Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We explored the possibility of simultaneous application of histochemical and immunohistochemical staining techniques on the same paraffin-embedded human tissue section. Conventional histological stains (PAS, Alcian, Alcian-PAS, Van Gieson, Gomori silver impregnation, and Giemsa) were used in association with a battery of markers (keratins, leucocyte common antigen, S-100 protein, Factor VIII-related antigen) that are widely employed in diagnostic and experimental studies. We found that the best procedure was to perform immunostaining before the histochemical reaction, as this enables all the other possible combinations to be carried out. In addition, several detection systems, such as peroxidase-anti-peroxidase (PAP), alkaline phosphatase-anti-alkaline phosphatase (APAAP), and avidin-biotin complex (ABC), were tested and all gave consistent results. Some minor modifications of the histological staining methods were necessary, but the current immunohistochemical techniques could be used as established. Preliminary findings indicate that immunohistochemistry can be combined with histochemistry techniques by means of a relatively simple procedure whose only disadvantage is the time required to carry out the double staining.
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PMID:Simultaneous visualization of immunodetected antigens and tissue components revealed by non-enzymatic histochemical stains. 128 Jun 67

Two endothelial cell lines were derived from grafts of the central nervous system using retrovirus mediated gene transfer to introduce the polyoma middle-T oncogene into fetal rat brain endothelial cells and transplantation of these cells into adult rat brain. In this report, we further characterize these cells and the effect of dexamethasone on the expression of specific enzymatic markers. These cells take up acetylated low density lipoprotein, leucine, and glucose, and express Factor VIII-related antigen, angiotensin converting enzyme, alkaline phosphatase, gamma-glutamyltranspeptidase, and as yet undescribed aminopeptidase A and B-like enzymes. When grown on semi-permeable membranes, these transformed cells do not spontaneously retain small hydrophilic molecules. In culture, one of the lines (EC 193) forms a confluent monolayer of spindle-shaped cells homogenously expressing gamma-glutamyltranspeptidase at a level comparable to primary cells. The other cell line (EC 219) grows as clusters of elongated cells, and gamma-glutamyltranspeptidase activity is expressed mainly in cells forming the clusters. This clustered pattern changes to a confluent one after culture on type-I collagen. Dexamethasone increases angiotensin-converting enzyme activity, and decreases the expression of gamma-glutamyltranspeptidase and aminopeptidase A, whereas the aminopeptidase B activity is little modified. Inhibition of aminopeptidase A activity by amastatin, potentiates angiotensin II effects on DNA synthesis. These results indicate that retrovirally transformed brain endothelial cells are a useful model for studying the blood-brain barrier in vitro and that dexamethasone, an agent with the potential to reduce brain edema, directly affects some blood-brain barrier properties in these endothelial cell lines.
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PMID:Dexamethasone selectively regulates the activity of enzymatic markers of cerebral endothelial cell lines. 135 67

A method to culture rat cerebral microvascular endothelial cells (RCMECs) was developed and adapted to concurrently obtain cultures of rat aortic endothelial cells (RAECs) without subculturing, cloning, or "weeding." The attachment and growth requirements of endothelial cell clusters from isolated brain microvessels were first evaluated. RCMECs required fetal bovine serum to attach efficiently. Attachment and growth also depended on the matrix provided (fibronectin approximately laminin much greater than gelatin greater than poly-D-lysine approximately Matrigel greater than hyaluronic acid approximately plastic) and the presence of endothelial cell growth supplement and heparin in the growth medium. Non-endothelial cells are removed by allowing these cells to attach to a matrix that RCMECs attach to poorly (e.g., poly-D-lysine) and then transferring isolated endothelial cell clusters to fibronectin-coated dishes. These cell cultures, labeled with 1,1'-dioctadecyl-3,3,3',3'-tetramethyl-indocarboxyamine perchlorate (DiI-Ac-LDL) and analyzed using flow cytometry, were 97.7 +/- 2.6% (n = 6) pure. By excluding those portions designed to isolate brain microvessels, the method was adapted to obtain RAEC cultures. RAECs do not isolate as clusters and have different morphology in culture, but respond similarly to matrices and growth medium supplements. RCMECs and RAECs have Factor VIII antigen, accumulate DiI-Ac-LDL, contain Weibel-Palade bodies, and have complex junctional structures. The activities of gamma-glutamyl transferase and alkaline phosphatase were measured as a function of time in culture. RCMECs had higher enzymatic activity than RAECs. In both RCMECs and RAECs enzyme activity decreased with time in culture. The function of endothelial cells is specialized depending on its location. This culture method allows comparison of two endothelial cell cultures obtained using very similar culture conditions, and describes their initial characterization. These cultures may provide a model system to study specialized endothelial cell functions and endothelial cell differentiation.
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PMID:A comparison of primary cultures of rat cerebral microvascular endothelial cells to rat aortic endothelial cells. 185 57

To demonstrate the degree of involvement of endothelial cells in cytomegalovirus (CMV) infection of the gastrointestinal tract we have stained sections from gastrointestinal specimens that showed inclusion bodies on hematoxylin-eosin staining. Factor VIII was first detected using a rabbit anti-factor VIII primary antibody and an alkaline phosphatase-labeled sheep anti-rabbit secondary antibody. The CMV was then visualized with a biotin-labeled CMV probe detected by a streptavidin peroxidase technique with aminoethyl carbazole as the chromogen. Factor VIII staining was a bright blue and CMV a brick red. The specimens included one small-bowel resection and four colonic resections, as well as an esophageal biopsy. The patients' diagnoses included bone marrow transplant recipient, acquired immunodeficiency syndrome, ulcerative colitis, and renal transplant recipient. Cells positive for both CMV and factor VIII ranged from 35% to 60% of positive cells in a representative section, and the relative percentages (mean +/- SE) for cell type for infected cells were: endothelial, 48.9 +/- 4.5; vascular luminal (factor VIII negative), 6.1 +/- 1.7; perivascular (factor VIII negative in vascular wall), 16.2 +/- 3.2; and other cell (non-vascular factor VIII negative), 28.9 +/- 5.1. These findings and clustering of infected cells around the vessels provide evidence that CMV infection of the gastrointestinal tract is primarily vasculitic and related to infection of endothelial cells.
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PMID:Cytomegalovirus infection of gastrointestinal endothelium demonstrated by simultaneous nucleic acid hybridization and immunohistochemistry. 254 Jul 25

This study reviews data on the histogenesis of Kaposi's sarcoma and angiosarcoma derived from clinical features, histology, electron microscopy, enzyme histochemistry, and immunochemistry of both diseases. Their hemorrhagic clinical appearance contrasts the predominantly lymphatic histologic features of vessels in early lesions. Investigations performed to resolve the debate whether these tumors arise from blood vessel or lymphatic endothelium show remarkably similar results for both conditions. Electron microscopy reveals Weibel - Palade bodies in a minority of cases, but features consistent with less well-differentiated blood vessel endothelium may be seen in a greater proportion of tumors. Enzyme histochemistry generally shows absence of adenosine triphosphatase and alkaline phosphatase in tumor cells; a pattern of enzymes similar to that found in normal lymphatic endothelium. Conflicting data arises from the large number of immunohistochemical studies performed on both conditions. Factor VIII-related antigen and Ulex europaeus agglutinin-I have been most frequently employed, but the specificity of these agents for blood vessel endothelium is debatable. Panendothelial markers show consistent labeling of both tumors, but marker studies employing a wide range of monoclonal antibodies specific for blood vessel endothelium have shown occasional positive labeling of tumor cells. A number of studies have claimed absence of labeling with specific blood vessel monoclonal antibodies, but at present no study employing a specific marker for lymphatic endothelium has been reported. Although the demonstration of specific markers for blood vessel endothelium in these tumors has been variable, the data would be compatible with lesions arising from undifferentiated stem cells that proliferate with varying degrees of differentiation toward blood vessel endothelium. An alternative hypothesis for the histogenesis of Kaposi's sarcoma would be one of multicentric hyperplasia containing lymphatic venular anastamoses with elements of both lymphatic and blood vessel endothelium.
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PMID:Histogenesis of Kaposi's sarcoma and angiosarcoma of the face and the scalp. 266 17

Tubular cells have been isolated, characterized and cultured from more than 70 adult cadaver kidneys (postmortem time less than or equal to 12 hr.). Confluent monolayers were observed at 7 days after seeding (10(6) cells/ml.) and cells demonstrating normal human karyotypes have been passaged up to 6 times. Primary isolates and monolayer cultures were negative for Factor VIII activity, and strongly positive for gamma-glutamyltransferase activity and keratin. Ultrastructurally primary isolates consisted of cells with numerous mitochondria, microvilli, cytoplasmic filaments and well-developed endocytotic apparati. Monolayer cultures examined at 7, 14, 21 and 72 days demonstrated less prominent microvilli and the additional structures of desmosomes and cell junctions. Membrane-associated and cytosolic enzyme activities were measured up to 28 days in culture. The membrane-associated enzymes gamma-glutamyltransferase and alkaline phosphatase both exhibited approximately 10-fold decreases in activity during the 1st 7 days in culture. There was an approximately 5-fold increase in pyruvate kinase activity during the same time period, while fructose-1,6-bisphosphatase activity exhibited a 5-fold decrease. Glucose-6-phosphatase activity did not change during the 28 day culture period examined. From 7 to 28 days no further changes were noted in any of the enzyme activities measured. Decreased membrane-associated enzyme activity corresponded to the ultrastructural observation of less prominent microvilli. Increases in glycolytic enzyme activity and decreases in gluconeogenic enzyme activity may reflect the presence of glucose in the culture medium. The morphologic and biochemical evidence suggests that primary isolates and cultures are proximal tubule cells which should provide a well-defined in vitro human system for future studies.
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PMID:Isolation, culture and characterization of human renal tubular cells. 285 5

Previous studies utilizing enzyme histochemistry, electron microscopy, and immunohistochemistry have failed to establish the cell of origin in Kaposi's sarcoma. The authors have rigorously tested the prevailing hypothesis that the lesion defined as Kaposi's sarcoma is derived from vascular endothelial cells. They use seven markers to characterize endothelial cells: three antigens (Factor VIII-related antigen, HLA-DR/Ia, macrophage/endothelial antigens), three enzymes (5'-nucleotidase, ATPase, alkaline phosphatase), and lectin binding (Ulex europaeus I). They applied the markers first to normal skin and lymph node, and then to biopsy specimens from 40 patients with Kaposi's sarcoma. Normal blood vessel endothelium was positive for all seven markers, but normal lymphatic endothelium was negative for all of the markers except 5'-nucleotidase and Ulex europaeus lectin. The neoplastic cells in 40 cases of Kaposi's sarcoma closely resembled those of normal lymphatic endothelium but not those of blood vessel endothelium. This suggests that Kaposi's sarcoma may originate in lymphatic endothelium.
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PMID:Evidence for the origin of Kaposi's sarcoma from lymphatic endothelium. 298 60

Human fetal lung homogenates contain an inactive form of renin which may be revealed by trypsin treatment. When activated, this form of renin has some biochemical similarities with fetal kidney renin: the pH optimum of fetal lung renin is approximately 6.5; it is bound by Affigel Blue affinity chromatography resin; and is inhibited by a monoclonal antibody (R-3-36-16) raised to human kidney renin. Inactive renin, partially-purified from both fetal kidney and lung, differs from this in that the renal form is of low-molecular weight (LMW, 45,000 daltons), whereas that from fetal lung is of high molecular weight (HMW, 58,000 daltons). Using a sensitive alkaline phosphatase-anti-alkaline phosphatase (APAAP) procedure with a polyclonal anti-renin antibody (R-15), immunoreactive renin in fetal lung was found in vessels in mesenchyme between airways. The pattern of staining was distributed similarly to Factor VIII-related antigen, suggesting localization in endothelial cells.
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PMID:Renin in human fetal lung--a biochemical and immunohistochemical study. 305 97

Previous studies on mice have revealed that the Griffonia simplicifolia I (GSI) lectin selectively binds to capillaries in a number of microvascular beds. These observations suggest that the lectin might be a suitable microvascular marker for physiological studies of skeletal muscle, particularly when fluorescent visualization of vessels is desired independently of their perfusion status. Since species and strain heterogeneity has been demonstrated for certain lectins associated with the microcirculatory vessels, lectin binding was studied in a number of muscles taken from the major species of mammals used for experimental purposes. Staining of cryostat sections confirmed the utility of GSI as a marker for capillaries from muscle of mice, rats, hamsters, rabbits, dogs, and monkeys. Differential staining of arterioles and veins was revealed by double labeling with GSI and antisera to Factor VIII-related antigen. Double labeling for GSI binding and alkaline phosphatase activity revealed that the GSI method detects many more capillaries and terminal arterioles than does the alkaline phosphatase method. GSI binding to unfixed whole mounts of thin skeletal muscles (hamster cheek pouch, mouse diaphragm, and rat cremaster) was studied to determine whether the GSI lectin would be a suitable marker for intravital studies. An extensive microvascular bed, including terminal arterioles, venules, and capillaries, was revealed which could be visualized in the complete absence of perfusion with fluorescent markers. These observations suggest that the GSI lectin may be extremely useful as a probe for the microcirculation of skeletal muscle in many types of physiological experiments.
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PMID:Griffonia simplicifolia I: fluorescent tracer for microcirculatory vessels in nonperfused thin muscles and sectioned muscle. 314

In contrast to the dose-rate independent X ray killing observed with human bone marrow hematopoietic stem cells, bone marrow adherent stromal cells from the same fresh marrow harvests demonstrate increased radiation resistance at low dose rate (LDR) (5 cGy/min), compared to high dose rate (HDR) irradiation (120-200 cGy/min). Physiologic changes observed in plateau phase bone marrow cells after LDR irradiation in vivo and in vitro suggested that marrow stromal cells might be heterogeneous in LDR irradiation repair. Five permanent clonal bone marrow stromal lines were derived from a single human marrow donor. Each cell line was positive for markers of fibroblasts including: immunohistochemically detectable fibronectin, collagen, acid phosphatase, and nonspecific esterase, and was negative for Factor VIII, alkaline phosphatase, lysozyme and several markers of marrow macrophages. The x-irradiation survival curve of each cell line was determined at LDR and HDR in vitro. Cell lines KM102, KM103, KM104, and KM105 each demonstrated a significant (p less than .05) increase in radioresistance at LDR (D0 = 142, n = 2.9; D0 = 131, n = 2.5; D0 = 145, n = 2.1 and D0 = 127, n = 2.1 respectively) compared to HDR: (D0 = 111, n = 2.1; D0 = 94, n = 3.5; D0 = 99, n = 3.5 and D0 = 95, n = 2.1 respectively). In contrast, cell line KM101 demonstrated no significant change in radiosensitivity relative to dose rate at LDR (D0 = 113, n = 3.3) compared to HDR, D0 = 114, n = 3.3. Cell line KM101 was more supportive than the other lines of cocultivated hemopoietic cells in vitro. Subclones of KM101 and KM104 selected by retroviral vector transfer of the neor gene for growth in the antibiotic neomycin-analogue G418, maintained the stably associated radiobiologic properties of each parent clonal line. These data indicate significant heterogeneity in the LDR irradiation response of clonal stromal cell lines derived from human bone marrow.
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PMID:Radiosensitivity of permanent human bone marrow stromal cell lines: effect of dose rate. 318 48


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