Assay of the Estrogenic Hormone in Hemophilic and Normal Male Urine

Abstract

Because hemophilia occurs only in males and is transmitted directly only by females, an assay of the urinary estrogenic hormone was undertaken.

The technic used was identical with that of Dr. Petersen of this institution in order that we might compare the results of our normal control group with his.

White female rats 2 months of age were used. Vaginal smears were made every 12 hours for 3 weeks to ascertain that estrous cycles were normal. Then a laparotomy was performed on each rat under ether anesthesia, the ovaries, broad ligaments and upper part of the cornua of the uterus being removed. The rats were then allowed 10 days to recover from the operation, following which vaginal smears were made at 12-hour intervals for 3 weeks to determine whether the estrous cycles had ceased.

One liter of urine from untreated hemophilic patients was used in each series of experiments. The urine, rendered acid to congo red, was extracted 3 times with benzene in a reflux condenser and evaporated to a syrup. Corn oil was then added and the remainder of the benzene evaporated. This leaves the hormone in the corn oil.

The rats were then divided into groups of 4. Three injections of urine extract of 1/6 cc each were made at 4-hour intervals into each of the 4 rats. Vaginal smears were made at 6-hour intervals for at least 48 hours or until activity ceased. If estrus was induced in 3 out of 4 rats, the test was considered positive.

The number of estrous cycles and the total hours of estrus were determined for the normal and hemophilic groups.

The urine of the first 5 hemophiliacs induced no estrual cycles. These results were recorded in a preliminary report in the J. Am. Med. Assn., Vol. 97, page 244.