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1.
Mario Castellucci Thomas Theelen Elena Pompili Lorenzo Fumagalli Gabriella De Renzis Judith Mühlhauser 《Cell and tissue research》1994,278(2):283-289
Proteases and their inhibitors play a pivotal role in developmental and differentiative processes. In the present report we investigated the immunohistochemical localization of 1-antitrypsin, 1-antichymotrypsin and inter--trypsin inhibitor in first trimester as well as in term human placentas. For this purpose polyclonal antibodies against these serine-protease inhibitors were used. All inhibitors were expressed in the villous syncytiotrophoblast of first and last trimester placentas. Placental fibrinoid was positively stained for 1-antitrypsin and inter--trypsin inhibitor throughout gestation. 1-Antitrypsin and 1-antichymotrypsin showed a strong immunostaining in the Hofbauer cells (first trimester and full term placentas). Extravillous cytotrophoblast was negative for the three protease inhibitors throughout gestation. The presence of the three inhibitors in the syncytiotrophoblast suggests a role in coagulative, invasive and immunomodulatory processes. Fibrinoid, staining for 1-antitrypsin and inter--trypsin inhibitor, could also have an important immunoprotective function. The presence of protease inhibitors in the Hofbauer cells suggests an involvement of these cells in villous remodelling and differentiative processes. 相似文献
2.
Our previous studies have demonstrated that a large quantity of oxytocin (OT)-like substance exists in human placental tissue. In the present study, the localization of the OT-like substance in the human placenta was investigated by the PAP (peroxidase and antiperoxidase complex) immunohistochemical method. The results demonstrated that the site of syncytiotrophoblast was positively stained by specific antiserum to OT, whereas the tissue was not significantly stained by normal rabbit serum (NRS). These results suggest that the OT-like substance might be localized in syncytiotrophoblast of the placental tissue. 相似文献
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Toshihiro Kobayashi Vadim S. Zinchuk Teruhiko Okada Eva Garcia del Saz H. Seguchi 《Histochemistry and cell biology》1998,110(4):395-406
Human neutrophils possess alkaline phosphatase-containing intracellular granules which are upregulated to the cell surface
upon stimulation. The mechanism that governs the intracellular dynamics of these granules is, however, poorly understood.
The aim of the present study was to investigate the possible participation of GTP-binding proteins in the reorganization and
exocytosis of the alkaline phosphatase-containing granules using electropermeabilized cells. Biochemical assays using intact
neutrophils showed that the alkaline phosphatase activity was upregulated and exocytosed into the extracellular space upon
stimulation with AlF4
– and N-formyl peptide. This upregulation was inhibited by treatment of cells with pertussis toxin and botulinum toxin. Alkaline
phosphatase activity was also upregulated in electropermeabilized cells stimulated with guanosine 5′-O-(3-thiotriphosphate) (GTPγS), but not with guanosine 5′-O-(2-thiodiphosphate) (GDPβS). Cytochemically, alkaline phosphatase-containing granules were dispersed throughout the cytoplasm
in unstimulated electropermeabilized neutrophils. Upon stimulation with GTPγS, but not with GDPβS, these granules fused to
form elongated tubular structures which eventually became associated with the plasma membrane. Nocodazole disturbed the reorganization
of the alkaline phosphatase-containing granules in cells stimulated with GTPγS. The results from this study indicate that
GTP-binding proteins participate in the reorganization and exocytosis of alkaline phosphatase-containing granules associated
with the microtubules in electropermeabilized human neutrophils.
Accepted: 31 March 1998 相似文献
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Isolation and characterization of human placenta fibronectin 总被引:3,自引:0,他引:3
M Isemura Y Yamaguchi H Munakata J Aikawa M Kan I Yamane Z Yosizawa 《Journal of biochemistry》1984,96(1):163-169
Fibronectin was isolated from human placenta tissues and compared with human plasma fibronectin. Placenta and plasma fibronectins had similar amino acid compositions, immunological properties, and cell attachment-promoting activities, but differed in apparent molecular weight on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, which could be accounted for at least partly by the difference in carbohydrate composition. Unlike plasma fibronectin, placenta fibronectin failed to form a precipitin line with concanavalin A in a double diffusion system. The non- or low-reactivity of placenta fibronectin with this lectin was also demonstrated by affinity chromatography with concanavalin A-agarose, in which more than 90% of the radiolabeled glycopeptides derived from placenta fibronectin was not retained on the gel. The two fibronectins also differed in the reactivity with Lens culinaris agglutinin of their glycopeptide fractions. These data indicate that placenta and plasma fibronectins are different in their carbohydrate structures and, therefore, suggest the presence of a tissue- or cell-specific mechanism for processing the carbohydrates of this glycoprotein. 相似文献
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Anthony J. Janckila Hadi Yaziji Sheron C. Lear Alvin W. Martin Lung T. Yam 《The Histochemical journal》1996,28(3):195-200
Summary Tartrate-resistant acid phosphatase is an inducible marker of cell differentiation and activation expressed by specialized cells of macrophage lineage and some activated lymphocytes. Clinically, this phosphatase is a diagnostic marker for hairy cell leukaemia and osteoclast activity. The cDNA for this enzyme has been cloned from a placental expression library, yet the cell(s) expressing the enzyme protein has not been determined with certainty. Our laboratories have developed a monoclonal antibody, 9C5, suitable for immunohistochemical localization of tartrate-resistant acid phosphatase in paraffin sections. The purpose of this study was to use antibody 9C5 to identify cells expressing tartrate-resistant acid phosphatase in sections of paraffin-embedded, normal, full-term placenta and to determine if those cells expressed other macrophage markers including CD68(PG-M1 antibody), LN5, lysozyme 1-antitrypsin and 1-antichymotrypsin. Histochemical localization of activity in frozen sections was compared with immunohistochemical localization in paraffin sections of the same tissue specimens. The activity and antigenicity of this enzyme were detected in decidual cells, syncytiotrophoblast, and some macrophages distributed throughout maternal and embryonic tissues, but not in neutrophils. Unlike other tissues previously examined, placenta contains significant numbers of the phosphate-positive cells that are not of macrophage origin. 相似文献
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Rao RM Dharmarajan AM Rao AJ 《Apoptosis : an international journal on programmed cell death》2000,5(1):53-60
Placenta is a transient feto-maternal association that develops during mammalian pregnancies. Human placental tissue during the first trimester of pregnancy is an actively dividing and differentiating tissue, while near term, it represents a fully differentiated unit performing many life-sustaining functions for the fetus. Previous studies have demonstrated that the percentage of placental cells that undergo apoptosis is greater at full term as compared to the first trimester of pregnancy. In this study, we undertook a study aimed at gaining an insight into the kind of genes expressed in the two developmentally distinct stages of gestation ie, the first trimester and term using Differential Display RT-PCR. Cloning and sequencing of one of the differentially expressed cDNAs from term placental tissue revealed that it is a novel gene, referred to as T-18 in the text. In this study, we also examined the regulation of this gene during apoptosis in the human placenta. A model for analysis of placental apoptosis was established by incubating placental villi in serum-free culture medium. It was observed that apoptosis occurred rapidly following incubation of placental villi without tropic support, and the proposed free-radical scavenger, superoxide dismutase (SOD) suppressed apoptosis in the placenta. Interestingly, the levels of T-18 mRNA increased significantly during spontaneous induction of apoptosis and decreased when apoptosis was blocked by SOD. These data clearly suggest that there is a strong correlation between the expression of T-18 and placental apoptosis and that T-18, may play a significant role in this process. Furthermore, the establishment of a defined in vitro explant culture model should facilitate elucidation of factors, which regulate apoptosis in human placenta. 相似文献
11.
Summary Lymphoepithelial complexes known as thymic nurse cells (TNC) have been isolated and described in the thymus of several animal species including man. Most of the investigations on TNC have been carried out in enzymatically digested thymuses in which TNC were isolated by differential sedimentation. In the present study we demonstrate TNC in immunohistochemically stained sections of human thymus as ring-shaped cells completely enclosing thymocytes and localized not only in the cortex, but also at the corticomedullary junction where they have not been previously described. TNC expressed epithelial markers [low and high molecular weight keratins identified by 35H11 and 34E12 monoclonal antibodies, a cortical antigen shared with neuroectodermal neoplasms recognized by the GE2 monoclonal antibody, and tissue polypeptide antigen (TPA:B1)], class II histocompatibility antigens (HLA-DR), and thymosin 1. Double staining experiments with the nuclear proliferation-associated antigen Ki-67 and the cortical epithelium marker GE2 showed that most thymocytes enclosed in these cortical TNC were not proliferating. The antigens expressed by TNC indicate that not only cortical, but also medullary epithelial cells are part of the TNC system. The possible role of TNC in the education and maturation of thymocytes is discussed. 相似文献
12.
Aromatase from human placenta has been purified to homogeneity (MW 55,000). Enzymatic activity can be reconstituted with reductase from pig liver in an aqueous buffer or after incorporation of the enzyme into liposomes. In both cases the enzyme converts androstenedione to estrone and testosterone to estradiol. Aromatase shows a typical CO-spectrum when reduced with dithionite and a type I spectral shift with both substrates. The NH2 terminal amino acid sequence is hydrophobic but shows no homology to that of other cytochromes P-450. Five cysteine peptides have been isolated by HPLC following tryptic digestion of the [14C]-carboxymethylated protein. Amino acid sequences of these peptides reveal that histidine is the carboxy-terminal amino acid of the protein and that significant homology exists with corresponding peptides from other cytochromes P-450. Unique oligonucleotides (62 and 30 MER) synthesized on the basis of a 45 amino acid sequence near the center of the molecular have been used to clone the aromatase gene from a cDNA expression library from human placenta in lambda gt11. 相似文献
13.
Isolation and characterization of thrombomodulin from human placenta 总被引:18,自引:0,他引:18
H H Salem I Maruyama H Ishii P W Majerus 《The Journal of biological chemistry》1984,259(19):12246-12251
Protein C, a plasma protein, is activated by thrombin to a protease (protein Ca) that functions as a physiological anticoagulant. We have isolated thrombomodulin, a cofactor required for the rapid activation of protein C, from human placenta. The purification to near homogeneity was achieved using a crude Triton-solubilized protein fraction from a placental particulate fraction as starting material. Chromatography on DEAE-Sepharose removed 95% of the protein and achieved a 3-fold purification. Thrombomodulin was then isolated by affinity chromatography on a column of thrombin-Sepharose wherein the thrombin had been previously inactivated with diisopropyl fluorophosphate. The final preparation was purified 7,900-fold over the membrane extract with a yield of 7%. We obtained 0.88 mg of thrombomodulin from 100 g of membrane extract derived from 5 kg of placenta. The protein was nearly homogeneous as judged by electrophoresis on 10% acrylamide sodium dodecyl sulfate gels in the presence of 2-mercaptoethanol with an apparent Mr = 105,000. Western blot analysis without 2-mercaptoethanol gave an apparent Mr = 75,000. The protein stimulated the rate of protein C activation by thrombin 800-fold to 10 mol of Ca formed/min/mol of thrombin. Thrombin and thrombomodulin appear to form a 1:1 stoichiometric complex as judged from experiments where we measured the effect of varying the concentration of thrombomodulin with respect to thrombin and the converse, on rates of protein C activation. An antibody directed against rabbit lung thrombomodulin inhibited the human placenta protein by 66%, and the amino acid composition of the proteins from the two species was similar indicating that the proteins are closely related. The apparent Michaelis constant of the thrombin-thrombomodulin complex for protein C is 9.8 microM. The protein C activation reaction requires calcium ions and is maximal at 1 mM Ca2+; higher concentrations inhibited the reaction. Coagulation factor Va and factor Va light chain both stimulate the activity of human thrombomodulin 2- to 3-fold. 相似文献
14.
Lysosomal beta-mannosidase was purified almost 10,000-fold from human placenta. The final preparation showed several protein bands on polyacrylamide gel electrophoresis. Its molecular mass was estimated to be 110 kDa, the optimal pH was 4.5, the Km was 0.56 mM, and the isoelectric point was 4.7. The enzyme was found to bind completely to Con A-Sepharose, and the pI was not changed after neuraminidase treatment. These results indicate that the purified enzyme represents a lysosomal form which contains high mannose type oligosaccharide chains and only a few sialic acids, if any. 相似文献
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An acid phosphatase species which is activated by Fe2+ was purified 3,700-fold from rat spleen by chromatography on columns containing Blue-Sepharose, concanavalin A-Sepharose, Sephadex G-100, and CM-Sephadex. The enzyme hydrolyzed aryl phosphates, nucleoside di- and triphosphates, phosphoproteins, and thiamine pyrophosphate with Km values of 10(-4) to 10(-3) M at an optimal pH of 5.0-5.8. Co-purification of the acid phosphatase and acid phosphoprotein phosphatase indicated that they were identical. The purified enzyme was glycoprotein in nature, showing four heterogeneous forms on acid polyacrylamide gel electrophoresis (pI values, 7.8, 8.0, 8.3, and 8.5), but it gave a molecular weight of 33,000 on sodium dodecyl sulfate-gel electrophoresis and gel permeation chromatography. The enzyme had a purple color (lambda max 545 nm) and contained 2 iron atoms per enzyme molecule. Among reductants, ascorbic acid and Fe2+ were the best activators, although their combined effect was not additive. Fe2+ and ascorbic acid both changed the purple enzyme into the same active form (lambda max 515 nm), giving almost the same kinetic constants for substrates and for inhibitors such as molybdate, phosphate and fluoride. However, low concentrations of Fe2+, from 0.01 mM to 1.0 mM, immediately and reversibly activated the enzyme, whereas high concentrations of ascorbic acid over 1 mM were required for maximal activation, which was slow and irreversible. 相似文献
17.
Purification and characterization of purple acid phosphatase from developing rat bone 总被引:1,自引:0,他引:1
Tartrate-resistant acid phosphatase active on nucleoside di- and triphosphate substrates was isolated from developing rat bone and purified 2500-fold. The enzyme concentration had a purple coloration and activity that was sensitive to reducing agents. Mild reducing agents such as ferrous ion and ascorbic acid caused loss of purple color and increased activity toward substrates severalfold; however, a strong reductant such as dithionite caused loss of both color and activity which were partially restored by addition of ferrous ion and ascorbic acid. Enzyme activity was homogeneous with protein during the final gel permeation steps of chromatography and gave an apparent molecular size of about 40,000 Da. Determination of iron in the most pure preparation revealed the presence of 1.3 atoms of iron per molecule of the tartrate-resistant enzyme E2. Other properties of the purified enzyme include a pI of approximately 9.5 and sensitivity to inhibition by ions of copper, zinc, fluoride, and molybdate. Antibody prepared to the pre-concanavalin A (Con A)-Sepharose purified enzyme reacted with all protein from the Con A step, but it did not react with tartrate-sensitive acid phosphatase from rat bone or with potato acid phosphatase. Purple acid phosphatase from rat bone has many properties that parallel the iron-containing purple acid phosphatases from rat spleen, bovine spleen, and pig uterine secretions. 相似文献
18.
The isolation and partial characterization of glycolipids of normal human leucocytes 总被引:3,自引:2,他引:1
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1. The lipids of purified human leucocytes were extracted with chloroform-methanol and the extract was washed with water. Glycolipids, isolated by Florisil chromatography, were subjected to mild alkaline hydrolysis and the alkali-resistant fraction was fractionated on a silicic acid column. 2. Three classes of glycolipid were separated. The less polar, containing 3.6% of the total glycolipid hexose as galactose, was tentatively identified as ceramide monohexoside. The major glycolipid fraction was characterized as ceramide dihexosides. The more polar glycolipids comprised 1.6% of the total glycolipid hexose as galactose and glucose (in the molar ratio 2:1) and were non-acidic. This class was separated as a mixture containing ninhydrin-positive glycolipids. 3. The ceramide dihexosides taken from two leucocyte preparations accounted for 15.2% and 16.4% by weight of the total lipids. 4. The carbohydrate moiety of the ceramide dihexosides contained galactose and glucose in the molar ratio 2:1. Partial acid hydrolysis and paper chromatography indicated that the hexoses are present as disaccharides, lactose being identified as one of them. 5. Palmitic acid (C(16:0)) and nervonic acid (C(24:1)) were the major fatty acids of this glycolipid. Hydroxy fatty acids were not detected. 相似文献
19.
Fu Sai Chuen Cheuk Yau Chuk Wong Yim Ping Wong Wan Nar Hung Leung Kim Chan Kai Ming 《The journal of histochemistry and cytochemistry》2004,52(9):1151-1157
Cells in tendons are conventionally identified as elongated tenocytes and ovoid tenoblasts, but specific markers for these cells are not available. The roles and interplay of these cells in tendon growth, remodeling, and healing are not well established. Therefore, we proposed to characterize these cells with respect to cell turnover, extracellular matrix metabolism, and expression of growth factors. Here we examined 14 healthy human patellar tendon samples for the expression of various proteins in tenocytes and tenoblasts, which were identified as elongated tendon cells and ovoid tendon cells, respectively. Matrix metalloproteinase 1 (MMP1), procollagen type I (procol I), heat shock protein 47 (hsp47), bone morphogenetic protein 12 (BMP12), 13 (BMP13), and transforming growth factor beta1 (TGFbeta1) were detected by immunohistochemistry (IHC). An image analysis of the IHC staining for proliferation cell nuclear antigen (PCNA) and apoptotic cells was performed to determine the proliferation index and the apoptosis index in elongated and ovoid tendon cells. The ovoid tendon cells expressed higher levels of procol I, hsp47, MMP1, BMP12, BMP13, and TGFbeta1 than the elongated tendon cells. Both the proliferation index and the apoptosis index of ovoid tendon cells were higher than those of the elongated tendon cells. The results suggested that ovoid tendon cells, conventionally recognized as tenoblasts, were more active in matrix remodeling. The expression of BMP 12, BMP13 and TGFbeta1 might be associated with the different cellular activities of tenoblasts and tenocytes. 相似文献