Oral, transdermal and vaginal combined contraceptives induce an increase in markers of chronic inflammation and impair insulin sensitivity in young healthy normal-weight women: a randomized study.

STUDY QUESTION: What is the effect of alternative administration routes of combined contraceptives (CCs) on androgen secretion, chronic inflammation, glucose tolerance and lipid profile? SUMMARY ANSWER: The use of oral, transdermal and vaginal CCs impairs glucose tolerance and induces chronic inflammation. WHAT IS KNOWN AND WHAT THIS PAPER ADDS: Oral CCs worsen insulin sensitivity and are associated with increased levels of circulating inflammatory markers, whereas the metabolic effects of transdermal and vaginal CCs have been reported to be minimal. This is the first study comparing three different administration routes of CCs on metabolic variables. STUDY DESIGN, SIZE AND DURATION: This randomized (computer-generated) open-label 9-week follow-up study was conducted at the Oulu University Hospital, Finland. Fasting blood samples were collected at baseline and thereafter at 5 and 9 weeks of treatment, and serum levels of 17-hydroxyprogesterone, androstenedione, testosterone, C-reactive protein (CRP), sex hormone-binding globulin (SHBG), glucose, insulin, C-peptide, total, low-density lipoprotein and high-density lipoprotein cholesterol and triglycerides were measured. Oral glucose tolerance tests were performed and plasma levels of pentraxin 3 (PTX-3) were measured at 0 and 9 weeks. The randomization list, with an allocation ratio of 1:1:1 and block size of six, was computer generated and constructed by a pharmacist at the Oulu University Hospital. The research nurse controlled the randomization list and assigned participants to their groups at the first visit. PARTICIPANTS AND SETTING: Forty-two of 54 healthy women who entered the study used oral contraceptive pills (n = 13), transdermal contraceptive patches (n = 15) or contraceptive vaginal rings (n = 14) continuously for 9 weeks. Inclusion criteria were regular menstrual cycles, at least a 2-month washout as regards hormonal contraceptives and no medication. MAIN RESULTS AND THE ROLE OF CHANCE: Serum levels of SHBG increased and consequently the free androgen index (FAI) decreased in all study groups from baseline to 9 weeks of treatment [FAI, oral: 1.3 (95% confidence interval, CI: 0.94; 1.62) to 0.40 (0.25; 0.54); transdermal: 1.2 (0.96; 1.4) to 0.36 (0.30; 0.43); vaginal: 1.6 (1.1; 2.1) to 0.43 (0.29; 0.58), P < 0.001 in all groups]. Insulin sensitivity was reduced at 9 weeks in all three groups according to the Matsuda index [oral: 7.3 (5.5; 9.0) to 5.6 (3.9; 7.3); transdermal: 9.1 (6.7; 11.4) to 6.6 (4.5; 8.8); vaginal: 7.7 (5.9; 9.5) to 5.4 (3.9; 7.0), P= 0.004-0.024]. Levels of HDL cholesterol, triglycerides and CRP rose in all three groups [CRP, oral: 0.70 (0.38; 1.0) to 5.4 (1.0; 9.9) mg/l; transdermal: 0.77 (0.45; 1.1) to 2.9 (1.4;4.4) mg/l; vaginal: 0.98 (0.52; 1.4) to 3.7 (-0.25; 7.7, a negative value due to skewed distribution to right) mg/l, P≤ 0.002 in all groups] and PTX-3 levels increased in the oral and transdermal study groups (P = 0.007 and P = 0.002). WIDER IMPLICATIONS OF THE FINDINGS: Although the long-term consequences of the present results remain undetermined, these findings emphasize the importance of monitoring glucose metabolism during the use of CCs, especially in women with known risks of type 2 diabetes or cardiovascular diseases. BIAS, LIMITATIONS, GENERALIZABILITY: The number of subjects was relatively low. Moreover, the 9-week exposure to CCs is too short to draw conclusions about the long-term health consequences. However, as the subjects were healthy, normal-weight young women, the possible alterations in the glucose and inflammatory profiles among women with known metabolic risks might be even greater. STUDY FUNDING/COMPETING INTERESTS: This work was supported by grants from the Academy of Finland, the Sigrid Jusélius Foundation, the Finnish Medical Foundation, the Research Foundation of Obstetrics and Gynecology, Oulu University Scholarship Foundation, the North Ostrobothnia Regional Fund of the Finnish Cultural Foundation, the Tyyni Tani Foundation of the University of Oulu and the Finnish-Norwegian Medical Foundation. No competing interests. TRIAL REGISTRATION NUMBER: NCT01087879.

Atrial natriuretic peptide (ANP) in rat gastrointestinal tract.

Acid extracts of rat stomach and small intestine contained 8.6 +/- 3.7 and 39 +/- 15 ng/g of immunoreactive atrial natriuretic peptide (ANP). When studied by gel filtration and reverse-phase high-performance liquid chromatography, the stomach immunoreactivity consisted of multiple components, whereas the small intestine contained mostly proANP and ANP 1-28-like material. These findings indicate that ANP may have a role in the physiology of the gastrointestinal tract, e.g. in the regulation of water and electrolyte absorption.

Role of putative neurotransmitters in the central gastric antisecretory effect of prostaglandin E2 in rats.

The role of putative neurotransmitters of the central nervous system in the central gastric antisecretory effect of prostaglandin E2 (PGE2) was investigated in pylorus-ligated rats. Pretreatment of the rats with an intracerebroventricular (i.c.v.) injection of 6-hydroxydopamine (6-OHDA) prevented the antisecretory effect of the i.c.v. administration of PGE2, whereas pretreatment with 5,6-dihydroxytryptamine (5,6-DHT) plus p-chlorophenylalanine (PCPA) had no effect. I.c.v.-administered phentolamine and idazoxan antagonized the inhibition of gastric secretion induced by i.c.v. PGE2, whereas prazosin, propranolol and sulpiride injected via the same route were ineffective. Diphenhydramine, cimetidine, naloxone and theophylline, all administered i.c.v., did not modify the antisecretory effect of i.c.v. PGE2. The results suggest that an activation of alpha 2-adrenoceptors in the brain is involved in the central gastric antisecretory effect of PGE2, whereas neither central 5-hydroxytryptamine receptors, alpha 1- or beta-adrenoceptors, D2-dopamine receptors, histamine or opioid receptors nor adenosine seem to play any role here.

Inhibition of gastric acid secretion by intracerebroventricularly administered prostaglandin e2 in anaesthetized rats.

1 The effect of intracerebroventricularly (i.c.v) administered prostaglandin E2 (PGE2) on gastric acid secretion was studied in anaesthetized rats. 2 Intracerebroventricular injections of 1 and 3 micrograms of PGE2 inhibited gastric acid secretion induced by insulin, electrical vagal stimulation and acetylcholine in a dose-dependent manner. 3 Intravenous injections of 1,3 and 10 micrograms of PGE2 had no effect on insulin-stimulated gastric acid output. 4 The results indicate that upon i.c.v. administration PGE2 inhibits gastric acid secretion by acting centrally and not peripherally after leakage from the central nervous system.

Central nervous system effects of arachidonic acid, PGE2, PGF2 alpha, PGD2 and PGI2 on gastric secretion in the rat.

The effects of arachidonic acid, prostaglandin E2 (PGE2), PGF2 alpha, PGD2 and PGI2 on gastric secretion (acid, pepsin and volume) after intracerebroventricular administration were investigated in conscious, pylorus-ligated rats. Arachidonic acid 30-1000 micrograms had no effect on gastric secretion. PGE2 3 and 10 micrograms, reduced gastric secretion as measured 1 hour after injection, although the inhibition induced by 3 micrograms disappeared by 2 h. PGF2 alpha 10 and 30 micrograms, inhibited gastric secretion as measured after 1 h, whereas no change was observed in the gastric contents collected 2 h after 10 micrograms of PGF2 alpha. Intramuscular injection of 30 micrograms of PGF2 alpha had no effect on gastric secretion. Intracerebroventricular administration of 1-30 micrograms of PGD2 or PGI2 had no effect on gastric secretion. The results indicate that PGE2 and PGF2 alpha have a potent central antisecretory action in conscious, pylorus-ligated rats, whereas arachidonic acid, PGD2 and PGI2 do not have any central effects on gastric secretion. It is suggested that PGE2 and PGF2 alpha may be involved in the central nervous system control of gastric secretion.

Effect of gastric secretagogues on the formation of inositol phosphates in isolated gastric cells of the rat.

The effects of compounds affecting gastric acid secretion were studied on the formation of inositol phosphates after prelabelling with [3H]-inositol in enriched gastric parietal cells of the rat, prepared by isopycnic centrifugation with Percoll. In cell preparations with 60 to 70% parietal cells, carbachol (10(-6)-10(-2) M) enhanced the accumulation of [3H]-inositol monophosphate ([3H]-IP1), [3H]-inositol bisphosphate ([3H]-IP2) and [3H]-inositol trisphosphate ([3H]-IP3) in a concentration-dependent manner, an effect which was antagonized by 10(-8) M atropine. Li+ (0.5-30 mM) enhanced the basal and carbachol-induced accumulation of all three [3H]-inositol phosphates, the formation of [3H]-IP1 being more sensitive to Li+ than those of [3H]-IP2 and [3H]-IP3. The concentration of Ca2+ in the incubation medium did not affect the relative stimulation of the accumulation of [3H]-inositol phosphates by carbachol, although the basal formation was higher in the presence of Ca2+ in the medium. In the absence of added Ca2+, the incorporation of [3H]-inositol into phospholipids was increased--an effect which was further enhanced by the addition of EGTA to the medium. Gastrin and pentagastrin (10(-8)-10(-5) M) enhanced the formation of [3H]-inositol phosphates, although they were clearly less effective than carbachol. Histamine (10(-6)-10(-3) M) had no effect of its own, but slightly attenuated the effect of carbachol. Cholecystokinin octapeptide (10(-9)-10(-6) M) slightly increased the formation of [3H]-inositol phosphates. Indomethacin (10(-4) M) had no consistent effect on the basal and carbachol-induced accumulation of [3H]-inositol phosphates, nor did prostaglandin E2 (10(-5) M) modify it. Adrenaline (10(-3) M), 5-hydroxytryptamine (10(-3) M), forskolin (10(-5) M), vasopressin (10(-5) M), angiotensin II (10(-5) M) and bombesin (10(-9)-10(-6) M) were all without effect. We suggest that the hydrolysis of inositol phospholipids may be involved in the signal transduction mechanism by which the activation of the muscarinic and gastrin receptors on the parietal cells leads to Ca2+ mobilization and the stimulation of hydrogen ion secretion.

Gastric mucosal blood flow in ethanol-induced mucosal damage in the rat.

Gastric mucosal blood flow in ethanol-induced mucosal damage was studied in urethane-anaesthetic rats by reference to 14C-aminopyrine clearance in the gastric mucosa. Irrigation of the stomach with 30% ethanol in acid saline (100 mM HCl plus 50 mM NaCl) for 40 min broke the gastric mucosal barrier, as indicated by an increased outflow of Na+ and K+ ions and back-diffusion of H+ ions. Gastric mucosal blood flow also increased about 2-fold decreasing after cessation of ethanol irrigation along with the net ion fluxes. The increase in gastric mucosal blood flow occasioned by 10, 20 and 30% ethanol in acid saline was directly proportional to the net fluxes of H+, Na+ and K+ ions. When the stomach was irrigated with 30% ethanol in a less acid medium (10 mM HCl, 90 mM choline chloride plus 50 mM NaCl) there was still a significant increase in the outflow of Na+ and K+ ions, but only a slight-back-diffusion of H+ ions. During this low rate of acid back-diffusion 30% ethanol reduced gastric mucosal blood flow by about 50%. The results suggest that ethanol-induced mucosal damage in the rat is associated with an increase in gastric mucosal blood flow only if combined with back-diffusion of H+ ions.

Effect of intragastric acid on pepsinogen secretion in the rat.

When the osmolarity of intragastric instillates was constant (200 mosmole/l) the changes in their hydrogen ion concentrations (maximally from 0.01 to 100 mM) did not significantly affect pepsinogen secretion in anaesthetised rats. Solutions of 100 mM HCl, 100 mM NaCl and 150 mM sucrose all produced a similar stimulation of pepsinogen secretion. The results do not support the view that the gastric mucosa contains receptors sensitive to hydrogen ions regulating pepsinogen secretion under physiological conditions.

Central inhibitory action of prostaglandin E2 on gastric secretion in the rat.

The central nervous system action of prostaglandin E2 (PGE2) on gastric secretion was studied in conscious pylorus-ligated rats. It was found that 1-10 micrograms of PGE2 given i.c.v. reduced the gastric output of acid, pepsin and fluid in a dose-dependent manner whereas 30-300 micrograms was required for inhibition when given subcutaneously. Given i.c.v. 400 micrograms of indomethacin, a potent inhibitor of prostaglandin biosynthesis, enhanced gastric secretion but not when given intramuscularly. It is concluded that PGE2 has a central inhibitory action on gastric secretion in conscious rats. In addition, the possibility that PGE2 may be involved as inhibitory modulator in the central control of gastric secretion warrants further attention.

Central gastric antisecretory action of adenosine in the rat.

The effects of intracerebroventricularly (i.c.v.) administered adenosine and some of its analogues on gastric secretion were studied in rats. The compounds inhibited the gastric output of acid, pepsin and fluid in pylorus-ligated rats in a dose-dependent manner with an order of potency: 5'-N-ethylcarboxamidoadenosine (NECA) greater than (-)N6-phenylisopropyladenosine (R-PIA) greater than (+)N6-phenylisopropyladenosine (S-PIA) greater than adenosine. Pretreatment with 10 and 30 mg/kg of theophylline i.v. or 5 mg/kg of 8-phenyltheophylline s.c. did not modify the antisecretory effect of 0.1 microgram of NECA i.c.v. NECA injected i.c.v. did not affect the secretion induced by carbachol in awake rats subjected to vagotomy or in anaesthetized rats with intact vagi. NECA i.c.v. had no effect on the serum concentration of gastrin. The depletion of brain monoamines (noradrenaline, dopamine and serotonin) with 6-OHDA i.c.v. significantly attenuated the inhibitory action of NECA. Pretreatment with 10 mg/kg of naloxone i.v. or indomethacin s.c. did not modify the antisecretory effect of NECA. The results indicate that adenosine inhibits gastric secretion in rats by a decrease in the stimulatory vagal impulses to the stomach, and that it acts in the brain via receptors insensitive to xanthines. Brain biogenic monoamines, but not opioid peptides or prostaglandins seem to be involved in the central gastric antisecretory action of adenosine.

Vagally mediated bradycardiac effect of prazosin in anaesthetized rats.

Prazosin at doses of 100 or 1000 nmol/kg i.v. induced bradycardia in urethane-anaesthetized rats. The bradycardia was completely abolished by atropine pretreatment or vagotomy. Prazosin also stimulated gastric acid secretion. This effect was completely abolished by vagotomy. The maximal hypotensive effect after prazosin i.c.v. occurred 10 min later than with i.v. administration. It is concluded that prazosin possesses vagal a stimulating property that may in part explain its modest tachycardiac effect seen in clinical use. The mechanism of vagal stimulation by prazosin remains unclear. The present findings suggest that prazosin does not exert any direct effects on central vagal structures accessible from the ventricular space.

Vagal-dependent stimulation of gastric acid secretion by intracerebroventricularly administered atrial natriuretic peptide in anaesthetized rats.

The intracerebroventricular (i.c.v.) administration of 10 micrograms of atrial natriuretic peptide (ANP) stimulated spontaneous gastric acid secretion in anaesthetized rats, whereas this dose of ANP was without any effect after i.v. injection. The secretagogue action of i.c.v. administered ANP was totally abolished by truncal vagotomy. The findings indicate that i.c.v. administration of ANP stimulates gastric acid secretion in anaesthetized rats by a central, vagal-dependent mechanism.

Centrally administered PGE2 inhibits gastric secretion in the rat by releasing vasopressin.

The pituitary mechanism of the central inhibitory effect of PGE2 on gastric secretion was investigated in rats. The intravenous (i.v.) injection of neurointermediate lobe extracts but not of anterior lobe extracts inhibited gastric secretion in pylorus-ligated rats. The i.c.v. administration of 3 micrograms of PGE2 increased the plasma level of vasopressin but had no effect on plasma beta-endorphin/beta-lipotropin. The i.v. administration of 10 micrograms d(CH2)5Tyr(Me) AVP, a vasopressin antagonist, prevented the inhibition of gastric secretion induced by i.c.v. administration of 3 micrograms PGE2 to pylorus-ligated rats. The results indicate that the central antisecretory action of PGE2 is due to the release of vasopressin from the pituitary gland.

Ethanol, indomethacin and gastric acid secretion in the rat.

In the anesthetized rat, 10% ethanol in the gastric lumen causes a total cessation of the secretion of gastric acid within 30--60 min. This effect is inhibited by pretreatment with indomethacin, which is a potent inhibitor of prostaglandin synthesis. Therefore, the inhibitory effect of local ethanol on gastric acid secretion in the rat may be mediated, at least partly, by increased synthesis of prostaglandins.

Ethanol-induced changes in gastric mucosal content of cyclic AMP and ATP in the rat.

The stomach of urethane-anaesthetised rats was perfused with 10% (v/v) ethanol. At 40 min, the secretion of acid was strongly inhibited and the contents of cyclic AMP and ATP were lowered in the superficial mucosa, but not in whole gastric mucosa. After discontinuation of the ethanol perfusion, the rate of gastric acid output as well as the cyclic AMP and ATP levels recovered. Most of the acid-secreting parietal cells were found in the superficial mucosa where the changes of cyclic AMP and ATP took place. The results suggest that lowering of the mucosal contents of cyclic AMP and ATP may be involved in the ethanol-induced inhibition of the gastric acid output.

Effects of ethanol on the cyclic AMP system of the dog gastric mucosa.

The effect of ethanol on the cyclic AMP system of the dog fundic mucosa was studied in vitro. The gastric mucosal content of cyclic AMP was increased by 2.5% ethanol, whereas 10 and 20% ethanol decreased the mucosal content of cyclic AMP. The activity of adenylate cyclase was increased by 2.5 and 5% ethanol, whereas 10% ethanol did not significantly affect it. The activity of cyclic AMP phosphodiesterase was inhibited by ethanol in a competitive manner. The increase in the gastric mucosal content of cyclic AMP, induced by low concentrations of ethanol, is apparently due to the stimulation of adenylate cyclase and inhibition of phosphodiesterase. Changes in the phosphodiesterase or adenylate cyclase activites do not explain the decrease of the mucosal content of cyclic AMP by higher concentrations of ethanol. The mechanism of the decrease is discussed.

Cardiovascular action of detomidine, a sedative and analgesic imidazole derivative with alpha-agonistic properties.

The cardiovascular effects of detomidine, a new veterinary sedative and analgesic imidazole derivative were studied in rats and cats using as reference compound xylazine, a widely employed veterinary antinociceptive and sedative drug with alpha-agonistic potency. Detomidine (1-30 micrograms/kg i.v.) and xylazine (10-1000 micrograms/kg i.v.) had both dose-dependent hypotensive and bradycardiac effects in anaesthetized rats. After i.v. administration of 3-100 micrograms/kg detomidine and 0.1-3 mg/kg xylazine to conscious rats, detomidine was more active in reducing the heart rate than in lowering the blood pressure. In anaesthetized cats, detomidine (1-30 micrograms/kg i.v.) was hypotensive and bradycardiac in a dose-dependent manner. A low dose of detomidine into the vertebral artery was more effective than i.v. application in reducing blood pressure. Idazoxan (0.3 mg/kg i.v. and 0.03 mg/kg into the vertebral artery) antagonized the hypotensive and bradycardiac effects of detomidine injected into the femoral vein or vertebral artery, respectively. In pithed rats, detomidine and xylazine stimulated presynaptic and postsynaptic alpha 2-adrenoceptors, and to a lesser extent postsynaptic alpha 1-adrenoceptors. The results indicate that detomidine is an agonist of central and peripheral alpha 2-adrenoceptors which exerts its hypotensive and bradycardiac effects via activation of the central alpha 2-adrenoceptors.

Centrally applied histamine increases gastric acid secretion in rats.

Administration of 10-100 micrograms of histamine into the lateral cerebral ventricle of anaesthetized rats stimulated gastric acid secretion in a dose-dependent manner, while subcutaneous (s.c.) injections of the same doses produced clearly less pronounced increases in the acid output. In vagotomized rats only a marginal response to histamine given into the lateral ventricle was observed. When injected into the third cerebral ventricle the doses of histamine needed for the stimulation of gastric acid secretion were 1-10 micrograms, the effect being totally abolished by vagotomy. The results indicate that histamine is capable of stimulating gastric acid secretion by a central, vagal-dependent mechanism.

Stimulation of central alpha-2 adrenoceptors inhibits gastric secretion in rats by releasing vasopressin.

The mechanism of the gastric antisecretory action of the stimulation of central alpha-2 adrenoceptors were studied in conscious, pylorus-ligated rats using intracerebroventricularly (i.c.v.) administered oxymetazoline as the model substance. I.c.v. administration of 10 micrograms of oxymetazoline strongly inhibited the secretion of acid, pepsin and fluid, whereas upon s.c. injection this dose was without any effect. Pretreatment with idazoxan abolished the antisecretory effect of i.c.v. administered oxymetazoline. I.c.v. injected oxymetazoline inhibited gastric secretion induced by carbachol in vagotomized rats, but the inhibitory effect was less pronounced than on the spontaneous secretion in rats with intact vagi. Hypophysectomy abolished the antisecretory effect of i.c.v. oxymetazoline, and pretreatment with the vasopressin antagonist, d(CH2)5Tyr(Me)AVP, significantly attenuated it. The results suggest that the inhibition of gastric secretion by the stimulation of central alpha-2 adrenoceptors in rats is mediated in part by vasopressin released from the pituitary gland.

Osmotic stimulation of pepsin secretion in the rat.

The effect of the osmolarity of intragastric instillates on pepsin secretion was studied in rats anaesthetised with urethane. Irrigation of the stomach with solutions of sucrose and NaCl, resp. caused a concentration-dependent increase in pepsin output. A stimulation was observed already by hypotonic solutions and the maximal effect was obtained by 300 m-osmole/l of sucrose and by 600 m-osmole/l of NaCl (13- and 10-fold stimulation resp.). A similar time course in the increase of pepsin output was produced by hyperosmotic solutions (600 m-osmole/l) of sucrose, urea, NaCl and choline chloride. Pepsin output was stimulated maximally within 30 min and decreased thereafter, but remained at about 4--6-fold higher levels than during the previous irrigation with distilled water. Replacement of hyperosmotic instillates by distilled water reduced pepsin secretion to the initial level. Hypertonic ethanol (600 m-osmole/l) increased pepsin output only slightly. Vagotomy, pretreatment with atropine (1 mg/kg i.v.) or cimetidine (5 mg/kg i.v.), local anesthesia of the gastric mucosa with 4% lidocaine or intravenous infusion of PGE2 (2 microgram/kg X min) did not antagonise the stimulation of pepsin output induced by hyperosmotic NaCl (600 m-osmole/l). The results indicate that the increase of the osmolarity of intragastric instillates stimulates pepsin secretion in the rat without involvement of neural (vagal or local cholinergic reflexes) or hormonal mechanisms (release of gastrin) which are known to stimulate gastric secretion in the gastric phase.