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41.
42.
Summary In the present study the cellular fine structure of freeze-substituted sporidia of the phytopathogenic fungusUstilago avenae is investigated by means of thin-section electron microscopy. A conventional embedding method using Spurr's low viscosity resin is compared with the recently developed methacrylate mixtures Lowicryl® K 4 M and HM 20 resin. Generally, freeze-substitution yields improved preservation of fine structural details of the fungus compared to previously applied conventional fixation methods. Using double fixation during freeze-substitution prior to conventional embedding the fungal membrane system (plasmalemma, endoplasmic reticulum), organelles (mitochondria, nucleus etc.) and other cytoplasmic features (ribosomes, cytoskeleton) appear well resolved and smoothly contoured. Aldehyde fixed and Lowicryl embedded sporidia ofU. avenae resemble these double fixed fungal specimens fairly closely. The complete low-temperature preparation produces visualization of distinct cellular details although contrast reversal of cellular membranes (er, mitochondria etc.) is sometimes observed. In particular, fine structure resolution is enhanced in Lowicryl HM 20 embedded fungal cells. This is due also to significant improvement in staining of the cellular membranes, cytoskeleton (microfilaments and microtubules) and Golgi apparatus-like areas, using tannic acid. In case of the fungusU. avenae, freeze-substitution in combination with mild glutaraldehyde fixation and final low-temperature embedding in HM 20 resin prove suitable for improved preservation of cellular ultrastructure.Abbreviations cw
cell wall
- cy
cytoplasm
- FS
freeze-substitution
- FS-A
GA/OsO4 freeze-substitution and Spurr's LV-embedding
- FS-B
GA freeze-substitution and Lowicryl K 4 M LT-embedding
- FS-C
GA freeze-substitution and Lowicryl HM 20 LT-embedding
- go
Golgi apparatus-like body
- GA
glutaraldehyde
- g
glycogen deposit
- l
lipid droplet
- LT
low temperature
- Lowicryl LT-embedding
Lowicryl low-temperature embedding
- Lowicryl LT-resin
Lowicryl low-temperature resin
- MeOH
methanol
- mf
microfilament
- mt
microtubule
- m
mitochondrion
- mvb
multivesicular body
- ne
nuclear envelope
- np
nuclear pore
- npl
nucleoplasm
- nu
nucleolus
- n
nucleus
- OsO4
osmium tetroxide
- pl
plasmalemma
- pr
polyribosomes
- Pb-citrate
Reynolds' lead citrate
- r
ribosome
- RT
room temperature
- rer
rough endoplasmic reticulum
- Spurr's LV-embedding
Spurr's low viscosity embedding
- Spurr's LV-resin
Spurr's low viscosity resin
- t
tonoplast
- Uac
uranyl acetate
- v
vacuole 相似文献
43.
W. Hermanns 《Histochemistry and cell biology》1987,86(3):225-227
Summary A double staining method is presented which allows the enzyme histochemical differentiation between osteoclasts (mono- and multinucleated forms) and mononuclear phagocytes (macrophages, multinucleated inflammatory giant cells). Osteoclasts are characterized by a strong acid phosphatase activity whereas macrophages and inflammatory giant cells show a variable non-specific esterase activity. The described method may be useful in studying the osteoclast origin and the extraosseus distribution of these cells.Supported by Deutsche Forschungsgemeinschaft, SFB 244,A1 相似文献
44.
Background
DNA methylation plays an important role in the process of tumorigenesis. Identifying differentially methylated genes or CpG islands (CGIs) associated with genes between two tumor subtypes is thus an important biological question. The methylation status of all CGIs in the whole genome can be assayed with differential methylation hybridization (DMH) microarrays. However, patient samples or cell lines are heterogeneous, so their methylation pattern may be very different. In addition, neighboring probes at each CGI are correlated. How these factors affect the analysis of DMH data is unknown. 相似文献45.
Rapid biomonitoring protocols, using biotic indices based on macroinvertebrate diversity to assess river ecosystem health, are widely used globally. Such quick assessment techniques are lauded for the rapid results obtained and the relatively easy protocol used to achieve an answer. However, do such quick assessments of water quality give enough information about ecosystems? Are important details being overlooked? When should a full faunal survey be used in preference? Important research programmes, including environmental impact studies, often misuse biomonitoring techniques, making influential management decisions using superficial, low-level data obtained using biomonitoring tools, inappropriate to address those management objectives. The value of using biomonitoring as a quick tool, versus a more detailed faunal assessment, is considered here. The assessment of teloganodid mayfly fauna occurring in South African rivers provides an example of the value of detailed studies versus superficial family level investigations, showing that a rapid biomonitoring approach should not be used as a shortcut when a more detailed survey is needed. Each situation should be assessed for its own merit in a given set of project circumstances. A checklist of criteria is presented, giving guidance on when rapid biomonitoring alone is valuable and when more detailed assessments would give a more relevant result. 相似文献
46.
47.
Stross C Radtke S Clahsen T Gerlach C Volkmer-Engert R Schaper F Heinrich PC Hermanns HM 《The Journal of biological chemistry》2006,281(13):8458-8468
Down-regulation of interleukin (IL)-6-type cytokine signaling has been shown to occur, among other mechanisms, via induction of the feedback inhibitor SOCS3 (suppressor of cytokine signaling 3). Binding of SOCS3 to the phosphorylated Tyr(759) in the cytoplasmic region of gp130, the common signal transducing receptor chain of all IL-6-type cytokines, is necessary for inhibition of Janus kinase-mediated signaling. In the present study, we analyzed the effect of SOCS3 on signal transduction by the proinflammatory cytokine oncostatin M (OSM), which signals through a receptor complex of gp130 and the OSM receptor (OSMR). OSM leads to a much stronger and prolonged induction of SOCS3 in HepG2 hepatoma cells and murine embryonal fibroblasts (MEF) compared with IL-6. A negative effect of SOCS3 on OSM signaling was confirmed using MEF cells lacking SOCS3. We can show that the OSMR-mediated signaling is inhibited by SOCS3 to a similar extent as previously described for gp130. However, the inhibition occurs independent of tyrosine motifs within the OSMR. Instead, SOCS3 interacts directly with JAK1 in a stimulation-dependent manner, a mechanism so far only known for SOCS1. 相似文献
48.
Maria Iris Hermanns Sabine Fuchs Michaela Bock Katja Wenzel Eckhard Mayer Kai Kehe Fernando Bittinger C. James Kirkpatrick 《Cell and tissue research》2009,336(1):91-105
In order to delineate individual pathomechanisms in acute lung injury and pulmonary toxicology, we developed a primary coculture
system to simulate the human alveolo-capillary barrier. Human pulmonary microvascular endothelial cells (HPMEC) were cocultivated
with primary isolated human type II alveolar epithelial cells (HATII) on opposite sides of a permeable filter support, thereby
constituting a bilayer. Within 7–11 days of coculture, the HATII cells partly transdifferentiated to type-I-like (HATI-like)
cells, as demonstrated by morphological changes from a cuboidal to a flattened morphology, the loss of HATII-cell-specific
organelles and the increase of HATI-cell-related markers (caveolin-1, aquaporin-5, receptor for advanced glycation end-products).
Immunofluorescent analysis detected type-II-like and type-I-like alveolar epithelial cells mimicking the heterocellular composition
of alveolar epithelium in vivo. The heterocellular epithelial monolayer showed a circumferential staining of tight-junctional
(ZO-1, occludin) and adherens-junctional (E-cadherin, β-catenin) proteins. HPMEC on the opposite side also developed tight
and adherens junctions (VE-cadherin, β-catenin). Under integral barrier properties, exposure to the proinflammatory cytokine
tumour necrosis factor-α from either the endothelial (basolateral) or the epithelial (apical) side caused a largely compartmentalized
release of the chemokines interleukin-8 and monocyte chemoattractant protein-1. Thus, the established coculture provides a
suitable in vitro model to examine barrier function at the distal lung, including the interaction of microvascular endothelial
cells with ATII-like and ATI-like epithelial cells. The compartmentalization of the barrier-forming bilayer also allows mechanisms
of lung injury to be studied in both the epithelial (intra-alveolar) and the endothelial (intravascular) compartments.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.
This study was supported by the BMVg Grant E/B41G/1G302/1A402 and the 6th Framework program of the European Union, NanoBioPharmaceutics. 相似文献
49.
Ana Cecilia AX De-Oliveira Renato S Carvalho Flavio HM Paixão Hellen S Tavares Luciana S Gueiros Carolina M Siqueira Francisco JR Paumgartten 《Malaria journal》2010,9(1):1-17
Background
The mechanisms by which malaria up and down-regulates CYP activities are not understood yet. It is also unclear whether CYP activities are modulated during non-lethal malaria infections. This study was undertaken to evaluate the time course of CYP alterations in lethal (Plasmodium berghei ANKA) and non-lethal (Plasmodium chabaudi chabaudi) murine malaria. Additionally, hypotheses on the association of CYP depression with enhanced nitric oxide (NO) production, and of CYP2a5 induction with endoplasmic reticulum dysfunction, enhanced haem metabolism and oxidative stress were examined as well.Methods
Female DBA-2 and C57BL/6 mice were infected with P.berghei ANKA or P. chabaudi and killed at different post-infection days. Infection was monitored by parasitaemia rates and clinical signs. NO levels were measured in the serum. Activities of CYP1a (ethoxyresorufin-O-deethylase), 2b (benzyloxyresorufin-O-debenzylase), 2a5 (coumarin-7-hydroxylase) and uridine-diphosphoglucuronyl-transferase (UGT) were determined in liver microsomes. Glutathione-S-transferase (GST) activity and concentrations of gluthatione (GSH) and thiobarbituric acid-reactive substances (TBARS) were determined in the liver. Levels of glucose-regulated protein 78 (GRP78) were evaluated by immunoblotting, while mRNAs of haemoxygenase-1 (HO-1) and inducible nitric oxide synthase (iNOS) were determined by quantitative RT-PCR.Results
Plasmodium berghei depressed CYP1a and 2b and induced 2a5 in DBA-2 mice. In P.berghei-infected C57BL/6 mice CYP activities remained unaltered. In both strains, GST and UGT were not affected by P.berghei. Plasmodium c. chabaudi depressed CYP1a and 2b and induced 2a5 activities on the day of peak parasitaemia or near this day. CYP2a5 induction was associated with over-expression of HO-1 and enhanced oxidative stress, but it was not associated with GRP78 induction, a marker of endoplasmic reticulum stress. Plasmodium chabaudi increased serum NO on days near the parasitaemia peak in both strains. Although not elevating serum NO, P.berghei enhanced iNOS mRNA expression in the liver.Conclusion
Down-regulation of CYP1a and 2b and induction of 2a5 occurred in lethal and non-lethal infections when parasitaemia rates were high. A contribution of NO for depression of CYP2b cannot be ruled out. Results were consistent with the view that CYP2a5 and HO-1 are concurrently up-regulated and suggested that CYP2a5 induction may occur in the absence of enhanced endoplasmic reticulum stress. 相似文献50.