RESUMO
AIM: To document satisfaction with root canal treatment procedures and outcomes among patients treated at Swedish public dental clinics. METHOD: The original material comprised 243 patients who began root canal treatment (RCT) at 20 public dental clinics in the county of Västra Götaland, Sweden. One to three years later, 236 (97.1%) were posted a questionnaire of eight items, rating patient perceptions of RCT-completion, present pain intensity and satisfaction with the RCT. To evaluate the reliability of the original responses, the first 50 respondents were mailed a follow-up questionnaire. Both descriptive and analytical statistics were used to compare respondents and non-respondents and tooth groups. RESULTS: One hundred and fifty-nine patients (67.4%) responded: 86 (54.1%) women and 73 (45.9%) men. The mean age 52.5 years, was higher than for non-respondents (P<0.001). A completed root filling was registered for the majority of the teeth (n = 112, 70.9%), but significantly fewer molars had been completed (n = 46, 59.7%, P = 0.02). Fifty percent (n = 59) of the patients reported current pain, mostly mild in intensity (n = 45, 38.1%). One hundred and twenty-three patients (80.9%) recalled experiencing pain during RCT. The highest satisfaction was registered for the item 'chewing ability' (mean = 1.6, SD = 1.9). The majority of patients (n = 114, 75.0%) stated that in retrospect they would still have chosen RCT. However, these patients belonged to the group which either registered the tooth as still present, or had not experienced much discomfort during or after RCT. Forty-four patients (88.0%) responded to the second questionnaire. The reliability of the responses was good. In summary, one to 3 years after beginning RCT at a public dental clinic, patient satisfaction was high, even though every fourth molar had been extracted or treatment had not been completed and half the patients reported persistent pain. The reliability of the patients' responses was considered to be good. CONCLUSIONS: The results indicate a need for further clinical observational studies of RCTs undertaken in general dental practice, with special reference to patient-centred outcomes.
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AIM: To study patient- and tooth-specific characteristics of teeth indicated for root canal treatment, in the public dental service of the county of Västra Götaland, Sweden. METHODOLOGY: During a designated 8-week period, general dental practitioners working at 20 different public dental clinics consecutively registered indications for undertaking root canal treatment. The patients' subjective level of pain was also registered (visual analogue scale) at the very first appointment. The following information was retrieved from computerized dental records and radiographs: gender, age, number of remaining teeth, tooth group, previous restoration, number of restored surfaces, dental caries and tooth substance loss. Gender and age were compared using both descriptive and analytical statistics. RESULTS: The material comprised 243 teeth in 243 patients: 128 (52.7%) women and 115 (47.3%) men, mean age 48.3 years. Molar teeth predominated (47.7%). Most of the teeth (83.5%) had previously been restored and exhibited significant loss of tooth substance, more than a third of the crown (71.3%). Dental caries was present in 127 teeth (62.9%). Dental trauma was implicated in only seven cases (2.9%). Initial treatment was frequently undertaken at an emergency visit, for relief of symptoms (64.9%). The most commonly registered indication was pulpal necrosis with apical periodontitis (38.1%), followed by pulpitis (37.7%). Retreatment of a root filled tooth was reported in 18 teeth (7.4%). CONCLUSIONS: In the general Public Dental Service of Sweden, root canal treatment is most frequently undertaken in molars. The primary indication is relief of symptoms. Retreatment of root filled teeth is uncommon.
Assuntos
Clínicas Odontológicas/estatística & dados numéricos , Padrões de Prática Odontológica/estatística & dados numéricos , Tratamento do Canal Radicular/estatística & dados numéricos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Coroas/estatística & dados numéricos , Cárie Dentária/epidemiologia , Cárie Dentária/terapia , Necrose da Polpa Dentária/epidemiologia , Necrose da Polpa Dentária/terapia , Restauração Dentária Permanente/estatística & dados numéricos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Dente Molar , Periodontite Periapical/epidemiologia , Periodontite Periapical/terapia , Pulpite/epidemiologia , Pulpite/terapia , Retratamento/estatística & dados numéricos , Obturação do Canal Radicular/estatística & dados numéricos , Inquéritos e Questionários , Suécia , Adulto JovemRESUMO
This position statement represents a consensus of an expert committee convened by the European Society of Endodontology (ESE) on Antibiotics in Endodontics. The statement is based on current scientific evidence as well as the expertise of the committee. The goal is to provide dentists and other healthcare workers with evidence-based criteria for when to use antibiotics in the treatment of endodontic infections, traumatic injuries of the teeth, revascularization procedures in immature teeth with pulp necrosis, and in prophylaxis for medically compromised patients. It also highlights the role that dentists and others can play in preventing the overuse of antibiotics. A recent review article provides the basis for this position statement and more detailed background information (International Endodontic Journal, 2017, https://doi.org/10.1111/iej.12741). Given the dynamic nature of research in this area, this position statement will be updated at appropriate intervals.
Assuntos
Antibacterianos/uso terapêutico , Endodontia/normas , Antibioticoprofilaxia , Contraindicações de Medicamentos , Humanos , Infecções/tratamento farmacológico , Abscesso Periapical/tratamento farmacológico , Traumatismos Dentários/tratamento farmacológico , Reimplante DentárioRESUMO
Femoroacetabular impingement (FAI) is a common cause of hip pain and dysfunction. The purpose of this study was to report outcome 2 years after the arthroscopic treatment of FAI using validated outcome measurements. Two hundred and eighty-nine patients (males = 190, females = 99) with a mean age of 37 years underwent arthroscopic surgery for FAI. Patients were included consecutively in a hip arthroscopy registry. The cohort was evaluated using online web-based validated health-related patient-reported outcomes measurements, including the iHOT-12, HAGOS, EQ-5D, HSAS for physical activity level, VAS for overall hip function and overall satisfaction. The mean follow-up time was 25.4 months. Pre-operative scores compared with those obtained at follow-up revealed statistically and clinically significant improvements (P < 0.05) for all measured outcomes; iHOT-12 (43 vs 66), VAS for global hip function (50 vs 71), HSAS (2.9 vs 3.6), EQ-5D index (0.58 vs 0.75), EQ-VAS (67 vs 75) and HAGOS different subscales (56 vs 76, 51 vs 69, 60 vs 78, 40 vs 65, 29 vs 57, 33 vs 58). At the 2-year follow-up, 236 patients (82%) reported they were satisfied with the outcome of surgery. We conclude that arthroscopic treatment for FAI resulted in statistically and clinically significant improvements in outcome parameters.
Assuntos
Artroscopia/métodos , Impacto Femoroacetabular/cirurgia , Fibrocartilagem/cirurgia , Articulação do Quadril/cirurgia , Sistema de Registros , Adulto , Estudos de Coortes , Exercício Físico , Feminino , Impacto Femoroacetabular/fisiopatologia , Fibrocartilagem/fisiopatologia , Seguimentos , Articulação do Quadril/fisiopatologia , Humanos , Masculino , Pessoa de Meia-Idade , Medidas de Resultados Relatados pelo Paciente , Satisfação do Paciente , Estudos Prospectivos , Resultado do Tratamento , Escala Visual Analógica , Adulto JovemRESUMO
AIM: Root canal treatment of teeth with necrotic pulps and apical periodontitis may be complicated by limited access to the root canals due to restorations and dystrophic calcifications. The objective of this study was to evaluate retrograde root canal as a primary treatment using a surgical approach as an alternative to conventional orthograde treatment. METHODOLOGY: Patients with apical periodontitis in the anterior region of the maxilla were consecutively recruited to the study over a period of 4 years. Fifty-seven patients met the inclusion criteria and received retrograde root canal treatment. A clinical and radiographic evaluation was made after 1 and 2 years postoperatively. RESULTS: Clinical and radiographically evaluation after 2 years revealed a successful outcome (as defined in this study) in 90% of the cases. CONCLUSION: Retrograde root canal as a primary treatment was a reliable alternative to treat apical periodontitis on single- and two-rooted teeth with limited orthograde access to the root canals in the maxilla.
Assuntos
Obturação Retrógrada/métodos , Tratamento do Canal Radicular/métodos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Maxila , Pessoa de Meia-Idade , Periodontite Periapical/diagnóstico por imagem , Periodontite Periapical/terapia , Estudos Prospectivos , Radiografia Dentária , Adulto JovemRESUMO
The overuse of antibiotics and the emergence of antibiotic-resistant bacterial strains is a global concern. This concern is also of importance in terms of the oral microbiota and the use of antibiotics to deal with oral and dental infections. The aim of this paper was to review the current literature on the indications and use of antibiotics and to make recommendations for their prescription in endodontic patients. Odontogenic infections, including endodontic infections, are polymicrobial, and in most cases, the prescription of antibiotics is empirical. This has led to the increasing use of broad-spectrum antibiotics even in cases where antibiotics are not indicated, such as symptomatic irreversible pulpitis, necrotic pulps and localized acute apical abscesses. In case of discrete and localized swelling, the primary aim is to achieve drainage without additional antibiotics. Adjunctive antibiotic treatment may be necessary in the prevention of the spread of infection, in acute apical abscesses with systemic involvement and in progressive and persistent infections. Medically compromised patients are more susceptible to complication arising from odontogenic infections and antimicrobials have a more specific role in their treatment. Therefore, antibiotics should be considered in patients having systemic diseases with compromised immunity or in patients with a localized congenital or acquired altered defence capacity, such as patients with infective endocarditis, prosthetic cardiac valves or recent prosthetic joint replacement. Penicillin VK, possibly combined with metronidazole to cover anaerobic strains, is still effective in most cases. However, amoxicillin (alone or together with clavulanic acid) is recommended because of better absorption and lower risk of side effects. In case of confirmed penicillin allergy, lincosamides such as clindamycin are the drug of choice.
Assuntos
Antibacterianos/uso terapêutico , Endodontia , Doenças Dentárias/tratamento farmacológico , Administração Tópica , Europa (Continente) , Infecção Focal Dentária/tratamento farmacológico , Humanos , Prescrição Inadequada/prevenção & controle , Prescrição Inadequada/estatística & dados numéricos , Padrões de Prática OdontológicaRESUMO
The purpose of the present work was to optimise the filtration and dose setting for double-contrast barium enema examinations using a Philips MultiDiagnost Eleva FD system. A phantom study was performed prior to a patient study. A CDRAD phantom was used in a study where copper and aluminium filtration, different detector doses and tube potentials were examined. The image quality was evaluated using the software CDRAD Analyser and the phantom dose was determined using the Monte Carlo-based software PCXMC. The original setting [100 % detector dose (660 nGy air kerma) and a total filtration of 3.5 mm Al, at 81 kVp] and two other settings identified by the phantom study (100 % detector dose and additional filtration of 1 mm Al and 0.2 mm Cu as well as 80 % detector dose and added filtration of 1 mm Al and 0.2 mm Cu) were included in the patient study. The patient study included 60 patients and up to 8 images from each patient. Six radiologists performed a visual grading characteristics study to evaluate the image quality. A four-step scale was used to judge the fulfillment of three image quality criteria. No overall statistical significant difference in image quality was found between the three settings (P > 0.05). The decrease in the effective dose for the settings in the patient study was 15 % when filtration was added and 34 % when both filtrations was added and detector dose was reduced. The study indicates that additional filtration of 1 mm Al and 0.2 mm Cu and a decrease in detector dose by 20 % from the original setting can be used in colon examinations with Philips MultiDiagnost Eleva FD to reduce the patient dose by 30 % without significantly affecting the image quality. For 20 exposures, this corresponds to a decrease in the effective dose from 1.6 to 1.1 mSv.
Assuntos
Sulfato de Bário/administração & dosagem , Colo/diagnóstico por imagem , Colonografia Tomográfica Computadorizada/métodos , Enema/métodos , Intensificação de Imagem Radiográfica/métodos , Colonografia Tomográfica Computadorizada/instrumentação , Meios de Contraste , Relação Dose-Resposta a Droga , Humanos , Imagens de Fantasmas , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
AIM: To investigate the technical feasibility and outcome of retrograde root canal treatment. SUMMARY: Endodontic access cavity preparation in abutment teeth may jeopardize the retention of the coronal restoration leading to prosthodontic failure. In such cases leaving the crown intact and performing retrograde root canal treatment might be an alternative approach. The potential to promote healing with retrograde endodontic treatment, and the technical feasibility to shape, clean and fill the canal was evaluated retrospectively. The study consisted of 21 incisors, canines and premolar teeth followed-up clinically and radiographically from 6 to 48 months. In 14 teeth the canals were completely negotiated. These cases were all judged as completely healed. In five cases no canal could be explored by files and a conventional ultrasonic root-end preparation and filling was performed. Two of these were classified as completely healed and three as 'uncertain'. In 2 two-rooted premolars a combination was performed with complete instrumentation of the buccal canal and the ultrasonic root-end preparation of the palatal root. One case was judged as a failure and the other was classified as completely healed. The results from this preliminary evaluation of retrograde root canal treatment are promising and merit a randomized clinical trial. KEY LEARNING POINTS: Abutment teeth with vital pulps may develop pulp necrosis and apical periodontitis in 10% of cases. Endodontic access preparation through an artificial crown may weaken its retention and jeopardize the longevity of a bridgework. Retrograde root canal treatment is often feasible in maxillary teeth. Results from this preliminary study suggest that treatment outcome for retrograde and orthograde root canal treatment is similar.
Assuntos
Dente Suporte , Periodontite Periapical/cirurgia , Obturação Retrógrada , Adulto , Necrose da Polpa Dentária/cirurgia , Estudos de Viabilidade , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Resultado do TratamentoRESUMO
BACKGROUND: Onychomycosis is a rare disorder in children. Few studies exist on the incidence or prevalence of onychomycosis in children. OBJECTIVE: To examine the epidemiology of childhood onychomycosis in Iceland during the period 1982-2000. METHODS: Results from all mycological samples taken from children in Iceland from 1982 to 2000 were examined. Information about the requesting physician, unique social security number, date of birth, sex, results of culture and microscopy were registered. Growth of a dermatophyte was taken as an indication of a case of onychomycosis. RESULTS: During the period 1982-2000 a total of 493 samples from 408 Icelandic children, aged 0-17 years, were examined. Dermatophytes were cultured from 148 (30.0%) samples. During the period 1982-85, the mean annual incidence of positive cultures was 1.65 per 100,000, increasing to 21.30 per 100,000 for the years 1996-2000. Trichophyton rubrum was the dominating organism and was found in 102/148 cases positive for the growth of a dermatophyte. The incidence of positive dermatophyte cultures increased with age and was found in eight children aged 0-4, and in 57 children aged 10-14 years. CONCLUSION: Onychomycosis is rare in children, but increases with age. It seems that onychomycosis increased during the study period, but it is not clear if this was due to a true increase in the prevalence of onychomycosis or an increased awareness of onychomycosis, or both.
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Onicomicose/epidemiologia , Adolescente , Arthrodermataceae/isolamento & purificação , Criança , Pré-Escolar , Feminino , Humanos , Islândia/epidemiologia , Incidência , Lactente , Masculino , PrevalênciaRESUMO
We describe a previously unreported case of cranial arterial dolichoectasia associated with spontaneous dissection of the petrous (C2) segment of the internal carotid artery (ICA) with 2 patent lumena. Dolichoectasia of the cranial arteries and different types of double lumen of ICA are discussed. A review of previously reported cases is included.
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Doenças das Artérias Carótidas/diagnóstico por imagem , Dissecação da Artéria Carótida Interna/diagnóstico por imagem , Artéria Carótida Interna/diagnóstico por imagem , Insuficiência Vertebrobasilar/diagnóstico por imagem , Artéria Basilar/diagnóstico por imagem , Doenças das Artérias Carótidas/complicações , Dissecação da Artéria Carótida Interna/etiologia , Dilatação Patológica , Humanos , Masculino , Pessoa de Meia-Idade , Radiografia , Insuficiência Vertebrobasilar/complicaçõesRESUMO
The authors present the case of a young woman, suffering from progressive tetraplegia, cranial nerve dysfunction, respiratory failure and lethargy due to radiographically demonstrated increased brainstem compression after coil embolization of an aneurysm of the midportion of the basilar artery. The patients condition improved dramatically after transclival decompression. The importance of understanding the effects of a rigid endovascular mass on adjacent neural structures is emphasized.
Assuntos
Fossa Craniana Posterior/cirurgia , Descompressão Cirúrgica/métodos , Embolização Terapêutica/efeitos adversos , Aneurisma Intracraniano/complicações , Aneurisma Intracraniano/terapia , Próteses e Implantes/efeitos adversos , Adulto , Artéria Basilar/diagnóstico por imagem , Artéria Basilar/patologia , Artéria Basilar/cirurgia , Angiografia Cerebral , Fossa Craniana Posterior/anatomia & histologia , Fossa Craniana Posterior/patologia , Feminino , Humanos , Aneurisma Intracraniano/fisiopatologia , Imageamento por Ressonância Magnética , Osso Occipital/anatomia & histologia , Osso Occipital/patologia , Osso Occipital/cirurgia , Paresia/etiologia , Paresia/fisiopatologia , Paresia/cirurgia , Ponte/irrigação sanguínea , Ponte/patologia , Ponte/fisiopatologia , Reoperação , Insuficiência Respiratória/etiologia , Insuficiência Respiratória/patologia , Insuficiência Respiratória/cirurgia , Tomografia Computadorizada por Raios X , Resultado do TratamentoRESUMO
The discovery of toxicity related to glucose degradation products (GDP) has initiated the development of new PD fluids with low GDP concentrations and higher, more physiological, pH levels. Cell numbers, differential counts and the respiratory burst responses of peritoneal leukocytes were compared between patients treated with the low GDP, high pH fluid Gambrosol-trio (n=10) and a conventional fluid (n=12). Effluents from over-night dwells were collected and leukocytes were evaluated morphologically and by luminol-amplified chemiluminescence (CL) after stimulation with opsonized zymosan. The frequency of necrosis and early apoptosis was quantified by means of annexin V binding and propidium iodide uptake. The Gambrosol-trio group produced significantly higher (p<5%) macrophage counts and stronger CL responses (p<10%) than did the conventional fluid group. The cell compositions did not differ significantly between the groups. Necrosis was significantly more common among the cells in the conventional fluid group. The occurrence of apoptosis did not differ between the fluids.
Assuntos
Soluções para Diálise/farmacologia , Glucose/metabolismo , Leucócitos/fisiologia , Diálise Peritoneal/métodos , Explosão Respiratória/efeitos dos fármacos , Adulto , Idoso , Apoptose , Sobrevivência Celular/efeitos dos fármacos , Soluções para Diálise/metabolismo , Feminino , Humanos , Concentração de Íons de Hidrogênio , Nefropatias/terapia , Leucócitos/metabolismo , Leucócitos/patologia , Masculino , Pessoa de Meia-Idade , Modelos Teóricos , NecroseRESUMO
Heat sterilization of glucose containing peritoneal dialysis (PD) fluids induces the production of cytotoxic glucose degradation products (GDPs), some of which are still unidentified. The present study was performed to characterize the kinetics and the dose-response of the respiratory burst inhibition of GDPs and to compare different fluids in this respect. The zymosan-induced respiratory burst of rat peritoneal neutrophils and macrophages was measured by chemiluminescence (CL) after incubation in vitro for 1, 2, and 4 hours in different homemade and commercially available PD fluids, followed by one hour of recovery in Hanks' buffer. Heat sterilized fluids were compared with their filter sterilized equivalents at two different pH levels. The results revealed that the inhibitory effect of heat sterilized fluids on the respiratory burst of peritoneal neutrophils is additive to that of low pH, but more fast-acting and, in contrast to the pH effect, similar in magnitude to its in vivo equivalent. The effect developed within 1 hour and had a linear dose response. The low GDP fluid Gambrosol-Bio was less toxic than the conventional fluid Gambrosol, but the difference was smaller than expected in relation to measured concentrations of known GDPs. Macrophages were less sensitive than neutrophils to the GDP effect.
Assuntos
Glucose/toxicidade , Temperatura Alta , Leucócitos/metabolismo , Diálise Peritoneal , Explosão Respiratória/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Glucose/metabolismo , Medições Luminescentes , Masculino , Ratos , Ratos Sprague-Dawley , EsterilizaçãoRESUMO
An integrated bioprocess has been developed suitable for production of recombinant peptides using a gene multimerization strategy and site-specific cleavage of the resulting gene product. The process has been used for production in E. coli of the human proinsulin C-peptide via a fusion protein BB-C7 containing seven copies of the 31-residues C-peptide monomer. The fusion protein BB-C7 was expressed at high level, 1.8 g l(-1), as a soluble gene product in the cytoplasm. A heat treatment procedure efficiently released the BB-C7 fusion protein into the culture medium. This step also served as an initial purification step by precipitating the majority of the host cell proteins, resulting in a 70% purity of the BB-C7 fusion protein. Following cationic polyelectrolyte precipitation of the nucleic acids and anion exchange chromatography, native C-peptide monomers were obtained by enzymatic cleavage at flanking arginine residues. The released C-peptide material was further purified by reversed-phase chromatography and size exclusion chromatography. The overall yield of native C-peptide at a purity exceeding 99% was 400 mg l(-1) culture, corresponding to an overall recovery of 56%. The suitability of this process also for the production of other recombinant proteins is discussed.
Assuntos
Biotecnologia/métodos , Peptídeo C/biossíntese , Peptídeo C/isolamento & purificação , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/isolamento & purificação , Reatores Biológicos , Peptídeo C/genética , Carboxipeptidase B , Carboxipeptidases/química , Carboxipeptidases/metabolismo , Precipitação Química , Cromatografia Líquida/métodos , Temperatura Alta , Humanos , Ácidos Nucleicos/isolamento & purificação , Engenharia de Proteínas/métodos , Proteínas Recombinantes de Fusão/genética , Tripsina/química , Tripsina/metabolismoRESUMO
Recent reports have demonstrated beneficial effects of proinsulin C-peptide in the diabetic state, including improvements of kidney and nerve function. To examine the background to these effects, C-peptide binding to cell membranes has been studied by using fluorescence correlation spectroscopy. Measurements of ligand-membrane interactions at single-molecule detection sensitivity in 0.2-fl confocal volume elements show specific binding of fluorescently labeled C-peptide to several human cell types. Full saturation of the C-peptide binding to the cell surface is obtained at low nanomolar concentrations. Scatchard analysis of binding to renal tubular cells indicates the existence of a high-affinity binding process with K(ass) > 3.3 x 10(9) M(-1). Addition of excess unlabeled C-peptide is accompanied by competitive displacement, yielding a dissociation rate constant of 4.5 x 10(-4) s(-1). The C-terminal pentapeptide also displaces C-peptide bound to cell membranes, indicating that the binding occurs at this segment of the ligand. Nonnative D-C-peptide and a randomly scrambled C-peptide do not compete for binding with the labeled C-peptide, nor were crossreactions observed with insulin, insulin-like growth factor (IGF)-I, IGF-II, or proinsulin. Pretreatment of cells with pertussis toxin, known to modify receptor-coupled G proteins, abolishes the binding. It is concluded that C-peptide binds to specific G protein-coupled receptors on human cell membranes, thus providing a molecular basis for its biological effects.
Assuntos
Peptídeo C/metabolismo , Túbulos Renais/metabolismo , Ligação Competitiva , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Humanos , Túbulos Renais/citologia , Túbulos Renais/efeitos dos fármacos , Cinética , Microscopia de Fluorescência , Toxina Pertussis , Fatores de Virulência de Bordetella/farmacologiaRESUMO
Hydrogen/deuterium (H/D) exchange measurements in low and moderate concentrations of GuHCI were conducted on the side chain H(N) atoms of the seven tryptophans of pseudo wild-type human carbonic anhydrase II. Tryptophans 5, 16 and 245, situated in or close to the N-terminal domain were found to have little protection against exchange. The H/D exchange results for Trp-123, Trp-192 and Trp-209 showed that a previously identified molten globule and the native state gave a similar protection against exchange. Global unfolding of the protein is necessary for the efficient exchange at Trp-97, which is located in the central part of the beta-sheet.
Assuntos
Anidrases Carbônicas/química , Deutério , Indóis , Ressonância Magnética Nuclear Biomolecular , Conformação Proteica , Triptofano , Deutério/química , Humanos , Hidrogênio , Indóis/química , Ressonância Magnética Nuclear Biomolecular/métodos , Desnaturação Proteica , Dobramento de Proteína , Triptofano/químicaRESUMO
A multimerization strategy to improve yields upon recombinant production of the 31-aa human proinsulin C-peptide is presented. Gene fragments encoding the C-peptide were assembled using specific head-to-tail multimerization. DNA constructs encoding one, three or seven copies of the C-peptide gene, fused to a serum albumin binding affinity tag, were expressed intracellularly in Escherichia coli. The three fusion proteins were produced at similar levels (approximately 50 mg/l) and were proteolytically stable during production. Enzymatic digestion by trypsin-carboxypeptidase B treatment of the fusion proteins was shown to efficiently release native C-peptide, as determined by mass spectrometry, reverse-phase chromatography and a radioimmunoassay. The quantitative yields of C-peptide obtained from the three different fusion proteins suggest that this multimerization strategy could provide a cost-efficient production scheme for the C-peptide, and that this strategy could be useful also for production of other recombinant peptides.
Assuntos
Peptídeo C/biossíntese , Peptídeo C/genética , Clonagem Molecular/métodos , Sequência de Aminoácidos , Peptídeo C/isolamento & purificação , Peptídeo C/metabolismo , Carboxipeptidase B , Carboxipeptidases/metabolismo , Cromatografia de Afinidade , DNA , Escherichia coli , Humanos , Dados de Sequência Molecular , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/isolamento & purificação , Proteínas Recombinantes de Fusão/metabolismo , Tripsina/metabolismoRESUMO
The aim of this study was to characterize the morphological changes in the peritoneum following experimental peritoneal dialysis of rats, and to compare the new high-pH and low glucose-derived degradation products (GDP)-level PD fluid PD-Bio with the conventional PD fluid Gambrosol at two different exposure frequencies. Rats were subjected to 10 mL intraperitoneal injections three times per day at 3-hour intervals daytime for 9 days (2 successive weeks, excluding weekends) or once daily for 4 weeks. Untreated animals and animals exposed to Gambrosol or PD-Bio were compared. Biopsy samples were taken from the diaphragm and prepared for light microscopy. Morphometric analysis was used to compare the thickness and the cell density of the sub-mesothelial connective tissue. Intraperitoneal leukocyte numbers were counted. Both fluids induced a significant thickening of the submesothelial connective tissue and an increase in intraperitoneal leukocyte numbers. After exposure three times per day, Gambrosol induced a significantly greater submesothelial thickening than PD-Bio. The submesothelial tissue was more cell-dense after exposure to PD-Bio than after exposure to Gambrosol. The present results indicate that the structural changes of the peritoneum that follow peritoneal dialysis may be dependent upon the chemical composition of the PD fluids.
Assuntos
Soluções para Diálise/química , Glucose/química , Diálise Peritoneal , Peritônio/patologia , Animais , Líquido Ascítico/citologia , Concentração de Íons de Hidrogênio , Contagem de Leucócitos , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
OBJECTIVE: To evaluate the in vivo effects of heat-sterilized peritoneal dialysis (PD) fluids on the respiratory burst response of rat peritoneal leukocytes. DESIGN: Rats were exposed to intraperitoneal injections of a laboratory-made PD fluid that was either heat-sterilized (H-PD) or filtered (F-PD). Control groups of animals were given Hank's buffer (HBSS) or saline (NaCl). Leukocytes were harvested by intraperitoneal lavage at different times in different animals and analyzed with respect to cell numbers, differential counts, and production of superoxide (chemiluminescence) in response to opsonized zymosan. The chemiluminescence responses of the macrophage and the neutrophil populations, respectively, were obtained by curve-fitting techniques from the responses of the mixed populations. RESULTS: All fluids induced a recruitment of neutrophils, the PD fluids causing a cell number increase that was more transient than that caused by NaCl and HBSS. Macrophage numbers were only slightly influenced, but were generally higher after NaCl and HBSS injections than after PD fluid injections. The H-PD exposure induced a significant inhibition of the macrophage chemiluminescence response after 2 and 12 hours, compared with the exposure to F-PD. The neutrophil chemiluminescence response was not significantly affected. CONCLUSION: The toxins produced by heat-sterilization of glucose-containing PD fluids inhibit in vivo the respiratory burst response of peritoneal macrophages.
Assuntos
Soluções para Diálise/efeitos adversos , Glucose/efeitos adversos , Macrófagos Peritoneais/efeitos dos fármacos , Esterilização/métodos , Animais , Soluções Tampão , Contagem de Células , Movimento Celular/efeitos dos fármacos , Filtração , Temperatura Alta , Indicadores e Reagentes , Contagem de Leucócitos , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , Medições Luminescentes , Luminol , Macrófagos Peritoneais/metabolismo , Masculino , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Lavagem Peritoneal , Ratos , Ratos Sprague-Dawley , Explosão Respiratória/efeitos dos fármacos , Cloreto de Sódio , Superóxidos/metabolismo , Zimosan/farmacologiaRESUMO
The refolding reaction of human carbonic anhydrase II has been characterized by use of seven variants in which tryptophan residues have been replaced by Phe or Cys, in each case giving proteins with six tryptophans. Intrinsic tryptophan fluorescence was used to monitor the refolding in the 2 ms-60 s time range, and kinetic traces showing the contributions from each particular tryptophan were obtained by calculation of differences between the wild-type protein and the variants. Earlier assignment [Mårtensson, L.-G., Jonasson, P., Freskgard, P.-O., Svensson, M., Carlsson, U., & Jonsson, B.-H. (1995) Biochemistry 34, 1011-1021] of specific fluorescence properties to each tryptophan, especially regarding energy transfer and intrinsic fluorescence quenching, has made it possible to use the kinetic data to describe the formation of tertiary structure at defined tryptophan residues. In summary, it was found that tertiary structure is formed earlier at those tryptophans that are associated with the central core of beta-strands than at tryptophan residues in the N-terminal minidomain.