemical and Petroleum Engineering and ‡Department of Pharmaceutical Chemistry, The University of Kansas, Lawrence, Kansas 66045, United States.

The focus of this work is to elucidate how phospholipid composition can modulate lipid nanoparticle interactions in phospholipid monolayer systems. We report on alterations in lipid domain formation induced by anionically engineered carbon nanodiamonds (ECNs) as a function of lipid headgroup charge and alkyl chain saturation. Using surface pressure vs area isotherms, monolayer compressibility, and fluorescence microscopy, we found that anionic ECNs induced domain shape alterations in zwitterionic phosphatidylcholine lipids, irrespective of the lipid alkyl chain saturation, even when the surface pressure vs area isotherms did not show any significant changes. Bean-shaped structures characteristic of dipalmitoylphosphatidylcholine (DPPC) were converted to multilobed, fractal, or spiral domains as a result of exposure to ECNs, indicating that ECNs lower the line tension between domains in the case of zwitterionic lipids. For membrane systems containing anionic phospholipids, ECN-induced changes in domain packing were related to the electrostatic interactions between the anionic ECNs and the anionic lipid headgroups, even when zwitterionic lipids are present in excess. By comparing the measured size distributions with our recently developed theory derived by minimizing the free energy associated with the domain energy and mixing entropy, we found that the change in line tension induced by anionic ECNs is dominated by the charge in the condensed lipid domains. Atomic force microscopy images of the transferred anionic films confirm that the location of the anionic ECNs in the lipid monolayers is also modulated by the charge on the condensed lipid domains. Because biological membranes such as lung surfactants contain both saturated and unsaturated phospholipids with different lipid headgroup charges, our results suggest that when studying potential adverse effects of nanoparticles on biological systems the role of lipid compositions cannot be neglected.
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http://dx.doi.org/10.1021/la504923jDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4702515PMC
May 2015

Monitoring phases and phase transitions in phosphatidylethanolamine monolayers using active interfacial microrheology.

Soft Matter 2015 May;11(17):3313-21

Department of Chemical Engineering, University of Kansas, Lawrence, KS 66045, USA.

Active interfacial microrheology is a sensitive tool to detect phase transitions and headgroup order in phospholipid monolayers. The re-orientation of a magnetic nickel nanorod is used to explore changes in the surface rheology of 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine (DLPE) and 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine (DMPE), which differ by two CH2 groups in their alkyl chains. Phosphatidylethanolamines such as DLPE and DMPE are a major component of cell membranes in bacteria and in the nervous system. At room temperature, DLPE has a liquid expanded (LE) phase for surface pressure, Π < ∼38 mN m(-1); DMPE has an LE phase for Π < ∼7 mN m(-1). In their respective LE phases, DLPE and DMPE show no measurable change in surface viscosity with Π, consistent with a surface viscosity <10(-9) N s m(-1), the resolution of our technique. However, there is a measurable, discontinuous change in the surface viscosity at the LE to liquid condensed (LC) transition for both DLPE and DMPE. This discontinuous change is correlated with a significant increase in the surface compressibility modulus (or isothermal two-dimensional bulk modulus). In the LC phase of DMPE there is an exponential increase in surface viscosity with Π consistent with a two-dimensional free area model. The second-order LC to solid (S) transition in DMPE is marked by an abrupt onset of surface elasticity; there is no measurable elasticity in the LC phase. A measurable surface elasticity in the S phase suggests a change in the molecular ordering or interactions of the DMPE headgroups that is not reflected in isotherms or in grazing incidence X-ray diffraction. This onset of measurable elasticity is also seen in DLPE, even though no indication of a LC-S transition is visible in the isotherms.
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http://dx.doi.org/10.1039/c4sm02900cDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4408260PMC
May 2015

Effect of lipid headgroup charge and pH on the stability and membrane insertion potential of calcium condensed gene complexes.

Langmuir 2015 Apr 30;31(14):4232-45. Epub 2015 Mar 30.

†Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047, United States.

Noncovalently condensed complexes of genetic material, cell penetrating peptides (CPPs), and calcium chloride present a nonviral route to improve transfection efficiency of nucleic acids (e.g., pDNA and siRNA). However, the exact mechanisms of membrane insertion and delivery of macromolecule complexes to intracellular locations as well as their stability in the intracellular environment are not understood. We show that calcium condensed gene complexes containing different hydrophilic (i.e., dTAT, K9, R9, and RH9) and amphiphilic (i.e., RA9, RL9, and RW9) CPPs formed stable cationic complexes of hydrodynamic radii 100 nm at neutral pH. However, increasing the acidity caused the complexes to become neutral or anionic and increase in size. Using zwitterionic and anionic phospholipid monolayers as models that mimic the membrane composition of the outer leaflet of cell membranes and intracellular vesicles and pHs that mimic the intracellular environment, we study the membrane insertion potential of these seven gene complexes (CPP/pDNA/Ca(2+) complexes) into model membranes. At neutral pH, all gene complexes demonstrated the highest insertion potential into anionic phospholipid membranes, with complexes containing amphiphilic peptides showing the maximum insertion. However, at acidic pH, the gene complexes demonstrated maximum monolayer insertion into zwitterionic lipids, irrespective of the chemical composition of the CPP in the complexes. Our results suggest that in the neutral environment the complexes are unable to penetrate the zwitterionic lipid membranes but can penetrate through the anionic lipid membranes. However, the acidic pH mimicking the local environment in the late endosomes leads to a significant increase in adsorption of the complexes to zwitterionic lipid headgroups and decreases for anionic headgroups. These membrane-gene complex interactions may be responsible for the ability of the complexes to efficiently enter the intracellular environment through endocytosis and escape from the endosomes to effectively deliver their genetic payload.
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http://dx.doi.org/10.1021/la504970nDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5704962PMC
April 2015

Dynamic measurements of membrane insertion potential of synthetic cell penetrating peptides.

Langmuir 2013 Dec 2;29(49):15336-49. Epub 2013 Dec 2.

Department of Pharmaceutical Chemistry, University of Kansas , Lawrence, Kansas 66047, United States.

Cell penetrating peptides (CPPs) have been established as excellent candidates for mediating drug delivery into cells. When designing synthetic CPPs for drug delivery applications, it is important to understand their ability to penetrate the cell membrane. In this paper, anionic or zwitterionic phospholipid monolayers at the air-water interface are used as model cell membranes to monitor the membrane insertion potential of synthetic CPPs. The insertion potential of CPPs having different cationic and hydrophobic amino acids were recorded using a Langmuir monolayer approach that records peptide adsorption to model membranes. Fluorescence microscopy was used to visualize alterations in phospholipid packing due to peptide insertion. All CPPs had the highest penetration potential in the presence of anionic phospholipids. In addition, two of three amphiphilic CPPs inserted into zwitterionic phospholipids, but none of the hydrophilic CPPs did. All the CPPs studied induced disruptions in phospholipid packing and domain morphology, which were most pronounced for amphiphilic CPPs. Overall, small changes to amino acids and peptide sequences resulted in dramatically different insertion potentials and membrane reorganization. Designers of synthetic CPPs for efficient intracellular drug delivery should consider small nuances in CPP electrostatic and hydrophobic properties.
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http://dx.doi.org/10.1021/la403370pDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3918496PMC
December 2013

Lysine221 is the general base residue of the isochorismate synthase from Pseudomonas aeruginosa (PchA) in a reaction that is diffusion limited.

Arch Biochem Biophys 2013 Oct 11;538(1):49-56. Epub 2013 Aug 11.

Molecular Biosciences, University of Kansas, Lawrence, KS 66045, United States.

The isochorismate synthase from Pseudomonas aeruginosa (PchA) catalyzes the conversion of chorismate to isochorismate, which is subsequently converted by a second enzyme (PchB) to salicylate for incorporation into the salicylate-capped siderophore pyochelin. PchA is a member of the MST family of enzymes, which includes the structurally homologous isochorismate synthases from Escherichia coli (EntC and MenF) and salicylate synthases from Yersinia enterocolitica (Irp9) and Mycobacterium tuberculosis (MbtI). The latter enzymes generate isochorismate as an intermediate before generating salicylate and pyruvate. General acid-general base catalysis has been proposed for isochorismate synthesis in all five enzymes, but the residues required for the isomerization are a matter of debate, with both lysine221 and glutamate313 proposed as the general base (PchA numbering). This work includes a classical characterization of PchA with steady state kinetic analysis, solvent kinetic isotope effect analysis and by measuring the effect of viscosogens on catalysis. The results suggest that isochorismate production from chorismate by the MST enzymes is the result of general acid-general base catalysis with a lysine as the base and a glutamic acid as the acid, in reverse protonation states. Chemistry is determined to not be rate limiting, favoring the hypothesis of a conformational or binding step as the slow step.
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http://dx.doi.org/10.1016/j.abb.2013.07.026DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3784010PMC
October 2013

Delivery and performance of surfactant replacement therapies to treat pulmonary disorders.

Ther Deliv 2013 Aug;4(8):951-80

Department of Pharmaceutical Chemistry, The University of Kansas, Lawrence, KS 66045, USA.

Lung surfactant is crucial for optimal pulmonary function throughout life. An absence or deficiency of surfactant can affect the surfactant pool leading to respiratory distress. Even if the coupling between surfactant dysfunction and the underlying disease is not always well understood, using exogenous surfactants as replacement is usually a standard therapeutic option in respiratory distress. Exogenous surfactants have been extensively studied in animal models and clinical trials. The present article provides an update on the evolution of surfactant therapy, types of surfactant treatment, and development of newer-generation surfactants. The differences in the performance between various surfactants are highlighted and advanced research that has been conducted so far in developing the optimal delivery of surfactant is discussed.
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http://dx.doi.org/10.4155/tde.13.72DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3840129PMC
August 2013

Interface-induced disassembly of a self-assembled two-component nanoparticle system.

Langmuir 2013 Mar 7;29(11):3654-61. Epub 2013 Mar 7.

Department of Chemical and Petroleum Engineering, The University of Kansas, Lawrence, Kansas 66046, USA.

We present a study of static and dynamic interfacial properties of self-assembled polyelectrolyte complex nanoparticles (size 110-120 nm) containing entrapped surfactant molecules at a fluid/fluid interface. Surface tension vs time measurements of an aqueous solution of these polyelectrolyte complex nanoparticles (PCNs) show a concentration-dependent biphasic adsorption to the air/water interface while interfacial microrheology data show a concentration-dependent initial increase in the surface viscosity (up to 10(-7) N·m/s), followed by a sharp decrease (10(-9) N·m/s). Direct visualization of the air/water interface shows disappearance of particles from the interface over time. On the basis of these observations, we propose that the PCNs at fluid/fluid interfaces exist in two states: initial accumulation of PCNs at the air/water interface as nanoparticles, followed by interface induced disassembly of the accumulated PCNs into their components. The lack of change in particle size, charge, and viscosity of the bulk aqueous solution of PCNs with time indicates that this disintegration of the self-assembled PCNs is an interfacial phenomenon. Changes in energy encountered by the PCNs at the interface lead to instability of the self-assembled system and dissociation into its components. Such systems can be used for applications requiring directed delivery and triggered release of entrapped surfactants or macromolecules at fluid/fluid interfaces.
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http://dx.doi.org/10.1021/la400062bDOI Listing
March 2013

Lipid-protein interactions alter line tensions and domain size distributions in lung surfactant monolayers.

Biophys J 2012 Jan 3;102(1):56-65. Epub 2012 Jan 3.

Department of Chemical Engineering, University of Kansas, Lawrence, Kansas, USA.

The size distribution of domains in phase-separated lung surfactant monolayers influences monolayer viscoelasticity and compressibility which, in turn, influence monolayer collapse and set the compression at which the minimum surface tension is reached. The surfactant-specific protein SP-B decreases the mean domain size and polydispersity as shown by fluorescence microscopy. From the images, the line tension and dipole density difference are determined by comparing the measured size distributions with a theory derived by minimizing the free energy associated with the domain energy and mixing entropy. We find that SP-B increases the line tension, dipole density difference, and the compressibility modulus at surface pressures up to the squeeze-out pressure. The increase in line tension due to SP-B indicates the protein avoids domain boundaries due to its solubility in the more fluid regions of the film.
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http://dx.doi.org/10.1016/j.bpj.2011.11.4007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3250676PMC
January 2012

Active interfacial shear microrheology of aging protein films.

Phys Rev Lett 2010 Jan 4;104(1):016001. Epub 2010 Jan 4.

Department of Chemical Engineering, University of California, Santa Barbara, California 93117, USA.

The magnetically driven rotation of 300 nm diameter rods shows the surface viscosity of albumin at an air-water interface increases from 10(-9) to 10(-5) N s/m over 2 h while the surface pressure saturates in minutes. The increase in surface viscosity is not accompanied by a corresponding increase in elasticity, suggesting that the protein film anneals with time, resulting in a more densely packed film leading to increased resistance to shear. The nanometer dimensions of the rods provide the same sensitivity as passive microrheology with an improved ability to measure more viscous films.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3407536PMC
http://dx.doi.org/10.1103/PhysRevLett.104.016001DOI Listing
January 2010

Comment on "osmotic propulsion: the osmotic motor".

Phys Rev Lett 2009 Apr 16;102(15):159801; discussion 159802. Epub 2009 Apr 16.

Institut für Experimentalphysik V, Universität Bayreuth, 95440 Bayreuth, Germany.

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http://dx.doi.org/10.1103/PhysRevLett.102.159801DOI Listing
April 2009

Immersion of charged nanoparticles in a salt solution/air interface.

J Phys Chem B 2008 Aug 22;112(32):9565-7. Epub 2008 Jul 22.

Electrostatic interactions strongly affect the immersion depth of nanoparticles into an interface. We prove this statement by measuring the diffusion constant of charged nanoparticles at a sodium chloride solution/air interface. Interfacial diffusion of nanoparticles slows down with increasing ionic strength of the sodium chloride solution. Hydrodynamic calculations are used to estimate the immersion depth from the diffusion constant, suggesting that nanoparticles with a carboxylate surface are only slightly immersed into a bare air/water interface. With increasing molarities of sodium chloride, the immersion depth increases to complete immersion for a 10(-2) molar solution. Our experiments show that the location of nanoparticles at interfaces is determined by an intricate interplay between the electrostatic properties of the solution/air interface, the solution/solid interface, and the classical contact angle.
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http://dx.doi.org/10.1021/jp805042jDOI Listing
August 2008

Curvature driven transport of mouse macrophages in a pulsating magnetic garnet film ratchet.

J Phys Chem B 2007 Nov 20;111(45):13097-100. Epub 2007 Oct 20.

Department of Chemistry and Biochemistry, The Florida State University, Tallahassee, Florida 32306, USA.

Magnetic fields varying on the colloidal length scale are used for the directed transport of magnetically labeled biological cells. The transport is achieved by using the ratchet effect which relies on an asymmetric, symmetry broken periodic potential where nonequilibrium fluctuations or oscillations generate a net cell current. Ferrofluid ingested mouse macrophages were placed on a magnetic garnet film with alternating stripe domain patterns, and a pulsating magnetic potential is provided by superposing an oscillating magnetic field normal to the film. The symmetry of the resulting periodic stripe potential is broken locally by the curvature of the stripes. We show, both experimentally and theoretically, the curvature of such stripes required for inducing directed transport of the macrophages in the ratchet. This may be useful for microfluidic devices such as a digital colloidal shift register for magnetically labeled biological cells.
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http://dx.doi.org/10.1021/jp0764485DOI Listing
November 2007

Orientations of overdamped magnetic nanorod-gyroscopes.

Nano Lett 2007 Apr 23;7(4):1010-2. Epub 2007 Mar 23.

Department of Chemistry and Biochemistry, The Florida State University, Tallahassee Florida 32306-4390, USA.

Overdamped magnetic nanorod-gyroscopes driven by a rotating magnetic field undergo a series of reorientations when sedimenting on top of a surface in a viscous liquid. By changing the amplitude and the rotation frequency of the driving magnetic field, the nanorod-gyroscope either synchronizes or desynchronizes with the field and rotates either around its long or short axis. The different regimes of motion are explained theoretically by coupling the nanorod-gyroscopes motion to the creeping flow equations of the surrounding fluid. It is shown that friction anisotropy plays an important role for the orientation of the nanorod-gyroscopes.
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http://dx.doi.org/10.1021/nl0701556DOI Listing
April 2007
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