Pain management during diagnostic office hysteroscopy in postmenopausal women: a randomized study.

Objective: This study aimed to compare efficacy and safety of lidocaine versus tramadol versus placebo in reducing the pain of diagnostic outpatient hysteroscopy (OH) in postmenopausal women.Materials and methods: This randomized double-blinded study included 156 menopausal women who received intrauterine lidocaine infusion or oral tramadol (50 mg) or placebo before diagnostic OH (52 women/group). Primary outcome was pain severity during the procedure using a 10-cm visual analog scale. Secondary outcomes were pain scores 10 and 30 min post procedure, satisfaction level, and ease of cervical entry.Results: Lidocaine had lower pain scores compared to placebo during and 10 min after the procedure (p < 0.001). Tramadol had lower pain scores than placebo during the procedure (p = 0.04), 10 min after the procedure (p = 0.03), and 30 min after the procedure (p = 0.04). Both lidocaine and tramadol resulted in an easier procedure than placebo (p < 0.001 and p = 0.04, respectively). Lidocaine had an easier cervical entry compared to tramadol (p = 0.004). Satisfaction scores in the lidocaine and tramadol groups were significantly higher than in the placebo group (p < 0.001).Conclusions: Lidocaine and tramadol were effective in reducing postmenopausal women-reported pain during and after diagnostic OH. However, lidocaine was better than tramadol in facilitating hysteroscope passage through the cervical canal and the reduction in pain levels with lidocaine was clinically relevant.Trial registration number: NCT03701984.

Dual-wavelength, mode-locked erbium-doped fiber laser employing a graphene/polymethyl-methacrylate saturable absorber.

Mode-locked fiber laser incorporating a saturable absorber is an attractive configuration due to its stability and simple structure. In this work, we demonstrate a dual-wavelength passively mode-locked erbium-doped fiber laser employing a graphene/polymethyl-methacrylate saturable absorber. A laser resonator is developed based on dual cavity architecture with unidirectional signal oscillation, which is connected by a fiber branch sharing a common gain medium and saturable absorber. Dual wavelength mode-locked fiber lasers are observed at approximately 1530 and 1560 nm with 22.6 mW pump power threshold. Soliton pulse circulates in the laser cavity with pulse duration of 900 and 940 fs at shorter and longer wavelengths, respectively. This work presents a viable option in developing a low threshold mode-locked laser source with closely spaced dual wavelength femtosecond pulses in the C-band wavelength region.

A new thin layer cell for battery related DEMS-experiments: the activity of redox mediators in the Li-O2 cell.

The reversibility of current Li-O2 batteries suffers from high charging overpotentials. To address this problem, the use of redox mediators has been proposed, which are supposed to improve the sluggish reaction kinetics of the oxygen evolution reaction via a solution mediated oxidation of lithium peroxide. In this study, we present a new thin layer cell for battery related differential electrochemical mass spectrometry (DEMS) experiments, which exhibits a high electrode surface area to electrolyte volume ratio which is closer to the situation in batteries other approaches/cells with their usually large electrolyte excess. The confined volume also allows a better distinction between the mediating activity of a redox system and a near continuous electrochemical reaction of this species. One further benefit of the new thin layer cell is that experiments can easily be performed under different O2-partial pressures. This new set-up allows the highly sensitive detection of volatile species formed during the OER. Therefore, small changes in the number of electrons transferred per oxygen molecule are observable. These changes help to identify side reactions and possible decomposition of the reaction products. During our experiments, we investigated the impact of TTF, TMPD, Fc and TEMPO on the oxidation of Li2O2. Within our experiments, we are able to precisely determine the potential at which the catalytic activity of the redox mediation starts. A comparison between the potential at which we observe the activity of the redox mediator to the half wave potential of the redox system could be explained with an outer sphere electron transfer for the oxidation of Li2O2 by a redox mediator. This observation is confirmed by a theoretical treatment of the redox mediation mechanism. Moreover, insights into the number of transferred electrons per oxygen molecule during the activity of the different redox mediators reveal the presence of side reactions. This finding is also underlined by an unexpected shift of the CO2 evolution onset for the redox mediator containing electrolytes. Our experiments also reveal that a Li-O2 cell, which contains a redox mediator, undergoes less fluctuation in its reversibility compared to a cell without a redox mediator.

Oxygen reduction and oxygen evolution in DMSO based electrolytes: the role of the electrocatalyst.

In the present paper the role of the electrode material in oxygen reduction in DMSO based electrolytes is elucidated using DEMS. We have found, employing platinum, gold, ruthenium rhodium, selenium decorated rhodium and boron doped diamond (BDD) as electrode materials, that the actual mechanism of oxygen reduction largely depends on the electrode material. At platinum, rhodium and selenium decorated rhodium the final reduction product, peroxide, is formed electrochemically. At gold and at low overpotentials oxygen is reduced to superoxide and peroxide is only formed by disproportionation of the latter. No oxygen reduction takes place at the diamond surface of the BDD-electrode, hence, showing unambiguously that oxygen reduction is an inner sphere reaction. Also, the rate of oxygen evolution varies with the electrode material, although the onset potential of oxygen evolution is not influenced. The amount of peroxide formed is limited to 1-2 monolayers. Contrary to intuition oxygen reduction and oxygen evolution from peroxide, therefore, are heterogeneous, electrocatalytic reactions. The finding of such an electrocatalytic effect is of great importance for the development and optimization of lithium-air batteries. Aside from the electrode material there are also effects of water as well as of the cation used in the electrolyte. This suggests an influence of the double layer at the interface between the electrode and the electrolyte on oxygen reduction in addition to the well-known higher stability of Na2O2 and K2O2. Electrospray ionization (ESI) results show that any effect of water in the Li(+) containing electrolyte is not due to an altered solvation of the cation.

Correction: Oxygen reduction and oxygen evolution in DMSO based electrolytes: the role of the electrocatalyst.

Correction for 'Oxygen reduction and oxygen evolution in DMSO based electrolytes: the role of the electrocatalyst' by C. J. Bondue et al., Phys. Chem. Chem. Phys., 2015, 17, 25593-25606.

A nucleic acid-based surface-enhanced Raman scattering of gold nanorods in N-gene integrated principal component analysis for COVID-19 detection.

A rapid, simple, sensitive, and selective point-of-care diagnosis tool kit is vital for detecting the coronavirus disease (COVID-19) based on the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strain. Currently, the reverse transcriptase-polymerase chain reaction (RT-PCR) is the best technique to detect the disease. Although a good sensitivity has been observed in RT-PCR, the isolation and screening process for high sample volume is limited due to the time-consuming and laborious work. This study introduced a nucleic acid-based surface-enhanced Raman scattering (SERS) sensor to detect the nucleocapsid gene (N-gene) of SARS-CoV-2. The Raman scattering signal was amplified using gold nanoparticles (AuNPs) possessing a rod-like morphology to improve the SERS effect, which was approximately 12-15 nm in diameter and 40-50 nm in length. These nanoparticles were functionalised with the single-stranded deoxyribonucleic acid (ssDNA) complemented with the N-gene. Furthermore, the study demonstrates method selectivity by strategically testing the same virus genome at different locations. This focused approach showcases the method's capability to discern specific genetic variations, ensuring accuracy in viral detection. A multivariate statistical analysis technique was then applied to analyse the raw SERS spectra data using the principal component analysis (PCA). An acceptable variance amount was demonstrated by the overall variance (82.4 %) for PC1 and PC2, which exceeded the desired value of 80 %. These results successfully revealed the hidden information in the raw SERS spectra data. The outcome suggested a more significant thymine base detection than other nitrogenous bases at wavenumbers 613, 779, 1219, 1345, and 1382 cm-1. Adenine was also less observed at 734 cm-1, and ssDNA-RNA hybridisations were presented in the ketone with amino base SERS bands in 1746, 1815, 1871, and 1971 cm-1 of the fingerprint. Overall, the N-gene could be detected as low as 0.1 nM within 10 mins of incubation time. This approach could be developed as an alternative point-of-care diagnosis tool kit to detect and monitor the COVID-19 disease.

Cyclic quantum teleportation of two-qubit entangled states by using six-qubit cluster state and six-qubit entangled state.

Cyclic quantum teleportation schemes requires at least the existence of three collaborators acting all as senders and receivers of quantum information, each one of them has an information to be transmitted to the next neighbour in a circular manner. Here, new cyclic quantum teleportation scheme is proposed for perfectly transmitting cyclically three arbitrary unknown two-qubit states ( α , ß and γ ) among the three collaborators. In this scheme, Alice can send to Bob the quantum information contained in her two-qubit state α and receive from Charlie the quantum information contained in the two-qubit state in his possession γ and similarly, Bob can transmit to Charlie the quantum information contained in his two-qubit state ß through a quantum channel of twelve-qubit state consisting of a six-qubit cluster state and a six-qubit entangled state by sequentially and cyclically performing Bell state measurements. Subsequently, each one of the three participants can afterwards retrieve his own desired two-qubit state using classical channel and by performing appropriate unitary Pauli operators and we have shown that our proposed scheme performs efficiently.

Development of inactivated heartwater (Ehrlichia ruminantium) vaccine in South Africa.

Heartwater, Ehrlichia ruminantium infection in cattle, sheep, goats, and some wild ruminants, is an economically important disease in Africa characterized by high mortality rates in susceptible populations. In South Africa, the current commercial heartwater vaccine is an infection and treatment type of immunization using virulent live E. ruminantium organisms generated from blood of infected sheep with subsequent treatment of the animals with antibiotics at specific times during the course of infection. This vaccine has several inherent problems preventing its wide use as the vaccine must be administered intravenously and it does not protect against all the South African field isolates. A vaccine based on inactivation of Zimbabwean E. ruminantium Mbizi strain organisms produced in endothelial cell cultures can be a sustainable option because it will not require antibiotic treatment and will be safe as there is no potential for reversion to virulence. Previous data generated in laboratory trials and under natural field setting provides support for this vaccine approach. Four inactivated vaccine formulations using the E. ruminantium Mbizi strain were tested for their efficacy in Merino sheep compared to an unvaccinated control group (11 sheep per group). Two vaccines were prepared by beta-propiolactone (BPL) inactivation, and two were inactivated with binary ethylenimine (BEI) while purification was done with both percoll and polyethylene glycol (PEG). The four vaccine preparations were formulated with Montanide ISA 50V2 adjuvant and administered twice subcutaneously (2 ml per dose) at an interval of 4 weeks. All groups were challenged with a virulent homologous cell-cultured E. ruminantium inoculated via the intra-venous route on day 56. The primary variable of efficacy was measured by the percentage survival rate or mortality between the Controls and Vaccine Groups. Three vaccine formulations (BEI/Percoll (Group 3), BEI/PEG (Group 4), BPL/Percoll, (Group 1) had a significantly higher percent of animal surviving challenge compared to the unvaccinated control (p-values 0.001, 0.035, 0.030, respectively). The highest number of survivors was obtained in Group 3 BEI/Percoll; 10/11 (91%). Groups 4 (BEI/PEG) and Group 1 (BPL/Percoll) produced similar percentage of survivals of 64%. In contrast, the lowest survival rate of 50% was observed in Group 2 (BPL/PEG) which was numerically different but not significantly different from the unvaccinated control which had an 18% survival rate (2/11). The inactivated vaccine using BEI or BPL as inactivating agents blended with ISA 50 adjuvant induced protective immunity against challenge. The BEI/Percoll (Group 3) vaccination regimen was most efficacious against a lethal heartwater challenge as it significantly protected sheep against mortality which is the most important aspect of heartwater infections. Future work should be directed towards improvement of this vaccine formulation especially from the down-stream processing point of view as the percoll method is not scalable for commercialization purposes.

MicroPulse® transscleral laser therapy in the management of glaucoma patients.

PURPOSE: To evaluate the efficacy and safety of MicroPulse® transscleral laser therapy (TLT) in the management of glaucoma patients. METHODS: A prospective, interventional, non-comparative case series was conducted in the department of ophthalmology, Ain Shams University Hospital on 61 eyes of 46 patients with various glaucoma types and of severity, ranging from mild to severe. In addition to best-corrected visual acuity (BCVA), intraocular pressure (IOP) and the number of glaucoma medications were recorded before and after treatment, along with the postoperative need for systemic carbonic anhydrase inhibitors (CAI), success rates, number of treatment sessions and postoperative complications. Success was defined as an IOP of 6-18 mmHg or at least a 30% decrease from preoperative IOP in the absence of any vision-threatening complications during the 6-month follow-up period. RESULTS: MicroPulse TLT was performed on 61 glaucomatous eyes. Eleven of the 61 eyes (18%) that did not achieve IOP between 6-18 mmHg, or at least a 30% decrease from baseline at 6 months, had a repeat MicroPulse TLT session. At 6 months follow-up post a single MicroPulse TLT session, the mean IOP reduction was 35.9 ± 14.2%; and 6 months after the second session, it was 36.2 ± 17.5% (p < 0.001). The success rate after the first session was 73.8% which increased to 78.7% after the second session. The mean anterior chamber (AC) cell reaction was +1.9 ± 0.8 at 1 day, +1.0 ± 0.7 at 1 week, and +0.2 ± 0.4 at 1 month postoperatively. No cells were detected in any of the cases at 3 and 6 months follow-up (p < 0.001). The average number of anti-glaucoma eye drops before MicroPulse TLT was 2.6 ± 1.0. Postintervention, the average number of anti-glaucoma eye drops was 1.7 ± 1.2, and sustained at 6 months follow-up after the last treatment session (p < 0.001). There were no significant complications were noted. One eye developed transient hypotony for 3 months after MicroPulse TLT. CONCLUSIONS: MicroPulse TLT is safe and effective in lowering IOP in a variety of glaucoma types and severity.

Lidocaine vs. magnesium: effect on analgesia after a laparoscopic cholecystectomy.

BACKGROUND: This double-blinded study aimed at evaluating and comparing the effects of magnesium and lidocaine on pain, analgesic requirements, bowel function, and quality of sleep in patients undergoing a laparoscopic cholecystectomy (LC). METHODS: Patients were randomized into three groups (n=40 each). Group M received magnesium sulfate 50 mg/kg intravenously (i.v.), followed by 25 mg/kg/h i.v., group L received lidocaine 2 mg/kg i.v., followed by 2 mg/kg/h i.v., and group P received saline i.v. Bolus doses were given over 15 min before induction of anesthesia, followed by an i.v. infusion through the end of surgery. Intraoperative fentanyl consumption and averaged end-tidal sevoflurane concentration were recorded. Abdominal and shoulder pain were evaluated up to 24 h using a visual analog scale (VAS). Morphine consumption was recorded at 2 and 24 h, together with quality of sleep and time of first flatus. RESULTS: Lidocaine or magnesium reduced anesthetic requirements (P<0.01), pain scores (P<0.05), and morphine consumption (P<0.001) relative to the control group. Lidocaine resulted in lower morphine consumption at 2 h [4.9 + or - 2.3 vs. 6.8 + or - 2.8 (P<0.05)] and lower abdominal VAS scores compared with magnesium (1.8 + or - 0.8 vs. 3.2 + or - 0.9, 2.2 + or - 1 vs. 3.6 + or - 1.6, and 2.1 + or - 1.4 vs. 3.3 + or - 1.9) at 2, 6, and 12 h, respectively (P<0.05). Lidocaine was associated with earlier return of bowel function and magnesium was associated with better sleep quality (P<0.05). CONCLUSION: I.v. lidocaine and magnesium improved post-operative analgesia and reduced intraoperative and post-operative opioid requirements in patients undergoing LC. The improvement of quality of recovery might facilitate rapid hospital discharge.

Neurobehavioral and Menstrual Disorders among Adolescent Females Environmentally Exposed to Pesticides, Menoufia Governorate, Egypt.

BACKGROUND: Adolescent females living in agricultural areas where crops are routinely sprayed by pesticides are expected to be environmentally exposed to pesticides' health hazards partially as those occupationally exposed. OBJECTIVE: to assess menstrual and neurobehavioral disorders among adolescent females environmentally exposed to pesticides. METHODS: This was a cross-sectional study conducted on 100 pesticide exposed adolescent females who had one or more of family members are pesticides' seasonal applicators and 50 non- exposed adolescent females matched for age and education, served as controls at Menoufia governorate, Egypt during the period of pesticide application season of cotton crop from the first days of May to the end of September 2017. A self-administered and a series of neurobehavioral tests were administered and serum Acetylcholinesterase (AChE) activity was assessed. RESULTS: A significant lower AChE activity levels were found in the exposed group than controls (Mean±SD=238.49± 23.83 vs 303.35±78.54 IU/L; respectively). There were significant higher mean scores of trail making test (parts 1 and 2) and significant lower mean scores of (similarities test, Benton visual retention test, block design test, Santa Ana dexterity test (dominant and non-dominant hands) and Beery visuo-motor imitation test in the exposed group than the controls (P<0.05). Also, the exposed group reported more prevalent irregular menstrual cycle (26.8%) and intermenstrual bleeding (28.2%) compared to the control participants (8.1% and 8.1%; respectively). CONCLUSION: Adolescent females living in agricultural areas and from families whose one or more members are pesticides' applicators have significantly lower neurobehavioral performance, report more prevalent menstrual irregularities and have lower levels of serum AChE compared to a control group. The neurobehavioral deficits demonstrated a dose-response relationship AChE levels in the exposed participants. This necessitates the need for implementation of health education programs to prevent or reduce health effects associated with pesticide exposure to adolescent females.

Ixodid ticks infesting domestic goats in communal land areas of Zimbabwe.

The purpose of this study was to determine the species spectrum of ticks infesting goats owned by resource-limited farmers in the state-owned communal land areas of Zimbabwe. Ticks were collected from goats at a single locality within each of 5 communal land areas, and a total of 14 ixodid tick species was recovered. The most numerous tick was Rhipicephalus evertsi evertsi, which was present in all areas at each sampling, and a Rhipicephalus sp. (near R. punctatus), which was most abundant on goats in the central regions of Zimbabwe during the March rainy season. Amblyomma hebraeum was present on goats in all areas sampled. In the eastern central region its distribution overlapped that of Amblyomma variegatum, while in the northwest it overlapped those of both Amblyomma marmoreum and A. variegatum. Hyalomma truncatum was present at all localities, whereas only a single Hyalomma rufipes was recovered. Rhipicephalus appendiculatus was collected from goats in the moist, slightly cooler regions, while the few Rhipicephalus zambeziensis recovered were present in the hotter, drier regions. Species recorded in lower numbers were Rhipicephalus (Boophilus) decoloratus, Rhipicephalus lunulatus, Rhipicephalus simus, Rhipicephalus tricuspis and Rhipicephalus turanicus. Attachment in the inter-digital space of adult A. hebreaum and H. truncatum was sometimes associated with lameness.

Occurrence of Theileria parva infection in cattle on a farm in the Ladysmith district, KwaZulu-Natal, South Africa.

Theileria parva causes widespread morbidity and mortality in cattle in endemic regions. An outbreak of theileriosis occurred on a farm near Ladysmith in KwaZulu-Natal, South Africa, which is not a declared Corridor disease-infected area. A survey of Red Brangus cattle from all age groups and areas of the farm was performed. Transmission of the parasite from infected animals on the farm to susceptible animals by tick transmission and tick-stabilate injection, was attempted. The survey indicated high numbers of animals with antibody titres to T. parva but only 6 infected animals, based on real-time PCR and RLB analysis. The transmission experiments failed to transmit the parasite. The study shows the difficulty in elucidating a source of infection and determining the dynamics of new infections in a herd where multiple possible sources are present and treatment with tetracyclines has taken place.

Efficacy of South African Babesia bovis vaccine against field isolates.

A high-passage Babesia bovis vaccine containing only one genotype population was, although protective, inferior compared to the immunity afforded by a lower passage of the same strain containing two populations. The 24 times serially passaged South African B. bovis S vaccine strain contain only a single parasite population (Bv80 allele A 558bp). Forty-four field isolates sampled were all found different with regard to the number and composition of the parasite populations present in each isolate. The extensive genotypic diversity in South Africa and the limited genotypic diversity observed in the S24 vaccine, raised the question on its ability to protect against such diverse populations. The 6 isolates selected for challenge in the current study originated from geographically distinct populations that also possessed thirteen unique genotypes based on the Bv80 gene and included strains that resulted in clinical disease. The strain coverage was therefore much greater than in previous studies on the protective ability of the S24 vaccine. Challenge of vaccinated cattle indicated that the vaccine gave adequate protection against 5/6 isolates. Protection against the remaining isolate proved inadequate. However, field observations in the region where this isolate originated from, showed only minor mortalities in vaccinated animals compared to losses experienced in unvaccinated herds. This study demonstrated the ability of the South African B. bovis S24 vaccine to protect cattle against challenge from local field isolates containing single or multiple parasite populations.

Sodium ion and the neutrotransmitter-stimulated 32P labelling of phosphoinositides and other phospholipids in the iris muscle.

The effects of Na+, other cations and the neurotransmitters, acetylcholine and norepinephrine on 32Pi incorporation into phospholipids of the rabbit iris smooth muscle were investigated [1]. The basal 32P-labelling of phospholipids including phosphatidic acid, phosphatidylinositol, phosphatidylcholine, phosphatidylethanolamine and the polyphosphoinositides increased with Na+ concentration [2]. The neurotransmitter-stimulated 32P labelling of phosphatidic acid, phosphatidylinositol and phosphatidylcholine is dependent on the presence of extracellular Na+ [3]. The monovalent cation requirement for Na+ specific. Of the monovalent cations Li+, NH+4, K+, Choline+ and Tris, only Li+ partially substituted for Na+ [4]. A significant decrease in 32P labelling of phospholipids in response to acetylcholine was observed when Ca2+ and/or K+ were added to an isoosmotic medium deficient of Na+ [5]. Ouabain, which blocks the Na+-pump, inhibited the basal 32Pi incorporation into phosphatidylcholine and the acetylcholine-stimulated 32P labelling of phosphatidic acid, phosphatidylinositol and phosphatidylcholine [6]. It was suggested that phosphoinositide breakdown is associated with Ca2+ influx as we have previously reported (Akhtar, R.A. and Abdel-Latif, A.A. (1978) J. Pharmacol. Exp. Ther. 204, 655-668) and that the enhanced 32P-labelling of phosphoinositides could be associated with Na+ outflux, via the Na+-pump mechanism.

Role of protein kinase C alpha in endothelin-1 stimulation of cytosolic phospholipase A2 and arachidonic acid release in cultured cat iris sphincter smooth muscle cells.

We have investigated the role and mechanism of protein kinase C (PKC) isoforms in endothelin-1 (ET-1)-induced arachidonic acid (AA) release in cat iris sphincter smooth muscle (CISM) cells. ET-1 increased AA release in a concentration (EC50=8 nM) and time-dependent (t1/2=1.2 min) manner. Cytosolic phospholipase A2 (cPLA2), but not phospholipase C (PLC), is involved in the liberation of AA in the stimulated cells. This conclusion is supported by the findings that ET-1-induced AA release is inhibited by AACOCF3, quinacrine and manoalide, PLA2 inhibitors, but not by U-73122, a PLC inhibitor, or by RHC-80267, a diacylglycerol lipase inhibitor. A role for PKC in ET-1-induced AA release is supported by the findings that the phorbol ester, PDBu, increased AA release by 96%, that prolonged treatment of the cells with PDBu resulted in the selective down regulation of PKCalpha and the complete inhibition of ET-1-induced AA release, and that pretreatment of the cells with staurosporine or RO 31-8220, PKC inhibitors, blocked the ET-1-induced AA release. Gö-6976, a compound that inhibits PKCalpha and beta specifically, blocked ET-1-induced AA release in a concentration-dependent manner with an IC50 value of 8 nM. Thymeatoxin (0.1 microM), a specific activator of PKCalpha, beta, and gamma induced a 150% increase in AA release. Treatment of the cells with ET-1 caused significant translocation of PKCalpha, but not PKCbeta, from cytosol to the particulate fraction. These results suggest that PKCalpha plays a critical role in ET-1-induced AA release in these cells. Immunochemical analysis revealed the presence of cPLA2, p42mapk and p44mapk in the CISM cells. The data presented are consistent with a role for PKCalpha, but not for p42/p44 mitogen-activated protein kinase (MAPK), in cPLA2 activation and AA release in ET-1-stimulated CISM cells since: (i) the PKC inhibitor, RO 31-8220, inhibited ET-1-induced AA release, cPLA2 phosphorylation and cPLA2 activity, but had no inhibitory effect on p42/p44 MAPK activation, (ii) genistein, a tyrosine kinase inhibitor, inhibited ET-1-stimulated MAPK activity but had no inhibitory effect on AA release in the ET-1-stimulated cells. We conclude that in CISM cells, ET-1 activates PKCalpha, which activates cPLA2, which liberates AA for prostaglandin synthesis.

Studies on the incorporation of [1-14C]arachidonic acid into glycerolipids and its conversion into prostaglandins by rabbit iris. Effects of anti-inflammatory drugs and phospholipase A2 inhibitors.

The effects of the anti-inflammatory drugs, indomethacin and aspirin, and the phospholipase A2 inhibitors, p-bromophenacyl bromide and mepacrine, on the in vitro metabolism of [1-14C]arachidonic acid by rabbit iris smooth muscle and iris microsomes were investigated. The incorporation of arachidonate into glycerolipids and its conversion into prostaglandins were rapid and time-dependent. About 65% of the total radioactivity was recovered in triacylglycerol, followed by that in phosphatidylcholine (20%), diacylglycerol (6%), phosphatidylethanolamine (5%) and phosphatidylinositol (3%), respectively. Time-course studies on arachidonate release from glycerolipids of prelabelled tissue showed that triacylglycerol, phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol are the major source for arachidonate in prostaglandin synthesis in this tissue. Arachidonate release from glycerolipids was not blocked by indomethacin and the effects of the phospholipase A2 inhibitors were nonspecific. p-Bromophenacyl bromide inhibited the labelling of glycerolipids in a dose-dependent manner. Mepacrine stimulated the labelling of phosphatidic acid, phosphatidylinositol and diacylglycerol, and inhibited that of phosphatidylcholine, phosphatidylethanolamine and triacylglycerol. At concentrations under 0.25 mM it stimulated prostaglandin synthesis in microsomes and at concentrations over 0.25 mM it inhibited their synthesis in both muscle and microsomes. Indomethacin and aspirin moderately increased the labelling of glycerolipids; however, both drugs inhibited prostaglandin synthesis by iris and iris microsomes in a dose-dependent manner. Possible explanations for mechanisms underlying these effects were presented. It is concluded that the phospholipase A2 inhibitors and the anti-inflammatory drugs exert profound effects on the incorporation of [1-14C]arachidonate into glycerolipids of the rabbit iris and on its conversion into prostaglandins by both iris and iris microsomes.

Studies on the properties of triphosphoinositide phosphomonoesterase and phosphodiesterase of rabbit iris smooth muscle.

The rabbit iris smooth muscle has been shown to contain triphosphoinositide phosphomonoesterase (phosphatidyl-myo-inositol-4,5-bisphosphate phosphohydrolase, EC 3.1.3.36) and phosphodiesterase (triphosphoinositide inositoltrisphosphohydrolase, EC 3.1.4.11) activities. Under our experimental conditions about 77% of the phosphomonoesterase and 61% of the phosphodiesterase activities were localized in the particulate fraction. The kinetic properties of the enzymes in the microsomal fraction were examined. The enzyme preparation was specific to polyphosphoinositides; it did not attack phosphatidylinositol under the present assay condition. The effects of Ca2+ and Mg2+ were also studied. Although the microsomal enzymes did not require added divalent cations for their activities, both the phosphomonoesterase and phosphodiesterase were appreciably inhibited by 1 mM EDTA. Phosphodiesterase and phosphomonoesterase were stimulated by Ca2+ and Mg2+, respectively. The demonstration of triphosphoinositide phosphodiesterase in the iris muscle, coupled with the findings that this enzyme is activated by Ca2+ and is not influenced by acetylcholine add further support to our previous conclusion (J. Pharmacol. Exp. Ther. (1978) 204, 655--668; J. Neurochem. (1978) 30, 517--525) that an increased Ca2+ influx, following the interaction between the neurotransmitter and its receptor, could act to stimulate the phosphodiesterase, thus leading to increased triphosphoinositide breakdown and increased phosphatidic acid via increased diacylglycerol.

Studies on the properties of a soluble phosphatidylinositol-phosphodiesterase of rabbit iris smooth muscle.

Some properties of the soluble phosphatidylinositol phosphodiesterase (monophosphatidylinositol inositolphosphohydrolase, EC 3.1.4.10) of rabbit iris smooth muscle are described. Studies on its subcellular distribution showed that in this tissue the phosphodiesterase is not exclusively cytosolic. Thus, under our experimental conditions about 58% of the enzyme activity was found in the soluble fraction and the remainder was particulate. When the latter was treated with deoxycholate about 59% of the enzyme activity, compared to 86% of that of ATPase, was still bound to the particulate fraction. The kinetic properties of the enzyme (30--50% (NH4)2SO4 fraction) were examined. Maximum breakdown was 7.7 mumol/h per mg protein and occurred at pH 5.6. The products of [14C]arachidonic acid-labelled phosphatidylinositol were 1,2-diacylglycerol and a mixture of 86% myoinositol 1-phosphate and 14% myoinositol 1,2-(cyclic)phosphate. The enzyme has an absolute requirement for Ca2+. Addition of Ba2+, La3+, Mg2+, Mn2+, EGTA or EDTA at 0.05--5 mM concentrations; Sr2+ at higher concentrations (greater than 0.25 mM) markedly inhibited the phosphodiesterase activity and this inhibition was completely reversed by Ca2+. The enzyme is specific for the phosphoinositides.

Studies on the properties of myo-inositol-1,4,5-trisphosphate 5-phosphatase and myo-inositol monophosphatase in bovine iris sphincter smooth muscle: effects of okadaic acid and protein phosphorylation.

In bovine iris sphincter, myo-inositol 1,4,5-trisphosphate (IP3) 5-phosphatase and myo-inositol 1-phosphate (IP1) monophosphatase are mainly localized in the microsomal and soluble fractions, respectively. Studies on the properties of these enzymes can be summarized as follows. (1) The microsomal IP3 5-phosphatase hydrolyzed IP3 to myo-inositol 1,4-bisphosphate with an apparent Km of 28 microM and Vmax of 32 nmol/min per mg protein. The IP1 monophosphatase in the soluble fraction hydrolyzed IP1 into free inositol with an apparent Km of 89 microM and Vmax of 7 nmol/min per mg protein. (2) IP3 5-phosphatase and IP1 monophosphatase had optimal pH values at 8.0 and 7.0, respectively. (3) Both enzymes required Mg2+ and their highest specific activities were at a cation concentration of 2 mM. (4) Ca2+ (> 0.5 microM) exerted an inhibitory effect on IP3 5-phosphatase activity, and marked inhibition (47%) was observed at a concentration of 10 microM. Higher concentrations of the cation (> 100 microM) were required to inhibit IP1 monophosphatase. (5) IP1 monophosphatase, but not IP3 5-phosphatase, was inhibited by Li+. Li+ had no effect on the contractile response in this smooth muscle. (6) Both enzymes were inhibited by ATP and by the thiol-blocking agent, disulfiram. In addition, thimerosal, a thiol reagent, also inhibited the IP3 5-phosphatase activity. (7) Protein phosphorylation of the microsomal and soluble fractions with PKA or PKC had no effect on the activities of these enzymes. (8) Okadaic acid, a protein phosphatase inhibitor, had no effect on the activity of IP3 5-phosphatase. However, in the intact iris sphincter the toxin significantly reduced the carbachol-induced IP3 production, 1,2-diacylglycerol formation, measured as phosphatidic acid, and caused muscle relaxation.