Untersuchung der Invasivität von primären Endometriosezellen

Background and aims Endometriosis is a chronic disease characterized by the presence of endome-trial tissue outside the uterus. Patients suffer of different kind of pain and com-pletely mixed dissemination of endometriosis. Due to heterogeneous clinical characteristics there exist multiple therapeutic options. Therefore the aim of the Clinic-meets-invasion-study (CMI-study) was to estab-lish a correlation between the most important clinical features of endometriosis (disease extent and pain levels) and primary cell lines, which were generated from the patients with the respective clinical characteristics. Methods This prospective study was designed as a 2 x 2 factorial case control study with patients having high pain levels vs. low pain levels (factor 1) and patients with a high degree of disease dissemination vs. a low degree of disease dissemination (factor 2). This design should result in 20 patients for each comparison and a total of 4 unique patient groups. For an explorative analysis 2 patient groups were added during the course of the study. Each group containing 5 patients with deep infiltrating endometriosis. One with lesions ≥ 5mm and ≤ 1cm diameter and the other with lesions ≥ 2 cm diameter. Patients were screened for the 6 groups at the Department of Gynecology and Obstetrics of the University Hospital in Erlangen. The pain level from each patient was determined with a standardized questionnaire. Endometriosis tissue of the fossa ovarica was obtained during schedueled laparoscopy and from each tissue endometriosis cells were isolated, cultivated and cell invasion was analyzed in a vertical gel invasion assay.   Results and observations No correlation was observed between the pain level and invasion depth in the invasion assay – although the cells of patients with no / lower pain invaded tendentially deeper into the gel than cells from patients with high pain. Comparing the cells of patients with dissemination vs those without dissemination, those with dissemination invaded tendentially deeper into the gel. This was especially prominent for the mean invasion of the median of gel invasion. There was a tendency of more of the not deep infiltrating endometriosis cells invading deeper into the gel than of those cell lines with deep infiltrating endometriosis. Conclusions The question arises whether it makes a difference for the endometriosis, in vivo, if more cells invade less deeply, or if fewer cells invade deeper. A tendency was observed in the CMI-study. More cells of patients with no/ low pain did invade into the gel, but less deep. Less cells of patients with high pain did invade into the gel, but deeper. Equally a tendency for deeper invasion was observed for cells of patients with high dissemination. The induction of epithelial-mesenchymal transition is possibly a process that could explain the mobility and invasive behavior of the endometriosis cells. Collective cell migration promotes potentially the invasion of deep infiltrating endometriosis. Perhaps due to a lack of factors promoting collective cell migra-tion or because of a lower number of cells in total the deep infiltrating cell lines do invade less deeply into the gel as the not deep infiltrating cell lines.Hintergrund und Ziele Unter dem Krankheitsbild der Endometriose versteht man das Vorkommen von endometriumartigen Zellverbänden außerhalb des cavum uteri. In der Clinic-meets-Invasion-Studie (CMI-Studie) sollte nun erstmalig, unter Verwendung von unterschiedlichen primären Endometriosezelllinien, im in vitro Zellinvasions-Assay untersucht werden, ob ein Zusammenhang zwischen den wichtigsten klinischen Merkmalen der Endometriose, dem Schmerz und der anatomischen Ausdehnung der Endometrioseherde, und dem Invasionsverhalten der Zellen besteht. Die aus den Versuchen gezogenen Erkenntnisse sollten unmittelbare Rück-schlüsse für die Entwicklung von Therapien gegen Endometriose ermöglichen. Methoden In dieser prospektiven Kohortenstudie der Frauenklinik des Universitätsklinikum in Erlangen wurden Patientinnen im 2 x 2 faktoriellen Design mit einem hohen bzw. niedrigen Schmerzlevel (Faktor 1) und Patientinnen mit anatomisch ausgedehnter bzw. anatomisch nicht ausgedehnter Endometriose (Faktor 2) eingeschleust. Jede der 4 resultierenden Gruppen sollte 5 Patientinnen umfassen. Der Schmerzwert jeder Patientin wurde mit Hilfe eines standardisierten Fragebogens ermittelt. Für eine zusätzliche explorative Analyse wurden im Verlauf der Studie weitere 2 Gruppen hinzugefügt. Dabei handelte es sich um insgesamt 10 Patientinnen mit tief infiltrierenden Endometrioseherden. 5 Patientinnen davon mit Läsionen ≥ 5mm und ≤ 1cm Durchmesser und 5 Patientinnen mit Läsionen ≥ 2 cm Durchmesser. Allen Patientinnen wurden an der Frauenklinik des Universitätsklinikums Erlangen intraoperativ Gewebeproben aus der Fossa ovarica entnommen. Anschließend wurden aus den Proben Endometriosezellen isoliert, kultiviert und in einem anschließenden Vertical Gel Invasion Assay das Invasionsverhalten der Zellen untersucht.   Ergebnisse und Beobachtungen Im Invasions Assay ergab sich keine positive Korrelation zwischen den Schmerzwerten und der Invasionstiefe der Zellen in das Kollagen. Es konnte jedoch eine Tendenz dahingehend beobachtet werden, dass mehr Zelllinien der Gruppe „kein/ wenig Schmerz“ mit ausgedehnter Endometriose tiefer in das Gel einwandern als Zellen der Gruppe „viel Schmerz“ mit nicht ausgedehnter Endometriose. Bei den tief infiltrierenden Endometrioseherden konnte beobachtet werden, dass tendenziell mehr Zelllinien der nicht tief infiltrierenden Endometrioseherde tiefer invadierten als die Zelllinien der tief infiltrierenden Endometrioseherde. Schlussfolgerung Es stellt sich die Frage, ob es in vivo für die Krankheit einen Unterschied macht, wenn mehr Zellen weniger tief einwandern oder wenn weniger Zellen tiefer einwandern. Bei der CMI-Studie ließ sich eine Tendenz erkennen. Bei den Patientinnen mit kein/ wenig Schmerz wanderten im Invasions Assay mehr Zellen ins Gel ein, aber dafür weniger tief. Bei den Patientinnen mit viel Schmerz, wanderten weniger Zellen ins Gel ein, dafür aber tiefer. Die Induktion der epithelial-mesenchymalen Transition ist möglicherweise ein Vorgang, der die Mobilität und das invasive Verhalten der Endometriosezellen erklären könnte. Bei der Invasion der tief infiltrierenden Endometriose fördert möglicherweise die kollektive Zellwanderung die Invasion. Womöglich wanderten deshalb in dem CMI Versuchsansatz die tief infiltrierenden Zelllinien weniger tief in das Gel ein als die nicht tief infiltrierenden, weil deren absolute Zellzahl vergleichsweise geringer war oder es an Faktoren mangelte, die die kollektive Zellwanderung fördern. Aufgrund der geringen Fallzahl in der CMI-Studie konnte nur eine einge-schränkte statistische Auswertung erfolgen. Es wäre daher sinnvoll in Zukunft Studien mit höheren Fallzahlen durchzuführen um sicher zu gehen, dass ge-samte Krankheitsbild zu erfassen und um eine umfassende statistische Auswertung zu gewährleisten

Immunization with a DNA vaccine cocktail encoding TgPF, TgROP16, TgROP18, TgMIC6, and TgCDPK3 genes protects mice against chronic toxoplasmosis

Toxoplasmosis is a zoonotic disease caused by the intracellular protozoan Toxoplasma gondii; and a major source of infection in humans is via ingestion of T. gondii tissue cysts. Ultimately, the goal of anti-toxoplasmosis vaccines is to elicit a sustainable immune response, capable of preventing formation of the parasite tissue cysts—or, at least, to restrain its growth. In this study, we formulated a cocktail DNA vaccine and investigated its immunologic efficacy as a protection against the establishment of T. gondii cysts in the mouse brain. This multicomponent DNA vaccine, encoded the TgPF, TgROP16, TgROP18, TgMIC6, and TgCDPK3 genes, which play key roles in the pathogenesis of T. gondii infection. Results showed that mice immunized via intramuscular injection three times, at 2-week intervals with this multicomponent DNA vaccine, mounted a strong humoral and cellular immune response, indicated by significantly high levels of total IgG, CD4+ and CD8+ T lymphocytes, and antigen-specific lymphocyte proliferation when compared with non-immunized mice. Immunization also induced a mixed Th1/Th2 response, with a slightly elevated IgG2a to IgG1 ratio. The increased production of proinflammatory cytokines gamma-interferon, interleukin-2, and interleukin-12 (p 0.05). The number of brain cysts in immunized mice was significantly less than those in non-immunized mice (643.33 ± 89.63 versus 3,244.33 ± 96.42, p < 0.0001), resulting in an 80.22% reduction in the parasite cyst burden. These findings indicate that a multicomponent DNA vaccine, encoding TgPF, TgROP16, TgROP18, TgMIC6, and TgCDPK3 genes, shows promise as an immunization strategy against chronic toxoplasmosis in mice, and calls for a further evaluation in food-producing animals

Fine-Scale Mapping of the 4q24 Locus Identifies Two Independent Loci Associated with Breast Cancer Risk

Background: A recent association study identified a common variant (rs9790517) at 4q24 to be associated with breast cancer risk. Independent association signals and potential functional variants in this locus have not been explored. Methods: We conducted a fine-mapping analysis in 55,540 breast cancer cases and 51,168 controls from the Breast Cancer Association Consortium. Results: Conditional analyses identified two independent association signals among women of European ancestry, represented by rs9790517 [conditional P = 2.51 × 10−4; OR, 1.04; 95% confidence interval (CI), 1.02–1.07] and rs77928427 (P = 1.86 × 10−4; OR, 1.04; 95% CI, 1.02–1.07). Functional annotation using data from the Encyclopedia of DNA Elements (ENCODE) project revealed two putative functional variants, rs62331150 and rs73838678 in linkage disequilibrium (LD) with rs9790517 (r2 ≥ 0.90) residing in the active promoter or enhancer, respectively, of the nearest gene, TET2. Both variants are located in DNase I hypersensitivity and transcription factor–binding sites. Using data from both The Cancer Genome Atlas (TCGA) and Molecular Taxonomy of Breast Cancer International Consortium (METABRIC), we showed that rs62331150 was associated with level of expression of TET2 in breast normal and tumor tissue. Conclusion: Our study identified two independent association signals at 4q24 in relation to breast cancer risk and suggested that observed association in this locus may be mediated through the regulation of TET2. Impact: Fine-mapping study with large sample size warranted for identification of independent loci for breast cancer risk

TheDebt of Love in the Theology of Richard of St. Victor:

Thesis advisor: Boyd Taylor CoolmanRichard of St. Victor is known primarily for two arguments from his book On the Trinity. The first is that the relationship between the Father, Son, and Holy Spirit can be understood from the logic of love, which tends toward sharing with others. The second is that a divine person (persona) is an “individual existence of a divine nature,” a revision of the definition of personhood given centuries earlier by Boethius. Fewer contemporary theologians are familiar with the third major innovation Richard makes in this work. In his description of love among the trinitarian persons, Richard organizes the three loving divine persons according to three modes of loving: gratuitous love (amor gratuitus) identical with the Father, owed love (amor debitus) identical with the Holy Spirit, and love mixed from both (amor permixtus) identical with the Son. The most striking—because the most apparently problematic—part of this scheme is the nomination of amor debitus, owed love, as a mode of loving proper to the Trinity. Presenting the apparently contradictory combination of owing and loving as a love proper to God as God, the conundra raised by Richard’s use of this phrase were interpreted away by his scholastic readers in a way that buried the potential of his insight for discussions about the nature of love called divine. My dissertation analyzes the meaning of amor debitus to reframe and re-present Richard’s arguments in On the Trinity with Richard’s radical claim about the nature of intra-trinitarian love in clearer view. I argue that the notion of owed love represents a class of experience which is distinct both from the obligation of a command (typically associated with debitum) and from the freedom of an unaffected choice (typically associated with amor). Amor debitus fuses the quality of the command or commanding conditions with a freedom that is not intrinsic to these prior conditions, but does not leave them behind. Richard’s link between owed love as a personal property in the Trinity and as the norm of human love for God gives Richard a fresh approach to the idea that “God’s love has been poured out into our hearts through the Holy Spirit” (Rom. 5:5), and therefore also to the longstanding Christian claim that human love is owed to God, ordered to neighborly service, and free.Thesis (PhD) — Boston College, 2024.Submitted to: Boston College. Graduate School of Arts and Sciences.Discipline: Theology

Method and study of the stresses around an opening under concentrated static loads using Stresscoat, A

Includes bibliographical references

Local and Peripheral Cues Driving Heterogeneous Populations of Brain-Infiltrating T Cells During Chronic Toxoplasma gondii Infection

T cells form a fundamental component of the protective response to chronic Toxoplasma gondii infection. The dire consequences of a failed T cell response in the protective response to this parasite are particularly apparent in AIDS patients, yet there must be a highly coordinated balance to the T cell response in the CNS. First, there must be a balance between pro-inflammatory and anti-inflammatory responses. Next, in a context of persistent antigen there must be a balance between the function of memory and effector subsets that simultaneously occupy the same niche. Finally, the T cell response must adapt to local conditions within the tissue. This dissertation explores these phenomena in the following ways:In Chapter 1, we discuss what is known about the T cell response to Toxoplasma gondii infection. We explore the signals that determine infiltration into the brain, behavior within the brain, and phenotypic diversity. In Chapter 2, we discuss the role of tissue resident memory T cells in the protective response to chronic Toxoplasma gondii infection. Here we use RNA-Seq to determine that the distinct transcriptional profile of brain infiltrating CD103+ CD8 T cells is consistent with tissue resident memory T cells, or TRM. We then use re-stimulation assays to establish that this population has a significantly increased capacity to produce pro-inflammatory cytokines as compared to other populations within the brain. This data supports the presence of a functional population of memory T cells in the response to chronic Toxoplasma gondii infection.In Chapter 3, we examine the requirement of lymph node homing dendritic cells expressing CCR7 for maintenance of effector T cells within the brain. This supports the requirement for ongoing recruitment of brain-infiltrating effector T cells. In Chapter 4 we examine how glutamate dysregulation within the CNS during chronic T. gondii infection impacts cytokine production and proliferation in activated T cells. We conclude in Chapter 5 by examining the common themes that emerge from the three primary chapters of the dissertation

Elizabeth Barrett Browning on the Death of Lord Byron

Elizabeth Barrett BrowningGeorge Gordon Lord Byron"Stanzas on the Death of Lord Byron"Romantic Poetry (ENG 4354