In vitro release of aldosterone and cortisol in human adrenal adenomas correlates to mRNA expression of steroidogenic enzymes for genes CYP11B2 and CYP17.

Adenomas of the adrenal cortex cause different disorders depending on the main steroid synthesized and released. The aim of this research is to increase our understanding of the pathophysiology of steroidogenesis in adrenocortical disorders by comparing the release of steroids from adrenocortical adenomas in vitro with the messenger RNA (mRNA) expression of steroid synthesizing enzymes. Fourteen patients with adrenal tumors were included in the present study; nine were diagnosed with primary aldosteronism and three with Cushing's syndrome. Two patients had an adrenal tumor discovered on computed tomography (CT) during workup for an unrelated disease. Serum cortisol, plasma aldosterone, and urinary catecholamines were normal. Tissue was taken for in vitro steroid release, and aldosterone and cortisol in the medium after a 1-hour incubation were determined. Oligonucleotide probes with sequences complementary to mRNAs encoding for the steroid synthesizing enzymes 11 beta-hydroxylase (CYP11B1), 18-hydroxylase (CYP11B2), 17 alpha-hydroxylase (CYP17), and 21-hydroxylase (CYP21) were synthesized (Genset, Paris, France) and in situ hybridization was performed. Moderate expression of CYP11B2 and low expression of CYP11B1 were seen in the zona glomerulosa. The zona fasciculata of the control adrenals expressed a high signal of CYP11B1, whereas the expression of CYP11B2 was very low. There was considerable variation in aldosterone release from the aldosteronomas, whereas the tumors from the Cushing patients showed no detectable release of aldosterone. In contrast, tumors from patients with primary aldosteronism, Cushing's syndrome, and no hyperfunction all had the ability to synthesize and release cortisol in vitro. The highest cortisol release was found in tumors from patients with Cushing's syndrome, but also the nonhyperfunctioning tumors and some of the aldosteronomas released significant amounts of cortisol. The two patients with highest release of aldosterone in vitro showed the highest expression of CYP11B2 and the lowest expression of CYP11B1 and CYP17. The remaining aldosteronomas had low expression of CYP11B2, similar to the two other groups. Expression of CYP11B1 was high as expected in the Cushing adenomas, but also the two nonhyperfunctioning tumors and some of the aldosteronomas showed a moderate expression. Adenomas from Cushing's syndrome, nonhyperfunctioning adenomas, and some of the aldosterone-producing adenomas had moderate to high expression of CYP17. This paper presents new means for functional characterization of adrenocortical tumors. Diagnosis of an aldosteronoma is often difficult, and with the advent of these methods it is possible to determine the functional capacity of a tumor, once it is removed. This is of special interest if the patient remains hypertensive postoperatively, and it is not clear whether the patient indeed had a functioning tumor.

Streptozocin and o,p'DDD in the treatment of adrenocortical cancer patients: long-term survival in its adjuvant use.

BACKGROUND: To evaluate the efficacy of streptozocin and o.p'DDD (SO) in adrenocortical cancer (ACC) patients since other chemotherapeutic regimens have limited effects. PATIENTS AND METHODS: We performed a phase II study with SO therapy in 40 ACC patients (median age 44 years). Oral o,p'DDD administration (1-4 g/d, every day) was given together with intravenous streptozocin (1 g/d for five days, thereafter 2 g once every three weeks). 5HT3-receptor blocker was used as standard premedication for streptozocin. RESULTS: The SO therapy was found to have significant effects on disease-free interval (P = 0.02) as well as on survival (P = 0.01) in adjuvantly treated cases (n = 17) in comparison to the patients who did not get any therapy after complete resection (n = 11). Complete or partial response was obtained in 36.4% of patients with measurable disease (n = 22). The overall two-year and five-year survival rates were 70% and 32.5%, respectively. The presence of metastases at diagnosis was identified as a poor prognostic factor (P = 0.02). CONCLUSIONS: The present study necessitates further randomized clinical study of SO therapy in the treatment of ACC, mainly as adjuvant treatment immediately after curative intended surgery, and could be developed into a regular treatment regimen.

[Acute abdomen calls for considerable care resources. Analysis of 3727 in-patients in the county of Stockholm during the first quarter of 1995]. / Akut buksjukdom kräver stora vårdresurser. Analys av 3,727 inlagda patienter första kvartalet 1995 i Stockholms län.

A total of 3,727 in-patients with acute abdominal symptoms were identified during the first quarter of 1995 at the surgical clinics of the nine hospitals with emergency departments in the county of Stockholm. The diagnoses were: non-specific abdominal pain 24%; cholecystitis 9%; appendicitis 8%; bowel obstruction 7%; intra-abdominal malignancy, diseases of the urinary tract and peptic ulcer 6% each; gastrointestinal hemorrhage, diverticulitis of the colon and pancreatitis 5% each; other diseases as a cause of abdominal symptoms, 19%. 1,601 operations were performed of which 47% were endoscopic procedures. The mean duration of hospital stay was 4.8 days. The length of stay increased significantly with age. The age-related relative frequency of hospitalization due to acute abdominal pain was also dramatically higher in the elderly cohorts. These facts and the prognosis of an 18% increase of inhabitants 50 years of age or older until 2010 in Greater Stockholm signal an increased need of hospital resources for this large group of patients in the coming years.

Value of diagnostic laparoscopy in fertile women with suspected appendicitis.

OBJECTIVE: To assess the value of diagnostic laparoscopy in women of child-bearing age with suspected appendicitis. DESIGN: Prospective, non-randomised study. SETTING: University hospital, Sweden. SUBJECTS: 94 women aged between 15 and 40 years with suspected appendicitis. INTERVENTION: Diagnostic laparoscopy. If necessary, the appendix was removed either laparoscopically or by open operation. The outcomes were compared with those during 1994, during which all appendicectomies were done through a laparotomy. MAIN OUTCOME MEASURES: Rates of unnecessary appendicectomy, operating time, and length of hospital stay. RESULTS: 77 had diagnostic laparoscopy, and 17 primary open appendicectomy. 73 appendixes were removed; 53 open and 20 laparoscopically. 12 appendicectomies (13%) were unnecessary compared with 27/80 (34%) during the year 1994. The duration of the open operation was significantly shorter (mean (SD) 35 (14) minutes) than the laparoscopic one (78 (18) minutes) (p<0.001), and there were no differences in duration of postoperative hospital stay. No patients developed complications. CONCLUSIONS: Diagnostic laparoscopy is safe and helpful in suspected cases of appendicitis and could substantially reduce the rate of unnecessary appendicectomies. However, the data suggest that laparoscopic appendicectomy offers no advantages over primary open appendicectomy in terms of postoperative course.

Long term experience after subtotal adrenalectomy for multiple endocrine neoplasia type IIa.

OBJECTIVE: To evaluate the long-term results after subtotal adrenalectomy in patients with multiple endocrine neoplasia type IIa (MEN IIa). DESIGN: Retrospective study. SETTING: University Hospital, Sweden. SUBJECTS: Five patients who underwent partial adrenalectomy between 1985 and 1989. INTERVENTIONS: Subtotal adrenalectomy with a rim of cortical tissue left in situ. MAIN OUTCOME MEASURES: Follow up by interview, measurement of cortisol and catecholamine excretion in urine, and cortisol concentration in serum in response to stimulation with ACTH. RESULTS: Three patients took no corticosteroids regularly, but during upper respiratory tract infections, or periods of severe stress they took 25 mg cortisone acetate daily. This is confirmed by their normal values of 24 hour urinary cortisol excretion and subnormal responses to an ACTH-stimulation test. The fourth and fifth patients had low concentrations of endogenous corticosteroids postoperatively, which is being replaced with 25 mg cortisone acetate daily. Postoperatively all five patients had low urinary adrenaline excretion. CONCLUSION: Subtotal adrenalectomy in patients with MEN IIa resulted in basal endogenous corticosteroids within the reference range in three of five patients. There was no evidence of reduced adrenocortical function with time, nor were there any signs of recurrence of the pheochromocytoma.

Delayed shoulder exercises in reducing seroma frequency after modified radical mastectomy: a prospective randomized study.

BACKGROUND: Seromas and impaired shoulder function are well-known complications after modified radical mastectomy for breast cancer. Early postoperative physiotherapy is a common treatment to avoid shoulder dysfunction. The aim of this study was to evaluate if the frequency of postoperative seromas could be reduced, without increasing shoulder dysfunction, by delayed postoperative shoulder exercises. METHODS: In a prospective study 163 patients with breast cancer undergoing modified radical mastectomy were randomized to physiotherapy starting on postoperative day 1 or day 7. Patients were seen by the surgeons and the physiotherapists during hospital stay and in the outpatient department. Seromas and other complications were registered by the surgeons. The physiotherapists instructed the patients pre- and postoperatively and assessed shoulder function. RESULTS: There was a significantly higher incidence of postoperative seromas in the group of patients that started physiotherapy postoperative day 1 (38%) compared to the group that started physiotherapy postoperative day 7 (22%) (p < 0.05). There was no significant difference between the groups in the late outcome of shoulder function. CONCLUSION: The incidence of seromas after modified radical mastectomy for breast cancer is reduced by delaying shoulder exercises one week postoperatively. Earlier postoperative physiotherapy is not necessary to avoid impaired shoulder function.

Ultrastructural localization of endothelin-1 in nonneoplastic, hyperplastic, and neoplastic adrenal gland.

Endothelin (ET)-1 is a 21-amino acid peptide with potent vasopressor and vasoconstrictive properties. Biochemical and recent histochemical studies have shown that this peptide is present in human adrenal cortex. This study was intended to determine ET-1 immunoreactivity in human adrenal cortex and cortical adenoma, and hyperplasia ultrastructurally. Light microscopical examination confirmed recent findings of ET-1 immunoreactivity in the three cortical zones (but not in the medulla) as well as in cortical adenoma and cortical hyperplasia. The immunoreactivity in the cortex and adenoma appeared in the cytoplasm in the form of vacuolar structures and grains. Focally, the cell membranes also showed immunoreactive staining. Electronmicroscopical investigation revealed ET-1 immunoreactive products adjacent to the outer surface of the membrane of lipid bodies, in mitochondria and rough endoplasmic reticulum and focally on the cell membrane, but no immunolabeling was seen in the medulla. The localization of ET-1 in the endoplasmic reticulum indicates that this peptide is synthesized in the cortical cells. Its localization in the membrane of the lipid bodies and in the mitochondria suggests that it takes part in synthesis and/or secretion of steroid hormones. The focally immunolabeled cell membranes may be dependent on ET-1 binding to ET receptors.

Immunohistochemical localization of endothelin-1 in non-neoplastic and neoplastic adrenal gland tissue.

Endothelin (ET)-1 is a 21-amino acid peptide with potent vasopressor and vasocontrictive properties. Biochemical studies suggest that this peptide occurs in adrenal glands, where it influences steroid hormone production. However, we have found no report of the topographical distribution of this peptide. The localization of ET-1 immunoreactivity in non-neoplastic (37 cases) and neoplastic adrenal glands (48 cases) was investigated with a sensitive immunohistochemical technique applied to routinely processed tissue specimens. ET-1 immunoreactivity was regularly seen in the cortex, especially in the zona fasciculata and to a varying extent also in the other two zones, but not in the medulla. The immunoreactive material appeared in the cytoplasm mostly in the form of vacuolar structures but also as grains. Focally, the cell membrane also showed immunoreactive staining. In the zona reticularis the immunoreactivity appeared mainly as cytoplasmic grains. Most cortical adenomas displayed numerous immunoreactive cells. The immunoreactivity in the tumour tissue appeared in the same forms as in normal cortex, but the reactive products were generally fewer in number. No obvious differences in immunostaining were seen between the aldosterone- and cortisol-producing adenomas or the non-functioning ones. Three of the ten carcinomas contained immunoreactive cells, but they were few, appearing focally and the ET-1 immunoreactive structures were seen as 'dust-like' material. The difference in immunoreactivity between the benign and the malignant cortical neoplasms may be of diagnostic value. Functionally our results support a relationship between ET-1 and steroid regulation in non-neoplastic cortical tissue.

Levels of cytochrome P-450, steroidogenesis and microsomal and cytosolic epoxide hydrolases in normal human adrenal tissue and corresponding tumors.

The levels of microsomal cytochrome P-450, steroidogenesis and microsomal and cytosolic epoxide hydrolase activities in normal human adrenal tissue (obtained from adult kidney transplant donors and autopsy material) and corresponding hyperplasia, adenomas and carcinomas (surgical biopsies) were determined. The increased steroid production demonstrated by most of the pathological tissue samples examined here was associated with either an unchanged or dramatically decreased specific microsomal content of cytochrome P-450. Furthermore, specific microsomal epoxide hydrolase activity was also found to be reduced in adrenocortical carcinomas, while the corresponding cytosolic activity was also decreased in at least two of these carcinomas. It is of interest to note in this connection that the level of microsomal epoxide hydrolase in slightly atropic adrenal cortex surrounding adrenocortical carcinomas was also found to be reduced. This would indicate that despite its appearance, this surrounding tissue is not normal in all respects. Thus, adrenocortical carcinomas fit into the common pattern in that their specific contents of microsomal cytochrome P-450 are dramatically decreased, but the simultaneous decrease in their microsomal epoxide hydrolase activity is more unusual.

Identification of hydrophobic fragments of alpha 1-antitrypsin and C1 protease inhibitor in human bile, plasma and spleen.

Hydrophobic peptides were isolated from the phospholipid fraction of human bile, plasma and spleen by exclusion chromatography in organic solvents. From plasma, the activation peptide of C1 protease inhibitor was recovered, from spleen the activation peptide of alpha 1-antitrypsin, and from bile, both these peptides, as well as a fragment generated by proteolytic cleavage of alpha 1-antitrypsin six residues N-terminal of the P1-P1' peptide bond. Cleavages in this region inactivate antiproteases but have previously not been reported to occur in vivo. These peptides in human bile may reflect physiological actions in regulation of antiproteolytic activity or bile secretion processes, and/or be of importance for the physicochemical state of cholesterol, phospholipids and bile acids in bile.

Plasma chromogranin A + B, neuropeptide Y and catecholamines in pheochromocytoma patients.

Plasma levels of chromogranin A + B, neuropeptide Y and catecholamines were analysed before, during and after surgery in seven patients with pheochromocytoma. The aim of the study was to determine the diagnostic sensitivity of these plasma amines and peptides, and to investigate their peroperative fluctuations. Chromogranin A + B in plasma was increased preoperatively in all patients, showed no significant increase during surgery, and normalized postoperatively. Neuropeptide Y, which alone can induce hypertension, was present in high levels in plasma from three patients preoperatively, increased further in four patients during surgery, and was postoperatively low in all patients. Fractionated plasma catecholamines were increased in five patients before surgery, increased in all patients during tumour dissection, and normalized postoperatively. It may be concluded that plasma chromogranin A + B exhibited as high a sensitivity for pheochromocytoma as fractionated urinary catecholamines in the patients studied.

Steroid profile in serum: increased levels of sulphated pregnenolone and pregn-5-ene-3 beta,20 alpha-diol in patients with adrenocortical carcinoma.

A serum steroid profile was determined in 11 patients with adrenocortical carcinoma, 5 with adrenocortical adenoma and 10 healthy controls. Seven of the patients with carcinoma had different forms of endocrine symptoms, and of those with adenoma 3 had Cushing's syndrome and 2 primary hyperaldosteronism. Sulphated steroids dominated in serum from both patients and controls, whereas the levels of free and glucuronated steroids were low. All patients with adrenocortical carcinoma had increased levels of sulphated pregn-5-ene-3 beta-ol-20-one (pregnenolone) and pregn-5-ene-3 beta,20 alpha-diol compared with healthy controls and patients with adrenocortical adenoma. Serum levels of 11-deoxycortisol and/or its glucuronated metabolite tetrahydro-11-deoxycortisol were clearly elevated in 8 of the patients with carcinoma. The results are in agreement with those previously found for conjugated urinary steroids in patients with adrenocortical carcinoma. Thus, an impaired function or deficiency of 3 beta-hydroxysteroid dehydrogenase/delta isomerase and in some cases also of 11 beta-hydroxylase could explain the findings. The serum levels of sulphated pregnenolone and/or pregn-5-ene-3 beta,20 alpha-diol, possibly together with free 11-deoxycortisol seem to be useful for preoperative discrimination between malignant and benign adrenocortical tumours.

Basal and ACTH-stimulated cortisol and aldosterone release from adrenocortical adenomas in vitro.

Basal and ACTH-stimulated release of aldosterone and cortisol was studied in slices of adrenocortical adenomas and adrenal cortex from 16 consecutive patients with Conn's syndrome (n = 7), Cushing's syndrome (n = 5) or no signs of steroid hypersecretion (n = 4). Clinical data were reviewed and histologic examination was repeated. The adrenal cortex and all tumours but one (non-hyperfunctioning) secreted cortisol. Cortisol release increased after addition of ACTH in 13/15 tumours and all cortex slices. Aldosterone was secreted from the adrenal cortex in all patients and was increased by ACTH in 75% of the tissue specimens. Adenomas from patients with hyperaldosteronism showed the highest basal aldosterone secretion. This secretion was further increased by ACTH. Tumours from patients without signs of corticosteroid excess also released steroids and responded to ACTH. It is concluded that in vitro studies of steroids released from adrenocortical tumours contribute to their diagnostic and functional evaluation.

Adrenocortical carcinoma. A retrospective study of a rare tumor with a poor prognosis.

A retrospective study was performed on 54 patients diagnosed as having adrenocortical carcinoma during 1974-1983. The initial symptoms were often diffuse: abdominal pain, weight loss, or fever, and more than 60% of the patients showed no evidence of overproduction of hormone. The median tumor diameter was 13 cm and almost half of the tumors had metastasized at diagnosis. A radical tumor resection could be performed in less than 50% of the patients, and at histopathological re-examination some tumors were not conclusively verified as malignant. Capsular invasion, nuclear pleomorphism and mitoses were found more commonly in patients who succumbed to the disease. Seven of 29 patients treated with chemotherapy showed an objective response and two of them are still alive and free of disease. The overall 5-year-survival rate was 19%, compared with 45% for patients with radically resected tumors. Patients with no biochemical signs of overproduction of adrenocortical hormone appeared to have a better prognosis than those with hormone excess. Together with increased use of ultrasound and computed tomography, a urinary steroid profile might hopefully contribute to earlier discovery of these often clinically silent tumors. However, it remains to be determined whether these diagnostic improvements, together with more aggressive surgery and adrenolytic chemotherapy, can improve the poor prognosis.

Steroid profile in urine: a useful tool in the diagnosis and follow up of adrenocortical carcinoma.

The urinary steroid profile was determined in 24 patients with adrenocortical carcinoma. Seventeen of the patients had Cushing's syndrome, virilization or feminization, and 7 had no signs of endocrine disease. Seven of the 11 patients still alive are free of disease, after a follow-up period of 5-75 months. The steroid profile varied widely between the patients with adrenocortical carcinoma. Patients with Cushing's syndrome had increased levels of cortisol metabolites and those with virilism had raised excretion of androgen metabolites. Six of the patients with adrenocortical carcinoma showed normal values of these metabolites. In 23 of the 24 patients the excretion of 3 beta-hydroxy-5-ene steroids and/or metabolites of cortisol precursors, such as tetrahydro-11-deoxycortisol, were significantly increased, compared with healthy controls or patients with adrenal adenomas. These findings suggest a relative deficit or low activity of 3 beta-hydroxysteroid dehydrogenase/delta isomerase and/or 11 beta-hydroxylase in tumour tissue. In the single patient where the steroid profile failed to indicate malignancy, hypercortisolism was seen and the tumour mass was small. The steroid excretion normalized after radical surgery and decreased in patients responding to chemotherapy. During recurred disease the metabolites of 3 beta-hydroxy-5-ene steroids and/or cortisol precursors increased, but in some patients the excretory pattern then was different from that seen before treatment.

Synthesis of alpha- and beta-calcitonin gene-related peptide by a human pheochromocytoma.

One histologically verified human pheochromocytoma, out of a total of nine such tumors examined, contained calcitonin gene-related peptide (CGRP)-like immunoreactivity (151 pmol/g wet weight) that was localized to a scattered population of tumor cells. The immunoreactivity was resolved by high performance ion-exchange chromatography into two components with the retention times of human alpha-CGRP and beta-CGRP in the approximate ratio 1:2. No pheochromocytoma in this series contained calcitonin-like immunoreactivity. In contrast, two neuroendocrine tumors of the pancreas contained calcitonin-like immunoreactivity (190 pmol/g and 125 pmol/g) but no CGRP-like immunoreactivity was detected. These results indicate a degree of tissue selectivity in the processing of the primary transcripts of the calcitonin/CGRP genes in human tumors.