Resolving the liver sinusoidal endothelial phenotype in health and disease

The burden of liver disease is continuously increasing globally, and this emphasises the need for the development of therapeutics. In order for this to be achieved, potential cellular and molecular targets need to be identified. Liver sinusoidal endothelial cells (LSECs) play a key role in maintaining liver homeostasis and their dysfunction drives liver disease pathophysiology and this role needs to be further elucidated. In order to identify phenotypic differences in LSECs in health and disease, a combination of analytical techniques such as immunohistochemistry and qPCR was applied on human tissue specimens. To confirm whether these changes are recapitulated in vitro, I isolated LSECs from human healthy and cirrhotic tissue specimens for the establishment of culture model of human LSECs. Validation of functional and phenotypic characteristics of LSECs in vitro was carried out using immunocytochemistry and qPCR. Furthermore, the development and optimisation of a super-resolution imaging protocol for the visualisation of LSEC fenestrations was performed. Altered expression and downregulation of scavenger receptors in LSECs was identified in diseased human tissue specimens compared to healthy specimens and this confirmed capillarisation of sinusoidal endothelial cells in liver disease. Expression of scavenger receptors and key regulatory molecules was maintained in LSECs in vitro. The phenotypic changes in LSECs identified in liver tissue specimens were partially recapitulated in LSECs in vitro. The application of pharmaceutical molecules for the enhancement of nitric oxide (NO) signalling in LSECs revealed an altered genotype in healthy and cirrhotic LSECs. Finally, fenestrations were visualised on the LSEC membrane using the developed super-resolution imaging protocol and improvement in LSEC porosity following the application of sildenafil citrate. Hence these findings emphasise the relevance of appropriate culture models and imaging approaches to study phenotypic changes in LSECs in relation to disease and highlight the therapeutic potential of sildenafil citrate in improving LSECs porosity

Accessory Gene Regulator Group Polymorphisms in Methicillin-Resistant Staphylococcus aureus: An Association with Clinical Significance

PURPOSE: Virulent gene expression in Staphylococcus aureus is controlled by regulators such as the accessory gene regulator (agr). Strains can be divided into four major agr groups (agr I-IV) on the basis of agrD and agrC polymorphisms. The purpose of this study was to define the proportion of agr I, II, and III polymorphisms and to compare the clinical characteristics between group I and non-group I polymorphisms of methicillin-resistant Staphylococcus aureus (MRSA) strains in a Korean tertiary care teaching hospital. MATERIALS AND METHODS: A total of 158 clinical isolates were evaluated by RFLPs (restriction fragment length polymorphisms). RESULTS: The mean age of the patients was 50.2 +/- 21.9 years old. There were 74 (49.3%), 66 (44.0%), 10 (6.7%), 7 (4.4%), and 1 (0.6%) strains in agr group I, II, III, I + II, and I + III polymorphisms, respectively. Only ear infections were a statistically significant clinical parameter according to univariate (p=0.001) and multivariate analysis (OR, 4.721 (1.273-17.508), p=0.020). CONCLUSION: This study suggests that agr group I is the most prevalent in Korea, and ear infections are correlated with the group I polymorphism, which is a different clinical trend from western countries. It can also be inferred that community-acquired MRSA correlates with agr group I.ope

Nationenbildung und Definitionsmöglichkeiten der Griechen im kolonialen Ägypten (1856-1919)

Table of contents 5 1\. The “giant national tree” 9 1.1 The Greeks in Egypt 10 1.2 Egyptian Greeks beyond the legacy 13 1.3 A project of “making Greece abroad” 16 1.4 Method and decisions 20 First part: The “westernmost part of the Greek world” 29 2\. The “story of Ulysses”: the return to “success” stories about the Greeks in Egypt 30 2.1 The most familiar foreigner of Egypt 31 2.2 An example of failure 35 2.3 The Greek “cosmopolitans” 37 2.4 The “nice Greek adventure” in Egypt 46 2.5 Summary 50 3\. Spread “like olive oil”: Greeks dispersed in the African interior 51 3.1 The “real migrant hero” 52 3.2 A “link between Europe and Africa” 57 3.3 The “art of speculation” in “semi- savage countries” 63 3.4 A “necessary period of exile” 70 3.5 Summary 73 4\. The Greeks which “have not really been civilized yet”: the increase of Greek urban migrants in Egypt 74 4.1 A growing Greek population in Egypt 75 4.2 Beyond the “success” stories 80 4.3 The “unseemly” whites 85 4.4 Different forms of “disgrace” for the Greek prestige in Egypt 92 4.4 Summary 100 5\. A “monster” named “cosmopolitanism”: “cosmopolitan” Greeks and the West 102 5.1 Instruments “of enrichment or domination” 103 5.2 A “Cosmopolis” of the long 19th century 107 5.3 “Cosmopolitanism” as a threat to the nation 113 5.4 Summary 122 6\. “Lessons” exchanged across the Mediterranean: Greeks, Egypt and the British in the long 19th century 123 6.1 Opposing violence with violence 124 6.2 The post-1882 “order” 132 6.3 “European” and other states in the Mediterranean 138 6.4 The “first conqueror of Egypt” 146 6.5 Summary 153 7\. Going “without reason beyond the ocean”: Greek human mobility at the turn of the 20th century 154 7.1 Models of Greek human mobility 155 7.2 Attempts to control Greek human mobility 164 7.3 Migration and colonial hierarchies 171 7.4 Summary 180 Second part: A global Greek national history 181 8\. The “tangible things”: nation and nationality in colonial Egypt 182 8.1 The “manufacturers of citizenship” 183 8.2 “Just one of many social boundaries” 190 8.3 Summary 195 9\. A “Greek Church” in Egypt: religious beliefs and identification to the Greek nation 196 9.1 The “guardian of religion” and the “guardian of nation” 197 9.2 “Descendants of the prophets and citizens of our nation” 208 9.3 Choosing to “change worlds” 217 9.4 Summary 228 10\. A “convenient amnesia”: the Egyptian experience and the delimitation of the Greek “omogeneis” 229 10.1 A Greek homogenizing process in the long 19th century 230 10.2 “Sacrificing” the “Arabic-speaking Greeks” for tobacco 236 10.3 From “brotherly races” to neighbors “beyond comparison inferior” 246 10.4 The “localistic spirit” 262 10.5 Summary 271 11\. Training “proper” Greeks: an advanced Greek image of communion 273 11.1 The broadening of Greek education 275 11.2 The “manly, athletic upbringing” of the nation 287 11.3 A global Greek public 293 11.4 “Discussing” through the gun 299 11.5 Summary 304 12\. The Greek state and the “real Greece” in Egypt: a story of mutual stereotypes and expectations 305 12.1 The “Profit-making Hermes” 306 12.2 The “voice” of the “Greeks Abroad” 312 12.3 The Greek state and the migrants in Egypt 320 12.4 Summary 332 13\. The emergence of the “Egyptiots”: Greek responses to early 20th century evolutions in Egypt 334 13.1 Greeks and Egyptian nationalism 335 13.2 Overcoming the boundaries? 342 13.3 The “Egyptiots” and the Egyptians 350 13.4 Summary 355 14\. A giant global “tree” 356 Maps 362 Short biographical note 365 German summary 366 Catalogue of literature and sources 371At the turn of the 20th century, there were more than 100.000 people, which in various sources and with different criteria could be defined as Greeks of Egypt. Greeks of the Egyptian (and Sudanese) province constituted already in the middle of the 19th century a first current of Greek migration. A second large wave came by the end of the century and was concentrated mainly in the two large Egyptian cities, Alexandria and Cairo, or in the cities neighboring to the Suez Channel. Especially since the end of the 20th century, many texts reproduce an idealized description of the “Greek diaspora” as a “success story”, especially in the so-called “Greek East”. Such representations of the past and of the “diaspora”, composed by “cosmopolitan”, i.e. “European”, and “patriotic” at the same time Greeks, served among others as a model juxtaposed to aspects of contemporary Greek society. Nevertheless, a great part of these migrants were not “successful”, whatever might be meant by this description. Furthermore, the “Western” influence in certain aspects were not an exclusive characteristic of “cosmopolitan” Egypt, but rather an indication of the broadening of a specific type of education by the new urban middle-strata in places with a big number of Greeks, not just in the “Greek East”, but even in Greece. More than an unproblematic acceptance of the West among the Greeks of Egypt, both by them as also by Greeks elsewhere, there were many different ways of selecting, of adopting, of negotiating or of rejecting various aspects classified as “Western”. The question of defining Greeks as such in Egypt also constitutes a major issue in this dissertation. Nationality was a factor, but the issue of recognizing an individual as a citizen of Greece was constantly a cause of dispute for Greek and Egyptian authorities and also a matter of negotiation between migrants and the Greek authorities. A further challenge for a historian dealing with the formation of a Greek national community in Egypt is related to the fact that, in spite of the great national role ascribed to Orthodox Christian institutions in the course of the 19th century, also in Egypt, Greeks were not necessarily identified with Orthodox Christians. And even among the Orthodox Christians of Egypt, it was not always self-evident whether someone would be defined or accepted by others as a Greek. The process of forming a Greek national community was multidirectional and included homogenization, negotiations, adjustments and exclusions until a certain, monocultural version of the Greek nation prevailed, with a part of it living in Egypt.Am Anfang des 20. Jahrhunderts gab es mehr als 100.000 Leute, die in verschiedenen Quellen und mit unterschiedlichsten Kriterien als die Griechen Ägyptens beschrieben wurden. Die Griechen der ägyptischen (und sudanesischen) Provinz bildeten schon in der Mitte des 19. Jahrhunderts eine Migrationswelle. Eine zweite große Welle von griechischen Migranten kam am Ende des 19. Jahrhunderts und blieb meistens in den zwei größten ägyptischen Städten, Alexandria und Kairo, oder in den Städten in der Nähe des Suezkanals. Besonders seit dem Ende des 20. Jahrhunderts gibt es eine Reihe von Texten, die erneut einer idealisierten Beschreibung der „griechischen Diaspora“ als einer „Erfolgsgeschichte“, besonders im „griechischen Osten“, anheimfallen. Unter anderem dienten solche Darstellungen der Vergangenheit und der „Diaspora“, deren Mitglieder „kosmopolitisch“, d. h. „europäisch“, und gleichzeitig „patriotisch“ gewesen sein sollen, als ein Gegenmodell zu Aspekten der heutigen griechischen Gesellschaft. Eine große Menge der griechischen Migranten in Ägypten aber war nicht „erfolgreich“, egal was darunter verstanden wurde. Außerdem waren die „westlichen“ Einflüsse in bestimmten Bereichen des Lebens nicht ein exklusives Merkmal eines „kosmopolitischen“ Ägyptens, sondern ein Zeichen der Verbreitung einer bestimmten Bildung bei den neuen Mittelschichten in Städten mit einer großen Zahl von griechischen Einwohnern, nicht nur im „griechischen Osten“, sondern auch in Griechenland. Anstelle einer unproblematischen Aufnahme des Westens bei den Griechen Ägyptens gab es bei diesen genauso wie unter den Griechen von anderen Orten verschiedene Arten, „westliche“ Aspekte auszuwählen, zu übernehmen, zu verhandeln oder abzulehnen. Die Frage der Definitionsmöglichkeiten der Griechen Ägyptens ist in der vorliegenden Dissertation auch zentral. Nationalität war ein Faktor, wer aber als ein Bürger Griechenlands in Ägypten anerkannt wurde, war ein dauerhaftes Thema von Streitigkeiten mit den ägyptischen Behörden und der Verhandlung der Migranten mit den griechischen Behörden. Eine weitere Herausforderung für Historiker besteht darin, dass die Griechen trotz ihrer im Laufe des 19. Jahrhunderts verstärkt zugeschriebenen nationalen Rolle in orthodoxen christlichen Institutionen, auch in Ägypten, nicht unbedingt identisch mit den Orthodoxen Christen waren. Auch bei den Orthodoxen Christen Ägyptens war es aber nicht immer selbstverständlich, wer als ein Grieche verstanden werden konnte. Aus verschiedenen Versuchen, eine homogene griechische Nation auszubilden, hat sich eine bestimmte, monokulturelle Version durchgesetzt

Investigation Of Human Cancer Immune Interaction Using In Vitro Assays

Cancer immunotherapy, including immune checkpoint blocking antibodies are important components for treatment of patients with various types of cancer as they enhance the ability of the immune system to fight tumours. However, tumor cells have the ability to develop resistance to a variety of transitional therapies such as chemotherapy. In this study, in vitro Tumour-Immune co-culture system (TICS) has been developed to evaluate the impact on the antitumor activity of the primary human lymphocytes and response to PD-1 (nivolumab) and PD-L1 (durvalumab) checkpoint blocking antibodies against acquired chemotherapy resistance cancer cell lines. Using paired ovarian and neuroblastoma cancer cell lines obtained prior to chemotherapy (naïve) and after chemotherapy resistance, the results show that resistant ovarian cancer cells have differential effect on activation of lymphocytes and respond poorly to nivolumab and durvalumab, compared to chemotherapy naïve cells. On the other hand, chemotherapy neuroblastoma resistance cells show to respond to PD-1/L1 blockade therapy in TICS. Furthermore, blocking important molecular interactions between cancer cells and human lymphocytes such as HLA-ABC, HLA-DR and IFN-γ receptor compromises response to immune checkpoint blockade. In accordance, deletion of programmed death ligand 1 (PD-L1) on cancer cells by the CRISPR/Cas9 system significantly increases antitumor activity of immune cells in TICS. Moreover, deletion of beta-2-microglobulin (B2M) on human cancer cells resulted in substantial downregulation of HLA-ABC, which influenced immune activation induced by PD-1 blockade. Together, these findings demonstrate that chemotherapy resistance in human cancer cells could limit efficient response of PD-1/L1 blockade and thus immune checkpoint therapy could be more effective in early stage cancers

Epidemic methicillin-resistant Staphylococcus aureus (EMRSA) in Canada, a phenotypic and genotypic perspective

grantor: University of TorontoEpidemic MRSA (EMRSA) are resistant to multiple antibiotics, have the ability to spread easily within the hospital, and cross large geographic distances. We hypothesize that epidemicity may be related to a strain's ability to efficiently colonize its host. The accessory gene regulator locus ('agr') of 'S. aureus' plays a major role in a strain's ability to colonize and invade tissues. Hypervariability within the 'agr'BDC locus of 'S. aureus' strains represents a bacterial interference mechanism that could promote the ability of EMRSA to colonize in the presence of resident microflora. We therefore assayed a collection of epidemic and sporadic MRSA (SMRSA) based on factors expressed under the control of ' agr'. Three Canadian EMRSA clones A, B and C as typed by PFGE have been identified and consist of 10, 5 and 2 strain subtypes, respectively. Restriction fragment length polymorphism (RFLP) of the 'agr'BDC locus revealed that Clones A and C belonged to previously identified groups II and I, respectively, which was also most prevalent amongst the SMRSA strains. Clone B was assigned to a different 'agr' group (Ia) that showed a similar RFLP profile to 'agr' group I. A few significant nucleotide and amino acid sequence differences between 'agr'BDC groups I and Ia suggest that Ia may represent a novel 'agr' type rather than a minor variant of group I. (Abstract shortened by UMI.)M.Sc

Investigation Of Human Cancer Immune Interaction Using In Vitro Assays

Cancer immunotherapy, including immune checkpoint blocking antibodies are important components for treatment of patients with various types of cancer as they enhance the ability of the immune system to fight tumours. However, tumor cells have the ability to develop resistance to a variety of transitional therapies such as chemotherapy. In this study, in vitro Tumour-Immune co-culture system (TICS) has been developed to evaluate the impact on the antitumor activity of the primary human lymphocytes and response to PD-1 (nivolumab) and PD-L1 (durvalumab) checkpoint blocking antibodies against acquired chemotherapy resistance cancer cell lines. Using paired ovarian and neuroblastoma cancer cell lines obtained prior to chemotherapy (naïve) and after chemotherapy resistance, the results show that resistant ovarian cancer cells have differential effect on activation of lymphocytes and respond poorly to nivolumab and durvalumab, compared to chemotherapy naïve cells. On the other hand, chemotherapy neuroblastoma resistance cells show to respond to PD-1/L1 blockade therapy in TICS. Furthermore, blocking important molecular interactions between cancer cells and human lymphocytes such as HLA-ABC, HLA-DR and IFN-γ receptor compromises response to immune checkpoint blockade. In accordance, deletion of programmed death ligand 1 (PD-L1) on cancer cells by the CRISPR/Cas9 system significantly increases antitumor activity of immune cells in TICS. Moreover, deletion of beta-2-microglobulin (B2M) on human cancer cells resulted in substantial downregulation of HLA-ABC, which influenced immune activation induced by PD-1 blockade. Together, these findings demonstrate that chemotherapy resistance in human cancer cells could limit efficient response of PD-1/L1 blockade and thus immune checkpoint therapy could be more effective in early stage cancers