Pharmacologic doses of interleukin 8 suppositories induce follicular maturation in rabbits.

Interleukin 8 (IL-8) is a neutrophil chemoattractant/activating factor that plays a role in the ovarian physiology leads to investigate the effects of IL-8 on follicular maturation. Experiments were conducted using suppositories containing 100 ng, 200 ng, 400 ng IL-8, 500 microl Witepsol-base (control), human menopausal gonadotropin (im) and conjugate of fluorescein isithiocyanate-labelled IL-8. The levels of IL-8 in ovarian fluid were also measured. Histology of ovaries treated with 200 ng IL-8 showed large antral follicles filled with follicular fluid. The theca layer was divided into an interna and an externa with large extracellular spaces. The granulosa cells were loosened and appeared to be detaching from the granulosa layer. Neutrophils were localized predominantly in the theca and medulla (P<0.0001, P<0.004), and relative collagen concentration was significantly decreased in ovaries of 200 ng IL-8 (P<0.0001) compared with controls. The IL-8 was detected in ovarian fluid after 6 h (P<0.0001), 12 h (P<0.001), and 18 h (P<0.01) compared with 0 h. Fluorescein isithiocyanate-labelled IL-8 conjugate was seen in the follicular wall and endometrium. We conclude that pharmacological dosage of exogenous IL-8 exerts an effect on follicular maturation through granulocyte chemotaxis and activation.

Increased concentrations of plasma neuropeptide Y in patients with eclampsia and preeclampsia.

OBJECTIVE: Epinephrine and norepinephrine are associated with the hyperstimulation of the sympathetic nervous system. Neuropeptide Y is a potent vasoconstrictive substance that is released in response to sympathetic nerve stimulation. STUDY DESIGN: The concentrations of plasma neuropeptide Y in pregnant patients with eclampsia (n = 8), preeclampsia (n = 8), and normotension (n = 8) were measured by radioimmunoassay on admission and 6 days after delivery. Correlations between plasma concentration of neuropeptide Y and mean arterial blood pressure were also evaluated in these patients on admission and 6 days after delivery. RESULTS: The plasma level of neuropeptide Y in women with eclampsia (P <.001) and preeclampsia (P <.003) was found to be significantly elevated with respect to that in normotensive pregnant women. At 6 days after delivery the concentration of plasma neuropeptide Y was significantly decreased in women with eclampsia, women with preeclampsia, and women with normotensive pregnancies compared with the value measured on admission (P <.0001, P <.0001, and P <.002, respectively). At admission the plasma neuropeptide Y level was positively correlated with mean arterial blood pressure in women with eclampsia and preeclampsia. However, no significant correlations were observed between plasma neuropeptide Y concentration and mean arterial blood pressure both at admission and 6 days after delivery in normotensive pregnant women and 6 days after delivery in women with eclampsia and preeclampsia. CONCLUSION: We have concluded that the level of neuropeptide Y in plasma is increased in women with eclampsia and preeclampsia. Elevated plasma neuropeptide Y levels may play a key role in the development of eclampsia and preeclampsia.

PIP: This study investigated the plasma concentrations of neuropeptide Y associated with the pre-eclamptic and eclamptic conditions. Subjects included patients in the third trimester of pregnancy admitted at Dhaka Medical College Hospital, Bangladesh, from January 1996 to March 1998 with untreated eclampsia (n = 8), pre-eclampsia (n = 8), and normotensive pregnancy (n = 8). Nonpregnant healthy volunteer women (n = 8) were also enrolled. Blood samples were collected and the concentrations of plasma neuropeptide Y were measured by radioimmunoassay on admission and 6 days after delivery. The correlations between plasma concentration of neuropeptide Y and mean arterial blood pressure were also evaluated in these patients. The findings showed that the plasma level of neuropeptide Y in women with eclampsia (P 0.001) and pre-eclampsia (P 0.003) was significantly elevated in comparison with normotensive pregnant women. At 6 days after delivery the concentration of plasma neuropeptide Y was significantly decreased in women with eclampsia, women with pre-eclampsia, and women with normotensive pregnancies compared with the value measured on admission (P 0.0001, P 0.0001, and P 0.002, respectively). At admission the plasma neuropeptide Y level was positively correlated with mean arterial blood pressure in women with eclampsia and pre-eclampsia. However, no significant correlations were observed between plasma neuropeptide Y concentration and mean arterial blood pressure both at admission and 6 days after delivery in normotensive pregnant women and 6 days after delivery in women with eclampsia and pre-eclampsia. The study concluded that the level of neuropeptide Y in plasma is increased in women with eclampsia and pre-eclampsia.

Induction of eclampticlike changes by stimulation of the celiac ganglion in rats.

OBJECTIVE: To examine the relation between the stimulation of the abdominal sympathetic nervous system and vasospasm of the brain in eclamptic seizures, we analyzed brain blood flow after stimulation of the celiac ganglion by lipopolysaccharide (LPS, 5, 50, or 500 mg/mL) or normal saline before and after denovation of sympathetic trunk in pregnant and nonpregnant rats. METHODS: The brain blood flow was measured after stimulation of the celiac ganglion with 50 microL (5 mg/mL) LPS in group I, 50 microL (50 mg/mL) LPS in group II, 50 microL saline in group III, and 50 microL (500 mg/mL) LPS (after denovation of the sympathetic trunk) in group IV. A sham control experiment was also done by stimulation of the abdominal peritoneum with 50 microL (500 mg/mL) LPS in group V. Changes in water content and histological findings in the brain were also studied in this protocol. RESULTS: A significant reduction in brain blood flow was observed in pregnant rats in groups I and II on stimulation of the celiac ganglion with LPS (p < 0.0001, p < 0.001) compared with before stimulation. Celiac ganglion stimulation with saline (group III) and LPS (group IV, after denovation of the sympathetic trunk) did not affect brain blood flow. Stimulation of the abdominal peritoneum with LPS (group V) could not induce any changes in brain blood flow. Repeated seizures occurred in 60% of pregnant rats and a remarkable increase in water content was observed after LPS stimulation of the celiac ganglion in groups I and II (p < 0.0001, p < 0.001). Histologically, we found that stimulation of the celiac ganglion with LPS caused widening of perivascular spaces with compression of the vessels leading to ischemic changes in brain tissues. There were no such findings observed in other groups. However, a lesser extent effect was noticed in nonpregnant than seen in pregnant rats. CONCLUSION: Stimulation of the abdominal sympathetic ganglions could induce vasoconstriction of the brain vessels, thus decreasing brain blood flow, which results in eclampsialike changes in rats.

Induction of preeclampsia like phenomena by stimulation of sympathetic nerve with cold and fasting stress.

The purpose of this study was to examine the effect of cold-stress, fasting stress and cold plus fasting stress on the sympathetic nerve activity. Pregnant and nonpregnant rats were kept in cold environment (0 degrees C), or fasting condition (12 h), and cold plus fasting condition for 2 weeks. Their plasma corticotrophin releasing factor (CRF), catecholamines, insulin levels, and platelets were measured, and histological examinations were performed. In cold plus fasting stress rats, a significant increased CRF, epinephrine (E), norepinephrine (NE), and insulin levels with decreased platelet count (P<0.0001) were observed compared with control. Histological study revealed that diffused enlarged glomeruli with fibrin deposition in the kidney, hemostasis, ischemic necrosis and fibrin deposition in liver and swelling along with hemorrhagic necrosis in adrenal gland of cold plus fasting stress rats. The biochemical and histological changes in cold plus fasting, cold-stressed or fasting rats were similar to human preeclampsia. The findings observed in cold plus fasting stress rats were more pronounced either than cold-stressed or fasting group. These results demonstrate that cold plus fasting stress is an intense stimulator of sympathetic nervous system than either cold stress or fasting.

The impact of vasoactive peptides on nitric oxide production in cultured sympathetic neurons.

The concentration of nitric oxide was found to be decreased in a hypersympathetic condition. We carried out experiments on cultured sympathetic neurons from 12-14-days-old chick embryos to investigate the role of vasoactive peptides and amine on nitric oxide production. Stimulation of cultured neurons with endothelin-1, norepinephrine and angiotensin-II initially increases nitric oxide production and subsequently decreases it in a dose-dependent manner (P<0.05, n = 7). Stimulation of Fura-2 acetoxymethyl ester-loaded neurons with endothelin-1, norepinephrine and angiotensin-II increases the calcium influx (within 30-90 s) and it is then restored to the initial level (P<0.05, n = 7). An additional observation was that specific stimulator L-arginine significantly increases the nitric oxide release and calcium influx into the cells, whereas N(W)-nitro-L-arginine methyl ester blunts nitric oxide release dose dependently (P<0.05, n = 7) and does not change the calcium concentration in the cells. We propose that vasoactive peptides and amines inhibit nitric oxide production in the cultured sympathetic neuron by regulation of intracellular calcium concentration.

Dehydroepiandrosterone sulphate promotes hyaluronic acid-induced cervical ripening in rabbits.

Hyaluronic acid (HA) stimulates the synthesis of interleukin (IL) 8, while dehydroepiandrosterone sulphate (DHEA-S) induces the expression of IL-8 and its receptor in the human cervical fibroblast. This has led us to investigate the effect of DHEA-S on HA-induced cervical ripening. Experiments were performed in pregnant rabbits using vaginal suppositories containing 1 mg HA, 30 mg DHEA-S, 30 mg DHEA-S + 0.1 mg HA, 30 mg DHEA-S + 1 mg HA, and 500 microl Witepsol-50 base (control). The effects were evaluated by measuring collagenase, gelatinase and elastase activities, water content, neutrophil infiltration, relative collagen concentration and histological assessment. The activities of collagenase, gelatinase and elastase were significantly increased in rabbits treated with DHEA-S + 1 mg HA compared with rabbits treated with DHEA-S + 0.1 mg HA (P < 0.009, P < 0.001, P < 0.009 respectively). Water content was markedly increased in rabbits treated with DHEA-S + 1 mg HA compared with DHEA-S + 0.1 mg HA treatment (P < 0.05). Neutrophil infiltration was markedly increased, while relative collagen concentration was significantly decreased with DHEA-S + 1 mg HA compared with the DHEA-S + 0.1 mg HA approach (P < 0.001, P < 0.002). The histology of cervices treated with DHEA-S + 1 mg HA showed the density of collagen to be markedly decreased, and collagen fibres irregularly separated. Increased vascularity with massive dilatation of blood vessels was also observed in these rabbits. We conclude that DHEA-S upregulates the HA-induced cervical ripening process.

Binding of interleukin-8 to heparan sulphate enhances cervical maturation in rabbits.

Cervical ripening is a cytokine-triggered process with substantial remodelling of the cervical extracellular matrix. Interleukin-8 (IL-8) is an important cytokine in cervical maturation. Glycosaminoglycans are also included in this process, but their role in not clearly understood. The effects of heparan sulphate (HS), hyaluronic acid (HA), IL-8, HS + IL-8 and HA + IL-8 on biochemical properties of the cervix were examined in non-pregnant rabbits. The changes in vascular pattern with collagen structure of the cervices and immunohistochemical studies, together with the relative collagen concentrations, were determined. A reduction in relative collagen concentration was significant after HS + IL-8, IL-8 and HA + IL-8 treatment (all P < 0.0001). Gel electrophoresis analysis showed that IL-8 bound preferentially to HS than to HA. Neutrophils were significantly increased in number (P < 0.0001) and located predominantly beneath the glandular epithelium and around the blood vessels after HS + IL-8 treatment. HS + IL-8 treatment caused cervices to increase their water content and become oedematous. The collagen fibres were considerably dissociated, the interfibrillar spaces markedly dilated, and the blood vessels notably increased and dilated. We conclude that binding to HS enhances the activity of IL-8 in inducing cervical maturation.

A new treatment of severe pre-eclampsia by long-term epidural anaesthesia.

We hypothesised that long term epidural anaesthesia suppresses sympathetic overactivity in pre-eclampsia. Two equal groups of severe pre-eclamptic patients were treated either with long-term epidural anaesthesia (epidural group) or bed rest, diet control, and an antihypertensive drug (control group). After 7 days of epidural block, mean arterial blood pressures decreased, platelet count and serum total protein increased in all cases while proteinuria decreased in four cases. All patients treated with a long-term epidural therapy continued their pregnancies for more than 3 weeks after admission. The infants in the epidural group had 1-min Apgar scores above 8, a body weight of 2240+/-310 g (mean +/- s.d.) and normal neonatal progress. In contrast, the pregnancies of eight patients in the control group were terminated within 2 weeks of admission due to severe pre-eclampsia or foetal distress, the birth weight of the infants was 1590+/-380 g (mean +/- s.d.) and four had neonatal distress. Progressive worsening in the mean arterial pressure, proteinuria, platelet count and serum total protein was found in these patients. Long-term epidural anaesthesia suppresses the sympathetic hyperactivity and thus improves pre-eclamptic condition which may open a new treatment in case of progressive severe pre-eclampsia.

Induction of hypercoagulability condition by chronic localized cold stress in rabbits.

To evaluate the effect of cold-induced stress on renal and hepatic blood flow and coagulation parameters, rabbits' soles were exposed to ice pad (0 degrees C). Renal and hepatic blood flow was measured after 1 h and 15 days of cold stress. Coagulation parameters (0, 8th and 15th days of stress) and histological studies were performed. Renal and hepatic blood flow was significantly reduced after cold-stress. Decreased platelet count, antithrombin III (AT III) activity, increased fibrinogen (Fbg) level, shortened activated partial thromboplastin time (aPTT) and prothrombin time (PT) was found after 8 and 15 days of cold-stress. Histology showed enlarged glomeruli with fibrin deposition in kidney, ischemic changes and fibrin deposition in liver and hemorrhagic necrosis in adrenal cortex. We conclude that undesirable localized cold induced sympathetic stimulation in daily life may be a predisposing factor for coagulopathy.

Urinary trypsin inhibitor suppresses vascular smooth muscle contraction by inhibition of Ca2+ influx.

Urinary trypsin inhibitor (UTI) and its precursor form inter-alpha trypsin inhibitor (ITI) are present in plasma. To determine the action of UTI on blood vessels, we performed isometric vascular muscle contraction tests, microcirculation studies and measurement of cytosolic free Ca2+ in vascular smooth muscle cells. An isometric vascular muscle contraction test showed that the contractions stimulated by endothelin-1 or norepinephrine were suppressed in the presence of UTI, and that the contractions were not inhibited in the presence of ITI. The microcirculation study showed that the contraction of mesenteric arterioles of WKY rats induced by norepinephrine were inhibited by treatment of UTI, and that they did not alter by treatment of ITI. Pre-incubation of UTI, but not ITI, with vascular smooth muscle cells inhibited the increase of cytosolic free Ca2+ induced by endothelin-1 or norepinephrine. Cell-binding study by biotinylated UTI showed that vascular smooth muscle cells have specific binding site for UTI, but not for ITI. We propose that circulating UTI converted from ITI has a regulatory effect on local vascular tone by regulation of Ca2+ influx into smooth muscle cells.

Vasospasms of cerebral arteries in hyperemesis gravidarum.

Hyperemesis gravidarum is associated with nausea and vomiting during the 1st trimester of pregnancy. We report 2 cases of hyperemesis gravidarum associated with vasospasms of cerebral arteries recognized by magnetic resonance imaging angiography. The hyperemetic condition of the patients improved after fluid and electrolyte replacement therapy.

Eclamptic plasma stimulates norepinephrine release in cultured sympathetic nerve.

The purpose of the present study was to evaluate the effect of plasma from eclamptic and preeclamptic patients on cultured sympathetic nerve. Sympathetic neurons from 12- to 14-day-old chick embryos were cultured; the neurons were then stimulated with 50% plasma from eclamptic, preeclamptic, hypertensive, normotensive pregnant, hypertensive, and normotensive nonpregnant women (n=7). Similarly, neurons were individually incubated with mixtures of 50% corresponding plasma with 0.25% bupivacaine or bupivacaine only (n=7). Furthermore, the effects of 1%, 10%, and 50% plasma from eclamptic, preeclamptic, and normotensive pregnant patients (n=7) were also evaluated. Norepinephrine concentrations were measured by high-performance liquid chromatography. Electron microscopic studies of nerve cells were also performed. Stimulation with plasma from eclamptic and preeclamptic women significantly increased norepinephrine concentration (P<0.0001) compared with control. The release of norepinephrine was found to be concentration-dependent. Conversely, norepinephrine secretion was significantly hampered by bupivacaine treatment (P<0.0001). Electron microscopic studies in eclamptic and preeclamptic plasma-stimulated nerve cells showed that perikarya were in close contact with each other and with nerve cell processes. After treatment with bupivacaine, nerve cells were irregular in shape and the cell membranes were demyelinated. These results suggest that eclamptic and preeclamptic plasma has an excitotoxic effect on sympathetic nerve via axoplasmic membrane depolarization, thus increasing norepinephrine secretion that is blocked by bupivacaine. A preeclamptic condition may be improved by depression of sympathetic nerve stimulation.

Decreased renal and hepatic blood flow with preeclampsia-like histologic changes was obtained by stimulation of the celiac ganglion with LPS.

Decreased renal and hepatic blood flow with preeclampsia-like histologic changes was developed by stimulation of the celiac ganglion with lipopolysaccharide (LPS) in pregnant rabbits. The renal and hepatic blood flow were measured after stimulation of the celiac ganglion with 10 microL (group 1), 100 microL (group II) and 300 microL (group III) of LPS (10 mg/mL conc.) and with 100 microL of normal saline (group IV). The bifurcation of the abdominal aorta was also stimulated with 300 microL of LPS (group V). A control experiment was also done in this study (group VI). Histopathological studies of kidney and liver tissue were also performed in this protocol. A significant reduction in renal and hepatic blood flow was observed in pregnant rabbits with the times and dosage dependent on stimulation of the celiac ganglion by LPS. Stimulation of the bifurcation of the abdominal aorta with LPS could not produce any changes in renal and hepatic blood flow. Preeclampsia-like histologic changes of kidney and liver tissue were also observed. These results suggest that exogenous stimulation of the celiac ganglion causes decreased renal and hepatic blood flow, resulting in preeclampsia-like histologic changes of kidney and liver in pregnant rabbits.