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1.
Acta Neurochir (Wien) ; 165(11): 3521-3527, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37715821

RESUMO

PURPOSE: Pedicle subtraction osteotomy (PSO) as an invasive procedure with high reoperation and complication rates in an often elderly population has often been questioned. The purpose of our study was to evaluate the impact of PSO for sagittal imbalance (SI) on patient-reported outcomes including self-reported satisfaction and health-related quality of life 2 years postoperatively. METHODS: Consecutive patients who underwent correction of their spinal deformity by thoracolumbar PSO were assessed using self-reporting questionnaires 2 years postoperatively. Outcome was measured by visual analogue scale (VAS) for back and leg pain, Oswestry Disability Index (ODI), and EQ-5D scores. Additionally, a Patient Satisfaction Index (PSI) rated in four grades (A: very satisfied to D: not satisfied), walking range, and the Timed Up and Go (TUG) Test were evaluated. RESULTS: Sixty-five patients were included, and each parameter was assessed preoperatively and 24 months postoperatively. The intervention led to significant improvements in back pain (8.1 ± 1.2 vs. 2.9 ± 1.9; p < 0.001), as well as ODI scores (57.7 ± 13.9 vs. 32.6 ± 18.9; p < 0.001), walking range (589 ± 1676 m vs. 3265 ± 3405 m; p < 0.001), and TUG (19.2 s vs. 9.7 s; p < 0.05). 90.7% of patients (n = 59/65) reported a PSI grade "A" or "B" 24 months postoperatively. CONCLUSION: Patient satisfaction 24 months after PSO for SI is high. Quality of life improved significantly by restoring sagittal balance.


Assuntos
Cifose , Fusão Vertebral , Humanos , Idoso , Qualidade de Vida , Osteotomia/efeitos adversos , Osteotomia/métodos , Satisfação do Paciente , Dor nas Costas , Caminhada , Estudos Retrospectivos , Fusão Vertebral/métodos , Resultado do Tratamento , Vértebras Lombares/cirurgia , Cifose/cirurgia , Vértebras Torácicas/cirurgia
2.
Pest Manag Sci ; 79(1): 134-139, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36114592

RESUMO

BACKGROUND: The spotted wing drosophila, Drosophila suzukii Matsumura, is a South-East Asian vinegar fly that is a serious worldwide economic threat to the small fruit industry. Typical control consists of weekly pesticide applications, which can have nontarget effects, increase residual pesticides and lead to the development of resistance within pest populations. One potential alternate method of control is the planting of aromatic intercrops to attract the natural enemies of D. suzukii and/or repel the flies directly. We intercropped strawberry rows with flowering sweet alyssum or ryegrass-clover (control) to evaluate their efficacy at mitigating D. suzukii infestation through the attraction of two specialized larval parasitoids, Leptopilina japonica (Novkovic and Kimura) and Ganaspis brasiliensis (Ihering). RESULTS: Our study did not demonstrate any significant effect of sweet alyssum intercropping on the infestation rate of D. suzukii in strawberries or parasitism level. However, we found that advanced sampling date and recorded numbers of D. suzukii larvae and parasitoids were positively correlated, indicating higher populations at the end of the strawberry-growing season. CONCLUSIONS: Sweet alyssum intercrops did not reduce D. suzukii infestation rates or increase parasitism levels, likely due to low population numbers in early season berry varieties. Aromatic intercrops may be more effective for increasing pest control in later season crops. © 2022 Society of Chemical Industry.


Assuntos
Drosophila , Lolium , Controle Biológico de Vetores , Vespas , Animais , Drosophila/parasitologia , Vespas/fisiologia , Controle Biológico de Vetores/normas , Fragaria/parasitologia , Lolium/fisiologia
3.
Pest Manag Sci ; 78(11): 4929-4938, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36054536

RESUMO

BACKGROUND: Invasive species threaten the productivity and stability of natural and managed ecosystems. Predicting the spread of invaders, which can aid in early mitigation efforts, is a major challenge, especially in the face of climate change. While ecological niche models are effective tools to assess habitat suitability for invaders, such models have rarely been created for invasive pest species with rapidly expanding ranges. Here, we leveraged a national monitoring effort from 543 sites over 3 years to assess factors mediating the occurrence and abundance of brown marmorated stink bug (BMSB, Halyomorpha halys), an invasive insect pest that has readily established throughout much of the United States. RESULTS: We used maximum entropy models to estimate the suitable habitat of BMSB under several climate scenarios, and generalized boosted models to assess environmental factors that regulated BMSB abundance. Our models captured BMSB distribution and abundance with high accuracy, and predicted a 70% increase in suitable habitat under future climate scenarios. However, environmental factors that mediated the geographical distribution of BMSB were different from those driving abundance. While BMSB occurrence was most affected by winter precipitation and proximity to populated areas, BMSB abundance was influenced most strongly by evapotranspiration and solar photoperiod. CONCLUSION: Our results suggest that linking models of establishment (occurrence) and population dynamics (abundance) offers a more effective way to forecast the spread and impact of BMSB and other invasive species than simply occurrence-based models, allowing for targeted mitigation efforts. Implications of distribution shifts under climate change are discussed. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.


Assuntos
Ecossistema , Heterópteros , Animais , Mudança Climática , Espécies Introduzidas , Dinâmica Populacional , Estados Unidos
4.
Environ Entomol ; 51(4): 670-678, 2022 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-35594566

RESUMO

Two species of larval parasitoids of the globally invasive fruit pest, Drosophila suzukii (Diptera: Drosophilidae), Leptopilina japonica, and Ganaspis brasiliensis (both Hymenoptera: Figitidae), were detected in British Columbia, Canada in 2016 and 2019, respectively. Both are presumed to have been unintentionally introduced from Asia; however, the extent of their establishment across different habitats with diverse host plants used by D. suzukii was unclear. In addition, there was no knowledge of the temporal dynamics of parasitism of D. suzukii by these two parasitoids. To address these gaps, we repeatedly sampled the fruits of known host plants of D. suzukii over the entire 2020 growing season in British Columbia. We documented the presence of L. japonica and G. brasiliensis and estimated the apparent percentage of D. suzukii parasitized among host plant species. Across a large region of southwestern British Columbia, both L. japonica and G. brasiliensis were found to be very common across a variety of mostly unmanaged habitats over the entire course of the season (May-October) in the fruits of most host plants known to host D. suzukii larvae. Parasitism of D. suzukii was variable (0-66% percent parasitism) and appeared to be time-structured. Our study demonstrates that the close association between the two larval parasitoids and D. suzukii that exists in Asia has evidently been reconstructed in North America, resulting in the highest parasitism levels of D. suzukii yet recorded outside of its area of origin.


Assuntos
Drosophila , Himenópteros , Animais , Colúmbia Britânica , Frutas , Controle de Insetos , Larva
5.
Eur Spine J ; 31(10): 2629-2638, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-35188587

RESUMO

BACKGROUND: Indications and outcomes in lumbar spinal fusion for degenerative disease are notoriously heterogenous. Selected subsets of patients show remarkable benefit. However, their objective identification is often difficult. Decision-making may be improved with reliable prediction of long-term outcomes for each individual patient, improving patient selection and avoiding ineffective procedures. METHODS: Clinical prediction models for long-term functional impairment [Oswestry Disability Index (ODI) or Core Outcome Measures Index (COMI)], back pain, and leg pain after lumbar fusion for degenerative disease were developed. Achievement of the minimum clinically important difference at 12 months postoperatively was defined as a reduction from baseline of at least 15 points for ODI, 2.2 points for COMI, or 2 points for pain severity. RESULTS: Models were developed and integrated into a web-app ( https://neurosurgery.shinyapps.io/fuseml/ ) based on a multinational cohort [N = 817; 42.7% male; mean (SD) age: 61.19 (12.36) years]. At external validation [N = 298; 35.6% male; mean (SD) age: 59.73 (12.64) years], areas under the curves for functional impairment [0.67, 95% confidence interval (CI): 0.59-0.74], back pain (0.72, 95%CI: 0.64-0.79), and leg pain (0.64, 95%CI: 0.54-0.73) demonstrated moderate ability to identify patients who are likely to benefit from surgery. Models demonstrated fair calibration of the predicted probabilities. CONCLUSIONS: Outcomes after lumbar spinal fusion for degenerative disease remain difficult to predict. Although assistive clinical prediction models can help in quantifying potential benefits of surgery and the externally validated FUSE-ML tool may aid in individualized risk-benefit estimation, truly impacting clinical practice in the era of "personalized medicine" necessitates more robust tools in this patient population.


Assuntos
Fusão Vertebral , Dor nas Costas/diagnóstico , Dor nas Costas/etiologia , Dor nas Costas/cirurgia , Feminino , Humanos , Vértebras Lombares/cirurgia , Masculino , Pessoa de Meia-Idade , Modelos Estatísticos , Prognóstico , Fusão Vertebral/métodos , Resultado do Tratamento
6.
J Econ Entomol ; 115(4): 922-942, 2022 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34984457

RESUMO

We provide recommendations for sampling and identification of introduced larval parasitoids of spotted-wing drosophila, Drosophila suzukii (Matsumura) (Diptera: Drosophilidae). These parasitoids are either under consideration for importation (aka classical) biological control introductions, or their adventive (presumed to have been accidentally introduced) populations have recently been discovered in North America and Europe. Within the context of the ecology of D. suzukii and its parasitoids, we discuss advantages and disadvantages of estimating larval parasitism levels using different methods, including naturally collected fruit samples and sentinel baits. For most situations, we recommend repeated sampling of naturally occurring fruit rather than using sentinel baits to monitor seasonal dynamics of host plant-Drosophila-parasitoid associations. We describe how to separate Drosophilidae puparia from host fruit material in order to accurately estimate parasitism levels and establish host-parasitoid associations. We provide instructions for identification of emerging parasitoids and include a key to the common families of parasitoids of D. suzukii. We anticipate that the guidelines for methodology and interpretation of results that we provide here will form the basis for a large, multi-research team sampling effort in the coming years to characterize the biological control and nontarget impacts of accidentally and intentionally introduced larval parasitoids of D. suzukii in several regions of the world.


Assuntos
Drosophila , Frutas , Animais , Europa (Continente) , Controle de Insetos/métodos , Larva , América do Norte
7.
Neurosurg Rev ; 45(1): 517-524, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33963469

RESUMO

Three-column osteotomy (3-CO) is a powerful technique in adult deformity surgery, and pedicle subtraction osteotomy (PSO) is the workhorse to correct severe kyphotic spinal deformities. Aging of the population, increasing cases of iatrogenic flat back deformities and understanding the importance of sagittal balance have led to a dramatic increase of this surgical technique. Surgery, however, is demanding and associated with high complication rates so that every step of the procedure requires meticulous technique. Particularly, osteotomy closure is associated with risks like secondary fracture, translation, or iatrogenic stenosis. This step is traditionally performed by compression or a cantilever maneuver with sometimes excessive forces on the screws or instrumentation. Implant loosening or abrupt subluxation resulting in construct failure and/or neurological deficits can result. The aim of this prospective registry study was to assess the efficacy and safety of our surgical PSO technique as well as the osteotomy closure by flexing a hinge-powered OR table. In a series of 84 consecutive lumbosacral 3-CO, a standardized surgical technique with special focus on closure of the osteotomy was prospectively evaluated. The surgical steps with the patients positioned prone on a soft frame are detailed. Osteotomy closure was achieved by remote controlled bending of a standard OR table without compressive or cantilever forces in all 84 cases. This technique carries a number of advantages, particularly the reversibility and the slow speed of closure with minimum force. There was not a single mechanical intraoperative complication such as vertebral body fracture, subluxation, or adjacent implant loosening during osteotomy closure, compared to external cohorts using the cantilever technique (p = 0.130). The feasibility of controlled 3-CO closure by flexing a standard OR table is demonstrated. This technique enables a safe, gentle closure of the osteotomy site with minimal risk of implant failure or accidental neurological injury.


Assuntos
Cifose , Mesas Cirúrgicas , Fusão Vertebral , Adulto , Humanos , Osteotomia , Estudos Prospectivos , Estudos Retrospectivos , Resultado do Tratamento
8.
Bioprocess Biosyst Eng ; 45(2): 297-307, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-34750672

RESUMO

In the field of therapeutic antibody production, diversification of fed-batch strategies is flourishing in response to the market demand. All manufacturing approaches tend to follow the generally accepted dogma of increasing titer since it directly increases manufacturing output. While titer is influenced by the biomass (expressed as IVCD), the culture time and the cell-specific productivity (qP), we changed independently each of these parameters to tune our process strategy towards adapted solutions to individual manufacturing needs. To do so, we worked separately on the increase of the IVCD as high seeding fed-batch capacity. Yet, as intensified fed-batch may not always be possible due to limited facility operational mode, we also separately increased the qP with the addition of specific media additives. Both strategies improved titer by 100% in 14 days relative to the standard fed-batch process with moderate and acceptable changes in product quality attributes. Since intensified fed-batch could rival the cell-specific productivity of a conventional fed-batch, we developed novel hybrid strategies to either allow for acceptable seeding densities without compromising productivity, or alternatively, to push the productivity the furthest in order to reduce timelines.


Assuntos
Técnicas de Cultura Celular por Lotes , Reatores Biológicos , Animais , Formação de Anticorpos , Células CHO , Cricetinae , Cricetulus
9.
J Neurotrauma ; 39(9-10): 613-626, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-34937399

RESUMO

Sensorimotor function of patients with spinal cord injury (SCI) is commonly assessed according to the International Standards for Neurological Classification of Spinal Cord Injury (ISNCSCI). From the ISNCSCI segmental motor and sensory assessments, upper and lower extremity motor scores (UEMS and LEMS), sum scores of pinprick (PP) and light touch (LT) sensation, the neurological level of injury (NLI) and the classification of lesion severity according to the American Spinal Injury Association Impairment Scale (AIS) grade are derived. Changes of these parameters over time are used widely to evaluate neurological recovery. Evaluating recovery based on a single ISNCSCI scoring or classification variable, however, may misestimate overall recovery. Here, we propose an Integrated Neurological Change Score (INCS) based on the combination of normalized changes between two time points of UEMS, LEMS, and total PP and LT scores. To assess the agreement of INCS with clinical judgment of meaningfulness of neurological changes, changes of ISNCSCI variables between two time points of 88 patients from an independent cohort were rated by 20 clinical experts according to a five-categories Likert Scale. As for individual ISNCSCI variables, neurological change measured by INCS is associated with severity (AIS grade), age, and time since injury, but INCS better reflects clinical judgment about meaningfulness of neurological changes than individual ISNCSCI variables. In addition, INCS is related to changes in functional independence measured by the Spinal Cord Independence Measure (SCIM) in patients with tetraplegia. The INCS may be a useful measure of overall neurological change in clinical studies.


Assuntos
Traumatismos da Medula Espinal , Humanos , Quadriplegia/complicações , Recuperação de Função Fisiológica , Sensação , Extremidade Superior
10.
Acta Neurochir (Wien) ; 163(1): 275-280, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33145630

RESUMO

BACKGROUND: Patients with intervertebral disc herniation undergo surgical removal of herniated disc material in cases of persisting symptoms and/or neurologic deficits. While motor deficits often prompt surgery, little is known about the optimal timing of surgery in these cases. The aim of this study was to prospectively evaluate the impact of timing of disc surgery on motor recovery. Does postponing surgical treatment worsen outcome? METHOD: In total, 120 patients with sciatica and/or sensorimotor deficits due to a lumbar disc herniation were surgically treated at the authors' center within a 3-month period. In 60 patients, motor deficits were present at the time of admission. Motor function was assessed using manual muscle testing and subdivided according to the Medical Research Council (MRC) scale. Patient demographics, neurologic deficits, duration of motor deficits, treatment characteristics, and outcome were assessed. At a minimum follow-up of 1 year, functional recovery and complications were collated. Patients were subdivided into groups according to the severity of the paresis (MRC ≤ 3/5 vs. MRC 4/5). Intra-group differences were compared based on the duration of the neurologic deficits. RESULTS: Patients with moderate and severe paresis (MRC ≤ 3/5) benefit from treatment within 72 h as they were shown to have a significantly higher complete recovery rate at 1-year follow-up (75% vs. 0%; p < 0.001). CONCLUSION: Immediate surgery should be offered to patients with moderate and severe motor deficits to increase the likelihood of neurologic recovery. This prospective data may have an impact on emergency triage in these patients.


Assuntos
Discotomia/métodos , Deslocamento do Disco Intervertebral/cirurgia , Vértebras Lombares/cirurgia , Complicações Pós-Operatórias/epidemiologia , Adulto , Discotomia/efeitos adversos , Feminino , Humanos , Deslocamento do Disco Intervertebral/patologia , Masculino , Pessoa de Meia-Idade , Movimento , Recuperação de Função Fisiológica
11.
Sci Rep ; 10(1): 19096, 2020 11 05.
Artigo em Inglês | MEDLINE | ID: mdl-33154398

RESUMO

Uncertainty about the taxonomic status and the specificity of a species commonly prevent its consideration as a candidate for biological control of pest organisms. Here we use a combination of molecular analysis and crossing experiments to gather evidence that the parasitoid wasp Ganaspis brasiliensis, a candidate for biological control of the invasive spotted wing drosophila Drosophila suzukii, is a complex of at least two cryptic species. Complementary experiments demonstrate that individuals from one genetic group readily parasitize several drosophila species regardless of their food source while individuals from the other one are almost exclusively specific to larvae feeding in ripening fruits. Because only D. suzukii attacks ripening fruits in its area of invasion, parasitoids from this second group appear to be well suited as a biological control agent. Our study demonstrates the need for a combination of biosystematics with biological and ecological investigations for the development of safe and efficient biological control programs.


Assuntos
Agentes de Controle Biológico , Drosophila/parasitologia , Drosophila/patogenicidade , Vespas/fisiologia , Vespas/patogenicidade , Animais , China , Ecossistema , Feminino , Frutas/parasitologia , Genética Populacional , Genoma de Inseto , Haplótipos , Interações Hospedeiro-Parasita/genética , Espécies Introduzidas , Japão , Masculino , Especificidade da Espécie , Vespas/genética
12.
Acta Neurochir (Wien) ; 162(11): 2927-2931, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32808086

RESUMO

Monostotic fibrous dysplasia (MFD) of the lumbar spine represents an exceedingly rare lesion. A 26-year-old patient presented with a progressive osteolytic lesion of the vertebral body L2 and the diagnosis of MFD. A minimally invasive left-sided eXtreme Lateral Interbody Fusion (XLIF) approach with resection of the vertebral body L2 with placement of a mesh cage was performed. No complications were observed perioperatively and the symptoms rapidly improved. Minimally invasive piecemeal resection with a combined dorsolateral approach showed a favorable clinical and radiological outcome and seems to be a safe and reliable technique for MFD.


Assuntos
Displasia Fibrosa Monostótica/cirurgia , Cifose/cirurgia , Fusão Vertebral/métodos , Adulto , Displasia Fibrosa Monostótica/complicações , Humanos , Cifose/etiologia , Vértebras Lombares/cirurgia , Masculino
13.
World Neurosurg ; 140: 664-673, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32445895

RESUMO

BACKGROUND: The desire to improve accuracy and safety and to favor minimally invasive techniques has given rise to spinal robotic surgery, which has seen a steady increase in utilization in the past 2 decades. However, spinal surgery encompasses a large spectrum of operative techniques, and robotic surgery currently remains confined to assistance with the trajectory of pedicle screw insertion, which has been shown to be accurate and safe based on class II and III evidence. The role of robotics in improving surgical outcomes in spinal pathologies is less clear, however. METHODS: This comprehensive review of the literature addresses the role of robotics in surgical outcomes in spinal pathologies with a focus on the various meta-analysis and prospective randomized trials published within the past 10 years in the field. RESULTS: It appears that robotic spinal surgery might be useful for increasing accuracy and safety in spinal instrumentation and allows for a reduction in surgical time and radiation exposure for the patient, medical staff, and operator. CONCLUSION: Robotic assisted surgery may thus open the door to minimally invasive surgery with greater security and confidence. In addition, the use of robotics facilitates tireless repeated movements with higher precision compared with humans. Nevertheless, it is clear that further studies are now necessary to demonstrate the role of this modern tool in cost-effectiveness and in improving clinical outcomes, such as reoperation rates for screw malpositioning.


Assuntos
Procedimentos Cirúrgicos Robóticos/instrumentação , Procedimentos Cirúrgicos Robóticos/tendências , Doenças da Coluna Vertebral/cirurgia , Cirurgia Assistida por Computador/instrumentação , Cirurgia Assistida por Computador/tendências , Fluoroscopia/instrumentação , Fluoroscopia/tendências , Humanos , Monitorização Neurofisiológica Intraoperatória/instrumentação , Monitorização Neurofisiológica Intraoperatória/tendências , Procedimentos Cirúrgicos Minimamente Invasivos/instrumentação , Procedimentos Cirúrgicos Minimamente Invasivos/tendências , Ensaios Clínicos Controlados Aleatórios como Assunto/métodos , Doenças da Coluna Vertebral/diagnóstico por imagem , Resultado do Tratamento
14.
Metab Eng ; 60: 77-86, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32247827

RESUMO

Maintaining a metabolic steady state is essential for an organism's fitness and survival when confronted with environmental stress, and metabolic imbalance can be reversed by exposing the organism to fasting. Here, we attempted to apply this physiological principle to mammalian cell cultures to improve cellular fitness and consequently their ability to express recombinant proteins. We showed that transient vitamin B5 deprivation, an essential cofactor of central cellular metabolism, can quickly and irreversibly affect mammalian cell growth and division. A selection method was designed that relies on mammalian cell dependence on vitamin B5 for energy production, using the co-expression of the B5 transporter SLC5A6 and a gene of interest. We demonstrated that vitamin B5 selection persistently activates peroxisome proliferator-activated receptors (PPAR), a family of transcription factors involved in energy homeostasis, thereby altering lipid metabolism, improving cell fitness and therapeutic protein production. Thus, stable PPAR activation may constitute a cellular memory of past deprivation state, providing increased resistance to further potential fasting events. In other words, our results imply that cultured cells, once exposed to metabolic starvation, may display an improved metabolic fitness as compared to non-exposed cells, allowing increased resistance to cellular stress.


Assuntos
Homeostase , Ácido Pantotênico/deficiência , Ácido Pantotênico/metabolismo , Proteínas Recombinantes/biossíntese , Animais , Células CHO , Divisão Celular , Células Cultivadas , Cricetinae , Cricetulus , Metabolismo Energético , Vetores Genéticos , Metabolismo dos Lipídeos/fisiologia , PPAR alfa/biossíntese , PPAR alfa/genética , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Estresse Fisiológico , Simportadores/metabolismo
15.
Biotechnol Bioeng ; 117(4): 1101-1116, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31956982

RESUMO

Despite extensive research conducted to increase protein production from Chinese hamster ovary (CHO) cells, cellular bottlenecks often remain, hindering high yields. In this study, a transcriptomic analysis led to the identification of 32 genes that are consistently upregulated in high producer clones and thus might mediate high productivity. Candidate genes were associated with functions such as signaling, protein folding, cytoskeleton organization, and cell survival. We focused on two engineering targets, Erp27, which binds unfolded proteins and the Erp57 disulfide isomerase in the endoplasmic reticulum, and Foxa1, a pioneering transcription factor involved in organ development. Erp27 moderate overexpression increased production of an easy-to-express antibody, whereas Erp27 and Erp57 co-overexpression increased cell density, viability, and the yield of difficult-to-express proteins. Foxa1 overexpression increased cell density, cell viability, and easy- and difficult-to-express protein yields, whereas it decreased reactive oxygen species late in fed-batch cultures. Foxa1 overexpression upregulated two other candidate genes that increased the production of difficult- and/or easy-to-express proteins, namely Ca3, involved in protecting cells from oxidative stress, and Tagap, involved in signaling and cytoskeleton remodeling. Overall, several genes allowing to overcome CHO cell bottlenecks were identified, including Foxa1, which mediated multiple favorable metabolic changes that improve therapeutic protein yields.


Assuntos
Engenharia Celular/métodos , Fator 3-alfa Nuclear de Hepatócito , Proteínas Recombinantes , Animais , Células CHO , Sobrevivência Celular , Cricetinae , Cricetulus , Fator 3-alfa Nuclear de Hepatócito/genética , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Dobramento de Proteína , Proteínas Recombinantes/análise , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
16.
Biotechnol Bioeng ; 117(4): 1117-1126, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31956990

RESUMO

In this study, we assessed the importance of cytoskeleton organization in the mammalian cells used to produce therapeutic proteins. Two cytoskeletal genes, Actin alpha cardiac muscle 1 (ACTC1) and a guanosine triphosphate GTPase-activating protein (TAGAP), were found to be upregulated in highly productive therapeutic protein-expressing Chinese hamster ovary (CHO) cells selected by the deprivation of vitamin B5. We report here that the overexpression of the ACTC1 protein was able to improve significantly recombinant therapeutic production, as well as to decrease the levels of toxic lactate metabolic by-products. ACTC1 overexpression was accompanied by altered as well as decreased polymerized actin, which was associated with high protein production by CHO cell cultured in suspension. We suggest that the depolymerization of actin and the possible modulation of integrin signaling, as well as changes in basal metabolism, may be driving the increase of protein secretion by CHO cells.


Assuntos
Citoesqueleto de Actina , Actinas , Proteínas Recombinantes , Citoesqueleto de Actina/química , Citoesqueleto de Actina/genética , Citoesqueleto de Actina/metabolismo , Actinas/genética , Actinas/metabolismo , Animais , Células CHO , Cricetinae , Cricetulus , Proteínas Ativadoras de GTPase/genética , Proteínas Ativadoras de GTPase/metabolismo , Proteínas Recombinantes/análise , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
17.
Biotechnol Bioeng ; 117(2): 466-485, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31631325

RESUMO

The Chinese hamster ovary (CHO) cells used to produce biopharmaceutical proteins are known to contain type-C endogenous retrovirus (ERV) sequences in their genome and to release retroviral-like particles. Although evidence for their infectivity is missing, this has raised safety concerns. As the genomic origin of these particles remained unclear, we characterized type-C ERV elements at the genome, transcriptome, and viral particle RNA levels. We identified 173 type-C ERV sequences clustering into three functionally conserved groups. Transcripts from one type-C ERV group were full-length, with intact open reading frames, and cognate viral genome RNA was loaded into retroviral-like particles, suggesting that this ERV group may produce functional viruses. CRISPR-Cas9 genome editing was used to disrupt the gag gene of the expressed type-C ERV group. Comparison of CRISPR-derived mutations at the DNA and RNA level led to the identification of a single ERV as the main source of the release of RNA-loaded viral particles. Clones bearing a Gag loss-of-function mutation in this ERV showed a reduction of RNA-containing viral particle release down to detection limits, without compromising cell growth or therapeutic protein production. Overall, our study provides a strategy to mitigate potential viral particle contaminations resulting from ERVs during biopharmaceutical manufacturing.


Assuntos
Células CHO/virologia , Retrovirus Endógenos , Mutagênese Sítio-Dirigida/métodos , RNA Viral , Vírion/genética , Animais , Sistemas CRISPR-Cas , Cricetinae , Cricetulus , Contaminação de Medicamentos/prevenção & controle , Retrovirus Endógenos/genética , Retrovirus Endógenos/metabolismo , Edição de Genes , Genoma Viral/genética , Mutação com Perda de Função/genética , RNA Viral/genética , RNA Viral/metabolismo
18.
BMJ Open ; 9(9): e030389, 2019 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-31501123

RESUMO

INTRODUCTION: Robotic guidance (RG) and computer-assisted navigation (NV) have seen increased adoption in instrumented spine surgery over the last decade. Although there exists some evidence that these techniques increase radiological pedicle screw accuracy compared with conventional freehand (FH) surgery, this may not directly translate to any tangible clinical benefits, especially considering the relatively high inherent costs. As a non-randomised, expertise-based study, the European Robotic Spinal Instrumentation Study aims to create prospective multicentre evidence on the potential comparative clinical benefits of RG, NV and FH in a real-world setting. METHODS AND ANALYSIS: Patients are allocated in a non-randomised, non-blinded fashion to the RG, NV or FH arms. Adult patients that are to undergo thoracolumbar pedicle screw instrumentation for degenerative pathologies, infections, vertebral tumours or fractures are considered for inclusion. Deformity correction and surgery at more than five levels represent exclusion criteria. Follow-up takes place at 6 weeks, as well as 12 and 24 months. The primary endpoint is defined as the time to revision surgery for a malpositioned or loosened pedicle screw within the first postoperative year. Secondary endpoints include patient-reported back and leg pain, as well as Oswestry Disability Index and EuroQOL 5-dimension questionnaires. Use of analgesic medication and work status are recorded. The primary analysis, conducted on the 12-month data, is carried out according to the intention-to-treat principle. The primary endpoint is analysed using crude and adjusted Cox proportional hazards models. Patient-reported outcomes are analysed using baseline-adjusted linear mixed models. The study is monitored according to a prespecified monitoring plan. ETHICS AND DISSEMINATION: The study protocol is approved by the appropriate national and local authorities. Written informed consent is obtained from all participants. The final results will be published in an international peer-reviewed journal. TRIAL REGISTRATION NUMBER: Clinical Trials.gov registry NCT03398915; Pre-results, recruiting stage.


Assuntos
Vértebras Lombares , Parafusos Pediculares/efeitos adversos , Complicações Pós-Operatórias , Reoperação , Procedimentos Cirúrgicos Robóticos , Doenças da Coluna Vertebral , Fusão Vertebral , Vértebras Torácicas , Europa (Continente)/epidemiologia , Feminino , Humanos , Vértebras Lombares/diagnóstico por imagem , Vértebras Lombares/cirurgia , Masculino , Pessoa de Meia-Idade , Estudos Multicêntricos como Assunto , Estudos Observacionais como Assunto , Avaliação de Resultados em Cuidados de Saúde , Complicações Pós-Operatórias/diagnóstico , Complicações Pós-Operatórias/etiologia , Complicações Pós-Operatórias/cirurgia , Reoperação/métodos , Reoperação/estatística & dados numéricos , Procedimentos Cirúrgicos Robóticos/efeitos adversos , Procedimentos Cirúrgicos Robóticos/instrumentação , Procedimentos Cirúrgicos Robóticos/métodos , Doenças da Coluna Vertebral/diagnóstico por imagem , Doenças da Coluna Vertebral/epidemiologia , Doenças da Coluna Vertebral/cirurgia , Fusão Vertebral/efeitos adversos , Fusão Vertebral/instrumentação , Fusão Vertebral/métodos , Cirurgia Assistida por Computador/efeitos adversos , Cirurgia Assistida por Computador/instrumentação , Cirurgia Assistida por Computador/métodos , Vértebras Torácicas/diagnóstico por imagem , Vértebras Torácicas/cirurgia
19.
J Biotechnol ; 305: 23-26, 2019 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-31442502

RESUMO

Cell counting and viability assessment is an integral part of mammalian cell line development. While manual counting with a hemocytometer is still the gold standard method, its subjectivity and high labor intensity has resulted in its reduced use in favor of automated systems. In addition, some of these automated systems offer multiwell plate based high throughput cell count, which is an asset for biopharmaceutical companies generating hundreds of high-performance cell lines per year. In this study, we used Chinese Hamster Ovary (CHO)-K1 cells cultured in suspension in order to evaluate two automated viable-cell counters, the Guava® easyCyte HT and the CytoFLEX®, for their performance in monitoring Viable Cell Density (VCD) and viability. Our results show that specificity, accuracy, precision and repeatability was comparable between the two systems and when compared to manual counting, thus providing efficient alternatives particularly when analyzing high sample numbers in a daily mode.


Assuntos
Contagem de Células/métodos , Técnicas de Cultura de Células/métodos , Ensaios de Triagem em Larga Escala/métodos , Animais , Células CHO , Sobrevivência Celular , Cricetulus
20.
Sci Rep ; 8(1): 11839, 2018 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-30087364

RESUMO

Drosophila suzukii is an invasive fly of East Asian origin that has become a serious fruit pest worldwide. Classical biological control through the introduction of parasitoids from Asia could help reduce populations of D. suzukii in invaded regions. Little is known about the native parasitoids of the fly in Asia. Therefore, surveys for larval parasitoids of D. suzukii were carried out in China and Japan between 2015 and 2017. Parasitoids of D. suzukii and other fruit-inhabiting drosophilids (D. pulchrella and D. subpulchrella) that are probably attacked by the same parasitoid complex were found in four Chinese provinces and four Japanese prefectures. Larval parasitoids were obtained at most sites where D. suzukii was found, with parasitism varying from 0.0 to 75.6%. At least eight parasitoid species were reared. The most abundant and frequent parasitoids were the Figitidae Ganaspis cf. brasiliensis and Leptopilina japonica, but another Leptopilina species and at least five Braconidae species belonging to the genera Areotetes, Asobara and Tanycarpa were obtained in low numbers. Due to its likely restricted host range, the most promising parasitoid for biological control is Ganaspis cf. brasiliensis. However, its exact specificity and taxonomic status require future research.


Assuntos
Formigas/fisiologia , Drosophila/fisiologia , Frutas/parasitologia , Controle Biológico de Vetores/métodos , Vespas/fisiologia , Animais , China , Drosophila/classificação , Geografia , Especificidade de Hospedeiro , Interações Hospedeiro-Parasita , Japão , Larva/fisiologia
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