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Amenorrhea: Diagnosis and Therapy
Amenorrhea: Diagnosis and Therapy HERBERT S. KUPPERMAN, M.D., PH.D. *
TREATMENT of the failure of menstruation, whether it is primary or secondary in nature, is predicated upon adequate diagnosis and understanding of the underlying mechanism triggering the aberration. One must be cognizant of a possible functional disturbance in the endocrine system as a major etiolgic factor. The intimate relationships existing between the pituitary, the master gland, and the subordinate glands of internal secretion necessitate proper identification of the organ or organ systems involved in the cessation of cyclic catamenia. The pituitary gland, while the dictator of the endocrine system, is to a certain extent governed in its activity by the glands it controls, much in the same manner that all dictators to a degree are controlled by the people to whom they dictate. Hence, in considering the problem of amenorrhea both with respect to etiology and therapy, one must understand the basic principles of the physiology of menstruation and realize that, while the pituitary gland may be an important entity in the etiology of amenorrhea, it can be adversely affected by the thyroid, adrenals and/or ovaries. In addition, the end organ itself, the endometrium or uterus, must also be evaluated before the dysfunction responsible for disturbing the mosaic pattern of menstrual function can be correctly identified. The primary menstrual axis-the pituitary, ovaries and uterus-must be virtually intact to permit normal menstrual function. The thyroid and the adrenal in their role as secondary factors may affect the primary axis by virtue of their effect upon pituitary function itself. The importance of the primary axis necessitates the relegation of thyroid and adrenal factors to the latter part of our discussion. From the Departments of Therapeutics and Obstetrics and Gynecology, New York University-BeUevue Medical Center and Obstetrical and Gynecological Service (Third Division), BeUevue Hospital, and Endocrine Clinic, Beth Israel Hospital, New York, N.Y. *Associate Professor of Medicine and Endocrinologist, New York University PostGraduate Medical College; Endocrinologist, Beth Israel Hospital, New York City, Methodist Hospital, Brooklyn, and the Margaret Sanger Research Bureau, New York City. 517
Herbert S. Kupperman
518 r-~
i PATIENT WITH LAMENORRHEA -~
I PROGES;ERONE
,,//C, 1! 8~~~0s ;::::fi:~-' PITUITARY
iJ
CYCLIC OR COMBINED ESTROGEN a PROGESTERONE
I!
I
\
.-------------~
GONADOTROPHIN ASSAY (FSH) OR PREGNANT MARE'S SERUM
I
HIGH FSH NO RESPONSE TO PMS
I
LON OR NORMAL FSH PMS RESPONSE I INCREASED BREAST SIZE. 2_ MATURATION OF VAGINAL SMEAR 3 MENSES AFTER PROGES-
INDICATES PITUITARY / INSUFFICIENCY'
,o,·a I) (\
TERONE, 100 MGM LM
Fig. 206. Chart demonstrating diagnostic work-up in patient with amenorrhea. AMENORRHEA DUE TO DISTURBANCES IN THE PRIMARY MENSTRUAL AXIS
The organ systems involved in the primary menstrual axis now will be considered in greater detaiL Three organs are necessary for adequate menstrual function to take place normally: a uterus or functioning endometrium, without which menstruation obviously cannot take place; ovaries capable of stimulating the endometrium; and a pituitary gland replete with properly proportioned gonadotropins for maintaining the ovary in a functioning state of rhythmic activity.1-3 The patient who presents herself with a primary complaint of amenorrhea must be correctly diagnosed in order to ascertain the presence or absence of gross disease, either functional or organic, in any of the three organ systems listed above. The initial step presumes that there may be a primary defect in anyone of these three organ systems in the primary menstrual axis. To that end, the patient is subjected to the initial diagnostic procedure in order to properly identify this defect. As the initial step in this work-up, the patient receives 100 mg. of progesterone in 2 cc. of oil administered intramuscularly, following which the time of onset of menstruation to take place is established (Fig. 206). If menstruation occurs, then we have irrefutable evidence that the patient has an endometrium capable of menstruating; secondly, that the endometrium has been adequately stimulated by sufficient endogenous estrogenic activity so that the exogenously administered progesterone can act to induce withdrawal bleeding. Progesterone cannot induce withdrawal bleeding, unless previous and adequate priming with estrogen has been accomplished. Thirdly, the patient must have a pituitary gland
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Fig. 207. Eighteen year old obese female with primary amenorrhea and mental retardation misdiagnosed as Frohlich's syndrome. Error in diagnosis established by occurrence of menstrual period after progesterone medication.
capable of secreting sufficient amounts of follicle-stimulating hormone to promote endogenous estrogenic production by the ovary. Thus, by the simple procedure of a single injection of progesterone and the presence of menstruation thereafter, one has eliminated any possible serious defect in the pituitary, ovary or uterine axis (Fig. 206). Menstruation should occur "'ithin two weeks after the progesterone injection and usually takes place within five to seven days. The patients who respond to this type of regimen ,yho have not menstruated before or who have previously exhibited secondary amenorrhea fall into a number of different categories. A specific example is a young female referred to our clinic with a diagnosis of Frohlich's syndrome (Fig. 207). The patient was short and obese, 18 years old, mentally retarded with inadequate development of her secondary sex characteristics. She was given 100 mg. of progesterone, following which she promptly menstruated. The resultant menstruation ruled out unequivocally the presence of Frohlich's syndrome inasmuch as individuals with this syndrome must by definition have pituitary insufficiency. The presence of
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Fig. 208. Two patients with pseudocyesis of some 8 months' duration. Diagnosis established by clinical findings and menstrual period taking place after progesterone administration.
menstruation after progesterone indicates there could not be significant pluriglandular insufficiency based upon the reasoning listed above. Other patients with amenorrhea in whom the etiological problem may be readily resolved are those in whom cessation of menstruation may be of psychosomatic origin. Thus, the young lady who desires pregnancy so strongly that she eventually believes she is pregnant and develops the classical picture of pseudocyesis, may menstruate very promptly after progesterone administration (Fig. 208). Menstruation so induced eliminates the need for any further extensive laboratory work-up. On the other hand, the woman who has been a bit promiscuous, fears pregnancy and as a result stops menstruating will also experience a menstrual flow after progesterone administration. Many other patients develop amenorrhea secondary to persistent follicular or proliferative activity associated with anovulatory cycles. In these females mental anxiety may be of sufficient extent to inhibit proper sequential secretion of the pituitary gonadotropic hormones. They may be made to menstruate very promptly after progesterone therapy. The primary underlying disturbance disrupting sequential pituitary function in such patients and some of those listed above many be in the hypothalamus. The hypothalamus is an im-
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portant factor in normal hypophyseal activity and is involved in the integration of one's emotions and pituitary function. Progesteroneinduced catamenia may be instrumental in re-establishing a more normal hypothalamic-hypophyseal relationship. Thus, menstruation occurring after the administration of progesterone is not only of diagnostic importance, but also may be of therapeutic advantage. Specific therapy in these patients will be considered below. If the patient does not menstruate after progesterone alone, we have no evidence of the functional integrity of the three organs mentioned above and the next step in our regimen is required. This demands the administration of cyclic therapy with estrogen and progesterone (Fig. 206, Step 2). Our usual procedure in this clinic involves the initial administration of an oral estrogen for three weeks. The following preparations have been employed: 0.05 mg. tablets of ethinyl estradiol or 1.25 mg. tablets of conjugated estrogens (equine) or comparable doses of other estrogens, all being administered twice a day for a period of three weeks. Single injections of parenteral estrogens may also be employed. Two such preparations, estradiol valerate and estradiol cyclopentylpropionate, may be administered as oil solutions-in single doses of 20 and 10 mg. respectively. The patient again receives 100 mg. of progesterone in oil intramuscularly following the three week course of oral estrogen therapy or two weeks after the single intramuscular injection of the longacting estrogen preparation. Here one of two responses occur: the patient either menstruates or she fails to menstruate. The failure of menstruation to occur indicts the uterus in the causation of the amenorrhea. The specific uterine factors that may be indicted in amenorrhea are the following: (1) Uterine agenesis. One such case has been seen in this clinic in which the cervix and fundus were represented merely by a fold of peritoneum. (2) After severe endometritis, particularly after an acidfast infection. This is a common cause of amenorrhea in the Near East. (3) Following destruction of the endometrium by radiation. (4) Following a dilatation and curettage for an incomplete abortion or following postpartum hemorrhage. In the latter cases, where the endometrium is soft and succulent, the sharp curette even in the most competent hands may remove enough of the endometrium so that further regeneration of endometrial tissue cannot occur. This may result in permanent amenorrhea. Prognosis for restoration of menstrual function in patients with uterine failure is poor in all categories with occasional exception in the last group. In those females in whom partial obliteration of the uterine cavity has been induced by the surgical removal of the endometrial lining, one may sometimes stimulate the remnant of endometrial tissue to proliferate sufficiently by administration of high doses of estrogenic substancessome three to four times those ordinarily used in priming. These high doses have been helpful in salvaging some patients with mechanical destruction of the endometrium.
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At times amenorrhea resistant to cyclic therapy with estrogen and progesterone may occur due to cervical stenosis following instrumentation or a surgical procedure, such as a dilatation and curettage. However, in these patients there is the tell-tale evidence of the history of amenorrhea associated with cyclic pain at monthly intervals, and usually accompanied by some uterine enlargement. A sanguineous discharge following fenestration of the stenotic os confirms the diagnosis. This procedure then is followed invariably by cyclic resumption of the catamenia. If the patient does menstruate after the combined cyclic therapy with estrogen and progesterone, one may assume that the integrity of the endometrium and uterus is intact and that further steps must be undertaken to establish the role of the pituitary or the ovary in the etiology of the amenorrhea. The patient is then subjected to the next diagnostic procedure (Fig. 206, Step 3). This involves the intramuscular administration of pregnant mares' serum gonadotropin and/or the performance of an urinary gonadotropin assay. The pregnant mares' serum is administered in doses of 500 I.D. three times a week for three weeks. This may result in ovarian stimulation. Stimulation of ovarian function would be characterized by: (1) increase of breast size; (2) increased vaginal secretion; (3) vaginal maturation as evidenced by vaginal smears taken before and after administration of the gonadotropin; (4) menstrual period occurring after the patient receives 100 mg. of progesterone in oil some three to five days after the last pregnant mares' serum injection. The presence of these signs of ovarian stimulation would indicate that the organ responsible for the amenorrhea is not the gonads, but in all probability is the pituitary gland. The failure of response to pregnant mares' serum would indict the ovaries for the absence of the catamenia and may indicate ovarian agenesis. The performance of a urinary gonadotropic assay would yield essentially confirmatory data to the pregnant mares' serum gonadotropin; i.e., a high follicle-stimulating hormone excretion would be noted in those patients who failed to respond to pregnant mares' serum. On the other hand, a low or normal gonadotropic excretion would be obtained in those females who show a response to pregnant mares' serum. Treatlllent
The treatment of a patient with amenorrhea based upon disturbances in the pituita-ry-ovarian-uterine axis may involve several modalities of therapy. In the individual who fails to menstruate spontaneously, but shows a catamenia following the administration of progesterone, the advisable form of therapy is to continue the progesterone medication at monthly intervals for the next three or four months. This form of therapy may aid in reinstituting periods which had been held in abeyance in the past. The mechanism of action whereby cyclic therapy with progesterone may initiate menses in these patients must rest on the obtuse explanation of the interrelation between the pituitary gland and the hormone prod-
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ucts of its end organs. Progesterone may induce certain regulatory effects upon pituitary secretion. In addition to the use of progesterone, we have found that the employment of one of the ataractic drugs may have a beneficial effect upon the induction of cyclic menstruation in these individuals. These ataractic drugs have the potentiality of inducing a more normal hypothalamicpituitary relationship by virtue of their effect upon the hypothalamus. It has been adequately demonstrated in animals, at least, that the hypothalamus is an important integral part in the cyclic functioning of the pituitary and ovary. Certain stimuli to the pituitary important in the induction of ovulation may be inhibited if they interfere with normal hypothalamic-hypophyseal relationships. Certain psychologic, physical or chemical stimuli may provoke such interference. Untoward hypothalamic effects upon pituitary function may be diminished by the use of one of the ataractic drugs. Hence, we feel that the combined use of cyclic progesterone therapy and one of the ataractic drugs may offer an effective means of management of a patient with secondary amenorrhea who menstruates promptly after the administration of progesterone. The following ataractic preparations and dosages have been used in our studies: Reserpine-o.l to 0.25 mg. t.i.d. Meprobamate-400 mg. t.i.d. Promazine-50 mg. t.i.d. Chlorpromazine-50 mg. t.i.d.
The monthly injection of progesterone, together with use of one of the ataractic drugs, has resulted in significant therapeutic salvage of these patients with amenorrhea. The prognosis with respect to therapy and salvage is poor in the female who fails to show any menstrual flow following progesterone alone, but who menstruates following cyclic therapy with estrogen and progesterone. This is especially true in patients with a high follicle-stimulating hormone secretion who fail to respond to pregnant mares' serum administration. The possibility of the induction of a spontaneous normal menstrual period is rather unlikely. However, if there is congenital ovarian failure or if ovarian failure occurs before the age of 30 to 35, it is wise to place the patient on cyclic therapy with estrogen and progesterone alone, not only to induce spontaneous periods, but to maintain adequate secondary sex development. This form of therapy will also prevent the peculiar aging process that these patients exhibit as a result of their lack of gonadal steroids. The prime benefit is due to the estrogen alone. However, since persistent estrogen therapy would result in endometrial hyperplasia, progesterone is used at monthly intervals to induce periodic desquamation of the endometrium. The doses used are identical to those employed in Step 2 of our diagnostic scheme (Fig. 206) and are repeated at four week intervals. In addition to the physiologic benefits achieved
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from cyclic therapy in these patients, the psychologic uplift following the resultant periodic catamenia is of untold benefit. The individual with low or normal follicle-stimulating hormone, associated with an adequate response to pregnant mares' serum in whom the cyclic therapy with estrogen and progesterone is effective in inducing menstrual flow, may be made to respond to constant doses of estrogens. Such a procedure may be conducive to initiation of spontaneous menstrual periods. The therapy has been as follows: Trianisylchloroethylene (chlorotrianisene) 12 mg. q.i.d. is administered for four weeks, followed by injection of 100 mg. of progesterone in oil. The progesterone is administered at monthly intervals without subsequent chlorotrianisene as long as menstrual periods occur. When menses fail to take place, the patient is reprimed with the chlorotrianisene capsules (12 mg. q.i.d.) for four weeks and reinjected again with progesterone at monthly intervals. Administration of estrogen in this manner apparently results in sustained effects. Such therapy has been effective in inducing restoration of ovarian function in an occasional patient. The estrogen in these cases may increase the response of the ovary to gonadotropin.4-7 AMENORRHEA DUE TO EXTRAGENITAL CAUSES
AlUenorrhea of Thyroid Origin
The thyroid and the adrenal glands play important contributory parts in inducing amenorrhea in many patients. Establishment of the role of the thyroid may be by clinical criteria, such as obesity, lethargy, constipation, intolerance to cold weather, dryness of the skin and hair, decrease or loss of axillary or pubic hair, and decrease or loss of hair on the lateral aspects of the eyebrow. There is usually an accompanying thinning of cephalic hair. In addition to the clinical findings, one may also observe a low basal metabolic rate and elevated cholesterol, a flat glucose tolerance (increased glucose tolerance), a low protein-bound iodine and a low radioactive iodine uptake. On the basis of the laboratory and clinical data, the diagnosis of hypothyroidism may be established. It is felt that hypothyroidism may result in sufficient pituitary alteration with respect to gonadotropic hormone secretion so that aberrant ovarian function occurs. This subsequently results in menstrual abnormalities, characterized by primary or secondary amenorrhea. Therapy revolves around replacement administration of thyroid substances. These include the standard U.S.P. thyroid preparation, thyroglobulin, sodium-I-thyroxine, and more recently, triiodothyronine. In our experience the following doses are approximately equivalent, although there is certainly individual patient variation: 65 mg. of U.S.P. thyroid is equivalent to 65 mg. of thyroglobulin, to 0.2 to 0.3 mg. of sodium-I-thyroxine and to 75 to 100 gamma of triiodothyronine. * We have found that the latter two crystalline preparations because of their stand* Refers to l-triiodothyronine.
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ard chemical nature may achieve a more predictable and definitive result in thyroid therapy. All thyroid preparations available to date, except triiodothyronine, when administered in effective doses do not achieve their maximal therapeutic action until after some two to three weeks of administration. Conversely, it should be noted that it takes a similar period of time for the preparation to be completely dissipated. As a result, laboratory tests to establish thyroid function must not be done unless the patient has been off all thyroid therapy for at least two to three weeks. Triiodothyronine is an exception to the above inasmuch as it achieves its full therapeutic effect in 48 to 72 hours and is as rapidly dissipated. Hence, toxic manifestations due to overdosage will persist much longer with the other thyroid preparations than with triiodothyronine. Amenorrhea in the Hypometabolic Syndrome
Amenorrhea or menstrual disturbances also may be observed in the hypometabolic syndrome. This syndrome is characterized clinically by a feeling of tiredness, lethargy, associated with some of the clinical manifestations of hypothyroidism plus the following laboratory findings: a· low basal metabolic rate, an elevated cholesterol, a flat glucose tolerance test (increased glucose tolerance) and a low 17-ketosteroid excretion in addition to the fact that one finds a normal protein-bound iodine and radioactive iodine uptake. It is thought that these patients are unable to break down thyroxine into triiodothyronine, the basic thyroid hormone which the cells are capable of utilizing. In patients with the hypometabolic syndrome triiodothyronine will produce results considerably superior to those achieved with other thyroid preparations. One last word with respect to thyroid therapy. It is generally accepted that the maximum thyroid dose should be administered to patients with proven hypothyroidism of the hypometabolic syndrome. Maximal dose is usually achieved by approaching the toxic dose and then reducing this toxic dose to the next highest nontoxic dose level. Adrenogenital Syndrome
The adrenal cortex also may play an essential role in the causation of amenorrhea. Adrenal cortical factors and syndromes important in menstrual abnormalities and amenorrhea are rather complex and space does not permit their elaboration in this report. Briefly, it is felt that excessive function of one part of the adrenal cortex may be sufficient to induce abnormal pituitary-gonadal function and result in the adrenogenital syndrome. It is presumed that the zona reticularis of the adrenal cortex, because of an enzymatic defect in the zona fasciculata, produces excessive amounts of 17-ketosteroid precursors under the influence of increased and uninhibited secretion of ACTH.8-11 These 17-ketosteroid precursors, being sex steroid in character, will inhibit or markedly suppress pituitary gonadotropic hormone. As a result, abnormal pituitary-gonadal relation-
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Fig. 209. Classical adrenogenital syndrome. Twenty-eight year old woman with primary amenorrhea, masculine habitus, enlarged clitoris and 17-ketosteroid excretion ranging between 70 to 80 mg./24 hours. The patient responded readily to corticoid therapy by a decrease in 17-ketosteroid levels to normal accompanied by feminization and initiation of cyclic catamenia.
ship ensues. In the diagnosis of the adrenogenital syndrome, the characteristic finding in these patients is the increased 17-ketosteroid and pregnanetriol excretion at times associated with a hypoglycemic response in the glucose tolerance test at the fourth or fifth hour and a low or lownormal 17-hydroxysteroid excretion. Clinically, these patients usually exhibit an increased degree of hirsutism, menstrual dysfunction that may result in cessation of the catamenia, plus other findings indicating increased androgenicity depending on the severity of the adrenogenital syndrome (Figs. 209, 210). Diagnosis of the classical adrenogenital syndrome requires little clinical acumen (Fig. 209). The masculine build, poor secondary sex development, enlargement of the clitoris and increased hirsutism provide adequate criteria for the clinician to warrant the use of the specific laboratory procedures listed above. On the other hand, a high suspicion index is required to consider the diagnosis in a young female, adequately endowed with secondary sex characteristics, who may show some of the following findings (Fig. 210): (a) slight or no menstrual irregularities; (b) ovulatory defects characterized by proliferative endometrium, monophasic basal body temperature curve, and infertility; (c) mild hirsutism and/ or acne. These patients may show a 17-ketosteroid value only at the upper limits of normal. A therapeutic response to corticoids according to
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Fig. 210. Subclassical adrenogenital syndrome in a 21 year old woman with welldeveloped secondary sex characteristics, some hirsutism and acne, and a history of irregular menstrual periods. 17-Ketosteroids were moderately elevated. Patient well controlled on oral corticoid therapy with resumption of regular, cyclic menstrual periods.
the dose scheme suggested below will establish the correctness of the diagnosis. The response clinically would be characterized by the resumption of greater regularity in the menstrual periods, occurrence of ovulation and suppression of the 17-ketosteroids to low-normal values. Treatment. Corticoid therapy in both the classical and nonclassical adrenogenital syndrome accomplishes its effect by suppressing the increased ACTH secretion, thereby decreasing the excessive 17-ketosteroid precursor production. Suppression of these precursors removes their inhibiting effect upon pituitary gonadotropin secretion. The average doses of the different corticoids that may be employed are as follows: Cortisone-12.5 mg. t.i.d. Hydrocortisone-lO mg. t.i.d. Prednisone-2.5 mg. three to four times a dayll
The dose of the particular corticoid is determined by its effectiveness in restoring normal menstrual function and depressing the excessive 17ketosteroid excretion. The reduction of hirsutism which usually occurs in these patients is a late outcome of therapy and may not be observed until after some nine to 12 months of therapy, if at all.
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Fig. 211. Twenty-five year old woman with a history of rather sudden increase in hirsutism and presence of irregular menstrual periods. The patient's elevated 17-ketosteroids were not suppressed with prednisone. An adrenal neoplasm was subsequently demonstrated and removed.
It should be emphasized that parenteral therapy is not required for the adequate management of the patient with the adrenogenital syndrome. Oral therapy provides for ready adjustment of dose and ease of administration. It must also be stated that the doses of corticoids listed above are not associated with any untoward effects. Consequently, there is no demand upon the patient to adhere to a special diet or limit her intake of salt. If salt retention may be a factor in a particularly sensitive individual, the theoretical advantage of prednisone in these patients should be considered. It has been shown that this steroid may at times promote sodium excretion. 12 If administration of the corticoids is not associated with a decrease in a significantly elevated 17-ketosteroid level then one should consider the possibility of an adrenal or extra-adrenal neoplasm as the cause of the masculinizing syndrome (Fig. 211). Thus, the corticoids are both of diagnostic and therapeutic value in patients suspected of having an adrenogenital syndrome. Diagnostically the corticoids can establish the presence of the sub classical adrenogenital syndrome in a patient showing few clinical stigmata. In addition, the failure of corticoids in adequate doses to suppress significantly the elevated 17-ketosteroids establishes the presence of a neoplasm--either adrenal or extra-adrenal in originand demands the need for surgical exploration. SUMMARY
This brief review of amenorrhea provides the surgeon with some workable methods for both diagnosis and treatment of patients with amenor-
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rhea-a clinical complaint which at times may be simply treated and diagnosed, but which may at times be of profound concern and distress to both the patient and attending physician. The suggested diagnostic plan and recommended therapeutic program offer the physician an effective, workable scheme that may lead to some success in this disturbing problem. REFERENCES 1. Kupperman, H. S., Blatt, M. H. G., Wiesbader, H. and Studdiford, W. E.: Progesterone in Menstrual Disturbances. Obst. & Gynec. 1 :650-662, 1953. 2. Kupperman, H. S. and Studdiford, W. E.: Endocrine Therapy in Gynecologic Disorders. Postgraduate Med. 14:410-425, 1953. 3. Kupperman, H. S. and Lefkovics, S. C.: Diagnostic and Therapeutic Use of Progesterone in Problems of Sterility. Fertil. & Steril. In press. 4. Zondek, B.: Some Problems Related to Ovarian Function and to Pregnancy. Recent Proc. Hormone Res. 10:395-423 (Academy Press), 1954, New York. 5. Pencharz, R. L.: Effect of Estrogens and Androgens Alone and in Combination with Chorionic Gonadotropin on the Ovary of the Hypophysectomized Rat. Science 91 :554-555, 1940. 6. Selye, H. and CoUip, J. B.: Fundamental Factors in the Interpretation of Stimuli Influencing Endocrine Glands. Endocrinology 20:667-672, 1936. 7. Simpson, M. E., Evans, H. M., Fraenkel-Conrat, H. and Li, C. C.: Synergism of Estrogens with Pituitary Gonadotropin in Hypophysectomized Rats. Endocrinology 28:37-41, 1941. 8. Jailer, J.: Virilism. Bull. New York Acad. Med. 29:377-394, 1953. 9. Bongiovanni, A. M.: Detection of Pregnanediol and Pregnanetriol in Urine of Patients with Adrenal Hyperplasia; Suppression with Cortisone. Bull. Johns Hopkins Hosp. 92:244-251, 1953. 10. Kupperman, H. S., Finkler, R. and Burger, J. Quantitative Effect of Cortisone upon Ketosteroid Excretion and Clinica11Picture of the Adrenogenital Syndrome. J. Clin. Endocrinol. & Metab. 13:1109-1117, 1953. 11. Kupperman, H. S. and Others: Comparat.ive Effects of Metacortandrocin, 9Alpha-fluorohydrocortisone and Hydrocortisone upon ACTH Secretion in Man. J. Clin. Endocrinol. & Metab. 15:911-922, 1955. 12. Bunim, J. J., Pechet, M. M. and Bollet, A. Z.: Studies on Metacortandralone and Metacortandracin in Rheumatoid Arthritis. J.A.M.A.157:311-318, 1955. 550 First Avenue Ncw York 16, N. Y.
TREATMENT of the failure of menstruation, whether it is primary or secondary in nature, is predicated upon adequate diagnosis and understanding of the underlying mechanism triggering the aberration. One must be cognizant of a possible functional disturbance in the endocrine system as a major etiolgic factor. The intimate relationships existing between the pituitary, the master gland, and the subordinate glands of internal secretion necessitate proper identification of the organ or organ systems involved in the cessation of cyclic catamenia. The pituitary gland, while the dictator of the endocrine system, is to a certain extent governed in its activity by the glands it controls, much in the same manner that all dictators to a degree are controlled by the people to whom they dictate. Hence, in considering the problem of amenorrhea both with respect to etiology and therapy, one must understand the basic principles of the physiology of menstruation and realize that, while the pituitary gland may be an important entity in the etiology of amenorrhea, it can be adversely affected by the thyroid, adrenals and/or ovaries. In addition, the end organ itself, the endometrium or uterus, must also be evaluated before the dysfunction responsible for disturbing the mosaic pattern of menstrual function can be correctly identified. The primary menstrual axis-the pituitary, ovaries and uterus-must be virtually intact to permit normal menstrual function. The thyroid and the adrenal in their role as secondary factors may affect the primary axis by virtue of their effect upon pituitary function itself. The importance of the primary axis necessitates the relegation of thyroid and adrenal factors to the latter part of our discussion. From the Departments of Therapeutics and Obstetrics and Gynecology, New York University-BeUevue Medical Center and Obstetrical and Gynecological Service (Third Division), BeUevue Hospital, and Endocrine Clinic, Beth Israel Hospital, New York, N.Y. *Associate Professor of Medicine and Endocrinologist, New York University PostGraduate Medical College; Endocrinologist, Beth Israel Hospital, New York City, Methodist Hospital, Brooklyn, and the Margaret Sanger Research Bureau, New York City. 517
Herbert S. Kupperman
518 r-~
i PATIENT WITH LAMENORRHEA -~
I PROGES;ERONE
,,//C, 1! 8~~~0s ;::::fi:~-' PITUITARY
iJ
CYCLIC OR COMBINED ESTROGEN a PROGESTERONE
I!
I
\
.-------------~
GONADOTROPHIN ASSAY (FSH) OR PREGNANT MARE'S SERUM
I
HIGH FSH NO RESPONSE TO PMS
I
LON OR NORMAL FSH PMS RESPONSE I INCREASED BREAST SIZE. 2_ MATURATION OF VAGINAL SMEAR 3 MENSES AFTER PROGES-
INDICATES PITUITARY / INSUFFICIENCY'
,o,·a I) (\
TERONE, 100 MGM LM
Fig. 206. Chart demonstrating diagnostic work-up in patient with amenorrhea. AMENORRHEA DUE TO DISTURBANCES IN THE PRIMARY MENSTRUAL AXIS
The organ systems involved in the primary menstrual axis now will be considered in greater detaiL Three organs are necessary for adequate menstrual function to take place normally: a uterus or functioning endometrium, without which menstruation obviously cannot take place; ovaries capable of stimulating the endometrium; and a pituitary gland replete with properly proportioned gonadotropins for maintaining the ovary in a functioning state of rhythmic activity.1-3 The patient who presents herself with a primary complaint of amenorrhea must be correctly diagnosed in order to ascertain the presence or absence of gross disease, either functional or organic, in any of the three organ systems listed above. The initial step presumes that there may be a primary defect in anyone of these three organ systems in the primary menstrual axis. To that end, the patient is subjected to the initial diagnostic procedure in order to properly identify this defect. As the initial step in this work-up, the patient receives 100 mg. of progesterone in 2 cc. of oil administered intramuscularly, following which the time of onset of menstruation to take place is established (Fig. 206). If menstruation occurs, then we have irrefutable evidence that the patient has an endometrium capable of menstruating; secondly, that the endometrium has been adequately stimulated by sufficient endogenous estrogenic activity so that the exogenously administered progesterone can act to induce withdrawal bleeding. Progesterone cannot induce withdrawal bleeding, unless previous and adequate priming with estrogen has been accomplished. Thirdly, the patient must have a pituitary gland
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Fig. 207. Eighteen year old obese female with primary amenorrhea and mental retardation misdiagnosed as Frohlich's syndrome. Error in diagnosis established by occurrence of menstrual period after progesterone medication.
capable of secreting sufficient amounts of follicle-stimulating hormone to promote endogenous estrogenic production by the ovary. Thus, by the simple procedure of a single injection of progesterone and the presence of menstruation thereafter, one has eliminated any possible serious defect in the pituitary, ovary or uterine axis (Fig. 206). Menstruation should occur "'ithin two weeks after the progesterone injection and usually takes place within five to seven days. The patients who respond to this type of regimen ,yho have not menstruated before or who have previously exhibited secondary amenorrhea fall into a number of different categories. A specific example is a young female referred to our clinic with a diagnosis of Frohlich's syndrome (Fig. 207). The patient was short and obese, 18 years old, mentally retarded with inadequate development of her secondary sex characteristics. She was given 100 mg. of progesterone, following which she promptly menstruated. The resultant menstruation ruled out unequivocally the presence of Frohlich's syndrome inasmuch as individuals with this syndrome must by definition have pituitary insufficiency. The presence of
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Herbert S. Kupperman
Fig. 208. Two patients with pseudocyesis of some 8 months' duration. Diagnosis established by clinical findings and menstrual period taking place after progesterone administration.
menstruation after progesterone indicates there could not be significant pluriglandular insufficiency based upon the reasoning listed above. Other patients with amenorrhea in whom the etiological problem may be readily resolved are those in whom cessation of menstruation may be of psychosomatic origin. Thus, the young lady who desires pregnancy so strongly that she eventually believes she is pregnant and develops the classical picture of pseudocyesis, may menstruate very promptly after progesterone administration (Fig. 208). Menstruation so induced eliminates the need for any further extensive laboratory work-up. On the other hand, the woman who has been a bit promiscuous, fears pregnancy and as a result stops menstruating will also experience a menstrual flow after progesterone administration. Many other patients develop amenorrhea secondary to persistent follicular or proliferative activity associated with anovulatory cycles. In these females mental anxiety may be of sufficient extent to inhibit proper sequential secretion of the pituitary gonadotropic hormones. They may be made to menstruate very promptly after progesterone therapy. The primary underlying disturbance disrupting sequential pituitary function in such patients and some of those listed above many be in the hypothalamus. The hypothalamus is an im-
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portant factor in normal hypophyseal activity and is involved in the integration of one's emotions and pituitary function. Progesteroneinduced catamenia may be instrumental in re-establishing a more normal hypothalamic-hypophyseal relationship. Thus, menstruation occurring after the administration of progesterone is not only of diagnostic importance, but also may be of therapeutic advantage. Specific therapy in these patients will be considered below. If the patient does not menstruate after progesterone alone, we have no evidence of the functional integrity of the three organs mentioned above and the next step in our regimen is required. This demands the administration of cyclic therapy with estrogen and progesterone (Fig. 206, Step 2). Our usual procedure in this clinic involves the initial administration of an oral estrogen for three weeks. The following preparations have been employed: 0.05 mg. tablets of ethinyl estradiol or 1.25 mg. tablets of conjugated estrogens (equine) or comparable doses of other estrogens, all being administered twice a day for a period of three weeks. Single injections of parenteral estrogens may also be employed. Two such preparations, estradiol valerate and estradiol cyclopentylpropionate, may be administered as oil solutions-in single doses of 20 and 10 mg. respectively. The patient again receives 100 mg. of progesterone in oil intramuscularly following the three week course of oral estrogen therapy or two weeks after the single intramuscular injection of the longacting estrogen preparation. Here one of two responses occur: the patient either menstruates or she fails to menstruate. The failure of menstruation to occur indicts the uterus in the causation of the amenorrhea. The specific uterine factors that may be indicted in amenorrhea are the following: (1) Uterine agenesis. One such case has been seen in this clinic in which the cervix and fundus were represented merely by a fold of peritoneum. (2) After severe endometritis, particularly after an acidfast infection. This is a common cause of amenorrhea in the Near East. (3) Following destruction of the endometrium by radiation. (4) Following a dilatation and curettage for an incomplete abortion or following postpartum hemorrhage. In the latter cases, where the endometrium is soft and succulent, the sharp curette even in the most competent hands may remove enough of the endometrium so that further regeneration of endometrial tissue cannot occur. This may result in permanent amenorrhea. Prognosis for restoration of menstrual function in patients with uterine failure is poor in all categories with occasional exception in the last group. In those females in whom partial obliteration of the uterine cavity has been induced by the surgical removal of the endometrial lining, one may sometimes stimulate the remnant of endometrial tissue to proliferate sufficiently by administration of high doses of estrogenic substancessome three to four times those ordinarily used in priming. These high doses have been helpful in salvaging some patients with mechanical destruction of the endometrium.
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At times amenorrhea resistant to cyclic therapy with estrogen and progesterone may occur due to cervical stenosis following instrumentation or a surgical procedure, such as a dilatation and curettage. However, in these patients there is the tell-tale evidence of the history of amenorrhea associated with cyclic pain at monthly intervals, and usually accompanied by some uterine enlargement. A sanguineous discharge following fenestration of the stenotic os confirms the diagnosis. This procedure then is followed invariably by cyclic resumption of the catamenia. If the patient does menstruate after the combined cyclic therapy with estrogen and progesterone, one may assume that the integrity of the endometrium and uterus is intact and that further steps must be undertaken to establish the role of the pituitary or the ovary in the etiology of the amenorrhea. The patient is then subjected to the next diagnostic procedure (Fig. 206, Step 3). This involves the intramuscular administration of pregnant mares' serum gonadotropin and/or the performance of an urinary gonadotropin assay. The pregnant mares' serum is administered in doses of 500 I.D. three times a week for three weeks. This may result in ovarian stimulation. Stimulation of ovarian function would be characterized by: (1) increase of breast size; (2) increased vaginal secretion; (3) vaginal maturation as evidenced by vaginal smears taken before and after administration of the gonadotropin; (4) menstrual period occurring after the patient receives 100 mg. of progesterone in oil some three to five days after the last pregnant mares' serum injection. The presence of these signs of ovarian stimulation would indicate that the organ responsible for the amenorrhea is not the gonads, but in all probability is the pituitary gland. The failure of response to pregnant mares' serum would indict the ovaries for the absence of the catamenia and may indicate ovarian agenesis. The performance of a urinary gonadotropic assay would yield essentially confirmatory data to the pregnant mares' serum gonadotropin; i.e., a high follicle-stimulating hormone excretion would be noted in those patients who failed to respond to pregnant mares' serum. On the other hand, a low or normal gonadotropic excretion would be obtained in those females who show a response to pregnant mares' serum. Treatlllent
The treatment of a patient with amenorrhea based upon disturbances in the pituita-ry-ovarian-uterine axis may involve several modalities of therapy. In the individual who fails to menstruate spontaneously, but shows a catamenia following the administration of progesterone, the advisable form of therapy is to continue the progesterone medication at monthly intervals for the next three or four months. This form of therapy may aid in reinstituting periods which had been held in abeyance in the past. The mechanism of action whereby cyclic therapy with progesterone may initiate menses in these patients must rest on the obtuse explanation of the interrelation between the pituitary gland and the hormone prod-
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ucts of its end organs. Progesterone may induce certain regulatory effects upon pituitary secretion. In addition to the use of progesterone, we have found that the employment of one of the ataractic drugs may have a beneficial effect upon the induction of cyclic menstruation in these individuals. These ataractic drugs have the potentiality of inducing a more normal hypothalamicpituitary relationship by virtue of their effect upon the hypothalamus. It has been adequately demonstrated in animals, at least, that the hypothalamus is an important integral part in the cyclic functioning of the pituitary and ovary. Certain stimuli to the pituitary important in the induction of ovulation may be inhibited if they interfere with normal hypothalamic-hypophyseal relationships. Certain psychologic, physical or chemical stimuli may provoke such interference. Untoward hypothalamic effects upon pituitary function may be diminished by the use of one of the ataractic drugs. Hence, we feel that the combined use of cyclic progesterone therapy and one of the ataractic drugs may offer an effective means of management of a patient with secondary amenorrhea who menstruates promptly after the administration of progesterone. The following ataractic preparations and dosages have been used in our studies: Reserpine-o.l to 0.25 mg. t.i.d. Meprobamate-400 mg. t.i.d. Promazine-50 mg. t.i.d. Chlorpromazine-50 mg. t.i.d.
The monthly injection of progesterone, together with use of one of the ataractic drugs, has resulted in significant therapeutic salvage of these patients with amenorrhea. The prognosis with respect to therapy and salvage is poor in the female who fails to show any menstrual flow following progesterone alone, but who menstruates following cyclic therapy with estrogen and progesterone. This is especially true in patients with a high follicle-stimulating hormone secretion who fail to respond to pregnant mares' serum administration. The possibility of the induction of a spontaneous normal menstrual period is rather unlikely. However, if there is congenital ovarian failure or if ovarian failure occurs before the age of 30 to 35, it is wise to place the patient on cyclic therapy with estrogen and progesterone alone, not only to induce spontaneous periods, but to maintain adequate secondary sex development. This form of therapy will also prevent the peculiar aging process that these patients exhibit as a result of their lack of gonadal steroids. The prime benefit is due to the estrogen alone. However, since persistent estrogen therapy would result in endometrial hyperplasia, progesterone is used at monthly intervals to induce periodic desquamation of the endometrium. The doses used are identical to those employed in Step 2 of our diagnostic scheme (Fig. 206) and are repeated at four week intervals. In addition to the physiologic benefits achieved
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from cyclic therapy in these patients, the psychologic uplift following the resultant periodic catamenia is of untold benefit. The individual with low or normal follicle-stimulating hormone, associated with an adequate response to pregnant mares' serum in whom the cyclic therapy with estrogen and progesterone is effective in inducing menstrual flow, may be made to respond to constant doses of estrogens. Such a procedure may be conducive to initiation of spontaneous menstrual periods. The therapy has been as follows: Trianisylchloroethylene (chlorotrianisene) 12 mg. q.i.d. is administered for four weeks, followed by injection of 100 mg. of progesterone in oil. The progesterone is administered at monthly intervals without subsequent chlorotrianisene as long as menstrual periods occur. When menses fail to take place, the patient is reprimed with the chlorotrianisene capsules (12 mg. q.i.d.) for four weeks and reinjected again with progesterone at monthly intervals. Administration of estrogen in this manner apparently results in sustained effects. Such therapy has been effective in inducing restoration of ovarian function in an occasional patient. The estrogen in these cases may increase the response of the ovary to gonadotropin.4-7 AMENORRHEA DUE TO EXTRAGENITAL CAUSES
AlUenorrhea of Thyroid Origin
The thyroid and the adrenal glands play important contributory parts in inducing amenorrhea in many patients. Establishment of the role of the thyroid may be by clinical criteria, such as obesity, lethargy, constipation, intolerance to cold weather, dryness of the skin and hair, decrease or loss of axillary or pubic hair, and decrease or loss of hair on the lateral aspects of the eyebrow. There is usually an accompanying thinning of cephalic hair. In addition to the clinical findings, one may also observe a low basal metabolic rate and elevated cholesterol, a flat glucose tolerance (increased glucose tolerance), a low protein-bound iodine and a low radioactive iodine uptake. On the basis of the laboratory and clinical data, the diagnosis of hypothyroidism may be established. It is felt that hypothyroidism may result in sufficient pituitary alteration with respect to gonadotropic hormone secretion so that aberrant ovarian function occurs. This subsequently results in menstrual abnormalities, characterized by primary or secondary amenorrhea. Therapy revolves around replacement administration of thyroid substances. These include the standard U.S.P. thyroid preparation, thyroglobulin, sodium-I-thyroxine, and more recently, triiodothyronine. In our experience the following doses are approximately equivalent, although there is certainly individual patient variation: 65 mg. of U.S.P. thyroid is equivalent to 65 mg. of thyroglobulin, to 0.2 to 0.3 mg. of sodium-I-thyroxine and to 75 to 100 gamma of triiodothyronine. * We have found that the latter two crystalline preparations because of their stand* Refers to l-triiodothyronine.
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ard chemical nature may achieve a more predictable and definitive result in thyroid therapy. All thyroid preparations available to date, except triiodothyronine, when administered in effective doses do not achieve their maximal therapeutic action until after some two to three weeks of administration. Conversely, it should be noted that it takes a similar period of time for the preparation to be completely dissipated. As a result, laboratory tests to establish thyroid function must not be done unless the patient has been off all thyroid therapy for at least two to three weeks. Triiodothyronine is an exception to the above inasmuch as it achieves its full therapeutic effect in 48 to 72 hours and is as rapidly dissipated. Hence, toxic manifestations due to overdosage will persist much longer with the other thyroid preparations than with triiodothyronine. Amenorrhea in the Hypometabolic Syndrome
Amenorrhea or menstrual disturbances also may be observed in the hypometabolic syndrome. This syndrome is characterized clinically by a feeling of tiredness, lethargy, associated with some of the clinical manifestations of hypothyroidism plus the following laboratory findings: a· low basal metabolic rate, an elevated cholesterol, a flat glucose tolerance test (increased glucose tolerance) and a low 17-ketosteroid excretion in addition to the fact that one finds a normal protein-bound iodine and radioactive iodine uptake. It is thought that these patients are unable to break down thyroxine into triiodothyronine, the basic thyroid hormone which the cells are capable of utilizing. In patients with the hypometabolic syndrome triiodothyronine will produce results considerably superior to those achieved with other thyroid preparations. One last word with respect to thyroid therapy. It is generally accepted that the maximum thyroid dose should be administered to patients with proven hypothyroidism of the hypometabolic syndrome. Maximal dose is usually achieved by approaching the toxic dose and then reducing this toxic dose to the next highest nontoxic dose level. Adrenogenital Syndrome
The adrenal cortex also may play an essential role in the causation of amenorrhea. Adrenal cortical factors and syndromes important in menstrual abnormalities and amenorrhea are rather complex and space does not permit their elaboration in this report. Briefly, it is felt that excessive function of one part of the adrenal cortex may be sufficient to induce abnormal pituitary-gonadal function and result in the adrenogenital syndrome. It is presumed that the zona reticularis of the adrenal cortex, because of an enzymatic defect in the zona fasciculata, produces excessive amounts of 17-ketosteroid precursors under the influence of increased and uninhibited secretion of ACTH.8-11 These 17-ketosteroid precursors, being sex steroid in character, will inhibit or markedly suppress pituitary gonadotropic hormone. As a result, abnormal pituitary-gonadal relation-
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Fig. 209. Classical adrenogenital syndrome. Twenty-eight year old woman with primary amenorrhea, masculine habitus, enlarged clitoris and 17-ketosteroid excretion ranging between 70 to 80 mg./24 hours. The patient responded readily to corticoid therapy by a decrease in 17-ketosteroid levels to normal accompanied by feminization and initiation of cyclic catamenia.
ship ensues. In the diagnosis of the adrenogenital syndrome, the characteristic finding in these patients is the increased 17-ketosteroid and pregnanetriol excretion at times associated with a hypoglycemic response in the glucose tolerance test at the fourth or fifth hour and a low or lownormal 17-hydroxysteroid excretion. Clinically, these patients usually exhibit an increased degree of hirsutism, menstrual dysfunction that may result in cessation of the catamenia, plus other findings indicating increased androgenicity depending on the severity of the adrenogenital syndrome (Figs. 209, 210). Diagnosis of the classical adrenogenital syndrome requires little clinical acumen (Fig. 209). The masculine build, poor secondary sex development, enlargement of the clitoris and increased hirsutism provide adequate criteria for the clinician to warrant the use of the specific laboratory procedures listed above. On the other hand, a high suspicion index is required to consider the diagnosis in a young female, adequately endowed with secondary sex characteristics, who may show some of the following findings (Fig. 210): (a) slight or no menstrual irregularities; (b) ovulatory defects characterized by proliferative endometrium, monophasic basal body temperature curve, and infertility; (c) mild hirsutism and/ or acne. These patients may show a 17-ketosteroid value only at the upper limits of normal. A therapeutic response to corticoids according to
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Fig. 210. Subclassical adrenogenital syndrome in a 21 year old woman with welldeveloped secondary sex characteristics, some hirsutism and acne, and a history of irregular menstrual periods. 17-Ketosteroids were moderately elevated. Patient well controlled on oral corticoid therapy with resumption of regular, cyclic menstrual periods.
the dose scheme suggested below will establish the correctness of the diagnosis. The response clinically would be characterized by the resumption of greater regularity in the menstrual periods, occurrence of ovulation and suppression of the 17-ketosteroids to low-normal values. Treatment. Corticoid therapy in both the classical and nonclassical adrenogenital syndrome accomplishes its effect by suppressing the increased ACTH secretion, thereby decreasing the excessive 17-ketosteroid precursor production. Suppression of these precursors removes their inhibiting effect upon pituitary gonadotropin secretion. The average doses of the different corticoids that may be employed are as follows: Cortisone-12.5 mg. t.i.d. Hydrocortisone-lO mg. t.i.d. Prednisone-2.5 mg. three to four times a dayll
The dose of the particular corticoid is determined by its effectiveness in restoring normal menstrual function and depressing the excessive 17ketosteroid excretion. The reduction of hirsutism which usually occurs in these patients is a late outcome of therapy and may not be observed until after some nine to 12 months of therapy, if at all.
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Fig. 211. Twenty-five year old woman with a history of rather sudden increase in hirsutism and presence of irregular menstrual periods. The patient's elevated 17-ketosteroids were not suppressed with prednisone. An adrenal neoplasm was subsequently demonstrated and removed.
It should be emphasized that parenteral therapy is not required for the adequate management of the patient with the adrenogenital syndrome. Oral therapy provides for ready adjustment of dose and ease of administration. It must also be stated that the doses of corticoids listed above are not associated with any untoward effects. Consequently, there is no demand upon the patient to adhere to a special diet or limit her intake of salt. If salt retention may be a factor in a particularly sensitive individual, the theoretical advantage of prednisone in these patients should be considered. It has been shown that this steroid may at times promote sodium excretion. 12 If administration of the corticoids is not associated with a decrease in a significantly elevated 17-ketosteroid level then one should consider the possibility of an adrenal or extra-adrenal neoplasm as the cause of the masculinizing syndrome (Fig. 211). Thus, the corticoids are both of diagnostic and therapeutic value in patients suspected of having an adrenogenital syndrome. Diagnostically the corticoids can establish the presence of the sub classical adrenogenital syndrome in a patient showing few clinical stigmata. In addition, the failure of corticoids in adequate doses to suppress significantly the elevated 17-ketosteroids establishes the presence of a neoplasm--either adrenal or extra-adrenal in originand demands the need for surgical exploration. SUMMARY
This brief review of amenorrhea provides the surgeon with some workable methods for both diagnosis and treatment of patients with amenor-
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rhea-a clinical complaint which at times may be simply treated and diagnosed, but which may at times be of profound concern and distress to both the patient and attending physician. The suggested diagnostic plan and recommended therapeutic program offer the physician an effective, workable scheme that may lead to some success in this disturbing problem. REFERENCES 1. Kupperman, H. S., Blatt, M. H. G., Wiesbader, H. and Studdiford, W. E.: Progesterone in Menstrual Disturbances. Obst. & Gynec. 1 :650-662, 1953. 2. Kupperman, H. S. and Studdiford, W. E.: Endocrine Therapy in Gynecologic Disorders. Postgraduate Med. 14:410-425, 1953. 3. Kupperman, H. S. and Lefkovics, S. C.: Diagnostic and Therapeutic Use of Progesterone in Problems of Sterility. Fertil. & Steril. In press. 4. Zondek, B.: Some Problems Related to Ovarian Function and to Pregnancy. Recent Proc. Hormone Res. 10:395-423 (Academy Press), 1954, New York. 5. Pencharz, R. L.: Effect of Estrogens and Androgens Alone and in Combination with Chorionic Gonadotropin on the Ovary of the Hypophysectomized Rat. Science 91 :554-555, 1940. 6. Selye, H. and CoUip, J. B.: Fundamental Factors in the Interpretation of Stimuli Influencing Endocrine Glands. Endocrinology 20:667-672, 1936. 7. Simpson, M. E., Evans, H. M., Fraenkel-Conrat, H. and Li, C. C.: Synergism of Estrogens with Pituitary Gonadotropin in Hypophysectomized Rats. Endocrinology 28:37-41, 1941. 8. Jailer, J.: Virilism. Bull. New York Acad. Med. 29:377-394, 1953. 9. Bongiovanni, A. M.: Detection of Pregnanediol and Pregnanetriol in Urine of Patients with Adrenal Hyperplasia; Suppression with Cortisone. Bull. Johns Hopkins Hosp. 92:244-251, 1953. 10. Kupperman, H. S., Finkler, R. and Burger, J. Quantitative Effect of Cortisone upon Ketosteroid Excretion and Clinica11Picture of the Adrenogenital Syndrome. J. Clin. Endocrinol. & Metab. 13:1109-1117, 1953. 11. Kupperman, H. S. and Others: Comparat.ive Effects of Metacortandrocin, 9Alpha-fluorohydrocortisone and Hydrocortisone upon ACTH Secretion in Man. J. Clin. Endocrinol. & Metab. 15:911-922, 1955. 12. Bunim, J. J., Pechet, M. M. and Bollet, A. Z.: Studies on Metacortandralone and Metacortandracin in Rheumatoid Arthritis. J.A.M.A.157:311-318, 1955. 550 First Avenue Ncw York 16, N. Y.