In this study, we aimed to determine the levels of cytokines IL-1β, IL-4, IL-10, IFN-γ, MIF and VEGF in tears derived from healthy subjects. We tested cytokines as potential markers of inflammation for their potential use in clinical practice. Having reliable method for measuring cytokine levels in tears would enable an early diagnosis of eye diseases. In two phases, cytokines in tears of healthy individuals were analyzed using Bio-Plex Cytokine Assay (Bio-Rad). We assessed the suitability of methods for diagnostic purposes as well as the suitability of our selected cytokines. Statistically significant positive correlations of cytokines were confirmed: IL-10 with IFN-γ (r = 0,81), MIF with VEGF (r = 0,42 / r = 0,49), IL-1β with IL-10 (r = 0,52), IL-1β with IFN-γ (r = 0,55), IL-1β with VEGF (r = 0,38), IFN-γ with VEGF (r = 0,45) and IL-4 with VEGF (r = 0,48) in healthy subjects in tears. IL-4 (r = -0,37) and IFN-γ (r = -0,42) correlate negatively with age. In healthy individuals, there seem to be no differences with regard to gender, BMI, body fat, time of meal consumption prior to tear collection, eye strain when using a computer, dry eyes. Thus, studied cytokines are suitable for diagnostic purposes. Significant differences in concentrations of four (IL-1β, IL-10, IFN-γ a VEGF) of the five...

Indiana University-Purdue University Indianapolis (IUPUI) / Background: Bacterial adherence to the acquired dental pellicle, important in caries, is mediated by receptor-adhesin interactions such as Streptococcus mutans antigen I/II (I/II). Ten I/II epitopes from the A, V, P and C regions were chosen to determine their reactivity in human saliva. Underlying the body’s ability to immunologically respond to bacteria that lead to caries are the human leukocyte antigen (HLA) genes, specifically HLA class II (HLA-II) genes that control antigen presentation. Previous studies suggested that a specific HLA biomarker group (HLA-DRB1*04) may have differential control of immune responses to I/II. However, it was not known whether secretory IgA (SIgA) responses to the selected epitopes from HLA-DRB1*04 positive subjects were different compared to their non-biomarker counterparts (negative), or across other caries factors, since no study to date had thus assessed these questions. Methods: Per IRB approval, the study population was divided into age, sex and race matched DRB1*04 positive (n=16) and negative groups (n=16). SIgA-epitope (and whole cell) reactivity was determined using ELISA. Other caries factors were measured. Subjects received a clinical exam by a trained examiner. ix Differences between DRB1*04 positive and negative groups were examined using a two-sided, two-sample t-test. Results: DRB1*04 positive subjects had numerically, but not statistically, higher reactivity to 9 out of 10 epitopes, the exception being residues 834-853 from the V and P regions of I/II across multiple measures. Though statistically insignificant, DRB1*04 positive subjects also exhibited 25-30 μg mL-1 less total IgA (TIgA) than negative counterparts. All clinical caries data proved inconclusive when comparing groups, likely due to exogenous factors and sample size. Conclusion: DRB1*04 positive subjects showed a trend toward lower TIgA. Moreover, they also showed a lower SIgA response across multiple measures to 834-853, the I/II V and P region epitope. This region forms a sort of functional epicenter involved in collaboration between domains along the entire I/II antigen, and governs the region involved in initial attachment to the acquired dental pellicle. This region may be involved in an in vivo discontinuous conformationally specific immunogenic epitope that serves as an HLA-II binding motif which remains elusive.

Recent studies have revealed extensive cortical demyelination in patients with progressive multiple sclerosis (MS). Demyelination in gray matter lesions is associated with activation of microglia. Macrophages and microglia are known to express myeloperoxidase (MPO) and generate reactive oxygen species during myelin phagocytosis in the white matter. In the present study we examined the extent of microglial activation in the cerebral cortex and the relationship of microglial activation and MPO activity to cortical demyelination. Twenty-one cases of neuropathologically confirmed multiple sclerosis, with 34 cortical lesions, were used to assess microglial activation. HLA-DR immunolabeling of activated microglia was significantly higher in demyelinated MS cortex than control cortex and, within the MS cohort, was significantly greater within cortical lesions than in matched non-demyelinated areas of cortex. In homogenates of MS cortex, cortical demyelination was associated with significantly elevated MPO activity. Immunohistochemistry revealed MPO in CD68-positive microglia within cortical plaques, particularly toward the edge of the plaques, but not in microglia in adjacent non-demyelinated cortex. Cortical demyelination in MS is associated with increased activity of MPO, which is expressed by a CD68-positive subset of activated microglia, suggesting that microglial production of reactive oxygen species is likely to be involved in cortical demyelination.

Die Arbeit beschreibt das phänotypische Verhalten von kultivierten Langerhanszellen, antigenpräsentierenden Zellen der Epidermis, sowie die Art und Weise ihrer Elimination. Hierfür wurden Zellkulturen von Langerhanszellen durch Migration aus normaler menschlicher Haut gewonnen. Die Langerhanszellen durchlaufen dabei die gleiche funktionelle Entwicklung, wie nach Antigenpräsentation in situ. Ziel der Untersuchung war es, die funktionellen und zellulären Eigenschaften und die Elimination von Langerhanszellen in der Zellkultur zu ermitteln. Die Anzahl viabler Zellen wurde mittels Trypanblauausschluß zu verschiedenen Zeitpunkten der Kultur ermittelt. Außerdem wurden die Zellen mittels Elektronenmikroskopie und Immuncytochemie untersucht. Die Befunde zeigen, daß die Zellen in der Kultur eine Veränderung ihres Phänotyps sowie funktionelle Änderungen im Sinne einer Ausreifung zu antigenpräsentierenden, T-Zell stimulierenden Zellen erfahren. Das in-vitro-Verhalten entspricht dem von Langerhanszellen in vivo nach Kontaktsensibilisierung. Mit Hilfe von eines für Apoptose spezifischen ELISA (= Enzyme-linked immunosorbent assay: eine Nachweisreaktion für Antigene bzw. Antikörper mithilfe von Enzymen) und Elektronenmikroskopie wurde nachgewiesen, daß die Zellen in unseren Kulturen durch Apoptose starben. Es gibt keinen Anhaltspunkt dafür, daß sich die Zellen nicht auch in vivo apoptotisch eliminieren. Die Verlauf der Funktionsmarker weist darauf hin, daß vorwiegend die maturierten Zellen von Apoptose betroffen waren, und die Apoptose über das CD 95/CD 95 L- System gesteuert wurde. Die Versuche zeigten insgesamt, daß Langerhanszellen durch Apoptose aus der Kultur eliminiert werden. Da sich die Zellen nach Migration in vitro wie Langerhanszellen nach Antigenpräsentation in vivo verhalten, scheint die Apoptose ein biologisches Regulativ für die Elimination von funktionell ausgereiften Langerhanszellen darzustellen. / This work describes the phenotypic behavior of cultivated Langerhans-cells, epidermal cells presenting antigenes and how they are eliminated . Therefore cultures of Langerhans-cells won by migration from normal human skin were used. The migrated Langerhans-cells have the same phenotypic features as Langerhans-cells after presentation of antigenes in situ. The aim of this work was to show the functional and cellular features of Langerhans-cells in culture and the way of their elimination. The cells still alive were count at distinct times using the Trypan-blue-exclusion-method. Additionally the cells were examined by electron microscopy and immuncytochemical methods. The findings show, that the cells in culture have the same characteristics of the phenotype and change of their function in the direction of developing to antigen-presenting, T-cell-stimulating cells. The in vitro behaviour is the same as of Langerhans-cells in vivo after contact-sensitization. With the help of an elisa (=Enzyme-linked immunosorbent assay) specific for apoptosis ( Cell Death Detection Elisa = CDDE) and with electron microscopy was shown, that the cultivated cells died by apoptosis. There is no reference point, that the cells do not do the same in vivo. The process of the functional markers shows, that predominantly the matured cells die by apoptosis and that it was controled by the CD 95/CD 95-L -system The investigations showed, that the Langerhans-cells were eliminated by apoptosis of the culture. The cells after migration in vitro behave in the same manner as after presentation of antigen in vivo. This indicates apoptosis to be the biologic regulation for the elimination of functional matured Langerhans-cells.