Localized regions of egg cytoplasm that promote expression of endoderm-specific alkaline phosphatase in embryos of the ascidian Halocynthia roretzi

Development ◽  
1993 ◽  
Vol 118 (1) ◽  
pp. 1-7 ◽  
Author(s):  
H. Nishida
Keyword(s):  
Alkaline Phosphatase ◽  
Histochemical Staining ◽  
Halocynthia Roretzi ◽  
Endoderm Differentiation ◽  
Initial Event ◽  
Cytoplasmic Factors ◽  
Early Embryos ◽  
Vegetal Pole ◽  
Cytoplasmic Determinants ◽  
Cytoplasmic Transfer

Embryogenesis in ascidians is known to be of the mosaic type, a property that suggests the presence of cytoplasmic factors in the egg which are responsible for specification of the developmental fates of early blastomeres. Endoderm cells are present in the trunk region of tadpole larvae, and these cells specifically express alkaline phosphatase (AP). Endoderm cells originate exclusively from blastomeres of the vegetal hemisphere of early embryos. To obtain direct evidence for cytoplasmic determinants of endoderm specification, we carried out cytoplasmic-transfer experiments by fusing blastomeres and cytoplasmic fragments from various regions. Initially, presumptive-epidermis blastomeres (blastomeres from the animal hemisphere) were fused to cytoplasmic fragments from various regions of blastomeres of 8-cell embryos of Halocynthia roretzi, and development of endoderm cells was monitored by histochemical staining for AP. AP activity was observed only when presumptive-epidermis blastomeres were fused with cytoplasmic fragments from the presumptive-endoderm blastomeres. The results suggest that cytoplasmic factors that promote the initial event of endoderm differentiation (endoderm determinants) are present in endoderm-lineage blastomeres. Next, to examine the presence and localization of endoderm determinants in the egg, cytoplasmic fragments from various regions of unfertilized and fertilized eggs were fused with the presumptive-epidermis blastomeres. The results suggest that endoderm determinants are already present in unfertilized eggs, and that they are segregated by movements of the ooplasm after fertilization. Initially, these determinants move to the vegetal pole of the egg. Then, prior to the first cleavage, their distribution extends in the equatorial direction, namely, to the entire vegetal hemisphere from which future endoderm-lineage blastomeres are formed.

Regionality of egg cytoplasm that promotes muscle differentiation in embryo of the ascidian, Halocynthia roretzi

Development ◽  
1992 ◽  
Vol 116 (3) ◽  
pp. 521-529 ◽  
Author(s):  
H. Nishida
Keyword(s):  
Muscle Cells ◽  
Muscle Differentiation ◽  
Posterior Region ◽  
Second Phase ◽  
Halocynthia Roretzi ◽  
Early Embryos ◽  
Vegetal Pole ◽  
Restricted Region ◽  
Ooplasmic Segregation ◽  
Cytoplasmic Determinants

Development of ascidians occurs in typical mosaic fashion: blastomeres isolated from early embryos differentiate into tissues according to their normal fates, an indication that cytoplasmic determinants exist in early blastomeres. To provide direct evidence for such cytoplasmic determinants, we have devised methods for fusing blastomeres and cytoplasmic fragments from various regions. (1) Presumptive-epidermis blastomeres were fused to cytoplasmic fragments from various regions of blastomeres of 8-cell embryos of Halocynthia roretzi and development of muscle cells was monitored by an antibody to ascidian myosin. Muscle differentiation was observed only when presumptive-epidermis blastomeres were fused with fragments from the posterior region of B4.1 (posterior-vegetal) blastomeres, the normal progenitor of muscle cells. The results indicate that muscle determinants are present and localized in the cytoplasm that enters muscle-lineage cells. (2) To investigate the presence and localization of muscle determinants in the egg, cytoplasmic fragments from various regions of unfertilized and fertilized eggs were fused with the presumptive- epidermis blastomeres, and formation of muscle cells was assessed by monitoring myosin, actin and acetylcholinesterase expression. These proteins were expressed only when cytoplasm from a restricted region of the eggs, i.e. the vegetal region, after the first phase of ooplasmic segregation, and posterior region, after the second phase of segregation, were fused. Based on these experiments, it is suggested that muscle determinants are segregated by ooplasmic movements after fertilization. They move initially to the vegetal pole of the egg and, prior to first cleavage, to the posterior region from whence future muscle-lineage blastomeres are formed. The inferred movements of muscle determinants correspond to those of the myoplasm, a microscopically visible portion of the egg cytoplasm.

Localization of egg cytoplasm that promotes differentiation to epidermis in embryos of the ascidian Halocynthia roretzi

Development ◽  
1994 ◽  
Vol 120 (2) ◽  
pp. 235-243 ◽  
Author(s):  
H. Nishida
Keyword(s):  
Specific Antigen ◽  
Equatorial Region ◽  
Epidermal Cells ◽  
Second Phase ◽  
Halocynthia Roretzi ◽  
Cytoplasmic Factors ◽  
Early Embryos ◽  
Ooplasmic Segregation ◽  
Anucleate Cell ◽  
Cell Fragments

Embryogenesis in ascidians is of the mosaic type. This property suggests the presence of cytoplasmic factors in the egg that are responsible for specification of the developmental fates of early blastomeres. The epidermal cells that surround the entire tadpole larva originate exclusively from blastomeres of the animal hemisphere of early embryos. To obtain direct evidence for cytoplasmic determinants of epidermis fate, we carried out cytoplasmic transfer experiments by fusing blastomeres and anucleate cell fragments from various regions of eggs and embryos. Initially, presumptive non-epidermis blastomeres (blastomeres from the vegetal hemisphere) were fused to cytoplasmic fragments from various regions of blastomeres of 8-cell embryos of Halocynthia roretzi, and development of epidermal cells was monitored by following the expression of an epidermis- specific antigen, as well as by observations of morphology and the secretion of larval tunic materials. Formation of epidermis was observed when vegetal blastomeres were fused with cytoplasmic fragments from the presumptive epidermis blastomeres. The results suggested that cytoplasmic factors that promoted epidermis differentiation (epidermis determinants) were present in epidermis progenitors. Vegetal blastomeres only manifested this change in fate when fused with cytoplasmic fragments of roughly equal or larger size. Next, to examine the presence and localization of epidermis determinants in the uncleaved egg, cytoplasmic fragments from various regions of unfertilized and fertilized eggs were fused with the vegetal blastomeres. The results suggested that epidermis determinants were already present in unfertilized eggs and that they were segregated by movements of the ooplasm after fertilization. After the first phase of ooplasmic segregation, these determinants were widely distributed, with the highest activity being located in the equatorial region. There were no indications of regional differences in the activity within the equatorial region of eggs at this stage. After the second phase of ooplasmic segregation, prior to the first cleavage, the activity moved in the animal direction, namely, to the animal hemisphere, from which future epidermis-lineage blastomeres are normally formed.

3D bioprinting of mesenchymal stem cells and endothelial cells in an alginate-gelatin-based bioink

2021 ◽  
Author(s):  
Sindhuja D Eswaramoorthy ◽  
Nandini Dhiman ◽  
Akshay Joshi ◽  
Subha N Rath
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Alkaline Phosphatase ◽  
Endothelial Cells ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Alkaline Phosphatase Activity ◽  
Histochemical Staining ◽  
Vital Role ◽  
3D Bioprinting

Aim: Bioink is one of the essential factors in 3D bioprinting that determines the fate of cells, in our case, umbilical cord-derived mesenchymal stem cells (UMSC). The aim was to determine if the presence of the osteoinductive factors in the bioink enhances osteodifferentiation as compared with adding them postprinting and if the UMSC and endothelial cells (EC) coculture result in better osteodifferentiation. Materials & methods: Alginate-gelatin along with UMSC–EC were bioprinted using an extrusion 3D bioprinter. Results & conclusion: The UMSC–EC interaction, as well as intrinsic addition of the differentiation components in the bioink, were observed to play a vital role in increasing the osteogenic differentiation as shown by the histochemical staining, alkaline phosphatase activity and gene expression of osteogenic markers.

Canine corticosteroid-induced alkaline phosphatase in serum was solubilized by phospholipase activity in vivo

1995 ◽  
Vol 269 (2) ◽  
pp. G278-G286 ◽  
Author(s):  
P. F. Solter ◽  
W. E. Hoffmann
Keyword(s):  
Alkaline Phosphatase ◽  
Polyacrylamide Gel Electrophoresis ◽  
Histochemical Staining ◽  
Gas Chromatography Mass Spectrometry ◽  
Phospholipase Activity ◽  
Glycosyl Phosphatidylinositol ◽  
Native Polyacrylamide Gel Electrophoresis ◽  
Membrane Attachment ◽  
Paired Serum

In this study, gas chromatography/mass spectrometry revealed the presence of stoichiometric amounts of myo-inositol in association with serum corticosteroid-induced isozyme of alkaline phosphatase (CALP) in canine serum. Such remnants are consistent with prior membrane attachment of serum CALP and its release into serum by endogenous phospholipase activity. Serum CALP was further shown to behave similarly to CALP released from hepatocyte membranes by glycosyl phosphatidylinositol phospholipase D (GPI-PLD) and differently from CALP solubilized by GPI-phospholipase C (PLC) on both native polyacrylamide gel electrophoresis and Western blot analysis using anti-cross-reacting determinant antibody. In addition to bile canalicular surfaces, CALP activity was found over hepatocyte sinusoidal surfaces by histochemical staining of canine liver sections. A significantly higher ratio of CALP to total alkaline phosphatase activity was observed in serum as opposed to bile in 10 of 11 paired serum and bile samples from dogs. This suggested that bile is not likely to be the source of serum CALP and is consistent with the release of CALP from hepatocyte basolateral surfaces directly into serum. It was concluded that serum CALP was once membrane bound and was released by phospholipase activity into serum. Our findings are consistent with release of CALP from the sinusoidal surfaces of hepatocytes into serum either by endogenous GPI-PLD activity or release by GPI-PLC followed by modification of the phosphatidylinositol remnant in vivo.

Kid-1 expression is high in differentiated renal proximal tubule cells and suppressed in cyst epithelia

1998 ◽  
Vol 275 (6) ◽  
pp. F928-F937 ◽  
Author(s):  
Ralph Witzgall ◽  
Nicholas Obermüller ◽  
Ulrike Bölitz ◽  
James P. Calvet ◽  
Benjamin D. Cowley ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Proximal Tubule ◽  
Rat Kidney ◽  
Histochemical Staining ◽  
Renal Development ◽  
Mrna Levels ◽  
Adult Rat ◽  
Renal Proximal Tubule Cells ◽  
Nephron Development ◽  
Brush Border Enzyme

The cDNA coding for the transcriptional repressor protein Kid-1 was cloned in a screen for zinc finger proteins, which are regulated during renal development and after renal ischemia. Kid-1 mRNA levels increase in the course of postnatal renal development and decrease after acute renal injury caused by ischemia or administration of folic acid. We have raised a monoclonal anti-Kid-1 antibody and demonstrate that the Kid-1 protein is strongly expressed in the proximal tubule of the adult rat kidney. During nephron development, the Kid-1 protein appears after the S-shaped body stage concomitantly with the brush-border enzyme alkaline phosphatase. In two animal models of polycystic kidney disease, the expression of Kid-1 is downregulated. The loss of expression of Kid-1 in cyst wall cells correlates with the loss of alkaline phosphatase histochemical staining. Kid-1 mRNA levels are also reduced in rodent renal cell carcinomas, another condition characterized by epithelial cell dedifferentiation and increased proliferation. We propose that Kid-1 plays an important role during the differentiation of the proximal tubule.

CCK-8 infusion increases plasma LMW alkaline phosphatase coincident with enterohepatic circulation of bile acids

1997 ◽  
Vol 273 (2) ◽  
pp. G381-G388 ◽  
Author(s):  
P. F. Solter ◽  
W. E. Hoffmann ◽  
M. D. Chambers ◽  
D. J. Schaeffer
Keyword(s):  
Alkaline Phosphatase ◽  
Bile Acids ◽  
Enterohepatic Circulation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Histochemical Staining ◽  
Gallbladder Emptying ◽  
Hydrophobic Membrane ◽  
Alp Activity ◽  
Exclusion Chromatography ◽  
Sinusoidal Surface

Bile acids may facilitate the release of liver alkaline phosphatase (ALP) from its hydrophobic membrane anchor. The purpose of this study was to determine whether such a facilitatory role could be observed during the enterohepatic circulation of bile acids in dogs. Increased hepatic ALP activity was induced in four dogs by daily injections of 4 mg.kg-1.day-1 of prednisone for 10 days. Intravenous infusions of cholecystokinin octapeptide (CCK-8) were given before treatment and on treatment days 3, 5, 7, and 10 to induce gallbladder emptying and the enterohepatic circulation of bile acids. Blood samples were taken hourly for 4 h before and for 4 h after CCK-8 infusion. These showed that plasma ALP activity increased significantly only after CCK-8 infusion. Gel exclusion chromatography, polyacrylamide gel electrophoresis, and octyl Sepharose phase separation showed that the increased ALP activity was a hydrophilic, low-molecular-weight (LMW) isoform, which is consistent with phospholipase release. Histochemical staining of endogenous ALP activity showed increased ALP activity over sinusoidal surfaces of prednisone-treated dogs. There was also an increased serum-to-tissue ratio of ALP activity in the prednisone-treated dogs, suggestive of increased release of ALP into blood. It was concluded that bile acids probably play a facilitatory role in the enzymatic release of ALP from the sinusoidal surface of hepatocytes, which may be accentuated by the presence of increased amounts of ALP on the sinusoidal surface in some disease states.

HISTOLOGICAL AND HISTOCHEMICAL OBSERVATIONS ON THE REPRODUCTIVE ORGANS OF Archachatina marginata ovum (GASTROPODA: ACHATINIDAE) AT DIFFERENT REPRODUCTIVE STATES

2017 ◽  
Vol 16 (1) ◽  
pp. 40-51
Author(s):  
S. I. OLA ◽  
O. AKINLADE ◽  
D. O. ADEYEMI
Keyword(s):  
Alkaline Phosphatase ◽  
Morphological Variation ◽  
Reproductive System ◽  
Growth And Development ◽  
Reproductive Organs ◽  
Histochemical Staining ◽  
Phosphatase Activities ◽  
Hermaphroditic Duct

The histological and histochemical variation in association with morphological variation in the repro-ductive system of Archachatina marginata ovum was the target of this investigation. Forty- five snails were dissected and categorized into 5 different reproductive stages (low mating, high mating, high egg, gravid and post reproductive). The reproductive tracts which include: hermaphroditic duct, albu-men gland, spermoviduct and spermatheca and the ovotestis were processed for histological and histochemical staining. There were some variations in the architecture of the reproductive organs be-tween the active (high mating, high egg and gravid) and non active stages (low mating and post repro-ductive) states. The active states were generally associated with colloidal or granular secretions. Gly-cogen and alkaline phosphatase activities were associated together throughout the epithelium of the reproductive system of A. marginata ovum and they were more strongly indicated in tissues that are intimately connected to the growth and development of gametes. It was concluded that morphological variation in the secreting glands of the reproductive system of A. marginata ovum is closely associated with changes in the functional secretory activities of the glands.

Mesoderm and endoderm differentiation in animal cap explants: identification of the HNF4-binding site as an activin A responsive element in the Xenopus HNF1alpha promoter

Development ◽  
1996 ◽  
Vol 122 (6) ◽  
pp. 1975-1984 ◽  
Author(s):  
H. Weber ◽  
B. Holewa ◽  
E.A. Jones ◽  
G.U. Ryffel
Keyword(s):  
Transcription Factor ◽  
Retinoic Acid ◽  
Gall Bladder ◽  
Binding Site ◽  
Activin A ◽  
Promoter Element ◽  
Gene Encoding ◽  
Endoderm Differentiation ◽  
Vegetal Pole ◽  
Responsive Element

The gene encoding the tissue-specific transcription factor HNF1alpha (LFB1) is transcriptionally activated shortly after mid-blastula transition in Xenopus embryos. We have now shown that the HNF1alpha protein is localized in the nuclei of the liver, gall bladder, gut and pronephros of the developing larvae. In animal cap explants treated with activin A together with retinoic acid, we induced HNF1alpha in pronephric tubules and epithelial gut cells, i.e. in mesodermal as well as in endodermal tissues. HNF1alpha can also be induced by activin A, but not by retinoic acid alone. To define the promoter element responding to the activin A signal, we injected various HNF1alpha promoter luciferase constructs into fertilized eggs and cultured the isolated animal caps in the presence of activin A. From the activity profiles of the promoter mutants used, we identified the HNF4-binding site as an activin-A-responsive element. As HNF4 is a maternal protein in Xenopus and localized in an animal-to-vegetal gradient in the cleaving embryo, we speculate that the activin A signal emanating from the vegetal pole cooperates with the maternal transcription factor HNF4 to define the embryonic regions expressing HNF1alpha.

scholarly journals Use of tissue-free glycol methacrylate sections as semi-permeable membranes: a simple way to shorten incubation times and to improve localization in enzyme histochemistry.

1989 ◽  
Vol 37 (2) ◽  
pp. 173-176 ◽  
Author(s):  
P O Gerrits ◽  
R W Horobin ◽  
M J Hardonk
Keyword(s):  
Alkaline Phosphatase ◽  
Azo Dye ◽  
Enzyme Histochemistry ◽  
Diazonium Salt ◽  
Histochemical Staining ◽  
Semipermeable Membranes ◽  
Large Molecules ◽  
Glycol Methacrylate ◽  
Tissue Sections ◽  
Antibody Staining

Placing 2-microns sections of tissue-free glycol methacrylate on top of tissue sections is a simple way of forming semipermeable membranes to enhance enzyme histochemical staining. For demonstrating alkaline phosphatase in glycol methacrylate-embedded kidney by a standard azo dye method, such membranes enabled incubation times to be reduced to 1-2 hr, with azo dye reaction product being more crisply localized as compared to sections stained without membranes. Such effects are possible because the membranes are highly permeable to small molecules (e.g., substrate and diazonium salt), slightly permeable to molecules of moderate size (e.g., the final reaction product), and impermeable to large molecules (e.g., alkaline phosphatase and other tissue biopolymers). The implications of these findings for enzyme histochemistry and for enzyme-labeled antibody staining are discussed.

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