Laparoendoscopic two-site myomectomy (LETS-M) using conventional laparoscopic instruments and the glove-port technique.

PURPOSE: To evaluate the perioperative outcome of laparoendoscopic two-site myomectomy (LETS-M). METHODS: The medical records of 204 women receiving LETS-M in a tertiary referral center, including 183 surgeries performed by the experienced surgeon and 21 surgeries performed by 3 well-supervised trainees were retrospectively reviewed. RESULTS: The age of the participants was 39.3 ± 6.4 years. The mean diameter of the largest myoma and the mean number of myomas were 8.5 ± 2.2 cm and 1.7 ± 1.1, respectively. Thirty-one (15%) operations removed more than 2 myomas larger than 5 cm in diameter. The mean weight of the myomas was 281.1 ± 183.1 g. The operation time was 97.6 ± 40.2 min, and the intraoperative blood loss was 99.3 ± 115.2 mL. There were 3 (1%) cases of excessive blood loss (more than 500 mL) and 2 (1%) of postoperative hematoma. The only significant difference between the experienced surgeon and trainees was the operation time (92.3 ± 32.2 min vs. 141.2 ± 54 min, p < .001), while the myoma number, myoma diameter, myoma weight, and intraoperative blood loss were not significantly different. The operation time did not differ among different myoma locations. In multivariate analysis, virginity, myoma number, more than 2 large myomas, and myoma size were independent variables for longer operation times. No patient experienced any major complications. CONCLUSION: LETS-M using conventional laparoscopic equipment is a minimally invasive surgical method that is safe, effective, and easy to learn for managing uterine myoma. It is useful to achieve a favorable perioperative outcome with acceptable operation time.

Role of Amino Acid Arginine and Nitric Oxide in Mechanisms of Cytoprotective Effect of Non-Opiate Leu-Enkephalin Analogue In Vitro.

Incubation of primary culture of pulmonary fibroblasts with non-opiate analogue of leuenkephalin (NALE; Phe-D-Ala-Gly-Phe-Leu-Arg, 0.1 µM) reduced generation of superoxide anion-radical (by 20.7%) and decreased the number of p53+ cells (by 40.2%) induced by exposure to H2O2 (60 µM). The cytoprotective effect of NALE was potentiated by NO synthase inhibitor L-NAME (1 mM): the number of p53+ cells decreased by 65.3% and morphometric parameters of the cell nuclei and nucleoli were improved. Incubation of pulmonary fibroblasts culture with peptide G (Phe-D-Ala-Gly-Phe-Leu-Gly, 0.1 µM) also significantly reduced the damaging effect of H2O2: the number of p53+ cells decreased by 73.5%, the area of cell nuclei returned to normal, and generation of superoxide anion-radical decreased by 18.4%. These results indicate that C-terminal amino acid Arg and activation of NO synthase are not involved in the direct cytoprotective effect of NALE.

Hepatoprotective Effect of Neonatal Administration of Non-Opioid Leu-Enkephalin Analogue in Adult Albino Rats Subjected to Antenatal Hypoxia.

We studied the effects of neonatal administration of non-opioid leu-enkephalin analogue peptide NALE) on the morphological parameters of the liver and redox status of 60-dayold albino rats subjected to antenatal hypoxia. In animals subjected to antenatal hypoxia, a decrease body weight and reduction of the area of hepatocytes and total area of their nucleoli were observed; these changes were accompanied by activation of free-radical processes and impairment of antioxidant defense in the liver and blood serum. Intraperitoneal administration of NALE (100 µg/kg, daily) during postnatal days 2-6 normalized body weight, hepatocyte area, and total area of their nucleoli, significantly reduced the intensity of ROS generation, and improved antioxidant protection in the liver and blood serum in 60-day-old animals subjected to antenatal hypoxia.

Effects of Opioid Peptides on Changes in Lipid Metabolism in Rats Subjected to Swimming Stress.

Blood levels of nonesterified fatty acids, total cholesterol, triglycerides, and LDL increased in rats subjected to forced swimming stress. Administration of opioid peptides dynorphin A(1-13), DSLET, or DAGO reduced stress-induced disturbances in lipid metabolism. Dynorphin A(1-13) and DAGO produced the most pronounced effects and prevented an increase in concentrations of nonesterified fatty acids, triglycerides, total cholesterol, and LDL as soon as 39 h after treatment. Only DSLET increased HDL content in the plasma of stressed rats. The observed effects can be explained by the stress-limiting effects of opioids, e.g. attenuation of the effect of catecholamines on the adipose tissue and inhibition of the generation LPO products suppressing activity of the cholesterol metabolizing enzyme.

Naltrindole derivatives with fluorinated ethyl substituents on the 17-nitrogen as δ opioid receptor inverse agonists.

We synthesized derivatives of the δ opioid receptor (DOR) antagonists naltrindole (NTI) and compound 1 that were modified with small alkyl or fluorinated ethyl substituents on the 17-nitrogen. Although the derivatives showed decreased binding affinities for the opioid receptors, their selectivities for the DOR were higher than the parent compounds NTI and compound 1. Surprisingly, 17-fluoroethyl NTI derivatives exerted DOR inverse agonistic activities. The DOR inverse agonism of compounds 4c-e was less efficacious but significant, as compared with a standard DOR inverse agonist ICI-174864. On the other hand, compound 1 and its derivatives with small alkyl or monofluoroethyl substituents were partial agonists, but the derivatives having di- or trifluoroethyl group showed neither agonistic nor inverse agonistic activities.

Synthesis and analgesic activity of alkylated, reduced and constrained oligoheterocyclic peptidomimetic analogs of Leu-enkephalin.

We report the design and the parallel solid phase synthesis of linear and oligoheterocyclic peptidomimetic analogs of Leu-enkephalin. The described peptidomimetics represent different unique scaffolds that distribute in the space the peptidyl side chains of amino acids essential for biological activity and mimic the bioactive conformation of the Leu-enkephalin peptide. All the compounds were screened in competitive radioligand binding assays to determine their affinities for µ-(MOR), and κ-(KOR) opioid receptors. A reduced analog of Leu-enkephalin TPI1879-26 with activity Ki=60 nM for the mu receptor was identified.

δ opioid receptor antagonist, ICI 174,864, is suitable for the early treatment of uncontrolled hemorrhagic shock in rats.

BACKGROUND: Fluid resuscitation is the essential step for early treatment of traumatic hemorrhagic shock. However, its implementation is greatly limited before hospital or during evacuation. The authors investigated whether δ opioid receptor antagonist ICI 174,864 was suitable for the early treatment of traumatic hemorrhagic shock. METHODS: With uncontrolled hemorrhagic-shock rats, the antishock effects of six dosages of ICI 174,864 (0.1, 0.3, 0.5, 1, 3, and 5 mg/kg) infused with or without a small volume of lactated Ringer's solution (LR) before bleeding controlled or bleeding cessation at different times were observed. RESULTS: ICI 174,864 (0.1-3 mg/kg) with or without 1/4 volume of LR infusion showed dose-dependent increase in the mean arterial blood pressure, and significantly prolonged the survival time and 8-h survival rate, as compared with ICI 174,864 plus 1/2 volume of LR infusion. The best effect was shown with 3 mg/kg of ICI 174,864. Bleeding cessation at 1, 2, or 3 h during infusion of ICI 174,864 (3 mg/kg) plus 1/4 volume of LR improved subsequent treatment (70% 24-h survival rate vs. 50 and 10% 24-h survival rate in hypotensive resuscitation and LR group, respectively). There was significant improvement in hemodynamic parameters, oxygen delivery, and tissue perfusion of hemorrhagic-shock rats with 3 mg/kg of ICI 174,864 plus 1/4 volume of LR infusion. CONCLUSION: δ Opioid receptor antagonist ICI 174,864 alone or with small volume of fluid infusion has good beneficial effect on uncontrolled hemorrhagic shock. Its early application can "buy" time for subsequent treatment of traumatic shock.

Micro-opioid receptor activation in the basolateral amygdala mediates the learning of increases but not decreases in the incentive value of a food reward.

The decision to perform, or not perform, actions known to lead to a rewarding outcome is strongly influenced by the current incentive value of the reward. Incentive value is largely determined by the affective experience derived during previous consumption of the reward-the process of incentive learning. We trained rats on a two-lever, seeking-taking chain paradigm for sucrose reward, in which responding on the initial seeking lever of the chain was demonstrably controlled by the incentive value of the reward. We found that infusion of the µ-opioid receptor antagonist, CTOP (d-Phe-Cys-Tyr-d-Trp-Orn-Thr-Pen-Thr-NH(2)), into the basolateral amygdala (BLA) during posttraining, noncontingent consumption of sucrose in a novel elevated-hunger state (a positive incentive learning opportunity) blocked the encoding of incentive value information normally used to increase subsequent sucrose-seeking responses. Similar treatment with δ [N, N-diallyl-Tyr-Aib-Aib-Phe-Leu-OH (ICI 174,864)] or κ [5'-guanidinonaltrindole (GNTI)] antagonists was without effect. Interestingly, none of these drugs affected the ability of the rats to encode a decrease in incentive value resulting from experiencing the sucrose in a novel reduced-hunger state. However, the µ agonist, DAMGO ([d-Ala2, NMe-Phe4, Gly5-ol]-enkephalin), appeared to attenuate this negative incentive learning. These data suggest that upshifts and downshifts in endogenous opioid transmission in the BLA mediate the encoding of positive and negative shifts in incentive value, respectively, through actions at µ-opioid receptors, and provide insight into a mechanism through which opiates may elicit inappropriate desire resulting in their continued intake in the face of diminishing affective experience.

Oxidative and nitrative modifications of enkephalins by human neutrophils: effect of nitroenkephalin on leukocyte functional responses.

Neutrophils play a major role in acute inflammation by generating reactive oxygen/nitrogen species. Opioid peptides, including enkephalins, are present at inflammation sites. Neutrophils contribute to protect against inflammatory pain by releasing opioid peptides. In this investigation, the ability of human polymorphonuclear cells to induce oxidative and nitrative modifications of Leu-enkephalin has been investigated in vitro. Activated human neutrophils mediate the oxidation of Leu-enkephalin resulting in the production of dienkephalin. In the presence of nitrite at concentrations observed during inflammatory and infectious process (10-50 µM), nitroenkephalin, a nitrated derivative of Leu-enkephalin, is additionally formed. The yield of nitroenkephalin increases with nitrite concentration and is significantly inhibited by the addition of catalase or 4-aminobenzoic acid hydrazide (ABAH), a specific inhibitor of peroxidases. These results suggest that neutrophils induce nitration of Leu-enkephalin by a mechanism that is dependent on myeloperoxidase activity and hydrogen peroxide. Oxidative/nitrative modifications of Leu-enkephalin have been also evidenced when cells were treated with the NO-donor molecule, DEANO. The nitrated enkephalin has been examined for its effect on leukocyte functional responses. The data reveal that nitroenkephalin at micromolar concentrations inhibits superoxide anion generation and degranulation of azurophilic granules of human polymorphonuclear cells. Moreover, nitroenkephalin inhibits spontaneous apoptosis of neutrophils, as evaluated by measuring caspase-3 activity. Collectively, our data indicate that the nitrated enkephalin attenuates neutrophil activation and promotes the short-term survival of these cells, suggesting a possible role of the nitrocompound in the efficiency and resolution of inflammatory processes.

Effect of opioid peptides on the content of LPO products and antioxidant enzyme activity in the liver of rats after restraint stress.

Administration of opioid peptides dynorphin A (1-13) and DSLET was followed by a decrease in the stress-induced activation of LPO and increase in SOD activity in the liver tissue of rats. DAGO produced a similar, but less pronounced effect. The observed changes can be related to a specific distribution of opioid receptors in the liver tissue and stress-limiting influence of these peptides in the whole body.

Lipophilic derivatives of leu-enkephalinamide: in vitro permeability, stability and in vivo nasal delivery.

Leu-enkephalin is an endogenous pain modulating opioid pentapeptide. Its development as a potential pharmaceutic has been hampered by poor membrane permeability and susceptibility to enzymatic degradation. The addition of an unnatural amino acid containing a lipidic side chain at the N-terminus and the modification of the C-terminus to a carboxyamide was performed to enhance the nasal delivery of the peptide. Two lipidic derivatives with varying side chain lengths (C(8)-Enk-NH(2) (1), C(12)-Enk-NH(2) (2)) and their acetylated analogues were successfully synthesised. Caco-2 cell monolayer permeability and Caco-2 cell homogenate stability assays were performed. C(8)-Enk-NH(2) (1) and its acetylated analogue Ac-C8-Enk-NH(2) (3) exhibited apparent permeabilities (mean±SD) of 2.51±0.75×10(-6)cm/s and 1.06±0.62×10(-6), respectively. C12-Enk-NH(2) (2) exhibited an apparent permeability of 2.43±1.26×10(-6) cm/s while Ac-C12-Enk-NH(2) (4) was not permeable through the Caco-2 monolayers due to its poor solubility. All analogues exhibited improved Caco-2 homogenate stability compared to Leu-Enk-NH(2) with t(½) values of: C8-Enk-NH(2) (1): 31.7 min, C(12)-Enk-NH(2) (2): 14.7 min, Ac-C8-Enk-NH(2) (3): 83 min, Ac-C(12)-Enk-NH(2) (4): 27 min. However, plasma stability assays revealed that the diastereoisomers of C8-Enk-NH(2) (1) did not degrade at the same rate, with the l isomer (t(1/2)=8.9 min) degrading into Leu-enkephalinamide and then des-Tyr-Leu-Enk-NH(2), whereas the d isomer was stable (t(1/2)=120 min). In vivo nasal administration of C(8)-Enk-NH(2) to male rats resulted in concentrations of 5.9±1.84×10(-2) µM in the olfactory bulbs, 1.35±1.01×10(-2) µM in the brain and 6.53±1.87×10(-3) µM in the blood 10 min after administration.

Peptide synthesis 'in water' by a solution-phase method using water-dispersible nanoparticle Boc-amino acid.

Regulatory pressure has compelled the chemical manufacturing industry to reduce the use of organic solvents in synthetic chemistry, and there is currently a strong focus on replacing these solvents with water. Here, we describe an efficient in-water solution-phase peptide synthesis method using Boc-amino acids. It is based on a coupling reaction utilizing suspended water-dispersible nanoparticle reactants. Using this method, peptides were obtained in good yield and with high purity.

Beta-endorphin inhibits the inflammatory response of bovine endometrial cells through δ opioid receptor in vitro.

Postpartum uterine infections are common reproductive diseases in postpartum cows. Evidence has shown that plasma ß-endorphins increase during bovine uterine inflammation. However, the effect of ß-endorphins on the inflammatory response in bovine endometrium has not been clarified. The aim of this study was to investigate the effect of ß-endorphins on the inflammatory response of bovine endometrial epithelial and stromal cells, and to explore the possible mechanism. The cells were treated with E. coli lipopolysaccharide (LPS) to simulate inflammation, which was characterized by the significant activation of NF-κB signaling pathway and the increased gene expression of the downstream proinflammatory cytokines (approximately 1.2- to 15-fold increase, P < 0.05). By using Western blot and qPCR techniques, we found that ß-endorphins inhibited the key protein expression of NF-κB pathway, and the gene expressions of TNF, IL1B, IL6, CXCL8, nitric oxide synthase 2, and prostaglandin-endoperoxide synthase 2 (P < 0.05). The co-treatment of ß-endorphins and opioid antagonists showed that the anti-inflammatory effect of ß-endorphins could be blocked (P < 0.05) by non-selective opioid antagonist naloxone or δ opioid receptor antagonist ICI 154129, but not the µ opioid receptor antagonist CTAP (P > 0.05). In conclusion, ß-endorphins may inhibit the inflammatory response of bovine endometrial epithelial and stromal cells through δ opioid receptor.

Analogues of both Leu- and Met-enkephalin containing a constrained dipeptide isostere prepared from a Baylis-Hillman adduct.

An efficient route was developed for the synthesis of the Fmoc-protected dipeptide 4, isostere of Gly-Gly containing an alpha-methylene beta-amino acid; the conformationally restricted analogues of Leu-enkephalin, 3a, and Met-enkephalin, 3b, respectively, were prepared by changing 4 for Gly(2)-Gly(3) in the native compounds 3a and 3b whose biological activities were significantly lower than the parent compounds.

A bioinspired synthetic approach for building metal-organic frameworks with accessible metal centers.

We demonstrate how a bioinspired synthetic approach can help organic linkers distinguish between different types of metal centers in metal-organic frameworks (MOFs). Modification of an organic building unit with methyl groups enables the unit to selectively coordinate to one of the two metal sites present in the MOFs. We report four new porphyrin-based, pillared-paddlewheel frameworks: PPF-11-Zn/Zn, -Co/Co, -Mn/Zn, and -Fe/Zn, where the first and second metals indicate the metal center for the porphyrin core and paddlewheel cluster, respectively. These compounds exhibit 3D MOFs in which 2D layers are pillared by a sterically controlled bipyridine, leaving the metal centers inside the porphyrin structurally unconnected.

δ Opioid Receptor Inverse Agonists and their In Vivo Pharmacological Effects.

The discovery of δ opioid receptor inverse agonist activity induced by ICI-174,864, which was previously reported as an δ opioid receptor antagonist, opened the door for the investigation of inverse agonism/constitutive activity of the receptors. Various peptidic or non-peptidic δ opioid receptor inverse agonists have since been developed. Compared with the reports dealing with in vitro inverse agonist activities of novel compounds or known compounds as antagonists, there have been almost no publications describing the in vivo pharmacological effects induced by a δ opioid receptor inverse agonist. After the observation of anorectic effects with the δ opioid receptor antagonism was discussed in the early 2000s, the short-term memory improving effects and antitussive effects have been very recently reported as possible pharmacological effects induced by a δ opioid receptor inverse agonist. In this review, we will survey the developed δ opioid receptor inverse agonists and summarize the possible in vivo pharmacological effects by δ opioid receptor inverse agonists. Moreover, we will discuss important issues involved in the investigation of the in vivo pharmacological effects produced by a δ opioid receptor inverse agonist.

Modification with organometallic compounds improves crossing of the blood-brain barrier of [Leu5]-enkephalin derivatives in an in vitro model system.

Enkephalin peptides are thought to be suitable vectors for the passage of the blood-brain barrier (BBB). Modifications that do not alter the amino acid sequence are often used to improve the permeation through living membrane systems. As a new type of modification we introduce organometallic compounds, in particular ferrocene carboxylic acid. Derivatives of [Leu5]enkephalin were synthesised and labelled with organometallic compounds by using solid-phase synthesis techniques. All new metal-peptide bioconjugates were comprehensively characterised by HPLC, NMR spectroscopy and mass spectrometry and found to be at least 95% pure. For the first time, permeation coefficients in a BBB model for organometal-peptide derivatives were determined in this work. The uptake and localisation of fluorescein-labelled enkephalins was monitored by fluorescence microscopy on three cancer cell lines. Octanol/H2O partition coefficients of the compounds were measured by HPLC. The introduction of the organometallic moiety enhances the uptake into cells and the permeation coefficient of [Leu5]-enkephalin. This could be due to an increase in lipophilicity caused by the organometallic label. The metal-peptide conjugates were found to be nontoxic up to mM concentrations. The low cytotoxicity encourages further experiments that could take advantage of the selectivity of enkephalin derivatives for opioid receptors.

Endogenous anticonvulsant substance in rat cerebrospinal fluid after a generalized seizure.

Cerebrospinal fluid taken from rats subjected to electroshock-induced seizures and injected into the cerebral ventricles of rats that had not been shocked increased the seizure threshold of the recipients. The anticonvulsant activity of the donor cerebrospinal fluid was antagonized by opioid antagonists and enhanced by peptidase inhibitors. These results suggest the existence of an endogenous anticonvulsant substance in rat cerebrospinal fluid, possibly opioid in nature, which is activated as a consequence of a seizure and which may play a critical role in postseizure inhibition.

Central administration of somatostatin stimulates feeding behavior in chicks.

The purpose of the present study was to determine if central administration of somatostatin influences feeding behavior in layer chicks. Five- to 7-day-old chicks that received intracerebroventricular (ICV) injections of 0.5 or 2 nmol somatostatin increased their food intake at 30 and 60 min after the injection, suggesting that central somatostatin serves as an orexigenic neuropeptide in chicks. This hypothesis was further supported since chicks ICV injected with 0.5 or 2 nmol cortistatin, which binds to somatostatin receptors, also had increased food intake at the same time. Somatostatin-associated feeding behavior was attenuated by co-administration of 20 nmol beta-funaltrexamine (an opioidergic mu-receptor antagonist) (to 31% of the orexigenic effect of somatostatin at 60 min after the injection) but not ICI-174,864 or nor-binaltorphimine (antagonists of opioidergic delta- and kappa-receptors, respectively). Co-administration of 13 nmol yohimbine, an adrenergic alpha-2 receptor antagonist, also attenuated the orexigenic effect of somatostatin (to 31% of the orexigenic effect of somatostatin at 60 min after the injection). These results suggest that somatostatin-associated feeding behavior is mediated by opioidergic mu- and adrenergic alpha-2-receptors in chicks.

Immune response of submissive and aggressive mice under conditions of opioid receptor activation.

Experiments on the model of paired sensory contact demonstrated that stimulation of immune response in aggressive CBA and C57Bl/6J mice with 10- and 20-day experience of victories, respectively, was prevented by selective δ(2)- and -opioid receptor agonists DSLET and κ-opioid receptor agonists rimorphin in a dose of 100 microg/kg. In C57Bl/6J mice with depression-like behavior, normalization (but not suppression) of the immune response under conditions of µ-opioid receptor stimulation (100 microg/kg) was observed. Selective modulation of activity of a certain type of opioid receptors can normalize the immune function modified during the formation of a certain behavioral type.