Demonstration of intracellular and secreted forms of large human chorionic gonadotrophin alpha subunit in cultures of normal placental tissue.

Free human chorionic gonadotrophin (hCG) alpha-subunit in tissue extracts and incubation media of normal placentae before and after four days of culture was investigated by Sephadex G-100 chromatography. Prior to incubation, tissue extracts of first-trimester and term placentae contained a large form and a small form of alpha-subunit relative to a reference urinary hCG alpha-subunit preparation. The large alpha-subunit comprised 30.6 +/- 1.4 per cent s.e.m. and the small alpha-subunit, 66.9 +/- 4.1 per cent of total intracellular alpha-subunit. After 96 h of incubation, the large alpha-subunit comprised 77.6 +/- 9.8 per cent of the total intracellular alpha-subunit, and the small 21.0 +/- 8.1 per cent. The incubation medium contained a single form of alpha-subunit which eluted with a larger apparent molecular weight than the urinary hCG alpha-subunit. Following neuraminidase treatment, the large alpha-subunit from tissue extracts eluted as a smaller molecule in the position of the urinary hCG alpha reference. A change in Ve/Vo of the small alpha-subunit was not detected. Identical neuraminidase treatment of the large alpha-subunit from incubation medium resulted in an increase in the Ve/Vo, but the large alpha-subunit continued to elute from Sephadex G-100 before the urinary hCG alpha-subunit. However, large alpha-subunit from culture medium did elute in the position of urinary hCG alpha following mild acid hydrolysis. These studies demonstrate that a large form of alpha-subunit is present in first-trimester and term placental tissue both before and after incubation in vitro and that the large form of alpha-subunit is secreted by the normal placenta. The differential effects of neuraminidase treatment on the intra- and extracellular forms of large alpha-subunit coupled with our findings from acid hydrolysis of the extracellular form suggest that alpha-subunit secreted into the culture medium may contain additional carbohydrate residues not present on the intracellular form.