Regulation of an important glycolytic enzyme, pyruvate kinase, through phosphorylation in the larvae of a species of freeze-tolerant insect, Eurosta solidaginis.

Larvae of the goldenrod gall fly, Eurosta solidaginis, rely on a freeze tolerance strategy to survive the sub-zero temperatures of Canadian winter. Critical to their survival is the accumulation of polyol cryoprotectants and global metabolic rate depression, both of which require the regulation of glycolysis and reorganization of carbohydrate metabolism. This study explored the role that pyruvate kinase (PK) regulation plays in this metabolic reorganization. PK was purified from control (5 °C-acclimated) and frozen (-15 °C-acclimated) larvae and enzyme kinetic properties, structural stability, and post-translational modifications were examined in both enzyme forms. The Km phosphoenolpyruvate (PEP) of frozen PK was 20% higher than that of control PK, whereas the Vmax of frozen PK was up to 50% lower than that of control PK at the lowest assay temperature, suggesting inhibition of the enzyme during the winter. Additionally, the activity and substrate affinity of both forms of PK decreased significantly at low assay temperatures, and both forms were regulated allosterically by a number of metabolites. Pro-Q™ Diamond phosphoprotein staining and immunoblotting experiments demonstrated significantly higher threonine phosphorylation of PK from frozen animals while acetylation and methylation levels remained constant. Together, these results indicate that PK exists in two structurally distinct forms in E. solidaginis. In response to conditions mimicking the transition to winter, PK appears to be regulated to support metabolic rate depression, the accumulation of polyol cryoprotectants, and the need for extended periods of anaerobic carbohydrate metabolism to allow the animal to survive whole-body freezing.

[Which place of antiphosphatidylethanolamine antibodies research in seronegative antiphospholipid syndrome suspicion?] / Quelle place pour la recherche des anticorps antiphosphatidyléthanolamine dans la suspicion du syndrome des antiphospholipides séronégatif ?

PURPOSE: Antiphospholipid syndrome (APS) is a clinico-biological syndrome, which associates vascular injury and persisting antiphospholipid antibodies (aPL). Patients with clinical symptoms of APS but without aPL are defined as "seronegative APS" (SNAPS). The aim of this study was to evaluate antiphosphatidylethanolamine antibody (aPE) investigation in patients with SNAPS suspicion. METHODS: This retrospective study was conducted in patients with SNAPS suspicion. A homemade enzyme-linked immunosorbent assay (ELISA) was used to search for aPE. The results of this homemade method were compared with those from a global screening ELISA. RESULTS: Two hundred twenty-eight patients with SNAPS suspicion were included. Among them, 58.3% had a thrombotic event. The homemade ELISA found positive persisting aPE in 23 patients (10%): 15 with a thrombotic event, 6 with obstetrical morbidity and 2 with a combined event. The global screening ELISA was positive in only 11 of these 23 patients (47.8%). CONCLUSION: These results suggest the implication of aPE in SNAPS.

Implantable electrophysiologic cardiac device infections: a risk factor analysis.

The risk of cardiac device infection (CDI) is rising significantly, with several risk factors identified. The purpose of this study is to determine the rate of CDI at our center and to assess the associated risk factors, in order to define appropriate measures to prevent this complication. We retrospectively reviewed all cases of patients with CDI at St. George Hospital between February 1999 and July 2010. Each case was matched with three controls. We performed a descriptive and bivariate analysis to identify significant risk factors. Eighteen case patients and 54 control subjects met the inclusion criteria. An organism was recovered in 58 % of the cases. Significant risk factors included previous history of CDI (p < 0.001), recent manipulation (p < 0.001), trauma to the site of implant (p = 0.003), having a dual chamber/dual lead pacemaker (p = 0.002), and development of post-procedural hematoma (p = 0.012). Our findings complement the results of previous studies. We recommend a pre-procedural risk assessment and a thorough post-implantation follow-up to prevent the development of infective complications.

Combined experimental and theoretical study on hydrogen-bonded complexes between cyclic ketones, lactones, and lactams with 3,4-dinitrophenol.

The interaction of 3,4 dinitrophenol (DNP) with cyclic ketones, lactones, and lactams was investigated by UV-visible spectroscopy and density functional theory (DFT) methods. Equilibrium constants K(HB) for 1:1 hydrogen bonded complexes were determined in solution in CCl(4) and C(6)H(12). For the entire range of studied bases, the pK(HB) scale, varying between 2.92 for beta-propiolactone to 5.53 for 1-methyl-epsilon-caprolactam, indicates that the basicity increases with the ring size. Geometries, energies, and vibrational characteristics of complexes were obtained by means of DFT calculations. For lactones and lactams, the energy difference between the two most stable conformers, cis and trans, with respect to the ring oxygen (nitrogen) atom, is relatively small, suggesting that the complex observed in solution is probably an equilibrium mixture of both forms. The good correlation between Gibbs free energies in solution and in the gas phase, computed at the B3LYP/6-311++G(3df,2p) level of theory, confirms the reliability of our results. The electron density of the complexes has been analyzed by means of the atoms in molecules (AIM) theory and the natural bond orbital (NBO) method have been used to characterize the orbital interactions. Our theoretical survey shows that the 1:1 complexes are stabilized by a network of conventional and/or nonconventional intermolecular hydrogen bonds.

Thermochemistry and gas-phase ion energetics of 2-hydroxy-4-methoxy-benzophenone (oxybenzone).

We have investigated the thermochemistry and ion energetics of the oxybenzone (2-hydroxy-4-methoxy-benzophenone, C14H12O3, 1H) molecule. The following parameters have been determined for this species: gas-phase enthalpy for the of neutral molecule at 298.15K, (Delta(f)H0(m)(g) = -303.5 +/- 5.1 kJ x mol-1), the intrinsic (gas-phase) acidity (GA(1H) = 1402.1 +/- 8.4 kJ x mol-1), enthalpy of formation for the oxybenzone anion (Delta(f)H0(m)(1-,g) = -402.3 +/- 9.8 kJ x mol-1). We also have obtained the enthalpy of formation of, 4-hydroxy-4'-methoxybenzophenone (Delta(f)H0(m)(g) = -275.4 +/- 10 kJ x mol-1) and 3-methoxyphenol anion (Delta(f)H0(m)(C7H7O2-,g) = -317.7 +/- 8.7 kJ x mol-1). A reliable experimental estimation of enthalpy related to intramolecular hydrogen bonding in oxybenzone has also been obtained (30.1 +/- 6.3 kJ x mol-1) and compared with our theoretical calculations at the B3LYP/6-311++G** level of theory, by means of an isodesmic reaction scheme. In addition, heat capacities, temperature, and enthalpy of fusion have been determined for this molecule by differential scanning calorimetry.

Use of octreotide, a long-acting somatostatin analogue, in the treatment of chylothorax of idiopathic etiology.

We describe a case of chylothorax of idiopathic etiology, treated with octreotide, a long-acting somatostatin analogue. A 30-year-old man with a left supraclavicular mass, and chylothorax, initially diagnosed by outpatient thoracentesis, underwent diagnostic surgery to ascertain the etiology of the pleural effusion. Biopsies of the left supraclavicular mass, lymphatic tissue and lymph nodes were benign. Triglyceride level in the pleural fluid was 396 mg/dL, diagnostic of chylothorax. Treatment included intravenous total parenteral nutrition (TPN) and a nil per os (NPO) diet. Subsequent surgical interventions included left lung decortication and glue-mediated control of chylothorax, combined with TPN and a strict low-fat diet. Given the persistency of chylothorax, thoracic duct ligation was also performed, and octreotide subcutaneous injections were started, with dramatic resolution of pleural effusion, after 1 week of treatment, in absence of any side effects. The patient fully recovered, and no relapse has been observed during a follow-up period of over 1 year. In conclusion, octreotide showed to be a valid and safe noninvasive approach for the treatment of chylothorax, whose early clinical use may also reduce the need for surgical intervention.

Metabolic and hemodynamic features of elderly patients undergoing beating heart coronary artery bypass graft surgery.

AIM: Coronary artery bypass graft (CABG) surgery, nowadays, is increasingly performed in patients who are older and have more comorbidities than subjects operated on a decade ago. In this study, we investigated metabolic and hemodynamic features of elderly patients with single vessel coronary artery disease (CAD), undergoing beating heart coronary artery bypass graft (BHCABG) surgery. METHODS: Twenty-five elderly patients, ages 73-78 years, with isolated left anterior descending artery (LAD) disease, were enrolled and compared to a younger similar group of 25 patients, mean age 48+/-1.2 years. A single vessel left internal mammary artery (LIMA) to LAD BHCABG was performed in all patients. Duration of temporary LAD occlusion was 9.8+/-0.5 min in the elderly group, and 10+/-0.4 min in the younger group. Myocardial arterial-venous differences in glucose, lactate, and creatine phosphokinase (CPK) were performed at different time points: preoperatively in the operating room (T0); at the end of the grafting procedure (T1); and before closing the chest (T2). Left ventricular stroke work index (LVSWI), as an indicator of global function of left ventricle, were recorded at T0, T1, T2, 6 (T3) and 48 (T4) hours postoperatively. RESULTS: Preoperative glucose extraction, observed in both groups, did not augment during and after surgery. In addition, neither lactate nor CPK were released in the coronary sinus during temporary LAD occlusion and following reperfusion in either group. Similarly, no significant changes in LVSWI were observed intra- and perioperatively between the two groups. CONCLUSIONS: Cardiac metabolism, hemodynamic parameters and global left ventricular function were not affected in either group by brief LAD occlusion during BHCABG, suggesting that BHCABG is a well-tolerated surgical approach, which can be safely attempted in patients of any age.

Glenoid Dysplasia: Radiographic, Direct MR Arthrographic and Arthroscopic Appearances.

Glenoid dysplasia is an uncommon developmental abnormality of the scapula that is frequently overlooked. We report a case of severe glenoid dysplasia in a 55 year old man and demonstrate its radiographic, direct MR arthrographic and arthroscopic appearances.

Density functional theory study of the hydrogen bond interaction between lactones, lactams, and methanol.

The structure and relative stability of methanol complexes with various cyclic ketones, lactones, lactams, and N-methyl lactams from three- to seven-membered rings have been investigated using the density functional theory method. The geometries, harmonic frequencies, and energies were calculated at the B3LYP/6-311+G(d,p) level. Three stable structures, cis-a, cis-b, and trans, with respect to the ring oxygen (nitrogen) atom, were found to be local minima of the potential energy surface. For lactones and N-methyl lactams, the most stable structure is trans; it is stabilized, as in cyclic ketones, through the conventional hydrogen bond (HB) interaction between the basic carbonyl oxygen and the acidic methanolic hydrogen and an unconventional HB interaction between the methanolic oxygen and the CH hydrogen, in the alpha position of the carbonyl group. For unsubstituted lactams, the cis-a structure, stabilized through a HB interaction between the NH group and the methanol oxygen in addition to the conventional HB interaction, is the most stable. The topological properties of the electron density ratify the existence of conventional (N,O-H. . .O) and unconventional (C-H. . .O) hydrogen bonding. A good correlation was found between the HB distances and the electron density at the HB critical point. The unsubstituted lactams yield more stable complexes with methanol than N-methyl lactams, lactones, and cyclic ketones. In the most stable complexes, both components behave simultaneously as a HB donor and as a HB acceptor.

CT and MR findings of a retrorectal cystic hamartoma confused with an adnexal mass on ultrasound.

We describe the imaging features of a tailgut cyst mistaken for an adnexal mass. A pelvic ultrasound in a 28-year-old woman showed a 10-cm hypoechoic left pelvic mass. Having not seen the left ovary, the radiologist concluded that the mass was an endometrioma. CT disclosed a retrorectal cystic lesion with wall calcifications and internal septa. MR confirmed the extra-ovarian location of the tumor, which was hyperintense on T2-weighted images and had an intermediate signal on T1-weighted images. Surgery revealed a retrorectal cystic hamartoma. Radiological diagnosis of a tailgut cyst requires first correct localization of the tumor and then differentiation from other retrorectal masses.

Cholesterol-dependent association of caveolin-1 with the transducin alpha subunit in bovine photoreceptor rod outer segments: disruption by cyclodextrin and guanosine 5'-O-(3-thiotriphosphate).

Evidence suggests that caveolins, 21-24 kDa cholesterol-binding proteins that generally reside in specialized detergent-resistant membrane microdomains, act as signaling scaffolds. Detergent-resistant membranes isolated from rod outer segments (ROS) have been previously shown to contain the photoreceptor G-protein, transducin. In this report we show, by subcellular fractionation, that caveolin-1 is an authentic component of purified ROS. We demonstrate that caveolin-1 in ROS almost exclusively resides in low-buoyant-density, cholesterol-rich, detergent-resistant membranes that can be disrupted by cholesterol depletion using methyl-beta-cyclodextrin (MCD). Cholesterol depletion was also observed to extract a pool of transducin alpha (Talpha) from ROS membranes. Immunoprecipitation with anti-caveolin-1 revealed the association of Talpha in the absence of Tbetagamma. Treatment of ROS with MCD resulted in a 2-fold decrease in recovery of Talpha in anti-caveolin-1 immunoprecipitates. This interaction was also completely disrupted when ROS were exposed to light in the presence of guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS), a nonhydrolyzable GTP analogue. In addition, caveolin-1/Talpha association in the immune complex was disrupted by a peptide based on the primary sequence of the caveolin-1 scaffolding domain. Finally, we confirm the colocalization of caveolin-1 and Talpha in photoreceptors by immunofluorescence microscopy. These results strongly suggest that the association between Talpha and caveolin-1 occurs in cholesterol-rich, detergent-resistant membranes and is likely to be dependent upon the activation state of Talpha.

Activation of focal adhesion kinase in adenovirus-infected human corneal fibroblasts.

PURPOSE: Focal adhesion kinase (FAK), a nonreceptor protein tyrosine kinase with protean downstream influences on cell cycle regulation, cytoskeletal dynamics, and cell attachment, is activated by integrin binding and aggregation. Adenoviruses, including those associated with human keratitis, enter permissive cells by an integrin-mediated mechanism. Hence, a possible relationship between adenovirus infection and tyrosine phosphorylation of FAK in human corneal cells was explored. METHODS: Human corneal fibroblasts (HCFs) derived from donor corneas were infected for various periods with adenovirus type 19 (Ad19) or were mock infected with virus-free medium. Parallel experiments included echistatin, which is a snake venom disintegrin and a partial inhibitor of FAK. For immunoblot analysis, Triton-X-soluble and Triton-X-insoluble fractions were analyzed by SDS-PAGE and immunoblotted with phosphospecific antibodies. Expression of the interleukin (IL)-8 gene was analyzed by RT-PCR and ELISA. RESULTS: Ad19 infection of HCFs induced tyrosine phosphorylation of a protein at 125 kDa, which was evident within 15 minutes after infection and was established by immunoprecipitation to be FAK. Immunoblot with antibody to FAK tyrosine-397 confirmed phosphorylation of this key binding site for downstream signaling proteins. Immunoblot analysis further suggested a shift in the intracellular location of phosphorylated FAK from the cytosol (Triton-X-soluble cell lysate fraction) to the cytoskeleton (Triton-X-insoluble cell lysate pellet) on infection. Finally, treatment of HCFs with echistatin reduced virus-induced expression of the neutrophil chemotaxin IL-8, previously implicated in adenoviral pathogenesis. CONCLUSIONS: Ad19 infection of HCFs induces rapid phosphorylation of FAK, and a dramatic change in its intracellular distribution. Activation of FAK may contribute to the inflammatory response to adenovirus infection of the human cornea.

Light-dependent association of Src with photoreceptor rod outer segment membrane proteins in vivo.

In vivo light exposure results in tyrosine phosphorylation of several rod outer segment (ROS) proteins (Ghalayini, A. J., Guo, X. X., Koutz, C. A, and Anderson, R. E. (1998) Exp. Eye Res. 66, 817-821). We now report the presence of Src in ROS and its increased association with bleached ROS membranes. Immunoprecipitation with anti-phosphotyrosine revealed that tyrosine kinase activity recovered from light-adapted ROS membranes was twice that recovered from dark-adapted ROS. Other experiments revealed the presence of both bleached rhodopsin and arrestin in immunoprecipitates of LROS, suggesting the formation of a multimeric complex containing Src, arrestin, and bleached rhodopsin. Additionally, when immobilized Src homology domains 2 and 3 (SH2 and SH3, respectively) were used to study the association of Src with ROS membranes, only bleached opsin and arrestin were found to associate with the SH2 domain of Src. These data strongly suggest that Src through its SH2 domain interacts with bleached rhodopsin and arrestin either directly or indirectly. Similar results were also obtained when dark-adapted and light-adapted retinas were used instead of ROS membranes. Our data strongly suggest that light exposure in vivo activates Src and promotes its association through its SH2 domain with a complex containing bleached rhodopsin and arrestin. A hypothesis for the functional significance of this phenomenon is presented.

Gonadotropin-releasing hormone for infertility in women with primary hypothalamic amenorrhea. Toward a more-interventional approach.

OBJECTIVE: To assess the effectiveness of a protocol of pulsatile gonadotropin releasing-hormone (GnRH) in treating infertility in women with primary hypothalamic amenorrhea. STUDY DESIGN: Retrospective analysis of 44 cycles treated at an infertility center. Twenty-four patients with primary hypothalamic amenorrhea were treated intravenously with pulsatile GnRH using 5 micrograms per bolus every 90 minutes. Ultrasound monitoring and cervical assessment by Insler's scoring system allowed timed injection of human chorionic gonadotropin (hCG) and intrauterine insemination if needed. Luteal support was provided with hCG. RESULTS: The ovulation rate was 95% with the 5-microgram dose. A single follicle was produced in 91% of cycles. The overall pregnancy rate per ovulatory cycle was 45%, and the pregnancy rate per patient was 83%. In patients treated previously with exogenous gonadotropins, poor results were observed. Only one case of mild overstimulation was reported. CONCLUSION: Pulsatile GnRH is an effective and safe method of treating infertility in women with primary hypothalamic amenorrhea, thus simulating normal ovulation; however, more-interventional management, including the qualitative estrogenic response, may lead to optimal results and increase the pregnancy rate.

[Radical surgical treatment of vulvar epidermoid cancer]. / Le traitement chirurgical radical du cancer épidermoïde de la vulve.

During an eighteen-year period, forty-nine patients with invasive epidermoid carcinoma of the vulva were treated by a radical vulvectomy at the Hôtel-Dieu de France hospital in Beirut. The age, the clinical presentation, and the lymph nodes involvement were close to what was published in the literature. The overall five-year survival rate was 72%. Those with stage I disease lesions had an excellent survival rate. A review of the literature would indicate that a conservative approach in the treatment of early invasive lesions can give satisfactory results, and that adjuvant radiotherapy is more beneficial than pelvic lymph nodes dissection when there is a high risk of pelvic involvement.

[Complications of laparoscopy in gynecology]. / Les complications de la coelioscopie en gynécologie.

In a review of the literature the authors describe the complications encountered during gynecological laparoscopy, insisting on the preparation phase, the conversion to laparotomy and the death cases. Vascular, digestive and urinary lesions, as well as anesthetic difficulties are discussed, aiming at a better prevention. Early recognition of the problem is an important prognostic factor for these patients.

[Cushing's syndrome and pregnancy: a case report and review of the literature]. / Syndrome de Cushing et grossesse: à propos d'un cas et revue de la littérature.

The association between pregnancy and Cushing syndrome is extremely rare. The diagnosis of Cushing syndrome during pregnancy is rendered difficult by hyperoestrogenic state that alters many of the classical tests. An early and precise diagnosis with adequate management will allow us to reduce maternal and fetal risks.

[Anatomoclinical correlations of endometriosis]. / Corrélations anatomocliniques de l'endométriose.

This is an epidemiological study of 1,498 patients who underwent laparoscopic surgery for different reasons. Between 1989 and 1996, 308 cases of endometriosis were identified among 1,498 patients who underwent laparoscopic surgery. One hundred and five patients were admitted for pelvic pain, 794 for infertility, 319 patients had both on admission, and 280 were admitted for non-gynecologic complaints. The incidence of endometriosis is related to the chief complaint on admission. This disease has different clinical manifestations, different locations and different stages. The mean age in our series is greater than that reported by the literature. The symptoms are related to the location of the lesions but not the stage of the disease. Unlike pelvic pain, infertility is correlated to the stages of the disease.