311 research outputs found
Acupuncture Muscle Channel in the Subcutaneous Layer of Rat Skin
AbstractUsing a mixed-dye injection technique, we found a novel kind of muscle fiber with a lumen, established its precise location in the subcutaneous muscle layer along the acupuncture muscle of the bladder line, and determined its detailed ultrastructure. The channels with flowing liquid were a novel kind of muscle fibers with lumens and they were located in the subcutaneous muscle layer of rat. Their detection was realized by using chrome-hematoxylin and a mixture of fluorescent nanoparticles and commercial Pelikan ink. These acupuncture muscle channels were hidden among the neighboring skin skeletal muscle fibers and were barely distinguishable from them with light microscopes. Only with a transmission electron microscope were their characteristic features shown to be different from normal skin skeletal muscle. These features included undifferentiated muscle fibers that resembled immature myofibrils without Z-lines and reassembled telophase nuclei
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Characterization of Mammalian Selenoprotein O: A Redox-Active Mitochondrial Protein
Selenoproteins exhibit diverse biological functions, most of which are associated with redox control. However, the functions of approximately half of mammalian selenoproteins are not known. One such protein is Selenoprotein O (SelO), the largest mammalian selenoprotein with orthologs found in a wide range of organisms, including bacteria and yeast. Here, we report characterization of mammalian SelO. Expression of this protein could be verified in HEK 293T cells by metabolic labeling of cells with 75Se, and it was abolished when selenocysteine was replaced with serine. A CxxU motif was identified in the C-terminal region of SelO. This protein was reversibly oxidized in a time- and concentration-dependent manner in HEK 293T cells when cells were treated with hydrogen peroxide. This treatment led to the formation of a transient 88 kDa SelO-containing complex. The formation of this complex was enhanced by replacing the CxxU motif with SxxC, but abolished when it was replaced with SxxS, suggesting a redox interaction of SelO with another protein through its Sec residue. SelO was localized to mitochondria and expressed across mouse tissues. Its expression was little affected by selenium deficiency, suggesting it has a high priority for selenium supply. Taken together, these results show that SelO is a redox-active mitochondrial selenoprotein
The selenoproteome of \u3ci\u3eClostridium\u3c/i\u3e sp. OhILAs: Characterization of anaerobic bacterial selenoprotein methionine sulfoxide reductase A
Selenocysteine (Sec) is incorporated into proteins in response to UGA codons. This residue is frequently found at the catalytic sites of oxidoreductases. In this study, we characterized the selenoproteome of an anaerobic bacterium, Clostridium sp. (also known as Alkaliphilus oremlandii) OhILA, and identified 13 selenoprotein genes, five of which have not been previously described. One of the detected selenoproteins was methionine sulfoxide reductase A (MsrA), an antioxidant enzyme that repairs oxidatively damaged methionines in a stereospecific manner. To date, little is known about MsrA from anaerobes. We characterized this selenoprotein MsrA which had a single Sec residue at the catalytic site but no cysteine (Cys) residues in the protein sequence. Its SECIS (Sec insertion sequence) element did not resemble those in Escherichia coli. Although with low translational efficiency, the expression of the Clostridium selenoprotein msrA gene in E. coli could be demonstrated by 75Se metabolic labeling, immunoblot analyses, and enzyme assays, indicating that its SECIS element was recognized by the E. coli Sec insertion machinery. We found that the Sec-containing MsrA exhibited at least a 20-fold higher activity than its Cys mutant form, indicating a critical role of Sec in the catalytic activity of the enzyme. Furthermore, our data revealed that the Clostridium MsrA was inefficiently reducible by thioredoxin, which is a typical reducing agent for MsrA, suggesting the use of alternative electron donors in this anaerobic bacterium that directly act on the selenenic acid intermediate and do not require resolving Cys residues
The selenoproteome of \u3ci\u3eClostridium\u3c/i\u3e sp. OhILAs: Characterization of anaerobic bacterial selenoprotein methionine sulfoxide reductase A
Selenocysteine (Sec) is incorporated into proteins in response to UGA codons. This residue is frequently found at the catalytic sites of oxidoreductases. In this study, we characterized the selenoproteome of an anaerobic bacterium, Clostridium sp. (also known as Alkaliphilus oremlandii) OhILA, and identified 13 selenoprotein genes, five of which have not been previously described. One of the detected selenoproteins was methionine sulfoxide reductase A (MsrA), an antioxidant enzyme that repairs oxidatively damaged methionines in a stereospecific manner. To date, little is known about MsrA from anaerobes. We characterized this selenoprotein MsrA which had a single Sec residue at the catalytic site but no cysteine (Cys) residues in the protein sequence. Its SECIS (Sec insertion sequence) element did not resemble those in Escherichia coli. Although with low translational efficiency, the expression of the Clostridium selenoprotein msrA gene in E. coli could be demonstrated by 75Se metabolic labeling, immunoblot analyses, and enzyme assays, indicating that its SECIS element was recognized by the E. coli Sec insertion machinery. We found that the Sec-containing MsrA exhibited at least a 20-fold higher activity than its Cys mutant form, indicating a critical role of Sec in the catalytic activity of the enzyme. Furthermore, our data revealed that the Clostridium MsrA was inefficiently reducible by thioredoxin, which is a typical reducing agent for MsrA, suggesting the use of alternative electron donors in this anaerobic bacterium that directly act on the selenenic acid intermediate and do not require resolving Cys residues
Bonghan Ducts as Possible Pathways for Cancer Metastasis
Objective: The present study has been designed to find a possible new route for the
metastasis of cancer cells on the fascia surrounding tumor tissue using a novel technique
of trypan blue staining.
Materials and Methods: Tumor tissues were grown in the skin of nude mice after subcutaneous
inoculation with human lung cancer cells. Trypan blue was recently identified
as a dye with specificity for Bonghan ducts (BHDs) and not other tissues, such as
blood or lymph vessels or nerves.
Results: We demonstrate that the trypan blue staining technique allows the first
visualization of BHDs which are connected to tumor tissues
Bonghan Ducts as Possible Pathways for Cancer Metastasis
Objective: The present study has been designed to find a possible new route for the
metastasis of cancer cells on the fascia surrounding tumor tissue using a novel technique
of trypan blue staining.
Materials and Methods: Tumor tissues were grown in the skin of nude mice after subcutaneous
inoculation with human lung cancer cells. Trypan blue was recently identified
as a dye with specificity for Bonghan ducts (BHDs) and not other tissues, such as
blood or lymph vessels or nerves.
Results: We demonstrate that the trypan blue staining technique allows the first
visualization of BHDs which are connected to tumor tissues
Use of Magnetic Nanoparticles to Visualize Threadlike Structures Inside Lymphatic Vessels of Rats
A novel application of fluorescent magnetic nanoparticles was made to visualize a new tissue which had not been detectable by using simple stereomicroscopes. This unfamiliar threadlike structure inside the lymphatic vessels of rats was demonstrated in vivo by injecting nanoparticles into lymph nodes and applying magnetic fields on the collecting lymph vessels so that the nanoparticles were taken up by the threadlike structures. Confocal laser scanning microscope images of cryosectioned specimens exhibited that the nanoparticles were absorbed more strongly by the threadlike structure than by the lymphatic vessels. Further examination using a transmission electron microscope revealed that the nanoparticles had been captured between the reticular fibers in the extracellular matrix of the threadlike structures. The emerging technology of nanoparticles not only allows the extremely elusive threadlike structures to be visualized but also is expected to provide a magnetically controllable means to investigate their physiological functions
Expression of Keratin 10 in Rat Organ Surface Primo-vascular Tissues
AbstractThe primo-vascular system is described as the anatomical structure corresponding to acupuncture meridians and has been identified in several tissues in the body, but its detailed anatomy and physiology are not well understood. Recently, the presence of keratin 10 (Krt10) in primo-vascular tissue was reported, but this finding has not yet been confirmed. In this study, we compared Krt10 expression in primo-vascular tissues located on the surface of rat abdominal organs with Krt10 expression on blood and lymphatic vessels. Krt10 protein (approximately 56.5 kDa) was evaluated by western blot analysis and immunohistochemistry. Krt10 (IR) in the primo-node was visualized as patchy spots around each cell or as a follicle-like structure containing a group of cells. Krt10 IR was also identified in vascular and lymphatic tissues, but its distribution was diffuse over the extracellular matrix of the vessels. Thus Krt10 protein was expressed in all three tissues tested, but the expression pattern of Krt10 in primo-vascular tissue differed from those of blood and lymphatic vascular tissues, suggesting that structural and the regulatory roles of Krt10 in primo-vascular system are different from those in blood and lymphatic vessels
Evaluation of an immunochromatographic assay for the detection of anti-hepatitis A virus IgM
<p>Abstract</p> <p>Background</p> <p>Hepatitis A virus (HAV) is a causative agent of acute hepatitis, which is transmitted by person-to-person contact and via the faecal-oral route. Acute HAV infection is usually confirmed by anti-HAV IgM detection. In order to detect anti-HAV IgM in the serum of patients infected with HAV, we developed a rapid assay based on immunochromatography (ICA) and evaluated the sensitivity of this assay by comparing it with a commercial microparticle enzyme immunoassay (MEIA) that is widely used for serological diagnosis.</p> <p>Results</p> <p>The newly developed ICA showed 100% sensitivity and specificity when used to test 150 anti-HAV IgM-positive sera collected from infected patients and 75 negative sera from healthy subjects. Also, the sensitivity of ICA is about 10 times higher than MEIA used in this study by determining end point to detect independent on infected genotype of HAV. In addition, the ICA was able to detect 1 positive sample from among 50 sera from acute hepatitis patients that had tested negative for anti-HAV IgM using the MEIA.</p> <p>Conclusion</p> <p>Conclusively, ICA for the detection of anti-HAV IgM will be very effective for rapid assay to apply clinical diagnosis and epidemiological investigation on epidemics due to the simplicity, rapidity and specificity.</p
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