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)

The study was aimed to establish a protocol of isolating and culturing adult mesenchymal stem cells (MSC) from human bone marrow aspirate and identify them by surface antigen analysis and committed differentiation in order to provide an experimental foundation for achieving a therapeutic benefit in applying MSC in hematopoietic stem cell transplantation. MSCs were obtained from fresh human bone marrow aspirate by gradient centrifugation with Percoll (1.073 g/ml) and anchoring culture in L-DMEM with 10% fetal bovine serum by a full medium exchange every 3 days. The MSC surface antigens, including CD34, CD45, CD73, CD105, CD166, were analyzed on FACScan flow cytometer. Under culture in conditioned medium for osteogenesis (the hormone cocktail containing 0.1 micromol/L dexamethasone, 10 mmol/L glycerol-2-phosphate and 50 micromol/L ascorbic acid) and adipogenesis (the cocktail containing 1 micromol/L dexamethasone, 5 mg/L insulin, 0.5 mmol/L 1-methyl-3-isobutylxanthine and 60 micromol/L indomethacin), MSCs committedly differentiated into osteoblasts and adipocytes. The differentiated mesenchymal stem cells were identified by morphological observation and immunohistochemical staining. The results showed that by gradient centrifugation and adhesion culture, MSCs could be isolated and culture-expanded from human bone marrow aspirate. These cells were uniformly negative for CD34, CD45 and positive for CD73, CD105 and CD166. The osteogenic differentiated cells were positive for alkaline phosphatase (ALP) and the adipogenic differentiated cells displayed accumulation of lipid vacuoles, as detected by oil red O. It is concluded that MSC can be isolated and expand-cultured from adult human bone marrow aspirate and committedly differentiate into osteoblasts and adipocytes. MSC primary identification can be accomplished by flow cytometry and induced differentiation. The set of methods in current experiment shows somewhat practical value for basic research and clinical application.
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PMID:[A modified method to isolate and identify the adult mesenchymal stem cells from human bone marrow]. 1680 Sep 42

We recently identified circulating osteoblastic cells using antibodies to osteocalcin (OCN) or alkaline phosphatase (AP). We now provide a more detailed characterization of these cells. Specifically, we demonstrate that 46% of OCN positive (OCN(pos)) cells express AP, and 37% also express the hematopoietic/endothelial marker CD34. Using two different anti-OCN antibodies and forward/side light scatter characteristics by flow cytometry, we find that OCN(pos) cells consist of two distinct populations: one population exhibits low forward/side scatter, consistent with a small cell phenotype with low granularity, and a second population has higher forward/side scatter (larger and more granular cell). The smaller, low granularity population also co-expresses CD34, whereas the larger, more granular cells are CD34 negative. Using samples from 26 male subjects aged 28 to 68 years, we demonstrate that the concentration of circulating OCN(pos) cells increases as a function of age (R=0.59, P=0.002). By contrast, CD34(pos) cells tend to decrease with age (R=-0.31, P=0.18); as a consequence, the ratio of OCN(pos):CD34(pos) cells also increase significantly with age (R=0.54, P=0.022). These findings suggest significant overlap between circulating cells expressing OCN and those expressing the hematopoietic/endothelial marker CD34. Further studies are needed to define the precise role of circulating OCN(pos) cells not only in bone remodeling but rather also potentially in the response to vascular injury.
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PMID:Characterization of circulating osteoblast lineage cells in humans. 1732 Apr 97

Pure white cell aplasia (PWCA) is a rare hematologic disorder characterized by agranulocytosis, a lack of virtually all neutrophil-lineage cells (from neutrophils to myeloblasts) in the bone marrow, and normal erythropoiesis and megakaryocy-topoiesis. We report the first case of PWCA that developed in a patient with primary biliary cirrhosis (PBC). An 83-year-old woman, who had had an elevated serum alkaline phosphatase level and shown positivity for serum antimitochondrial antibodies for 10 years, was referred to us because of a perianal abscess. She had severe neutropenia, and her bone marrow showed typical findings of PWCA. Although methylprednisolone pulse therapy induced complete neutrophil recovery, this effect was transient. She died of infection, and the autopsy confirmed the diagnosis of PBC. In vitro investigations showed that factors inhibitory to normal CD34 cell-derived granulopoiesis were present in the patient's serum.
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PMID:Pure white cell aplasia: report of the first case associated with primary biliary cirrhosis. 1732 84

Adipose tissue contains a stromal vascular fraction (SVF) that is a rich source of adipose tissue-derived stem cells (ASCs). ASCs are multipotent and in vitro-expanded ASCs have the capacity to differentiate, into amongst others, adipocytes, chondrocytes, osteoblasts, and myocytes. For tissue engineering purposes, however, it would be advantageous to use the whole SVF, which can be transplanted without further in vitro selection or expansion steps. Because little is known about the freshly isolated ASCs in the SVF, we phenotypically characterized human freshly isolated ASCs, using flow cytometry. In addition, we investigated whether freshly isolated ASCs have functional properties comparable to cultured ASCs. For this, the differentiation potential of both freshly isolated ASCs and cultured ASCs into the osteogenic pathway was analyzed. Freshly isolated ASCs slightly differed in immunophenotype from cultured ASCs. Contrary to cultured ASCs, freshly isolated ASCs were shown to be highly positive for CD34, and positive for CD117 and HLA-DR. On the other hand, expression of CD105 and especially CD166 on the freshly isolated ASCs was relatively low. After osteogenic stimulation of freshly isolated ASCs, both Runx-2 and CollaI gene expression were significantly increased (p < 0.05). However, there was a difference in the kinetics of gene expression between freshly isolated and cultured ASCs and also between the different SVF isolates tested. There was no difference in alkaline phosphatase activity between freshly isolated ASCs and cultured ASCs. In addition, freshly isolated ASCs stained positive for osteonectin and showed matrix mineralization. We conclude that although there are minor differences in phenotype and kinetics of differentiation between freshly isolated ASCs and cultured ASCs, the use of freshly isolated ASCs for tissue engineering purposes involving bone repair is potentially applicable.
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PMID:Phenotypical and functional characterization of freshly isolated adipose tissue-derived stem cells. 1734 7

Multiorgan dysfunction ensuing from severe heatstroke includes hypotension, hepatic and renal failure, hypercoagulable state, activated inflammation, and cerebral ischemia and injury. We attempted to assess whether human umbilical cord blood-derived CD34+ cell therapy improves survival during experimental heatstroke by attenuating multiorgan dysfunction. Anesthetized rats, immediately after the onset of heatstroke, were divided into 2 major groups and given CD34- or CD34+ cells (1 x 10(5)-5 x 10(5)/mL/kg body weight) i.v. They were exposed to ambient temperature of 43 degrees C to induce heatstroke. Another group of rats were exposed to room temperature (26 degrees C) and used as normothermic controls. Hypotension, hepatic and renal failure (evidenced by increased serum urea nitrogen, creatinine, aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase levels in plasma), hypercoagulable state (evidenced by increased prothrombin time, activated partial thromboplastin time, and D-dimer, and decreased platelet count and protein C in plasma), activated inflammation (evidence by increased TNF-alpha levels in serum), and cerebral dysfunction (evidenced by intracranial hypertension, cerebral hypoperfusion and hypoxia, and cerebral ischemia and injury) were monitored. When the CD34- cell-treated or untreated rats underwent heat stress, their survival time values were found to be 19 to 23 min. Resuscitation with CD34+ cells significantly improved survival time (duration, 63-291 min). As compared with normothermic controls, all CD34- cell-treated heatstroke animals displayed hypotension, hepatic and renal failure, hypercoagulable state, activated inflammation, and cerebral ischemia and injury. However, CD34+ cell therapy significantly caused attenuation of all the above-mentioned heatstroke reactions. In addition, the levels of IL-10 in plasma and glial cell line-derived neurotrophic factors in brain were all significantly increased after CD34+ cell therapy during heatstroke. Our data indicate that CD34+ cell therapy may resuscitate persons who had a heatstroke by reducing multiorgan dysfunction or failure.
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PMID:Human umbilical cord blood-derived CD34+ cells cause attenuation of multiorgan dysfunction during experimental heatstroke. 1750 7

A 14-year-old female domestic shorthair cat was presented to Tehran University Veterinary Teaching Hospital for a persistent fever, anorexia, intermittent vomiting, weight loss and weakness. The main clinical signs were pale mucous membranes, dehydration and splenomegaly. The complete blood count and serum biochemistry tests revealed non-regenerative anaemia, thrombocytopenia and increased alkaline phosphatase (ALP) activity. An enzyme-linked immunosorbent assay (ELISA) test for feline leukaemia virus was negative. Blood film and bone marrow examination revealed a large number of immature eosinophils with variable sizes and numbers of faintly azurophilic granules. Cytochemical staining of blood film demonstrated 70% positive cells for ALP activity. Four percent CD34 positive cells were detected by flow cytometry. As eosinophilic leukaemia is difficult to identify by light microscopy, well-defined diagnostic criteria and the use of flow cytometry and cytochemical staining can improve the ability to correctly diagnose this type of leukaemia in cats.
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PMID:Eosinophilic leukaemia in a cat. 1766 77

Human embryonic stem cells (hESC) face ethical sensitivities and the problem of teratoma formation. Although Wharton's jelly stem cells (WJSC), also of embryonic origin, may not face such ethical concerns, it is not definitely known whether under hESC culture conditions they would be as pluripotent as hESC. WJSC grown on plastic showed two types of morphology (epithelioid and short fibroblastic) in primary culture depending on the culture medium used, and only fibroblastic morphology when passaged. When grown in the presence of hESC medium on mouse feeder cells, they produced atypical colonies containing hESC-like cells with high-nuclear cytoplasmic ratios and prominent nucleoli. They were positive for the hESC markers Tra-1-60, Tra-1-81, SSEA-1, SSEA-4, Oct-4 and alkaline phosphatase, negative for SSEA-3, showed normal karyotypes, developed embryoid body (EB)-like structures, did not produce teratomas in SCID mice and differentiated into neuronal derivatives. They were also positive for the mesenchymal CD markers (CD105, CD90, CD44), negative for CD34 and HLA, and although nine out of 10 embryonic stem cell genomic markers were detectable, these were expressed at low levels. WJSC are thus not as pluripotent as hESC but widely multipotent, and have the advantages of being able to be scaled up easily and not inducing teratomas.
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PMID:Comparative growth behaviour and characterization of stem cells from human Wharton's jelly. 1806 71

Numerous studies support the concept that the nonhemopoietic cells of the bone marrow (BM), are derived from a population of multipotent bone marrow stromal stem cells (BMSSCs), which reside in perivascular niches within the bone marrow. These BMSSCs are thought to give rise not only to more cells that are phenotypically and functionally identical but also differentiated, lineage-committed mesenchymal progeny, including chondrocytes, smooth muscle cells, adipocytes, and osteoblasts. Recently, we have generated a novel monoclonal antibody (mAb) (designated STRO-3) that reacts with a minor subset of STRO-1(+) cells contained within adult BM aspirates and does not react with CD34(+) hemopoietic stem cells. Our results also show that STRO-3 identifies a high proportion of BMSSCs that possess extensive proliferative and multilineage differentiative capacity. Using retroviral expression cloning, we determined that STRO-3 binds to tissue nonspecific alkaline phosphatase (TNSALP), a cell-surface glycoprotein usually associated with cells of the osteoblast lineage. Studies presented here suggest that in addition to being expressed by osteoblasts, TNSALP may also represent a marker of immature BMSSCs in vivo. Finally, these studies suggest that antibodies to TNSALP may be used as an effective single marker of enrichment of BMSSCs from various tissues.
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PMID:A novel monoclonal antibody (STRO-3) identifies an isoform of tissue nonspecific alkaline phosphatase expressed by multipotent bone marrow stromal stem cells. 1841 17

Here we describe the derivation of a novel human embryonic stem (hES) cell line, Endeavour-1 (E1), its four new clonal lines (E1C1, E1C2, E1C3, E1C4), and their characterization. E1 and its clonal lines are propagated on human fetal fibroblasts (HFFs) derived and grown in a largely serum-free medium. Seven inner cell masses were isolated from 34 donated human embryos (27 survived), and one new hES cell line was obtained. E1 has been in culture for over 1 year and possesses all the typical features of stem cells, i.e., expression of stem cell surface markers (stage-specific embryonic antigens SSEA-3 and SSEA-4, and tumor recognition antigens TRA-1-60 and TRA-1-81), staining for alkaline phosphatase, and the presence of the pluripotent gene marker (nanog). This line shows pluripotency both under in vitro and in vivo conditions. E1 has a normal karyotype (46XX). Using our optimized procedure for cloning, four new clonal lines were derived from E1: E1C1, E1C2, E1C3, and E1C4. These clonal lines show normal characteristics: karyotype of that of the parent line (46XX) except for E1C3, which showed reciprocal translocation involving chromosomes 15 and 17; stem cell surface markers SSEA-4, TRA-1-60, and TRA-1-81; and gene expression for pluripotency (Nanog). All of these clonal lines formed embryoid bodies (EBs) in suspension cultures. After seeding, the EBs differentiated, forming cell lineages derived from all three germ layers as indicated by immunolocalization for the ectodermal marker beta-III tubulin, the mesodermal marker CD34, and the endodermal marker alpha-fetoprotein (AFP). There were subtle differences in the expression of these markers between clones. These clonal lines showed pluripotency in vivo. E1 and its clonal lines can differentiate to definitive endoderm after treatment with activin A, and, as indicated by expression of SOX17, FOXa2, and GATA-4 by RT-PCR, there are some subtle differences between these clonal lines. This may help in selecting clonal lines for specific lineage specification and for developing future cell therapy for various diseases.
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PMID:Derivation of a new human embryonic stem cell line, endeavour-1, and its clonal propagation. 1827 99

Currently, mesenchymal stem cells (MSCs) are considered as the most eligible cells for skeletal tissue engineering. However, factors such as difficult stimulation and control of differentiation in vivo hamper their clinical use. In contrast, periosteum or periosteum-derived cells (PCs) are routinely clinically applied for bone and cartilage repair. PCs have often been named MSCs but, although cells of osteochondrogenic lineages arise from MSCs, it is unclear whether periosteum really contains MSCs. Our aim was to investigate the MSC-like character of PCs derived from the periosteum of mastoid bone. Harvesting of periosteum from mastoid bone is easy, so mastoid represents a good source for the isolation of PCs. Therefore, we analysed the MSC-like growth behaviour and the expression of embryonic, ectodermal, endodermal and mesodermal markers by microarray and FACS technology, and the multilineage developmental capacity of human PCs. Regarding clinical relevance, experiments were performed in human serum-supplemented medium. We show that PCs do not express early embryonic stem cell markers such as Oct4 and Nanog, or the marker of haematopoietic stem cells CD34, but express some other MSC markers. Osteogenesis resulted in the formation of calcified matrix, increased alkaline phosphatase activity, and induction of the osteogenic marker gene osteocalcin. Staining of proteoglycans and deposition of type II collagen documented chondrogenic development. As shown for the first time, adipogenic stimulation of mastoid-derived PCs resulted in the formation of lipid droplets and expression of the adipogenic marker genes aP2 and APM1. These results suggest MSC-like PCs from mastoid as candidates for therapy of complex skeletal defects.
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PMID:Human mastoid periosteum-derived stem cells: promising candidates for skeletal tissue engineering. 1838 54


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