Endothelial cells of extremely premature infants display impaired immune response after proinflammatory stimulation.

BackgroundEndothelial cells (ECs) exert immunological functions such as production of proinflammatory cytokines/chemokines as well as facilitation of extravasation of immune cells into infected tissue. Limited data are available on the functionality of ECs from extremely preterm neonates during infection. Accordingly, the aim of our study was to investigate the immune response of premature ECs after proinflammatory stimulation.MethodsCell adhesion receptors' expression and function, nuclear factor 'kappa-light-chain-enhancer' of activated B-cells (NFκB) signaling, and chemokine production were analyzed in umbilical cord ECs from extremely preterm and term neonates after proinflammatory stimulation.ResultsP-selectin and E-selectin surface expression as well as NFκB signaling were lower after lipopolysaccharide (LPS) stimulation in premature ECs. Preterm ECs exhibited lower, but significant, cell-adhesive functions after LPS stimulation compared with term ECs. CCL2/CXCL8 chemokine secretion was significantly upregulated after proinflammatory stimulation in both groups. CXCL10 production was significantly increased in term but not in preterm ECs upon stimulation with tumor necrosis factor compared with unstimulated ECs.ConclusionExtremely premature ECs showed partly reduced expression levels and function of cell adhesion molecules. Both NFκB signaling and chemokine/cytokine production were reduced in premature ECs. The diminished endothelial proinflammatory immune response might result in impaired infection control of preterm newborns rendering them prone to severe infection.

Femtosecond Laser-Processing of Pre-Anodized Ti-Based Bone Implants for Cell-Repellent Functionalization.

Microstructures and nanostructures can be used to reduce the adhesion of the cells on the auxiliary material. Therefore, the aim of our work was to fabricate laser-induced hierarchical microstructures and nanostructures by femtosecond laser-treatment (wavelength 1040 nm, pulse length 350 fs, repetition rates in the kHz range) to reduce the cell adhesion. Additionally, surface chemistry modification by optimized electrochemical anodization was used to further reduce the cell adhesion. For testing, flat plates and bone screws made of Ti-6Al-4V were used. Bone-forming cells (human osteoblasts from the cell line SAOS-2) were grown on the bone implants and additional test samples for two to three weeks. After the growth period, the cells were characterized by scanning electron microscopy (SEM). While earlier experiments with fibroblasts had shown that femtosecond laser-processing followed by electrochemical anodization had a significant impact on cell adhesion reduction, for osteoblasts the same conditions resulted in an activation of the cells with increased production of extracellular matrix material. Significant reduction of cell adhesion for osteoblasts was only obtained at pre-anodized surfaces. It could be demonstrated that this functionalization by means of femtosecond laser-processing can result in bone screws that hinder the adhesion of osteoblasts.

Retinal analysis of a mouse model of Alzheimer's disease with multicontrast optical coherence tomography.

Significance. Recent Alzheimer's disease (AD) patient studies have focused on retinal analysis, as the retina is the only part of the central nervous system that can be imaged noninvasively by optical methods. However, as this is a relatively new approach, the occurrence and role of retinal pathological features are still debated. Aim. The retina of an APP/PS1 mouse model was investigated using multicontrast optical coherence tomography (OCT) in order to provide a documentation of what was observed in both transgenic and wild-type mice. Approach. Both eyes of 24 APP/PS1 transgenic mice (age: 45 to 104 weeks) and 15 age-matched wild-type littermates were imaged by the custom-built OCT system. At the end of the experiment, retinas and brains were harvested from a subset of the mice (14 transgenic, 7 age-matched control) in order to compare the in vivo results to histological analysis and to quantify the cortical amyloid beta plaque load. Results. The system provided a combination of standard reflectivity data, polarization-sensitive data, and OCT angiograms. Qualitative and quantitative information from the resultant OCT images was extracted on retinal layer thickness and structure, presence of hyper-reflective foci, phase retardation abnormalities, and retinal vasculature. Conclusions. Although multicontrast OCT revealed abnormal structural properties and phase retardation signals in the retina of this APP/PS1 mouse model, the observations were very similar in transgenic and control mice.

Evaluating cellularity and structural connectivity on whole brain slides using a custom-made digital pathology pipeline.

AIMS: In brain research, the histopathological examination of coronar whole-brain slides provides important insights into spatial disease characteristics. Regarding brain tumor research, this enables visualization of tumor heterogeneity, infiltration patterns and the relationship with the surrounding brain parenchyma. The precise correlation between radiological imaging and post-mortem brains is of special interest. NEW METHOD: We developed a wide-field slide scanner, comprising a microscope, a high-precision remotely controllable x-y-stage, a camera and a computer workstation, for automatically scanning uncommonly large formats. We analyzed whole brain slides of three patients and constructed cellularity heatmaps and fiber tract maps using a custom-made image processing pipeline. RESULTS: The obtained cellularity heatmaps allow for distinguishing compact tumor (5714 ± 1786 cells/mm², mean ± standard deviation) from white matter (3581 ± 828 cells/mm²) and grey matter (2473 ± 716 cells/mm²). Compared to magnetic resonance imaging, the proposed histopathological work-up (i) reveals a larger zone of tumor infiltration around the compact tumor areas and (ii) shows how pre-existing tracts are displaced by the tumor bulk. Moreover, we highlight differences in the histological tumor growth pattern of two different radiological progression subtypes. COMPARISON WITH EXISTING METHOD(S): Compared to existing (commercial) solutions, our slide scanning solution is adaptable and cost-efficient. Moreover, we showcase potential clinical applications by mapping whole brain histology to magnetic resonance imaging. CONCLUSIONS: We herein provide instructions on how to (i) construct a custom-built slide scanner capable of scanning arbitrary slide formats, (ii) automatically evaluate the cell density and (iii) perform fiber tracking on whole brain slides.

Revealing brain pathologies with multimodal visible light optical coherence microscopy and fluorescence imaging.

We present a multimodal visible light optical coherence microscopy (OCM) and fluorescence imaging (FI) setup. Specification and phantom measurements were performed to characterize the system. Two applications in neuroimaging were investigated. First, curcumin-stained brain slices of a mouse model of Alzheimer's disease were examined. Amyloid-beta plaques were identified based on the fluorescence of curcumin, and coregistered morphological images of the brain tissue were provided by the OCM channel. Second, human brain tumor biopsies retrieved intraoperatively were imaged prior to conventional neuropathologic work-up. OCM revealed the three-dimensional structure of the brain parenchyma, and FI added the tumor tissue-specific contrast. Attenuation coefficients computed from the OCM data and the florescence intensity values were analyzed and showed a statistically significant difference for 5-aminolevulinic acid (5-ALA)-positive and -negative brain tissues. OCM findings correlated well with malignant hot spots within brain tumor biopsies upon histopathology. The combination of OCM and FI seems to be a promising optical imaging modality providing complementary contrast for applications in the field of neuroimaging.

Polarization-sensitive optical coherence tomography imaging of the anterior mouse eye.

Polarization-sensitive optical coherence tomography (PS-OCT) enables noninvasive, high-resolution imaging of tissue polarization properties. In the anterior segments of human eyes, PS-OCT allows the visualization of birefringent and depolarizing structures. We present the use of PS-OCT for imaging the murine anterior eye. Using a spectral domain PS-OCT setup operating in the 840-nm regime, we performed in vivo volumetric imaging in anesthetized C57BL/6 mice. The polarization properties of murine anterior eye structures largely replicated those known from human PS-OCT imagery, suggesting that the mouse eye may also serve as a model system under polarization contrast. However, dissimilarities were found in the depolarizing structure of the iris which, as we confirmed in postmortem histological sections, were caused by anatomical differences between both species. In addition to the imaging of tissues in the anterior chamber and the iridocorneal angle, we demonstrate longitudinal PS-OCT imaging of the murine anterior segment during mydriasis as well as birefringence imaging of corneal pathology in an aged mouse.

Assessment of pathological features in Alzheimer's disease brain tissue with a large field-of-view visible-light optical coherence microscope.

We implemented a wide field-of-view visible-light optical coherence microscope (OCM) for investigating ex-vivo brain tissue of patients diagnosed with Alzheimer's disease (AD) and of a mouse model of AD. A submicrometer axial resolution in tissue was achieved using a broad visible light spectrum. The use of various objective lenses enabled reaching micrometer transversal resolution and the acquisition of images of microscopic brain features, such as cell structures, vessels, and white matter tracts. Amyloid-beta plaques in the range of 10 to 70 µm were visualized. Large field-of-view images of young and old mouse brain sections were imaged using an automated x-y-z stage. The plaque load was characterized, revealing an age-related increase. Human brain tissue affected by cerebral amyloid angiopathy was investigated and hyperscattering structures resembling amyloid beta accumulations in the vessel walls were identified. All results were in good agreement with histology. A comparison of plaque features in both human and mouse brain tissue was performed, revealing an increase in plaque load and a decrease in reflectivity for mouse as compared with human brain tissue. Based on the promising outcome of our experiments, visible light OCM might be a powerful tool for investigating microscopic features in ex-vivo brain tissue.

Beyond backscattering: optical neuroimaging by BRAD.

Optical coherence tomography (OCT) is a powerful technology for rapid volumetric imaging in biomedicine. The bright field imaging approach of conventional OCT systems is based on the detection of directly backscattered light, thereby waiving the wealth of information contained in the angular scattering distribution. Here we demonstrate that the unique features of few-mode fibers (FMF) enable simultaneous bright and dark field (BRAD) imaging for OCT. As backscattered light is picked up by the different modes of a FMF depending upon the angular scattering pattern, we obtain access to the directional scattering signatures of different tissues by decoupling illumination and detection paths. We exploit the distinct modal propagation properties of the FMF in concert with the long coherence lengths provided by modern wavelength-swept lasers to achieve multiplexing of the different modal responses into a combined OCT tomogram. We demonstrate BRAD sensing for distinguishing differently sized microparticles and showcase the performance of BRAD-OCT imaging with enhanced contrast for ex vivo tumorous tissue in glioblastoma and neuritic plaques in Alzheimer's disease.

Spectroscopic imaging with spectral domain visible light optical coherence microscopy in Alzheimer's disease brain samples.

A visible light spectral domain optical coherence microscopy system was developed. A high axial resolution of 0.88 µm in tissue was achieved using a broad visible light spectrum (425 - 685 nm). Healthy human brain tissue was imaged to quantify the difference between white (WM) and grey matter (GM) in intensity and attenuation. The high axial resolution enables the investigation of amyloid-beta plaques of various sizes in human brain tissue and animal models of Alzheimer's disease (AD). By performing a spectroscopic analysis of the OCM data, differences in the characteristics for WM, GM, and neuritic amyloid-beta plaques were found. To gain additional contrast, Congo red stained AD brain tissue was investigated. A first effort was made to investigate optically cleared mouse brain tissue to increase the penetration depth and visualize hyperscattering structures in deeper cortical regions.