Revealing the Chemical Composition of Birch Pollen Grains by Raman Spectroscopic Imaging

The investigation of the biochemical composition of pollen grains is of the utmost interest for several environmental aspects, such as their allergenic potential and their changes in growth conditions due to climatic factors. In order to fully understand the composition of pollen grains, not only is an in-depth analysis of their molecular components necessary but also spatial information of, e.g., the thickness of the outer shell, should be recorded. However, there is a lack of studies using molecular imaging methods for a spatially resolved biochemical composition on a single-grain level. In this study, Raman spectroscopy was implemented as an analytical tool to investigate birch pollen by imaging single pollen grains and analyzing their spectral profiles. The imaging modality allowed us to reveal the layered structure of pollen grains based on the biochemical information of the recorded Raman spectra. Seven different birch pollen species collected at two different locations in Germany were investigated and compared. Using chemometric algorithms such as hierarchical cluster analysis and multiple-curve resolution, several components of the grain wall, such as sporopollenin, as well as the inner core presenting high starch concentrations, were identified and quantified. Differences in the concentrations of, e.g., sporopollenin, lipids and proteins in the pollen species at the two different collection sites were found, and are discussed in connection with germination and other growth processes

Intestinal epithelial barrier integrity investigated by label-free techniques in ulcerative colitis patients

The intestinal epithelial barrier, among other compartments such as the mucosal immune system, contributes to the maintenance of intestinal homeostasis. Therefore, any disturbance within the epithelial layer could lead to intestinal permeability and promote mucosal inflammation. Considering that disintegration of the intestinal epithelial barrier is a key element in the etiology of ulcerative colitis, further assessment of barrier integrity could contribute to a better understanding of the role of epithelial barrier defects in ulcerative colitis (UC), one major form of chronic inflammatory bowel disease. Herein, we employ fast, non-destructive, and label-free non-linear methods, namely coherent anti-Stokes Raman scattering (CARS), second harmonic generation (SHG), two-photon excited fluorescence (TPEF), and two-photon fluorescence lifetime imaging (2P-FLIM), to assess the morpho-chemical contributions leading to the dysfunction of the epithelial barrier. For the first time, the formation of epithelial barrier gaps was directly visualized, without sophisticated data analysis procedures, by the 3D analysis of the colonic mucosa from severely inflamed UC patients. The results were compared with histopathological and immunofluorescence images and validated using transmission electron microscopy (TEM) to indicate structural alterations of the apical junction complex as the underlying cause for the formation of the epithelial barrier gaps. Our findings suggest the potential advantage of non-linear multimodal imaging is to give precise, detailed, and direct visualization of the epithelial barrier in the gastrointestinal tract, which can be combined with a fiber probe for future endomicroscopy measurements during real-time in vivo imaging

Label-free multimodal imaging of infected Galleria mellonella larvae

Non-linear imaging modalities have enabled us to obtain unique morpho-chemical insights into the tissue architecture of various biological model organisms in a label-free manner. However, these imaging techniques have so far not been applied to analyze the Galleria mellonella infection model. This study utilizes for the first time the strength of multimodal imaging techniques to explore infection-related changes in the Galleria mellonella larvae due to massive E. faecalis bacterial infection. Multimodal imaging techniques such as fluorescent lifetime imaging (FLIM), coherent anti-Stokes Raman scattering (CARS), two-photon excited fluorescence (TPEF), and second harmonic generation (SHG) were implemented in conjunction with histological HE images to analyze infection-associated tissue damage. The changes in the larvae in response to the infection, such as melanization, vacuolization, nodule formation, and hemocyte infiltration as a defense mechanism of insects against microbial pathogens, were visualized after Enterococcus faecalis was administered. Furthermore, multimodal imaging served for the analysis of implant-associated biofilm infections by visualizing biofilm adherence on medical stainless steel and ePTFE implants within the larvae. Our results suggest that infection-related changes as well as the integrity of the tissue of G. mellonella larvae can be studied with high morphological and chemical contrast in a label-free manner

Exposure to allergens of different cattle breeds and their relevance in occupational allergy

Introduction Cattle are an important source of allergens in the working area of farmers. Asthma caused by cow allergens is a significant occupational problem. Yet in allergological testing, the results of in vivo and in vitro diagnostic tests are often inconsistent even in cases with clearly cattle-related symptoms. Objectives and methods The aim of this study was to investigate four different commercial cow allergen extracts and to compare them with self prepared extracts of different cattle breeds by means of SDS-PAGE and immunoblotting using the sera of 42 German farmers with asthma and rhino-conjunctivitis caused by cattle contact. Results The commercial extracts investigated in this study showed only minor differences in protein pattern. Using sera in immunoblotting experiments distinct bands were found for all symptomatic farmers, even in 13 farmers with a negative result in commercially available serological allergy tests. Bands with molecular weights in the range between about 11 and 67 kDa were observed; reactivity with the major allergen Bos d 2 at about 20 kDa was detected in all farmers, although it was not the strongest band in all cases. Conclusions We demonstrate for the first time the allergenic relevance of additional proteins with molecular weights of 14, 30, 55 and approx. 67-97 kDa in more than 50% of farmers with cattle related symptoms. One of our most striking results was that 32% of the investigated farmers with cattle related symptoms showed negative results with commercial serological tests but distinct reactions with cow allergen in immunoblotting experiments. The Bos d 2 content in hair showed differences between certain breeds whereas German Brown and Simmental had particularly higher quantities of Bos d 2 in their hair than breeds such as Holstein-Friesian. These results strongly support the following recommendation: test results with commercial extracts that are contradictory to the clinical symptoms should be supplemented by skin tests using extracts of the hair of the farmers' own cattle

Gesundheitliche Bewertung von Maßnahmen und Energieträgern im Rahmen der Energiewende aus Umweltmedizinischer Sicht

Heutelbeck A, Hornberg C, Herr C, Dott W, Eikmann T, Schümann M. Gesundheitliche Bewertung von Maßnahmen und Energieträgern im Rahmen der Energiewende aus Umweltmedizinischer Sicht. Bundesgesundheitsblatt. 2017