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Biochemical and biophysical research communications

Alkaline phosphatase retained in HepG2 hepatocarcinoma cells vs. alkaline phosphatase released to culture medium: difference of aberrant glycosylation.


PMID 15796897

Abstract

Liver tissue is the source of 90% of serum alkaline phosphatase (AP). The serum levels and structures of tumor marker proteins change under many disease conditions as well as cancer. The study was aimed at determining the type of alkaline phosphatase (AP) present in HepG2 hepatocellular carcinoma cell line. Alkaline phosphatase rich extracts of healthy human liver, HepG2 hepatocarcinoma cells, as well as the condition medium of HepG2 cells were prepared by extraction with 40% n-butanol and 30-50% acetone precipitation, and subjected to various chromatographic procedures. Lectin affinity chromatography of the samples with concanavalin A-Sepharose 4B showed considerable differences in the elution patterns. Non-denaturing polyacrylamide gel electrophoresis of the culture medium yielded a relatively slow migrating band of activity that coincided with none of the three bands of activity produced by the normal liver extract, nor with the bands of the cell pellet extract. Inhibition patterns were established by measuring the enzyme activities in the presence of varying concentrations of L-phenylalanine, L-leucine, L-homoarginine, and levamisole. The APs from the cell line were neuraminidase sensitive. According to the results the main AP produced and released to the medium by HepG2 cell line is an aberrantly glycosylated tissue non-specific AP. In addition, the differences between the cell-pellet AP and the culture medium AP seemed to stem from different sugar moieties in their structures.

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