Browsing by Author "Berbari, Edward J."
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Item Adipose stromal cells enhance keratinocyte survival and migration in vitro, and graft revascularization in mouse wound healing model(2013-12-11) Knowles, Kellen Alexander; Berbari, Edward J.; March, Keith Leonard, 1963-; Ji, JulieIn the US, more than 1 million burn injuries are reported annually. About 45,000 injuries due to fires and burns result in hospitalization and ten percent of these result in death every year. Advances in burn treatment have led to a reduction in mortality rate over the last decades. Since more patients are surviving the initial resuscitation phase even with very large areas of skin being burned away, wound care has become increasingly important to ensure continued patient survival and improvement. While currently a common treatment for third degree burn wounds, skin grafts have several drawbacks. The availability of donor sites for autografts may be limited, especially in incidences of extensive skin loss. The rejection associated with the use of allografts and xenografts may render them inadequate or undesirable. Even if a suitable graft is found, poor retention due to infection, hematoma, and low vascularity at the recipient site are other drawbacks associated with the use of skin grafts as a primary treatment for severe burn wounds. As such, research has been done into alternative treatments, which include but are not limited to artificial skin, cell therapy, and growth factor application. We propose the delivery of adipose derived stem cells (ASC) in combination with endothelial progenitor cells (EC) via Integra Dermal Regenerative Template (DRT) to promote faster graft vascularization and thus faster healing of wounds. Integra DRT is an acellular skin substitute that consists of a dermal layer composed of bovine collagen and chondroitin-6-sulfate glycosaminoglycan, and an "epidermal" layer, which consists of silicone polymer. This silicone layer is removed after the collagen matrix is adequately vascularized (usually takes 2-3 weeks), and then a thin layer autograft is applied to the top of the neo-dermis. ASC are derived from the stromal-vascular fraction (SVF) of adipose tissue and are a readily available, pluripotent, mesenchymal cell known to promote angiogenesis. They are being explored as a treatment for a myriad of diseases and conditions, including wound healing. In combination with ECs, they form stable microvessel networks in vitro and in vivo. In our work, we found that ASC+EC form stable microvessel networks when cultured on Integra DRT. Also, ASC and ASC+EC conditioned media promoted both survival and migration of human epidermal keratinocytes compared to control medium. In a full thickness wound healing model, using healthy NSG mice, the ASC+EC case showed a significantly higher rate of wound closure compared to control. Based on best linear unbiased estimates (BLUE), the difference between the healing rates of ASC alone treatment and the Control treatment group is -0.45 +/- 0.22 mm²/day (p=0.041), which is not less than 0.025 and thus not statistically significant (Bonferroni Adjusted). However, the BLUE for the difference between the ASC+EC group and the Control group healing rates is -0.55 +/- 0.28 mm²/day (p = 0.017<0.025, Bonferroni Adjusted), which is statistically significant. Histology revealed a significantly higher number of vessels compared to control in both ASC alone and ASC+EC case. CD31 staining revealed the presence of human vessels in ASC+EC treatment scaffolds. We conclude that the combination of ASC and EC can be used to accelerate healing of full-thickness wounds when delivered to site of the wound via Integra. This result is especially compelling due to the fact that the mice used were all healthy. Thus our treatment shows an improvement in healing rate even compared to normal wound healing.Item Analysis of the Bioelectric Impedance of the Tissue-Electrode Interface Using a Novel Full-Spectrum Approach(2014-01-15) Sempsrott, David Robert; Yoshida, Ken; Salama, Paul; Berbari, Edward J.Non-invasive surface recording of bioelectric potentials continues to be an essential tool in a variety of research and medical diagnostic procedures. However, the integrity of these recordings, and hence the reliability of subsequent analysis, diagnosis, or recommendations based on the recordings, can be significantly compromised when various types of noise are allowed to penetrate the recording circuit and contaminate the signals. In particular, for bioelectric phenomena in which the amplitude of the biosignal is relatively low, such as muscle activity (typically on the order of millivolts) or neural traffic (microvolts), external noise may substantially contaminate or even completely overwhelm the signal. In such circumstances, the tissue-electrode interface is typically the primary point of signal contamination since its impedance is relatively high compared to the rest of the recording circuit. Therefore, in the recording of low-amplitude biological signals, it is of paramount importance to minimize the impedance of the tissue-electrode interface in order to consistently obtain low-noise recordings. The aims of the current work were (1) to complete the development of a set of tools for rapid, simple, and reliable full-spectrum characterization and analytical modeling of the complex impedance of the tissue-electrode interface, and (2) to characterize the interfacial impedance and signal-to-noise ratio (SNR) at the surface of the skin across a variety of preparation methods and determine a factor or set of factors that contribute most effectively to the reduction of tissue-electrode impedance and noise contamination during recording. Specifically, we desired to test an initial hypothesis that surface abrasion is the principal determining factor in skin preparation to achieve consistently low-impedance, low-noise recordings. During the course of this master’s study, (1) a system with portable, battery-powered hardware and robust acquisition/analysis software for broadband impedance characterization has been achieved, and (2) the effects of skin preparation methods on the impedance of the tissue-electrode interface and the SNR of surface electromyographic recordings have been systematically quantified and compared in human subjects. We found our hypothesis to be strongly supported by the results: the degree of surface abrasion was the only factor that could be correlated to significant differences in either the interfacial impedance or the SNR. Given these findings, we believe that abrasion holds the key to consistently obtaining a low-impedance contact interface and high-quality recordings and should thus be considered an essential component of proper skin preparation prior to attachment of electrodes for recording of small bioelectric surface potentials.Item Biomechanical and morphological characterization of common iliac vein remodeling: Effects of venous reflux and hypertension(2014) Brass, Margaret Mary; Kassab, Ghassan S. (Ghassan Sleewa), 1965-; Na, Sungsoo; Berbari, Edward J.The passive properties of the venous wall are important in the development of venous pathology. Increase in venous pressure due to retrograde flow (reflux) and obstruction of venous flow by intrinsic and extrinsic means are the two possible mechanisms for venous hypertension. Reflux is the prevailing theory in the etiology of venous insufficiency. The objective of this thesis is to quantify the passive biomechanical response and structural remodeling of veins subjected to chronic venous reflux and hypertension. To investigate the effects of venous reflux on venous mechanics, the tricuspid valve was injured chronically in canines by disrupting the chordae tendineae. The conventional inflation-extension protocol in conjunction with intravascular ultrasound (IVUS) was utilized to investigate the passive biomechanical response of both control common iliac veins (from 9 dogs) and common iliac veins subjected to chronic venous reflux and hypertension (from 9 dogs). The change in thickness and constituent composition as a result of chronic venous reflux and hypertension was quantified using multiphoton microscopy (MPM) and histological evaluation. Biomechanical results indicate that the veins stiffened and became less compliant when exposed to eight weeks of chronic venous reflux and hypertension. The mechanical stiffening was found to be a result of a significant increase in wall thickness (p < 0.05) and a significant increase in the collagen to elastin ratio (p < 0.05). After eight weeks of chronic reflux, the circumferential Cauchy stress significantly reduced (p < 0.05) due to wall thickening, but was not restored to control levels. This provided a useful model for development and further analysis of chronic venous insufficiency and assessment of possible intervention strategies.Item Developing a Neural Signal Processor Using the Extended Analog Computer(2013-08-21) Soliman, Muller Mark; Yoshida, Ken; Eberhart, Russell C.; Mills, Jonathan W. (Jonathan Wayne); Berbari, Edward J.Neural signal processing to decode neural activity has been an active research area in the last few decades. The next generation of advanced multi-electrode neuroprosthetic devices aim to detect a multiplicity of channels from multiple electrodes, making the relatively time-critical processing problem massively parallel and pushing the computational demands beyond the limits of current embedded digital signal processing (DSP) techniques. To overcome these limitations, a new hybrid computational technique was explored, the Extended Analog Computer (EAC). The EAC is a digitally confgurable analog computer that takes advantage of the intrinsic ability of manifolds to solve partial diferential equations (PDEs). They are extremely fast, require little power, and have great potential for mobile computing applications. In this thesis, the EAC architecture and the mechanism of the formation of potential/current manifolds was derived and analyzed to capture its theoretical mode of operation. A new mode of operation, resistance mode, was developed and a method was devised to sample temporal data and allow their use on the EAC. The method was validated by demonstration of the device solving linear diferential equations and linear functions, and implementing arbitrary finite impulse response (FIR) and infinite impulse response (IIR) linear flters. These results were compared to conventional DSP results. A practical application to the neural computing task was further demonstrated by implementing a matched filter with the EAC simulator and the physical prototype to detect single fiber action potential from multiunit data streams derived from recorded raw electroneurograms. Exclusion error (type 1 error) and inclusion error (type 2 error) were calculated to evaluate the detection rate of the matched filter implemented on the EAC. The detection rates were found to be statistically equivalent to that from DSP simulations with exclusion and inclusion errors at 0% and 1%, respectively.Item A digitally invertible universal amplifier for recording and processing of bioelectric signals(2014-01-03) Mauser, Kevin Alton; Yoshida, Ken; Berbari, Edward J.; Salama, PaulThe recording and processing of bioelectric signals over the decades has led to the development of many different types of analog filtering and amplification techniques. Meanwhile, there have also been many advancements in the realm of digital signal processing that allow for more powerful analysis of these collected signals. The issues with present acquisition schemes are that (1) they introduce irreversible distortion to the signals and may ultimately hinder analyses that rely on the unique morphological differences between bioelectric signal events and (2) they do not allow the collection of frequencies in the signal from direct-current (DC) to high-frequencies. The project put forth aims to overcome these two issues and present a new scheme for bioelectric signal acquisition and processing. In this thesis, a system has been developed, verified, and validated with experimental data to demonstrate the ability to build an invertible universal amplifier and digital restoration scheme. The thesis is primarily divided into four sections which focus on (1) the introduction and background information, (2) theory and development, (3) verification implementation and testing, and (4) validation implementation and testing. The introduction and background provides pertinent information regarding bioelectric signals and recording practices for bioelectric signals. It also begins to address some of the issues with the classical and present methods for data acquisition and make the case for why an invertible universal amplifier would be better. The universal amplifier transfer function and architecture are discussed and presented along with the development and optimization of the characterization and the inversion, or restoration, filter process. The developed universal amplifier, referred to as the invertible universal amplifier (IUA), while the universal amplifier and the digital restoration scheme together are referred to as the IUA system. The IUA system is then verified on the bench using typical square, sine, and triangle waveforms with varying offsets and the results are presented and discussed. The validation is done with in-vivo experiments showing that the IUA system may be used to acquire and process bioelectric signals with percent error less than to 6% when post-processed using estimated characteristics of and when compared to a standard flat bandwidth high-pass cutoff amplifier.Item Exploration of Sinusoidal Low Frequency Alternating Current Stimulation to Block Peripheral Nerve Activity(2024-05) Horn, Michael Ryne; Yoshida, Ken; Ward, Mathew P.; Berbari, Edward J.; Schild, John H.Sinusoidal low frequency alternating current (LFAC) stimulation is a novel mode of electrical modulation observed in the Bioelectroics Lab in 2017. LFAC is capable of blocking the single fiber action potentials (APs) of the earthworm with only a few 100’s of µA. The goal of this dissertation was to further explore and characterize the LFAC waveform to determine it’s feasibility as a method for block in the mammalian peripheral nervous system (PNS). To better understand the mechanisms of LFAC block (LFACb), a blend of in-silico modeling work was explored and the predictions were validated with ex-vivo and in-vivo experiments. This dissertation is divided into five chapters. The first chapter will explore the history of bioelectricity, the current state of in-silico modeling and methods of nerve block used in the PNS. The second chapter explores a major modeling assumption, the conductivity and permittivity of the nerve laminae of a mammalian nerve bundle. Four point electrochemical impedance spectroscopy (EIS) was performed on excised canine vagus nerve to evaluate the electrical properties of the perineurium and epineurium. This study’s result, found that the corner frequency of the perineurium (2.6kHz) and epineurium (370Hz) were much lower than previously assumed. This explain a major difference between LFACb and the more established kilohertz frequency alternating current (kHFAC) block. The third chapter revisits the initial earthworm experiments during the discovery of LFACb. The effect of conduction slowing was observed in these earthworm experiments and were also seen in a mammalian canine vagus nerve and in the Horn-Yoshida-Schild (HYS) autonomic unmyelinated axon mode. These experiments showed that LFACb occurs as a cathodic block in which the sodium channels are held inactive. Chapter 4 explored the window between LFACb and LFAC activation (LFACa). The window between the two states was describes by LFAC amplitude and LFAC frequency in an in-vivo rat sciatic nerve and an in-silico model of a myelinated motor neuron, the McIntyre-Richardson-Grill (MRG) axon model. Geometrical effects were also observed by varying the bipolar pair of contacts used to deliver the LFACb waveform from an asymmetrical tripolar cuff electrode. Plantar flexor force measurements and electromyography (EMG) of the lateral gastrocnemius (LG) and soleus (Sol) were used to quantify the effects of the LFAC waveform. Convergence between in-silico modeling and in-vivo results showed promise that modeling efforts could be used with confidence to explore the LFAC block-activation more completly. LFACa was found to be highly dependent on frequency with increasing frequency lowering the threshold of activation. LFACb was shown to be mostly invariant to frequency. The final chapter takes the information found in this dissertation and summarizes it. Future work on LFAC is also proposed and the hypothesized results presented with the findings from this dissertation and available literature.Item In Vitro and In Silico Analysis of Osteoclastogenesis in Response to Inhibition of De-phosphorylation of EIF2alpha by Salubrinal and Guanabenz(2013) Tanjung, Nancy Giovanni; Yokota, Hiroki, 1955-; Ji, Julie; Na, Sungsoo; Berbari, Edward J.An excess of bone resorption over bone formation leads to osteoporosis, resulting in a reduction of bone mass and an increase in the risk of bone fracture. Anabolic and anti-resorptive drugs are currently available for treatment, however, none of these drugs are able to both promote osteoblastogenesis and reduce osteoclastogenesis. This thesis focused on the role of eukaryotic translation initiation factor 2 alpha (eIF2alpha), which regulates efficiency of translational initiation. The elevation of phosphorylated eIF2alpha was reported to stimulate osteoblastogenesis, but its effects on osteoclastogenesis have not been well understood. Using synthetic chemical agents such as salubrinal and guanabenz that are known to inhibit the de-phosphorylation of eIF2alpha, the role of phosphorylation of eIF2alpha in osteoclastogenesis was investigated in this thesis. The questions addressed herein were: Does the elevation of phosphorylated eIF2alpha (p-eIF2alpha) by salubrinal and guanabenz alter osteoclastogenesis? If so, what regulatory mechanism mediates the process? It was hypothesized that p-eIF2alpha could attenuate the development of osteoclast by regulating the transcription factor(s) amd microRNA(s) involved in osteoclastogenesis. To test this hypothesis, we conducted in vitro and in silico analysis of the responses of RAW 264.7 pre-osteoclast cells to salubrinal and guanabenz. First, the in vitro results revealed that the elevated level of phosphorylated eIF2alpha inhibited the proliferation, differentiation, and maturation of RAW264.7 cells and downregulated the expression of NFATc1, a master transcription factor of osteoclastogenesis. Silencing eIF2alpha by RNA interference suppressed the downregulation of NFATc1, suggesting the involvement of eIF2alpha in regulation of NFATc1. Second, the in silico results using genome-wide expression data and custom-made Matlab programs predicted a set of stimulatory and inhibitory regulator genes as well as microRNAs, which were potentially involved in the regulation of NFATc1. RNA interference experiments indicated that the genes such as Zfyve21 and Ddit4 were primary candidates as an inhibitor of NFATc1. In summary, the results showed that the elevation of p-eIF2alpha by salubrinal and guanabenz leads to attenuation of osteoclastogenesis through the downregulation of NFATc1. The regulatory mechanism is mediated by eIF2alpha signaling, but other signaling pathways are likely to be involved. Together with the previous data showing the stimulatory role of p-eIF2alpha in osteoblastogenesis, the results herein suggest that eIF2alpha-mediated signaling could provide a novel therapeutic target for treatment of osteoporosis by promoting bone formation and reducing bone resorption.Item Monte Carlo Simulation to Study Propagation of Light through Biological Tissues(2012-09-20) Prabhu Verleker, Akshay; Berbari, Edward J.; Stantz, Keith; Yoshida, KenPhotoacoustic Imaging is a non-invasive optical imaging modality used to image biological tissues. In this method, a pulsating laser illuminates a region of tissues to be imaged, which then generates an acoustic wave due to thermal volume expansion. This wave is then sensed using an acoustic sensor such as a piezoelectric transducer and the resultant signal is converted into an imaging using the back projection algorithm. Since different types of tissues have different photo-acoustic properties, this imaging modality can be used for imaging different types of tissues and bodily organ systems. This study aims at quantifying the process of light conversion into the acoustic signal. Light travels through tissues and gets attenuated (scattered or absorbed) or reflected depending on the optical properties of the tissues. The process of light propagation through tissues is studied using Monte Carlo simulation software which predicts the propagation of light through tissues of various shapes and with different optical properties. This simulation gives the resultant energy distribution due to light absorption and scattering on a voxel by voxel basis. The Monte Carlo code alone is not sufficient to validate the photon propagation. The success of the Monte Carlo code depends on accurate prediction of the optical properties of the tissues. It also depends on accurately depicting tissue boundaries and thus the resolution of the imaging space. Hence, a validation algorithm has been designed so as to recover the optical properties of the tissues which are imaged and to successfully validate the simulation results. The accuracy of the validation code is studied for various optical properties and boundary conditions. The results are then compared and validated with real time images obtained from the photoacoustic scanner. The various parameters for the successful validation of Monte Carlo method are studied and presented. This study is then validated using the algorithm to study the conversion of light to sound. Thus it is a significant step in the quantification of the photoacoustic effect so as to accurately predict tissue properties.Item A Novel Approach to Peripheral Nerve Activation Using Low Frequency Alternating Currents(2020-08) Al Hawwash, Awadh Mubarak M.; Yoshida, Ken; Berbari, Edward J.; Schild, John H.The standard electrical stimulation waveform used for electrical activation of nerve is a rectangular pulse or a charge balanced rectangular pulse, where the pulse width is typically in the range of ∼100 µsec through ∼1000 µsec. In this work, we explore the effects of a continuous sinusoidal waveform with a frequency ranging from 5 through 20 Hz, which was named the Low Frequency Alternating Current (LFAC) waveform. The LFAC waveform was explored in the Bioelectronics Laboratory as a novel means to evoke nerve block. However, in an attempt to evoke complete nerve block on a somatic motor nerve, increasing the amplitude of the LFAC waveform unexpectedly produced nerve activation, and elicited a strong non-fatiguing muscle contraction in the anesthetized rabbit model (unpublished observation). The present thesis aimed to further explore the phenomenon to measure the effect of LFAC waveform frequency and amplitude on nerve activation. In freshly excised canine cervical vagus nerve (n=3), it was found that the LFAC waveform at 5, 10, and 20 Hz produced burst modulated activity. Compound action potentials (CAP) synchronous to the stimuli was absent from the electroneurogram (ENG) recordings. When applied in-vivo, LFAC was capable of activating the cervical vagus nerve fibers in anaesthetized swine (n=5) and induced the Hering-Breuer reflex. Additionally, when applied in-vivo to anesthetized Sprague Dawley rats (n=4), the LFAC waveform was able to activate the left sciatic nerve fibers and induced muscle contractions. The results demonstrate that LFAC activation was stochastic, and asynchronous to the stimuli unlike conventional pulse stimulation where nerve and muscle response simultaneously and synchronously to stimulus. The activation thresholds were found to be frequency dependent. As the waveform frequency increases the required current amplitude decreases. These experiments also implied that the LFAC phenomenon was most likely to be fiber type-size dependent but that more sophisticated exploration should be addressed before reaching clinical applications. In all settings, the LFAC amplitude was within the water window preventing irreversible electrochemical reactions and damages to the cuff electrodes or nerve tissues. This thesis also reconfirms the preliminary LFAC activation discovery and explores multiple methods to evaluate the experimental observations, which suggest the feasibility of the LFAC waveform at 5, 10, and 20 Hz to activate autonomic and somatic nerve fibers. LFAC appears to be a promising new technique to activate peripheral nerve fibers.Item PREPARATION AND EVALUATION OF NOVEL ANTIBACTERIAL DENTAL RESIN COMPOSITES(2013-07-12) Chong, Voon Joe; Xie, Dong; Na, Sungsoo; Li, Jiliang; Berbari, Edward J.Both quaternary ammonium bromide (QAB) and furanone derivatives were synthesized, characterized and formulated into dental resin composites for improved antibacterial properties. Compressive strength (CS) and S. mutans viability were used to evaluate the mechanical strength and antibacterial activity of the restoratives. The effects of chain length, loading, saliva and aging on CS and S. mutans viability were investigated. Chapter 2 describes how we studied and evaluated the formulated antibacterial resin composites by incorporating the synthesized QAB-containing oligomers into the formulation. The results show that all the QAB-modified resin composites showed significant antibacterial activity and mechanical strength reduction. Increasing chain length and loading significantly enhanced the antibacterial activity but dramatically reduced the CS as well. The 30-day aging study showed that the incorporation of the QAB accelerated the degradation of the composite, suggesting that the QAB may not be well suitable for development of antibacterial dental resin composites or at least the QAB loading should be well controlled. Chapter 3 describes how we studied and evaluated the formulated antibacterial resin composite by incorporating the synthesized furanone derivative into the formulation. The results show that the modified resin composites showed a significant antibacterial activity without substantially decreasing the mechanical strengths. With 5 to 30% addition of the furanone derivative, the composite kept its original CS unchanged but showed a significant antibacterial activity with a 16-68% reduction in the S. mutans viability. Further, the antibacterial function of the new composite was found not to be affected by human saliva. The aging study indicates that the composite may have a long-lasting antibacterial function. In summary, we have developed a novel QAB- and furanone-containing antibacterial system for dental restoratives. Both QAB- and furanone-modified resin composites have demonstrated significant antibacterial activities. The QAS-modified experimental resin composite may not be well suitable for development of antibacterial dental resin composites due to its accelerated degradation in water unless the QAB loading is well controlled. The furanone-modified resin composite shows nearly no reduction in mechanical strength after incorporation of the antibacterial furanone derivative. It appears that the furanone-modified resin composite is a clinically attractive dental restorative that can be potentially used for long-lasting restorations due to its high mechanical strength and permanent antibacterial function.