Angiotensin-sodium interaction in blood pressure maintenance of renal hypertensive and normotensive rats.

A specific inhibitor of angiotensin II was used in rats to investigate whether angiotensin is involved in the maintenance of blood pressure in one-kidney Goldblatt hypertension, in which plasma renin levels are not usually increased. The inhibitor produced marked falls in blood pressure, often down to normal levels in the hypertensive animals only when they were depleted in sodium and not after sodium repletion. Much lesser but still significant falls in blood pressure were also produced in normotensive sodium-depleted rats but not in repleted rats. We conclude that the importance of angiotensin for maintaining blood pressure is largely determined by its relation to available sodium or fluid volume, since the renin component in maintenance of either the hypertensive or the normotensive state could be exposed only by sodium deprivation. Therefore, volume expansion per se or other pressor factors may be involved in maintaining blood pressure of these sodium-replete normotensive or hypertensive animals.

Activity of (des-Aspartyl1)-angiotensin II and angiotensin II in man. Differences in blood pressure and adrenocortical response during normal and low sodium intake.

This study was designed to compare the effect of [des-Aspartyl(1)]-angiotensin II ([des-Asp]-A II) and angiotensin II (A II) on blood pressure and aldosterone production in man under conditions of normal and low sodium (Na) intake. Seven normal male subjects in balance on constant normal Na intake (U(Na) V 160.3+/-5.0 meq/24 h) for 5 days received A II and [des-Asp]-A II infusions on two consecutive days; 1 mo later they were restudied after 5 days of low Na intake (U(Na) V 10.5+/-1.6 meq/24 h). Each dose was infused for 30 min, sequentially. During normal Na intake, [des-Asp]-A II from 2 to 18 pmol/kg per min increased mean blood pressure from 85.2+/-3 to 95.3+/-5 mm Hg and plasma aldosterone concentration from 5.2+/-1.1 to 14.3+/-1.9 ng/100 ml. During low Na intake, the same dose of [des-Asp]-A II increased mean blood pressure from 83.7+/-3 to 86.7+/-3 mm Hg and plasma aldosterone concentration from 34.4+/-6.0 to 51.0+/-8.2 ng/100 ml. In contrast, A II from 2 to 6 pmol/kg per min during normal Na intake increased mean blood pressure from 83.3+/-4 to 102.3+/-4 mm Hg and plasma aldosterone concentration from 7.0+/-2.2 to 26.8+/-2.0 ng/100 ml; during low Na intake, A II increased mean blood pressure from 83.0+/-3 to 96.0+/-4 mm Hg and plasma aldosterone concentration from 42.0+/-9.7 to 102.2+/-15.4 ng/100 ml. A II and [des-Asp]-A II were equally effective in suppressing renin release. Plasma cortisol and Na and K concentration did not change. The effects of two doses (2 and 6 pmol/kg per min) of each peptide on blood pressure and aldosterone production were evaluated. During normal Na intake, [des-Asp]-A II had 11-36% of the pressor activity and 15-30% of the steroidogenic activity of A II. Na deprivation attenuated the pressor response and sensitized the adrenal cortex to both peptides, but the increase in steroidogenesis was greater with [des-Asp]-A II than with A II. The dose-response curves for [des-Asp]-A II with respect to blood pressure and aldosterone production were not parallel, and although no maximum was established for A II, [des-Asp]-A II was less efficacious.In summary, (a) [des-Asp]-A II has biologic activity in man, (b) [des-Asp]-A II is less efficacious than A II in stimulating aldosterone production, (c) Na deprivation sensitizes the adrenal cortex more markedly to [des-Asp]-A II than A II, and (d) dose-response curves for the two peptides differ, suggesting the possibility that they act at different receptor sites in vascular smooth muscle and the adrenal cortex.

Activity of [des-aspartyl1]-angiotensin II in primary aldosteronism.

This study describes the effects of [des-Aspartyl(1)]-angiotensin II ([des-Asp]-AII) on blood pressure and aldosterone production in patients with primary aldosteronism due to aldosterone-producing adrenal adenoma (APA) and idiopathic adrenal hyperplasia (IHA), and in normotensive control subjects. 10 patients with primary aldosteronism, 7 with APA and 3 with IHA, and 6 normotensive control subjects were placed on a constant 150-meq sodium diet for 4 days. [des-Asp]-AII was infused for 30 min at 6, 12, and 18 pmol/kg per min. Three groups of patients were identified on the basis of aldosterone response to [des-Asp]-AII. Group I, composed of normotensive control subjects, showed incremental increases in plasma aldosterone concentration from 6+/-1 to 14+/-3 ng/100 ml (P < 0.01) with [des-Asp]-AII infusion. Group II, composed of patients with primary aldosteronism, showed incremental increases in plasma aldosterone concentration from 33+/-8 to 65+/-13 ng/100 ml (P < 0.05) with 12 pmol/kg per min of [des-Asp]-AII. Group III, also composed of patients with primary aldosteronism, showed no increase of plasma aldosterone concentration with [des-Asp]-AII. Groups I and II showed similar percentage increases in plasma aldosterone concentration (P = NS). Group III showed significantly lower aldosterone responses than group I (P < 0.01). Group II included all patients with IHA and two patients with APA. Group III included only patients with APA. The blood pressure responses to [des-Asp]-AII of subjects in group I did not differ significantly from those of groups II or III.Thus, patients with IHA and a subgroup of patients with APA showed responsiveness to [des-Asp]-AII which was limited to adrenal cortical stimulation of aldosterone biosynthesis. This suggests that adrenal responsiveness to angiotensin is a major control mechanism in some forms of primary aldosteronism. The differential adrenal responsiveness to [des-Asp]-AII in patients with APA indicates either that there are two distinct subpopulations of APA, or that alteration in tumor response to angiotensin occurs during the natural progression of the disease history.

A questionnaire survey of current UK practice for adjuvant radiotherapy following surgery for oral and oropharyngeal squamous cell carcinoma.

A postal questionnaire was sent to 281 members of the British Association of Head and Neck Oncologists (BAHNO) to survey, which patients should receive adjuvant radiotherapy following primary surgery for oral and oropharyngeal squamous cell carcinoma (O&OSCC). Two hundred and one clinicians were involved in decision making for adjuvant radiotherapy in O&OSCC, of which, 132 (66%) responded. Apart from general agreement that patients with involved margins or extracapsular spread (ECS) should have adjuvant radiotherapy and that in patients with small tumours with clear margins and no neck metastasis, radiotherapy should be avoided, opinion was divided. Considerable variation in opinion in the UK was identified for a subgroup of intermediate risk patients as to whether they should have adjuvant radiotherapy. The majority of respondents (95%) would consider submitting patients to a prospective multi-centre trial. There is a need for research regarding adjuvant radiotherapy for O&OSCC patients at intermediate risk of relapse following primary surgery.

An assessment of deprivation as a factor in the delays in presentation, diagnosis and treatment in patients with oral and oropharyngeal squamous cell carcinoma.

This study explores the relationship between deprivation and patient and professional delays in presentation and treatment of oral and oropharyngeal squamous cell carcinoma. The cohort comprised 559 consecutive previously untreated patients presenting to the Regional Maxillofacial Unit, Liverpool from 1 January 1992 to 31 December 2002. All had primary surgery. The head and neck database was searched together with a review of casenotes. Deprivation was scored using the Index of Multiple Deprivation 2000 (IMD 2000) from patient post codes. PATIENT DELAY: Similar numbers of patients presented to general dental and general medical practitioners. The predominant presenting symptom was either an ulcer or swelling and 38% had symptoms for 3 or more months. Patients with shorter duration of symptoms tended to be smokers, drinkers, with lower gum and floor of mouth tumours, and more advanced disease. Primary health professional, patient age, gender, marital status, and deprivation showed no obvious correlation with patient delay. PROFESSIONAL DELAY: For 78% of patients a referral letter from GPs and GDPs was sent to the MFU on the same day as the primary consultation. There was on average about 3 weeks from referral to definitive diagnosis and about another 3 weeks before having surgery. Professional delay was shorter in patients with more advanced tumours and for patients living in the most deprived of wards. Deprivation did not seem to significantly lengthen presentation or referral however it may be that it is associated with more rapidly growing tumours.

A comparison of outcomes for patients with oral squamous cell carcinoma at intermediate risk of recurrence treated by surgery alone or with post-operative radiotherapy.

Controversy remains about which patients at intermediate risk of recurrence of oral squamous cell carcinoma would benefit from radiotherapy. A retrospective review of computerised database and medical records for 462 consecutive patients at the Regional Maxillofacial Unit in Liverpool who were treated with primary surgery with or without post-operative radiotherapy was carried out. We classified 29% (134) of patients as being at 'low' risk of disease recurrence (pT1-2, N0 with clear margins), 29% (135) at 'high' risk (involved margins or lymph node extracapsular spread) and the remaining 42% (193) at 'intermediate' risk. Of those at intermediate risk, 41% (80/193) received adjuvant radiotherapy and their 5 year survival (SE) was 54% (6%) compared to 71% (5%) for those with primary surgery alone (P=0.002). A higher proportion of patients having radiotherapy had loco-regional recurrence (19/80 24%) compared to those treated by surgery alone (17/113 15%). The improved salvage rate for recurrent disease in the surgery alone group (8/17 53%), compared to those receiving radiotherapy (2/19 13%, P=0.05), indicates an advantage in withholding radiotherapy for patients at intermediate risk of recurrence. This study indicates a potential disadvantage associated with the use of postoperative radiotherapy for patients at intermediate risk of recurrence. A randomised trial comparing a watch and wait policy to postoperative radiotherapy in patients with an intermediate risk of recurrence is required to confirm the trend indicated in this retrospective data.

The influence of hyperbaric oxygen on the outcome of patients treated for osteoradionecrosis: 8 year study.

This study was a retrospective review of treatment outcomes of osteoradionecrosis (ORN) of the mandible over an 8-year period, with specific reference to the role of hyperbaric oxygen therapy (HBOT). The presentation and management of 23 patients treated for ORN was studied by categorising them into three grades according to the severity of clinical and radiographic involvement. At presentation there were 13 patients with grade I ORN, six patients with grade II ORN and four patients with grade III ORN. HBOT was given to 10 patients in the grade I group, four patients in the grade II group and two patients in the grade III group. Overall eight patients (62%) with grade I, three patients (50%) with grade II and two patients (50%) with grade III were cured. In the patients who received HBOT the cure rate was 12.5% whilst in those without HBOT it was 86%. Although the cohort was small it seems that HBOT was of little benefit. HBOT is demanding for patients and has cost implications for the NHS; hence further clinical outcome data are urgently required with regard to its role in the management of ORN.

Inhibitors of 5alpha-reductase in the treatment of benign prostatic hyperplasia.

The description of 5alpha-reductase deficiency in male pseudohermaphroditism, characterization of type-1 and type-2 isoenzymes of 5alpha-reductase, and development of 4-aza steroid competitive inhibitors of 5alpha-reductase were milestones in the development of 5alpha-reductase inhibitors, a class of drugs approved for the treatment of symptomatic benign prostatic hyperplasia (BPH). Stromal and epithelial hyperplasia in the region of the prostate that surrounds the urethra begins in the fourth decade of life and by the sixth decade, the prevalence is 50%. Benign prostatic hyperplasia is a frequent cause of lower urinary tract symptoms, urinary tract infection, and acute urinary retention requiring surgical intervention. Medical options for treatment of symptomatic BPH include 1) the 5alpha-reductase inhibitors finasteride and dutasteride, 2) the alpha1-adrenergic antagonists doxazocin, terazosin, tamsulosin, and alfuzosin, and 3) the combination of a 5alpha-reductase inhibitor and a alpha1-adrenergic antagonist. By inhibiting the production of dihydrotestosterone (DHT) locally within the prostate gland, 5alpha-reductase inhibitors have the effect of reducing prostate volume, improving lower urinary tract symptoms, increasing peak urinary flow, and decreasing the risk of acute urinary retention and need for surgical intervention. Alpha-1 adrenergic antagonists relax the smooth muscle of the bladder neck and prostate, thereby decreasing the resistance to urine flow and increasing peak urinary flow and improving lower urinary tract symptoms. The alpha1-adrenergic antagonists are effective in the short-term, and reduce clinical progression of BPH, but do not reduce the long-term risk of urinary retention or need for surgical intervention. The 5alpha-reductase inhibitors are effective in the long-term, especially in men with large prostates, and reduce the clinical progression of BPH, and further reduce the long-term risk of urinary retention and need for surgical intervention. The combination of a 5alpha-reductase inhibitor and a alpha1-adrenergic antagonist significantly reduces the clinical progression of BPH over either drug class alone.

Trends in head and neck microvascular reconstructive surgery in Liverpool (1992-2001).

Microvascular reconstructive techniques in head and neck surgery are well established, but we are now entering an era of modification exemplified by perforator and free style free flaps. We present a review of the database introduced into the unit in 1992 over a 10-year period, during which time 977 patients with malignant disease were operated on and 620 defects were reconstructed with free flaps. There were 358 radial forearm flaps, 78 composite radial forearm flaps, 84 iliac crest flaps, 43 fibular flaps, 24 from the scapula, 26 from the latissimus dorsi, 4 from the rectus abdominis, and 3 from the lateral arm. The main changes over this time have been the use of more bulky flaps for larger resections of the tongue and the preference for iliac crest flaps over those from the fibula and forearm for composite reconstructions. Improving reliability of tissue transfer remains an important aim, and further development of reliable objective methods of monitoring of flaps is required.

Timing and presentation of recurrent oral and oropharyngeal squamous cell carcinoma and awareness in the outpatient clinic.

The aim of this study was to assess the timing of outpatient review appointments in relation to tumour recurrence. A retrospective review of 278 consecutive previously untreated patients with oral and oropharyngeal squamous cell carcinoma (SCC) between 1995 and 1999 was performed. Information on the time of recurrence, site, presentation, treatment and outcome was collected. There were 54 (19%) patients who developed recurrent disease. Recurrence occurred at a median time of 8 months after the initial operation and most (49/54) within 2 years. Thirty-five patients (65%) presented with a new lump (7 local, 22 regional and 5 locoregional). Our policy is to review patients once a month for the first year and every other month for the second year. Patients were seen less frequently than expected, and one in five patients attended half or less than half as frequently as intended in the first year. Although 20 patients were aware of new symptoms from their recurrent disease fewer than half (9) brought their appointment forward. This study has emphasised the need for close clinical follow-up of patients previously treated for oral/oropharyngeal SCC if recurrent tumours are to be discovered and treated at the earliest opportunity.

p53 mutations in squamous cell carcinoma of the head and neck predominate in a subgroup of former and present smokers with a low frequency of genetic instability.

We examined the p53 mutational profile of 65 squamous cell carcinomas of the head and neck (SCCHNs) from patients living in northwest England. Twenty-three p53 mutations were detected in 20 samples (31%). GC-->AT transitions were the predominant type of mutation. The p53 mutational profile of SCCHN tumors was similar to that of non-small cell lung tumors from patients within the same geographical area, supporting the idea of a common model for carcinogenesis in the upper respiratory tract. Statistical analysis showed that the incidence of p53 mutations among present and former smokers was significantly higher than that in nonsmokers (P < 0.02). In addition, p53 mutations were found to predominate in a group of SCCHN patients with low genetic damage, as indicated by the fractional allelic loss value. The above findings suggest an early initiating role for p53 and imply that at least two separate carcinogenic pathways may be involved in the development of SCCHN.

Synchronous oral carcinomas: independent or common clonal origin?

Second primary tumors in patients with head and neck cancer have a detrimental impact on long-term survival; at least 15% of patients develop additional tumors. Originally, it was hypothesized that multiple tumors developed independently after widespread epithelial exposure to carcinogens (the field cancerization theory), but recent molecular studies now support the alternative theory of a common clonal origin. If multiple tumors originate from the same clone, early genetic alterations in these cells should be common to all of the tumors. We have compared the pattern of allelic imbalance in paired tumors from five male patients with two synchronous oral squamous cell carcinomas and in peripheral dysplasia using microsatellite markers on chromosomes 3p, 9p, and 17p. Discordance, usually through loss of alternate alleles at the same microsatellite loci, was detected in two patients. The remaining three patients had identical alterations in their tumors. The changes identified occurred early in tumorigenesis, because, with only one exception, these were also present in the associated dysplasia. Thus, we provide evidence that synchronous oral squamous cell carcinomas are of independent origin in some patients but may be of common clonal origin in others.

Analysis of a mouse monoclonal antibody that reacts with a specific region of the human proximal tubule and subsets renal cell carcinomas.

Murine monoclonal antibody (mAb) F31 detects a heat-stable antigen (URO-8) found in the acidic lipid fraction of renal cancer cell extracts. Serological analysis of mAb F31 reactivity was assayed on 176 human cell lines. mAb F31 reacted with 38 of 45 renal cancers, a subpopulation of cells in primary cultures of normal renal epithelia, and two of 13 colon, two of 15 lung, and four of five ovarian cancers. No other epithelial, hematopoietic, or neuroectodermal cell lines tested were reactive. Immunofluorescence and immunoperoxidase analyses of fresh frozen tissue sections revealed mAb F31 reactivity in kidney, gastrointestinal tract, biliary canaliculi, bronchial epithelium, and skin. Within the kidney, mAb F31 immunoreactivity was confined to the straight portion of the proximal tubule. A panel composed of previously characterized mAbs as well as mAb F31 defines the antigenic phenotype of proximal convoluted tubular cells as URO-2+/URO-3+/URO-4+/URO-10+/URO-8-/URO-5-; proximal straight tubular cells as URO-2+/URO-3+/URO-4+/URO-10-/URO-8+/URO-5-; and cells of the descending thin limb of Henle as URO-2-/URO-3+ or -/URO-4+/URO-10-/URO-8-/URO-5+. While adult proximal tubular cells demonstrated reciprocal expression of URO-8 and URO-10, fetal kidney proximal tubule progenitor cells coexpressed both antigens (URO-10+/URO-8+). Fifty renal cancer specimens were typed with these antibodies. Fourteen cases were URO-10+/URO-8-, ten cases were URO-10-/URO-8+, and 25 cases expressed both antigens (URO-10+/URO-8+). These phenotypes are consistent with derivation of these particular subsets from the proximal convoluted tubule, the pars recta, or a proximal tubule progenitor cell, respectively. Only one specimen failed to express either URO-8 or URO-10.

Elevated expression of the c-myc oncoprotein correlates with poor prognosis in head and neck squamous cell carcinoma.

We quantitated c-myc oncoprotein in 44 squamous cell carcinomas of the head and neck using an enzyme-linked immunosorbence assay. The clinicopathological parameters of these patients were followed up for between 3 and 60 months and analysed for any correlations with observed levels of c-myc protein using the Kruskal-Wallis one-way analysis of variance method. Although no statistical correlation was found between different clinicopathological parameters (patient age, sex, TNM staging, number of lymph nodes invaded, extracapsular rupture of the tumour, its histopathological differentiation, or its site), the survival periods of patients with tumours possessing elevated levels of c-myc protein were found to be statistically shorter than those with lower levels of c-myc expression, (P less than 0.02). This indicates that c-myc expression may be an effective prognostic indicator in head and neck cancer.

Nipple differentiation in fetal male rats treated with an inhibitor of the enzyme 5 alpha-reductase: definition of a selective role for dihydrotestosterone.

In the rat, androgens are responsible for sexually dimorphic nipple differentiation. Nipples are expressed in the female, while in the male, the nipple anlage regress prenatally. Mammary gland development is present in both sexes. Treatment of pregnant Sprague-Dawley rats from days 12-21 of gestation with the aza-steroid 17 beta-N,N-diisopropylcarbamoyl-4-aza-5 alpha-androstan-3-one, a competitive inhibitor of the enzyme 5 alpha-reductase, resulted in nipple development in male offspring. Additionally, there was feminization of the external genitalia, with urethral displacement to the base of the phallus. The role of androgens in suppression of nipple anlage in the male rat fetus is known. This study, however, suggests for the first time a selective role for 5 alpha-dihydrotestosterone in regression of the nipple anlage in utero. Thus, 5 alpha-dihydrotestosterone may be critical not only for masculinization of the external genitalia, but also for inhibition of nipple development in the male rat fetus.

Responses of blood pressure and angiotensin-converting enzyme activity to acute captopril administration in normotensive and hypertensive rats.

The purpose of these experiments ws to determine the dose of captopril which resulted in essentially complete blockade of tissue and plasma converting enzyme activity (CEA) and to correlate the effect of this dose of inhibitor on blood pressure and CEA in a number of normotensive and hypertensive rat models. Oral administration of captopril (0.3-10 mg/kg) induced a dose-related attenuation of CEA in plasma freshly drawn from normotensive conscious rats. After storage at -20 or 4 C before assay, both captopril-treated and untreated plasmas displayed markedly greater CEA. The converting enzyme inhibitor induced a parallel shift to the right of angiotensin I-induced blood pressure responses, reaching a 100-fold displacement of dose-dependent responses in the presence of 10 mg/kg captopril. Sixty minutes after oral administration of 10 mg/kg captopril, plasma CEA was blocked completely in all normo- and hypertensive models studied. This dose of the inhibitor reduced blood pressure in the sodium-deplete normotensive rat, the spontaneously hypertensive rat, and the initial phase two-kidney, one-clip Goldblatt rat but not in the sodium-replete normotensive rat, the spontaneously hypertensive rat, the mineralocorticoid hypertensive rat, or the chronic phase of two-kidney, one-clip Goldblatt rat. It is concluded that acute administration of captopril at a dose which results in complete blockade of plasma converting enzyme and severe attenuation of tissue converting enzyme reduces blood pressure in animals with high PRA but not in animals in which the PRA is low. (Endocrinology 108: 536, 1981)

Formation of [Des-Asp1]angiotensin I by the perfused rat lung.

Bioassay studies have indicated that angiotensin I (Ang I), but not angiotensin II (Ang II), is degraded by the intact lung. The present study was an attempt to isolate and quantify pulmonary metabolites of Ang I. The pulmonary vascular bed of the rat was isolated and perfused at 4-6 ml/min (< 20 mm Hg) with oxygenated Krebs buffer containing 5% bovine serum albumin. [3H-Leu10]Ang I (3-10 ng) was administered, and pulmonary effluent samples were collected every 15 sec for 5 min. Peptides were isolated by Dowex chromatography and separated by thin layer chromatography. 3H-labeled peptides were eluted from the thin layer chromatography plate, and the Ang I and [des-Asp1]Ang I levels were estimated by RIA. These peptides were shown to be hydrolyzed by purified converting enzyme with the release of His-3H-Leu. About 25% of the [3H]Ang I administered was isolated as [3H-des-Asp1]Ang I, and this percentage increased to 33% after treatment with teprotide. These studies clearly demonstrate that [des-Asp1]Ang I is a major pulmonary metabolite of Ang I and suggest the presence of a pulmonary aminopeptidase which hydrolyzes Ang I but not Ang II.

Demonstration of a primary antidiuretic action of angiotensin II: effects of intrarenal converting enzyme inhibition in the conscious dog.

This study was designed to determine whether angiotensin II of renal origin physiologically mediates free water production by the kidney. The effect of renal arterial infusion of the angiotensin-converting enzyme inhibitor, teprotide, on renal function was studied under conditions of hydropenia and hydration in uninephrectomized conscious dogs after 5 days of equilibration on a normal or low sodium diet. In low sodium animals, intrarenal infusion of teprotide (0.25 microgram/kg.min) after 12 h of water restriction elicited marked increases in the glomerular filtration rate (GFR) and free water formation. After 24 h of water restriction, vasopressin (10 microU/min) was infused iv to achieve an effective maximal urine concentration. Under these conditions, the inhibitor again increased the GFR and free water formation. After the production of a state of stable hydration, intrarenal infusion of teprotide also increased the GFR and free water formation. In the normal sodium animals, intrarenal infusion of the converting enzyme inhibitor (2.5 microgram/kg.min) in 12-h hydropenic dogs failed to change the GFR but increased free water formation, while the inhibitor decreased effective renal plasma flow and free water formation in normal sodium, hydrated animals. These data suggest that angiotensin II of renal origin plays an important role in the control of free water production by the kidney.

Comparison of the effects of the 5 alpha-reductase inhibitor finasteride and the antiandrogen flutamide on prostate and genital differentiation: dose-response studies.

Studies were performed to compare the effects of 5 alpha-reductase inhibition and antiandrogen receptor blockade on differentiation of male internal and external genital structures and prostate in the rat. Dose-response studies were performed on male rats treated in utero during the period of sexual differentiation with either the potent 5 alpha-reductase inhibitor finasteride or the antiandrogen flutamide. The treated animals were raised to adulthood and killed, and genital structures were evaluated. Treatment with the 5 alpha-reductase inhibitor finasteride at a dose of 25 mg/kg.day resulted in significant feminization of the external genitalia. There was no further feminization of the genitalia at doses up to 300 mg/kg.day. Wolffian ductal differentiation occurred at all doses evaluated. Seminal vesicle weight, however, significantly decreased at 25 mg/kg.day, but without a further decrease at higher doses of the 5 alpha-reductase inhibitor. Vas deferens and epididymal weights were unchanged at all doses evaluated. There was a significant decrease in prostate size at 25 and 50 mg/kg.day, with no further decrease at higher doses. In flutamide-treated animals, complete feminization of the genitalia occurred at 24 mg/kg.day in all animals. At 18 mg/kg.day, Wolffian ductal differentiation occurred, but seminal vesicle weight was decreased. At dosages of 100, 200, and 300 mg/kg.day flutamide, the vas deferens was absent unilaterally or bilaterally, with small remnants of epididymal head and tail present. At dosages of 24 mg/kg.day and above, the prostate was absent. Studies with the 5 alpha-reductase inhibitor finasteride demonstrate the dependency of prostate and male external genital differentiation on dihydrotestosterone (DHT). However, unlike androgen receptor blockade with flutamide, finasteride did not totally abolish prostate differentiation or completely feminize the external genitalia, despite increasingly higher doses. Since there is no evidence of multiple 5 alpha-reductase isoenzymes to date in the rat, these results suggest that testosterone (T) can compensate for DHT to some degree at the level of the androgen receptor. Wolffian differentiation, however, was not affected by inhibition of DHT, demonstrating its T dependency, but seminal vesicle growth was impaired. Thus, inhibition of 5 alpha-reductase activity limits seminal growth potential in adulthood. Studies with the antiandrogen flutamide show that at doses significantly above that required to completely block prostate differentiation and cause genital feminization, Wolffian ductal differentiation is significantly impaired. Thus, higher doses of flutamide are needed to block the paracrine effect of T on the Wolffian ducts.

The development of a male pseudohermaphroditic rat using an inhibitor of the enzyme 5 alpha-reductase.

Incomplete masculinization of the external genitalia occurred in male Sprague-Dawley rats treated with a potent inhibitor of enzyme 5 alpha-reductase at the critical period of sexual differentiation in utero. The studies were performed using the 5 alpha-reductase inhibitor, 4-methyl-4-aza-5-pregnan-3-one-20[s] carboxylate, one of a series of aza steroids known to competitively inhibit the enzyme 5 alpha-reductase. The degree of inhibition of male external genital development was dependent upon the dose of the inhibitor, and at a dose of 36 mg/kg X day, there was complete feminization of the external genitalia of the male animal with a urogenital sinus and a pseudovagina. These studies provide conclusive evidence for the hypothesis that 5 alpha-reductase activity and dihydrotestosterone (17 beta-hydroxy-5 alpha-androstan-3-one) formation are essential for normal differentiation of male external genitalia. Epididymidis, vasa deferentia, and seminal vesicles were present at all doses of the inhibitor, suggesting testosterone dependency. However, confirmation of the testosterone dependency of Wolffian ductal differentiation awaits further studies, particularly comparison studies with the rabbit and dog, since Wolffian ductal differentiation in the rat, unlike the rabbit and dog, is not abolished with the antiandrogen, cyproterone acetate. The presence of prostatic buds, despite complete external genital feminization, was unexpected and suggests that these structures may have different thresholds of response for dihydrotestosterone. Prostatic differentiation may have a much lower threshold, requiring less dihydrotestosterone for differentiation.