Studies on the rat ovarian augmentation method for follicle stimulating hormone.
Studies on the rat ovarian augmentation method for follicle stimulating hormone.
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促卵泡激素大鼠卵巢增大方法的研究。
DOI：
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发表时间：
1972
期刊：
影响因子：
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通讯作者：
P. Christiansen
中科院分区：
文献类型：
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作者：
P. Christiansen
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).
