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Isolation, Identification, and Purification of Murine Thymic Epithelial Cells
Published on: August 8, 2014
Initial characterization of human thymocyte sialidase activity: evidence that this enzymatic system is not altered
A Greffard1, J C Pairon, H Terzidis-Trabelsi
1Inserm U 139, Hôpital Henri Mondor, Creteil, France.
Insights
Human thymocytes possess an acid-active, membrane-bound sialidase. A unique acidic soluble sialidase in T-cells may not be involved in thymic maturation, despite its presence in circulating T-cells.
Area of Science:
- Biochemistry
- Immunology
- Cell Biology
Background:
- Sialidases are enzymes that cleave sialic acids, crucial for cell surface interactions.
- Mammalian soluble sialidases are typically active at neutral pH, while membrane-bound forms often exhibit acidic activity.
- The role of sialidase activity in T-cell development and function is not fully understood.
Purpose of the Study:
- To characterize the sialidase activity in human thymocytes.
- To investigate the properties and cellular localization of thymocyte sialidase.
- To determine the potential role of sialidase in thymic maturation.
Main Methods:
- Fluorogenic assay to measure sialidase activity.
- Cell fractionation (lysosomal, microsomal, cytosolic) to determine enzyme localization.
- Analysis of thymocyte subpopulations and mature T-cells.
Main Results:
- Human thymocyte sialidase activity is primarily acid-active and membrane-bound, with significant recovery in lysosomal and microsomal fractions.
- A unique, weakly active soluble sialidase with an acidic pH optimum was detected in thymocytes and mature T-cells.
- This acidic soluble sialidase activity remained consistent across thymic maturation stages.
- The enzymatic system did not appear to be involved in the observed fluctuations of cell surface sialic acid during thymic maturation.
Conclusions:
- Human thymocytes exhibit a predominantly membrane-bound, acid-active sialidase.
- A distinct acidic soluble sialidase is characteristic of the human T-cell lineage.
- This acidic soluble sialidase is unlikely to be involved in regulating cell surface sialic acid levels during thymic maturation.
Abstract:
1. The sialidase activity of human thymocyte was examined by a fluorogenic assay. 2. These studies revealed that human thymocyte sialidase activity is essentially acid-active and membrane-bound since 59.6% and 33% of the total activity was recovered in the lysosome-enriched and microsomal fractions, respectively. 3. A weak activity was also detected in the cytosolic fraction. 4. However, the acidic optimum pH of this soluble sialidase was at variance with the general concept of mammalian soluble sialidases which are known to be optimally active at more neutral pH. 5. This acidic soluble sialidase seems to be a general characteristic of the human T-cell lineage since examination of mature circulating T-cells revealed that they contain a soluble sialidase activity similar to that observed in thymocytes. 6. Analysis of mature and immature thymocyte subpopulation obtained by differential PNA agglutination indicated that this enzymatic system was not altered during the course of thymic maturation. 7. These results suggest that unlike in T-cell activation where changes in the level of sialidase activity were shown to influence the extent of cell surface sialylation and thereby the cell physiology, this enzymatic system seems not to be involved in the fluctuation of cell surface sialic acid content observed during thymic maturation.
