Summary
Several types of cultured cells release glycolytic enzymes into their suspending medium. This effect is most obvious with tumor cells, especially with their ascites forms. Erythrocytes do not release glycolytic enzymes. The total extracellular phosphoglucose isomerase activity consists of two components. One part is dissolved in the medium, the other one is sedimentable at 150× g together with the cells. The latter seems to be localized at the cell surface.
At densities of about 106 cells/ml maximum activity in the medium is reached within 5–10 min. After that no further release of enzyme activity can be observed.
Serum reduces the rate of enzyme release considerably. This effect can be reversed by washing with protein free media. Treatment with trypsin leads to high extracellular phosphoglucose isomerase activities of the cells which originally show low external enzyme activity. Erythrocytes do not show any effect with trypsin, ascites tumor cells do not alter their high extracellular enzyme activity.
At a density of 105 cells/ml, Yoshida acites tumor cells, cultured in vitro, release about 12% of originally intracellular phosphoglucose isomerase activity by 5 elutions with fresh medium. The process of enzyme release shows a certain selectivity in respect to different glycolytic enzymes. Aldolase exhibits the highest activity in the medium in relation to its homogenate activity.
Zusammenfassung
Zellen verschiedenen Ursprungs setzen in vitro glykolytische Enzyme in das umgebende Medium frei. Dieser Prozeß ist besonders deutlich bei Tumorzellen, am ausgeprägtesten bei den Aszitesformen. Erythrozyten zeigen diesen Effekt nicht.
Die insgesamt nachweisbare extrazelluläre Phosphoglucose-Isomerase-Aktivität besteht aus zwei Anteilen: einem, der im Medium gelöst ist und einem zweiten, der bei 150×g mit den Zellen sedimentierbar ist. Der letztere scheint an der Zelloberfläche lokalisiert zu sein.
Bei Zelldichten von 106 Zellen/ml wird das Maximum der Enzymfreisetzung innerhalb von 5–10 min erreicht. Danach bleibt die Enzymaktivität konstant. In Gegenwart von Serum ist die Geschwindigkeit der Enzymfreisetzung erheblich vermindert. Dieser Prozeß ist reversibel nach Waschungen mit Protein-freien Medien.
Behandlung mit Trypsin führt zu hohen extrazellulären Phosphoglucose-Isomerase-Aktivitäten bei den Zellen, die ursprünglich geringe Enzymaktivitäten aufweisen. Erythrozyten zeigen keinen Effekt. Die hohe extrazelluläre Phosphoglucose-Isomerase-Aktivität der Aszites-Tumorzellen ändert sich durch Trypsin nicht.
Bei einer Zelldichte von 105 Zellen/ml können ca. 12% der intrazellulären Phosphoglucose-Isomerase-Aktivität durch 5 Elutionen mit jeweils frischen Medien freigesetzt werden.
Die Freisetzung verschiedener glykolytischer Enzyme unterliegt einer deutlich nachweisbaren Selektivität. Im Vergleich zu anderen glykolytischen Enzymen weist die Aldolase die höchste Aktivität im Medium im Verhältnis zu ihrer Homogenataktivität auf.
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Abbreviations
- FCS:
-
Fetal calf serum
- Fru-6-P:
-
Fructose 6-phosphate
- Glc-6-P:
-
Glucose 6-phosphate
- Hanks BSS:
-
Hank's balanced salt solution
- HEPES:
-
2-(4-(2-Hydroxyethyl)-piperazinyl-(1))-ethansulfonic acid
- MEM:
-
Minimum essential medium (Eagle)
- Tricine:
-
N-(Tris-(hydroxymethyl)-methyl)-glycine
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Keller, K., Kolbe, H., Lange, K. et al. Release of glycolytic enzymes from cultivated tumor cells. Z. Krebsforsch. 92, 275–286 (1978). https://doi.org/10.1007/BF00461650
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DOI: https://doi.org/10.1007/BF00461650