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Title: PARASYMPATHETIC NERVE DERIVED EXOSOMES INHIBIT HYPERGLYCEMIA INDUCED APOPTOSIS IN CARDIOMYOBLAST CELLS.
Creator: Singla, Reetish K, Cheng, Zixi (Jack), University of Central Florida
Abstract / Description: Diabetic cardiomyopathy involves both forms of cardiac cell cell death such as apoptosis and necrosis. However, this remains unknown whether hyperglycemia induced apoptosis in the cell culture system is inhibited by parasympathetic nerve derived exosomes. We isolated parasympathetic and sympathetic nerves and derived exosomes. We developed hyperglycemia induced apoptosis in H9c2 cells. H9c2 cells were divided into 4 groups: 1) Control, 2) H9c2+ Glucose 100 mmol, 3) H9c2+ Glucose ...
Show moreDiabetic cardiomyopathy involves both forms of cardiac cell cell death such as apoptosis and necrosis. However, this remains unknown whether hyperglycemia induced apoptosis in the cell culture system is inhibited by parasympathetic nerve derived exosomes. We isolated parasympathetic and sympathetic nerves and derived exosomes. We developed hyperglycemia induced apoptosis in H9c2 cells. H9c2 cells were divided into 4 groups: 1) Control, 2) H9c2+ Glucose 100 mmol, 3) H9c2+ Glucose +parasympathetic-exo, 4) H9c2+ Glucose+sympathetic-exo. We determined cell proliferation and viability with MTT assay kit and apoptosis with TUNEL staining and cell death detection ELISA kit. Data was further confirmed with pro-apoptotic proteins caspase-3 and BAX and anti-apoptotic protein Bcl2. High glucose exposed H9c2 cells significantly reduced cell viability which is improved by parasympathetic-exo but not by sympathetic-exo. Increased apoptosis in hyperglycemia in H9c2 cells confirmed with TUNEL staining and cell death ELISA was significantly (p
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Date Issued: 2018
Identifier: CFH2000378, ucf:45833
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFH2000378

Title: MICROSCOPIC ANALYSIS OF SYMPATHETIC AND PARASYMPATHETIC DISTRIBUTION, TERMINAL MORPHOLOGY, AND INTERACTION IN WHOLE-MOUNT ATRIA OF C57BL/6 MICE.
Creator: Harden, Scott, Cheng, Zixi (Jack), University of Central Florida
Abstract / Description: The sympathetic (SNS) and parasympathetic (PSNS) branches of the autonomic nervous system (ANS) innervate the heart, exerting excitatory and inhibitory influences (respectively) over cardiac functions (heart rate, AV conduction velocity, and contractility). However, the distribution and structure of SNS and PSNS innervation has not yet been well studied. Detailed characterization of the distributional organization and structural morphology of the SNS and PSNS in normal states is essential to...
Show moreThe sympathetic (SNS) and parasympathetic (PSNS) branches of the autonomic nervous system (ANS) innervate the heart, exerting excitatory and inhibitory influences (respectively) over cardiac functions (heart rate, AV conduction velocity, and contractility). However, the distribution and structure of SNS and PSNS innervation has not yet been well studied. Detailed characterization of the distributional organization and structural morphology of the SNS and PSNS in normal states is essential to the study of pathological autonomic remodeling. The present study utilized double immunohistochemical labeling techniques to examine tyrosine hydroxylase (TH) immunoreactive (IR) SNS and vesicular acetylcholine transporter (VAChT) IR PSNS axons and terminal structures in whole-mount atria of C57BL/6 mice. We found that: (1) The atria contain a dense network of ANS axons. TH-IR, VAChT-IR, and dual cholinergic/dopaminergic TH+VAChT-IR axons travel together in bundles on the epicardium before branching into differentiated terminal structures. (2) Parallel TH-IR and VAChT-IR axons often diverge from epicardial bundles and travel in parallel (less than 1μm apart) before forming terminal structures in the epicardium and myocardium. Such parallel SNS/PSNS axons interdigitize and have large alternating varicosities along their length adjacent to one other, suggesting possible antagonistic communication between both branches of the ANS at the prejunctional level. (3) Intrinsic cardiac ganglia (ICG) are targets for extrinsic sympathetic nerves which travel through ICG without forming large synaptic varicosities around cardiac principal neurons (PNs). (4) Small intensely fluorescent (SIF) cells (presumably chemoreceptors and/or interneurons) exist near SNS bundles, inside ICG, and in the epicardium unaccompanied by ganglia and nerve bundles. (5) The subpopulation of TH+VAChT-IR PNs within ICG form loose terminals in the atria and do not project to other PNs. (6) Both TH-IR and VAChT-IR axons innervate atrial vasculature. (7) TH-IR axons innervate fat pads adjacent to the heart. (8) SNS/PSNS parallelism is not exclusive to the atria. Similar structures exist in the esophagus, right ventricle, and small intestine. This study provides a novel and overall view of the innervation and interaction of the SNS and PSNS in the atria. This will underlie a foundation for future physiological, pharmacological, and anatomical studies of SNS/PSNS innervation, interaction, and remodeling in pathological states (such as aging, intermittent hypoxia and diabetes).
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Date Issued: 2009
Identifier: CFE0002561, ucf:47647
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0002561

Title: EFFICACY AND TOLERABILITY OF ATOMOXETINE USE FOR PATIENTS WITH AUTISM SPECTRUM DISORDERS AND ATTENTION-DEFICIT/HYPERACTIVITY DISORDER (ADHD) SYMPTOMS: A SYSTEMATIC REVIEW AND META-ANALYSIS.
Creator: El-Said, Angie, Cheng, Zixi, University of Central Florida
Abstract / Description: Introduction: Patients with autism spectrum disorders (ASD) and attention-deficit/hyperactivity disorder (ADHD) show more symptoms of ADHD. Since there are more adverse events caused by psychostimulants compared to non-psychostimulants, the use of a non-psychostimulant such as atomoxetine might prove more beneficial for younger patients and/or those with comorbid ADHD. Objective: The aim of this thesis is to determine the efficacy and tolerability of atomoxetine in ASD patients presenting...
Show moreIntroduction: Patients with autism spectrum disorders (ASD) and attention-deficit/hyperactivity disorder (ADHD) show more symptoms of ADHD. Since there are more adverse events caused by psychostimulants compared to non-psychostimulants, the use of a non-psychostimulant such as atomoxetine might prove more beneficial for younger patients and/or those with comorbid ADHD. Objective: The aim of this thesis is to determine the efficacy and tolerability of atomoxetine in ASD patients presenting with ADHD, by examining (a) differences in ADHD symptoms for participants receiving atomoxetine versus those receiving placebos, and (b) risk differences in adverse events between these participants. Methods: An electronic search of both PubMed.gov and ClinicalTrials.gov were conducted. To be deemed eligible, studies had to (a) be randomized, double-blind, placebo-controlled trials comparing atomoxetine with a placebo, (b) administer atomoxetine for at least 1 week, and (c) include data on either ADHD outcomes or adverse events. Effect sizes for ADHD outcomes were calculated using Cohen's d, whereas risk differences were calculated for adverse events. For each of these two meta-analyses, effect sizes were aggregated across studies using a random effects method. Results: Overall ADHD outcomes were better for participants who received atomoxetine than for participants who received placebo, =0.297. Participants who received atomoxetine also demonstrated better outcomes in terms of attention and hyperactivity-impulsivity symptoms, =0.345 and 0.393, respectively. Though there were more adverse events for patients taking atomoxetine than placebo, the results were not statistically significant. Discussion: This thesis extends the findings of previous meta-analyses of pharmacological treatments for ASD and ADHD, while addressing the concerns raised in the critique of existing meta-analyses presented in this thesis, e.g., limited studies, length of treatment weeks, and dichotomization of data. It provides evidence that atomoxetine improves ADHD symptoms, with an overall frequency of adverse events that did not sufficiently differ from placebo beyond chance.
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Date Issued: 2019
Identifier: CFH2000534, ucf:45644
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFH2000534

Title: INNERVATION, DISTRIBUTION AND MORPHOLOGY OF CALCITONIN GENE RELATED PEPTIDE AND SUBSTANCE P IMMUNOREACTIVE AXONS IN THE WHOLE-MOUNT ATRIA OF FVB MICE.
Creator: Li, Liang, Cheng, Zixi, University of Central Florida
Abstract / Description: Degeneration of nociceptive afferent axons and terminals in the heart is associated with painless sudden cardiac death. However, innervation, distribution and morphological structures of sympathetic cardiac nociceptive afferent axons and terminals have not yet been fully characterized. The aim of the present study is to characterize the density, arrangement, and structural features of differentiated sympathetic afferent axons and terminals in whole-mount FVB mouse atria. FVB mice (3-6 months...
Show moreDegeneration of nociceptive afferent axons and terminals in the heart is associated with painless sudden cardiac death. However, innervation, distribution and morphological structures of sympathetic cardiac nociceptive afferent axons and terminals have not yet been fully characterized. The aim of the present study is to characterize the density, arrangement, and structural features of differentiated sympathetic afferent axons and terminals in whole-mount FVB mouse atria. FVB mice (3-6 months old) were perfused and the tissues were fixed. The right and left atria were processed with immunohistochemistry. Calcitonin gene-related peptide (CGRP) and substance P (SP) are two neuropeptides which have been widely used to label sympathetic nociceptive afferent axons in many tissues. CGRP (rabbit anti-CGRP) and SP (Goat anti-SP) primary antibodies were applied, followed by Alexa Fluor 594 and 660 conjugated secondary antibodies. Whole-mount preparations of right and left atria were examined using a laser scanning confocal microscope. We found that 1) CGRP immunoreactive (IR) axon bundles innervated the right and left atria including the auricle and entrance area of the superior vena cava, the inferior vena cava, left precaval vein and pulmonary veins. Large axon bundles entered the area from the major veins and bifurcated into smaller axon bundles and single axon fibers to form terminal end-nets and free endings in the epicardium at each region with a similar pattern. In the atrial muscle layer, varicose CGRP-IR axons had close contacts with muscle fibers. In addition, CGRP-IR axons terminated in the intrinsic cardiac ganglia (ICGs) with varicosities surrounding individual ganglionic principle neurons (PNs). In the aortic arch, the CGRP-IR fibers exhibited similar terminal structures to those seen in the atria. 2) SP-IR axons also projected to the right and left atria and aorta. Similar to CGRP-IR axons, these SP-IR axons also formed end-nets and free endings in these areas. In cardiac ganglia, SP-IR axons formed varicose endings around many individual PNs. However, a salient difference was found: There appeared to be fewer SP-IR axons and terminals than CGRP-IR axons and terminals in the atria. 3) None of the cardiac PNs in ICG were CGRP-IR or SP-IR. 4) Many SP-IR axon terminals around PNs within ICGs and atrial muscles were found to have colocalized expression of CGRP-IR. Collectively, our data for the first time documented the distribution patterns and morphology of sympathetic afferent axons and terminals in each region of the atria in the mouse model. This will provide a foundation for future analysis of the pathological changes of sympathetic afferent nerves in the atria in different disease models (e.g., diabetes, sleep apnea, and aging). This study was supported by NIH R01 HL-79636.
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Date Issued: 2010
Identifier: CFE0003536, ucf:48965
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0003536

Title: AMELIORATION OF AMYLOID BURDEN IN ADVANCED HUMAN AND MOUSE ALZHEIMER'S DISEASE BRAINS BY ORAL DELIVERY OF MYELIN BASIC PROTEIN BIOENCAPSULATED IN PLANT CELLS.
Creator: Kohli, Neha, Daniell, Henry, Kim, Yoon-Seong, Cheng, Zixi, University of Central Florida
Abstract / Description: One of the pathological hallmarks of Alzheimer's disease (AD) is the amyloid plaque deposition in aging brains by aggregation of amyloid-? (A?) peptides. In this study, the effect of chloroplast derived myelin basic protein (MBP) fused with cholera toxin subunit B (CTB) was investigated in advanced diseased stage of human and mouse AD brains. The CTB-fusion protein in chloroplasts facilitates transmucosal delivery in the gut by the natural binding ability of CTB pentameric form with GM1...
Show moreOne of the pathological hallmarks of Alzheimer's disease (AD) is the amyloid plaque deposition in aging brains by aggregation of amyloid-? (A?) peptides. In this study, the effect of chloroplast derived myelin basic protein (MBP) fused with cholera toxin subunit B (CTB) was investigated in advanced diseased stage of human and mouse AD brains. The CTB-fusion protein in chloroplasts facilitates transmucosal delivery in the gut by the natural binding ability of CTB pentameric form with GM1 receptors on the intestinal epithelium. Further, bioencapsulation of the MBP within plant cells confers protection from enzymes and acids in the digestive system. Here, 12-14 months old triple transgenic AD mice were fed with CTB-MBP bioencapsulated in the plant cells for 3 months. A reduction of 67.3% and 33.3% amyloid levels in hippocampal and cortical regions, respectively were observed by immunostaining of brain sections with anti- A? antibody. Similarly, 70% decrease in plaque number and 40% reduction of plaque intensity was observed through thioflavin S (ThS) staining that specifically stains amyloid in the AD brain. Furthermore, ex vivo 3xTg AD mice brain sections showed up to 45% reduction of ThS stained amyloid levels when incubated with enriched CTB-MBP in a concentration dependent manner. Similarly, incubation of enriched CTB-MBP with ex vivo postmortem human brain tissue sections with advanced stage of AD resulted up to 47% decrease of ThS stained amyloid plaque intensity. Lastly, lyophilization of plant material facilitates dehydration and long term storage of capsules at room temperature, in addition to increasing CTB-MBP concentration by 17 fold. These observations offer a low cost solution for treatment of even advanced stages of the AD by facilitating delivery of therapeutic proteins to central nervous system to address other neurodegenerative disease.
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Date Issued: 2012
Identifier: CFE0004564, ucf:49237
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0004564

Title: Dissertation Title: Development of molecular and cellular imaging tools to evaluate gene and cell based therapeutic strategies in vivo.
Creator: Xia, Jixiang, Ebert, Steven, Khaled, Annette, Cheng, Zixi, Daniell, Henry, University of Central Florida
Abstract / Description: Molecular imaging modalities are important tools to evaluate the efficacy of gene delivery systems and cell-based therapies. Development and application of these modalities will advance our understanding of the mechanism of transgene expression and cell fate and functions. Physical gene transfer methods hold many advantages over viral vectors among gene therapeutic strategies. Here, we evaluated the efficacy of biolistic ((")gene gun(")) gene targeting to tissues with non-invasive...
Show moreMolecular imaging modalities are important tools to evaluate the efficacy of gene delivery systems and cell-based therapies. Development and application of these modalities will advance our understanding of the mechanism of transgene expression and cell fate and functions. Physical gene transfer methods hold many advantages over viral vectors among gene therapeutic strategies. Here, we evaluated the efficacy of biolistic ((")gene gun(")) gene targeting to tissues with non-invasive bioluminescence imaging (BLI) methods. Plasmids carrying the firefly luciferase reporter gene were transfected into mouse skin and liver using biolistics, and BLI was measured at various time points after transfer. With optimized DNA loading ratio (DLRs), reporter gene expression reached to peak 1day after transfer to mouse skin, and the maximum depth of tissue penetration was between 200-300?m. Similar peak expression of reporter gene was found in mouse liver but the expression was relatively stable 4-8 days post-biolistic gene transfer and remained for up to two weeks afterward. Our results demonstrated BLI was an efficient strategy for evaluation of reporter gene expression in the same animals over a period of up to two weeks in vivo. Different tissues showed different expression kinetics, suggesting that this is an important parameter to consider when developing gene therapy strategies for different target tissues. We also employed BLI to measure differentiation of mouse embryonic stem (ES) cells into beating cardiomyocytes in vitro and in vivo. A subset of these cardiomyocytes appears to be derived from an adrenergic lineage that ultimately contribute to substantial numbers of cardiomyocytes primarily on the left side of the heart. At present, it is unclear what the precise role of these cardiac adrenergic cells is with respect to heart development, though it is known that adrenergic hormones (adrenaline and noradrenaline) are essential for embryonic development since mice lacking them die from apparent heart failure during the prenatal period. To identify and characterize cardiac adrenergic cells, we developed a novel mouse genetic model in which the nuclear-localized enhanced green fluorescent protein (nEGFP) reporter gene was targeted to the first exon of the Phenylethanoamine N-transferase (Pnmt) gene, which encodes for the enzyme that converts noradrenaline to adrenaline, and hence serves as a marker for adrenergic cells. Our results demonstrate this knock-in strategy effectively marked adrenergic cells in both fetal and adult mice. Expression of nEGFP was found in Pnmt-positive cells of the adult adrenal medulla, as expected. Pnmt-nEGFP expression also recapitulated endogenous Pnmt expression in the embryonic mouse heart. In addition, nEGFP and Pnmt expression were induced in parallel during differentiation of pluripotent mouse ES cells into beating cardiomyocytes. This new mouse genetic model provides a useful new tool for studying the properties of adrenergic cells in different tissues. We also identified two limitations of the Pnmt-nEGFP model. One is that the amount of nEGFP expressed within individual adrenergic cells was highly variable. Secondly, expression of nEGFP in the embryonic heart was of low abundance and difficult to distinguish from background autofluorescence. To overcome these limitations, we developed two alternative genetic models to investigate adrenergic cells: (1) Mouse embryonic stem cells, which have been previously targeted with Pnmt-Cre recombinase gene, were additionally targeted with a dual reporter plasmid which covered both a loxP-flanked cDNA of red fluorescence protein (HcRed) and also EGFP. Under the undifferentiated status, cells emit red fluorescence as transcription stops before EGFP coding sequence. After differentiation into beating cardiomyoctyes, some cells switch fluorescence from red to green, indicating that excision of loxP-flanked sequences by Cre since Pnmt had been activated. (2) A surface marker, truncated low-affinity nerve growth factor receptor (?LNGFR) was used as the reporter gene as cells expressing this marker can be enriched by magnetic-activated cell sorting (MACS), a potentially efficient way to yield highly purified positive cells at low input abundance in a population. Through a series of subcloning steps, the targeting construct, Pnmt-?LNGFR-Neo-DTA was created and electroporated into 7AC5EYFP embryonic stem cells. Correctly targeted cells were selected by positive and negative screening. These cells provide a new tool with which to identify, isolate, and characterize the function of adrenergic cells in the developing heart, adrenal gland, and other tissues where adrenergic cells make important contributions.
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Date Issued: 2011
Identifier: CFE0004491, ucf:49287
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0004491

Title: Evaluation of Intestinal Microbial Diversity and a New Antibiotic Regimen in Crohn's Disease Patients.
Creator: Alcedo, Karel, Naser, Saleh, Cheng, Zixi, Siddiqi, Shadab, University of Central Florida
Abstract / Description: Crohn's disease (CD) is a chronic granulomatous inflammatory bowel disease involving Mycobacterium avium subspecies paratuberculosis (MAP). Other microorganisms such as adherent-invasive Escherichia coli (AIEC) have also been proposed in CD association. To date, only one study investigated both MAP and AIEC simultaneously using peripheral blood but not in affected intestinal tissues. A standardized and effective antibiotic therapy against MAP and/or AIEC is needed for better treatment. Three...
Show moreCrohn's disease (CD) is a chronic granulomatous inflammatory bowel disease involving Mycobacterium avium subspecies paratuberculosis (MAP). Other microorganisms such as adherent-invasive Escherichia coli (AIEC) have also been proposed in CD association. To date, only one study investigated both MAP and AIEC simultaneously using peripheral blood but not in affected intestinal tissues. A standardized and effective antibiotic therapy against MAP and/or AIEC is needed for better treatment. Three antibiotic drugs (-) Clarithromycin (CLA), Rifabutin (RIF), and Clofazimine (CLO) have been used to treat CD patients suspected with MAP infection. However, the outcome has been controversial. The treatment dosage is high, the duration is long, and the reported drug side effects resulted in patient non-compliance; therefore, a lower and effective drug dosage is needed. In this study, we developed two aims 1) to evaluate RHB 104, a drug formula comprised of low dosages of CLA, RIF, and CLO, against clinical MAP strains in-vitro using fluorescence quenching method, and 2) to develop a fluorescence in-situ hybridization method to detect both MAP and AIEC simultaneously in intestinal tissues of CD patients. A total of 16 clinical MAP strains and 19 non-MAP strains were tested against varied concentrations of RHB 104, CLA, RIF, and CLO. Although the MIC for all drugs ranged between 0.5-20 ?g/ml, the MIC for RHB 104 was significantly lower against most MAP strains. The effect of RHB 104 against MAP was bactericidal. Unlike RHB-104 formula, CLA, CLO, and RIF dosage similar to those in RHB-104 did not inhibit MAP growth when trialed individually and in dual-drug combinations. The data illustrated the presence of synergistic anti-MAP activity of low dosage of the three antibiotics in RHB-104. We also developed a rapid and sensitive multicolor in-situ hybridization technique that can detect MAP and AIEC using tagged-oligonucleotide probes. Non-pathogenic Escherichia coli (npEC) was used as a control for the study. Specifically, cultured MAP and npEC were fixed and hybridized with MAP488 and EC647 probes, respectively. Confocal laser scanning microscope (CLSM) revealed specific signals at 488nm for MAP and 647nm for npEC, indicating probe binding to each bacteria. This was confirmed with hybridization of MAP with EC647 and npEC with MAP488 resulting in absence of signals. Intestinal tissue samples from 9 CD patients were then analyzed using our technique. Preliminary data indicated positive results in 6/6 samples for MAP, 6/6 for npEC, 3/3 for AIEC, and 2/2 for both MAP and AIEC with MAP being more dominant. This protocol shortened the FISH procedure from multiple days to short-hours. The protocol allows the investigation of more than one pathogen simultaneously in the same clinical sample. A quantitative measurement of the signals is needed.
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Date Issued: 2015
Identifier: CFE0005917, ucf:50831
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005917

Title: BMP-7 Inhibits p38 and JNK Pathways and Increases M2 Macrophage Differentiation to Reduce Atherosclerosis in Apolipoprotein E-/- Mice.
Creator: Shoulders, Heidi, Singla, Dinender, Cheng, Zixi, Naser, Saleh, University of Central Florida
Abstract / Description: We have previously shown that treating atherosclerosis with bone morphogenetic protein-7 (BMP-7) affects the presence of macrophage subtypes in vitro, however it remains unknown whether BMP-7 treatment affects development and progression of atherosclerosis in vivo at an early and mid-stage of the disease. We therefore performed a Day 5 (D5) and Day 28 (D28) study to examine BMP-7's potential to affect monocyte differentiation. Atherosclerotic plaque formation was developed using our standard...
Show moreWe have previously shown that treating atherosclerosis with bone morphogenetic protein-7 (BMP-7) affects the presence of macrophage subtypes in vitro, however it remains unknown whether BMP-7 treatment affects development and progression of atherosclerosis in vivo at an early and mid-stage of the disease. We therefore performed a Day 5 (D5) and Day 28 (D28) study to examine BMP-7's potential to affect monocyte differentiation. Atherosclerotic plaque formation was developed using our standard method and ApoE-/- mice were sacrificed at D5 and D28 post-surgery. Treatment animals received intravenous injections of BMP-7 at 200(&)#181;g/kg of bodyweight. Hematoxylin and Eosin morphological stain shows that BMP-7 is capable of significantly reducing plaque accumulation at D28 post-surgery vs. PLCA group, p(<)0.05. At D5, plaque formation was reduced but not significant. Immunohistochemistry staining was performed to determine BMP-7's effect on monocytes (CD14), inflammatory M1 (iNOS) and anti-inflammatory M2 (CD206, Arginase-1) macrophages. Immunohistochemistry results show BMP-7 administration reduced pro-inflammatory monocytes and M1 macrophages at D5 and D28 compared to PLCA animals; however, monocytes were not statistically lower at D28. The anti-inflammatory M2 macrophage population was significantly less in PLCA animals compared to SHAM animals at D5 and D28. There was no significant difference in M2 macrophages between PLCA and PLCA + BMP7 animals at D5, however, by D28, PLCA + BMP7 animals showed a significant increase in M2 macrophages compared to PLCA animals. Western blot analysis confirms a significant increase in pro-survival kinase ERK and a significant reduction in pro-inflammatory kinases p38 and JNK in BMP-7 treated mice (D5 and D28, p(<)0.05). ELISA showed a significant reduction in pro-inflammatory cytokines IL-6, MCP-1, and TNF-? (D5 and D28, p(<)0.05) and a significant increase in anti-inflammatory cytokine IL-10 in BMP-7 treated mice (D5 and D28, p(<)0.05). In summary, our data indicate BMP-7 treatment induces monocyte to M2 macrophage differentiation, increases anti-inflammatory cytokine levels (IL-1ra and IL-10), and improves blow flow velocity (D5 and D28, p(<)0.05) compared to untreated animals. The mechanisms of monocyte to M2 macrophage differentiation appear to be mediated by the p38, JNK, and ERK pathways. This study suggests BMP-7 is capable of reducing inflammation and slowing progression of atherosclerosis at both an early and mid-stage of the disease.
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Date Issued: 2016
Identifier: CFE0006504, ucf:51388
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0006504

Title: Plasticity of Central and Peripheral Nervous System: Effects of Oxygen-Glucose Deprivation (OGD), Chronic Intermittent Hypoxia (CIH) and hSOD1 Overexpression.
Creator: Chen, Jin, Cheng, Zixi, Naser, Saleh, Singla, Dinender, University of Central Florida
Abstract / Description: Transient receptor potential canonical 6 (TRPC6) channels are permeable to Na+ and Ca2+ and are widely expressed in the brain. In this study, we investigated the role of TRPC6 following ischemia/reperfusion (I/R) and oxygen-glucose deprivation (OGD). We found that TRPC6 expression was increased in wild type (WT) mice cortical neurons following I/R and in primary neurons with OGD, and that deletion of TRPC6 reduced the I/R-induced brain infarct in mice and the OGD- /neurotoxin-induced neuronal...
Show moreTransient receptor potential canonical 6 (TRPC6) channels are permeable to Na+ and Ca2+ and are widely expressed in the brain. In this study, we investigated the role of TRPC6 following ischemia/reperfusion (I/R) and oxygen-glucose deprivation (OGD). We found that TRPC6 expression was increased in wild type (WT) mice cortical neurons following I/R and in primary neurons with OGD, and that deletion of TRPC6 reduced the I/R-induced brain infarct in mice and the OGD- /neurotoxin-induced neuronal death. Using live-cell imaging to examine intracellular Ca2+ levels ([Ca2+]i), we found that OGD induced a significant higher increase in glutamate-evoked Ca2+ influx compared to untreated control and such an increase was reduced by TRPC6 deletion. Enhancement of TRPC6 expression using AdCMV-TRPC6-GFP infection in WT neurons increased [Ca2+]i in response to glutamate application compared to AdCMV-GFP control. Inhibition of N-methyl-d-aspartic acid receptor (NMDAR) with MK801 decreased TRPC6-dependent increase of [Ca2+]i, indicating that such a Ca2+ influx was NMDAR dependent. Furthermore, TRPC6-dependent Ca2+ influx was blunted by blockade of Na+ entry. Finally, OGD-enhanced Ca2+ influx was reduced, but not completely blocked, in the presence of voltage dependent Na+ channel blocker tetrodotoxin (TTX) and dl???amino?3?hydroxy?5?methyl?4?isoxazole propionic acid (AMPA) blocker CNQX. Altogether, we concluded that I/R-induced brain damage was, in part, due to upregulation of TRPC6 in cortical neurons. We postulate that overexpression of TRPC6 following I/R may induce neuronal death partially through TRPC6-dependent Na+ entry which activated NMDAR, thus leading to a damaging Ca2+ overload. These findings may provide a potential target for future intervention in stroke-induced brain damage. Obstructive sleep apnea (OSA) is a highly prevalent sleep disorder that is associated with many cardiovascular complications, such as autonomic dysfunctions, stroke and heart failure. Chronic intermittent hypoxia (CIH) is a prominent feature of OSA. In CIH exposed rodents (a model for OSA), CIH induces the similar cardiovascular complications as seen in OSA patients. In particular, OSA impairs baroreflex control of the heart rate (HR), which is used as an independent indicator for heart failure. Since the baroreflex control arc includes the aortic depressor nerve (ADN), vagal efferent and central components, we hypothesize that CIH induces dysfunctions of all three components. Since mice can be genetically manipulated, an understanding of the effects of CIH on multiple neural components in the baroreflex arc in wild type mice may lead to a future study of treatments. In this study, we have examined the effects of CIH on baroreceptor afferent, central and vagal efferent components of the baroreflex circuitry in normal wild type C57BL/6J mice. Mice (4-5 months) were exposed to room air (RA) or CIH for 35-50 days and were then anesthetized with isoflurane, ventilated and catheterized for measurement of mean arterial blood pressure (MAP) and HR. Baroreceptor function was characterized by measuring percent changes of integrated ADN activity (Int ADNA) relative to the baseline value in response to the vasodilator sodium nitroprusside and the vasoconstrictor phenylephrine-induced changes in MAP. Data were fitted to a sigmoid logistic function curve. HR responses to electrical stimulation of the left ADN and the right vagus nerve were assessed under anesthesia. Compared with RA controls, CIH significantly increased maximum baroreceptor gain or maximum slope, maximum Int ADNA, and Int ADNA range (maximum-minimum Int ADNA). In addition, CIH maintained the maximum amplitude of the bradycardic response to vagal efferent stimulation. In contrast, CIH significantly reduced the maximum amplitude of bradycardic response to left ADN stimulation. Thus, CIH decreased central mediation of the baroreflex, but augmented the baroreceptor afferent function and maintained vagal efferent control of HR in mice. Excessive reactive oxygen species (ROS) (such as the superoxide radical) is commonly associated with cardiac autonomic dysfunctions. Though superoxide dismutase 1 (SOD1) overexpression may protect against ROS damage to the autonomic nervous system, superoxide radical reduction may change normal physiological functions. Previously, we demonstrated that human SOD1 (hSOD1) overexpression did not change baroreflex bradycardia and tachycardia, but increased aortic depressor nerve activity (ADNA) in responses to arterial pressure changes in C57B6SJL-Tg (SOD1)2 Gur/J mice. Since the barorelfex arc includes afferent, central and efferent components, the objective of this study was to determine whether hSOD1 overexpression alters the central and vagal efferent mediation of the heart rate (HR) responses. Our data indicate that SOD1 overexpression decreased HR responses to vagal efferent nerve stimulations but did not change HR responses to aortic nerve stimulation. Along with the previous study, we suggest that SOD1 overexpression preserves the normal baroreflex function but may alter the functions of aortic depressor nerve, vagal efferent and central components differently. While SOD1 overexpression likely enhanced aortic depressor nerve function and central mediation of bradycardia, it decreased vagal efferent control of HR. Currently, we are using the hSOD1 overexpressing mouse model to determine whether hSOD1 overexpression can preserve normal afferent, efferent, and central components of the baroreflex arc in the CIH model of sleep apnea.
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Date Issued: 2017
Identifier: CFE0006576, ucf:51334
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0006576

Title: TIMP-1 ACTIVATES A UNIQUE CARDIAC STEM CELL POPULATION, CD63+ve/C-KIT+ve, THEREBY ENHANCING CARDIAC DIFFERENTIATION, AND PROTECTS THE HEART FROM ADVERSE CARDIAC REMODELING FOLLOWING MYOCARDIAL INFARCTION.
Creator: Abdelli, Latifa, Singla, Dinender, Cheng, Zixi, Parthasarathy, Sampath, Jewett, Mollie, University of Central Florida
Abstract / Description: We previously demonstrated that embryonic stem (ES) cells over-expressing tissue inhibitor of metalloproteinase-1 (TIMP-1) have increased potential to engraft and differentiate into cardiac myocytes following transplantation into the infarcted heart. However, the ability of TIMP-1 to activate endogenous stem cells and enhance their differentiation into cardiac regenerative cell types is still unknown. We postulate that TIMP-1 may additionally activate a stem cell population that enhances...
Show moreWe previously demonstrated that embryonic stem (ES) cells over-expressing tissue inhibitor of metalloproteinase-1 (TIMP-1) have increased potential to engraft and differentiate into cardiac myocytes following transplantation into the infarcted heart. However, the ability of TIMP-1 to activate endogenous stem cells and enhance their differentiation into cardiac regenerative cell types is still unknown. We postulate that TIMP-1 may additionally activate a stem cell population that enhances cardiac cell type differentiation in the infarcted myocardium. To prove this hypothesis, we isolated c-kit+ve cells from four weeks old C57BL/6 mice and cultured them in vitro in presence of ES conditioned media (ESCM), ES-TIMP-1-CM or TIMP-1. Our immunostaining data validate the existence of a novel CD63+ve/c-kit+ve cells. When treated with TIMP-1, these cells showed significantly (p(<)0.05) increased proliferation and differentiation into cardiac myocytes, vascular smooth muscle cells, and endothelial cells. Western blot analysis revealed significantly (p(<)0.05) increased expression of CD63, phosphorylated and total ?-catenin proteins. Furthermore, our RT-PCR data showed increased cardiac gene expression (GATA-4, Mef2C, and Nkx-2.5) when compared to ESCM and control cells. Based on the in vitro findings, we investigated the effect of intramyocardial delivery of TIMP-1 on endogenous CD63+ve/c-kit+ve cells following myocardial infarction (MI). C57BL/6 and TIMP-1 KO mice underwent coronary artery ligation followed by intramyocardial delivery of 20(&)#181;l of culture media (CC), ESCM, ES-TIMP-1-CM or TIMP-1. Subsequent immunohistochemistry analysis demonstrated the presence of a CD63+ve/c-kit+ve cell population within the peri-infarct area and confirmed intramyocardial delivery of ES-TIMP-1-CM or TIMP-1 significantly (p(<)0.05) enhanced their proliferation. Percentage of CD63+ve/c-kit+ve cells was significantly (p(<)0.05) lower in TIMP-1 KO mice compared to C57BL/6 animals. RT-PCR analysis revealed TIMP-1 KO animals expressed significantly less CD63 and TIMP-1 mRNAs compared to C57BL/6 mice. Activated CD63+ve/c-kit+ve cells were also able to differentiate into major cardiac cell types as previously shown in vitro. The differentiation potential of these cells was however higher in C57BL/6 mice compared to TIMP-1 KO mice. We also demonstrate that CD63+ve/c-kit+ve cells differentiation is regulated by CD63/?-catenin pathway in vivo. Additionally, we provide evidence that TIMP-1 protects the heart from adverse cardiac remodeling through inhibition of cardiac apoptosis and fibrosis leading to significantly (p(<)0.05) improved contractile function. Collectively, our data show TIMP-1 plays a dual protective role in the MI heart. It activates a unique stem cell population, CD63+ve/c-kit+ve, which proliferates and differentiates into functional myocytes, smooth muscle cells and endothelial cells mediated through CD63/?-catenin pathway. TIMP-1 also protects the heart from adverse cardiac remodeling. Increased cardiac regeneration and inhibition of adverse cardiac remodeling consequently lead to restored cardiac function. ?
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Date Issued: 2015
Identifier: CFE0005750, ucf:50108
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005750

Title: Defective Dynamics of Mitochondria in Amyotrophic Lateral Sclerosis and Huntington's Disease.
Creator: Song, Wenjun, Bossy-Wetzel, Ella, Fernandez-Valle, Cristina, Cheng, Zixi, Self, William, University of Central Florida
Abstract / Description: Mitochondria play important roles in neuronal function and survival, including ATP production, Ca2+ buffering, and apoptosis. Mitochondrial dysfunction is a common event in the pathogenesis of neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and Huntington's disease (HD); however, what causes the mitochondrial dysfunction remains unclear. Mitochondrial fission is mediated by dynamin-related protein 1 (DRP1) and fusion by mitofusin 1/2 (MFN1/2) and optic atrophy 1 (OPA1),...
Show moreMitochondria play important roles in neuronal function and survival, including ATP production, Ca2+ buffering, and apoptosis. Mitochondrial dysfunction is a common event in the pathogenesis of neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS) and Huntington's disease (HD); however, what causes the mitochondrial dysfunction remains unclear. Mitochondrial fission is mediated by dynamin-related protein 1 (DRP1) and fusion by mitofusin 1/2 (MFN1/2) and optic atrophy 1 (OPA1), which are essential for mitochondrial function. Mutations in the mitochondrial fission and fusion machinery lead to neurodegeneration. Thus, whether defective mitochondrial dynamics participates in ALS and HD requires further investigation.ALS is a fatal neurodegenerative disease characterized by upper and lower motor neuron loss. Mutations in Cu/Zn superoxide dismutase (SOD1) cause the most common familiar form of ALS by mechanisms not fully understood. Here, a new motor neuron-astrocyte co-culture system was created and live-cell imaging was used to evaluate mitochondrial dynamics. Excessive mitochondrial fission was observed in mutant SOD1G93A motor neurons, correlating with impaired axonal transport and neuronal cell death. Inhibition of mitochondrial fission restored mitochondrial dynamics and protected neurons against SOD1G93A-induced mitochondrial fragmentation and neuronal cell death, implicating defects in mitochondrial dynamics in ALS pathogenesis.HD is an inherited neurodegenerative disorder caused by glutamine (Q) expansion in the polyQ region of the huntingtin (HTT) protein. In the current work, mutant HTT caused mitochondrial fragmentation in a polyQ-dependent manner in both primary cortical neurons and fibroblasts from human patients. An abnormal interaction between DRP1 and HTT was observed in mutant HTT mice and inhibition of mitochondrial fission or promotion of mitochondrial fusion restored mitochondrial dynamics and protected neurons against mutant HTT-induced cell death. Thus, mutant HTT may increase mitochondrial fission by elevating DRP1 GTPase activity, suggesting that mitochondrial dynamics plays a causal role in HD.In summary, rebalanced mitochondrial fission and fusion rescues neuronal cell death in ALS and HD, suggesting that mitochondrial dynamics could be the molecular mechanism underlying these diseases. Furthermore, DRP1 might be a therapeutic target to delay or prevent neurodegeneration.
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Date Issued: 2012
Identifier: CFE0004444, ucf:49356
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0004444

Title: Synthesis of Fluorescent Molecules and their Applications as Viscosity Sensors, Metal Ion Indicators, and Near-Infrared Probes.
Creator: Wang, Mengyuan, Belfield, Kevin, Campiglia, Andres, Miles, Delbert, Frazer, Andrew, Cheng, Zixi, University of Central Florida
Abstract / Description: The primary focus of this dissertation is the development of novel fluorescent near-infrared molecules for various applications. In Chapter 1, a compound dU-BZ synthesized via Sonogashira coupling reaction methodology is described. A deoxyuridine building block was introduced to enhance hydrophilic properties and reduce toxicity, while an alkynylated benzothiazolium dye was incorporated for near-IR emission and reduce photodamage and phototoxicity that is characteristic of common fluorphores...
Show moreThe primary focus of this dissertation is the development of novel fluorescent near-infrared molecules for various applications. In Chapter 1, a compound dU-BZ synthesized via Sonogashira coupling reaction methodology is described. A deoxyuridine building block was introduced to enhance hydrophilic properties and reduce toxicity, while an alkynylated benzothiazolium dye was incorporated for near-IR emission and reduce photodamage and phototoxicity that is characteristic of common fluorphores that are excited by UV or visible light. A 30-fold enhancement of fluorescence intensity of dU-BZ was achieved in a viscous environment. Values of fluorescence quantum yields in 99% glycerol/1% methanol (v/v) of varying temperature from 293 K to 343 K, together with fluorescence quantum yields, radiative and nonradiative rate constants and fluorescence lifetimes in glycerol/methanol solutions of varying viscosities from 4.8 to 950 cP were determined. It was found that both fluorescence quantum yields and fluorescence lifetimes increased with increasing viscosity, which is consistent with results predicted by theory. This suggests that the newly designed compound dU-BZ is capable of functioning as a probe of local microviscosity, and was later confirmed by in vitro bioimaging experiments.In Chapter 2, a new BAPTA (O,O'-bis(2-aminophenyl)ethyleneglycol-N,N,N',N'-tetra acetic acid) and BODIPY (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene)-based calcium indicator, BAPBO-3, is reported. A new synthetic route was employed to simplify both synthesis and purification, which tend to be low yielding and cumbersome for BAPTA derivatives. Upon excitation, a 1.5-fold increase in fluorescence intensity in buffer containing 39 ?? Ca2+ and a 3-fold increase in fluorescence intensity in buffer containing 1 M Ca2+ was observed; modest but promising fluorescence turn-on enhancements.In Chapter 3, a newly-designed unsymmetrical squaraine dye, SQ3, was synthesized. A one-pot synthesis was employed resulting in a 10% yield, a result that is generally quite favorable for the creation of unsymmetrical squaraines Photophysical and photochemical characterization was conducted in various solvents, and a 678 nm absorption maximum and a 692 nm emission maximum were recorded in DMSO solution with a fluorescence quantum yield of 0.32. In vitro cell studies demonstrated that SQ3 can be used as a near-IR probe for bioimaging.
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Date Issued: 2014
Identifier: CFE0005900, ucf:50863
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005900

Title: Investigating New Guaiazulenes and Diketopyrropyrroles for Photonic Applications.
Creator: Ghazvini Zadeh, Ebrahim, Belfield, Kevin, Campiglia, Andres, Yuan, Yu, Zou, Shengli, Cheng, Zixi, University of Central Florida
Abstract / Description: ?-Conjugated systems have been the focus of study in recent years in order to understand their charge transport and optical properties for use in organic electronic devices, fluorescence bioimaging, sensors, and 3D optical data storage (ODS), among others. As a result, several molecular building blocks have been designed, allowing new frontiers to be realized. While various successful building blocks have been fine-tuned at both the electronic and molecular structure level to provide advanced...
Show more?-Conjugated systems have been the focus of study in recent years in order to understand their charge transport and optical properties for use in organic electronic devices, fluorescence bioimaging, sensors, and 3D optical data storage (ODS), among others. As a result, several molecular building blocks have been designed, allowing new frontiers to be realized. While various successful building blocks have been fine-tuned at both the electronic and molecular structure level to provide advanced photophysical and optoelectronic characteristics, the azulene framework has been under-appreciated despite its unique electronic and optical properties. Among several attributes, azulenes are vibrant blue naturally occurring hydrocarbons that exhibit large dipolar character, coupled with stimuli-responsive behavior in acidic environments. Additionally, the non-toxic nature and the accompanying eco-friendly feature of some azulenes, namely guaiazulene, may set the stage to further explore a more (")green(") route towards photonic and conductive materials.The first part of this dissertation focuses on exploiting guaiazulene as a natural building block for the synthesis of chromophores with varying stimuli-responsiveness. Results described in Chapter 1 show that extending the conjugation of guaiazulene through its seven-membered ring methyl group with aromatic substituents dramatically impacts the optical properties of the guaiazulenium carbocation. Study of these ?(-)stabilized tropilium ions enabled establishing photophysical structure-property trends for guaiazulene-terminated ?-conjugated analogs under acidic conditions, including absorption, emission, quantum yield, and optical band gap patterns. These results were exploited in the design of a photosensitive polymeric system with potential application in the field of three dimensional (3D) optical data storage (ODS).Chapter 2 describes the use of guaiazulene reactive sites (C-3 and C-4 methyl group) to generate a series of cyclopenta[ef]heptalenes that exhibit strong stimuli-responsive behavior. The approach presents a versatile route that allows for various substrates to be incorporated into the resulting cyclopenta[ef]heptalenes, especially after optimization that led to devising a one-pot reaction toward such tricyclic systems. Examining the UV-vis absorption profiles in neutral and acidic media showed that the extension of conjugation at C(4) of the cyclopenta[ef]heptalene skeleton results in longer absorption maxima and smaller optical energy gaps. Additionally, it was concluded that these systems act as sensitizers of a UV-activated ((<) 300 nm) photoacid generator (PAG), via intermolecular photoinduced electron transfer (PeT), upon which the PAG undergoes photodecomposition resulting in the generation of acid.In a related study, the guaiazulene methyl group at C-4 was employed to study the linear and nonlinear optical properties of 4-styrylguaiazulenes, having the same ?(-)donor with varying ?-spacer. It was realized that the conjugation length correlates with the extent of bathochromic shift of the protonated species. On the other hand, a trend of decreasing quantum yield was established for this set of 4-styrylguaiazulenes, which can be explained by the increasingly higher degree of flexibility.The second part of this dissertation presents a comprehensive investigation of the linear photophysical, photochemical, and nonlinear optical properties of diketopyrrolopyrrole (DPP)-based derivatives, including two-photon absorption (2PA), femtosecond transient absorption, stimulated emission spectroscopy, and superfluorescence phenomena. The synthetic feasibility, ease of modification, outstanding robustness, and attractive spectroscopic properties of DPPs have motivated their study for fluorescence microscopy applications, concluding that the prepared DPP's are potentially suitable chromophores for high resolution stimulated emission depletion (STED) microscopy.
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Date Issued: 2015
Identifier: CFE0006034, ucf:50986
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0006034

Title: Synthesis and Characterization of New Probes for use in Fluorescence and X-ray CT Bioimaging.
Creator: Tang, Simon, Belfield, Kevin, Miles, Delbert, Campiglia, Andres, Zou, Shengli, Cheng, Zixi, University of Central Florida
Abstract / Description: The pursuit of more suitable drugs intended for possible biological applications are a continuously growing topic of research within the scientific community. One of these suitable qualities includes the need for hydrophilicity and or some appropriate delivery system for the drug to enter into biological systems. A system of analyzing and following these compounds would then, however, be necessary to conduct any kind of mechanistic or interaction studies for he said drug within the biological...
Show moreThe pursuit of more suitable drugs intended for possible biological applications are a continuously growing topic of research within the scientific community. One of these suitable qualities includes the need for hydrophilicity and or some appropriate delivery system for the drug to enter into biological systems. A system of analyzing and following these compounds would then, however, be necessary to conduct any kind of mechanistic or interaction studies for he said drug within the biological system. Just to name a few, fluorescence and X-ray computed tomography (CT) methods allow for imaging of biological systems but require the need of compounds with specific qualities. Finally, even with a means of entering and following a oaded drug, it would not be complete without a way of targeting its intended location. Herein, the first chapter reports the synthesis and characterization of a fluorene-based pyridil bis-?-diketone compound with suitable one- and two-photon fluorescent properties and its encapsulation into Pluronic F127 micelles for the possible application of tracking lysosomes. Next the synthesis and characterization of a BODIPY-based fluorophore with excellent fluorescence ability is reported. This compound was conjugated to two triphenylphosphine (TPP) groups and is shown as a potential mitochondria probe within HCT-116 cells. Finally, the synthesis and characterization of diatrizoic acid (DA) based derivatives conjugated to silica nanoparticles, as well as unconjugated, are reported as potential CT contrast agents. The derivatives were also functionalized with maleimide moieties facilitating subsequent potential bioconjugation of a targeting protein via a thiol group.
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Date Issued: 2015
Identifier: CFE0006056, ucf:50961
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0006056

Title: In Actu Et In Silicio: Linear and Nonlinear Photophysical Characterization of a Novel Europium Complex, and Incorporating Computational Calculations in the Analysis of Novel Organic Compounds.
Creator: Woodward, Adam, Belfield, Kevin, Campiglia, Andres, Harper, James, Frazer, Andrew, Cheng, Zixi, University of Central Florida
Abstract / Description: Despite not being a tangible substance, light is becoming an increasingly valuable tool in numerous areas of science and technology: the use of laser excitation of a fluorescent probe can generate incredibly detailed images of cellular structures without the need for large amounts of dissection; new types of solar cells are being produced using organic dyes to harvest light; computer data can be stored by inducing a chemical change in a compound through irradiation with light. However, before...
Show moreDespite not being a tangible substance, light is becoming an increasingly valuable tool in numerous areas of science and technology: the use of laser excitation of a fluorescent probe can generate incredibly detailed images of cellular structures without the need for large amounts of dissection; new types of solar cells are being produced using organic dyes to harvest light; computer data can be stored by inducing a chemical change in a compound through irradiation with light. However, before any of these materials can be applied in such a way, their properties must first be analyzed for them to be deemed viable.The focus of this dissertation is the photophysical characterization, linear and nonlinear, of a several novel organic compounds, and a europium complex, as well as using quantum chemical calculation techniques to understand some of the phenomena that are witnessed and begin to develop predictive capability. The nonlinear characterization of compounds utilizes wavelengths outside of their linear absorption range, where a focused beam can achieve the same excitation as one at half the wavelength, though this effect has a quadratic dependence on power.The potential for nonlinear excitation, or two-photon absorption (2PA), is becoming of increasing interest and importance for organic chromophores. Exciting only a small volume of material at a focal point makes it possible to nondestructively image samples in 3-dimensions, record data in multiple layers, and fabricate intricate structures through photopolymerization reactions.Lanthanides such as europium are known to exhibit sharp emission bands when excited, typically through an antenna effect due to the low probability of achieving direct excitation. This emission is long-lived, and through gating systems can readily be separated from background noise and autofluorescence (often observed in biological samples) that have much shorter lifetimes. Thus, one of the foci of this dissertation is the photophysical investigation of a series of novel lanthanide complexes, with particular attention to a europium complex.
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Date Issued: 2014
Identifier: CFE0005908, ucf:50891
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005908

Title: Functional Identification of Nucleus Tractus Solitarius (NTS) Barosensitive Neurons: Effect of Chronic Intermittent Hypoxia (CIH).
Creator: Kolpakova, Jenya, Cheng, Zixi, Naser, Saleh, Kim, Yoon-Seong, Ebert, Steven, University of Central Florida
Abstract / Description: Chronic Intermittent Hypoxia (CIH) is a model used to study obstructive sleep apnea (OSA). Previously, we showed that baroreflex control of heart rate (HR) (baroreflex sensitivity) is reduced in CIH rats. While afferent function and HR in response to vagal efferent stimulationare enhanced, the effect of CIH on the central components, in particular NTS, is still notcompletely understood. F344 rats (3-4 mo) were exposed either to CIH or room air (RA) for 35-50 days. Following CIH exposure, rats...
Show moreChronic Intermittent Hypoxia (CIH) is a model used to study obstructive sleep apnea (OSA). Previously, we showed that baroreflex control of heart rate (HR) (baroreflex sensitivity) is reduced in CIH rats. While afferent function and HR in response to vagal efferent stimulationare enhanced, the effect of CIH on the central components, in particular NTS, is still notcompletely understood. F344 rats (3-4 mo) were exposed either to CIH or room air (RA) for 35-50 days. Following CIH exposure, rats were anaesthetized with Ket/Ace. Using single-unitextracellular recording technique, we recorded NTS barosensitive neurons in response to arterialpressure (AP) changes induced by descending aorta occlusion. Our data indicated that 1) themean arterial pressure and HR were similar in RA control and CIH groups. 2) The majority ofneurons from RA and CIH NTS neurons increased firing rate, whereas other neurons decreasedfiring upon AP elevation. 3) In 27 RA and 31 CIH NTS neurons with increased firing rate, 15 RA and 15 CIH neurons were activated at a low ?MAP at the early phase of AP increase (early neurons); whereas 12 RA neurons and 16 CIH neurons were activated at a late phase of AP increase (late neurons). The early neurons rapidly increased their firing during the rising phase of MAP, whereas late neurons did not increase their firing until the ?MAP reached its peak. 4) Early neuron activity-?MAP relationship was further characterized by the logistic sigmoid function curve. CIH significantly increased the maximal gain of the neuron activity-?MAP curve and the range of the response. In addition, CIH early neurons had a significantly higher firingrate than RA early neurons, whereas CIH did not change the firing rate in late neurons. 5) Forlate neurons, HR reduction correlated with neuronal activity. HR reduction-neuronal activityincrease curve was shifted to the right in CIH neurons, indicating that CIH decreased HR control in response to NTS firing increase. Collectively, our data suggest that NTS barosensitive neuronshave both early and late neurons, CIH selectively enhances neuron activity in response to APchanges in NTS early neurons and attenuate the baroreflex bradycardia. Along our previous workthat CIH-induced the cell loss in the nucleus ambiguus (NA), we conclude that CIH attenuatesthe functions of NA, whereas enhances the NTS functions to compensate for the loss of functionin NA
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Date Issued: 2015
Identifier: CFE0005967, ucf:50806
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005967

Title: Application of Two-Photon Absorbing Fluorene-Containing Compounds in Bioimaging and Photodyanimc Therapy.
Creator: Yue, Xiling, Belfield, Kevin, Campiglia, Andres, Miles, Delbert, Frazer, Andrew, Cheng, Zixi, University of Central Florida
Abstract / Description: Two-photon absorbing (2PA) materials has been widely studied for their highly localized excitation and nonlinear excitation efficiency. Application of 2PA materials includes fluorescence imaging, microfabrication, 3D data storage, photodynamic therapy, etc. Many materials have good 2PA photophysical properties, among which, the fluorenyl structure and its derivatives have attracted attention with their high 2PA cross-section and high fluorescence quantum yield.Herein, several compounds with...
Show moreTwo-photon absorbing (2PA) materials has been widely studied for their highly localized excitation and nonlinear excitation efficiency. Application of 2PA materials includes fluorescence imaging, microfabrication, 3D data storage, photodynamic therapy, etc. Many materials have good 2PA photophysical properties, among which, the fluorenyl structure and its derivatives have attracted attention with their high 2PA cross-section and high fluorescence quantum yield.Herein, several compounds with 2PA properties are discussed. All of these compounds contain one or two fluorenyl core units as part of the conjugated system. The aim of this dissertation is to discuss the application of these compounds according to their photophysical properties. In chapters 2 to 4, compounds were investigated for cell imaging and tissue imaging. In chapter 5, compounds were evaluated for photodynamic therapy effects on cancer cells. Chapters 2 and 3 detail compounds with quinolizinium and pyran as core structures, respectively. Fluorene was introduced into structures as substituents. Quinolizinium structures exhibited a large increase in fluorescence when binding with Bovine Serum Albumin (BSA). Further experiments in cell imaging demonstrated a fluorescence turn-on effect in cell membranes, indicating the possibility for these novel compounds to be promising membrane probes. Pyran structures were conjugated with arginylglycylaspartic acid peptide (RGD) to recognize integrin and introduced in cells and an animal model with tumors. Both probes showed specific targeting of tumor vasculature. Imaging reached penetration as deep as 350 ?m in solid tumors and exhibited good resolution. These results suggest the RGD-conjugated pyran structure should be a good candidate probe for live tissue imaging. Chapter 4 applied a fluorene core structure conjugated with RGD as well. Application of this fluorenyl probe compound is in wound healing animal models. Fluorescence was collected from vasculature and fibroblasts up to ? 1600 ?m within wound tissue in lesions made on the skin of mice. The resolution of images is also high enough to recognize cell types by immunohistochemical staining. This technology can be applied for reliable quantification and illustration of key biological processes taking place during tissue regeneration in the skin. Chapter 5 describes three fluorenyl core structures with photoacid generation properties. One of the structures showed excellent photo-induced toxicity. Cancer cells underwent necrotic cell death due to pH decrease in lysosomes and endosomes, suggesting a new mechanism for photodynamic therapy.
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Date Issued: 2014
Identifier: CFE0005565, ucf:50276
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005565

Title: Synthesis of Fluorene-based derivatives, Characterization of Optical properties and their Applications in Two-photon Fluorescence Imaging and Photocatalysis.
Creator: Githaiga, Grace, Belfield, Kevin, Patino Marin, Pedro, Chumbimuni Torres, Karin, Zou, Shengli, Cheng, Zixi, University of Central Florida
Abstract / Description: The two-photon absorption (2PA) phenomenon has attracted attention from various fields ranging from chemistry and biology to optics and engineering. Two of the common NLO applications in which organic materials have been used are three-dimensional (3D) fluorescence imaging and optical power limiting. Two-photon absorbing materials are, therefore, in great demand to meet the needs of emerging technologies. Organic molecules show great promise to meet this need as they can be customized through...
Show moreThe two-photon absorption (2PA) phenomenon has attracted attention from various fields ranging from chemistry and biology to optics and engineering. Two of the common NLO applications in which organic materials have been used are three-dimensional (3D) fluorescence imaging and optical power limiting. Two-photon absorbing materials are, therefore, in great demand to meet the needs of emerging technologies. Organic molecules show great promise to meet this need as they can be customized through molecular engineering, and as the development of two-photon materials that suit practical application intensifies, so does research to meet this need. However, there remains some uncertainty in the particulars of design criteria for molecules with large 2PA cross sections at desired wavelengths, as such research to understand structure-property relationships is matter of significant importance. As a result, the full potential of 2PA materials has not been fully exploited. Several strategies to enhance the magnitude and tune the wavelength of 2PA have been reported for ?-conjugated organic molecules. On this account, we have designed novel fluorophores using the fluorene moiety and modified it to tune the properties of the compounds.Chapter 2 of this dissertation reports the successful application of fluorene-based compounds in photocatalysis; a process that involves the decomposition of organic compounds into environmentally friendly carbon dioxide and water attesting to the photostability of the fluorene moiety. A facile organic nanoparticle preparation method is reported in chapter 3 using the reprecipitation method, whose surface was then modified using a naturally occurring surfactant, Lecithin, and were then successfully used in fluorescence cell imaging. Chapter 4 reports the design and synthesis of a fluorene-based compound using an acceptor, s-indacene-1, 3, 5, 7(2H, 6H)-tetra one, or Janus Dione, a moiety that is relatively new and that has not been fully exploited despite its very attractive features. Owing to the hydrophobicity of this compound, notwithstanding its unprecedented 2PA cross section, it was not applicable in fluorescence cell imaging but provided the tenets for the design of related derivative. This limitation was circumvented in the concluding chapter by tuning the compound's hydrophilicity. The hydrophilic Janus dione probe was then used as envisioned for cell imaging as the dual prerequisites for fluorescence imaging probes; large 2PA cross sections and high fluorescence quantum yields were met.
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Date Issued: 2015
Identifier: CFE0005620, ucf:50207
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005620

Title: Overexpression of human Cu/Zn Superoxide Dismutase in Mice: A Model to Study the Effect of Increased Superoxide Scavenging on the Autonomic Control of the Heart.
Creator: Hatcher, Jeffrey, Cheng, Zixi, Bossy-Wetzel, Ella, Fernandez-Valle, Cristina, Belfield, Kevin, University of Central Florida
Abstract / Description: Dysregulation of the autonomic cardiovascular control is a complication of diseases including diabetes, hypertension, sleep apnea, and aging. A common factor in these conditions is an increase in reactive oxygen species (ROS) in neural, cardiac, and endothelial tissues. Cu/Zn superoxide dismutase (SOD1) is an intracellular anti-oxidant enzyme that catalyzes dismutation of the superoxide anion (O2.-) to hydrogen peroxide (H2O2). Expression and function of this enzyme are diminished in...
Show moreDysregulation of the autonomic cardiovascular control is a complication of diseases including diabetes, hypertension, sleep apnea, and aging. A common factor in these conditions is an increase in reactive oxygen species (ROS) in neural, cardiac, and endothelial tissues. Cu/Zn superoxide dismutase (SOD1) is an intracellular anti-oxidant enzyme that catalyzes dismutation of the superoxide anion (O2.-) to hydrogen peroxide (H2O2). Expression and function of this enzyme are diminished in pathologies that impair cardiovascular autonomic control. This study employed mice overexpressing a transgene for human SOD1 (hSOD1) to determine if its overexpression would alter autonomic regulation of BP, HR, and BRS in healthy animals, and if this animal line (C57B6SJL-Tg (SOD1)2 Gur/J) could be used in future studies to determine if hSOD1 overexpression can preserve cardiac autonomic function in disease models. To accomplish this aim, using anesthetized SOD1 and C57 (control) mice, we recorded HR, and aortic depressor nerve (ADN) activity changes in response to pharmacologically-induced BP changes in order to measure baroreflex and baroreceptor sensitivity, respectively. In order to identify any alterations in central, efferent, and cardiac components of the baroreflex arc, we electrically stimulated the left ADN and left cervical vagus and compared the reductions in BP and HR between the C57 and SOD1 mice. Time- and frequency-domain analysis of heart rate variability (HRV) was performed using pulse pressure recordings prior to pharmacologic or surgical procedures. We found that hSOD1 overexpression in the SOD1 mouse line, in comparison to C57 controls did not significantly affect resting HR (C57: 558 (&)#177; 8 vs. SOD1:553 (&)#177; 13 beats-per-minute) or blood pressure (C57: 88.8 (&)#177; 2.9 vs.SOD1: 85.8 (&)#177; 2.1 mmHg). hSOD1 overexpression did not affect the decrease in average mean arterial pressure (MAP) following injection of sodium nitroprusside (SNP) (C57: 38.7 (&)#177; 1.4 vs. SOD1: 39.5 (&)#177; 1.3 mmHg) or increase in average MAP (C57: 135.8 (&)#177; 3.1 vs. SOD1: 136.6 (&)#177; 3.5 mmHg) following injection of phenylephrine (PE). BRS, as measured by the averaged regression lines for ?HR/?MAP for the SNP-induced tachycardic baroreflex (C57: 0.57 (&)#177; 0.06 bpm/mmHg, SOD1: 0.61 (&)#177; 0.08 bpm/mmHg)) and the PE-induced bradycardic baroreflex (C57: -2.9 (&)#177; 0.57 bmp/mmHg, SOD1: -4.3 (&)#177; 0.84 bpm/mmHg) are not significantly different between C57 and SOD1. Baroreceptor activation showed a significant increase in gain (C57: 5.4 (&)#177; 0.3 vs. SOD1: 7.4 (&)#177; 0.5 %/mmHg, P (<) 0.01) in the SOD1 transgenic mice. Heart rate depression in response to electrical stimulation of the left ADN and cervical vagus was comparable between C57 and SOD1, though MAP reduction in response to ADN stimulation is slightly, but significantly increased at 50 Hz in SOD1 animals. Time- domain analysis of HRV did not reveal any significant difference in beat-to-beat variability between SOD1 and C57 (SDNN: C57: 2.78 (&)#177; 0.20, SOD1: 2.89 (&)#177; 0.27), although frequency-domain analysis uncovered a significant reduction in the low-frequency power component of the HRV power spectral distribution (C57: 1.19 (&)#177; 0.11, SOD1: 0.35 (&)#177; 0.06, P (<) 0.001). This study shows that although hSOD1 overexpression does not affect overall baroreflex function, it does potentiate baroreceptor sensitivity and brain stem control of arterial pressure, and reduces low-frequency beat-to-beat variations in HR, without affecting total HRV.
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Date Issued: 2015
Identifier: CFE0005803, ucf:50025
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005803

Title: Squaraine dyes, design and synthesis for various functional materials applications.
Creator: Zhang, Yuanwei, Belfield, Kevin, Campiglia, Andres, Zou, Shengli, Frazer, Andrew, Cheng, Zixi, University of Central Florida
Abstract / Description: This dissertation contains the synthesis and characterization of squaraine based new functional materials. In the first part of this thesis work, a water soluble benzothiazolium squaraine dye was synthesized with pyridium pendents, and controlled aggregation properties were achieved. After formation of partially reversible J-aggregation on a polyelectrolyte (poly(acryl acid) sodium salt) template, the nonlinear, two-photon absorption cross section per repeat unit was found to be above 30-fold...
Show moreThis dissertation contains the synthesis and characterization of squaraine based new functional materials. In the first part of this thesis work, a water soluble benzothiazolium squaraine dye was synthesized with pyridium pendents, and controlled aggregation properties were achieved. After formation of partially reversible J-aggregation on a polyelectrolyte (poly(acryl acid) sodium salt) template, the nonlinear, two-photon absorption cross section per repeat unit was found to be above 30-fold enhanced compared with nonaggregate and/or low aggregates. Using a similar strategy, sulfonate anions were introduced into the squaraine structure, and the resulting compounds exhibited good water solubilities. A 'turn on' fluorescence was discovered when these squaraine dyes interacted with bovine serum albumin (BSA), titration studies by BSA site selective reagents show these squaraine dyes can bind to both site I and II of BSA, with a preference of site II. Introduction of these squaraine dyes to BSA nanoparticles generated near-IR protein nano fabricates, and cell images were collected. Metal sensing properties were also studied using the sulfonates containing a benzoindolium squaraine dye, and the linear response of the absorption of the squaraine dye to the concentration of Hg2+ makes it a good heavy metal-selective sensing material that can be carried out in aqueous solution. Later, a squaraine scaffold was attached to deoxyribonucleosides by Sonogashira coupling reactions, in which the reaction conditions were modified. Iodo-deoxyuridine and bromo-deoxyadenosine were used as the deoxyribonucleosides building blocks, and the resulting squaraine dye-modified deoxyribonucleosides exhibited near-IR absorption and emission properties due to the squaraine chromophore. Interestingly, these non-natural deoxyribonucleosdies showed viscosity dependent photophysical properties, which make them nice candidates for fluorescence viscosity sensors at the cellular level. After incubation with cells, these viscosity sensors were readily uptaken by cell, and images were obtained showing regions of high viscosity in cells.
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Date Issued: 2013
Identifier: CFE0005451, ucf:50369
Format: Document (PDF)
PURL: http://purl.flvc.org/ucf/fd/CFE0005451

two-photon absorption (3)	+ -
Neurodegenerative disease (2)	+ -
Two-photon absorption (2)	+ -
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Adherent-invasive Escherichia coli (1)	+ -

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University of Central Florida (20)	+ -
Cheng, Zixi (18)	+ -
Belfield, Kevin (8)	+ -
Campiglia, Andres (6)	+ -
Frazer, Andrew (4)	+ -

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Zou, Shengli (4)	+ -
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Cheng, Zixi (Jack) (2)	+ -
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