RESUMO
The natural aminocarboxylic acid product ethylenediamine- N, N'-disuccinic acid [( S, S)-EDDS] is able to form a stable complex with metal ions, making it an attractive biodegradable alternative for the synthetic metal chelator ethylenediaminetetraacetic acid (EDTA), which is currently used on a large scale in numerous applications. Previous studies have demonstrated that biodegradation of ( S, S)-EDDS may be initiated by an EDDS lyase, converting ( S, S)-EDDS via the intermediate N-(2-aminoethyl)aspartic acid (AEAA) into ethylenediamine and two molecules of fumarate. However, current knowledge of this enzyme is limited because of the absence of structural data. Here, we describe the identification and characterization of an EDDS lyase from Chelativorans sp. BNC1, which has a broad substrate scope, accepting various mono- and diamines for addition to fumarate. We report crystal structures of the enzyme in an unliganded state and in complex with formate, succinate, fumarate, AEAA, and ( S, S)-EDDS. The structures reveal a tertiary and quaternary fold that is characteristic of the aspartase/fumarase superfamily and support a mechanism that involves general base-catalyzed, sequential two-step deamination of ( S, S)-EDDS. This work broadens our understanding of mechanistic diversity within the aspartase/fumarase superfamily and will aid in the optimization of EDDS lyase for asymmetric synthesis of valuable (metal-chelating) aminocarboxylic acids.
Assuntos
Proteínas de Bactérias/metabolismo , Carbono-Nitrogênio Liases/metabolismo , Etilenodiaminas/metabolismo , Phyllobacteriaceae/enzimologia , Succinatos/metabolismo , Proteínas de Bactérias/química , Carbono-Nitrogênio Liases/química , Cristalografia por Raios X , Etilenodiaminas/química , Formiatos/química , Formiatos/metabolismo , Fumaratos/química , Fumaratos/metabolismo , Modelos Moleculares , Phyllobacteriaceae/química , Phyllobacteriaceae/metabolismo , Conformação Proteica , Especificidade por Substrato , Succinatos/químicaRESUMO
The iron binding siderophore pyoverdine constitutes a major adaptive factor contributing to both virulence and survival in fluorescent pseudomonads. For decades, pyoverdine production has allowed the identification and classification of fluorescent and nonfluorescent pseudomonads. Here, we demonstrate that PvdP, a periplasmic enzyme of previously unknown function, is a tyrosinase required for the maturation of the pyoverdine chromophore in Pseudomonas aeruginosa. PvdP converts the nonfluorescent ferribactin, containing two iron binding groups, into a fluorescent pyoverdine, forming a strong hexadentate complex with ferrous iron, by three consecutive oxidation steps. PvdP represents the first characterized member of a small family of tyrosinases present in fluorescent pseudomonads that are required for siderophore maturation and are capable of acting on large peptidic substrates.
Assuntos
Proteínas de Bactérias/metabolismo , Regulação Enzimológica da Expressão Gênica/fisiologia , Monofenol Mono-Oxigenase/metabolismo , Oligopeptídeos/metabolismo , Pseudomonas aeruginosa/enzimologia , Proteínas de Bactérias/genética , Domínio Catalítico , Regulação Bacteriana da Expressão Gênica/fisiologia , Modelos Moleculares , Monofenol Mono-Oxigenase/genética , Oligopeptídeos/genética , Filogenia , Conformação Proteica , Transporte Proteico , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/metabolismoRESUMO
Enzymatic amino acid synthesis: Kinetic resolution and asymmetric synthesis of various valuable 3-substituted aspartic acids, which were obtained in fair to good yields with diastereomeric ratio values of up to >98:2 and enantiomeric excess values of up to >99 %, by using engineered methylaspartate ammonia lyases are described. These biocatalytic methodologies for the selective preparation of aspartic acid derivatives appear to be attractive alternatives for existing chemical methods.
Assuntos
Amônia-Liases/metabolismo , Ácido Aspártico/biossíntese , Aminação , Substituição de Aminoácidos , Amônia-Liases/genética , Ácido Aspártico/química , Biocatálise , Cinética , EstereoisomerismoRESUMO
Members of the aspartase/fumarase superfamily share a common tertiary and quaternary fold, as well as a similar active site architecture; the superfamily includes aspartase, fumarase, argininosuccinate lyase, adenylosuccinate lyase, δ-crystallin, and 3-carboxy-cis,cis-muconate lactonizing enzyme (CMLE). These enzymes all process succinyl-containing substrates, leading to the formation of fumarate as the common product (except for the CMLE-catalyzed reaction, which results in the formation of a lactone). In the past few years, X-ray crystallographic analysis of several superfamily members in complex with substrate, product, or substrate analogues has provided detailed insights into their substrate binding modes and catalytic mechanisms. This structural work, combined with earlier mechanistic studies, revealed that members of the aspartase/fumarase superfamily use a common catalytic strategy, which involves general base-catalyzed formation of a stabilized aci-carboxylate (or enediolate) intermediate and the participation of a highly flexible loop, containing the signature sequence GSSxxPxKxN (named the SS loop), in substrate binding and catalysis.
Assuntos
Aspartato Amônia-Liase/química , Aspartato Amônia-Liase/metabolismo , Fumarato Hidratase/química , Fumarato Hidratase/metabolismo , Adenilossuccinato Liase/química , Adenilossuccinato Liase/genética , Adenilossuccinato Liase/metabolismo , Sequência de Aminoácidos , Argininossuccinato Liase/química , Argininossuccinato Liase/genética , Argininossuccinato Liase/metabolismo , Aspartato Amônia-Liase/genética , Catálise , Domínio Catalítico , Sequência Conservada , Escherichia coli/enzimologia , Escherichia coli/genética , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Fumarato Hidratase/genética , Humanos , Liases Intramoleculares/química , Liases Intramoleculares/genética , Liases Intramoleculares/metabolismo , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Quaternária de Proteína , Estrutura Terciária de Proteína , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Homologia de Sequência de Aminoácidos , Especificidade por Substrato , delta-Cristalinas/química , delta-Cristalinas/genética , delta-Cristalinas/metabolismoRESUMO
Methylaspartate ammonia lyase (MAL; EC 4.3.1.2) catalyzes the reversible addition of ammonia to mesaconate to give (2S,3S)-3-methylaspartate and (2S,3R)-3-methylaspartate as products. MAL is of considerable biocatalytic interest because of its potential use for the asymmetric synthesis of substituted aspartic acids, which are important building blocks for synthetic enzymes, peptides, chemicals, and pharmaceuticals. Here, we have cloned the gene encoding MAL from the thermophilic bacterium Carboxydothermus hydrogenoformans Z-2901. The enzyme (named Ch-MAL) was overproduced in Escherichia coli and purified to homogeneity by immobilized metal affinity chromatography. Ch-MAL is a dimer in solution, consisting of two identical subunits (â¼49 kDa each), and requires Mg(2+) and K(+) ions for maximum activity. The optimum pH and temperature for the deamination of (2S,3S)-3-methylaspartic acid are 9.0 and 70°C (k (cat) = 78 s(-1) and K (m) = 16 mM). Heat inactivation assays showed that Ch-MAL is stable at 50°C for >4 h, which is the highest thermal stability observed among known MALs. Ch-MAL accepts fumarate, mesaconate, ethylfumarate, and propylfumarate as substrates in the ammonia addition reaction. The enzyme also processes methylamine, ethylamine, hydrazine, hydroxylamine, and methoxylamine as nucleophiles that can replace ammonia in the addition to mesaconate, resulting in the corresponding N-substituted methylaspartic acids with excellent diastereomeric excess (>98% de). This newly identified thermostable MAL appears to be a potentially attractive biocatalyst for the stereoselective synthesis of aspartic acid derivatives on large (industrial) scale.
Assuntos
Amônia-Liases/metabolismo , Bactérias Gram-Positivas/enzimologia , Amônia-Liases/genética , Cromatografia de Afinidade , Clonagem Molecular , Ativadores de Enzimas/metabolismo , Estabilidade Enzimática , Escherichia coli/genética , Expressão Gênica , Bactérias Gram-Positivas/química , Bactérias Gram-Positivas/genética , Concentração de Íons de Hidrogênio , Magnésio/metabolismo , Peso Molecular , Potássio/metabolismo , Multimerização Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Especificidade por Substrato , TemperaturaRESUMO
3-Methylaspartate ammonia-lyase (MAL) catalyzes the reversible amination of mesaconate to give both (2S,3S)-3-methylaspartic acid and (2S,3R)-3-methylaspartic acid as products. The deamination mechanism of MAL is likely to involve general base catalysis, in which a catalytic base abstracts the C3 proton of the respective stereoisomer to generate an enolate anion intermediate that is stabilized by coordination to the essential active-site Mg(II) ion. The crystal structure of MAL in complex with (2S,3S)-3-methylaspartic acid suggests that Lys331 is the only candidate in the vicinity that can function as a general base catalyst. The structure of the complex further suggests that two other residues, His194 and Gln329, are responsible for binding the C4 carboxylate group of (2S,3S)-3-methylaspartic acid, and hence are likely candidates to assist the Mg(II) ion in stabilizing the enolate anion intermediate. In this study, the importance of Lys331, His194, and Gln329 for the activity and stereoselectivity of MAL was investigated by site-directed mutagenesis. His194 and Gln329 were replaced with either an alanine or arginine, whereas Lys331 was mutated to a glycine, alanine, glutamine, arginine, or histidine. The properties of the mutant proteins were investigated by circular dichroism (CD) spectroscopy, kinetic analysis, and (1)H NMR spectroscopy. The CD spectra of all mutants were comparable to that of wild-type MAL, and this indicates that these mutations did not result in any major conformational changes. Kinetic studies demonstrated that the mutations have a profound effect on the values of k(cat) and k(cat)/K(M); this implicates Lys331, His194 and Gln329 as mechanistically important. The (1)H NMR spectra of the amination and deamination reactions catalyzed by the mutant enzymes K331A, H194A, and Q329A showed that these mutants have strongly enhanced diastereoselectivities. In the amination direction, they catalyze the conversion of mesaconate to yield only (2S,3S)-3-methylaspartic acid, with no detectable formation of (2S,3R)-3-methylaspartic acid. The results are discussed in terms of a mechanism in which Lys331, His194, and Gln329 are involved in positioning the substrate and in formation and stabilization of the enolate anion intermediate.
Assuntos
Amônia-Liases/química , Amônia-Liases/genética , Amônia-Liases/metabolismo , Domínio Catalítico , Clostridium tetanomorphum/enzimologia , Cinética , Magnésio/química , Mutagênese Sítio-Dirigida , Proteínas Recombinantes/metabolismo , EstereoisomerismoRESUMO
In order to examine, whether the lobeline-induced cough is a true reflex or a voluntary effort to get rid of its irritating sensations in the upper respiratory tract, we systematically studied the cough response to lobeline, of subjects who were unable to make conscious discriminations i.e. were either comatose (n=4) or anaesthetized (n=5). 8 microg/kg lobeline injected into the right atrium of one and 29 microg/kg intravenously (i.v.) into another evenly and spontaneously breathing comatose subject produced a cough after 4s and 12s, respectively. Cough was repeatable and showed a dose response relationship i.e., its latency decreasing and its duration/intensity increasing with the dose. In a third subject, capable only of weak spontaneous respiration, a relatively high dose injected into the right atrium (44 microg/kg) generated a pronounced cough-like respiratory movement superimposed on the artificial ventilation and also during the apnoea after disconnecting the pump. No respiratory response was evoked in a fourth subject who had no evidence of brainstem reflexes. In five normals, cough was elicited with a mean dose of 35+/-5 microg/kg i.v. (latency 14+/-2 s; duration 10+/-3 s). After thiopental anaesthesia, injecting 41+/-7 microg/kg produced a cough within 13+/-2 s that lasted for 12+/-2 s. It may be noted that neither the later dose nor the latency or duration of cough that it produced were significantly different from the pre anaesthesia values (P>0.05). These two sets of results show unequivocally that the lobeline-induced cough is evoked reflexly; its magnitude in the conscious state could vary by subjective influences. We discuss the likelihood of its origin from juxtapulmonary capillary receptors.
Assuntos
Tosse/induzido quimicamente , Injeções Intravenosas/métodos , Lobelina/efeitos adversos , Medicamentos para o Sistema Respiratório/efeitos adversos , Ureia/análogos & derivados , Adulto , Lesões Encefálicas/complicações , Lesões Encefálicas/tratamento farmacológico , Peróxido de Carbamida , Coma/tratamento farmacológico , Coma/etiologia , Relação Dose-Resposta a Droga , Combinação de Medicamentos , Feminino , Humanos , Inalação/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Peróxidos/sangue , Tempo de Reação/efeitos dos fármacos , Testes de Função Respiratória/métodos , Fatores de Tempo , Ureia/sangueRESUMO
INTRODUCTION: Sutureless and glue-free conjunctival autograft as a treatment modality for primary pterygium is recently gaining popularity but conventional technique of suturing conjunctival autograft is still practised widely. AIM: To compare the outcome of sutureless and glue-free technique with sutures for limbal conjunctival autografting in management of primary pterygium. MATERIALS AND METHODS: A prospective interventional study was carried out in 50 consecutive eyes with primary nasal pterygium requiring surgical excision. Simple excision under local anaesthesia was performed followed by closure of the bare sclera by sutureless and glue-free conjunctival autograft in 25 eyes of 25 patients (group 1) and by the conventional method of suturing conjunctival autograft using interrupted 10-0 nylon sutures in 25 eyes of 25 patients (group 2), followed by bandaging for 24 hours in both the groups. Surgical time was recorded for both the techniques. Postoperative discomfort was assessed using preformed questionnaires. The patients were followed up for 6 months. During follow up, graft related complications and recurrence if any were noted. RESULTS: Mean surgical time for group 1 (23.20±1.55 minutes) was significantly less as compared to group 2 (37.76±1.89 minutes); (p=0.001). Postoperative symptoms were seen in less number of patients (20%) and were of shorter duration (2 weeks) in group 1 as compared to group 2 with 20 (80%) patients having symptoms lasting for 4 weeks; (p<0.001). Recurrence rate and conjunctival granuloma formation rate for group 1 (0%) and for group 2 (4%) were statistically insignificant. CONCLUSION: Sutureless and glue-free conjunctival autograft technique is simple, easy, safe, effective and less time consuming than sutured limbal autograft technique with less postoperative discomfort and adverse events encountered with the use of suture material. Postoperative results of both techniques are comparable. Hence sutureless and glue-free conjunctival autografting is a good technique for the treatment of primary pterygium.
RESUMO
A simple, three-step chemoenzymatic synthesis of l-threo-3-benzyloxyaspartate (l-TBOA), as well as l-TBOA derivatives with F, CF3, and CH3 substituents at the aromatic ring, starting from dimethyl acetylenedicarboxylate was investigated. These chiral amino acids, which are extremely difficult to prepare by chemical synthesis, form an important class of inhibitors of excitatory amino acid transporters involved in the regulation of glutamatergic neurotransmission. In addition, a new chemical procedure for the synthesis of racemic mixtures of TBOA and its derivatives was explored. These chemically prepared racemates are valuable reference compounds in chiral-phase HPLC to establish the enantiopurities of the corresponding chemoenzymatically prepared amino acids.
RESUMO
Since there is evidence that lobeline-induced sensations, associated with discomfort in the mouth, throat and chest arise by stimulating juxtapulmonary or J receptors, we were interested in investigating if similar sensations are felt by patients with left ventricular dysfunction (LVD) in whom a natural stimulation of these receptors would occur by transient interstitial oedema or during augmentation of the stimulus, by increased pulmonary blood flow during exercise. Threshold doses of lobeline produced three or more respiratory sensations simultaneously in 9 out of 10 patients, which was greater than the response of the controls (P < 0.01). With mild exercise, a greater number of patients (7) than controls (1) reported feeling two or more sensations (P < 0.01); in fact half the controls did not express a respiratory sensation with equivalent exercise (P < 0.05). The predominant lobeline-like sensations reported by patients with exercise were a feeling of heat or burning and pressure in the throat or chest (P < 0.05). The presence of cough in three patients and in none of the controls was noteworthy. The mean latency with which sensations appeared during exercise in patients (4.4 +/- 0.3 min) was almost half that in controls (7.4 +/- 0.2 min) (P < 0.005). Since, respiratory sensations in response to lobeline and exercise were intensified in patients compared to controls and since both lobeline and exercise-induced sensations were similar (P < 0.05), we speculate that a common origin exists. Despite important caveats, that we discuss, in our view these respiratory sensations and cough arise from stimulation of J receptors.
Assuntos
Exercício Físico/fisiologia , Lobelina/farmacologia , Agonistas Nicotínicos/farmacologia , Sensação/efeitos dos fármacos , Disfunção Ventricular Esquerda , Adulto , Idoso , Tosse/induzido quimicamente , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Faringe/fisiopatologia , Sistema Respiratório/efeitos dos fármacos , Sensação/fisiologia , Tórax/fisiopatologiaRESUMO
Methylaspartate ammonia-lyase (MAL; EC 4.3.1.2) catalyzes the reversible addition of ammonia to mesaconate to yield l-threo-(2S,3S)-3-methylaspartate and l-erythro-(2S,3R)-3-methylaspartate as products. In the proposed minimal mechanism for MAL of Clostridium tetanomorphum, Lys-331 acts as the (S)-specific base catalyst and abstracts the 3S-proton from l-threo-3-methylaspartate, resulting in an enolate anion intermediate. This enolic intermediate is stabilized by coordination to the essential active site Mg(2+) ion and hydrogen bonding to the Gln-329 residue. Collapse of this intermediate results in the release of ammonia and the formation of mesaconate. His-194 likely acts as the (R)-specific base catalyst and abstracts the 3R-proton from the l-erythro isomer of 3-methylaspartate, yielding the enolic intermediate. In the present study, we have investigated the importance of the residues Gln-73, Phe-170, Gln-172, Tyr-356, Thr-360, Cys-361 and Leu-384 for the catalytic activity of C. tetanomorphum MAL. These residues, which are part of the enzyme surface lining the substrate binding pocket, were subjected to site-directed mutagenesis and the mutant enzymes were characterized for their structural integrity, ability to catalyze the amination of mesaconate, and regio- and diastereoselectivity. Based on the observed properties of the mutant enzymes, combined with previous structural studies and protein engineering work, we propose a detailed catalytic mechanism for the MAL-catalyzed reaction, in which the side chains of Gln-73, Gln-172, Tyr-356, Thr-360, and Leu-384 provide favorable interactions with the substrate, which are important for substrate binding and activation. This detailed knowledge of the catalytic mechanism of MAL can serve as a guide for future protein engineering experiments.
RESUMO
Juxta-pulmonary capillary (J or pulmonary C fiber) receptors are stimulated by an increase in pulmonary blood flow and give rise to respiratory acceleration and related sensations and inhibit exercise. However, the reverse, i.e., the effect of reducing pulmonary blood flow on their reflexes, is as yet not known. This was investigated by carrying out a placebo-controlled study on the acute effects of a single dose (0.4 mg) of sublingual glyceryl nitrate (GTN), known to shift blood from the central to the peripheral circulation, on the respiratory parameters of exercising healthy subjects and on their responses to i.v. lobeline. In 10 subjects, GTN use delayed the first appearance of respiratory sensations from 9.08 ± 0.9 min to 11 min (P=0.002), reduced the increase in minute ventilation by the end of 10 min of exercise (P=0.003) and increased its duration by 1-4s and doubled it in the remaining one subject. In a majority of 8 of them, the effect of GTN on i.v. lobeline-induced respiratory reflexes and sensations was a significant increase in the dose required (P=0.006) for producing threshold effects and in the latency of their appearance (P=0.003). The latter feature points to a reduction in blood flow in the lung parenchyma where these receptors are located and to which they are sensitive. As this would have led to a reduced stimulation of these receptors, it would account for the delayed appearance of respiratory symptoms, a reduction in ventilatory increase and prolongation of exercise duration. We demonstrated a mechanism of reducing the stimulus level of J receptors by reducing pulmonary blood flow by means of pharmacological sequestration with GTN use, which then led to a reduction in the magnitude of respiratory and viscerosomatic reflexes, while noting at the same time that changes in blood flow in the pulmonary bed do not directly influence limb muscles, tendons and joints which also determine exercise output.
Assuntos
Dispneia/prevenção & controle , Exercício Físico/fisiologia , Nitroglicerina/farmacologia , Respiração/efeitos dos fármacos , Células Receptoras Sensoriais/efeitos dos fármacos , Vasodilatadores/farmacologia , Adaptação Fisiológica/efeitos dos fármacos , Administração Sublingual , Adulto , Pressão Sanguínea , Relação Dose-Resposta a Droga , Humanos , Lobelina/administração & dosagem , Pulmão/irrigação sanguínea , Pulmão/efeitos dos fármacos , Pulmão/inervação , Masculino , Valores de Referência , Reflexo/efeitos dos fármacos , Fluxo Sanguíneo Regional/efeitos dos fármacos , Medicamentos para o Sistema Respiratório/administração & dosagem , Adulto JovemRESUMO
Leptospirosis, long thought to be a disease of sewage workers and found post-flooding is re-emerging as a potentially life threatening infectious disease in urban India during monsoon and immediately after due to poor sanitation and drainage. However, it is often ignored and under diagnosed due to a low index of suspicion. We present a fatal case of leptospirosis with acute liver failure, respiratory failure and jaundice and encephalopathy.
Assuntos
Leptospirose/diagnóstico , Falência Hepática Aguda/microbiologia , Insuficiência Respiratória/microbiologia , Evolução Fatal , Feminino , Humanos , Icterícia/microbiologia , Leptospirose/complicações , Pessoa de Meia-IdadeRESUMO
Ammonia lyases catalyze the formation of α,ß-unsaturated bonds by the elimination of ammonia from their substrates. This conceptually straightforward reaction has been the emphasis of many studies, with the main focus on the catalytic mechanism of these enzymes and/or the use of these enzymes as catalysts for the synthesis of enantiomerically pure α-amino acids. In this Review aspartate ammonia lyase and 3-methylaspartate ammonia lyase, which represent two different enzyme superfamilies, are discussed in detail. In the past few years, the three-dimensional structures of these lyases in complex with their natural substrates have revealed the details of two elegant catalytic strategies. These strategies exploit similar deamination mechanisms that involve general-base catalyzed formation of an enzyme-stabilized enolate anion (aci-carboxylate) intermediate. Recent progress in the engineering and application of these enzymes to prepare enantiopure l-aspartic acid derivatives, which are highly valuable as tools for biological research and as chiral building blocks for pharmaceuticals and food additives, is also discussed.
Assuntos
Amônia-Liases/metabolismo , Aspartato Amônia-Liase/metabolismo , Amônia/química , Amônia-Liases/química , Aspartato Amônia-Liase/química , Bactérias/enzimologia , Biocatálise , Domínio Catalítico , Modelos Moleculares , Engenharia de ProteínasRESUMO
The redesign of enzymes to produce catalysts for a predefined transformation remains a major challenge in protein engineering. Here, we describe the structure-based engineering of methylaspartate ammonia lyase (which in nature catalyses the conversion of 3-methylaspartate to ammonia and 2-methylfumarate) to accept a variety of substituted amines and fumarates and catalyse the asymmetric synthesis of aspartic acid derivatives. We obtained two single-active-site mutants, one exhibiting a wide nucleophile scope including structurally diverse linear and cyclic alkylamines and one with broad electrophile scope including fumarate derivatives with alkyl, aryl, alkoxy, aryloxy, alkylthio and arylthio substituents at the C2 position. Both mutants have an enlarged active site that accommodates the new substrates while retaining the high stereo- and regioselectivity of the wild-type enzyme. As an example, we demonstrate a highly enantio- and diastereoselective synthesis of threo-3-benzyloxyaspartate (an important inhibitor of neuronal excitatory glutamate transporters in the brain).
Assuntos
Aminoácidos/síntese química , Amônia-Liases/química , Catálise , Domínio Catalítico , Cristalografia por Raios X , Modelos Moleculares , Mutagênese Sítio-DirigidaRESUMO
In 15 healthy subjects, the effect of 60 mg oral codeine and placebo was examined on intravenously injected lobeline-elicited respiratory reflexes and sensations. Its influence was also studied on ventilation and appearance of distressful respiratory sensations with modest but incremental exercise. After placebo, tachypnoea and respiratory sensations were evoked with 12.1 ± 1.9 µg/kg of lobeline i.v. (mean threshold dose) and after codeine, by significantly higher doses i.e., 18.0 ± 3.1 µg/kg (P < 0.05). Additionally after codeine, in response to incremental doses of lobeline the respiratory reflex was notably attenuated and the magnitude of respiratory sensations, subdued. Dry cough seen in 66% of the subjects with suprathreshold doses of lobeline i.e., 22.0 ± 3.4 µg/kg (mean), appeared post codeine, with significantly higher doses i.e., 27.0 ± 3.9 µg/kg (mean) (P < 0.05) and in a fewer subjects (60%). Mean increase in minute ventilation at the end of 8 min of incremental treadmill walking after codeine was 21% less than after placebo (P < 0.05); 60% of the subjects continued to walk for an additional 4 min and the onset of respiratory discomfort was delayed by 1-5 min. This is the first report of an attenuation of lobeline-elicited respiratory reflexes and sensations that are attributable to J receptors (pulmonary C fibres) by a pharmacological entity. It also suggests that codeine decreased these receptors' known contribution to respiratory augmentation and motor inhibition during exercise, which was seen as a delay in the onset of, and a decrease in the magnitude of respiratory discomfort during treadmill walking and an increase in the duration walked by more than half the subjects.
Assuntos
Analgésicos Opioides/farmacologia , Codeína/farmacologia , Exercício Físico/fisiologia , Lobelina/farmacologia , Reflexo/efeitos dos fármacos , Medicamentos para o Sistema Respiratório/farmacologia , Sistema Respiratório/efeitos dos fármacos , Sensação/efeitos dos fármacos , Adulto , Método Duplo-Cego , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Respiração/efeitos dos fármacos , Adulto JovemRESUMO
Using a physiological model of acutely increasing venous return into the lungs, i.e. by applying and then releasing lower body negative pressure (LBNP) to mimic the natural stimulus of juxtapulmonary capillary (J) or pulmonary C fibre receptors, produced an immediate and significant reduction in the amplitude of the Hoffman (H) reflex by 81±4% (P=0.001) in a majority of subjects 70% (n=5). Accompanying this was a notable change in the respiratory pattern with tidal volume (V(T)) increasing in all subjects from (mean) 0.462±.038 to 0.777±.061l/min (P=0.001) and the respiratory rate (F(R)) in 40% from 14±1 to 24±0.8 breaths/min. A feeling of pressure in throat, upper chest was reported by all and a shortness of breath-by 70% of the subjects. These were similar in nature to the respiratory sensations felt with threshold doses of intravenous lobeline, a well-established chemical stimulant of J receptors. All effects lasted for 15-20s and within a minute the parameters resumed their earlier control values. In animals, respiratory augmentation and locomotion inhibition are well-established reflexes of J receptors - this simultaneous though transitory reduction in H reflex amplitude reflecting change in the excitability of the motoneurone pool and appearance of respiratory effects, is the first demonstration in human subjects of the two reflexes appearing in response to a sudden increase in pulmonary blood flow that mimics the natural stimulus of these receptors.
Assuntos
Pressão Negativa da Região Corporal Inferior/métodos , Músculo Esquelético/fisiologia , Reflexo Anormal/fisiologia , Respiração , Sensação/fisiologia , Adulto , Relação Dose-Resposta a Droga , Eletromiografia , Feminino , Humanos , Lobelina/farmacologia , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/efeitos dos fármacos , Reflexo Anormal/efeitos dos fármacos , Respiração/efeitos dos fármacos , Medicamentos para o Sistema Respiratório/farmacologia , Sensação/efeitos dos fármacos , Limiar Sensorial/efeitos dos fármacos , Fatores de Tempo , Adulto JovemRESUMO
A prospective interventional self-control study to analyse the profile and outcome of suture less and glue free limbal conjunctival autograft for the management of primary pterygium was carried out in 80 eyes with primary nasal pterygium requiring surgical excision. Pterygium excision with limbal conjunctival autografting without using glue or sutures was performed in all the eyes followed by bandaging for 24 hours. The patients were followed up post operatively after 24 hrs., on 3rd day, after 1, 3, 6 weeks and at 3 months. They were examined for visual acuity, hemorrhage, wound gape, graft retraction, graft edema, recurrence or any other complication among the study population. The mean age of the patients was 40.78 years (range 19-80), 56.25% of which were females. Graft retraction occurred in 3 eyes (3.75%) and recurrence was seen in 1 eye (1.25%). Hemorrhage was seen in 26 (32.5%) eyes at 24 hours, which persisted in only 06(7.5%) eyes at 3 weeks and resolved completely in 100% of eyes at 6 weeks. Edema was also noted in 4 (5%) eyes, persisted only in 1(1.25%) eye at 3rd day and resolved completely by 1 week. At 6 weeks postoperatively, three (3.75%) eyes showed gain in best corrected visual acuity by one line and one (1.25%) eye by three lines on snellen's drum. No other complication was noted. Suture less and glue free limbal conjunctival autografting following pterygium excision is an effective and safe option for the management of primary pterygium.
RESUMO
Respiratory sensations of eight patients with mitral stenosis in response to i.v. lobeline and 6-min walk before percutaneous mitral valvulotomy (PMV) were 'being short of breath', pressure in chest, tracheo-bronchial irritation, a desire to cough, persistent dry cough, chest pain and were qualitatively similar amongst 75% (P=0.005) of the patients. A week after PMV lobeline evoked similar sensations but the threshold dose decreased from 32.4+/-3.8 to 24.1+/-3.2 microg/kg (P=0.001) and pulmonary artery wedge pressure (PAwP), signifying reduction in pulmonary congestion, from 23.1+/-1.4 to 14.3+/-3.4 mm Hg (P<0.001). Distance walked in 6 min increased from 217+/-58 to 319+/-51.6m; and mitral valve area from 0.63+/-0.01 to 1.43+/-0.26 cm(2) (P<0.001). A fall in lobeline-sensation threshold dose indicated reduction in pulmonary congestion and stimulus to juxtapulmonary/J (or pulmonary C fibre) receptors which suggests that they had contributed to the respiratory and viscerosomatic symptoms seen before PMV.