Hepatosplenomegaly in liver cirrhosis is caused by reactive oxygen species formation, an increase in apoptosis and autophagy, and pronounced autoimmune reactions

Various factors of infectious and toxic genesis can lead to the liver cirrhosis, often accompanied by complications such as recurrent bleeding due to portal hypertension against the background of hepatosplenomegaly. Metabolic changes and disturbances in immunoreactivity occur in the liver and spleen. To substantiate the choice of personalized treatment tactics for patients with hepatosplenomegaly, we investigated individual metabolic predictors and immunopathological processes in patients with: liver cirrhosis and hepatitis B (HBV) and/or hepatitis C (HCV) viruses (I group, n = 52); with herpes viruses CMV (cytomegalovirus) and EBV (Epstein-Barr virus) (II group, n = 48), and with splenomegaly and frequent recurrent bleeding associated with hereditary enzymopathies (III group, n = 15). We used the methods of immunoturbidimetry; enzyme immunoassay; light, fluorescence and confocal microscopy. In group I (HBV/HCV), we revealed a decrease in the C4 component; a significant increase in the phagocytic index and phagocytic number, a reduced number of active phagocytes and the digestion index; a decrease in the IL-1β content and an increase in IL-18 and IL-6. In group II (CMV/EBV), we revealed a high activity of the C3 and a low activity of the C4 component against the background of a high level of ROS in neutrophils; the antineutrophil antibodies (ANCA) formation in 85.7% of patients (71.4% –perinuclear antibodies (pANCA) to myeloperoxidase; 14.3% – cytoplasmic antibodies (CANCA) to proteinase 3). Also, in group II, an increased level of pro-inflammatory cytokines IFN-γ, IL-1β, TNF-α, IL-18 and anti-inflammatory IL-6 was detected. Changes in links of immunity in II group led to the formation of autoimmune reactions in 64.7% of patients, which was expressed in the development of a broad range of antinuclear antibodies ANA (11 specificities, including ANA to chromatin and chromatin-associated proteins, to proteins cytoskeleton, enzymes and enzyme complexes). In group III, we revealed a low absorption capacity of neutrophils, a high frequency of antineutrophil antibodies pANCA occurrence and cANCA (in 67.2% of the examined), and low concentration of TNF-α. The developed model of the stepwise change of immunological markers makes it possible to substantiate the choice of a complex targeted treatment, including antiviral and immunotropic therapy.

Coming together—symbiont acquisition and early development in deep-sea bathymodioline mussels

How and when symbionts are acquired by their animal hosts has a profound impact on the ecology and evolution of the symbiosis. Understanding symbiont acquisition is particularly challenging in deep-sea organisms because early life stages are so rarely found. Here, we collected early developmental stages of three deep-sea bathymodioline species from different habitats to identify when these acquire their symbionts and how their body plan adapts to a symbiotic lifestyle. These mussels gain their nutrition from chemosynthetic bacteria, allowing them to thrive at deep-sea vents and seeps worldwide. Correlative imaging analyses using synchrotron-radiation based microtomography together with light, fluorescence and electron microscopy revealed that the pediveliger larvae were aposymbiotic. Symbiont colonization began during metamorphosis from a planktonic to a benthic lifestyle, with the symbionts rapidly colonizing first the gills, the symbiotic organ of adults, followed by all other epithelia of their hosts. Once symbiont densities in plantigrades reached those of adults, the host's intestine changed from the looped anatomy typical for bivalves to a straightened form. Within the Mytilidae, this morphological change appears to be specific to Bathymodiolus and Gigantidas , and is probably linked to the decrease in the importance of filter feeding when these mussels switch to gaining their nutrition largely from their symbionts.

Cell wall characteristics during sexual reproduction of Mougeotia sp. (Zygnematophyceae) revealed by electron microscopy, glycan microarrays and RAMAN spectroscopy

Mougeotia spp. collected from field samples were investigated for their conjugation morphology by light-, fluorescence-, scanning- and transmission electron microscopy. During a scalarifom conjugation, the extragametangial zygospores were initially surrounded by a thin cell wall that developed into a multi-layered zygospore wall. Maturing zygospores turned dark brown and were filled with storage compounds such as lipids and starch. While M. parvula had a smooth surface, M. disjuncta had a punctated surface structure and a prominent suture. The zygospore wall consisted of a polysaccharide rich endospore, followed by a thin layer with a lipid-like appaerance, a massive electron dense mesospore and a very thin exospore composed of polysaccharides. Glycan microarray analysis of zygospores of different developmental stages revealed the occurrence of pectins and hemicelluloses, mostly composed of homogalacturonan (HG), xyloglucans, xylans, arabino-galactan proteins and extensins. In situ localization by the probe OG7-13AF 488 labelled HG in young zygospore walls, vegetative filaments and most prominently in conjugation tubes and cross walls. Raman imaging showed the distribution of proteins, lipids, carbohydrates and aromatic components of the mature zygospore with a spatial resolution of ~ 250 nm. The carbohydrate nature of the endo- and exospore was confirmed and in-between an enrichment of lipids and aromatic components, probably algaenan or a sporopollenin-like material. Taken together, these results indicate that during zygospore formation, reorganizations of the cell walls occured, leading to a resistant and protective structure.

Antibacterial Potential by Rupture Membrane and Antioxidant Capacity of Purified Phenolic Fractions of Persea americana Leaf Extract

The present research focused on evaluating the antibacterial effect and the mechanism of action of partially purified fractions of an extract of Persea americana. Furthermore, both its antioxidant capacity and composition were evaluated. The extract was fractionated by vacuum liquid chromatography. The antimicrobial effect against Staphylococcus aureus (ATCC 6538), Escherichia coli (ATCC 11229), Pseudomonas aeruginosa (ATCC 15442), and Salmonella choleraesuis (ATCC 1070) was analyzed by microdilution and the mechanism of action by the Sytox green method. The antioxidant capacity was determined by DPPH, FRAP, and ABTS techniques and the composition by Rp-HPLC-MS. All fractions showed a concentration-dependent antibacterial effect. Fractions F3, F4, and F5 (1000 µg/mL) showed a better antibacterial effect than the extract against the bacteria mentioned. The F3 fraction showed inhibition of 95.43 ± 3.04% on S. aureus, F4 showed 93.30 ± 0.52% on E. coli, and F5 showed 88.63 ± 1.15% on S. choleraesuis and 86.46 ± 3.20% on P. aeruginosa. The most susceptible strain to the treatment with the extract was S. aureus. Therefore, in this strain, the bacterial membrane damage induced by the extract and fractions was evidenced by light fluorescence microscopy. Furthermore, the extract had better antioxidant action than each fraction. Finally, sinensitin was detected in F3 and cinnamoyl glucose, caffeoyl tartaric acid, and cyanidin 3-O-(6′′-malonyl-3′′-glucosyl-glucoside) were detected in F4; esculin and kaempferide, detected in F5, could be associated with the antibacterial and antioxidant effect.

X-ray microscopy enables multiscale high-resolution 3D imaging of plant cells, tissues, and organs

Capturing complete internal anatomies of plant organs and tissues within their relevant morphological context remains a key challenge in plant science. While plant growth and development are inherently multiscale, conventional light, fluorescence, and electron microscopy platforms are typically limited to imaging of plant microstructure from small flat samples that lack direct spatial context to, and represent only a small portion of, the relevant plant macrostructures. We demonstrate technical advances with a lab-based X-ray microscope (XRM) that bridge the imaging gap by providing multiscale high-resolution 3D volumes of intact plant samples from the cell to whole plant level. Serial imaging of a single sample is shown to provide sub-micron 3D volumes co-registered with lower magnification scans for explicit contextual reference. High quality 3D volume data from our enhanced methods facilitate more sophisticated and effective computational segmentation and analyses than have previously been employed for X-ray based imaging. Advances in sample preparation make multimodal correlative imaging workflows possible, where a single resin-embedded plant sample is scanned via XRM to generate a 3D cell-level map, and then used to identify and zoom in on sub-cellular regions of interest for high resolution scanning electron microscopy. In total, we present the methodologies for use of XRM in the multiscale and multimodal analysis of 3D plant features using numerous economically and scientifically important plant systems.

In Vitro Activity of Sertraline, an Antidepressant, Against Antibiotic-Susceptible and Antibiotic-Resistant Helicobacter pylori Strains

Antibiotic resistance of Helicobacter pylori, a spiral bacterium associated with gastric diseases, is a topic that has been intensively discussed in last decades. Recent discoveries indicate promising antimicrobial and antibiotic-potentiating properties of sertraline (SER), an antidepressant substance. The aim of the study, therefore, was to determine the antibacterial activity of SER in relation to antibiotic-sensitive and antibiotic-resistant H. pylori strains. The antimicrobial tests were performed using a diffusion-disk method, microdilution method, and time-killing assay. The interaction between SER and antibiotics (amoxicillin, clarithromycin, tetracycline, and metronidazole) was determined by using a checkerboard method. In addition, the study was expanded to include observations by light, fluorescence, and scanning electron microscopy. The growth inhibition zones were in the range of 19–37 mm for discs impregnated with 2 mg of SER. The minimal inhibitory concentrations (MICs) and minimal bactericidal concentrations (MBCs) counted for 2–8 µg/mL and 4–8 µg/mL, respectively. The time-killing assay showed the time-dependent and concentration-dependent bactericidal activity of SER. Bacteria exposed to MBCs (but not sub-MICs and MICs ≠ MBCs) underwent morphological transformation into coccoid forms. This mechanism, however, was not protective because these cells after a 24-h incubation had a several-fold reduced green/red fluorescence ratio compared to the control. Using the checkerboard assay, a synergistic/additive interaction of SER with all four antibiotics tested was demonstrated. These results indicate that SER may be a promising anti-H. pylori compound.

Sites of secretion of bioactive compounds in leaves of Dracocephalum moldavica L.: anatomical, histochemical, and essential oil study

Dracocephalum moldavica L. is an aromatic plant emitting intense lemon scent. The aboveground parts of the plants constitute raw material for medicine and food industry. In contrast to the comprehensively investigated trichomes, there are only few studies of the histochemical characteristics of the leaves of essential oil-bearing plants from the family Lamiaceae. The present study shows the micromorphology, anatomy, and histochemistry of the leaves of the analysed species. The research aimed to determine the location of essential oil and other specialised metabolites in leaf tissues. The investigations of fresh and fixed material were carried out with the use of light, fluorescence, and scanning electron microscopy. Additionally, the content and composition of essential oil in the leaves were determined with the GC/MS method. The leaf epidermis had non-glandular unbranched trichomes and three types of glandular trichomes: peltate as well as long and short capitate trichomes. The results of the histochemical assays showed positive reactions to lipids and to some secondary compounds such as essential oil, terpenes, phenolic compounds, and flavonoids in all types of the glandular and non-glandular trichomes. The same compounds were found in the epidermis cells of the leaves. The results of the present study indicate that the intense smell of the leaves is associated with emission of essential oil not only by the glandular and non-glandular trichomes but also by the leaf epidermis cells. The main components in the essential oil (0.10%) include geranial, neral, geraniol, nerol, and trans-myrtanol acetate. Since D. moldavica has been used as an adulterant of Melissa officinalis L., the anatomical traits of leaves and the essential oil composition in both species were compared in the study.

Microstructural and histochemical analysis of aboveground organs of Centaurea cyanus used in herbal medicine

Centaurea cyanus L. is a valuable source of many different bioactive substances. It is used in herbal medicine, but the structure of its organs used as raw material and secretory tissues has been insufficiently examined. The aim of this paper was to investigate the microstructure of C. cyanus flowers, bracts, stems and leaves with particular emphasis on secretory structures. Moreover, the main classes of secondary metabolites present in the secretion were identified and the taxonomic value of some micromorphological and anatomical features was analysed. Histochemical, micromorphological and ultrastructural analyses of aboveground organs of C. cyanus were carried out using light, fluorescence, scanning and transmission electron microscopy. The analyses revealed the presence of petal papillae and a characteristic cuticular pattern on the petals, stamens and stylar hairs. There were four types of non-glandular trichomes on the bracts, leaves and stem surfaces. The epidermal cells of the bracts contained prismatic calcium oxalate crystals. Two kinds of secretory structures, i.e. glandular trichomes and ducts, were observed in the C. cyanus organs. The glandular trichomes were located on the bract and stem surfaces, and the ducts were detected in the leaves and stems. Ultrastructural analyses of the epithelium of the ducts showed the presence of strongly osmiophilic insoluble phenolic material in vacuoles as well as moderately osmiophilic insoluble lipidic material in elaioplasts and vesicles. The results of histochemical assays showed a heterogeneous nature of the duct secretion, which contained essential oil, lipids, flavonoids, tannins and terpenes containing steroids.