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Proteoglycans synthesized by human polymorphonuclear leucocytes in vitro
P M Bartold1, D G Harkin, L P Bignold
1Department of Pathology, University of Adelaide, Australia.
Abstract:
Polymorphonuclear leucocytes (PMN) were assessed in vitro for their ability to synthesize and secrete proteoglycans. The PMN were isolated from human peripheral blood and were found to contain less than 5% mononuclear cells. Following 24 h incubation in the presence of (35S)-sulfate, significant quantities of 35S-labelled macromolecules were detected both within the culture medium and cells. Although the PMN preparations contained some platelets (approximately five platelets:one PMN), culture of platelets alone did not result in the detection of any 35S-labelled macromolecules in either the medium or platelets. 35S/3H-labelled macromolecules from the PMN cultures were identified as proteoglycans on the basis of their degradation by papain, alkaline sodium borohydride, chondroitinase ACII, chondroitinase ABC and nitrous acid. The labelled proteoglycans isolated from the medium and cells eluted from Sepharose CL-4B with a Kav of 0.63; this indicated a small size compared with many other proteoglycans. The glycosaminoglycans associated with the proteoglycans were identified as heparan sulfate, chondroitin sulfate and dermatan sulfate, with chondroitin sulfate being the principal component. The average molecular weight of the glycosaminoglycans was determined to be 16,000. Therefore, the data from this study demonstrate the ability of human PMN to synthesize and secrete proteoglycans in vitro which appear to differ from those synthesized by mesenchymal cells with respect to molecular size and glycosaminoglycan composition.
Insights
Human polymorphonuclear leucocytes (PMN) can synthesize and secrete proteoglycans in vitro. These novel proteoglycans are smaller and have a different glycosaminoglycan composition than those from mesenchymal cells.
Area of Science:
- Biochemistry
- Cell Biology
- Immunology
Background:
- Polymorphonuclear leucocytes (PMN) are key immune cells.
- The extracellular matrix composition and synthesis by immune cells are not fully understood.
Purpose of the Study:
- To investigate the in vitro synthesis and secretion of proteoglycans by human PMN.
- To characterize the biochemical properties of PMN-derived proteoglycans.
Main Methods:
- Human peripheral blood PMN were isolated and cultured for 24 hours with (35S)-sulfate.
- Radiolabeled macromolecules were analyzed using enzymatic degradation (papain, chondroitinase, nitrous acid) and size exclusion chromatography (Sepharose CL-4B).
- Glycosaminoglycan composition and molecular weight were determined.
Main Results:
- Significant amounts of 35S-labeled proteoglycans were detected in both PMN cell cultures and the culture medium.
- Platelet-only cultures did not yield detectable labeled macromolecules, confirming PMN as the source.
- PMN proteoglycans were characterized as small in size (Kav 0.63) and composed primarily of chondroitin sulfate, with heparan and dermatan sulfate also present.
- The average molecular weight of the glycosaminoglycans was 16,000 Da.
Conclusions:
- Human PMN possess the capability to synthesize and secrete proteoglycans in vitro.
- PMN-derived proteoglycans exhibit distinct characteristics, including smaller molecular size and a unique glycosaminoglycan profile, compared to those synthesized by mesenchymal cells.
- This finding contributes to understanding the role of PMN in tissue remodeling and extracellular matrix dynamics.