RESUMO
PURPOSE: Image-free handheld robotic-assisted total knee arthroplasty (RATKA) has shown to achieve desired limb alignment compared to conventional jig-based instrumented total knee arthroplasty (CTKA). The aim of this prospective randomized controlled trial (RCT) was to evaluate the accuracy of a semi-autonomous imageless handheld RATKA compared to CTKA in order to achieve the perioperative planned target alignment of the knee postoperatively. METHODS: Fifty-two patients with knee osteoarthritis were randomized in 1:1 ratio to undergo unilateral CTKA or an imageless handheld RATKA. A full-length lower limb CT-scan was obtained pre- and 6-week postoperative. The primary outcomes were radiologic measurements of achieved target hip-knee-ankle axis (HKA-axis) and implant component position including varus and external rotation and flexion of the femur component, and posterior tibial slope. The proportion of outliers in above radiographic outcomes, defined as > 3° deviation in postoperative CT measurements as compared to perioperative planned target, were also noted. Knee phenotypes were compared with use of the Coronal Plane Alignment of the Knee (CPAK) classification. RESULTS: Baseline conditions were comparable between both groups. The overall proportion and percentage of outliers (n = 38, 24.4% vs n = 9, 5.8%) was statistically significant (p < 0.001) in favor of RATKA. The achieved varus-valgus of the femoral component (varus 1.3° ± 1.7° vs valgus - 0.1° ± 1.9°, p < 0.05) with statistically significant less outliers (0% vs 88.5%, p < 0.01), the achieved HKA-axis (varus 0.4° ± 2.1° vs valgus - 1.2° ± 2.1°, p < 0.05) and the posterior tibial slope (1.4° ± 1.1° vs 3.2° ± 1.8°, p < 0.05) were more accurate with RATKA. The most common postoperative CPAK categories were type II (50% CTKA vs 61.5% RATKA), type I (3.8% CTKA vs 23.1% RATKA) and type V (26.9% CTKA vs 15.4% RATKA). CPAK classification III was only found in CTKA (19.2%). Type VI, VII, VIII, and IX were rare in both populations. CONCLUSIONS: The present trial demonstrates that an imageless handheld RATKA system can be used to accurately perioperatively plan the desired individual component implant positions with less alignment outliers whilst aiming for a constitutional alignment. LEVEL OF EVIDENCE: I.
Assuntos
Artroplastia do Joelho , Prótese do Joelho , Osteoartrite do Joelho , Procedimentos Cirúrgicos Robóticos , Humanos , Articulação do Joelho/diagnóstico por imagem , Articulação do Joelho/cirurgia , Osteoartrite do Joelho/diagnóstico por imagem , Osteoartrite do Joelho/cirurgia , Tomografia Computadorizada por Raios XRESUMO
PURPOSE: Despite widespread adoption of NAVIO robotic-assisted total knee arthroplasty (NAVIO RATKA) in clinical practice, clinical outcome in terms of adverse events and complications remains unclear. The purpose of this study was to compare adverse events, length of stay, surgical time, hemoglobin drop, early readmission rate and revision rate between conventional TKA (CTKA) and NAVIO RATKA. METHODS: This single-centre retrospective cohort analysis compared 230 NAVIO RATKA patients to 489 CTKA patients with a minimal follow-up of 12 months. Baseline demographic and comorbidity parameters were collected, as well as length of stay, revision rate and reason for revision, early readmission rate (< 6w) and reason for readmission, post-operative hemoglobin levels, adverse events, surgical time and operating room time. Data were compared using Mann-Whitney U test for continuous data without normal distribution and ordinal data, categorical variables were compared using the Chi-square or Fisher exact test. RESULTS: There were no clinically relevant baseline demographic or comorbidity differences between groups. CTKA had shorter length of stay than NAVIO RATKA (5.0 days vs 5.4 days, p = 0.010) but trended towards a higher reoperation rate (4.1% vs 1.7%, p = .144, n.s). No differences were found in hemoglobin drop, readmission rate or overall incidence of adverse events, but CTKA showed more hematoma formation (1.6% vs 0%, p = .044) and higher incidence of periprosthetic joint infection (PJI) (1% vs 0%, p = n.s.), whilst NAVIO RATKA showed more periprosthetic fractures and persistent wound drainage (0.4% vs 2.2%, p = .038 and 0.6% vs 4.3%, p = .001, respectively). Surgical time remained significantly longer in NAVIO RATKA during all 230 cases (87 min vs 67.6 min) and showed a continuous downward trend. CONCLUSIONS: This study further validates the usage of NAVIO RATKA as a safe method to perform TKA, with comparable short term outcomes to CTKA in terms of early revisions and adverse events. Surgeons should be mindful of the differing adverse event profile in NAVIO RATKA and adjust their patient selection accordingly to ensure optimal outcomes. In addition, surgeons using NAVIO RATKA should expect a linear learning curve and a surgical time exceeding that of CTKA. LEVEL OF EVIDENCE: Level III (therapeutic retrospective cohort study).
RESUMO
For prosthetic joint infections, antibiotic loaded poly methyl methacrylate (PMMA) spacer or beads can be used to release high concentrations of antibiotics locally at the infection site, while minimizing systemic toxicity. The aim of this study is to determine in vitro and in vivo pharmacokinetic release profile of antibiotics from PMMA spacers and PMMA beads. For the in vitro experiment, the PMMA spacers or beads were submerged in phosphate-buffered saline and gentamicin concentrations were determined from collected specimen at several times points, measured with enzyme-linked immunosorbent assays (ELISA). To assess the in vivo antibiotic release profile of different spacers, wound drainage fluid samples were collected after implantation of a spacer over a period of maximum 14 days. After 48 h, the burst gentamicin concentration elution was 9862 ± 1782 ng/ml (mean ± SD) from spacers versus 38,394 ± 7071 ng/ml (mean ± SD) for beads. Over 35 days, spacers had eluted a cumulative mean concentration of 13,812 ± 3548 versus 55,048 ± 12,006 ng/ml for beads (p < 0.001). Clinical samples of patients with a Vancogenx® spacer showed higher gentamicin release than Refobacin™ spacers (p < 0.001). This is the first study that measured the release data of local antibiotics with ELISA. Compare to spacers, the exact release values of gentamicin from PMMA beads are more than 10 times higher and reached a maximum much later than spacers. This makes the use of PMMA beads more preferable to use for treatment of the infection itself.
Assuntos
Antibacterianos , Infecções Relacionadas à Prótese , Humanos , Antibacterianos/uso terapêutico , Polimetil Metacrilato , Cimentos Ósseos , Gentamicinas/farmacocinética , Drenagem , Ensaio de Imunoadsorção Enzimática , Infecções Relacionadas à Prótese/tratamento farmacológicoRESUMO
Literature into the short-term follow-up of total knee arthroplasty (TKA) using a handheld image- free robotic system are scarce. The purpose of this study was to compare the clinical outcomes and patient-reported outcome measures (PROMs) between patients operated for TKA with an image- free robotic system (robot group) or conventionally TKA (conventional group) 2 years postoperatively. A total of 147 patients were evaluated after TKA, respectively 73 in the robot and 74 in conventional group. Outcome measures included adverse events (AEs), hospital readmission rate, patient satisfaction and the following PROMs: Pain Visual Analogue Score (VAS), Oxford Knee Score (OKS), Forgotten Joint Score Knee (FJS-12) and the EuroQOL-5D (EQ-5D). There were no statistically significant differences in the number of AEs; 8 (10.8%) in the conventional group versus 7 (9.7%) in the robot group. The FJS (p ≤ 0.05) and OKS (p ≤ 0.05) differed statistically in favour of the robot group. The EQ-5D and EQ-5D VAS did not statistically differed between the groups (p=0.231 and p=0.373 respectively). The VAS pain improved statically significant in both groups when comparing the pre- and postoperative values (5.8 points). Patients operated with a handheld image-free robotic system have the ability to forget their artificial knee joint in everyday life as measured with the FJS-12 at short-term follow-up.
Assuntos
Artroplastia do Joelho , Osteoartrite do Joelho , Procedimentos Cirúrgicos Robóticos , Artroplastia do Joelho/efeitos adversos , Humanos , Articulação do Joelho/cirurgia , Osteoartrite do Joelho/cirurgia , Dor/etiologia , Medidas de Resultados Relatados pelo Paciente , Procedimentos Cirúrgicos Robóticos/efeitos adversosRESUMO
Osteochondritis dissecans as a pathology is pre-dominantly described in the knee, elbow and ankle. Osteochondritis dissecans of the humeral head is a more uncommon reported injury. We present a case of a bilateral osteochondritis dissecans of the humeral head in a 16-year-old soccer player and an algorithm for treatment of OCD of the humeral head. To our knowledge this has never been described so specifically in literature before.
Assuntos
Articulação do Cotovelo , Osteocondrite Dissecante , Humanos , Adolescente , Ombro , Osteocondrite Dissecante/diagnóstico por imagem , Osteocondrite Dissecante/cirurgia , Cabeça do Úmero , Articulação do Joelho/patologiaRESUMO
Although the individuality of whole-body movements has been suspected for years, the scientific proof and systematic investigation that individuals possess unique movement patterns did not manifest until the introduction of the criteria of uniqueness and persistence from the field of forensic science. Applying the criteria of uniqueness and persistence to the individuality of motor learning processes requires complex strategies due to the problem of persistence in the learning processes. One approach is to examine the learning process of different movements. For this purpose, it is necessary to differentiate between two components of movement patterns: the individual-specific component and the discipline-specific component. To this end, a kinematic analysis of the shot put, discus, and javelin throwing movements of seven high-performance decathletes during a qualification competition was conducted. In total, joint angle waveforms of 57 throws formed the basis for the recognition task of individual- and discipline-specific throwing patterns using a support vector machine. The results reveal that the kinematic throwing patterns of the three disciplines could be distinguished across athletes with a prediction accuracy of up to 100% (57 of 57 throws). In addition, athlete-specific throwing characteristics could also be identified across the three disciplines. Prediction accuracies of up to 52.6% indicated that up to 10 out of 19 throws of a discipline could be assigned to the correct athletes, based on only knowing these athletes from the kinematic throwing patterns in the other two disciplines. The results further suggest that individual throwing characteristics across disciplines are more pronounced in shot put and discus throwing than in javelin throwing. Applications for training and learning practice in sports and therapy are discussed. In summary, the chosen approach offers a broad field of application related to the search of individualized optimal movement solutions in sports.
RESUMO
PURPOSE: The aim of this study was to develop and internally validate a multivariable model for accurate prediction of surgical site infection (SSI) after instrumented spine surgery using a large cohort of a Western European academic center. METHOD: Data of potential predictor variables were collected in 898 adult patients who underwent instrumented posterior fusion of the thoracolumbar spine. We used logistic regression analysis to develop the prediction model for SSI. The ability to discriminate between those who developed SSI and those who did not was quantified as the area under the receiver operating characteristic curve (AUC). Model calibration was evaluated by visual inspection of the calibration plot and by computing the Hosmer and Lemeshow goodness-of-fit test. RESULTS: Sixty patients (6.7%) were diagnosed with an SSI. After backward stepwise elimination of predictor variables, we formulated a model in which an individual's risk of an SSI can be computed. Age, body mass index, ASA score, degenerative or revision surgery and NSAID use appeared to be independent predictor variables for the risk of SSI. The AUC was 0.72 (95% CI 0.65-0.79), indicating reasonable discriminative ability. CONCLUSIONS: We developed and internally validated a prediction model for SSI after instrumented thoracolumbar spine surgery using predictor variables of standard clinical practice that showed reasonable discriminative ability and calibration. Identification of patients at risk for SSI allows for individualized patient risk assessment with better patient-specific counseling and may accelerate the implementation of multi-disciplinary strategies for reduction of SSI. These slides can be retrieved under Electronic Supplementary Material.
Assuntos
Vértebras Lombares/cirurgia , Medição de Risco/métodos , Infecção da Ferida Cirúrgica/diagnóstico , Vértebras Torácicas/cirurgia , Adulto , Idoso , Índice de Massa Corporal , Estudos de Coortes , Feminino , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Curva ROC , Reoperação , Estudos Retrospectivos , Fatores de Risco , Infecção da Ferida Cirúrgica/etiologiaRESUMO
Background: There is no generally established treatment algorithm for the management of surgical site infection (SSI) and non-union after instrumented spinal surgery. In contrast to infected hip- and knee- arthroplasties, the use of a local gentamicin impregnated carrier in spinal surgery has not been widely reported in literature. Patients and methods: We studied 48 deep SSI and non-union patients after instrumented spine surgery, treated between 1999 and 2016. The minimum follow-up was 1.5 years. All infections were treated with a treatment-regimen consisting of systemic antibiotics and repetitive surgical debridement, supplemented with local gentamicin releasing carriers. We analysed the outcome of this treatment regimen with regard to healing of the infection, as well as patient- and surgery-characteristics of failed and successfully treated patients. Results: 42 of the 48 (87.5%) patients showed successful resolution of the SSI without recurrence with a stable spine at the end of treatment. 36 patients' SSI were treated with debridement, local antibiotics, and retention or eventual restabilization of the instrumentation in case of loosening. 3 patients were treated without local antibiotics because of very mild infection signs during the revision operation. 3 patients were treated with debridement, local antibiotics and removal of instrumentation. One of these patients was restabilized in a second procedure. Infection persisted or recurred in 6 patients. These patients had a worse physical status with a higher ASA-score. Staphylococcus aureus was the most frequent causative microorganism. Interpretation: Debridement and retention of the instrumentation, in combination with systemic antibiotics and the addition of local antibiotics provided a successful treatment for SSI and non-union after instrumented spinal fusion.
RESUMO
BACKGROUND: A prediction model for surgical site infection (SSI) after spine surgery was developed in 2014 by Lee et al. This model was developed to compute an individual estimate of the probability of SSI after spine surgery based on the patient's comorbidity profile and invasiveness of surgery. Before any prediction model can be validly implemented in daily medical practice, it should be externally validated to assess how the prediction model performs in patients sampled independently from the derivation cohort. METHODS: We included 898 consecutive patients who underwent instrumented thoracolumbar spine surgery. To quantify overall performance using Nagelkerke's R2 statistic, the discriminative ability was quantified as the area under the receiver operating characteristic curve (AUC). We computed the calibration slope of the calibration plot, to judge prediction accuracy. RESULTS: Sixty patients developed an SSI. The overall performance of the prediction model in our population was poor: Nagelkerke's R2 was 0.01. The AUC was 0.61 (95% confidence interval (CI) 0.54-0.68). The estimated slope of the calibration plot was 0.52. CONCLUSIONS: The previously published prediction model showed poor performance in our academic external validation cohort. To predict SSI after instrumented thoracolumbar spine surgery for the present population, a better fitting prediction model should be developed.
Assuntos
Vértebras Lombares/cirurgia , Modelos Teóricos , Infecção da Ferida Cirúrgica/diagnóstico , Infecção da Ferida Cirúrgica/epidemiologia , Vértebras Torácicas/cirurgia , Adulto , Idoso , Estudos de Coortes , Europa (Continente)/epidemiologia , Feminino , Previsões , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Fatores de RiscoRESUMO
Background and purpose - A 2-stage revision is the most common treatment for late deep prosthesis-related infections and in all cases of septic loosening. However, there is no consensus about the optimal interval between the 2 stages. Patients and methods - We retrospectively studied 120 deep infections of total hip (n = 95) and knee (n = 25) prostheses that had occurred over a period of 25 years. The mean follow-up time was 5 (2-20) years. All infections had been treated with extraction, 1 or more debridements with systemic antibiotics, and implantation of gentamicin-PMMA beads. There had been different time intervals between extraction and reimplantation: median 14 (11-47) days for short-term treatment with uninterrupted hospital stay, and 7 (3-22) months for long-term treatment with temporary discharge. We analyzed the outcome regarding resolution of the infection and clinical results. Results - 88% (105/120) of the infections healed, with no difference in healing rate between short- and long-term treatment. 82 prostheses were reimplanted. In the most recent decade, we treated patients more often with a long-term treatment but reduced the length of time between the extraction and the reimplantation. More reimplantations were performed in long-term treatments than in short-term treatments, despite more having difficult-to-treat infections with worse soft-tissue condition. Interpretation - Patient, wound, and infection considerations resulted in an individualized treatment with different intervals between stages. The 2-stage revision treatment in combination with local gentamicin-PMMA beads gave good results even with difficult prosthesis infections and gentamicin-resistant bacteria.
Assuntos
Artroplastia de Quadril/efeitos adversos , Artroplastia do Joelho/efeitos adversos , Previsões , Gentamicinas/administração & dosagem , Prótese de Quadril/efeitos adversos , Prótese do Joelho/efeitos adversos , Polimetil Metacrilato , Infecções Relacionadas à Prótese/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/administração & dosagem , Desbridamento/métodos , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Infecções Relacionadas à Prótese/diagnóstico , Reoperação , Estudos Retrospectivos , Resultado do TratamentoRESUMO
BACKGROUND: Hallux valgus deformity is a common musculoskeletal foot disorder with a prevalence of 3.5% in adolescents to 35.7% in adults aged over 65 years. Radiographic measurements of hallux valgus angles (HVA) are considered to be the most reproducible and accurate assessment of HVA. However, in European countries, many podiatrists do not have direct access to radiographic facilities. Therefore, alternative measurements are desired. Such measurements are computerised plantar pressure measurement and clinical goniometry. The purpose of this study was to establish the agreement of these techniques and radiographic assessments. METHODS: HVA was determined in one hundred and eighty six participants suffering from diabetes. Radiographic measurements of HVA were performed with standardised static weight bearing dorsoplantar foot radiographs. The clinical goniometry for HVA was measured with a universal goniometer. Computerised plantar pressure measurement for HVA was executed with the EMED SF-4® pressure platform and Novel-Ortho-Geometry software. The intra-class correlation coefficients (ICC) and levels of agreement were analysed using Bland & Altman plots. RESULTS: Comparison of radiographic measurements to clinical goniometry for HVA showed an intraclass correlation coefficient (ICC) of 0.81 (95% confidence interval, 0.76 to 0.86; p<0.001). Radiographic measurement versus computerised plantar pressure measurement showed an ICC of 0.59 (95% confidence interval, 0.49 to 0.68; p<0.001). In addition, clinical goniometry versus computerised plantar pressure measurement showed an ICC of 0.77 (95% confidence interval, 0.70 to 0.82; p<0.001). The systematic difference of the computerised plantar pressure measurement compared with radiographic measurement and clinical goniometry was 7.0 degrees (SD 6.8) and 5.2 degrees (SD 5.0), respectively. The systemic difference of radiographic measurements compared with clinical goniometry was 1.8 degrees (SD 5.0). CONCLUSIONS: The agreement of computerised plantar pressure measurement and clinical goniometry for HVA compared to radiographic measurement of HVA is unsatisfactory. Radiographic measurements of HVA and clinical goniometry for HVA yield better agreement compared to radiographic measurements and computerised plantar pressure measurement. The traditional radiographic measurement techniques are strongly recommended for the assessment of HVA.
RESUMO
The achievement of mastery in playing a composition by means of a musical instrument typically requires numerous repetitions and corrections according to the keys and notations of the music piece. Nevertheless, differences in the interpretation of the same music piece by highly skilled musicians seem to be recognizable. The present study investigated differences within and between skilled flute players in their finger and body movements playing the same piece several times on the same and on different days. Six semiprofessional and four professional musicians played an excerpt of Mozart's Flute Concerto No. 2 several times on three different days. Finger and body movements were recorded by 3D motion capture and analyzed by linear and nonlinear classification approaches. The findings showed that the discrete and continuous movement timing data correctly identified individuals up to 100% by means of their finger movements and up to 94% by means of their body movements. These robust examples of identifying individual movement patterns contradict the prevailing models of small, economic finger movements that are favored in the didactic literature for woodwind players and question traditional recommendations for teaching the learning of motor skills.
Assuntos
Dedos/fisiologia , Individualidade , Atividade Motora/fisiologia , Música , Postura/fisiologia , Desempenho Psicomotor/fisiologia , Adulto , Algoritmos , Aptidão/fisiologia , Fenômenos Biomecânicos/fisiologia , Feminino , Humanos , Articulações/fisiologia , Masculino , Pessoa de Meia-Idade , Redes Neurais de Computação , Psicofisiologia , Adulto JovemRESUMO
BACKGROUND: Deep postoperative and hematogenous prosthesis infections may be treated with retention of the prosthesis, if the prosthesis is stable. How long the infection may be present to preclude a good result is unclear. PATIENTS AND METHODS: We retrospectively studied 89 deep-infected stable prostheses from 69 total hip replacements and 20 total knee replacements. There were 83 early or delayed postoperative infections and 6 hematogenous. In the postoperative infections, treatment had started 12 days to 2 years after implantation. In the hematogenous infections, symptoms had been present for 6 to 9 days. The patients had been treated with debridement, prosthesis retention, systemic antibiotics, and local antibiotics: gentamicin-PMMA beads or gentamicin collagen fleeces. The minimum follow-up time was 1.5 years. We investigated how the result of the treatment had been influenced by the length of the period the infection was present, and by other variables such as host characteristics, infection stage, and type of bacteria. RESULTS: In postoperative infections, the risk of failure increased with a longer postoperative interval: from 0.2 (95% CI: 0.1-0.3) if the treatment had started ≥ 4 weeks postoperatively to 0.5 (CI: 0.2-0.8) if it had started at ≥ 8 weeks. The relative risk for success was 0.6 (CI: 0.3-0.95) if the treatment had started ≥ 8 weeks. In the hematogenous group, 5 of 6 infections had been treated successfully. INTERPRETATION: A longer delay before the start of the treatment caused an increased failure rate, but this must be weighed against the advantage of keeping the prosthesis. We consider a failure rate of < 50% to be acceptable, and we therefore advocate keeping the prosthesis for up to 8 weeks postoperatively, and in hematogenous infections with a short duration of symptoms.
Assuntos
Antibacterianos/administração & dosagem , Infecções Bacterianas/tratamento farmacológico , Prótese de Quadril/efeitos adversos , Prótese do Joelho/efeitos adversos , Infecções Relacionadas à Prótese/tratamento farmacológico , Administração Tópica , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/uso terapêutico , Artroplastia de Quadril , Artroplastia do Joelho , Infecções Bacterianas/microbiologia , Infecções Bacterianas/cirurgia , Terapia Combinada , Desbridamento , Portadores de Fármacos , Feminino , Gentamicinas/administração & dosagem , Gentamicinas/uso terapêutico , Prótese de Quadril/microbiologia , Humanos , Prótese do Joelho/microbiologia , Masculino , Metilmetacrilatos/administração & dosagem , Metilmetacrilatos/uso terapêutico , Pessoa de Meia-Idade , Período Pós-Operatório , Falha de Prótese , Infecções Relacionadas à Prótese/microbiologia , Infecções Relacionadas à Prótese/cirurgia , Estudos Retrospectivos , Medição de Risco/métodos , Resultado do TratamentoRESUMO
A 74-year-old woman presented with a nonreducible, painful bulge of 10 cm in the right groin-region, which had been present for 5 days. She was not feeling ill. During the CT-scan the patient performed a Valsalva maneuver. The scan demonstrated a right femoral hernia with incarcerated peritoneal fat, infiltrate and an inflamed appendix. Peroperative findings confirmed the diagnosis "hernia De Garengeot".
Assuntos
Apendicite/diagnóstico , Hérnia Femoral/diagnóstico , Idoso , Apendicite/cirurgia , Diagnóstico Diferencial , Feminino , Virilha/diagnóstico por imagem , Virilha/patologia , Hérnia Femoral/cirurgia , Humanos , Radiografia , Manobra de ValsalvaRESUMO
BACKGROUND: Quantitative real-time PCR (qPCR) is a commonly used technique to quantify gene expression levels. Validated normalization is essential to obtain reliable qPCR data. In that context, normalizing to multiple reference genes has become the most popular method. However, expression of reference genes may vary per tissue type, developmental stage and in response to experimental treatment. It is therefore imperative to determine stable reference genes for a specific sample set and experimental model. The present study was designed to validate potential reference genes in hippocampal tissue from rats that had experienced early-life febrile seizures (FS). To this end, we applied an established model in which FS were evoked by exposing 10-day old rat pups to heated air. One week later, we determined the expression stability of seven frequently used reference genes in the hippocampal dentate gyrus. RESULTS: Gene expression stability of 18S rRNA, ActB, GusB, Arbp, Tbp, CycA and Rpl13A was tested using geNorm and Normfinder software. The ranking order of reference genes proposed by geNorm was not identical to that suggested by Normfinder. However, both algorithms indicated CycA, Rpl13A and Tbp as the most stable genes, whereas 18S rRNA and ActB were found to be the least stably expressed genes. CONCLUSIONS: Our data demonstrate that the geometric averaging of at least CycA, Rpl13A and Tbp allows reliable interpretation of gene expression data in this experimental set-up. The results also show that ActB and 18S rRNA are not suited as reference genes in this model.
Assuntos
Ciclofilina A/genética , Giro Denteado/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas Ribossômicas/genética , Convulsões Febris/genética , Software , Proteína de Ligação a TATA-Box/genética , Ar , Algoritmos , Animais , Animais Recém-Nascidos , Expressão Gênica , Perfilação da Expressão Gênica , Genes Essenciais , Temperatura Alta , Masculino , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real/normas , Padrões de ReferênciaRESUMO
PURPOSE: Febrile seizures (FS), the most frequent seizure type during childhood, have been linked to temporal lobe epilepsy (TLE) in adulthood. Yet, underlying mechanisms are still largely unknown. Altered γ-aminobutyric acid (GABA)ergic neurotransmission in the dentate gyrus (DG) circuit has been hypothesized to be involved. This study aims at analyzing whether experimental FS change inhibitory synaptic input and postsynaptic GABA(A) R function in dentate granule cells. METHODS: We applied an immature rat model of hyperthermia (HT)-induced FS. GABA(A) R-mediated neurotransmission was studied using whole-cell patch-clamp recordings from dentate granule neurons in hippocampal slices within 6-9 days post-HT. KEY FINDINGS: Frequencies of spontaneous inhibitory postsynaptic currents (sIPSCs) were reduced in HT rats that had experienced seizures, whereas sIPSC amplitudes were enhanced. Whole-cell GABA responses revealed a doubled GABA(A) R sensitivity in dentate granule cells from HT animals, compared to that of normothermic (NT) controls. Analysis of sIPSCs and whole-cell GABA responses showed similar kinetics in postsynaptic GABA(A) Rs of HT and NT rats. quantitative real-time polymerase chain reaction (qPCR) experiments indicated changes in DG GABA(A) R subunit expression, which was most pronounced for the α3 subunit. SIGNIFICANCE: The data support the hypothesis that FS persistently alter neuronal excitability.
Assuntos
Giro Denteado/fisiologia , Receptores de GABA-A/fisiologia , Convulsões Febris/fisiopatologia , Transmissão Sináptica/fisiologia , Fatores Etários , Animais , Potenciais Pós-Sinápticos Inibidores/fisiologia , Masculino , Ratos , Ratos Sprague-Dawley , Ácido gama-Aminobutírico/fisiologiaRESUMO
The spatio-temporal membrane behavior of glycine receptors (GlyRs) is known to be of influence on receptor homeostasis and functionality. In this work, an elaborate fluorimetric strategy was applied to study the GlyR α3K and L isoforms. Previously established differential clustering, desensitization and synaptic localization of these isoforms imply that membrane behavior is crucial in determining GlyR α3 physiology. Therefore diffusion and aggregation of homomeric α3 isoform-containing GlyRs were studied in HEK 293 cells. A unique combination of multiple diffraction-limited ensemble average methods and subdiffraction single particle techniques was used in order to achieve an integrated view of receptor properties. Static measurements of aggregation were performed with image correlation spectroscopy (ICS) and, single particle based, direct stochastic optical reconstruction microscopy (dSTORM). Receptor diffusion was measured by means of raster image correlation spectroscopy (RICS), temporal image correlation spectroscopy (TICS), fluorescence recovery after photobleaching (FRAP) and single particle tracking (SPT). The results show a significant difference in diffusion coefficient and cluster size between the isoforms. This reveals a positive correlation between desensitization and diffusion and disproves the notion that receptor aggregation is a universal mechanism for accelerated desensitization. The difference in diffusion coefficient between the clustering GlyR α3L and the non-clustering GlyR α3K cannot be explained by normal diffusion. SPT measurements indicate that the α3L receptors undergo transient trapping and directed motion, while the GlyR α3K displays mild hindered diffusion. These findings are suggestive of differential molecular interaction of the isoforms after incorporation in the membrane.
Assuntos
Membrana Celular/metabolismo , Receptores de Glicina/química , Receptores de Glicina/metabolismo , Linhagem Celular , Células HEK293 , Humanos , Isoformas de Proteínas/química , Análise Espectral/métodosRESUMO
The inhibitory neurotransmitter glycine is known to enhance microglial nitric oxide production. However, up to now, the mechanism is undocumented. Since calcium is an important second messenger in both immune and glial cells, we studied the effects of glycine on intracellular calcium signaling. We found that millimolar concentrations of glycine enhance microglial intracellular calcium transients induced by 100 µM ATP or by 500 nM thapsigargin. This modulation was unaffected by the glycine receptor antagonist strychnine and could not be mimicked by glycine receptor agonists such as taurine or ß-alanine, indicating glycine receptor independency. The modulation of calcium responses could be mimicked by several structurally related amino acids (e.g., serine, alanine, or glutamine) and was inhibited in the presence of the neutral amino acid transporter substrate α-aminoisobutyric acid (AIB). We correlated these findings to immunofluorescence glycine uptake experiments which showed a clear glycine uptake which was inhibited by AIB. Furthermore, all amino acids that were shown to modulate calcium responses also evoked AIB-sensitive inward currents, mainly carried by sodium, as demonstrated by patch clamp experiments. Based on these findings, we propose that sodium-coupled neutral amino acid transporters are responsible for the observed glycine modulation of intracellular calcium responses.
Assuntos
Sistema A de Transporte de Aminoácidos/fisiologia , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/fisiologia , Glicina/farmacologia , Microglia/efeitos dos fármacos , Microglia/fisiologia , Ácidos Aminoisobutíricos/farmacologia , Animais , Linhagem Celular , Glicina/metabolismo , Glicinérgicos/farmacologia , Camundongos , Modelos Animais , Técnicas de Patch-Clamp , Receptores de Glicina/agonistas , Receptores de Glicina/antagonistas & inibidores , Receptores de Glicina/fisiologia , Estricnina/farmacologia , Taurina/farmacologia , beta-Alanina/farmacologiaRESUMO
The aim of the study was to train and test support vector machines (SVM) and self-organizing maps (SOM) to correctly classify gait patterns before, during and after complete leg exhaustion by isokinetic leg exercises. Ground reaction forces were derived for 18 gait cycles on 9 adult participants. Immediately before the trials 7-12, participants were required to completely exhaust their calves with the aid of additional weights (44.4±8.8kg). Data were analyzed using: (a) the time courses directly and (b) only the deviations from each individual's calculated average gait pattern. On an inter-individual level the person recognition of the gait patterns was 100% realizable. Fatigue recognition was also highly probable at 98.1%. Additionally, applied SOMs allowed an alternative visualization of the development of fatigue in the gait patterns over the progressive fatiguing exercise regimen.
Assuntos
Marcha/fisiologia , Fadiga Muscular/fisiologia , Máquina de Vetores de Suporte , Adulto , Fenômenos Biomecânicos , Humanos , Individualidade , Masculino , Dinâmica não Linear , Reconhecimento Automatizado de Padrão , Levantamento de Peso/fisiologia , Adulto JovemRESUMO
In insects, inhibitory neurotransmission is generally associated with members of the cys-loop ligand-gated anion channels, such as the glutamate-gated chloride channel (GluCl), the GABA-gated chloride channels (GABACl), and the histamine-gated chloride channels (HisCl). These ionotropic receptors are considered established target sites for the development of insecticides, and therefore it is necessary to obtain a better insight in their distribution, structure, and functional properties. Here, by combining electrophysiology and molecular biology techniques, we identified and characterized GluCl, GABACl, and HisCl in dorsal unpaired median (DUM) neurons of Locust migratoria. In whole cell patch-clamp recordings, application of glutamate, GABA, or histamine induced rapidly activating ionic currents. GluCls were sensitive to ibotenic acid and blocked by picrotoxin and fipronil. The pharmacological profile of the L. migratoria GABACl fitted neither the vertebrate GABA(A) nor GABA(C) receptor and was similar to the properties of the cloned Drosophila melanogaster GABA receptor subunit (Rdl). The expression of Rdl-like subunit-containing GABA receptors was shown at the molecular level using RT-PCR. Sequencing analysis indicated that the orthologous GABACl of D. melanogaster CG10357-A is expressed in DUM neurons of L. migratoria. Histamine-induced currents exhibited a fast onset and desensitized completely on continuous application of histamine. In conclusion, within the DUM neurons of L. migratoria, we identified three different cys-loop ligand-gated anion channels that use GABA, glutamate, or histamine as their neurotransmitter.