Doctor of Philosophy

dissertationThe chemotaxis signaling pathway of Escherichia coli is the best studied signal transduction mechanism in biology. Better understanding of this signal-processing machinery at the molecular level will foster new therapies for pathogenic infections and new designs of highly specific and sensitive biosensors. A sensory adaptation system plays a critical role in this chemotactic behavior. Sensory adaptation is regulated by covalent modifications of the chemoreceptors, mediated by CheR and CheB enzymes. This PhD research project explores the sensory adaptation mechanism of the serine receptor (Tsr) in E. coli. In this study, I showed that all adaptation sites of Tsr, including the fifth unorthodox site, worked in a similar way to regulate Tsr signal output. I also found that site 5 (Tsr-E502) and site 3 (Tsr-Q311) have differential signaling effects, mainly due to their different localizations on the methylation helices. Finally, I discovered unexpected signaling effects of CheR and CheB, the two adaptation enzymes. In summary, this thesis provides important insights into the sensory adaptation system and receptor input-output control in bacterial chemotaxis

PERKEMBANGAN KEMAMPUAN MEMBACA PADA ANAK USIA PRASEKOLAH : Studi Observasi Terhadap Seorang Anak Usia Prasekolah yang Belajar Membaca Berdasarkan Pendekatan Pengalaman Berbahasa

The increasing popularity of essential oils for skincare has led to investigation of their biological effects in human skin cells. In this study, we investigated the biological activities of three commercially available essential oils, i.e. rosemary oil, wild orange oil, and a blend (commercial name: Deep Blue) composed of oils from wintergreen, camphor, peppermint, blue tansy, German chamomile, Helichrysum, and Osmanthus, in a pre-inflamed human dermal fibroblast culture model, simulating chronic inflammation. The impact of essential oils on proteins associated with inflammation and tissue remodeling and on the genome-wide expression of 21,224 genes was investigated. The three essential oils diversely modulated global gene expression. Ingenuity Pathway Analysis showed that the oils affected numerous critical genes and signaling pathways. Specifically, rosemary oil influenced processes involved in cancer signaling and metabolism; orange oil affected processes related to cancer signaling, immunomodulation, and metabolism; the blend influenced inflammation, immunomodulation, and wound healing. These findings are largely consistent with the existing literature, supporting the beneficial biological activities of these essential oils. Our study provides the first evidence indicating how these essential oils affect genome-wide gene expression in human skin cells and establishes a basis for further research into their biological mechanisms of action

Autophagic Degradation Deficit Involved in Sevoflurane-Induced Amyloid Pathology and Spatial Learning Impairment in APP/PS1 Transgenic Mice

The adverse effects of anesthetics on elderly people, especially those with brain diseases are very concerning. Whether inhaled anesthetics have adverse effects on Alzheimer’s disease (AD), which is the most common form of dementia with brain degenerative changes, remains controversial. Autophagy, a crucial biological degradation process, is extremely important for the pathogenesis of AD. In this study, the inhaled anesthetic sevoflurane elicited many enlarged autolysosomes and impaired the overall autophagic degradation in the hippocampus of an AD mouse model, which is involved in the accumulation of amyloid-β (Aβ) and spatial learning deficits. However, rapamycin treatment counteracted all these effects. The results suggested that inhaled anesthetics may accelerate the pathological process of AD, and enlarged autolysosomes may be a new marker for prediction and diagnosis of the neurotoxicity of anesthetics in AD

TFEB Probably Involved in Midazolam-Disturbed Lysosomal Homeostasis and Its Induced β-Amyloid Accumulation

Alzheimer’s disease (AD) is one of the most common neurodegenerative diseases, and β-amyloid (Aβ) plays a leading role in the pathogenesis of AD. The transcription factor EB (TFEB), a main regulating factor of autophagy and lysosome biosynthesis, is involved in the pathogenesis of AD by regulating autophagy-lysosomal pathways. To date, the choice of anesthetics during surgery in patients with neurodegenerative diseases and evaluation of the effects and underlying mechanisms in these patients have rarely been reported. In this study, the HEK293-APP cells overexpressing APP and Hela cells were used. The cells were treated with midazolam at different concentrations and at different times, then lysosomes were stained by lysotracker and their morphology was observed under a fluorescence microscope. The number and size of lysosomes were analyzed using the ImageJ software. The levels of TFEB in the nucleus and APP-cleaved intracellular proteins were detected by nuclear separation and Western Blot. Finally, ELISA was used to detect the levels of Aβ40 and Aβ42 in the cells after drug treatment. We found that 30 μM midazolam decreased the number of lysosomes and increased its size in HEK293 and HeLa cells. However, 15 μM midazolam transiently disturbed lysosomal homeostasis at 24 h and recovered it at 36 h. Notably, there was no significant difference in the extent to which lysosomal homeostasis was disturbed between treatments of different concentrations of midazolam at 24 h. In addition, 30 μM midazolam prevents the transport of TFEB to the nucleus in either normal or starved cells. Finally, the intracellular C-terminal fragment β (CTFβ), CTFα, Aβ40 and Aβ42 levels were all significantly elevated in 30 μM midazolam-treated HKE293-APP cells. Collectively, the inhibition of TFEB transport to the nucleus may be involved in midazolam-disturbed lysosomal homeostasis and its induced Aβ accumulation in vitro. The results indicated the risk of accelerating the pathogenesis of AD by midazolam and suggested that TFEB might be a candidate target for reduction of midazolam-dependent neurotoxicity

<i>In vitro</i> biological activities of Douglas fir essential oil in a human skin disease model

<p>Although essential oils from Douglas fir are popular topical skincare products, research regarding their biological effects on human skin cells is scarce. Here, we studied the biological activity of a commercially available Douglas fir (<i>Pseudotsuga menziesii</i>) essential oil (DEO) in a human dermal fibroblast model of chronic inflammation and fibrosis induced by stimulation with cytokines. Chemical analysis of DEO indicated that its major chemical components (i.e. >5%) were beta-pinene (23%), sabinene (17%), terpinolene (14%), delta-3-carene (11%), and alpha-pinene (9%). We analyzed the effect of DEO on the levels of 17 important protein biomarkers associated with inflammation, immune system modulation, and tissue remodeling. DEO exhibited significant anti-proliferative activity in human fibroblasts. DEO also significantly inhibited the production of vascular cell adhesion molecule 1, collagen III, and plasminogen activator inhibitor 1. We also observed that DEO robustly modulated global gene expression levels in diverse ways. In particular, DEO affected the expression of genes involved in immune modulation and cancer signaling. This study provides the first evidence of biological activity of DEO in human dermal fibroblasts. Our results suggest that DEO may modulate immune responses and tumor signaling processes. Further research about the biological and pharmacological mechanisms of DEO action is recommended.</p

Biological activity of vetiver (Vetiveria zizanioides) essential oil in human dermal fibroblasts

Vetiver (Vetiveria zizanioides) essential oil (VEO) has a long history of use. However, research on its biological activity in human skin cells is scarce. In this study, we investigated the biological activity of VEO in a pre-inflamed human dermal fibroblast model, which was designed to mimic the disease biology of chronic inflammation and fibrosis. We analyzed the impact of VEO on the levels of 17 important protein biomarkers pertinent to immune response and tissue remodeling. VEO exhibited strong antiproliferative activity in these cells and significantly inhibited the production of collagen III, an important molecule for skin and tissue remodeling processes. We also studied the effect of VEO on regulating genome-wide gene expression. VEO robustly impacted many genes and signaling pathways that are closely related to tissue remodeling and metabolism, among others. Specifically, VEO significantly impacted pathways for cholesterol synthesis and metabolism. This study provides the first evidence of the biological activity of VEO in human dermal fibroblasts. Though a definite conclusion remains elusive, the data suggest that VEO has therapeutic potential for both cosmetic and metabolic health care products. Further research into VEO’s biological and pharmacological mechanisms of action is recommended

Anti-inflammatory activity of Juniper (Juniperus communis) berry essential oil in human dermal fibroblasts

Although juniper (Juniperus communis) berry essential oil (JEO) has been used in skin care products, research on its biological activity in human skin cells is scarce. In the current study, we explored the biological activity of JEO (with alpha-pinene as the major active component) in pre-inflamed human dermal fibroblasts, which were designed to mimic the disease biology of chronic inflammation and fibrosis. We analyzed the levels of 17 important protein biomarkers pertinent to inflammation and tissue remodeling. JEO exhibited robust antiproliferative activity and significantly inhibited the increased production of the proinflammatory chemokines interferon gamma-induced protein 10 (IP-10) and interferon-inducible T-cell alpha chemoattractant (I-TAC). Additionally, JEO significantly inhibited tissue remodeling biomarkers, namely collagen I, collagen III, and plasminogen activator inhibitor 1 (PAI-I). Macrophage colony-stimulating factor (M-CSF), an immunomodulatory protein molecule, was also significantly downregulated by JEO. Moreover, we found that JEO robustly modulated global gene expression. Ingenuity Pathway Analysis also showed that JEO affected many important signaling pathways that are closely related to metabolism, inflammation, immune response, wound healing, and cancer biology. This study provides the first evidence of the biological activity of JEO in human dermal fibroblasts. Thus, JEO is a promising therapeutic candidate for inflammatory conditions in the skin

Anti-inflammatory activity of clove (Eugenia caryophyllata) essential oil in human dermal fibroblasts

Context: Clove (Eugenia caryophyllata Thunb. [Myrtaceae]) essential oil (CEO) has been shown to possess antimicrobial, antifungal, antiviral, antioxidant, anti-inflammatory and anticancer properties. However, few studies have focused on its topical use. Objective: We investigated the biological activity of a commercially available CEO in a human skin disease model. Materials and methods: We evaluated the effect of CEO on 17 protein biomarkers that play critical roles in inflammation and tissue remodelling in a validated human dermal fibroblast system, which was designed to model chronic inflammation and fibrosis. Four concentrations of CEO (0.011, 0.0037, 0.0012, and 0.00041%, v/v) were studied. The effect of 0.011% CEO on genome-wide gene expression was also evaluated. Results and discussion: CEO at a concentration of 0.011% showed robust antiproliferative effects on human dermal fibroblasts. It significantly inhibited the increased production of several proinflammatory biomarkers such as vascular cell adhesion molecule-1 (VCAM-1), interferon γ-induced protein 10 (IP-10), interferon-inducible T-cell α chemoattractant (I-TAC), and monokine induced by γ interferon (MIG). CEO also significantly inhibited tissue remodelling protein molecules, namely, collagen-I, collagen-III, macrophage colony-stimulating factor (M-CSF), and tissue inhibitor of metalloproteinase 2 (TIMP-2). Furthermore, it significantly modulated global gene expression and altered signalling pathways critical for inflammation, tissue remodelling, and cancer signalling processes. CEO significantly inhibited VCAM-1 and collagen III at both protein and gene expression levels. Conclusions: This study provides important evidence of CEO-induced anti-inflammatory and tissue remodelling activity in human dermal fibroblasts. This study also supports the anticancer properties of CEO and its major active component eugenol

Cardamom (Elettaria cardamomum) essential oil significantly inhibits vascular cell adhesion molecule 1 and impacts genome-wide gene expression in human dermal fibroblasts

Cardamom (Elettaria cardamomum) essential oil (CEO) is popular in skin care, although no studies have reported its biological activity in human skin cells. We studied its effect on 17 protein biomarkers closely related to inflammation, immune responses, and tissue remodeling using a dermal fibroblast cell culture system designed to model chronic inflammation. CEO significantly inhibited the proliferation of skin cells and the expression of vascular cell adhesion molecule 1 (VCAM-1) and macrophage colonystimulating factor (M-CSF). The CEO-induced inhibition of the production of these protein biomarkers suggests its anti-inflammatory and immunomodulatory potential, which has been largely attributed to its major active component, eucalyptol. We further studied the effect of CEO on the expression of 21,224 genes in the same cell culture system. Ingenuity pathway analysis showed that CEO affected critical genes and signaling pathways closely involved in inflammation, immune responses, and tissue remodeling. The observed overall CEO-induced inhibition of these genes and pathways supports its anti-inflammatory and immunomodulatory properties. This study provides important evidence of the biological activity of CEO in human skin cells. Further research into the mechanism of action of CEO in human skin and other systems is recommended