Stem cell-based therapies are promising in regenerative medicine. However, the success of cell therapy is greatly limited by the low engraftment rate to the target tissues. The present study demonstrated that human mesenchymal stem cells (hMSCs) were subjected to a self selection process via microencapsulation in collagen barrier when they were induced to migrate out from this barrier. While retaining the immuophenotype and self renewal capacity, the selected hMSCs showed a significantly better in vitro migratory response of than those cultured in traditional monolayer. The migratory response could be controlled by varying the fabrication parameters of the collagen barrier, including initial collagen concentration and cells seeding density. Affinity to adhere on endothelial cells layer is another engraftment related property. Significant difference was observed between these selected hMSCs and hMSCs in monolayer culture. In order to investigate the engraftment potential of the selected hMSCs, an animal model was performed. The selected hMSCs were transplanted intravenously into NOD/SCID mice under partial hepatectomy. Presence of human cells in the residual liver was determined by the presence of human HLA-ABC using flow cytometry after 48 hours, 1 week and 1 month. Engraftment of the selected hMSCs was significantly higher than that of monolayer cultured hMSCs in time point of 1 month. It demonstrated that the selected hMSCs favor the engraftment to the injured liver. Further investigation is required to determine the fate of the engrafted hMSCs in order to truly confirm their therapeutic potential. The current work demonstrated that collagen-hMSCs microsphere could act as a barrier to select hMSCs with enhanced in vitro migratory response and in vivo engraftment properties. These findings may contribute towards the development of better stem cell therapies. / published_or_final_version / Mechanical Engineering / Master / Master of Philosophy

Background: Organ transplantation is getting more important and the demand increased exponentially in the past decade. However, due to the shortage of organ supply, some people died while waiting for an organ and the other may rely on all forms of organ support to live. Hong Kong has the lowest organ donation rate among the developed world despite large-scale organ donation enhancement activities implemented for decade. Since consent from family members is mandatory, it is important to understand the reasons relatives given for consenting or refusing to donation. Objectives: (1) To investigate the change of attitudes towards organ donation in Hong Kong during 2004 to 2012. (2) To understand the difficulties of transplant coordinators encountered during counseling Methods: This is a cross-sectional survey looking at the recent change of reasons of the families of the potential donors to make decision about donation. Transplant coordinators of all 7 clusters completed questionnaires after counseling each potential donor referred during the 7 months study period. Demographic data and reasons of refusal or consent of donation were recorded and analyzed and compared with previous results. Difficulties of transplant coordinators encountered during counseling were recorded. Results: Totally 52 questionnaires were completed within 7 months. The conversion rate were similar when compared to that in 2004 (44.3% in 2004 vs 40% in 2012). The proportion of older potential donor in the refusal group is higher than that of consent group (> 60 years old 51.6% vs 38%,). The demographic data was neither statistical significantly different between consent and refusal group nor correlated with refusal of donation. From 2004 to this survey, the most common reason to refusal remained to be the traditional belief of buried intact. Both were 33%. The “expressed wish of the deceased to object donation while alive” increased from 13% in 2004 to 20% in 2012 (p=0.378), and “no consensus from family” dropped from 26% in 2004 to 11% in 2012 (p=0.083). The reason “no expression of any wish” remained static when compared to that in 2004 (15% vs 16%, p=0.982). The reason for consenting to donation changed a lot. Although the most common reason remained the “wish to help others” (94% vs 41%, p<0.0001), the second and third reason changed to “feeling good by being charitable” (21% vs 0%, p=0.0071) and “not to waste useful organs” (18% vs 3%, p=0.0455) while the fourth reason was “respect deceased’s choice”(14% vs 0%, p=0.0278). The reasons of refusal and consenting donation are explained in the context of the decision-making process. The main obstacles the transplant coordinators have encountered were knowledge gap, emotion, time pressure and family or social disharmony. Conclusion: The major reasons for consenting and refusing donation did not only change in position but also change in proportion in last decade. The importance of some reasons increased significantly and strategies to improve conversion rate should be adjusted according to the changing reasons. Education to change the traditional belief, encourage expression of donation wish, create position image of donation and the benefit of transplant are all important. / published_or_final_version / Public Health / Master / Master of Public Health

Intermedin (IMD) is a peptide hormone discovered in 2004 belonging to the calcitonin/calcitonin gene-related peptide superfamily. It signals through a Gprotein coupled receptor by the coupling of a calcitonin receptor-like receptor (CRLR) and one of the receptor activity-modifying proteins (RAMPs) 1-3. Due to its similarity to adrenomedullin in structure and functions, IMD is also known as adrenomedullin 2 (ADM2). Among members of the superfamily, IMD shares the highest degree of homology with ADM, which is a multifunctional vasodilator ubiquitously expressed in various tissues and organs and has been studied by our group for its reproductive functions. It is hypothesized that IMD may be present in the reproductive systems of the rat and exert some effects on reproductive functions. The objectives of this study were to investigate the expression of IMD and its receptor components in the male and female reproductive systems of the rat, the changes in expression across the oestrous cycle, and its effect on uterine contraction. The gene expression levels of Imd and its receptor components and peptide levels of IMD were measured by RT-PCR and enzyme immunoassay respectively. The effect of IMD on the uterine contraction was studied by the organ bath technique. Imd mRNA and IMD levels were detected in the testis, epididymis, ventral prostate, coagulating gland, and seminal vesicle of the male rat and the ovary, oviduct, and uterus of the female rat, suggesting possible roles for IMD in both the male and female reproductive systems. In the male, the Imd mRNA levels were the highest in the seminal vesicle but lowest in the testis and the epididymis and IMD peptide levels were the highest in the coagulating gland but lowest in the epididymis. In the female, the Imd mRNA and IMD peptide levels were the highest in the oviduct and the uterus respectively while both the Imd and IMD levels were the lowest in the ovary. Imd mRNA and IMD levels displayed cyclic changes in various female reproductive tissues across the oestrous cycle. In the ovary, positive immunostaining was detected in the follicles and corpora lutea with more staining in the latter. The Imd mRNA level was significantly higher at prooestrus than dioestrus while the IMD peptide level was significantly higher at metoestrus than dioestrus. In the oviduct, the Imd mRNA level was the lowest at dioestrus but the IMD peptide level was the highest at dioestrus. Positive immunostaining was observed in the ciliated epithelial cells. Uterine Imd mRNA level was the highest at prooestrus while the IMD level was the highest at dioestrus. IMD was found in the luminal and glandular epithelia. IMD significantly reduced the uterine contraction amplitude and frequency but not the basal tone. CGRP receptor antagonist hCGRP8-37 and ADM receptor antagonist hADM22-52 partially abolished the inhibitory effect of IMD on uterine contraction while the IMD specific receptor antagonist hIMD17-47 completely blocked the actions. Enzyme inhibitors of NO (L-NAME) and PI3K (Wortmannin) pathways diminished the IMD-mediated effects on uterine contraction while cAMP/PKA blocker KT5720 had no effect. / published_or_final_version / Physiology / Master / Master of Philosophy

A safe, potent and broad-spectrum antiviral is urgently needed to combat emerging viral respiratory diseases such as avian influenza H5N1 and H7N9, severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS). Previous studies carried out by PhD students in our lab found that mouse β-defenisn 4 (mBD4) shows highly antiviral activity in vitro. However, the recombinant mBD4 (rmBD4) expressed by E.coli is limited to very small scale of production and is very expensive. Thus, in this study, we firstly screened 16 short peptides derived from mBD4 and other mouse and human β-defensins for identifying their antiviral effects. One short peptide P9 (30 amino acids), derived from mBD4, exhibited potent and broad-spectrum antiviral effects against multiple respiratory viruses, including influenza A viruses H1N1, H3N2, H5N1, H7N7 and H7N9, SARS coronavirus (SARS-CoV)and MERS coronavirus (MERS-CoV). This P9 showed very high selectivity index (970), which was higher than that of the full-length peptide of synthetic mBD4 (smBD4) and rmBD4 in vitro. Secondly, the prophylactic and therapeutic effects of P9 against the infection of H1N1 virus were further detected in animal model. The survival rate of P9-pretreated mice challenged by lethal dose of H1N1 virus was 100%. The therapeutic effects of P9 protecting mice from lethal challenge of H1N1 virus were also statistically significant. The survival rate of mice could reach up to 67% by intranasal inoculation and 56% by intraperitoneal injection, respectively. To investigate the antiviral mechanism, we firstly elucidated that P9 could inhibit viral infection but not viral replication or release. Secondly, we detected whether P9 inhibited viral infection by binding to the surface of target cells or viral particles. The results showed that P9 only bound to viral particles but not to the cell surface. It was further identified that P9 bound to viral surface glycoprotein HA but not NA. Thirdly, we demonstrated that P9 did not inhibit virus binding to its receptor and block the virus entry into cells by endocytosis. Instead, P9 inhibited the acidification in late endosomes and thusP9 blocked virus-membrane fusion and subsequent viral disassembly and viral RNA release. Finally, we elucidated that the antiviral activity of P9 was attributed to its high binding affinity to viral HA and the abundance of basic amino acids in its composition. In this study, we have demonstrated that a short peptide P9, which is derived from mBD4, showed potent antiviral activity against multiple respiratory viruses. This peptide can be developed to a new promising prophylactic and therapeutic agent with broad-spectrum antiviral activity and low possibility to cause drug resistance. Moreover, this study has also revealed a novel antiviral mechanism for P9 and paved a path for the development of new antiviral agents with broad-spectrum antiviral activity against emerging respiratory viruses, such as avian influenza H5N1 and H7N9, as well as SARS-CoV and MERS-CoV. / published_or_final_version / Microbiology / Doctoral / Doctor of Philosophy

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This thesis is concerned with the chemical control of breathing during exercise in humans. Chapter 1 reviews some of the relevant studies in animals and humans. Chapter 2 describes the experimental apparatus and the technique of dynamic end-tidal forcing performed using a computer-controlled gas-mixing system. Chapter 3 describes a study of the effects of sustained hypoxia on ventilation during steady exercise. The acute ventilatory response to hypoxia (AHR) was increased during exercise as compared with rest, but the magnitude of the subsequent decline in ventilation (HVD), expressed as a fraction of the AHR, was reduced. A simple model of the hypoxic peripheral chemoreflex is proposed, in which the mechanisms underlying AHR and HVD are functionally separate and can be independently modulated by external factors. Chapter 4 assesses changes in peripheral chemoreflex sensitivity to hypoxia in terms of the degree of decline in AHR measured in the resting periods shortly after prior conditioning periods of hypoxia and/or exercise. At rest, a second AHR measured 6 min after a period of sustained hypoxia had declined by 30% as compared with the initial AHR. In contrast, the AHR measured in the resting period after a period of sustained hypoxic exercise was only 11% smaller in magnitude than the AHR measured after a period of euoxic exercise. The results suggest that the degree to which hypoxic sensitivity declines during sustained hypoxia is genuinely attenuated, rather than masked, by exercise. Chapter 5 describes the changes in respiration during prolonged exercise breathing air with and without added CO₂. During prolonged poikilocapnic exercise, ventilation remained constant, but metabolic CO₂ production, respiratory quotient and end-tidal P_CO2 declined; a result which suggests that in man, ventilation can be dissociated from the CO₂ flux. During hypercapnic exercise, ventilation progressively increased; this was interpreted as being due to a correction by end-tidal forcing of the natural tendency for end-tidal CO₂ to decline, together with an independent effect of CO₂ per se on the ventilation. Chapter 6. Electrical muscle stimulation was used as means of inducing non-volitional exercise. Electrically-induced exercise increased the AHR as compared with rest, and with voluntary exercise at matched external work rate. The AHRs during electrical stimulation and voluntary exercise matched to the internal work rate were similar. Chapter 7. Electrical muscle stimulation was used in paraplegic subjects in whom there would be no neural control of exercise. Electrically-induced exercise increased the AHR as compared with rest. When compared with the data from Chapter 6, the results suggest that the observed increase in AHR during normal voluntary exercise can be wholly accounted for by the increase in metabolic CO₂ production, or closely related factors. Chapter 8 presents a brief summary of the findings in this thesis.

published_or_final_version / Medical Sciences / Master / Master of Medical Sciences

published_or_final_version / abstract / toc / Obstetrics and Gynaecology / Doctoral / Doctor of Philosophy

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