Enhanced Melanization of Harding-Passey Mouse Melanoma Cells Following Treatment with Exogenous Melanosomes in Monolayer Culture

1979 ◽  
Vol 34 (1-2) ◽  
pp. 124-130 ◽  
Author(s):  
H. B. Leising ◽  
D.O. Schachtschabel
Keyword(s):  
Monolayer Culture ◽  
Actinomycin D ◽  
Melanoma Cells ◽  
Tyrosinase Activity ◽  
The Other ◽  
Monolayer Cultures ◽  
Protein Portion ◽  
Inhibitor Treatment

Abstract Purified melanosomes isolated from subcutaneously growing Harding-Passey melanomas of NMRI-mice were labeled either in vitro with [14C] tyrosine or [14C]DOPA in the melanin portion, or in vivo in the melanin and protein portion following i. p. injection of [14C] tyrosine. Treatment of monolayer cultures of Harding-Passey melanoma cells (HPM-73 line) with such labeled melanosomes resulted in rapid uptake of label during the first 4 h which leveled off thereafter. A portion of the “incorporated” label could be removed by a 15 min chase with unlabeled melanosomes.Uptake of labeled melanosomes by HPM-73 cells was followed by increased cellular melaniza­tion which was not only due to melanin derived from incorporated melanosomes but primarily to newly formed melanin. Tyrosinase activity was elevated in melanosome-treated cells. Tyrosinase activity of control cells was significantly reduced following a 24 h exposure to actinomycin D or cycloheximide. On the other side, the same inhibitor treatment of melanosome-pretreated cells resulted in less inhibition of tyrosinase activity.The present findings suggest “melanophagic” properties of cultured melanoma cells resulting in enhanced melanogenesis after phagocytotic uptake of functionally active exogenous melanosomes.

scholarly journals ANTIGENIC MODULATION

1968 ◽  
Vol 127 (3) ◽  
pp. 523-539 ◽  
Author(s):  
Lloyd J. Old ◽  
Elisabeth Stockert ◽  
Edward A. Boyse ◽  
Jae Ho Kim
Keyword(s):  
Actinomycin D ◽  
Progressive Increase ◽  
The Other ◽  
Antigenic Modulation ◽  
Cytotoxic Test ◽  
Cell Divisions ◽  
Wide Range ◽  
D Region

Antigenic modulation (the loss of TL antigens from TL+ cells exposed to TL antibody in the absence of lytic complement) has been demonstrated in vitro. An ascites leukemia, phenotype TL.1,2,3, which modulates rapidly and completely when incubated with TL antiserum in vitro, was selected for further study of the phenomenon. Over a wide range of TL antibody concentrations modulation at 37°C was detectable within 10 min and was complete within approximately 1 hr. The cells were initially sensitized to C' by their contact with antibody, thereafter losing this sensitivity to C' lysis together with their sensitivity to TL antibody and C' in the cytotoxic test. The capacity of the cells to undergo modulation was abolished by actinomycin D and by iodoacetamide, and by reducing the temperature of incubation to 0°C. Thus modulation apparently is an active cellular process. Antigens TL. 1,2, and 3 are all modulated by anti-TL.1,3 serum and by anti-TL.3 serum. This modulation affects all three TL components together, even when antibody to one or two of them is lacking. aAnti-TL.2 serum does not induce modulation and in fact impairs modulation by the other TL antibodies. The influence of the TL phenotype of cells upon the demonstrable content of H-2 (D region) isoantigen, first shown in cells modulated in vivo, has been observed with cells modulated in vitro. Cells undergoing modulation show a progressive increase in H-2 (D region) antigen over a period of 4 hr, with no change in H-2 antigens of the K region. Restoration of the TL+ phenotype of modulated cells after removal of antibody is less rapid than TL+ → TL- modulation and may require several cell divisions.

scholarly journals Tyrosinase activity and melanogenic effects of Lespedeza bicolor extract in vitro and in vivo

BioResources ◽  
2020 ◽  
Vol 15 (3) ◽  
pp. 6244-6261
Author(s):  
Si Young Ha ◽  
Ji Young Jung ◽  
Hee Young Kang ◽  
Tae-Heung Kim ◽  
Jae-Kyung Yang
Keyword(s):  
Ethanol Extract ◽  
Melanoma Cells ◽  
B16 Melanoma ◽  
Tyrosinase Activity ◽  
Melanin Content ◽  
B16 Melanoma Cells ◽  
Notable Increase ◽  
Lespedeza Bicolor

Lespedeza bicolor (L. bicolor) is used for medicinal purposes because of its various biological and pharmacological activities. In this study, the effects of L. bicolor ethanol extract on the treatment of vitiligo were investigated. The determination of melanin content in melanocytes was measured using B16 melanoma cells and C57BL/6J Ler-vit/vit mice. Finally, the quercetin content in L. bicolor were qualitatively analyzed using HPLC. The results obviously indicated that the L. bicolor extract enhanced melanogenesis and increased tyrosinase activity in cultured melanoma cells and C57BL/6J Ler-vit/vit mice. Treatment with L. bicolor extract led to a higher content of melanin and eumelanin in C57BL/6J Ler-vit/vit mice hair than in control (untreated) mice, which demonstrated the therapeutic effect of hair-graying associated with vitiligo. There was a notable increase in melanocytes in the skin of C57BL/6J Ler-vit/vit mice treated with L. bicolor extract compared with the control. L. bicolor extract was a potent tyrosinase and melanin synthesis activator in B16 melanoma cells. C57BL/6J Ler-vit/vit mice treated with L. bicolor extract had significantly higher melanin content in hair than the untreated control. The results suggest that L. bicolor extract is a potential alternative treatment for improvement of vitiligo.

scholarly journals Studies of the ethylation of rat liver transfer ribonucleic acid after administration of l-ethionine

1972 ◽  
Vol 128 (1) ◽  
pp. 59-68 ◽  
Author(s):  
A. E. Pegg
Keyword(s):  
Rat Liver ◽  
Ribonucleic Acid ◽  
Major Part ◽  
Actinomycin D ◽  
Major Product ◽  
The Other ◽  
Methyl Donor ◽  
Principal Product

1. The ethylated nucleosides present in tRNA isolated from the livers of rats treated with 0.5g of l-ethionine/kg body wt. were investigated. Evidence that this tRNA contained N2-ethylguanine, N2N2-diethylguanine, N2-ethyl-N2-methylguanine, 7-ethylguanine, two ethylated pyrimidines and ethylated ribose groups was obtained. 2. Ethylation of bacterial tRNA was catalysed by extracts containing tRNA methylases prepared from rat liver by using S-adenosyl-l-ethionine as an ethyl donor, but the rate of ethylation was 20 times less than the rate of methylation with S-adenosyl-l-methionine as a methyl donor. 3. The principal product of such ethylation in vitro was N2-ethylguanine and traces of the other ethylated guanines and pyrimidines found in tRNA isolated from rats treated with ethionine in vivo were also found. 1-Ethyladenine was not formed, although 1-methyl-adenine is a major product of methylation of bacterial tRNA by these extracts, and 1-ethyladenine was not present in the rat liver tRNA isolated from ethionine-treated animals. 4. After injection of actinomycin D (15mg/kg body wt.) or l-methionine (1.0g/kg body wt.) before the ethionine, ethylation of tRNA was diminished by about 80% but not completely abolished. Administration of 1-aminocyclopentanecarboxylic acid (2.5g/kg body wt.) to inhibit the formation of S-adenosyl-l-ethionine inhibited ethylation of tRNA by 44%. 5. These results suggest that not all of the ethylation of tRNA that occurs in the livers of rats treated with ethionine is mediated by the action of tRNA methylases acting with S-adenosyl-l-ethionine as a substrate, but that this pathway does occur and accounts for a major part of the observed ethylation. 6. The results are discussed with reference to ethionine-induced hepatocarcinogenesis.

Tyrosinase activity in primary cell culture of amelanotic melanoma cells

1983 ◽  
Vol 3 (11) ◽  
pp. 1027-1034 ◽  
Author(s):  
A. Słomiński ◽  
P. W. D. Scisłowski ◽  
A. Bomirski
Keyword(s):  
Gel Electrophoresis ◽  
Primary Cell Culture ◽  
Soluble Fraction ◽  
Calf Serum ◽  
Melanoma Cells ◽  
Amelanotic Melanoma ◽  
Tyrosinase Activity ◽  
Logarithmic Phase

After transfer of the Ab amelanotic melanoma cells from in vivo to in vitro growth conditons tyrosinase activity in their soluble fraction rapidly increased. This increase lasted to the middle of the logarithmic phase of growth and was followed by a decrease of tyrosinase activity, which was accompanied by accumulation of melanin in the cells. Calf serum stimulated simul-taneously tyrosinase activity, melanin synthesis, and proliferation of the melanoma ceils. Acrylamide-gel electrophoresis patterns of soluble tyrosinase from the Ab melanoma cells cultured in vitro consisted of two bands, similarly as soluble tyrosinase from the Ma melanotic melanoma cells freshly isolated from solid tumors.

Modification of muscle cell phenotype in monolayer culture by different media

Development ◽  
1972 ◽  
Vol 28 (3) ◽  
pp. 559-570
Author(s):  
Oscar Ramírez ◽  
Victor Alemán
Keyword(s):  
Monolayer Culture ◽  
Phenotypic Expression ◽  
Calcium Content ◽  
Cell Phenotype ◽  
Rich Medium ◽  
Cell Stage ◽  
Monolayer Cultures ◽  
Fibre Development

Conditions for monolayer cultures of chick skeletal muscle in a rich medium showed that non-muscle contaminants were only 20% of the population. Although the calcium content of this rich medium was 0·95 mm, multinucleate and long fibres were observed after 8 days in culture. Either in rich or in restricted media, gelatin organized the pattern of cell and fibre development in the Petri plates. Cells grown outside the gelatin boundaries looked fibroblast-shaped and many were vacuolated. Gelatin did not fully prevent the growth of fibroblast-like population. Calcium in restricted media appeared to be very important for the acquisition of a definite elongated shape. The possibility of the existence of myofibro-myoblasts was supported by the finding of multinucleated fibroblast-like cells during culture in restricted medium. Epigenetic factors, such as different media in a given culture or the origin of a serum batch utilized as a component of those media, affected the fate of the cultures and might also explain the myoblastic variance observed in this and other studies reported. The capacity of phenotypic expression during the modulation change of muscle cells, in vitro, depends not only upon their genotypic origin but also on parameters such as the cell stage during this process, the type of nutrient media used and the interplay of both parameters; in vivo it depends upon specific cytological interactions.

Ancylostoma caninum Anticoagulant Peptide Blocks Metastasis In Vivo and Inhibits Factor Xa Binding to Melanoma Cells In Vitro

1998 ◽  
Vol 79 (05) ◽  
pp. 1041-1047 ◽  
Author(s):  
Kathleen M. Donnelly ◽  
Michael E. Bromberg ◽  
Aaron Milstone ◽  
Jennifer Madison McNiff ◽  
Gordon Terwilliger ◽  
...  
Keyword(s):  
Active Site ◽  
Thrombin Generation ◽  
Factor Xa ◽  
Coagulation Factor ◽  
Melanoma Cells ◽  
Factor V ◽  
Ancylostoma Caninum ◽  
Metastatic Activity

SummaryWe evaluated the in vivo anti-metastatic activity of recombinant Ancylostoma caninum Anticoagulant Peptide (rAcAP), a potent (Ki = 265 pM) and specific active site inhibitor of human coagulation factor Xa originally isolated from bloodfeeding hookworms. Subcutaneous injection of SCID mice with rAcAP (0.01-0.2 mg/mouse) prior to tail vein injection of LOX human melanoma cells resulted in a dose dependent reduction in pulmonary metastases. In order to elucidate potential mechanisms of rAcAP’s anti-metastatic activity, experiments were carried out to identify specific interactions between factor Xa and LOX. Binding of biotinylated factor Xa to LOX monolayers was both specific and saturable (Kd = 15 nM). Competition experiments using antibodies to previously identified factor Xa binding proteins, including factor V/Va, effector cell protease receptor-1, and tissue factor pathway inhibitor failed to implicate any of these molecules as significant binding sites for Factor Xa. Functional prothrombinase activity was also supported by LOX, with a half maximal rate of thrombin generation detected at a factor Xa concentration of 2.4 nM. Additional competition experiments using an excess of either rAcAP or active site blocked factor Xa (EGR-Xa) revealed that most of the total factor Xa binding to LOX is mediated via interaction with the enzyme’s active site, predicting that the vast majority of cell-associated factor Xa does not participate directly in thrombin generation. In addition to establishing two distinct mechanisms of factor Xa binding to melanoma, these data raise the possibility that rAcAP’s antimetastatic effect in vivo might involve novel non-coagulant pathways, perhaps via inhibition of active-site mediated interactions between factor Xa and tumor cells.

Enhancement of ADP-induced Platelet Aggregation by Adrenaline in Vivo and Its Prevention

1973 ◽  
Vol 29 (02) ◽  
pp. 490-498 ◽  
Author(s):  
Hiroh Yamazaki ◽  
Itsuro Kobayashi ◽  
Tadahiro Sano ◽  
Takio Shimamoto
Keyword(s):  
Platelet Count ◽  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
The Other ◽  
Endothelial Surface ◽  
Important Interaction ◽  
Blood Coagulability ◽  
The Difference

SummaryThe authors previously reported a transient decrease in adhesive platelet count and an enhancement of blood coagulability after administration of a small amount of adrenaline (0.1-1 µg per Kg, i. v.) in man and rabbit. In such circumstances, the sensitivity of platelets to aggregation induced by ADP was studied by an optical density method. Five minutes after i. v. injection of 1 µg per Kg of adrenaline in 10 rabbits, intensity of platelet aggregation increased to 115.1 ± 4.9% (mean ± S. E.) by 10∼5 molar, 121.8 ± 7.8% by 3 × 10-6 molar and 129.4 ± 12.8% of the value before the injection by 10”6 molar ADP. The difference was statistically significant (P<0.01-0.05). The above change was not observed in each group of rabbits injected with saline, 1 µg per Kg of 1-noradrenaline or 0.1 and 10 µg per Kg of adrenaline. Also, it was prevented by oral administration of 10 mg per Kg of phenoxybenzamine or propranolol or aspirin or pyridinolcarbamate 3 hours before the challenge. On the other hand, the enhancement of ADP-induced platelet aggregation was not observed in vitro, when 10-5 or 3 × 10-6 molar and 129.4 ± 12.8% of the value before 10∼6 molar ADP was added to citrated platelet rich plasma (CPRP) of rabbit after incubation at 37°C for 30 second with 0.01, 0.1, 1, 10 or 100 µg per ml of adrenaline or noradrenaline. These results suggest an important interaction between endothelial surface and platelets in connection with the enhancement of ADP-induced platelet aggregation by adrenaline in vivo.

CONTRIBUTION TO THE EXISTENCE OF REGULATORY MECHANISMS AT THE ADRENAL LEVEL

1972 ◽  
Vol 70 (4) ◽  
pp. 741-757
Author(s):  
Otto Linèt
Keyword(s):  
Orotic Acid ◽  
Adrenal Glands ◽  
Actinomycin D ◽  
Delay Period ◽  
Regulatory Mechanisms ◽  
Leucine Incorporation ◽  
Total Period ◽  
Free Media

ABSTRACT Rat adrenal glands atrophied by the administration of cortisol acetate in vivo were used as a model for the study of early metabolic processes occurring in vitro. Atrophied adrenals incubated in the presence of 14C-leucine incorporated subnormal quantities of this amino acid per mg of protein for the first 120 min. When the incubation lasted for a total period of 180 or 240 min a supranormal rise in the 14C-leucine incorporation was observed. Similar changes occurred with some delay with regard to corticosterone production as expressed per 100 mg of tissue. No differences in 14C-leucine incorporation were observed between the control and atrophied adrenals in vivo. Homogenates from atrophied glands incorporated 14C-leucine to a greater extent than the control homogenates. The in vitro incorporation of 14C-orotic acid into the RNA was also higher in atrophied adrenals. The in vitro use of actinomycin D, cycloheximide and amphenone indicated that corticosterone production depended on the incorporation of 14C-leucine. The addition of cortisol to the incubation media markedly decreased the enhancement of 14C-lysine incorporation into the protein of atrophied adrenals. These, as well as additional results suggest rebound phenomena: once atrophic adrenals are transferred to cortisol-free media, reparative processes begin after a delay period. Such phenomena seem to be mediated by regulatory mechanisms at the adrenal level.

EFFECT OF ACTINOMYCIN D ON TSH-INDUCED STIMULATION OF THYROIDAL PROTEIN SYNTHESIS IN THE RAT

1974 ◽  
Vol 77 (1) ◽  
pp. 64-70 ◽  
Author(s):  
Gustav Wägar
Keyword(s):  
Protein Synthesis ◽  
Actinomycin D ◽  
The Other ◽  
Leucine Incorporation ◽  
Short Term ◽  
Other Hand ◽  
Thyroid Glands ◽  
Translational Level ◽  
Stimulation Of

ABSTRACT Whether the short-term regulation of thyroidal protein synthesis by TSH occurs at the transcriptional or the translational level was tested by measuring the effect of actinomycin D (act D) on the TSH-induced stimulation of L-14C-leucine incorporation into the thyroidal proteins of rats. TSH was injected 6 h before the rats were killed. The thyroid glands were then removed and incubated in vitro in the presence of L-14C-leucine for 2 h. The pronounced stimulation of leucine incorporation in the TSH-treated animals was depressed as compared with controls but still significant even when the animals had been pre-treated with 100 μg act D 24 and 7 h before sacrifice. On the other hand, act D strongly decreased incorporation of 3H-uridine into RNA. Short-term regulation of thyroidal protein synthesis by TSH appears to be partly but not wholly dependent on neosynthesis of RNA. Hence regulation may partly occur at the translation level of protein synthesis.

Oxytocin analogues with non-coded amino acid residues in position 8: [8-Neopentylglycine]oxytocin and [8-cycloleucine]oxytocin

1987 ◽  
Vol 52 (9) ◽  
pp. 2317-2325 ◽  
Author(s):  
Jan Hlaváček ◽  
Jan Pospíšek ◽  
Jiřina Slaninová ◽  
Walter Y. Chan ◽  
Victor J. Hruby
Keyword(s):  
Amino Acid ◽  
Solid Phase Synthesis ◽  
Solid Phase ◽  
The Other ◽  
Amino Acid Residues ◽  
Phase Synthesis ◽  
Other Hand ◽  
Fragment Condensation

[8-Neopentylglycine]oxytocin (II) and [8-cycloleucine]oxytocin (III) were prepared by a combination of solid-phase synthesis and fragment condensation. Both analogues exhibited decreased uterotonic potency in vitro, each being about 15-30% that of oxytocin. Analogue II also displayed similarly decreased uterotonic potency in vivo and galactogogic potency. On the other hand, analogue III exhibited almost the same potency as oxytocin in the uterotonic assay in vivo and in the galactogogic assay.

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