RESUMO
Vaccinia virus (VV) egress has been studied using confocal, video, and electron microscopy. Previously, intracellular-enveloped virus (IEV) particles were proposed to induce the polymerization of actin tails, which propel IEV particles to the cell surface. However, data presented support an alternative model in which microtubules transport virions to the cell surface and actin tails form beneath cell-associated enveloped virus (CEV) particles at the cell surface. Thus, VV is unique in using both microtubules and actin filaments for egress. The following data support this proposal. (a) Microscopy detected actin tails at the surface but not the center of cells. (b) VV mutants lacking the A33R, A34R, or A36R proteins are unable to induce actin tail formation but produce CEV and extracellular-enveloped virus. (c) CEV formation is inhibited by nocodazole but not cytochalasin D or 4-amino-5-(4-methylphenyl)-7-(t-butyl)pyrazolo(3,4-d)pyrimidine (PP1). (d) IEV particles tagged with the enhanced green fluorescent protein fused to the VV B5R protein moved inside cells at 60 microm/min. This movement was stop-start, was along defined pathways, and was inhibited reversibly by nocodazole. This velocity was 20-fold greater than VV movement on actin tails and consonant with the rate of movement of organelles along microtubules.
Assuntos
Proteínas de Transporte , Membrana Celular/metabolismo , Membrana Celular/virologia , Endorribonucleases , Peptídeos e Proteínas de Sinalização Intracelular , Microtúbulos/metabolismo , Microtúbulos/virologia , Vaccinia virus/metabolismo , Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Animais , Transporte Biológico Ativo/efeitos dos fármacos , Transporte Biológico Ativo/fisiologia , Linhagem Celular , Membrana Celular/ultraestrutura , Inibidores Enzimáticos/farmacologia , Proteínas de Fluorescência Verde , Humanos , Proteínas Luminescentes/genética , Glicoproteínas de Membrana/genética , Microscopia Confocal , Microscopia Eletrônica , Microscopia de Fluorescência , Microscopia de Contraste de Fase , Microtúbulos/ultraestrutura , Nocodazol/farmacologia , Fosfoproteínas Fosfatases , Fosforilação/efeitos dos fármacos , Proteínas de Ligação a RNA/farmacologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Fatores de Tempo , Tirosina/metabolismo , Vaccinia virus/genética , Vaccinia virus/ultraestrutura , Proteínas do Envelope Viral/genética , Proteínas Estruturais Virais/metabolismoRESUMO
Attention is brought to a unique case of an anterior dislocation of the proximal tibiofibular joint detected 1 month following closed reduction of a posterior knee dislocation. Open reduction and internal fixation were necessary to achieve a stable proximal tibiofibular joint. Additional attention should be paid to the proximal tibiofibular joint when evaluating acute or chronic knee dislocation.
Assuntos
Acidentes de Trânsito , Fíbula , Luxações Articulares/diagnóstico por imagem , Traumatismos do Joelho/diagnóstico por imagem , Motocicletas , Tíbia , Adolescente , Artroscopia , Pinos Ortopédicos , Transplante Ósseo , Seguimentos , Humanos , Luxações Articulares/fisiopatologia , Luxações Articulares/cirurgia , Traumatismos do Joelho/complicações , Traumatismos do Joelho/fisiopatologia , Traumatismos do Joelho/cirurgia , Radiografia , Amplitude de Movimento Articular , Retalhos Cirúrgicos , Fatores de TempoRESUMO
BACKGROUND: The shoulder is the most frequently dislocated joint in the body. Multiple causes and pathologies account for the various types of shoulder instability. Multi-directional instability (MDI) and multi-directional laxity with antero-inferior instability (MDL-AII) are similar in pathology, less common and more difficult to treat. These instabilities are caused by ligamentous capsular redundancy. When non-operative management fails for these patients, quality of life is significantly impaired and surgical treatment is required to tighten the ligaments and joint capsule. The current reference (gold) standard treatment for MDI/MDL-AII is an open inferior capsular shift (ICS) surgical procedure. An alternative treatment involves arthroscopic thermal shrinkage of redundant capsular tissue to tighten the joint. However, there is a lack of scientific evidence to support the use of this technique called, electrothermal arthroscopic capsulorrhaphy (ETAC). This trial will compare the effectiveness of ETAC to open ICS in patients with MDI and MDL-AII, using patient-based quality of life outcome assessments. METHODS: This study is a multi-centre randomized clinical trial with a calculated sample size of 58 patients (p = 0.05, 80% power). Eligible patients are clinically diagnosed with MDI or MDL-AII and have failed standardized non-operative management. A diagnostic shoulder arthroscopy is performed to confirm eligibility, followed by intra-operative randomization to the ETAC or ICS surgical procedure. The primary outcome is the disease-specific quality of life questionnaire (Western Ontario Shoulder Instability Index), measured at baseline, 3, 6, 12 and 24 months. Secondary outcomes include shoulder-specific measures (American Shoulder and Elbow Surgeons Score and Constant Score). Other outcomes include recurrent instability, complications and operative time. The outcome measurements will be compared on an intention-to-treat basis, using two-sample independent t-tests to assess statistical significance. A Generalized Estimated Equations (GEE) analysis will determine whether there is an effect over time. DISCUSSION: This ongoing trial has encountered unexpected operational and practical issues, including slow patient enrollment due to high intra-operative exclusion rates. However, the authors have a greater understanding of multi-directional laxity in the shoulder and anticipate the results of this trial will provide the medical community with the best scientific clinical evidence on the efficacy of ETAC compared to open ICS.
RESUMO
Fifty-seven subjects with shoulder pain, the cause of which was not clear on clinical examination, underwent magnetic resonance imaging (MRI) and arthroscopy. With arthroscopy as the standard of reference, the accuracy of MRI for diagnosing shoulder disease was assessed. Pathologic conditions were organized into 3 groups: (1) labral tears, (2) rotator cuff disease, and (3) other pathologic conditions. At arthroscopic evaluation there were 15 labral tears, 24 rotator cuff tears, and 27 other abnormalities. MRI identified 11 of the labral injuries, with accuracy of 62%, sensitivity 73%, and specificity 58%. Rotator cuff tears were identified by MRI with accuracy of 68%, sensitivity 96%, and specificity 49%. When partial rotator cuff tears were differentiated from complete tears, only 16 of 24 injuries were identified by MRI. Nonrotator cuff, nonlabral disease was identified with accuracy of 75%, sensitivity 63%, and specificity 87%. In our center MRI does not appear to be an accurate effective tool for assessing shoulder pathologic conditions in patients in whom the clinical picture is not clear and therefore may not be of assistance in surgical planning for patients with these difficult conditions.
Assuntos
Artralgia/diagnóstico , Artroscopia , Imageamento por Ressonância Magnética , Articulação do Ombro/patologia , Adulto , Idoso , Feminino , Humanos , Escala de Gravidade do Ferimento , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Lesões do OmbroRESUMO
Thermal denaturation of collagen in ligament tissue has the potential to enhance arthroscopic shoulder stabilization. Previous studies have shown that laser energy produces significant capsular shortening without alteration of viscoelastic properties, but little information is available on the effects of radio frequency electrothermal energy. We assessed the acute effects of radio frequency shrinkage with use of the lapine medial collateral ligament model, in which the tibial insertion was shifted proximally to produce abnormal laxity. Thermal treatment resulted in restoration of laxity from 3.33 +/- 0.25 mm to 0.66 +/- 0.31 mm, which was not significantly different from medial collateral ligaments replaced anatomically (0.50 +/- 0.34 mm). When tested at 4.1 megapascals, cyclic and static creep strains were increased twofold to threefold in thermally-treated ligaments (P <.01), and partial failure occurred in 2 of 8 cases. We conclude that radio frequency electrothermal shrinkage is effective at reducing laxity but significantly alters viscoelastic properties, posing a risk of recurrent stretching-out at "physiological" loads.
Assuntos
Artroscopia/métodos , Ligamentos Colaterais/fisiopatologia , Temperatura Alta , Articulação do Joelho/fisiopatologia , Lasers , Análise de Variância , Animais , Colágeno/química , Feminino , Instabilidade Articular , Modelos Animais , Probabilidade , Coelhos , Distribuição Aleatória , Sensibilidade e EspecificidadeRESUMO
The purposes of this report are (1) to describe the development of a quality-of-life outcome measure for rotator cuff disease (RC-QOL) and a new simple test of rotator cuff function, the Functional Shoulder Elevation Test (FSET), and (2) to compare these with the SF-36 and the American Shoulder and Elbow Surgeons Assessment (ASESA) in a cohort of 86 patients with surgically treated large and massive rotator cuff tears. The RC-QOL is a 34-item self-administered questionnaire that has demonstrated excellent reliability, face validity, and ability to discriminate between large and massive cuff tears; it has also confirmed the construct of higher correlations with the other shoulder outcome measures than with the SF-36. The FSET is a simple test of rotator cuff function that uses 5% body weight and measurement of pain and discomfort on a 100-point visual analog scale. It was highly correlated with the RC-QOL and ASESA and discriminated between large and massive cuff tears. The study achieved an 82% follow-up at an average of 3.5 years. Large cuff tears had statistically significantly improved outcome scores in comparison with massive cuff tears on these 3 outcome measures. The SF-36, however, was not able to discriminate between tear sizes. The RC-QOL and the FSET are recommended outcome measures for rotator cuff disease.
Assuntos
Qualidade de Vida , Lesões do Manguito Rotador , Manguito Rotador/cirurgia , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Recuperação de Função Fisiológica , Manguito Rotador/fisiopatologia , Inquéritos e Questionários , Fatores de TempoRESUMO
We describe a detailed morphological characterization of the endocytic pathway in differentiating chicken erythroblasts transformed by a temperature-sensitive mutant of avian erythroblastosis virus (AEV). These cells express high levels of transferrin receptors (TfR) when induced to differentiate at 42 degrees C. Biochemical analysis showed that most (approximately 90%) of the internalized 125I-Tf recycled within approximately 30 min while a smaller fraction of 125I-Tf required up to 2 h for recycling. By immunocytochemistry, the bulk of Tf and TfR was localized at the plasma membrane and in tubuloreticular early endosomes. This structure contained coated buds that labelled with an antibody specific for the clathrin light chain. Decreasing amounts of both Tf and TfR were detected in two distal compartments, spherical endosome vesicles resembling multivesicular bodies and the prelysosomal compartment (PLC) enriched in cation-independent mannose 6-phosphate receptor. As shown by fluorescent (FITC-Tf) labelling of living cells, the movement of Tf/TfR complex into these late structures was accompanied by a significant drop in pH from about 6, the value displayed by early endosomes, to values below pH 5.0. Since no detectable 125I-Tf degradation was observed during a 4 h period we believe that the Tf/TfR detected in these late endocytic structures avoids degradation and recycles back to the cell surface. The addition of an anti-TfR monoclonal antibody to the culture medium of these cells blocks their differentiation. Under this condition the antibody-TfR complex was trapped in an early endosome compartment that enlarged to more than twice its normal size. However, this condition did not affect the transport kinetics of horseradish peroxidase from the medium to the PLC.
Assuntos
Endocitose/fisiologia , Eritropoese/fisiologia , Transferrina/metabolismo , Alpharetrovirus , Animais , Anticorpos Monoclonais , Compartimento Celular/fisiologia , Linhagem Celular Transformada , Galinhas , Clatrina/metabolismo , Invaginações Revestidas da Membrana Celular/metabolismo , Eritroblastos/metabolismo , Eritroblastos/ultraestrutura , Concentração de Íons de Hidrogênio , Microscopia Eletrônica , Receptores da Transferrina/antagonistas & inibidores , Receptores da Transferrina/imunologia , Receptores da Transferrina/metabolismoRESUMO
The subcellular distribution of the regulatory (RII) subunit of cyclic AMP-dependent protein kinase (PK-A) was analyzed at the electron-microscopical level using thawed cryo-sections of Madin Darby Bovine Kidney (MDBK) cells. The highest density of labelling for RII was found on membranes of the prelysosomal compartment (PLC; marked with the cation-independent mannose 6-phosphate receptor, MPR) and the trans-Golgi network, TGN (at 20 degrees C, marked with the G protein of vesicular stomatitis virus, VSV), as well as in coated buds on the latter. Significant labelling was also localized to the cytoplasmic surface of the plasma membrane, including clathrin-coated pits and microvilli, and to early endosomes (identified using internalized HRP). In contrast, no significant label was seen over the Golgi compartments proximal to the TGN, the endoplasmic reticulum (ER) or over lysosomes. From these results we conclude that PK-A type II is associated with the membranes of precisely those subcellular compartments that are active in endocytosis and recycling of cell surface receptors. We believe these findings to be related to the well-established role of cyclic AMP in signal transduction. In particular, we propose that activation of PK-A in endocytic compartments may contribute to regulation (via phosphorylation) of the subcellular distribution of internalized surface receptors or their functional coupling to effector systems involved in signal propagation.
Assuntos
Proteínas de Transporte/análise , Membrana Celular/química , Endocitose , Peptídeos e Proteínas de Sinalização Intracelular , Animais , Proteínas de Transporte/ultraestrutura , Bovinos , Linhagem Celular , Membrana Celular/ultraestrutura , Complexo de Golgi/química , Complexo de Golgi/ultraestrutura , Lisossomos/química , Lisossomos/ultraestrutura , Receptor IGF Tipo 2 , Receptores de Superfície Celular/metabolismoRESUMO
This study was undertaken to compare the fixation of the semitubular blade plate with that of the AO T plate. Cadaveric humeri (n=12 pairs) from an elderly population (41 to 89 years) had either a blade plate (n=12) or a T plate (n=12) fixed to them, subsequent to which a transverse osteotomy was performed. Mechanical testing in tension was performed in 1 series (n=5, blade plate; n=5, T plate) by applying a single maximal load to failure and in a second series (n=7, blade plate; n=7, T plate) by applying submaximal cyclic loading before failure was induced. Results showed that the fixation provided by the semitubular blade plate was significantly better (P < .05) than that of the T plate in those specimens subjected to submaximal cyclic loading before failure (series 2). This latter testing method contains some component of the clinical situation compared with monotonic distraction to failure. Based on these results plus favorable clinical results reported in the literature, the semitubular blade plate is possibly a better alternative to the T plate in the management of proximal humeral fractures that require open reduction and internal fixation.
Assuntos
Placas Ósseas , Fixação Interna de Fraturas/instrumentação , Fraturas do Ombro/cirurgia , Adulto , Idoso , Idoso de 80 Anos ou mais , Cadáver , Análise de Falha de Equipamento , Humanos , Pessoa de Meia-Idade , Fraturas do Ombro/fisiopatologia , Estresse Mecânico , Resistência à TraçãoRESUMO
The consequence of redirecting the vaccinia virus (VV) B5R protein to the endoplasmic reticulum (ER) has been investigated by the addition of an ER retrieval signal KKSL (K(2)X(2)) to the B5R C-terminus. This mutant B5R gene and a version of the gene with the inactive ER retrieval sequence KKSLAL (K(2)X(4)) were inserted into the thymidine kinase locus of a VV mutant lacking the B5R gene, vDeltaB5R. Similar levels of B5R protein were made by each virus, but the B5R-K(2)X(2) protein remained sensitive to endoglycosidase H and colocalised with protein disulphide isomerase in the ER. In contrast, the B5R-K(2)X(4) protein colocalised with 1, 4-galactosyltransferase in the trans-Golgi network. Electron microscopy revealed that even when the B5R protein was redirected to the ER, intracellular mature virus particles were wrapped by cellular membranes to form intracellular enveloped virus particles, although more incompletely wrapped particles were evident compared with wild type. These intracellular enveloped virus particles were, however, unable to efficiently induce the polymerisation of actin and the plaque size formed by vB5R-K(2)X(2) was small. Nevertheless, the amount and specific infectivity of EEV produced by vB5R-K(2)X(2) were similar to those of wild type, despite the dramatic reduction in the amount of B5R protein present in vB5R-K(2)X(2) EEV.