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1.
Eur Spine J ; 26(6): 1782-1788, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-27942937

RESUMO

PURPOSE: The minimal clinically important difference (MCID) of the Appearance domain of the SRS-22 questionnaire is an increase ≥1.0 in surgically treated patients with adolescent idiopathic scoliosis (AIS). However, no study has sought to identify the factors associated with an SRS-22 Appearance score increase greater than the MCID at 2 years. METHODS: A retrospective analysis was performed on a prospectively collected multicenter database of 1020 surgically treated AIS patients with a minimum 2-year follow-up. Patients were divided into two cohorts: "I" = Improved after surgery (Δ Appearance ≥1.0) and "NI" = Not improved after surgery (Δ Appearance <1.0). Univariate regression was used to find a significant difference between the cohorts for individual measures. Multivariate logistic regression was used to find continuous predictors. RESULTS: 663 (65%) patients were improved greater than the MCID, and 357 were not improved (35%). The improved cohort trended toward a greater percentage of underweight patients (p = 0.074) with lower preoperative SRS Appearance scores (p < 0.001) and larger preoperative trunk shifts (p = 0.033). Postoperatively, those patients with greater percent correction of thoracic (p = 0.021) and lumbar (p = 0.003) Cobb angles, smaller apical lumbar translation (p = 0.006), and a greater correction in trunk shift (p = 0.003) were most likely to attain the MCID. CONCLUSION: Several factors influence which patients are most likely to attain the MCID following surgery for AIS. Factors such as preoperative appearance scores and body weight are patient specific; other factors such as percent correction of the thoracic and lumbar Cobb angles, trunk shift, and lumbar apical translation may be influenced by the surgeon. LEVEL OF EVIDENCE: II.


Assuntos
Diferença Mínima Clinicamente Importante , Escoliose/cirurgia , Adolescente , Peso Corporal , Feminino , Humanos , Masculino , Análise Multivariada , Estudos Retrospectivos
2.
Eur Spine J ; 24(7): 1533-9, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25510515

RESUMO

PURPOSE: This retrospective chart review evaluates the clinical and radiographic outcomes of anterior vertebral body tethering (VBT) at 1-year follow-up. Anterior VBT offers a fusionless treatment option for skeletally immature patients with adolescent idiopathic scoliosis. It is a growth-modulation technique, which utilizes patients' growth to attain progressive scoliosis correction. Numerous animal models support its promise; however, clinical data remain sparse. METHODS: Clinical and radiographic data were retrospectively analyzed. We reviewed 32 patients who underwent thoracic VBT with a minimum one-year follow-up. Pertinent clinical and radiographic data were collected. ANOVA, Student's t test and Fisher's exact test were utilized to compare different time points. RESULTS: 32 patients with thoracic idiopathic scoliosis (72 % female) with a minimum one-year follow-up were identified; mean age at surgery was 12 years. All patients were considered skeletally immature pre-operatively; mean Risser score 0.42, mean Sanders score 3.2. Patients underwent tethering of an average of 7.7 levels (range 7-11). Median blood loss was 100 cc. The mean pre-operative thoracic curve magnitude was 42.8° ± 8.0° which corrected to 21.0° ± 8.5° on first erect and 17.9° ± 11.4° at most recent. The pre-operative lumbar curve of 25.2° ± 7.3° demonstrated progressive correction (first erect = 18.0° ± 7.1°, 1 year = 12.6° ± 9.4°, p < 0.00001). Thoracic axial rotation measured 13.4° pre-operatively and 7.4° at the most recent measurement (p < 0.00001). One patient experienced prolonged atelectasis which required a bronchoscopy; otherwise, no major complications were observed. CONCLUSIONS: Our early results indicate that anterior VBT is a safe and potentially effective treatment option for skeletally immature patients with idiopathic scoliosis. These patients experienced an improvement of their scoliosis with minimal major complications. However, longer term follow-up of this cohort will reveal the true benefits of this promising technique. LEVEL OF EVIDENCE: IV.


Assuntos
Procedimentos Ortopédicos/métodos , Escoliose/cirurgia , Vértebras Torácicas/cirurgia , Adolescente , Criança , Estudos de Coortes , Feminino , Humanos , Masculino , Radiografia , Estudos Retrospectivos , Escoliose/diagnóstico por imagem , Vértebras Torácicas/diagnóstico por imagem , Resultado do Tratamento
3.
Spine Deform ; 2024 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-38997612

RESUMO

INTRODUCTION: The evolution of MCGR technique has led to modifications in the configuration of the proximal construct to decrease the incidence of implant-related complications (IRC) and revision surgeries. However, there is no data characterizing the performance of the most used configurations reducing the risk of complications. METHODS: 487 patients were identified from an international multicenter EOS database. INCLUSION CRITERIA: EOS patients, primary dual MCGR, complete radiographs, and minimum of 2-year follow-up. 76 patients had incomplete X-rays, 5 had apical fusions, and 18 had inconclusive complications, leaving 388 patients for review. A digital spine template was created to document UIV; number of levels; number, type, and location of anchors; as well as implant configuration. First available postoperative and latest follow-up radiographs were reviewed by two senior surgeons and two spine fellows. UPROR due to IRC was defined as any change in proximal anchors between the postoperative and final follow-up radiographs. RESULTS: The most common proximal construct configuration: UIV at T2 (50.0%) with 17.5% UPROR, followed by T3 (34.0%) with 12.1% UPROR; number of levels was three (57.1%) with 16.8% UPROR and two (26.0%) with 17.0% UPROR; number of proximal anchors was six (49.9%) with 14.1% UPROR and four (27.0%) with 18.3% UPROR. The most common anchors were all screws (42.0%) with 9.9% UPROR, and all hooks (26.4%) with 31.4% UPROR (P < 0.001). The construct with the lowest rate of UPROR was a UIV at T2, with six anchors (all screws) across three levels (42 cases), with 0% UPROR. Other construct combinations that yielded 0% UPROR rates were UIV of T3, six anchors (all screws) across three levels (25 cases), and a UIV of T3 with six anchors (screws and hooks) across three3 levels (9 cases). CONCLUSION: Proximal anchor configuration impacts the incidence of UPROR due to IRC in MCGR. UIV at T2 and T3 compared to T4, and the use of all screws or combination of screws and hooks compared to all hooks were associated with a lower UPROR rate. The most common construct configuration was T2 UIV, three levels, six anchors, and all screws. The use of a combination of six anchors (screws or screws and hooks) across three levels with a UIV at T2 or T3 was associated with a lower UPROR rate. Additional research is needed to further evaluate the variables contributing to configuration selection and their association with IRC.

4.
J Environ Qual ; 42(3): 704-12, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23673937

RESUMO

There is currently much interest in developing crop management practices that will decrease NO emissions from agricultural soils. Many different approaches are being investigated, but to date, no studies have been published on how microbial inoculants affect NO emissions. This study was conducted to test the hypothesis that microbial-based inoculants known to promote root growth and nutrient uptake can reduce NO emissions in the presence of N fertilizers under controlled conditions. Carbon dioxide and CH fluxes were also measured to evaluate microbial respiration and determine the aerobic and anaerobic conditions of the incubated soil. The microbial-based treatments investigated were SoilBuilder (SB), a metabolite extract of SoilBuilder (SBF), and a mixture of four strains of plant growth-promoting spp. Experiments included two different N fertilizer treatments, urea and urea-NHNO 32% N (UAN), and an unfertilized control. Emissions of NO and CO were determined from soil incubations and analyzed with gas chromatography. After 29 d of incubation, cumulative NO emissions were reduced 80% by SB and 44% by SBF in soils fertilized with UAN. Treatment with spp. significantly reduced NO production on Days 1 and 2 of the incubation in soils fertilized with UAN. In the unfertilized treatment, cumulative emissions of NO were significantly reduced 92% by SBF. Microbial-based treatments did not reduce NO emissions associated with urea application. Microbial-based treatments increased CO emissions from soils fertilized with UAN, suggesting a possible increase in microbial activity. Overall, the results demonstrated that microbial-based inoculants can reduce NO emissions associated with N fertilizer application, and this response varies with the type of microbial-based inoculant and fertilizer.


Assuntos
Fertilizantes , Óxido Nitroso , Inoculantes Agrícolas , Dióxido de Carbono , Metano , Nitrogênio/química , Óxido Nitroso/química , Solo/química , Ureia
5.
PeerJ ; 11: e16215, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37872950

RESUMO

Illegal, unreported, and unregulated (IUU) fishing is a major problem worldwide, often made more challenging by a lack of at-sea and shoreside monitoring of commercial fishery catches. Off the US West Coast, as in many places, a primary concern for enforcement and management is whether vessels are illegally fishing in locations where they are not permitted to fish. We explored the use of supervised machine learning analysis in a partially observed fishery to identify potentially illicit behaviors when vessels did not have observers on board. We built classification models (random forest and gradient boosting ensemble tree estimators) using labeled data from nearly 10,000 fishing trips for which we had landing records (i.e., catch data) and observer data. We identified a set of variables related to catch (e.g., catch weights and species) and delivery port that could predict, with 97% accuracy, whether vessels fished in state versus federal waters. Notably, our model performances were robust to inter-annual variability in the fishery environments during recent anomalously warm years. We applied these models to nearly 60,000 unobserved landing records and identified more than 500 instances in which vessels may have illegally fished in federal waters. This project was developed at the request of fisheries enforcement investigators, and now an automated system analyzes all new unobserved landings records to identify those in need of additional investigation for potential violations. Similar approaches informed by the spatial preferences of species landed may support monitoring and enforcement efforts in any number of partially observed, or even totally unobserved, fisheries globally.


Assuntos
Conservação dos Recursos Naturais , Pesqueiros , Animais , Humanos , Caça , Pesquisadores , Aprendizado de Máquina
6.
Protein Expr Purif ; 75(1): 1-14, 2011 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-20692346

RESUMO

Multi-protein complexes are involved in essentially all cellular processes. A protein's function is defined by a combination of its own properties, its interacting partners, and the stoichiometry of each. Depending on binding partners, a transcription factor can function as an activator in one instance and a repressor in another. The study of protein function or malfunction is best performed in the relevant context. While many protein complexes can be reconstituted from individual component proteins after being produced individually, many others require co-expression of their native partners in the host cells for proper folding, stability, and activity. Protein co-expression has led to the production of a variety of biological active complexes in sufficient quantities for biochemical, biophysical, structural studies, and high throughput screens. This article summarizes examples of such cases and discusses critical considerations in selecting co-expression partners, and strategies to achieve successful production of protein complexes.


Assuntos
Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Animais , Expressão Gênica , Humanos , Complexos Multiproteicos/isolamento & purificação , Mapeamento de Interação de Proteínas , Proteínas/genética , Proteínas/isolamento & purificação , Proteínas/metabolismo
7.
Int J Spine Surg ; 15(3): 562-569, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33963036

RESUMO

BACKGROUND: Venous thromboembolism (VTE) is known to occur at high rates in patients who suffer spinal cord injury (SCI). Large population studies in the United States have shown a prevalence of approximately 4-5%, with some studies suggesting higher prevalence. While the specific mechanism behind increased VTE incidence in the SCI population is unknown, it is likely tied to immobility and functional decline. Previous studies have also linked social determinants of health to higher VTE prevalence in certain populations. The purpose of this study is to determine the prevalence of VTE after acute SCI in the inner urban city population and to identify factors that place patients at increased VTE risk. METHODS: Patients who suffered SCI between 2014 and 2019 were identified from one inner city urban hospital. A retrospective chart review was completed to record the development of VTE after SCI. Medical comorbidities, type of thromboprophylaxis used, patient demographics, injury characteristics, and postinjury sequelae were compared between individuals who did and did not develop VTE. RESULTS: A total of 148 patients were included. These patients were from a low socioeconomic demographic when compared with the larger US population. Average household income based on zip code data for included patients was $56 647, $30 315 below the national average. The prevalence of VTE in this patient population was 19.59%. Weight, deep vein thrombosis history, chemoprophylaxis use, mobility impairment postinjury, neurologic level of SCI, malignancy history, and history of smokeless tobacco were associated with VTE (all P < .05). Low molecular weight heparin (LMWH) use was associated with fewer occurrences of VTE (P < .001). CONCLUSION: Patients from our urban inner city hospital have a higher VTE prevalence after SCI than shown in previous US-based studies. These patients may be at increased risk due to increased numbers of medical comorbidities, social factors, or undiagnosed medical conditions. Thromboprophylaxis with LMWH appears to lower the risk of VTE after SCI. LEVEL OF EVIDENCE: 3. CLINICAL RELEVANCE: Patients with the previously mentioned risk factors are at increased risk for VTE development during their acute recovery process. These patients should have a much lower screening threshold for VTE evaluation and likely would benefit from more routine screening to prevent complications related to VTE development or progression. Furthermore, these patients should, when medically appropriate, be treated with low molecular weight heparin for VTE prophylaxis as previously recommended by the Consortium for Spinal Cord Medicine.

8.
Indian J Orthop ; 55(4): 879-885, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34194642

RESUMO

BACKGROUND: Fat embolism syndrome (FES) is commonly reported in the setting of long bone and pelvic fractures, but the etiology and pathogenesis are unclear. The aim of this study was to identify clinical characteristics and laboratory findings that may place orthopedic trauma patients at a higher risk of developing FES. METHODS: Electronic medical records were reviewed of all patients aged 18-89 years from 2015 to 2020 with a mention of FES in the patient chart who met Gurd and Wilson's criteria for diagnosis after experiencing orthopedic trauma. A 3:1 matched pair analysis was performed between FES patients and those with similar age, gender, and FES-associated fracture (femur, tibia, humerus, or pelvis fracture). RESULTS: 18 patients with FES who met inclusion criteria were identified. Hypomagnesemia (OR = 7.43), hyperphosphatemia (OR = 6.24), hypoalbuminemia (OR = 3.78), blunt traumatic mechanism of injury (OR = 7.16) and a greater number of bones fractured (Avg/SD = 2.89/1.53) were seen more often in FES patients (all p-values < 0.05). CONCLUSION: Findings of this study suggest that patients with hypomagnesemia, hyperphosphatemia, hypoalbuminemia, a blunt trauma mechanism of injury, and an increased number of bones fractured are at increased risk for the development of FES. This may be related to their roles in physiologic oncotic pressure and inflammatory response, and thus further investigation of these variables is necessary for the evaluation of FES prevention. LEVEL OF EVIDENCE: Level 3.

9.
J Bone Joint Surg Am ; 103(17): 1611-1619, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34185722

RESUMO

BACKGROUND: Anterior vertebral body tethering (aVBT) has emerged as a novel treatment option for patients with idiopathic scoliosis. We present the results from the first U.S. Food and Drug Administration (FDA) Investigational Device Exemption (IDE) study on aVBT. METHODS: In this prospective review of a retrospective data set, eligible patients underwent aVBT at a single center from August 2011 to July 2015. Inclusion criteria included skeletally immature patients with Lenke type-1A or 1B curves between 30° and 65°. Clinical and radiographic parameters were collected, with the latter measured by an independent reviewer. RESULTS: Fifty-seven patients (49 girls and 8 boys), with a mean age (and standard deviation) of 12.4 ± 1.3 years (range, 10.1 to 15.0 years), were enrolled in the study. The patients had a mean of 7.5 ± 0.6 levels tethered, the mean operative time was 223 ± 79 minutes, and the mean estimated blood loss was 106 ± 86 mL. The patients were followed for an average of 55.2 ± 12.5 months and had a mean Risser grade of 4.2 ± 0.9 at the time of the latest follow-up. The main thoracic Cobb angle was a mean of 40.4° ± 6.8° preoperatively and was corrected to 18.7° ± 13.4° at the most recent follow-up. In the sagittal plane, T5-T12 kyphosis measured 15.5° ± 10.0° preoperatively, 17.0° ± 10.1° postoperatively, and 19.6° ± 12.7° at the most recent follow-up. Eighty percent of patients had curves of <30° at the most recent follow-up. The most recent Scoliosis Research Society (SRS) scores averaged 4.5 ± 0.4, and scores on the self-image questionnaire averaged 4.4 ± 0.7. No major neurologic or pulmonary complications occurred. Seven (12.3%) of 57 patients had a revision: 5 were done for overcorrection and 2, for adding-on. CONCLUSIONS: Anterior VBT is a promising technique that has emerged as a treatment option for patients with immature idiopathic scoliosis. We present the results from the first FDA-approved IDE study on aVBT, which formed the basis for the eventual Humanitarian Device Exemption approval. The findings affirm the safety and efficacy of this technique and suggest opportunities for improvement, particularly with respect to reoperation rates. LEVEL OF EVIDENCE: Therapeutic Level IV. See Instructions for Authors for a complete description of levels of evidence.


Assuntos
Procedimentos Ortopédicos/instrumentação , Escoliose/cirurgia , Adolescente , Perda Sanguínea Cirúrgica , Parafusos Ósseos , Criança , Aprovação de Equipamentos , Feminino , Humanos , Cifose/cirurgia , Masculino , Duração da Cirurgia , Procedimentos Ortopédicos/métodos , Estudos Prospectivos , Reoperação/estatística & dados numéricos , Testes de Função Respiratória , Escoliose/diagnóstico por imagem , Coluna Vertebral/cirurgia , Toracoscopia/métodos , Tronco/fisiologia , Estados Unidos , United States Food and Drug Administration
10.
Protein Expr Purif ; 65(2): 251-60, 2009 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-19297698

RESUMO

We describe here two strategies to produce biologically active chemokines with authentic N-terminal amino acid residues. The first involves producing the target chemokine with an N-terminal 6xHis-SUMO tag in Escherichia coli as inclusion bodies. The fusion protein is solubilized and purified with Ni-NTA-agarose in denaturing reagents. This is further followed by tag removal and refolding in a redox refolding buffer. The second approach involves expressing the target chemokine with an N-terminal 6xHis-Trx-SUMO tag in an engineered E. coli strain that facilitates formation of disulfide bonds in the cytoplasm. Following purification of the fusion protein via Ni-NTA and tag removal, the target chemokine is refolded without redox buffer and purified by reverse phase chromatography. Using the procedures, we have produced more than 15 biologically active chemokines, with a yield of up to 15 mg/L.


Assuntos
Quimiocinas/biossíntese , Quimiocinas/metabolismo , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/metabolismo , Quimiocinas/isolamento & purificação , Clonagem Molecular , Escherichia coli/genética , Escherichia coli/metabolismo , Oxirredução , Reação em Cadeia da Polimerase , Engenharia de Proteínas , Dobramento de Proteína , Proteínas Recombinantes de Fusão/isolamento & purificação
11.
Methods Mol Biol ; 552: 199-211, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19513651

RESUMO

BacMam viruses are modified baculoviruses that contain mammalian expression cassettes for viral gene delivery and transient expression in mammalian cells. They are easily, inexpensively, and rapidly generated and provide a versatile solution for G protein-coupled receptor (GPCR) cell-based assay development. Using BacMam technology, target gene expression levels are easily controlled and simultaneous delivery of multiple genes is possible, for example, coexpression of a receptor and a G protein or a reporter gene. BacMam viruses are compatible with the GPCR cell-based assay formats typically used in high-throughput screening and provide an unparalleled level of experimental flexibility that is simply not possible when using stable recombinant cell lines.


Assuntos
Baculoviridae/genética , Neoplasias Ósseas/metabolismo , Técnicas de Transferência de Genes , Osteossarcoma/metabolismo , Receptores Acoplados a Proteínas G/genética , Neoplasias Ósseas/genética , Vetores Genéticos , Humanos , Osteossarcoma/genética , Receptores Acoplados a Proteínas G/metabolismo , Transdução Genética , Células Tumorais Cultivadas
12.
J Am Acad Orthop Surg ; 27(10): e462-e472, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30407981

RESUMO

Scheuermann's kyphosis (SK) is a rigid structural deformity of the thoracic spine defined radiographically as three or more contiguous vertebrae with at least 5° of wedging anteriorly. Prevalence of the disease is thought to be between 0.4% and 10%. The true cause of SK remains unclear; however, various theories include growth irregularities, mechanical factors, genetic factors, and/or poor bone quality as the causes. Patients with mild disease (less than 70°) generally have a favorable prognosis with good clinical outcomes. Most patients with SK are successfully treated nonsurgically with observation, anti-inflammatory medications, and physical therapy. Surgical intervention is indicated in patients with greater than 70° to 75° thoracic curves, greater than 25° to 30° thoracolumbar curves, intractable pain, neurologic deficit, cardiopulmonary compromise, or poor cosmesis. Because of advances in posterior spinal instrumentation, surgery can typically be performed through a posterior-only approach. When surgical treatment is planned, appropriate selection of the upper- and lower-instrumented vertebrae is important to achieve a well-balanced spine, preserve motion segments, and reduce the risk of junctional kyphosis.


Assuntos
Doença de Scheuermann/diagnóstico , Doença de Scheuermann/terapia , Fusão Vertebral/métodos , Vértebras Torácicas/cirurgia , Anti-Inflamatórios/uso terapêutico , Tratamento Conservador , Humanos , Modalidades de Fisioterapia , Radiografia , Doença de Scheuermann/etiologia , Doença de Scheuermann/patologia , Fusão Vertebral/efeitos adversos , Vértebras Torácicas/diagnóstico por imagem , Vértebras Torácicas/patologia
13.
Mol Biotechnol ; 39(2): 127-34, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18327553

RESUMO

Cortisol is an important glucocorticoid in humans that regulates many physiological processes. Human 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) converts cortisone to cortisol in vivo and has emerged as an appealing therapeutic target for treating metabolic diseases. Here, we report a sensitive and robust high-throughput (HT) cell-based assay for screening 11beta-HSD1 inhibitors. This assay utilizes a HEK293 cell line transduced by a BacMam virus expressing human 11beta-HSD1. The enzyme activity in the cells was measured by quantifying cortisol levels released into the cell culture supernatant via a competitive homogenous time-resolved fluorescence (HTRF) method. We show that 11beta-HSD1 activity in supernatant of BacMam-transduced HEK293 cells increases with 11beta-HSD1 BacMam virus load in a dose-dependent manner, and is comparable to the enzyme activity detected in differentiated mouse adipocytes. In addition, we show that co-expression of hexose-6-phosphate dehydrogenase (H6PDH) is not required for the enzyme to function effectively as an oxo-reductase. This assay has been developed in low-volume 384-well format and it is sensitive, robust, and amenable to HT screening.


Assuntos
Fluorimunoensaio/métodos , Rim/enzimologia , Transdução Genética/métodos , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/antagonistas & inibidores , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/metabolismo , Células 3T3-L1 , Adipócitos/enzimologia , Animais , Desidrogenases de Carboidrato/metabolismo , Sobrevivência Celular , Cortisona/metabolismo , Meios de Cultura/análise , Humanos , Hidrocortisona/metabolismo , Camundongos
14.
Drug Discov Today ; 12(9-10): 396-403, 2007 May.
Artigo em Inglês | MEDLINE | ID: mdl-17467576

RESUMO

Membrane protein targets constitute a key segment of drug discovery portfolios and significant effort has gone into increasing the speed and efficiency of pursuing these targets. However, issues still exist in routine gene expression and stable cell-based assay development for membrane proteins, which are often multimeric or toxic to host cells. To enhance cell-based assay capabilities, modified baculovirus (BacMam virus) gene delivery technology has been successfully applied to the transient expression of target proteins in mammalian cells. Here, we review the development, full implementation and benefits of this platform-based gene expression technology in support of SAR and HTS assays across GlaxoSmithKline.


Assuntos
Baculoviridae/genética , Desenho de Fármacos , Transfecção/métodos , Animais , Indústria Farmacêutica/métodos , Expressão Gênica , Vetores Genéticos/genética , Humanos , Modelos Biológicos , Tecnologia Farmacêutica/métodos
15.
Methods Mol Biol ; 388: 95-114, 2007.
Artigo em Inglês | MEDLINE | ID: mdl-17951767

RESUMO

BacMams are modified baculoviruses that contain mammalian expression cassettes for gene delivery and expression in mammalian cells. The BacMam system combines the advantages of viral transient expression, ease in generation, and a wide cell tropism. It enables rapid, facile, and flexible gene over-expression experiments to be performed in a variety of mammalian cell lines. Conversion of baculovirus vectors to BacMam vectors involves replacement of the viral specific expression cassette with a mammalian expression cassette or the addition of a mammalian expression cassette. Viruses are produced using standard methods in a few weeks. Mammalian cells transduced with the BacMam viruses have been routinely used as substitutes for stable cell lines.


Assuntos
Baculoviridae/genética , Expressão Gênica , Vetores Genéticos/genética , Nucleopoliedrovírus/genética , Animais , Linhagem Celular , DNA Viral/genética , Citometria de Fluxo , Humanos , Luciferases/genética , Luciferases/metabolismo , Mariposas/virologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Transfecção/métodos
16.
Spine J ; 17(10): 1406-1411, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28412564

RESUMO

BACKGROUND CONTEXT: A paucity of data exists studying outcomes of patients with syringomyelia undergoing spinal deformity correction. The literature does not stratify patients by syrinx size, which is likely a major contributor to outcomes. PURPOSE: The study aimed to compare differences in outcomes between patients with large (≥4 mm) and small syrinxes (<4 mm) undergoing spinal deformity correction. DESIGN: This is a retrospective review. PATIENT SAMPLE: The sample included 28 patients (11 with large syrinx [LS, >4 mm] and 17 with small syrinx [SS, <4 mm]). OUTCOME MEASURES: The outcome measures were radiographic, operative, and neurophysiological measures. METHODS: We retrospectively reviewed 28 patients with syringomyelia who underwent spine deformity surgery with 2-year follow-up. Demographic, surgical, and radiographic data were collected and compared preoperatively and at 2 years. RESULTS: The LS group (11 patients) trended toward more left-sided thoracic curves (36% vs. 18%, p=.38) and was more likely to have had a Chiari decompression (45% vs. 12%, p=.08). The LS patients had larger preoperative major curves (LS=66° vs. SS=57°, p=.05), more thoracic kyphosis (LS=42°, SS=24°, p<.01), and greater rib prominences (LS=16°, SS=13°, p=.04). The LS patients had more levels fused (LS=12.2, SS=11.2, p=.05), higher estimated blood loss (EBL) (LS=1068 cc, SS=832 cc, p=.04), and a trend toward less percent correction of the major curve (LS=57%, SS=65%, p=.18). Four of 11 LS patients (36%) did not have somatosensory evoked potentials, and one of these also did not have motor evoked potentials. Neuromonitoring changes occurred in 3 of 11 (27%) LS patients and in none of the SS patients, with no postoperative deficits. CONCLUSIONS: Outcomes of patients with syringomyelia undergoing spine deformity surgery are dependent on the size of the syrinx. Those with large syringomyelia are fused longer with more EBL and less correction. Spine surgeons should be aware that these patients are more likely to have less reliable neuromonitoring, with a higher chance of experiencing a change.


Assuntos
Descompressão Cirúrgica/efeitos adversos , Complicações Pós-Operatórias/etiologia , Medula Espinal/patologia , Siringomielia/cirurgia , Adolescente , Adulto , Potencial Evocado Motor , Potenciais Somatossensoriais Evocados , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Complicações Pós-Operatórias/fisiopatologia , Siringomielia/patologia
17.
Br J Pharmacol ; 148(2): 173-90, 2006 May.
Artigo em Inglês | MEDLINE | ID: mdl-16547525

RESUMO

Several peptidic urotensin-II (UT) receptor antagonists exert 'paradoxical' agonist activity in recombinant cell- and tissue-based bioassay systems, likely the result of differential urotensin-II receptor (UT receptor) signal transduction/coupling efficiency between assays. The present study has examined this phenomenon in mammalian arteries and recombinant UT-HEK (human embryonic kidney) cells.BacMam-mediated recombinant UT receptor upregulation in HEK cells augmented agonist activity for all four peptidic UT ligands studied. The nominal rank order of relative intrinsic efficacy was U-II>urantide ([Pen(5)-DTrp(7)-Orn(8)]hU-II(4-11))>SB-710411 (Cpa-c[DCys-Pal-DTrp-Lys-Val-Cys]-Cpa-amide)>>GSK248451 (Cin-c[DCys-Pal-DTrp-Orn-Val-Cys]-His-amide) (the relative coupling efficiency of recombinant HEK cells was cat>human>>rat UT receptor). The present study further demonstrated that the use of high signal transduction/coupling efficiency isolated blood vessel assays (primate>cat arteries) is required in order to characterize UT receptor antagonism thoroughly. This cannot be attained simply by using the rat isolated aorta, an artery with low signal transduction/coupling efficiency in which low-efficacy agonists appear to function as antagonists. In contrast to the 'low-efficacy agonists' urantide and SB-710411, GSK248451 functioned as a potent UT receptor antagonist in all native isolated tissues studied (UT receptor selectivity was confirmed in the rat aorta). Further, GSK248451 exhibited an extremely low level of relative intrinsic activity in recombinant HEK cells (4-5-fold less than seen with urantide). Since GSK248451 (1 mg kg(-1), i.v.) blocked the systemic pressor actions of exogenous U-II in the anaesthetized cat, it represents a suitable peptidic tool antagonist for delineating the role of U-II in the aetiology of mammalian cardiometabolic diseases.


Assuntos
Fragmentos de Peptídeos/farmacologia , Peptídeos Cíclicos/farmacologia , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Urotensinas/farmacologia , Animais , Artérias/efeitos dos fármacos , Artérias/fisiologia , Ligação Competitiva/efeitos dos fármacos , Pressão Sanguínea/efeitos dos fármacos , Cálcio/metabolismo , Gatos , Linhagem Celular , Relação Dose-Resposta a Droga , Haplorrinos , Humanos , Técnicas In Vitro , Masculino , Fragmentos de Peptídeos/metabolismo , Peptídeos Cíclicos/química , Ensaio Radioligante , Ratos , Ratos Sprague-Dawley , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Proteínas Recombinantes/antagonistas & inibidores , Proteínas Recombinantes/metabolismo , Urotensinas/metabolismo , Vasoconstrição/efeitos dos fármacos , Vasoconstritores/farmacologia
18.
Methods Mol Biol ; 1350: 95-116, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26820855

RESUMO

BacMams are modified baculoviruses that contain mammalian expression cassettes for gene delivery and expression in mammalian cells. BacMams have become an integral part of the recombinant mammalian gene expression toolbox in research labs worldwide. Construction of transfer vectors is straightforward using basic molecular biology protocols. Virus generation is based on common methods used with the baculovirus insect cell expression system. BacMam transduction of mammalian cells requires minimal modifications to familiar cell culture methods. This chapter highlights the BacMam transfer vector pHTBV.


Assuntos
Baculoviridae/genética , Engenharia Genética/métodos , Animais , Baculoviridae/isolamento & purificação , Células CHO , Clonagem Molecular , Cricetinae , Cricetulus , Enzimas de Restrição do DNA/genética , DNA Complementar/genética , Expressão Gênica , Vetores Genéticos/genética , Células HEK293 , Humanos , Técnicas de Amplificação de Ácido Nucleico , Células Sf9 , Spodoptera , Transformação Genética
19.
J Bone Joint Surg Am ; 98(17): 1478-83, 2016 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-27605692

RESUMO

BACKGROUND: Confidence in intraoperative neurophysiologic monitoring (IONM) data can allow scoliosis surgeons to proceed with surgery even after a monitoring alert, assuming the recovery of signals. We sought to determine the outcomes of surgical treatment of adolescent idiopathic scoliosis (AIS) after a notable IONM alert. METHODS: We identified 676 patients who underwent arthrodesis with use of IONM for the treatment of AIS. The patients were divided into 2 cohorts: those who experienced a lower-extremity IONM alert and those who did not. An alert was defined as a notable change in IONM data, specifically, a ≥50% drop in somatosensory evoked potentials (SSEPs) and/or in transcranial motor evoked potentials (tcMEPs). RESULTS: Of the 676 patients, 36 (5.3%) experienced IONM alerts. Those patients had a larger preoperative major Cobb angle (mean of 61° ± 13° compared with 55° ± 12° for the no-alert group; p < 0.01), a greater number of levels fused (mean of 12 ± 2 compared with 11 ± 2; p < 0.01), a longer operative duration (mean of 357 ± 157 minutes compared with 298 ± 117 minutes; p < 0.01), a higher estimated blood loss (1,857 ± 1,323 mL compared with 999 ± 796 mL; p < 0.01), and a greater volume of autologous blood transfused (mean of 527 ± 525 mL compared with 268 ± 327 mL; p < 0.01). Among patients who experienced an alert and had a completed operation (34 of 36 patients), mean postoperative radiographic measurements were similar to those of the no-alert group in terms of the percentage of correction of the major Cobb angle (alert, 66% ± 13%; no alert, 64% ± 19%; p = 0.53) and of rib prominence (alert, 49% ± 36%; no alert, 47% ± 46%; p = 0.83) and measurement of thoracic kyphosis (alert, 23° ± 10°; no alert, 22° ± 2°; p = 0.58). The Scoliosis Research Society (SRS)-22 outcome scores were also similar between the 2 cohorts. CONCLUSIONS: Notable IONM changes occurred in 5.3% of the patients who underwent arthrodesis for AIS. Those patients had larger preoperative deformity, a longer operative duration, a greater number of levels fused, a higher estimated blood loss, and a greater volume of autologous blood transfused. Return of IONM data guided the surgeon to safely complete the procedure in 34 of 36 patients, with correction similar to that of patients who did not experience an alert. LEVEL OF EVIDENCE: Therapeutic Level III. See Instructions for Authors for a complete description of levels of evidence.


Assuntos
Potenciais Somatossensoriais Evocados/fisiologia , Monitorização Neurofisiológica Intraoperatória/métodos , Escoliose/cirurgia , Fusão Vertebral/métodos , Adolescente , Feminino , Humanos , Masculino , Escoliose/fisiopatologia , Resultado do Tratamento
20.
J Neurosurg Spine ; 22(2): 185-91, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25415486

RESUMO

OBJECT: Almost all pediatric patients who incur a spinal cord injury (SCI) will develop scoliosis, and younger patients are at highest risk for curve progression requiring surgical intervention. Although the use of pedicle screws is increasing in popularity, their impact on SCI-related scoliosis has not been described. The authors retrospectively reviewed the radiographic outcomes of pedicle screw-only constructs in all patients who had undergone SCI-related scoliosis correction at a single institution. METHODS: Medical records and radiographs from Shriner's Hospital for Children-Philadelphia for the period between November 2004 and February 2011 were retrospectively reviewed. RESULTS: Thirty-seven patients, whose mean age at the index surgery was 14.91±3.29 years, were identified. The cohort had a mean follow-up of 33.2±22.8 months. The mean preoperative coronal Cobb angle was 65.5°±25.7°, which corrected to 20.3°±14.4°, translating into a 69% correction (p<0.05). The preoperative coronal balance was 24.4±22.6 mm, with a postoperative measurement of 21.6±20.7 mm (p=1.00). Preoperative pelvic obliquity was 12.7°±8.7°, which corrected to 4.1°±3.8°, translating into a 68% correction (p<0.05). Preoperative shoulder balance, as measured by the clavicle angle, was 8.2°±8.4°, which corrected to 2.7°±3.1° (67% correction, p<0.05). Preoperatively, thoracic kyphosis measured 44.2°±23.7° and was 33.8°±11.5° postoperatively. Thoracolumbar kyphosis was 18.7°±12.1° preoperatively, reduced to 8.1°±7.7° postoperatively, and measured 26.8°±20.2° at the last follow-up (p<0.05). Preoperatively, lumbar lordosis was 35.3°±22.0°, which remained stable at 35.6°±15.0° postoperatively. CONCLUSIONS: Pedicle screw constructs appear to provide better correction of coronal parameters than historically reported and provide significant improvement of sagittal kyphosis as well. Although pedicle screws appear to provide good radiographic results, correlation with clinical outcomes is necessary to determine the true impact of pedicle screw constructs on SCI-related scoliosis correction.


Assuntos
Vértebras Lombares/cirurgia , Parafusos Pediculares , Escoliose/cirurgia , Traumatismos da Medula Espinal/cirurgia , Fusão Vertebral , Adolescente , Criança , Feminino , Humanos , Cifose/cirurgia , Masculino , Estudos Retrospectivos , Escoliose/complicações , Traumatismos da Medula Espinal/etiologia , Fusão Vertebral/métodos , Resultado do Tratamento , Adulto Jovem
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