Regulation of pancreatic islet cell replication: an inquiry into the controversy regarding the effects of steroid hormones.

The controversial issue of the effects of prednisolone and 17 beta-estradiol on replication of fetal rat pancreatic islets in culture was studied using 32P and [3H]thymidine as probes for studying DNA synthesis. DNA synthesis was not affected by the steroid hormones, as was evident from the rate of incorporation of 32P into total DNA. Decreased incorporation of [3H]thymidine into DNA found in islets treated with either of these steroids seemed to reflect an inhibitory effect of these hormones on thymidine kinase, leading to decreased phosphorylation of labeled thymidine. In addition, the hormones stimulated the activity of thymidylate synthetase, thus enhancing the endogenous synthesis of thymidine and thereby diluting the specific activity of the [3H]thymidine added to the cultured islets. Further support for a lack of inhibition of growth of islet cells treated with steroid hormones was provided by the observation that prednisolone increased uridine kinase activity and RNA biosynthesis, both of which may participate in the growth of cells preceding mitosis and (the latter) in protein hormone biosynthesis.

Modulation of rat pancreatic islet cell replication and insulin release by glibenclamide.

Glibenclamide significantly stimulated the incorporation of [3H]thymidine into DNA of fetal rat pancreatic islets at a physiological (5.5 mM) but not at a high (22 mM) glucose concentration. There was no significant stimulation of insulin release under these conditions. In contrast, the drug-stimulated insulin release from adult islets cultured at 5.5 mM glucose but had no effect on their DNA synthesis. The observations suggest that insulin secretion and DNA synthesis may be dissociated in rat pancreatic islets.

Islet cell growth and function. A reappraisal of the role of progesterone and prednisolone.

The mechanism of the inhibitory effect of steroid hormones, progesterone and prednisolone on the incorporation of [3H]-thymidine into pancreatic islet cell DNA was investigated. Treatment with either hormone had no effect on the incorporation of 32P-orthophosphate into islet cell DNA. Both prednisolone (10 microM) and progesterone (3 microM) markedly stimulated the activity of the enzyme thymidylate synthetase of islet cells possibly leading to increased synthesis of endogenous thymidine which resulted in dilution of the [3H]-thymidine added to the islets in tissue culture. Prednisolone (10 microM) significantly increased both insulin biosynthesis and release, while at 5 microM it was effective in increasing only insulin release. In contrast, progesterone at the two concentrations employed did not affect insulin biosynthesis or release. The smaller doses of both hormones markedly stimulated the total protein biosynthesis.

Effect of LDL and serum from diabetic subjects on DNA synthesis in HIT-cells in culture.

The effect of lipoprotein-deficient serum (LPDS) with and without added low-density lipoprotein (LDL), isolated from diabetic subjects, on the replication of SV40-transformed islet cells (HIT cells) was investigated. Whole serum as well as LPDS preparations stimulated DNA synthesis maximally when added to the culture medium at a final concentration of 0.1%. The addition of LDL at 25 and 175 micrograms protein/ml medium did not cause further stimulation. On the contrary, the higher concentrations resulted in a significant inhibition. These results suggest that previously observed stimulation of DNA synthesis in smooth muscle cells by LDL from diabetic subjects is most likely due to the presence of growth factors in the serum of these patients and not to LDL per se.

Validation of the use of 32P-orthophosphate as a probe for monitoring DNA replication in pancreatic islet cells.

This study aims at validating the use of 32P-orthophosphate as a probe for monitoring DNA replication instead of [3H]-thymidine where the specific activity of the latter is affected by perturbation of the cellular environment. It was found that to obtain a clean DNA fraction for determining the incorporation of the label, protein must be removed by the use of Proteinase K, and RNA by alkaline hydrolysis. Alkaline hydrolysis of RNA was found superior to RNAse digestion in not leaving behind large RNA fragments that may contaminate the DNA fraction.

Kinetics of ribosomal protein S6 phosphorylation by HepG-2 cells in response to insulin.

The effect of insulin on the phosphorylation of ribosomal protein S6 was studied in a human liver cell line (HepG-2), using [32P] inorganic phosphate. Increased rate of protein S6 phosphorylation was detected 8 min following the addition of insulin to serum starved cells. Maximum enhancement of phosphorylation was observed at 80 nM insulin. Minimum level of insulin required to produce measurable increase of S6 phosphorylation was 20 nM. Radioactivity of protein S6 increased most in the native subunit and polysome fractions. Significant increase in radioactivity of this protein was not observed in the monosome fraction during the first 30 min of insulin stimulation. Increase in the specific radioactivity of native 40S subunit was higher than that of polysomes. These results suggest that phosphorylation takes place in the subunit compartment and moves preferentially into the polysomes.

Biliary lipids in ox and sheep.

1. The lipids of gallbladder bile from sheep are more concentrated than in ox (P less than 0.001). 2. More than 90% of bile acids in both species are conjugated. 3. Taurocholate predominates in sheep while glycocholate predominates in bovine gallbladder bile. 4. The concentration of free bile acids i.e. lithocholate and (deoxycholate + chenodeoxycholate) is higher in bovine than in sheep gallbladder bile (P less than 0.001). 5. The ratio of conjugated to free bile acids is greater in sheep than in bovine gallbladder bile (P less than 0.010). 6. The ratio of total conjugated dihydroxy to trihydroxy bile acids is greater in bovine than in sheep gallbladder bile (P less than 0.001). 7. The ratio of the tauroconjugates of dihydroxy to trihydroxybile acids is higher in bovine than in sheep gallbladder bile (P less than 0.001). 8. It is concluded that the high fibre content of the diet of the two species studied stimulates the liver to form bile acids and hence the low lithogenicity of bile observed.

Effect of recombinant IL-1 isomers on fetal and adult islet cell function and replication.

Natural human interleukin and the recombinant human IL-1 alpha and beta isomers were tested for effects on both fetal and adult rat isolated pancreatic islets. IL-1 alpha inhibited both replication and insulin secretion and decreased the insulin content of both islets. In contrast, the beta isomer did not affect fetal islets. It is concluded that the two isomers may have different avidities for the same receptors, or that receptors are expressed at different stages of differentiation.

The aetiology of galactorrhoea in women with regular menstruation and normal prolactin levels.

Fourteen primary infertile women with expressible galactorrhoea associated with regular ovulatory cycles and normal basal prolactin levels (group A) were matched for age and weight with 14 infertile women with regular menstruation but no galactorrhoea (group B). Both groups showed equivalent increases in prolactin levels after stimulation with 200 micrograms thyrotrophin-releasing hormone (TRH) during the follicular and luteal phases of the menstrual cycle. Patients in group A had a greater increase in luteinizing hormone levels after 100 micrograms i.v. injection of a luteinizing hormone-releasing hormone during the follicular phase (P less than 0.05). Following a 60 mg oral dose of buspirone hydrochloride on day 22 of the menstrual cycle, patients in group A had a greater increase in prolactin levels than patients in group B (P less than 0.01). This reflects hyper-responsive 5-hydroxytryptamine type 1A (5HT1A) receptors in group A patients and may explain the presence of galactorrhoea in these patients despite normal basal and post-TRH prolactin levels.

Effect of a hypoglycaemic sulphonylurea (HB419) and a non-metabolizable amino acid (BCH) on the insulin release and endogenous substrate metabolism of rat pancreatic islets.

The effects of the hypoglycaemia sulphonylurea glibenclamide (HB419) and the non-metabolizable leucine analogue beta-2-aminobicyclo(2.2.1)heptane-2-carboxylic acid (BCH) on insulin release and endogenous substrate metabolism were studied in isolated rat islets. Pre-labelling of the endogenous islet substrate was performed with [14C]glucose (20 mM) or [14C]glutamine (10 mM) during a 24 h tissue culture period before measurements of insulin release or 14CO2 production in short-term incubations. Both HB419 and BCH stimulated the insulin release of the cultured islets, although BCH only after culture of islets with glutamine. The rate of labelling of the islets with [14C]glucose reached an apparent plateau after 16 h in culture and the total islet accumulation of glucose carbon over the 24 h period averaged 12.9 +/- 3.0 nmol/25 islets. Less than 0.5% of the glucose residues was converted to glycogen whereas lipids represented about 2.5%. Fractionation of lipids showed 67% phospholipids, 18% triacylglycerols, 11% diacylglycerols and 6% non-esterified fatty acid. The islet accumulation of glutamine during 24 h corresponded to 11.5 +/- 1.5 nmol/25 islets. After pre-labelling of islets with [14C]glucose there was no effect on the 14CO2-evolution over a 30 min incubation period of either HB419 or BCH. There was also no effect of HB419 after pre-labelling with [14C]glutamine, whereas, in this latter situation, a significant stimulation was observed with BCH. It is concluded that the effects on the pancreatic B-cells by antidiabetic sulphonylureas are not mediated via nutrient receptors.

The structure and hormone content of the endocrine pancreas of the one-humped camel (Camelus dromedarius).

Little is known of the blood sugar regulation in the camel and the morphology and function of its endocrine pancreas. The present paper describes the light microscopic structure and hormone content of the endocrine pancreas of the one-humped camel. Staining of pancreatic sections with haematoxylin-eosin or aldehyde-fuchsin showed numerous islets evenly distributed in all parts of the pancreas. Immunocytochemical staining for insulin or glucagon indicated that islets were predominantly composed of centrally located B-cells, surrounded by a peripheral rim of A-cells. Corresponding stainings for somatostatin or pancreatic polypeptide (PP) demonstrated that D-cells comprised only a small part of the islet volume while PP-cells were common both within and outside the islets. There were no obvious differences between the frequency of the various islet cells in different pancreatic regions. The pancreatic hormone concentrations roughly corresponded to the frequency of the different islet cell types. Insulin appeared most abundant followed by glucagon, PP and somatostatin in decreasing order. The concentrations of each of the hormones were similar in different regions of the gland. It is concluded that the endocrine pancreas of the one-humped camel is dispersed into islets of the same size and cellular composition as has been described in many other mammalian species.

Effects of steroid hormones on DNA synthesis and insulin production of isolated foetal rat pancreatic islets in tissue culture.

This study describes the effects of prednisolone, oestradiol-17B and progesterone on DNA replication and insulin biosynthesis and release of cultured foetal rat islets. Prednisolone significantly inhibited the incorporation of [3H]-thymidine into DNA of islets cultured at a physiological (5.5 mmol/l) but not at a high (22 mmol/l) glucose concentration. It also increased insulin biosynthesis and release of islets cultured at 5.5 mmol/l glucose. Oestradiol-17B reduced the incorporation of [3H]-thymidine into islet DNA at both glucose concentrations, but had no effect on insulin biosynthesis and release. Progesterone had no effect on either the growth or the function of the cultured foetal islets. The observations show a clear dissociation between the action of prednisolone on islet growth versus islet function. They also support the view that neither progesterone nor oestradiol is directly involved in the high rate of B-cell replication previously observed in islets of pregnant rats.

Ultrastructure of the intrahepatic bile duct system of the one-humped camel (Camelus dromedarius).

The ultrastructure of the intrahepatic bile duct system of the camel was studied. Bile canaliculi are present as cavities containing numerous microvilli between adjacent hepatic cells. They are lined with cuboidal cells surrounded by a basal lamina. The lateral plasma membranes of adjacent cells form extensive evaginations below tight junctions at the lumenal surface. The basal surface is open. Collagen bundles are arranged mainly in parallel to the duct.

Ovarian electrocautery: responders versus non-responders.

The endocrine and biophysical attributes of 29 patients with polycystic ovarian disease (PCOD) were examined before and after laparoscopic ovarian electrocautery to establish the criteria which determined their clinical response. Patients with high luteinizing hormone (LH) levels (> 12 IU/l) had a better response than patients with lower LH values but a high LH: follicle stimulating hormone (FSH) ratio (> or = 2:1). However, the body mass index, ovarian volume or pretreatment testosterone values were not helpful in predicting the clinical outcome. Both responders (n = 22) and non-responders (n = 7) showed a decline in LH and testosterone and an increase in FSH 6-h mean values, following 15-min blood sampling 1 month after surgery, compared to the corresponding pretreatment levels. The magnitude of change was significantly higher for LH (p < 0.01) in responders but there was no difference in the corresponding values of the other hormones between the two groups (p > 0.05). Accordingly, unsupplemented ovarian electrocautery is better reserved for the treatment of patients with PCOD and high LH values. Four of the 17 patients who had second-look diagnostic laparoscopy showed minimal to mild pelvic adhesions and all patients had patent Fallopian tubes. The performance of a second-look diagnostic laparoscopy is not indicated as a routine procedure in all cases following surgery.

Polycystic ovaries: do these represent a specific endocrinopathy?

Serum pituitary gonadotrophins, oestradiol, testosterone and insulin pulse patterns were examined at 15-min intervals for 6 h in 40 women with a previous diagnosis of polycystic ovarian disease (PCO) based on clinical, endocrinological and ultrasound data. Age, duration of symptoms, body mass index (BMI) and ovarian volume showed no correlation with the 6-h mean value of any hormone and testosterone blood levels did not correlate with those of insulin. Some patients had high and others low LH pulse pattern components, and few had an inverted LH:FSH ratio. Morphological polycystic ovarian changes may be a reflection of various rather than a single pattern of gonadotrophin secretion.

Implications of ultrasonically diagnosed polycystic ovaries. I. Correlations with basal hormonal profiles.

The incidence of ultrasonically diagnosed polycystic ovaries (PCO) was studied in 389 Arab patients with different types of menstrual dysfunction and 100 normal women with regular menstruation. Two-hundred-and-forty-six patients (63.2%) were found to have PCO but only 206 (53.0%) were confirmed as cases of polycystic ovarian disease (PCOD) on endocrine grounds. Polycystic ovaries were diagnosed in 50% of patients with hyperprolactinaemia, 36.4% with hypothyroidism, 23.7% with hypothalamic dysfunction, 100% with adrenal 21-hydroxylase deficiency and in 16.0% of normal women. More women with PCOD presented with oligomenorrhoea or dysfunctional uterine bleeding (77.7%) and hirsutism (72.3%) but obesity had no discriminating value between the groups with different diagnoses. Ultrasonic diagnosis of PCO should be supplemented with an endocrine biochemical assessment to prevent overdiagnosis of PCOD and to exclude other endocrine dysfunctions.

Implications of ultrasonically diagnosed polycystic ovaries. II. Studies of dynamic and pulsatile hormonal patterns.

Studies of 6-h hormone pulse patterns distinguished patients with polycystic ovarian disease (PCOD) from those with hyperprolactinaemia or hypothyroidism associated with ultrasonically diagnosed polycystic ovaries (PCO). No specific derangement in the gonadotrophin pulse pattern was responsible for these changes, as shown in patients with and without PCO in the latter two groups. These changes may reflect an abnormal ovarian response to normal or abnormal gonadotrophic drive. Out of 26 patients with PCO and elevated dehydroepiandrosterone sulphate (DHEA-S) levels, only three patients (11.5%) proved to have adrenal 21-hydroxylase deficiency. Ultrasonic visualization of polycystic ovaries must be supplemented with an endocrine biochemical assessment. Moreover, mild elevation of DHEA-S, without a concurrently high 17 alpha-hydroxyprogesterone level was not diagnostic of adrenal hyperplasia.

The Effect of Short Term Use of Indomethacin, a Non-Steroidal Anti--inflammatory Drug, on Peritoneal Dialysis Patients.

In this study, the effect of indomethacin a nonsteroidal anti-inflammatory drug (NSAID) was studied to evaluate the effect of its short-term use on peritoneal clearance of urea and creatinine as well as protein excretion in nine chronic renal failure patients on intermittent peritoneal dialysis therapy. Four of them were males and five were females with mean age of 54.4 years. Clearance values and protein excretion in the dialysate effluents were measured before and after administering the indomethacin. There was no significant effect on the urea and creatinine clearance after the use of this drug. However, the mean +/- SE protein concentration in the dialysate effluents decreased significantly after the use of indomethacin from 43.1 + 4.5 to 37.0 + 3.5 mg/dl (p< 0.01). This reduction of protein excretion in the dialysate after the use of indomethacin might enhance the nutritional status of patients on peritoneal dialysis.

Hormonal changes in patients with polycystic ovarian disease after ovarian electrocautery or pituitary desensitization.

Eleven patients with polycystic ovarian disease (PCO) were treated by laparoscopic ovarian electrocautery and nine with a long-acting luteinizing hormone releasing agonist (LHRH-A) for 8 weeks. Both groups showed equivalent significant decreases in their 6-h mean values of luteinizing hormone (LH) and testosterone (T) measured in 25 samples collected every 15 min. Patients treated with ovarian electrocautery showed significant increases in their 6-h mean values of follicle stimulating hormone (FSH) and insulin with variable oestradiol (E2) responses. The magnitude of change following treatment was significantly greater for LH than for FSH. Buserelin medication did not cause persistent significant changes in the levels of insulin or FSH but it did cause a significant reduction in the 6-h mean values of E2. We conclude that LH is the gonadotrophin primarily affected after pituitary desensitization and ovarian electrocautery. Furthermore, there is no direct correlation between the levels of circulating insulin and testosterone in patients with PCO.