Thesis (Ph. D.)--University of Hong Kong, 2002. / Includes bibliographical references (leaves 110-140).

Thesis (Ph. D.)--University of Washington, 2003. / Vita. Includes bibliographical references (leaves 69-103).

published_or_final_version / Psychiatry / Doctoral / Doctor of Philosophy

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

Melanoma is a malignant type of skin cancer arising from the combined effects of genetic alteration and extrinsic signaling, resulting in transformation of neural crest (NC)-derived melanocytes into metastatic melanoma. Current therapies against metastatic melanoma are merely effective with less than 5% 5-year survival rate of patients. Understanding the underlying molecular mechanism of how melanoma acquires metastatic behavior could formulate strategies for new therapeutic options. Features of metastatic melanoma resemble NC cells undergoing an epithelial-mesenchymal transition (EMT) suggesting similar regulators might be in place to control the process. Our previous studies showed that SoxE transcription factors (Sox8/9/10) play a crucial role in NC development, in particular Sox9 transactivates expression of Snail2 and co-operates with it to induce features of EMT. To examine the role of SOXE proteins in melanoma development and whether they regulate SNAIL expression, we first investigated the expression profile of SOXE and SNAIL in a human melanoma tissue array. The data showed that SOX8, SOX10, and SNAIL genes are highly expressed in metastatic melanoma whereas SOX9 and SNAIL2 transcript levels are low. Moreover, SNAIL transcript level was shown to have a positive correlation with SOX8 and SOX10 expression levels. SNAIL is well-known to be the key regulator of tumor invasiveness in various cancers. Our data raised the possibility that SOXE proteins may also regulate SNAIL expression in initiating melanoma metastatic behavior. The human metastatic melanoma cell line A375 exhibits similar SOXE and SNAIL expression profiles as the tissue array. Knockdown of SNAIL in A375 reduced its migratory ability and in vivo tumorigenecity, suggesting that SNAIL plays a crucial role in melanoma metastasis. How SNAIL transcription is regulated in melanoma has been poorly understood. Previous studies have identified a minimal enhancer region downstream of the SNAIL locus which contains YY1 and SOX consensus binding sequences. Chromatin immunoprecipitation assay revealed that SOX8 and SOX10 proteins could bind to the SNAIL 3’ minimal enhancer region specifically. Mutation of the SOX consensus binding sequence reduced the enhancer activity while mutations in both SOX and YY1 binding sites resulted in further reduction suggesting that YY1 and SOX protein binding is required and important for enhancer activity and SNAIL transcription. These findings provide a molecular basis to examine further whether metastasis of melanoma is regulated by SOXE proteins in which one of the potential mechanisms could act through regulation of SNAIL expression. / published_or_final_version / Biochemistry / Master / Master of Philosophy

Background: Breast cancer is the most frequent female cancer in both developed and developing world which comprising 16% of all female cancer according to WHO GLOBOCAN 2008. The statistic from Hong Kong Cancer Registry reported that breast cancer is the third commonest cause of female death in Hong Kong. Breast cancer incidence varies remarkably among developed countries. The high dietary consumption of soy isoflavones has been hypothesized to explain the lower breast cancer incidence among women in Asian countries in observational studies, but whether soy isoflavones exert estrogenic or anti-estrogenic in breast tissue remains uncertain. Objective: This systematic review was to assess the effects of isoflavone-rich soy consumption on breast cancer risk in pre- and post-menopausal women Methods: Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines for conducting and reporting randomized controlled trials were followed. Data sources: A systematic review of randomized controlled trials was conducted through searching databases: MEDLINE, PubMed and Cochrane Library (2002 until March 2012). Keywords for electronic searches included: [(soy OR isoflavones) AND (breast cancer OR breast neoplasms)] limited study types to human & randomized controlled studies. Study selection: RCTs of the effects of isoflavones or supplement versus placebo or control diet among pre- and post-menopausal participants who were currently free from breast cancer. Outcome measurements: serum sex hormones and IGF profile, mammographic density and menstruation cycle length Results: 15 RCTs (1527 women) compared isoflavones with placebo or control diet for study duration ranged from 2 months to 2 years. No significant effect was found on serum sex hormones, IGF profile, mammographic density or menstrual cycle length. The effect of menstrual cycle on mammographic densities was noticed. Conclusion: The results of the systematic review did not support the hypothesis that short-term isoflavones exposure has an effect on modulating breast cancer risk. The effect of menstrual cycle on mammographic densities probably reflects the effect of hormonal changes. Null results did not necessarily contradict the inverse association between soy intake and breast cancer risk from the results of epidemiologic studies. The absence of conclusive data on the effects might be attributable to the insufficient exposure duration in the RCTs. Longer duration of soy exposure and early life exposure might be a scope for future research. / published_or_final_version / Public Health / Master / Master of Public Health

published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

Neural stem cells (NSCs) exist in both embryonic and adult neural tissues and are characterized by their self-renewal capacity and multipotency that contribute to the generation of three major cell types in the vertebrate central nervous system (CNS):neurons, oligodendrocytes and astrocytes. The tremendous therapeutic potential of NSCs to treat the neurodegenerative diseases and repair brain injuries has provoked intensive study in the molecular regulation of their induction, maintenance and differentiation. Previous study reported that Sox9, a member of high-mobility-group(HMG) containing SoxE transcription factors family, plays important roles in regulating the formation and maintenance of NSCs in both mouse and chick CNS, as well as the cell fate switch between neuronal and glial. Whether it plays similar roles in human NSCs (hNSCs)is still unknown. My RT-qPCR analysis showed that SOX9is expressed at a basal level in human embryonic stem cells (hESCs) and up-regulated upon commitment into neural lineage and maintained at a high level in hESCs-derived hNSCs. I therefore hypothesized that SOX9 might also be involved in the induction, maintenance and differentiation of hNSCs. To test this, two stable hESC lines(HES2)were generated with each constitutively expressing short hairpin RNA (shRNA) against SOX9andGL2 Luciferase (Luc, as control) respectively. Upon neural induction, SOX9-knock-down(KD) hESCs were able to commit neural lineage and differentiate into NSCs/neurospheres (NSPs), however, these NSCs exhibited reduced multipotency and glial marker (GALC, CD44) expressions but enhanced self-renewal compared to the shLuc NSCs. Hence, SOX9 is required for both the induction and maintenance of multipotent hNSCs. Strikingly, extensive TUJ1+ neurites and advance groupings of these neurites into bundles were observed in SOX9-KD NSPs after three days and seven days neuronal differentiation respectively, suggesting premature neurogenesis as a result of SOX9 ablation. In addition, RT-qPCR analysis revealed down-regulated expression of NSC marker HES1but induced proneural basic helix-loop-helix transcription factor MASH1in shSOX9-1208 NSCs. The inhibitory role of HES1 on the expression and functions of MASH1 has been reported to be essential for the timely generation of neurons. Hence, ablation of SOX9 is likely to relieve the inhibition on MASH1activity via down-regulated HES1expression and leads to early neuronal differentiation. Expression of the potent neurite blocker NG2 was also found to be reduced in SOX9-KD NSCs which may explain the extensive neurite network observed. Altogether, similar to previous studies in mouse NSCs, SOX9 is also required for the induction and maintenance of hNSCs. However, this study further reveals a putative novel role of SOX9 in preventing premature neuronal differentiation by regulating the expressions of HES1 to counteract MASH1 function and NG2 to control neurite outgrowth. / published_or_final_version / Biochemistry / Master / Master of Philosophy

Background Radiological examination is a common diagnostic practice in modern medicine, they are not uncommonly performed during pregnancy or childhood. The potential biological effects of radiation to both the developing fetus and children are not always clear and remained controversial over many years. Physicians who care for these patients always find it difficult to evaluate the risk, and have misconceptions regarding the use of ionized radiation in pregnancy and children, which may delay the management process. Objective This study has reviewed all recent published observational studies, and analyse any possible association of prenatal or postnatal X-ray exposure from diagnostic imaging and childhood cancer risk. Methods Eligible epidemiological studies published between January 2000 and June 2013 were reviewed. These studies were found through electronic searches using Medline, PubMed, Embase, and Cochrane Database. Predetermined inclusion and exclusion criteria were applied to the identified articles Results Twenty-five articles with fourteen million participants were recruited. 17 out of 25 were case-control studies and 8 were cohort studies. All studies tried to prove an association between X-ray or CT scan exposure, and cancer of the haematopoietic system, brain and soft tissue regions. Results were summarized separately for their study methods, mode of radiation exposure and for each cancer outcome. The quality of the articles was accessed with the Newcastle-Ottawa scale. The overall OR estimate from case-control studies showed postnatal X-ray exposure positively and significantly associated with leukaemia risk (OR 1.21; 95% CI: 1.10-1.32; I2 = 3%). Cancer risk other than leukaemia are lacking in case-control studies. Recent cohort studies also showed a small but significant increase risk of leukaemia and brain tumour from childhood CT scan exposure. Conclusion This analysis had shown a small but significant increase cancer risk from X-ray or CT scans exposure in postnatal period. Varies measures should be used to minimize the radiation dose in children during radiation exposure. As long as the radiological imaging is clinically indicated and performed using appropriate scanning protocol, the benefits of radiological imaging should far outweigh the small radiation risk. / published_or_final_version / Public Health / Master / Master of Public Health

Circadian rhythmsare biochemical, physiological, and behavioral processes display oscillations of oughly 24-hour, which existing in both prokaryotes and eukaryotes. Circadian rhythms improve fitness of organisms in both constant and changing environments. The cryptochrome (CRY)and PAS-containing proteins are light sensors and key elements of the circadian system in eukaryotic organisms. Photolyases and cryptochromes are evolutionarily related flavoproteins which perform distinct physiological functions. Photolyases are evolutionarily ancient enzymes that activated by light and repairing UV-induced DNA damage. Although cryptochromes share structural similarity with the DNA photolyases, they lack the DNA repair activity. CRYs are key elements of mammal circadian system, and play roles in light sensing in insects and plants to entrain circadian rhythms. The PAS domains are widely distributed in proteins across all kingdoms of life and act as signal modules. They are common in photoreceptors and transcriptional regulators of eukaryotic circadian clock components including bHLH-PAS proteins (BMAL, CYC,CLK and NPAS2) and PER in animals, PHY and ZTL in plants, WC-1, 2and VVD in fungi. They are mainly involved in protein-protein interaction and light sensing functions. The CRY/PHR superfamily consists of 7 major subfamilies: CPD class I and CPD class II PHRs, (6-4) PHR, CRY-DASH, plant PHR2, plant CRY and animal CRY. Although the superfamily evolved primarily under strong purifying selection (average ω = 0.0178), it experienced strong episodic positive selection at some periods of evolution. The level of variation is subfamily-and domain-specific. The homologs with apparent circadian functions (i.e., plant and animal CRY) are significantly more conserved than the other photolyases. Photolyases were lost in eukaryotic groups like placental mammals, suggesting that natural selection apparently became weaker in the late stage of evolutionary history. The phylogenetic trees of fish Cry features two major clusters, which correspond to Cry1and Cry2. Teleost species possess extra copies of Cry1 due to fish-specific genome duplication (FSGD), and formed 3 clades of Cry1. Clade1B of Cry1(π= 0.129 ±0.062) is more conserved than the other paralogs (πrange from 0.173to 0.195). Test of positive selection revealed that fish cryptochromes evolved under strong purifying selection (average ω= 0.0066).Different fishes preserved different Cry duplicates that associated with reciprocal gene loss, thus generated the diverse circadian molecular mechanisms. The level of DNA variation in the PAS-containing proteins appears to be subfamily-specific. The animal PAS-containing homologs are more polymorphic than the plant and fungal homologs. Although the whole superfamily evolved primarily under strong purifying selection (average ω range from 0.0030to 0.1164), it experienced strong positive selection at some periods of the evolution. Although the PAS domains from different proteins vary in sequence and length, they maintain a fairly conserved 3D structure. The 3D fold of PAS domains is determined by only 8 conserved residues which shared by all subfamilies. The evolutionary time estimates showed that plant and animal Cry, WC-1& 2, bHLH-PAS proteins and Per originated in the Neoproterozoic Era (~1000 –542 Mya), plant Phy and ZTL evolved in the Paleozoic (541 –252 Mya), which might be a result of adaptation to the global climate and light regime changes. / published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy