Showing papers by "Peter Christiansen published in 1972"

Urinary follicle stimulating hormone and luteinizing hormone in normal adult men

Abstract: The normal range of the excretion of follicle stimulating hormone (FSH) and luteinizing hormone (LH), as well as the relation between these values and those in aging subjects was investigated by specific bioassays in 50 normal adult men with an age range of 21 to 82 years. The FSH rose significantly but did not reach the levels observed in postmenopausal women. The LH showed no significant rise and consequently the FSH/LH ratio rose. Normal values and the 95 % fiducial limits in the decades from 21 to 70 years are shown. In contrast to the many reports about FSH and LH excretion in women, little has been published about FSH and LH in men and especially in normal men We therefore studied this problem and it is the purpose of this report to show the excretion of FSH and LH in normal men as measured by specific bio¬ assays. The development of radioimmunoassays in recent years has made it possible to investigate the plasma levels of FSH and LH from hour to hour and day to day. Peterson et al. (1968) found in 4 normal men no systematic day to day variation, as occurs in normal women and found in 2 normal men examined every hour for 24 hours no special diurnal rhythm. However, Saxena et al. (1968) found in 3 normal men, morning values of FSH higher than evening values and in 2 of the men a similar variation of LH levels. In our study any influence of such variations was eliminated as combined extracts from urine collected over 12-14 days were assayed. Downloaded from Bioscientifica.com at 11/28/2018 03:47:28PM via free access MATERIAL AND METHODS Subjects All the subjects were healthy adult males with an age range of 21-82 years. They had testes of normal size, were normally virile without any sign of endocrine disorder and those who were married and wanted children had proved their fertility by having at least 1 child. Altogether 50 subjects were investigated. All the collected 12-14 24 hour urine samples were extracted by the method of Johnsen (1958), and then pooled and divided into 3 portions for the 3 bioassays. Bioassays 1. Total urinary hypophyseal gonadotrophins (HG) were measured by the mouse uterus test (Johnsen 1958) and expressed in mouse uterus units (MUU) per 24 hours. One ampoule of the 2. international reference preparation for human menopausal gonadotrophin (2. IRP-HMG) contains 40 IU FSH, 40 IU LH and 133 MUU. 2. The urinary follicle stimulating hormone (FSH) was measured by the rat ovarian augmentation test (Steelman 8c Pohley 1953) and performed in the following way: 26-28 days old female rats of the Serum Institute strain (originating from the Wistar strain), weighing 45—55 g were injected twice daily for 3 days subcutaneously, each time with a volume of 0.5 ml. Autopsy was performed on the 4th day. Both ovaries were dissected free of surrounding tissue and weighed on a torsion balance to the nearest 0.5 mg. The test material together with human chorionic gonadotrophin (Physex® LEO) was mixed in vitro in such a way that each rat got 20 IU HCG as total dose for augmentation. The extracts were assayed against the 2. IRP-HMG with a 3 + 3 design expressing the activity in IU per 24 hours. Five rats per dose. 3. The urinary luteinizing hormone (LH) was measured by the ventral prostate weight method (VPW) (Greep 1941) and performed as previously described (Christian¬ sen 1967). The extracts were assayed against the 2. IRP-HMG in a 2 + 2 design ex¬ pressing the activity in IU per 24 hours. 5-8 rats were used per dose. The bioassays were calculated according to a computer programme for bioassays (McArthur et al. 1966). Only statistically valid assays were accepted. Significant responses were obtained with all urine samples.

Studies on the rat ovarian augmentation method for follicle stimulating hormone.

Abstract: The rat ovarian augmentation method for the quantitative determination of follicle stimulating hormone (FSH) has been systematically studied. Assays with different strains and in rats of different ages have been performed, as well as assays with hypophysectomized rats. The effects of variations of the injection schedule and in the amounts of HCG used for augmentation have been examined. Accordingly some modifications of the original method of Steelman & Pohley (1953) have been made. The best results were obtained with 26\\p=n-\\28days old rats of the SS strain (breed of Statens Seruminstitut, originating from the Wistar strain), weighing 45\\p=n-\\55 g, and injected twice daily sc for 3 days, with autopsy on the 4th day. The most suitable dose of HCG for augmentation was 20 IU as the total dose per rat. A 3 + 3 design with 3 rats per dose gave a satisfactory precision, the index of precision being between 0.10 and 0.15. The sensitivity was about 2 IU of the 2. IRP-HMG. The course of the curves for human and ovine FSH are found to be identical. The reproducibility of the method during 1 year has been studied and is satisfactory. Since the follicle stimulating hormone (FSH) was discovered many attempts have been made to quantify this hormone. Evans et al. (1939) used a histological assay based on the production of normal follicles in hypophysectomized female rats, Greep et al. (1942) used Downloaded from Bioscientifica.com at 10/03/2018 06:07:59PM via free access the increase in testicular weight in hypophysectomized male rats and Fevold et al. (1940) used the increase in ovarian weight in immature, intact female rats. Other investigators used the increase in uterine weight in intact, immature mice or rats. When it became known that human chorionic gonadotrophin (HCG) augments the effect of FSH on the ovaries, Simpson et al. (1951), in hypophysectomized HCG-treated female rats showed that a dose-response curve could be obtained, and Steelman 8c Pohley (1953) found that immature intact HCG-treated rats could be used instead of hypophysectomized rats and that their ovaries were more sensitive to FSH. Later Brown (1955), using the same principle, employed mice instead of rats and got a more sensitive assay. How¬ ever, the method seems to be highly dependent on the strain of mice used. Igarashi 8c McCann (1964) published a quantal assay for FSH using the principle that small amounts of HCG sentisize the mouse uterus to FSH. The method was sensitive but highly unspecific (Uberoi 8c Meyer 1967). When some two years ago we needed a bioassay to measure the excretion of FSH in urine from human subjects, preliminary experiments indicated that the Steelman-Pohley method was the most promising for the animals we had at our disposal. In work on bioassays it is well known that even small alterations in the practical performance of the assay can give remarkable improvements. We therefore systematically studied the method, and the purpose of this paper is to present the results obtained. The specificity of the method will be dis¬ cussed in a following paper (Christiansen 1972). MATERIALS AND METHODS Animals Infantile female rats of the Holtzman strain and of the SS strain (a breed of Statens Seruminstitut, originating from the Wistar strain); intact, as well as hypo¬ physectomized rats were used. Altogether 760 rats were used for the assays. Preparations A) Human chorionic gonadotrophin. A commercial preparation, Physex®, con¬ taining 1500 IU per ampoule, prepared by LEO Pharmaceutical Products, Copenhagen. B) Preparations containing follicle stimulating hormone. 1) Ovine: NIH-FSH-S-1, NIH-FSH-S-2 and NIH-FSH-S-3, kindly supplied by the National Institutes of Health, Bethesda, USA. 2) Human: a) The second international reference preparation for human menopausal gonadotrophin (2. IRP-HMG) established 1964, containing 40 IU FSH and 40 IU luteinizing hormone (LH) per ampoule, b) HMG LEO 63051 and 64021, two commercial preparations made by LEO Pharmaceutical Products, Copen¬ hagen. The method used was the rat ovarian augmentation test (Steelman Se Pohley 1953). In all assays the test material, together with the HCG was dissolved in borate buffer (pH = 9.2) and injected subcutaneously. Between the injections the solutions were stored at + 4°C. Downloaded from Bioscientifica.com at 10/03/2018 06:07:59PM via free access Fig. 1. 26-28 days old female rats of the SS strain. X-X 26-28 days old female rats of the Holtzman strain. Mean values and total doses per rat are referred to. 5 rats per dose, 10 rats as controls. Augmented with a total of 20 IU HCG se per rat, 5 injections sc during 3 days, the volume per injection being 0.5 ml. Autopsy on the 4th day. The ordinate indicates the weight of both ovaries. The vertical bars indicate the standard deviations. The mean values in the different curves have been displaced a little in order to show the standard deviations separately. The rats were killed with ether, the ovaries immediately removed and dissected free of surrounding fat tissue and weighed on a torsion balance to the nearest 0.5 mg. Statistical calculations were done according to McArthur et al. (1966).