RESUMO
Obstructive sleep apnoea syndrome (OSAS) is associated with several sleep disorders and sleep-related problems. Therefore, the aim of this study was to compare the effects of a mandibular advancement device (MAD) with those of nasal continuous positive airway pressure (nCPAP) on self-reported symptoms of common sleep disorders and sleep-related problems in mild and moderate OSAS patients. In this randomised placebo-controlled trial, sixty-four OSAS patients (52·0 ± 9·6 years) were randomly assigned to an MAD, nCPAP or an intra-oral placebo appliance in a parallel design. All participants filled out the validated Dutch Sleep Disorders Questionnaire (SDQ) twice: one before treatment and one after six months of treatment. With 88 questions, thirteen scales were constructed, representing common sleep disorders and sleep-related problems. Linear mixed model analyses were performed to study differences between the groups for the different SDQ scales over time. The MAD group showed significant improvements over time in symptoms corresponding with 'insomnia', 'excessive daytime sleepiness', 'psychiatric sleep disorder', 'periodic limb movements', 'sleep apnoea', 'sleep paralysis', 'daytime dysfunction', 'hypnagogic hallucinations/dreaming', 'restless sleep', 'negative conditioning' and 'automatic behaviour' (range of P values: 0·000-0·014). These improvements in symptoms were, however, not significantly different from the improvements in symptoms observed in the nCPAP and placebo groups (range of P values: 0·090-0·897). It can be concluded that there is no significant difference between MAD and nCPAP in their positive effects on self-reported symptoms of common sleep disorders and sleep-related problems in mild and moderate OSAS patients. These beneficial effects may be a result of placebo effects.
Assuntos
Pressão Positiva Contínua nas Vias Aéreas , Avanço Mandibular , Autorrelato , Apneia Obstrutiva do Sono/complicações , Apneia Obstrutiva do Sono/terapia , Transtornos do Sono-Vigília/complicações , Transtornos do Sono-Vigília/fisiopatologia , Adulto , Feminino , Humanos , Masculino , Avanço Mandibular/instrumentação , Pessoa de Meia-Idade , Países Baixos , Polissonografia , Apneia Obstrutiva do Sono/fisiopatologia , Inquéritos e Questionários , Resultado do TratamentoRESUMO
AIM: The present study aims to compare the host immune responses induced by benign (meningiomas) and malignant (gliomas) brain tumors. METHODS: Peripheral blood samples from 8 meningioma and 7 glioma patients collected pre- and post operatively were assessed for cell-mediated immunity, humoral immunity and IL-6, IL-8 and TNF-a expression. Apoptosis and necrosis of circulating lymphocytes and monocytes were evaluated by Annexin/PI, while DNA analysis was applied to trace circulating cells with an abnormal DNA content. RESULTS: Cell-mediated immunity was similar in the two groups either pre- or post- operatively. However, differences in the apoptosis and necrosis of circulating lymphocytes and monocytes were observed. Menigioma patients were characterized by increased percentage of apoptotic lymphocytes and necrotic monocytes pre-operatively and apoptotic monocytes postoperatively. In contrast glioma patients showed an increase in necrotic monocytes postoperatively. Humoral immunity and cytokine expression were at comparable levels both pre- and post-operatively. IL-6 expression was significant elevated after surgery in both groups. Circulating aneuploid cells were identified in three glioma patients pre-operatively, by DNA analysis. CONCLUSION: The presented data indicate that meningioma and glioma tumors trigger comparable systemic host immunity response mediated by impairments in cell-mediated immunity due to alternations in apoptosis and necrosis that also influence their shift towards the Th2 immunity profile. Moreover, the presented evidences on the circulation of aneuploid cells in glioma patients may substantiate further the immunosuppressive phenotype detected in these patients and offer a mechanism for the rare cases that extra- neural dissemination was observed without previous surgical intervention.
Assuntos
Apoptose/fisiologia , Glioma/imunologia , Linfócitos/citologia , Neoplasias Meníngeas/imunologia , Meningioma/imunologia , Monócitos/citologia , Adulto , Idoso , Neoplasias Encefálicas/imunologia , Neoplasias Encefálicas/patologia , Movimento Celular , Feminino , Glioma/patologia , Humanos , Linfócitos/imunologia , Masculino , Neoplasias Meníngeas/patologia , Meningioma/patologia , Pessoa de Meia-Idade , Monócitos/imunologia , Necrose/metabolismoRESUMO
Lung cancer with intramedullary spinal cord metastasis (ISCM) is a rare event exhibiting dismal prognosis. In the present paper, we describe a 74-year-old male who developed bilateral leg weakness with associated backache and non-productive cough. Chest imaging evaluation demonstrated pronounced bilateral mediastinal lymphadenopathy and a nodular opacity in the right lower lobe. The patient was diagnosed with small cell lung cancer through bronchoscopic procedures. Magnetic resonance imaging of the spinal cord with contrast-enhancement revealed an intramedullary lesion consistent with metastasis at the T5-T6 level. Despite chemotherapy and thoracic spine radiotherapy, he eventually succumbed to the disease 3 months after diagnosis. A brief overview of the current literature is also provided laying emphasis on the therapeutic strategies of this unusual extrathoracic metastatic disease.
Assuntos
Carcinoma de Células Pequenas/secundário , Neoplasias Pulmonares/patologia , Neoplasias da Medula Espinal/secundário , Idoso , Broncoscopia , Carcinoma de Células Pequenas/diagnóstico , Diagnóstico Diferencial , Evolução Fatal , Humanos , Neoplasias Pulmonares/diagnóstico , Imageamento por Ressonância Magnética , Masculino , Neoplasias da Medula Espinal/diagnóstico , Tomografia Computadorizada por Raios XRESUMO
It has recently been suggested that wearing a maxillary occlusal splint (i.e. a hard acrylic resin dental appliance that covers the occlusal surfaces of the maxillary dentition and that is being indicated for the treatment of, e.g. temporomandibular pain) may be associated with a risk of aggravating obstructive sleep apnoea (OSA). The present study tested the hypothesis that raising the bite without mandibular protrusion in OSA patients is associated with an increase in the apnoea-hypopnoea index (AHI). Eighteen OSA patients (13 men; 49·5 ± 8·1 years old) received a mandibular advancement device in 0% protrusion of the mandible (0%MAD). The MAD caused a bite rise of 6 mm as measured interincisally. Polysomnographic recordings were obtained at baseline and with the 0%MAD in situ. No statistically significant difference in AHI was noted between the baseline night and the 0%MAD night. However, nine patients had an aggravation in AHI during the night they used the 0%MAD. Taking into account the previously established smallest detectable difference of 12·8 in AHI, the AHI increased in only two of the patients. The outcomes of this study suggest that an increased jaw gape without mandibular protrusion might be associated with a risk of aggravation of OSA for some, but not for all OSA patients. Dental practitioners should be aware of this possible association when treating patients with oral devices that raise the bite.
Assuntos
Avanço Mandibular/instrumentação , Placas Oclusais/efeitos adversos , Apneia Obstrutiva do Sono/etiologia , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Monitorização Ambulatorial/métodos , Países Baixos , Polissonografia/métodos , Resultado do TratamentoRESUMO
Plasma membranes of boar sperm from caput, corpus and cauda of the epididymis were purified by differential- and sucrose-density equilibrium centrifugation and were found to yield a single band at a density of 1.13 g/cm3. This fraction was enriched in acid and alkaline phosphatase, 5'-nucleotidase and (Na+ + K+)-ATPase activities, whereas it contained minimal amounts of hyaluronidase and N-acetylglucosaminidase and no succinic acid dehydrogenase activities. The plasma membrane of caput, corpus and cauda sperm had the same phospholipid/protein and cholesterol/phospholipid ratios but yielded different amounts of protein and individual lipid classes. Several changes in the plasma membrane were observed during transit of sperm through the epididymis. Within the phospholipid class a decrease in the percentage of phosphatidylethanolamine, phosphatidylserine and phosphatidylinositol was detected accompanied by an increase in amount of phosphatidylcholine, sphingomyelin and polyphosphoinositides. In the other lipid classes there was a decrease in the amount of free fatty acid and the major glycolipid. The amount of cholesterol decreased, while the amount of desmosterol and cholesterol sulfate increased. There was an increase in the amount of diacylglycerol. In addition, the changes in the fatty acid composition of the total membrane lipid and each phospholipid were determined. The above changes in the lipid composition of the plasma membrane during epididymal maturation may help to explain the decreased resistance to cold shock and changes in membrane fluidity of sperm during transit in the epididymis.
Assuntos
Epididimo/crescimento & desenvolvimento , Lipídeos de Membrana/análise , Maturidade Sexual , Envelhecimento , Animais , Fracionamento Celular , Membrana Celular/análise , Membrana Celular/ultraestrutura , Cromatografia Gasosa , Cromatografia em Camada Fina , Ácidos Graxos não Esterificados/análise , Glicerídeos/análise , Glicolipídeos/análise , Masculino , Fosfolipídeos/análise , Esteróis/análise , Suínos , Distribuição TecidualRESUMO
Oil-polluted sediment bioremediation depends on both physicochemical and biological parameters, but the effect of the latter cannot be evaluated without the optimization of the former. We aimed in optimizing the physicochemical parameters related to biodegradation by applying an ex-situ landfarming set-up combined with biostimulation to oil-polluted sediment, in order to determine the added effect of bioaugmentation by four allochthonous oil-degrading bacterial consortia in relation to the degradation efficiency of the indigenous community. We monitored hydrocarbon degradation, sediment ecotoxicity and hydrolytic activity, bacterial population sizes and bacterial community dynamics, characterizing the dominant taxa through time and at each treatment. We observed no significant differences in total degradation, but increased ecotoxicity between the different treatments receiving both biostimulation and bioaugmentation and the biostimulated-only control. Moreover, the added allochthonous bacteria quickly perished and were rarely detected, their addition inducing minimal shifts in community structure although it altered the distribution of the residual hydrocarbons in two treatments. Therefore, we concluded that biodegradation was mostly performed by the autochthonous populations while bioaugmentation, in contrast to biostimulation, did not enhance the remediation process. Our results indicate that when environmental conditions are optimized, the indigenous microbiome at a polluted site will likely outperform any allochthonous consortium.
Assuntos
Consórcios Microbianos , Poluição por Petróleo , Petróleo/metabolismo , Animais , Biodegradação Ambiental , Hidrólise , Paracentrotus , Petróleo/toxicidade , Testes de Toxicidade , VibrioRESUMO
Bacillus megaterium QM B1551 spore lipids were extracted by an improved technique, and the phospholipid and fatty acid compositions were determined. Phospholipids accounted for 65% of the total fatty acids; the neutral lipid fraction contained 15% and the remaining fatty acids were in the interphase, aqueous phase and pellet from the lipid extraction. Each phospholipid had similar fatty acid compositions as did the delipidated pellet. However, the aqueous phase and, to some extent, the interphase had unique fatty acid compositions. Also, fatty acids were found acylated to proteins, which was observed by electrophoresis of delipidated proteins from spores grown in [1-14C]palmitate. Therefore, spores contain unique non-phosphatide fatty acid components that can now be analyzed.
Assuntos
Bacillus megaterium/análise , Lipídeos/análise , Acilação , Autorradiografia , Proteínas de Bactérias/análise , Eletroforese , Ácidos Graxos/análise , Concentração de Íons de Hidrogênio , Lipídeos de Membrana/análise , Fosfolipídeos/análise , Esporos Bacterianos/análiseRESUMO
Prior to fertilization, mammalian sperm must undergo the acrosome reaction, which involves modifications of the plasma and outer acrosomal membranes followed by vesiculation and release of the membranes. The membrane fraction that was released from caudal boar sperm undergoing an in vitro acrosome-like reaction was isolated and characterized with respect to density, marker enzymes and lipid composition. This membrane had a lower phospholipid/protein ratio (mg/mg) than the sperm plasma membrane, whereas both membranes had similar molar sterol/phospholipid ratios. The major phospholipid was sphingomyelin, followed by phosphatidylethanolamine and phosphatidylcholine, whereas in the plasma membrane the order was reversed; the two major phosphoglycerides contained alkylacyl and alkenylacyl species in addition to the diacyl species. The released membrane also contained lower amounts of cholesterol sulfate and unsaturated fatty acids than the plasma membranes. These results, in combination with our studies on the changes of the sperm membranes during maturation and acrosome reaction, will allow a better understanding of the mechanism of the sperm acrosome reaction.
Assuntos
Acrossomo/fisiologia , Lipídeos de Membrana/análise , Fosfolipídeos/análise , Espermatozoides/fisiologia , Animais , Membrana Celular/análise , Membrana Celular/enzimologia , Diglicerídeos/análise , Epididimo , Ácidos Graxos não Esterificados/análise , Glicolipídeos/análise , Hidrolases/metabolismo , Masculino , Esteróis/análise , SuínosRESUMO
Oil spills are treated as a widespread problem that poses a great threat to any ecosystem. Following first response actions, bioremediation has emerged as the best strategy for combating oil spills and can be enhanced by the following two complementary approaches: bioaugmentation and biostimulation. Bioaugmentation is one of the most controversial issues of bioremediation. Studies that compare the relative performance of bioaugmentation and biostimulation suggest that nutrient addition alone has a greater effect on oil biodegradation than the addition of microbial products because the survival and degradation ability of microbes introduced to a contaminated site are highly dependent on environmental conditions. Microbial populations grown in rich media under laboratory conditions become stressed when exposed to field conditions in which nutrient concentrations are substantially lower. There is increasing evidence that the best approach to overcoming these barriers is the use of microorganisms from the polluted area, an approach proposed as autochthonous bioaugmentation (ABA) and defined as a bioaugmentation technology that exclusively uses microorganisms indigenous to the sites (soil, sand, and water) slated for decontamination. In this work, we examined the effectiveness of strategies combining autochthonous bioaugmentation with biostimulation for successful remediation of polluted marine environments. Seawater was collected from a pristine area (Agios Onoufrios Beach, Chania) and was placed in a bioreactor with 1% v/v crude oil to facilitate the adaptation of the indigenous microorganism population. The pre-adapted consortium and the indigenous population were tested in combination with inorganic or lipophilic nutrients in the presence (or absence) of biosurfactants (rhamnolipids) during 90-day long experiments. Chemical analysis (gas chromatography-mass spectrometry) of petroleum hydrocarbons confirmed the results of previous work demonstrating that the biodegradation processes were enhanced by the addition of lipophilic fertilizers (uric acid and lecithin) in combination with biosurfactants (rhamnolipids), resulting in increased removal of petroleum hydrocarbons as well as reduction of the lag phase within 15 days of treatment. Considering this outcome and examining the results, the use of biostimulation additives in combination with naturally pre-adapted hydrocarbon-degrading consortia (bioaugmentation) has proved to be an effective treatment and is a promising strategy that could be applied specifically when an oil spill approaches near a shore line and an immediate hydrocarbon degradation effort is needed.
Assuntos
Recuperação e Remediação Ambiental/métodos , Consórcios Microbianos , Poluição por Petróleo , Petróleo/análise , Água do Mar/microbiologia , Microbiologia da Água , Poluentes Químicos da Água/análise , Biodegradação Ambiental , Petróleo/metabolismo , Água do Mar/química , Tensoativos , Poluentes Químicos da Água/metabolismoRESUMO
Mediterranean coastal regions are particularly exposed to oil pollution due to extensive industrialization, urbanization and transport of crude and refined oil to and from refineries. Bioremediation of contaminated beach sand through landfarming is both simple and cost-effective to implement compared to other treatment technologies. The purpose of the present study was to investigate the effect of alternative nutrients on biodegradation of crude oil contaminated beach sand in an effort to reduce the time required for bioremediation employing only indigenous hydrocarbon degraders. A natural sandy soil was collected from Agios Onoufrios beach (Chania, Greece) and was contaminated with weathered crude oil. The indigenous microbial population in the contaminated sand was tested alone (control treatment) or in combination with inorganic nutrients (KNO3 and K2HPO4) to investigate their effects on oil biodegradation rates. In addition, the ability of biosurfactants (rhamnolipids), in the presence of organic nutrients (uric acid and lecithin), to further stimulate biodegradation was investigated in laboratory microcosms over a 45-day period. Biodegradation was tracked by GC/MS analysis of aliphatic and polycyclic aromatic hydrocarbons components and the measured concentrations were corrected for abiotic removal by hopane normalizations. It was found that the saturated fraction of the residual oil is degraded more extensively than the aromatic fraction and the bacterial growth after an incubation period of approximately 3 weeks was much greater from the bacterial growth in the control. The results show that the treatments with inorganic or organic nutrients are equally effective over almost 30 days where C12-C35n-alkanes were degraded more than 97% and polyaromatic hydrocarbons with two or three rings were degraded more than 95% within 45 days. The results clearly show that the addition of nutrients to contaminated beach sand significantly enhanced the activity of indigenous microorganisms, as well as the removal of total recoverable petroleum hydrocarbons (TRPH) over a 45-day study period.
Assuntos
Recuperação e Remediação Ambiental/métodos , Poluição por Petróleo , Petróleo/metabolismo , Poluentes Químicos da Água/metabolismo , Biodegradação Ambiental , Glicolipídeos , Grécia , Nitratos/química , Petróleo/análise , Fosfatos/química , Hidrocarbonetos Policíclicos Aromáticos/análise , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Compostos de Potássio/química , Poluentes Químicos da Água/análiseRESUMO
This is a prospective study to assess a complementary treatment for genital warts after laser vaporization. 62 patients were enrolled in two randomized groups: Al: laser vaporization alone. A2: laser vaporization, followed with Pidotimod plus vitamin C for 2 1/2 months. The latter treatment shortened the time of warts remission and marginally decreased the rate of the warts' recurrence: 81% versus 67% (N.S.). Despite the non-significant difference, this complementary treatment seems to have some efficiency.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Ácido Ascórbico/administração & dosagem , Condiloma Acuminado/tratamento farmacológico , Condiloma Acuminado/cirurgia , Terapia a Laser , Ácido Pirrolidonocarboxílico/análogos & derivados , Tiazolidinas/administração & dosagem , Administração Oral , Adolescente , Adulto , Terapia Combinada , Condiloma Acuminado/imunologia , Feminino , Humanos , Estudos Prospectivos , Ácido Pirrolidonocarboxílico/administração & dosagem , Adulto JovemRESUMO
OBJECTIVE: To compare the most recent commercial interferon-gamma release assay (IGRA), the QuantiFERON-TB Gold In-Tube (QFT-GIT), with the tuberculin skin test (TST) in Greek army recruits who were bacille Calmette-Guérin (BCG) vaccinated during childhood and had no history of tuberculosis (TB) exposure. METHOD: We conducted a cross-sectional comparison study of 1750 young army recruits. TST was performed on all participants, while QFT-GIT was performed in all subjects with TST > 0 mm and in 18 TST-negative controls (TST = 0 mm). RESULTS: Among the study subjects, 5.4% (96/1750) had TST indurations of >or=10 mm, and 3.4% (59/1750) had indurations of >or=15 mm. Among subjects with a positive TST, 11.4% (11/96) tested positive on QFT-GIT. All those with QFT-GIT positivity had TST indurations of >or=15 mm, and none of those with TST indurations of 10-14 mm were positive by QFT-GIT. The overall agreement between TST and QFT-GIT was poor (kappa = 0.02). CONCLUSION: We found a significant discordance between TST and QFT-GIT in BCG-vaccinated Greek army recruits consistent with previous studies showing that BCG received after infancy produces false-positive TST reactions. Our findings underline the need for a two-step approach in diagnosing latent TB infection in all BCG-vaccinated individuals: initial TST screening, followed by an IGRA to confirm TST positivity.
Assuntos
Vacina BCG/administração & dosagem , Interferon gama , Teste Tuberculínico/métodos , Tuberculose/diagnóstico , Adjuvantes Imunológicos/administração & dosagem , Adolescente , Adulto , Estudos Transversais , Feminino , Grécia , Humanos , Masculino , Militares , Tuberculose/imunologia , Adulto JovemAssuntos
Síndrome de Behçet/imunologia , Citocinas/sangue , Células Th1/imunologia , Células Th2/imunologia , Adolescente , Adulto , Antígenos CD/sangue , Síndrome de Behçet/sangue , Feminino , Humanos , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Valores de Referência , Linfócitos T/imunologiaRESUMO
AIMS: (1) To identify factors at 1 week of age which put infants at risk of failing to sleep through the night at 12 weeks of age. (2) To assess whether a behavioural programme increases the likelihood that these infants will sleep through the night at 12 weeks of age. METHODS: A community sample of 316 newborn infants was employed to identify the risk factors at 1 week of age which increased the likelihood of failing to sleep through the night at 12 weeks of age. Infants who met these risk criteria and were randomly assigned to a behavioural programme were compared with at risk infants in the control group on measures of sleeping, crying, and feeding at 12 weeks of age. RESULTS: Infants who had a high number (>11) of feeds in 24 hours at 1 week were 2.7 times (95% CI 1.5 to 4.8) more likely than other control group infants to fail to sleep through the night at 12 weeks of age. At 12 weeks, 82% of these at risk infants assigned to the behavioural programme, compared to 61% in the control group, slept through the night. The findings were similar in breast and bottle feeders. CONCLUSIONS: Preventing infant sleeping problems should be more cost effective than treating them after they have arisen. This study provides evidence that it is possible to identify infants who are at risk of failing to sleep through the night at an early age, and that a simple, three step, preventive behavioural programme increases the number who sleep through the night by 21%.
Assuntos
Terapia Comportamental/métodos , Transtornos do Sono-Vigília/prevenção & controle , Alimentação com Mamadeira , Aleitamento Materno , Estudos de Casos e Controles , Etnicidade , Seguimentos , Humanos , Lactente , Fenômenos Fisiológicos da Nutrição do Lactente , Recém-Nascido , Ensaios Clínicos Controlados Aleatórios como Assunto , Análise de Regressão , Fatores de Risco , Transtornos do Sono-Vigília/diagnósticoRESUMO
An in vitro acrosome-like reaction was induced in spermatozoa from the boar cauda epididymis by incubation in Tyrode's solution containing 1 mg/ml fatty acid-free bovine serum albumin. Plasma membranes were isolated from the spermatozoa at different times during the incubation and analyzed for their lipid composition. The total lipid, phospholipid, and glycolipid content of the membranes did not change during the acrosome-like reaction, whereas the amount of diacylglycerols and free fatty acids increased. Within the phospholipid class, a decrease of the inositol phospholipid and and sphingomyelin content was observed, whereas the other phospholipids of the plasma membranes did not decrease significantly after 2 h of incubation. Changes in the sterol composition of the membranes were also observed. The onset of the lipid changes was correlated with the uptake of extracellular calcium by the spermatozoa. These results for the lipid changes in isolated sperm plasma membranes during an in vitro acrosome reaction provide the first direct evidence that a modulation of the plasma membrane lipid composition is involved in an acrosome-like reaction of mammalian spermatozoa.
Assuntos
Acrossomo/fisiologia , Fusão de Membrana , Lipídeos de Membrana/metabolismo , Espermatozoides/metabolismo , Espermatozoides/fisiologia , Animais , Membrana Celular/metabolismo , Diglicerídeos/metabolismo , Ácidos Graxos não Esterificados/metabolismo , Glicolipídeos/metabolismo , Técnicas In Vitro , Masculino , Consumo de Oxigênio , Fosfolipídeos/metabolismo , Esteróis/metabolismo , SuínosRESUMO
Boar sperm plasma membranes and the membranes released during an in vitro acrosome-like reaction were capable of autophosphorylation. The purified membranes were incubated in Tyrode's buffer containing [32P]ATP with or without Ca2+ and/or diacylglycerol. In both membrane fractions, Ca2+ plus diacylglycerol stimulated the autophosphorylation of several sperm membrane proteins. These results suggest a protein kinase C activity is present in sperm membranes and could play a role in the acrosome reaction.
Assuntos
Trifosfato de Adenosina/metabolismo , Proteínas de Membrana/metabolismo , Espermatozoides/metabolismo , Animais , Cloreto de Cálcio/farmacologia , Membrana Celular/metabolismo , Diglicerídeos/farmacologia , Eletroforese em Gel de Poliacrilamida , Masculino , Proteínas de Membrana/isolamento & purificação , Radioisótopos de Fósforo , Fosforilação , SuínosRESUMO
Phospholipase C from human platelets was found to catalyze the Ca2+-dependent degradation of phosphatidylinositol (PI), phosphatidylinositol 4'-phosphate (DPI), and phosphatidylinositol 4',5'-bisphosphate (TPI) at Ca2+ concentrations from 150 microM to 5 mM. Both DPI and TPI inhibited the hydrolysis of [2-3H]inositol-labeled PI (250 microM) in a concentration-dependent manner. The use of DPI and TPI from beef brain, both of which have fatty acid compositions different from that of soybean PI, permitted an assessment of the inhibitory effect of polyphosphoinositides on the hydrolysis of PI by phospholipase C. Fatty acid analysis of the diacylglycerols formed demonstrated that DPI and TPI, when incubated in mixture with PI, were competitive substrates for PI hydrolysis. Increasing the DPI/PI ratio from 0 to 0.3 caused a shift in the degradation of PI to DPI without greatly affecting the formation of 1,2-diacylglycerol. TPI alone, or in mixture with PI, was a poor substrate for phospholipase C. Increasing the TPI/PI ratio from 0 to 0.21, on the other hand, inhibited both PI degradation (greater than or equal to 95%) and overall formation of 1,2-diacylglycerol (greater than or equal to 82%). Kinetic analysis revealed that TPI acts as a mixed-type inhibitor with a Ki of about 10 microM. The Ka for Ca2+ in PI hydrolysis was profoundly increased from 5 to 180 microM when TPI (36 microM) was included with PI (250 microM). Optimum PI degradation under these conditions was only attained when the calcium concentration approached 4 mM. Analysis of phospholipids from unstimulated human platelets from five different donors revealed DPI/PI and TPI/PI ratios of 0.42 and 0.16, respectively. These findings, combined with the observed inhibition of PI hydrolysis by TPI at a TPI/PI ratio of 0.16, would suggest that in unstimulated platelets phospholipase C activity may be inhibited by greater than or equal to 75%. Changes in 33P-prelabeled phospholipids of intact platelets upon stimulation with thrombin indicated a transient decline in 33P label of both TPI and DPI (15 s) followed by an increase in [33P]phosphatidic acid but no change in [33P]PI. The finding that DPI is selectively degraded by phospholipase C in mixture with PI at DPI/PI ratios determined to be present in unstimulated platelets indicates that DPI may be more important than PI in the formation of 1,2-diacylglycerol which is believed to serve as precursor of arachidonic acid for thromboxane biosynthesis. Furthermore, the results suggest that in human platelets TPI may serve as modulator for the formation of 1,2-diacylglycerol from inositol phospholipids.
Assuntos
Plaquetas/enzimologia , Fosfatos de Fosfatidilinositol , Fosfatidilinositóis/farmacologia , Fosfolipases/sangue , Fosfolipases Tipo C/sangue , Cálcio/farmacologia , Humanos , Hidrólise , Cinética , Fosfatidilinositol 4,5-Difosfato , Fosfatidilinositóis/metabolismo , Fosfolipases Tipo C/antagonistas & inibidoresRESUMO
Ninety Enterobacter cloacae strains isolated from 12 Greek hospitals were examined in terms of epidemiological types and resistance mechanisms. Using O serotyping 69% of the strains were assigned to a specific serotype and overall 16 different serotypes were identified. The combination of serotyping, phagetyping and biotyping efficiently discriminated most of the strains, indicating that single epidemic strains were not prevalent, although serotypes 3, 7, and group II predominated. Eight representative strains, all resistant to gentamicin, tobramycin, amikacin and netilmicin, were further examined for transferability and mechanisms of resistance. Aminoglycoside resistance was found to be transferable in most strains, and 13 R plasmids of 40-120 MDa molecular weight were detected. The enzymes detected consisted of three enzymes active against gentamicin [ANT(2h'), AAC(3)-I and AAC(3)-V]; three active against tobramycin [ANT(2"), AAC(3)-V and AAC(6')-I]; two active against netilmicin [AAC(3)-V and AAC(6')-I]; and one active against amikacin [AAC(6')-I]. APH(3') and ANT (3"), which modify neomycin and streptomycin plus spectinomycin respectively, were also found. Overall up to five aminoglycoside modifying enzymes were detected on the same R plasmid, AAC(6')-I plus ANT(2") being the most prevalent. The high incidence of multiresistance in Enterobacter cloacae and the fact that resistance is due to enzymatic inactivation of the antibiotics, indicate that in Greece this species might act as a gene pool for the spread of resistance to other bacteria of clinical relevance.