The development of adrenocorticotrophin-sensitive adenylate cyclase activity in the foetal rabbit adrenal: a correlated biochemical and morphological study.

Proliferation of the smooth endoplasmic reticulum, the site of some hydroxylating steroidogenic enzymes in foetal adrenocortical cells, is the first major change in the process of their differentiation into steroidogenic tissue. This was observed in our ultrastructural studies on foetal rabbit adrenals to begin at about day 19 of development. Morphological changes in the mitochondria, the site of production of other steroidogenic enzymes, occurred at about day 24. The elongated or rod-shaped forms of the earlier stages became flattened and rounded by this time, while the cristae were transformed from a flattened lamellar type of the earlier stages to the tubulo-vesicular form of the adult. Other changes observed included an increase in microvilli and in cell size, with a concomitant increase in thickness of the gland. Adenylate cyclase activity in foetal adrenal homogenates was assessed in response to sodium fluoride (NaF) and ACTH. All preparations responded to NaF. While good responses to ACTH were observed at days 24, 27, 28 and in the neonate, there was a lack of any significant response in the day 19 gland. Foetal ACTH was depressed by administration of cortisol, and the effects of this treatment on both the morphological changes and adenylate cyclase activity was reassessed. The response of foetal adrenals to ACTH was depressed by this treatment and differentiation of the mitochondria was arrested. These results suggest a circumscribed period for the development of ACTH-sensitive adenylate cyclase coinciding with the time at which final differentiation of the mitochondria is completed. Furthermore, both the differentiation of the mitochondria and the development of ACTH-sensitive adenylate cyclase in the foetal adrenal may be dependent on foetal ACTH secretion.

Assessment of new radioimmunoassay kit for determining urinary albumin at low concentrations: comparison with radial immunodiffusion.

The assay characteristics of a new radioimmunoassay kit for determining urinary albumin at low concentrations were studied. The sensitivity for urinary albumin was 2 mg/l, the analytical range 2 to 40 mg/l, and interassay coefficient of variation less than 12%. In a method comparison study entailing diabetic urine samples covering an albumin concentration of 2 to 150 mg/l the kit compared adequately with radial immunodiffusion (mean difference between methods = 2 mg/l; residual standard deviation = 4.6 mg/l), absolute variation between methods increasing with the concentration. The kit required much less skill than radial immunodiffusion but its capital and running cost were higher.

Modifying effects of amlodipine on cyclosporin A-induced changes in renal function in patients with psoriasis.

OBJECTIVE: To assess the benefit of amlodipine, a calcium channel blocker, on renal function and vasoactive hormones in individuals with normal kidneys and blood pressure and who were being treated with cyclosporin A for refractory psoriasis. DESIGN: An open-label, two-stage, longitudinal study was employed. METHODS: Patients were divided into two groups: Group I received cyclosporin A, 5 mg/kg per day for 6 months titrated down to 2.5-3.5 mg/kg per day for 6 months, then concomitant amlodipine 5 mg/day for 6 months; and Group II received concomitant cyclosporin A, 5 mg/kg per day, and amlodipine, 5 mg/day, for 6 months. Blood pressure, serum creatinine, glomerular filtration rate, urinary magnesium, plasma renin activity and urinary kallikrein excretion were measured before and after treatment. RESULTS: Eighteen patients were enrolled, and 12 completed the study. In Group I (n = 7), 12 months of cyclosporin A therapy significantly increased systolic blood pressure and significantly decreased glomerular filtration rate, plasma renin activity and active urinary kallikrein. Amlodipine reversed these changes. In Group II (n = 5), 6 months of concomitant cyclosporin A and amlodipine significantly reduced active urinary kallikrein levels. No significant changes occurred in the other measured parameters in either group. CONCLUSIONS: Cyclosporin A produces a sustained and significant fall in glomerular filtration rate and urinary kallikrein excretion, even in patients with normal kidneys and blood pressure. Amlodipine is potentially capable of reversing these nephrotoxic effects.

Final results from the Betaseron (interferon ß-1b) Pregnancy Registry: a prospective observational study of birth defects and pregnancy-related adverse events.

OBJECTIVE: Women with multiple sclerosis are often diagnosed and treated during their reproductive years. Limited data are available on the safety of treatment during pregnancy. The Betaseron Pregnancy Registry prospectively monitored women exposed to interferon ß-1b (IFNß-1b) during pregnancy to estimate the rates of birth defects, spontaneous abortions (SABs) and other negative outcomes in this population. DESIGN: From 2006 to 2011, this observational registry enrolled women exposed prior to conception or during pregnancy (but prior to or without abnormalities on prenatal screening). Follow-up continued from enrolment through the 4-month paediatric visit. SETTING: Patients in the USA who met these criteria were enrolled in the registry. RESULTS: The registry enrolled 99 pregnant women; 3 were lost to follow-up. The earliest exposure to IFNß-1b occurred during the first trimester for 95 pregnancies and in the third trimester for 1 pregnancy. There were 99 birth outcomes (3 twins), including 86 (86.9%) live births, 11 (11.1%) SABs and 2 (2%) stillbirths. Birth defects were reported in five (5.1%) cases. Rates of birth defects and SAB were not significantly different from population comparators. No developmental concerns were identified at the 4-month paediatric visit. CONCLUSIONS: The small sample size limits the ability to draw definitive conclusions; however, there was no pattern to suggest increased negative outcomes with IFNß-1b. CLINICAL TRIALS REGISTRATION NUMBER: NCT00317564.

A simplified method for monitoring blood concentration of cyclosporin by a finger-stab technique.

We have developed a simple method of sample collection which allows the measurement of cyclosporin (CsA) to be made from blood obtained from a finger stab and collected onto filter paper. CsA concentrations estimated from blood spots and venous samples obtained at the same time showed good correlation. This method has various practical applications: peak and trough blood CsA concentrations (as well as profiles) may be routinely measured from samples collected by patients at home; it may be of particular value in children and other patients with poor venous access.

Kallikrein and kinin release using superfused disaggregated cortical cells from rat and human kidney: effects of AVP and dopamine.

Potential regulators of the renal Kallikrein-Kinin System are poorly defined. We have therefore examined the effect of arginine vasopressin and dopamine on the release of kallikrein and kinin from collagenase-dispersed rat and human renal cortical cells.

Kallikrein release from the kidney: in vitro effects of AVP and DDAVP in three species.

Kallikrein released during superfusion of rat, monkey and human kidney cortical slices was mainly in the inactive form. Arginine vasopressin and the nonpressor synthetic analogue des-amino-D-arginine vasopressin increased the release of inactive kallikrein to a similar extent in all species. No detectable change in active kallikrein release occurred. These results suggest a role for the kallikrein-kinin system in the regulation of the hydro-osmotic effect of arginine vasopressin.

Pharmacokinetics of cyclosporin in man following a single oral dose: relationship to body fat content.

We have investigated the pharmacokinetics of the immunosuppressive drug cyclosporin in 11 patients receiving regular dialysis therapy. After a single oral dose of cyclosporin, whole blood concentrations were measured at regular intervals for 24 h. Body fat content was estimated in each subject from the measurement of skin-fold thickness and mid-arm circumference. In ten of the 11 patients there was a relatively uniform response in the increase of cyclosporin blood concentration to a peak within the first 6 h. Calculated area under the blood concentration curve did not correlate with total bodyweight, but correlated strongly with body fat content as well as fat-free mass. This study suggests that more predictable blood concentrations of cyclosporin might be achieved if the initial dose were calculated on the basis of fat-free mass rather than total bodyweight.

Gender differences in urinary kallikrein excretion in man: variation throughout the menstrual cycle.

1. Urinary kallikrein excretion was measured in healthy male subjects and in healthy pre- and post-menopausal females. 2. Urinary kallikrein excretion was shown to be constant throughout a 24 h period. Individual male subjects showed little fluctuation in urinary kallikrein excretion; within-subject variance accounted for 1.65% of the total. 3. Female subjects with ovulatory menstrual periods excreted significantly more kallikrein than post-menopausal females and males. 4. Pre-menopausal females showed a much greater within-subject variation in urinary kallikrein excretion and this could be related to the stage of the menstrual cycle, with significantly greater urinary kallikrein excretion in the luteal phase than in the follicular phase. 5. Plasma renin activity and plasma aldosterone concentration also showed a menstrual variation, with concentrations in the luteal phase being significantly higher than those in the follicular phase. 6. The rise in urinary kallikrein excretion in the luteal phase could be abolished by oral administration of the aldosterone antagonist spironolactone. 7. Urinary kallikrein excretion in post-menopausal females was similar to the range found in males, and showed no cyclic changes over a 4 week period. 8. Gender and menstrual status should be taken into account in studies of the physiological role of tissue kallikreins.

Bartter's syndrome and diabetes mellitus.

We here report a case of Bartter's syndrome occurring in association with diabetes mellitus. The patient, an insulin-dependent diabetic, presented with hypokalaemia, inappropriate kaliuresis and metabolic alkalosis. He had high plasma renin activity, relatively low plasma aldosterone, and resistance to infused angiotensin II. A high potassium diet raised total body potassium and serum potassium, did not affect plasma renin activity, but raised plasma aldosterone significantly and did not alter the resistance to angiotensin II. Indomethacin administered acutely reduced urinary potassium and kallikrein excretion and, on chronic administration, lowered plasma renin activity, urinary chloride excretion, and raised serum potassium. Salt restriction resulted in a prompt and significant reduction in urinary sodium and chloride excretion. Urinary kallikrein excretion was very high throughout, increased with sodium restriction, and decreased with sodium loading. Oral potassium supplementation partially corrected the hypokalaemia, but did not affect blood sugar control. In this patient the primary defect appears to have been primary urinary potassium wasting, rather than sodium or chloride wasting. The striking effects of indomethacin suggest that prostaglandins may play a fundamental role in the genesis of the syndrome.

Cyclic AMP responses to parathyroid hormone and glucagon during lithium treatment.

Inhibition of adenylate cyclase has been proposed as a mechanism for hypothyroidism and nephrogenic diabetes insipidus occurring during lithium treatment, but these disorders are rarely found in the same patients. We have measured plasma levels of adenosine 3':5'-cyclic monophosphate (cyclic AMP) after an intravenous injection of glucagon in eight patients receiving long term lithium treatment and in six control subjects. Urinary cyclic AMP levels after an intravenous injection of bovine parathyroid hormone (PTH) were also measured in the patients. The plasma cyclic AMP response to glucagon in the patient group was significantly lower than that of the controls. No correlation was demonstrated between the plasma cyclic AMP response after glucagon and the urinary cyclic AMP response after PTH. We have previously shown that impairment of the response to PTH correlates with reduced urine concentrating ability during lithium treatment. In contrast, there was no correlation between the responses to PTH and glucagon in individual patients. These results are consistent with the hypothesis that inhibition of adenylate cyclase is an important factor in lithium-induced endocrine dysfunction.

Impairment of cyclic AMP response to bovine parathyroid hormone in patients on chronic lithium therapy with diminished renal urine-concentrating ability.

1. Urinary and plasma levels of adenosine 3':5'-cyclic monophosphate (cyclic AMP) after an intravenous injection of bovine parathyroid hormone (PTH) were measured in 12 patients on long-term lithium treatment and in nine control subjects. The maximum urine osmolality (Umax.) after an intravenous injection of desamino-D-arginine vasopressin (DDAVP) was also measured. 2. In all the control subjects and six of the patients, the Umax. after DDAVP exceeded 700 mosmol/kg. The cyclic AMP responses in these two groups did not differ significantly. 3. In the remaining six patients whose Umax. did not reach 700 mosmol/kg, the cyclic AMP response to PTH was significantly less than that of the controls. 4. A strong correlation was demonstrated in the patients between the urinary cyclic AMP response after PTH and the maximum osmolality after the administration of DDAVP. 5. These observations are consistent with the hypothesis that reduced adenylate cyclase activity contributes to the development of nephrogenic diabetes insipidus in patients on long-term lithium treatment.

The effects of potassium, 5-hydrocytryptamine, adrenocorticotrophin and angiotensin II on the concentration of adenosine 3':5'-cyclic monophosphate in suspensions of dispersed rat adrenal zona glomerulosa and zona fasciculata cells.

Dispersed rat adrenal cells prepared from both the capsule and the decapsulated gland were used to investigate the effects on cyclic AMP accumulation of known stimuli of steroidogenesis [ACTH (adrenocorticotrophin), angiotensin II, K(+) ions and 5-hydroxytryptamine]. Since glomerulosa-cell preparations from capsular strippings are normally contaminated with a proportion of fasciculata cells, cells purified by fractionation on a bovine serum albumin gradient were also used. The results showed that: (1) ACTH and angiotensin II stimulated cyclic AMP accumulation in both fractionated and unfractionated zona fasciculata cells; (2) 5-hydroxytryptamine and an increased extracellular K(+) concentration (from 3.6 to 8.4mm) had no effect on cyclic AMP concentrations in fasciculata cell preparations; (3) the addition of ACTH, angiotensin II, 5-hydroxytryptamine or K(+) to the incubation medium resulted in increased cyclic AMP concentrations in unpurified zona glomerulosa cell preparations; (4) fractionation and hence the virtual elimination of fasciculata contamination, did not affect the response to 5-hydroxytryptamine and increased K(+) concentration. However, the responses to ACTH and angiotensin II were markedly lowered but not abolished. These results strongly suggest a link between cyclic AMP production and steroidogenesis in the zone of the adrenal gland that specifically secretes aldosterone. All four agents used stimulated both steroid output and cyclic AMP accumulation. However, at certain doses of 5-hydroxytryptamine, K(+) and angiotensin II the significant increases in corticosterone output were not accompanied by measurable increases in cyclic AMP accumulation.

Biological assay for tissue kallikrein: comparison with the synthetic substrate S2266.

A highly sensitive biological assay for tissue kallikrein is described, using human kininogen as substrate; and quantitation, by radioimmunoassay, of generated kinins. Using purified human urinary kallikrein as a reference standard we have correlated the kininogenase activity of kallikrein with amidase activity as measured by cleavage of the synthetic substrate S2266.