Molecular genetics of the immotile short tail sperm defect

Hedelmättömyyttä aiheuttavan siittiöiden puolihäntävian molekyyligenetiikka Suomalaisissa Yorkshire karjuissa yleistyi 1990-luvun lopulla autosomaalisesti ja resessiivisesti periytyvä hedelmättömyyttä aiheuttava siittiöiden puolihäntävika (ISTS, immotile short tail sperm). Sairaus aiheuttaa normaalia lyhyemmän ja täysin liikkumattoman siittiön hännän muodostuksen. Muita oireita sairailla karjuilla ei ole havaittu ja emakot ovat oireettomia. Tämän tutkimuksen tarkoituksena oli kartoittaa siittiöiden puolihäntävian aiheuttava geenivirhe ja kehittää DNA-testi markkeri- ja geeniavusteiseen valintaan. Koko genomin kartoituksessa vian aiheuttava alue paikannettiin sian kromosomiin 16. Paikannuksen perusteella kahden geenimerkin haplotyyppi kehitettiin käytettäväksi markkeri-avusteisessa valinnassa. Sairauteen kytkeytyneen alueen hienokartoitusta jatkettiin geenitestin kehittämiseksi kantajadiagnostiikkaan. Vertailevalla kartoituksella oireeseen kytkeytynyt alue paikannettiin 2 cM:n alueelle ihmisen kromosomiin viisi (5p13.2). Tällä alueella sijaitsevia geenejä vastaavista sian sekvensseistä löydetyn muuntelun perusteella voitiin tarkentaa sairauteen kytkeytyneitä haplotyyppejä. Haplotyyppien perusteella puolihäntäoireeseen kytkeytynyt alue rajattiin kahdeksan geenin alueelle ihmisen geenikartalla. Alueelle paikannetun kandidaattigeenin (KPL2) sekvensointi paljasti introniin liittyneen liikkuvan DNA-sekvenssin, Line-1 retroposonin. Tämä retroposoni muuttaa geenin silmikointia siten, että sitä edeltävä eksoni jätetään pois tai myös osa introni- ja inserttisekvenssiä liitetään geenin mRNA tuotteeseen. Molemmissa tapauksissa tuloksena on lyhentynyt KPL2 proteiini. Tähän retroposoni-inserttiin perustuva geenitesti on ollut sianjalostajien käytössä vuodesta 2006. KPL2 geenin ilmenemisen tarkastelu sialla ja hiirellä paljasti useita kudosspesifisiä silmikointimuotoja. KPL2 geenin pitkä muoto ilmenee pääasiassa vain kiveksessä, mikä selittää geenivirheen aiheuttamat erityisesti siittiön kehitykseen liittyvät oireet. KPL2 proteiinin ilmeneminen hiiren siittiön hännän kehityksen aikana ja mahdollinen yhteistoiminta IFT20 proteiinin kanssa viittaavat tehtävään proteiinien kuljetuksessa siittiön häntään. Mahdollisen kuljetustehtävän lisäksi KPL2 saattaa toimia myös siittiön hännän rakenneosana, koska se paikannettiin valmiin siittiön hännän keskiosaan. Lisäksi KPL2 proteiini saattaa myös toimia Golgin laitteessa sekä Sertolin solujen ja spermatidien liitoksissa, mutta nämä havainnot kuitenkin vaativat lisätutkimuksia. Tämän tutkimuksen tulokset osoittavat, että KPL2 geeni on tärkeä siittiön hännän kehitykselle ja sen rakennemuutos aiheuttaa siittiöiden puolihäntäoireen suomalaisilla Yorkshire karjuilla. KPL2 proteiinin ilmeneminen ja paikannus siittiön kehityksen aikana antaa viitteitä proteiinin toiminnasta. Koska KPL2 geenisekvenssi on erittäin konservoitunut, nämä tulokset tuovat uutta tietoa kaikkien nisäkkäiden siittiöiden kehitykseen ja urosten hedelmättömyyteen syihin.

Design and production of protein nanostructures for biomolecular detection

Particulate nanostructures are increasingly used for analytical purposes. Such particles are often generated by chemical synthesis from non-renewable raw materials. Generation of uniform nanoscale particles is challenging and particle surfaces must be modified to make the particles biocompatible and water-soluble. Usually nanoparticles are functionalized with binding molecules (e.g., antibodies or their fragments) and a label substance (if needed). Overall, producing nanoparticles for use in bioaffinity assays is a multistep process requiring several manufacturing and purification steps. This study describes a biological method of generating functionalized protein-based nanoparticles with specific binding activity on the particle surface and label activity inside the particles. Traditional chemical bioconjugation of the particle and specific binding molecules is replaced with genetic fusion of the binding molecule gene and particle backbone gene. The entity of the particle shell and binding moieties are synthesized from generic raw materials by bacteria, and fermentation is combined with a simple purification method based on inclusion bodies. The label activity is introduced during the purification. The process results in particles that are ready-to-use as reagents in bioaffinity. Apoferritin was used as particle body and the system was demonstrated using three different binding moieties: a small protein, a peptide and a single chain Fv antibody fragment that represents a complex protein including disulfide bridge.If needed, Eu3+ was used as label substance. The results showed that production system resulted in pure protein preparations, and the particles were of homogeneous size when visualized with transmission electron microscopy. Passively introduced label was stably associated with the particles, and binding molecules genetically fused to the particle specifically bound target molecules. Functionality of the particles in bioaffinity assays were successfully demonstrated with two types of assays; as labels and in particle-enhanced agglutination assay. This biological production procedure features many advantages that make the process especially suited for applications that have frequent and recurring requirements for homogeneous functional particles. The production process of ready, functional and watersoluble particles follows principles of “green chemistry”, is upscalable, fast and cost-effective.

Foodstuffs and medicines as legal categories in the EU and China. Functional foods as a borderline case

The thesis discusses the regulation of foodstuffs and medicines, and particularly the regulation of functional foods. Legal systems investigated are the EU and China. Both are members of the WTO and Codex Alimentarius, which binds European and Chinese rules together. The study uses three Chinese berries as case examples of how product development faces regulation in practice. The berries have traditional uses as herbal medicines. Europe and China have similar nutrition problems to be resolved, such as obesity, cardiovascular disease, and diabetes. The three berries might be suitable raw materials for functional foods. Consumer products with health-enhancing functions, such as lowering blood pressure, might legally be classifi ed either as foodstuffs or medicines. The classifi cation will depend on functions and presentation of the product. In our opinion, food and medicine regulation should come closer together so the classifi cation issue would no longer be an issue. Safety of both foodstuffs and medicines is strictly regulated. With medicines, safety is a more relative concept, where benefi ts of the product are compared to side-effects in thorough scientifi c tests and trials. Foods, on the other hand, are not allowed to have side-effects. Hygiene rules and rules on the use of chemicals apply. In China, food safety is currently at focus as China has had several severe food scandals. Newly developed foods are called novel foods, and are specifi cally regulated. The current European novel food regulation from 1997 treats traditional third country products as novel. The Chinese regulation of 2007 also defi nes novel foods as something unfamiliar to a Chinese consumer. The concepts of novel food thus serve a protectionist purpose. As regards marketing, foods are allowed to bear health claims, whereas medicines bear medicinal claims. The separation is legally strict: foods are not to be presented as having medicinal functions. European nutrition and health claim regulation exists since 2006. China also has its regulation on health foods, listing the permitted claims and how to substantiate them. Health claims are allowed only on health foods. The European rules on medicines include separate categories for herbal medicines, traditional herbal medicines, and homeopathic medicines, where there are differing requirements for scientifi c substantiation. The scientifi c and political grounds for the separate categories provoke criticism. At surface, the Chinese legal system seems similar to the European one. To facilitate trade, China has enacted modern laws. Laws are needed as the country moves from planned economy to market economy: ‘rule of law’ needs to replace ‘rule of man’. Instead of being citizens, Chinese people long were subordinates to the Emperor. Confucius himself advised to avoid confl ict. Still, Chinese people do not and cannot always trust the legal system, as laws are enforced in an inconsistent manner, and courts are weak. In China, there have been problems with confl icting national and local laws. In Europe, the competence of the EU vs. the competence of the Member States is still not resolved, even though the European Commission often states that free trade requires harmonisation. Food and medicine regulation is created by international organisations, food and medicine control agencies, standards agencies, companies and their organisations. Regulation can be divided in ‘hard law’ and ‘soft law’. One might claim that hard law is in crisis, as soft law is gaining importance. If law is out of fashion, regulation certainly isn’t. In the future, ‘law’ might mean a process where rules and incentives are created by states, NGOs, companies, consumers, and other stakeholders. ‘Law’ might thus refer to a constant negotiation between public and private actors. Legal principles such as transparency, equal treatment, and the right to be heard would still be important.

Conversioni al cattolicesimo a Roma tra Sei e Settecento. La presenza degli scandinavi nell’Ospizio dei Convertendi.

In my PhD dissertation, I have examined a group of people of Scandinavian origin received by Ospizio dei Convertendi. This group has been hitherto largely unknown to historical research. The Ospizio was an institute founded by the Oratorian Congregation in Rome in 1673 to provide religious instruction and material aid to both recent and aspirant converts to Roman Catholicism. My research traces the profile of converts and a typology of motives, examining different factors which influenced the conversion process. I show that the key factors were often of a social rather than a religious nature. Moreover, I have analyzed the hospice in the context of Counter-Reformation charity as well. In terms of numbers, the Scandinavians formed a somewhat marginal yet not insignificant group within the Roman hospice. Out of a total of 2203 guests received between 1673 and 1706, 4.6 % were Scandinavians: 74 Swedes (including Finland and Livonia) and 27 Danes (including Norway). They came from a rigorously Protestant region which reacted to Catholicism with severe legislative measures. Converts to Catholicism risked confiscation of their goods, expulsion or even capital punishment. Since both Sweden and Denmark were practically impenetrable to Catholicism at the time and clandestine missionary attempts often failed before they had even properly started, the Roman Catholic Church shifted its interest towards Northerners arriving in Rome, a preferred destination for young noblemen, artists and migrant craftsmen. The material related to Ospizio dei Convertendi, conserved in the Vatican archives, is a scarcely known yet unusually rich source, not only for the religious history of our continent, but also for social history and the study of migration in early modern Europe. It contains a wealth of information about members of the subordinate classes, of their travels and lives in Europe. The profile delineated in these documents is of individuals who had a wide range of different professions and different aspirations. These documents encompass a vast social spectrum that was highly mobile on a continent which by that time had become pluriconfessional. Therefore, these migrants faced the complex religious reality in their everyday life. The principal corpus of my research consists of two types of manuscript sources created for administrative and in a way also for apologetic purposes of the Roman Catholic Church. My starting point is the Primo registro generale of Ospizio dei Convertendi. This is a volume in which the following information about each guest was registered: name, nationality, city of origin, age, sex, profession, confession professed before converting, date of arrival, departure, abjuration and baptism. Typically, the convert was male, originating from Stockholm or Copenhagen, from 21 to 30 years of age. The biggest occupational groups in descending order were soldiers, noblemen, craftsmen and sailors. Thus the data reflects a multiform reality of interurban and long distance migration, ideals regarding the education of young noblemen and gentry as well as the need of European armies to hire foreign mercenaries in their various campaigns. Against this background the almost total absence of women is hardly surprising: there is only one woman in the material I have studied. The second main source, Nota degl’ospiti ricevuti e spese fatte per essi, sheds more light on the choices of the converts, their motivations and their lives outside Scandinavia before reaching Rome. This narrative material permits an analysis which completes but also goes far beyond the columns of the Institute’s general register. This material consists of reports written by Catholic priests based on an interview conducted upon each guest’s arrival. The material frequently includes information on what the converts would do following their departure from the Institute as well. These sources have a specific narrative form and contain short biographies, list reasons for converting and information about the journey from the North to the Mediterranean - a journey which in many cases took several years. Moreover, they show that certain unorthodox practices such as calling on the saints and pleading for help from them were not uncommon in the Protestant popular religion. The recording of information on conversions from Protestantism to Catholicism reflects both religious and social interest on the part of the receiving institute. The information obtained was used for the purposes of religious teaching, for finding adequate ways of inserting the convert into Italian society so that he could earn a living, and to find effective methods to convert others with a similar cultural and geographical background. The stories recorded were based on interviews with the newly-arrived, information obtained from a travel companion or fellow countrymen, or from written documents the aspirant converts carried with them. These sources illustrate, although sometimes in rather simplified ways, the circumstances and motivations which were relevant to the choice of changing one’s confession. In addition, I have examined petitions addressed to the hospice and other Roman authorities in order to get financial aid. These petitions were written by Italian scrittori, and they contain certain conventions and topoi of presenting the conversion with the purpose of improving the chances of obtaining financial aid. It is through these filters, which may seem initially almost invisible, that the remote voice of the converts reaches us. The results of the analysis are particularly interesting because they disagree with some of the principal conclusions of previous work on the subject. First, earlier research has focused almost exclusively on the conversions of noblemen, and has argued, second, that the Queen Christina of Sweden was the driving force behind their change of confession. The sources examined for this dissertation present a profile of long-distance migrants, many of them members of the subordinate classes, who were looking for ways to make their living in Europe. These people had in many cases left their country of origin several years earlier and not for religious reasons, so, crucially, we are not dealing with confessional migration in these cases. Rather, conversion was a complex process, intricately tied up with strategies of survival, integration and upward social mobility. At the same time, while these components are significant on their own right, they do not necessarily point to the absence of motivations of a more clearly religious nature.