Super7 passaging method to improve Chinese hamster ovary cell fed-batch performance.

Chinese hamster ovary (CHO) cells are widely used to manufacture biopharmaceuticals, most of all monoclonal antibodies (mAbs). Some CHO cell lines exhibit production instability, where the productivity of the cells decreases as a function of time in culture. To counter this, we designed a passaging strategy that, rather than maximizing the time spent in log-growth phase, mimics the first 7 days of a fed-batch production process. Cultures passaged using this method had lower net growth rates and were more oxidative throughout 6 weeks of passaging. Fed-batch cultures inoculated by cells passaged using this method had increased net growth rates, oxidative metabolism, and volumetric productivity compared to cells passaged using a conventional strategy. Cells from unstable cell lines passaged by this new method produced 80%-160% more mAbs per unit volume than cells passaged by a conventional method. This new method, named Super7, provides the ability to mitigate the impact of production instability in CHO-K1 cell lines without a need for further cell line creation, genetic engineering, or medium development.

Opposing global change drivers counterbalance trends in breeding North American monarch butterflies.

Many insects are in clear decline, with monarch butterflies (Danaus plexippus) drawing particular attention as a flagship species. It is well documented that, among migratory populations, numbers of overwintering monarchs have been falling across several decades, but trends among breeding monarchs are less clear. Here, we compile >135,000 monarch observations between 1993 and 2018 from the North American Butterfly Association's annual butterfly count to examine spatiotemporal patterns and potential drivers of adult monarch relative abundance trends across the entire breeding range in eastern and western North America. While the data revealed declines at some sites, particularly the US Northeast and parts of the Midwest, numbers in other areas, notably the US Southeast and Northwest, were unchanged or increasing, yielding a slightly positive overall trend across the species range. Negative impacts of agricultural glyphosate use appeared to be counterbalanced by positive effects of annual temperature, particularly in the US Midwest. Overall, our results suggest that population growth in summer is compensating for losses during the winter and that changing environmental variables have offsetting effects on mortality and/or reproduction. We suggest that density-dependent reproductive compensation when lower numbers arrive each spring is currently able to maintain relatively stable breeding monarch numbers. However, we caution against complacency since accelerating climate change may bring growing threats. In addition, increases of summer monarchs in some regions, especially in California and in the south, may reflect replacement of migratory with resident populations. Nonetheless, it is perhaps reassuring that ubiquitous downward trends in summer monarch abundance are not evident.

Complex life histories predispose aphids to recent abundance declines.

Many animals change feeding habits as they progress through life stages, exploiting resources that vary in space and time. However, complex life histories may bring new risks if rapid environmental change disrupts the timing of these switches. Here, we use abundance times series for a diverse group of herbivorous insects, aphids, to search for trait and environmental characteristics associated with declines. Our meta dataset spanned three world regions and >300 aphid species, tracked at 75 individual sites for 10-50 years. Abundances were generally falling, with median changes of -8.3%, -5.6%, and -0.1% per year in the central USA, northwestern USA, and United Kingdom, respectively. Aphids that obligately alternated between host plants annually and those that were agricultural pests exhibited the steepest declines, relative to species able to persist on the same host plant year-round or those in natural areas. This suggests that host alternation might expose aphids to climate-induced phenology mismatches with one or more of their host plant species, with additional risks from exposure to insecticides and other management efforts. Warming temperatures through time were associated with milder aphid declines or even abundance increases, particularly at higher latitudes. Altogether, while a warming world appeared to benefit some aphid species in some places, most aphid species that had time-sensitive movements among multiple host plants seemed to face greater risk of decline. More generally, this suggests that recent human-induced rapid environmental change is rebalancing the risks and rewards associated with complex life histories.

Recent climate change is creating hotspots of butterfly increase and decline across North America.

Some insect populations are experiencing dramatic declines, endangering the crucial ecosystem services they provide. Yet, other populations appear robust, highlighting the need to better define patterns and underlying drivers of recent change in insect numbers. We examined abundance and biodiversity trends for North American butterflies using a unique citizen-science dataset that has recorded observations of over 8 million butterflies across 456 species, 503 sites, nine ecoregions, and 26 years. Butterflies are a biodiverse group of pollinators, herbivores, and prey, making them useful bellwethers of environmental change. We found great heterogeneity in butterfly species' abundance trends, aggregating near zero, but with a tendency toward decline. There was strong spatial clustering, however, into regions of increase, decrease, or relative stasis. Recent precipitation and temperature appeared to largely drive these patterns, with butterflies generally declining at increasingly dry and hot sites but increasing at relatively wet or cool sites. In contrast, landscape and butterfly trait predictors had little influence, though abundance trends were slightly more positive around urban areas. Consistent with varying responses by different species, no overall directional change in butterfly species richness or evenness was detected. Overall, a mosaic of butterfly decay and rebound hotspots appeared to largely reflect geographic variability in climate drivers. Ongoing controversy about insect declines might dissipate with a shift in focus to the causes of heterogeneous responses among taxa and sites, with climate change emerging as a key suspect when pollinator communities are broadly impacted.

LncRNA Meg3 protects endothelial function by regulating the DNA damage response.

The role of long non-coding RNAs (lncRNAs) in regulating endothelial function through the DNA damage response (DDR) remains poorly understood. In this study, we demonstrate that lncRNA maternally expressed gene 3 (Meg3) interacts with the RNA binding protein polypyrimidine tract binding protein 3 (PTBP3) to regulate gene expression and endothelial function through p53 signaling ─ a major coordinator of apoptosis and cell proliferation triggered by the DDR. Meg3 expression is induced in endothelial cells (ECs) upon p53 activation. Meg3 silencing induces DNA damage, activates p53 signaling, increases the expression of p53 target genes, promotes EC apoptosis, and inhibits EC proliferation. Mechanistically, Meg3 silencing reduces the interaction of p53 with Mdm2, induces p53 expression, and promotes the association of p53 with the promoters of a subset of p53 target genes. PTBP3 silencing recapitulates the effects of Meg3 deficiency on the expression of p53 target genes, EC apoptosis and proliferation. The Meg3-dependent association of PTBP3 with the promoters of p53 target genes suggests that Meg3 and PTBP3 restrain p53 activation. Our studies reveal a novel role of Meg3 and PTBP3 in regulating p53 signaling and endothelial function, which may serve as novel targets for therapies to restore endothelial homeostasis.

Cut points of the Actigraph GT9X for moderate and vigorous intensity physical activity at four different wear locations.

Accelerometer cut points are an important consideration for distinguishing the intensity of activity into categories such as moderate and vigorous. It is well-established in the literature that these cut points depend on a variety of factors, including age group, device, and wear location. The Actigraph GT9X is a newer model accelerometer that is used for physical activity research, but existing cut points for this device are limited since it is a newer device. Furthermore, there is not existing data on cut points for the GT9X at the ankle or foot locations, which offers some potential benefit for activities that do not involve arm and/or core motion. A total of N = 44 adults completed a four-stage treadmill protocol while wearing Actigraph GT9X sensors at four different locations: foot, ankle, wrist, and hip. Metabolic Equivalent of Task (MET) levels assessed by indirect calorimetry along with Receiver Operating Characteristic (ROC) curves were used to establish cut points for moderate and vigorous intensity for each wear location of the GT9X. Area under the ROC curves indicated high discrimination accuracy for each case.

Potential for Reclamation of Abandoned Gas Wells to Restore Ecosystem Services in the Fayetteville Shale of Arkansas.

Unconventional oil and gas (UOG) drilling has expanded rapidly across the United States, including in the Fayetteville Shale formation in north-central Arkansas where drilling began in 2004. As one of the oldest regions of UOG activity in the United States, this area has experienced significant land-use changes, specifically development of natural habitat and agricultural land for gas infrastructure. In recent years, drilling of new wells has stopped and production has declined. By 2017, 1038 wells had ceased production and been abandoned, which makes them eligible for land reclamation. However, most of these sites (80%) have not been reclaimed and continue to cause losses in ecosystem services. If reclamation was performed on lands associated with abandoned infrastructure, we estimate more than $2 million USD annually in agricultural, timber, and carbon sequestration values would be gained. These benefits far outweigh the costs of reclamation, especially since the benefits accrue over time and reclamation is a short-term cost. Our estimates indicate a 2-4 year break-even time period when cumulative ecosystem services benefits will outweigh reclamation costs. We predicted a well-abandonment rate of 155 per year until 2050 when 98% of wells will be abandoned, which indicates great potential for future ecosystem services restoration. Thus, we recommend that Arkansans at the government and citizen level work to restore lands impacted by UOG development in the Fayetteville Shale region so that their value to landowners and society can be recovered, which will enhance long-term economic and environmental benefits.

Tibial Accelerations During the Single-Leg Hop Test: Influence of Fixation.

CONTEXT: Performance on single-leg hopping (SLH) assessments is commonly included within return-to-sport criteria for rehabilitating athletes. Triaxial accelerometers have been used to quantify impact loading in a variety of movements, including hopping; however, they have never been attached to the tibia during SLH, and their method of fixation has not been investigated. OBJECTIVE: The purpose of this study was to quantify triaxial accelerations and evaluate the influence of the fixation method of a lightweight inertial measurement unit (Blue Trident) mounted to the tibia during SLH performance. DESIGN: Single cohort, repeated-measures experimental design. PARTICIPANTS: Sixteen healthy participants (10 females and 6 males; 20 [0.9] y; 1.67 [0.08] m; 66.0 [8.5] kg) met the inclusion criteria, volunteered, and completed this study. INTERVENTIONS: Participants performed 2 sets of 3 SLH trials with an inertial measurement unit (1500 Hz) fixated to the tibia, each set with 1 of 2 attachment methods (double-sided tape [DST] with athletic tape and silicon strap [SS] with Velcro adhesion). MAIN OUTCOME MEASURES: Hop distance, peak tibial acceleration (PTA), time to PTA, and the acceleration slope were assessed during each hop landing. RESULTS: Repeated-measures analysis of variance determined no significant effect of the attachment method on hop metrics (P = .252). Across 3 trials, both fixation methods (DST and SS) had excellent reliability values (intraclass correlation coefficient: .868-.941) for PTA and acceleration slope but not for time to PTA (intraclass correlation coefficient: .397-.768). The PTA for DST (27.22 [7.94] g) and SS (26.21 [10.48] g) was comparable and had a moderate, positive relationship (DST: r = .72, P < .01; SS: r = .77, P < .01) to SLH distance. CONCLUSIONS: Tibial inertial measurement units with triaxial accelerometers can reliably assess PTA during performance of the SLH, and SS is a viable alternative tibial attachment to DST.

Accuracy of Functional Movement Screen Deep Squat Scoring and the Influence of Optimized Scoring Criteria: A 3-Dimensional Kinematic Approach.

CONTEXT: The deep squat (DS) test is a component of the functional movement screen, which is used to assess the quality of fundamental movement patterns; however, the accuracy of the DS has not been studied. The DS is a complex, total body movement pattern with evaluation required at several points along the kinematic chain. OBJECTIVE: To assess the accuracy of DS scoring by an athletic trainer, physical therapist, and exercise science professional via a comparative analysis with kinematic data (KD) and to identify scoring criteria that would improve agreement between raters and KD scores. DESIGN: Cross-sectional study. SETTING: Motion analysis laboratory. PARTICIPANTS: A rater from each of 3 movement science disciplines rated the DS of 23 male college athletes (20.3 [1.2] y; 70.5 [3.5] kg). INTERVENTIONS: Subjects were outfitted with reflective markers and asked to perform the DS. The DS performance was scored by 3 raters and kinematic analysis. Subsequently, the optimal set of criteria that minimized the difference between mode rater score and KD was determined via a Nelder-Mead simplex optimization routine. MAIN OUTCOME MEASURES: Intraclass correlation coefficients (ICCs) were calculated using SPSS (version 23; IBM, Armonk, NY) to determine tester agreement with the KD score and between the mode score and KD score. RESULTS: Agreement was poor for the athletic trainer (ICC = .387), physical therapist (ICC = .298), exercise science professional (ICC = .378), and raters' DS scores when compared with the KD. Agreement was poor for the mode score when compared with KD prior to optimization and good following optimization (ICC = .830), thereby allowing identification of specific scoring errors. CONCLUSIONS: Agreement for DS scores is poor when compared with KD; however, it may be improved with optimization of DS scoring criteria.

Reliability of Smartphone Inclinometry to Measure Upper Cervical Range of Motion.

CONTEXT: Assessment of upper cervical range of motion (UCROM) and mobility is commonly performed in the clinical setting for patients suffering from headache, neck pain, and vestibular dysfunction. Reliable and reproducible measurement of this motion is often difficult or too expensive to perform in the clinical setting. Smartphone applications using the device's internal gyroscope offer an easy and inexpensive means of measuring UCROM, but their reliability has not been reported in the literature. OBJECTIVE: To assess the reliability of an inclinometer application installed on 2 different devices (iPhone 6 [IP] and android [AN]) and to measure UCROM in a healthy population. DESIGN: Two examiners assessed passive UCROM. Each examiner was assigned to a specific smartphone, and a repeated-measures design consisting of 3 trials for each examiner-phone was performed. The order of testing was randomized, and the examiners were blinded to UCROM measures. SETTING: Laboratory. PARTICIPANTS: A total of 38 subjects (19 females and 19 males; age: 23.8 [1.2] y) without pain or injury to the neck or spine for at least 3 months. INTERVENTION: Each examiner passively flexed the head fully, rotated the head fully in 1 direction, and then in another. Peak rotation measures were recorded from each smartphone. Three trials were performed for each phone, with a 2-minute break between examiners/phones. MAIN OUTCOME MEASURES: Intraclass correlation coefficient using a 2-way mixed, absolute agreement model was obtained (1) between each examiner-phone and (2) within each examiner-phone for the measurements in each rotation direction. RESULTS: Interphone/examiner reliability comparing average peak and total UCROM for each device was excellent (.87, .81). Intraphone/examiner reliability, determined across 3 trials, was also excellent (AN right rot. = .91, AN left rot. = .96; IP right rot. = .98, IP left rot. = .95). CONCLUSION: UCROM can be reliably measured using a smartphone inclinometer application.

Tracking Steps on Apple Watch at Different Walking Speeds.

KEY POINTS: QUESTION: How accurate are the step counts obtained from Apple Watch? FINDINGS: In this validation study, video steps vs. Apple Watch steps (mean ± SD) were 2965 ± 144 vs. 2964 ± 145 steps; P < 0.001. Lin's concordance correlation coefficient showed a strong correlation (r = 0.96; P < 0.001) between the two measurements. There was a total error of 0.034% (1.07 steps) for the Apple Watch steps when compared with the manual counts obtained from video recordings. MEANING: Our study is one of the initial studies to objectively validate the accuracy of the step counts obtained from Apple watch at different walking speeds. Apple Watch tested to be an extremely accurate device for measuring daily step counts for adults.

The role of interactions of long non-coding RNAs and heterogeneous nuclear ribonucleoproteins in regulating cellular functions.

Long non-coding RNAs (lncRNAs) are emerging as critical regulators of various biological processes and human diseases. The mechanisms of action involve their interactions with proteins, RNA and genomic DNA. Most lncRNAs display strong nuclear localization. Heterogeneous nuclear ribonucleoproteins (hnRNPs) are a large family of RNA-binding proteins that are important for multiple aspects of nucleic acid metabolism. hnRNPs are also predominantly expressed in the nucleus. This review discusses the interactions of lncRNAs and hnRNPs in regulating gene expression at transcriptional and post-transcriptional levels or by changing genomic structure, highlighting their involvements in glucose and lipid metabolism, immune response, DNA damage response, and other cellular functions. Toward the end, several techniques that are used to identify lncRNA binding partners are summarized. There are still many questions that need to be answered in this relatively new research area, which might provide novel targets to control the biological outputs of cells in response to different stimuli.

Impact of Direction of Unloading Influence on Template Rate of Perceived Exertion.

Greer, BK, Young, PR, Thompson, B, Rickert, BJ, and Moran, MF. Impact of direction of unloading influence on template rate of perceived exertion. J Strength Cond Res 32(12): 3407-3413, 2018-It is suggested that exercisers engage in a process of teleoanticipation and create an exercise template based on previous experience with the exercise task that guides their perceptions of the amount of effort required for task completion. This study examined how altering workload intensity during a positive-pressure treadmill task may impact Rating of Perceived Exertion (RPE). In a counterbalanced design, 15 collegiate cross-country runners (7 men and 8 women) performed 2 25-minute runs at a constant velocity, while body mass (BM) was either increased from 60 to 100% (low-to-high progression trial [INC]) or decreased from 100 to 60% (high-to-low progression trial) in 5-minutes increments. Oxygen consumption (V[Combining Dot Above]O2), heart rate (HR), and respiratory exchange ratio (RER) were collected. RPE was recorded at the end of each stage, and energy expenditure (EE) was calculated with V[Combining Dot Above]O2 and RER data. There were no significant differences between direction of loading conditions for V[Combining Dot Above]O2, EE, HR, and RER (p > 0.05). Between-trial differences in RPE at 100, 90, and 80% BM were statistically significant (p < 0.001), with higher RPEs observed during the INC. Differences in RPE observed between conditions cannot be explained by physiological mechanisms. These findings suggest that RPE is a multifaceted construct that can be impacted by subjectively based anticipatory factors such as exercise intensity.

A Comparison of the Impacts of Wind Energy and Unconventional Gas Development on Land-use and Ecosystem Services: An Example from the Anadarko Basin of Oklahoma, USA.

The United States energy industry is transforming with the rapid development of alternative energy sources and technological advancements in fossil fuels. Two major changes include the growth of wind turbines and unconventional oil and gas. We measured land-use impacts and associated ecosystem services costs of unconventional gas and wind energy development within the Anadarko Basin of the Oklahoma Woodford Shale, an area that has experienced large increases in both energy sectors. Unconventional gas wells developed three times as much land compared to wind turbines (on a per unit basis), resulting in higher ecosystem services costs for gas. Gas wells had higher impacts on intensive agricultural lands (i.e., row crops) compared to wind turbines that had higher impacts on natural grasslands/pastures. Because wind turbines produced on average less energy compared to gas wells, the average land-use-related ecosystem cost per gigajoule of energy produced was almost the same. Our results demonstrate that both unconventional gas and wind energy have substantial impacts on land use, which likely affect wildlife populations and land-use-related ecosystem services. Although wind energy does not have the associated greenhouse gas emissions, we suggest that the direct impacts on ecosystems in terms of land use are similar to unconventional fossil fuels. Considering the expected rapid global expansion of these two forms of energy production, many ecosystems are likely to be at risk.

Tibial Acceleration and Spatiotemporal Mechanics in Distance Runners During Reduced-Body-Weight Conditions.

CONTEXT: Treadmills that unload runners via a differential air-pressure (DAP) bladder (eg, AlterG Anti-Gravity Treadmill) are commonly used to reduce effective body weight (BW) in a clinical setting. However, the relationship between the level of unloading and tibial stress is currently unknown. OBJECTIVE: To determine the relationship between tibial impact acceleration and level of BW unloading during running. DESIGN: Cross-sectional. SETTING: University motion-analysis laboratory. PARTICIPANTS: 15 distance runners (9 male, 6 female; 20.4 ± 2.4 y, 60.1 ± 12.6 kg). MAIN OUTCOME MEASURES: Peak tibial acceleration and peak-to-peak tibial acceleration were measured via a uniaxial accelerometer attached to the tibia during a 37-min continuous treadmill run that simulated reduced-BW conditions via a DAP bladder. The trial began with a 10-min run at 100% BW followed by nine 3-min stages where BW was systematically reduced from 95% to 60% in 5% increments. RESULTS: There was no significant relationship between level of BW and either peak tibial acceleration or peak-to-peak tibial acceleration (P > .05). Both heart rate and step rate were significantly reduced with each 5% reduction in BW level (P < .01). CONCLUSIONS: Although ground-reaction forces are reduced when running in reduced-BW conditions on a DAP treadmill, tibial shock magnitudes are unchanged as an alteration in spatiotemporal running mechanics (eg, reduced step rate) and may nullify the unloading effect.

Medium-term patient-reported outcomes after total hip replacement for displaced hip fractures.

INTRODUCTION: The aim of the present study was to define the medium-term outcomes following total hip replacement (THR) for hip fracture. METHODS: We prospectively followed up 92 patients who underwent THR for a displaced hip fracture over a 3-year period between 2007 and 2010. These patients were followed up at 5 years using the Oxford Hip Score, Short-Form 12 (SF-12) questionnaire and satisfaction questionnaire. These outcomes were compared to the short-term outcomes previously reported at 2 years to determine any significant differences. RESULTS: Mean follow-up was at 5.4 years with a mean age at follow-up of 76.5 years. Seventy-four patients (80%) responded. Patients reported excellent functional outcomes and satisfaction (mean Oxford Hip Score 40.3; SF-12 Physical Health Composite Score 44.0; SF-12 Mental Health Composite Score 46.2; mean satisfaction 90%). The rates of dislocation (2%), deep infection (2%) and revision (3%) were comparable to those quoted for elective THR. When compared with 2-year follow-up, there were no statistically significant adverse changes in outcome parameters. CONCLUSIONS: Medium-term outcomes for THR after hip fracture in fit older patients are excellent, and these results demonstrate that the early proven benefits of this surgery are sustained into the midterm.

Trabecular Metal Use in Paprosky Type 2 and 3 Acetabular Defects: 5-Year Follow-Up.

BACKGROUND: The reconstructive challenge of achieving a stable acetabulum in revision total hip arthroplasties in the presence of major osteolytic lesions has led to debate about the most appropriate surgical strategy to minimize implant-related failures. Trabecular metal (TM) implants have become popular but ongoing surveillance of their performance is required. METHODS: We reviewed the clinical and radiological outcome of a consecutive series of 52 patients (55 hips) who had undergone revision total hip arthroplasty for Paprosky type 2 or 3 acetabular defects with TM revision acetabular shells between 2002 and 2008. RESULTS: Four implant failures occurred (2 infections and 2 dislocations). Eleven patients from this cohort died (representing 12 hips) before the 5-year follow-up period giving us a follow-up of 78.2%. Implant survival at 5 years was 92% (95% confidence interval: 80.2%-96.9%). There were no cases of radiological loosening. The mean Oxford hip score was 34 (range, 5-48) at a mean follow-up of 63 months (range, 34-105 months). CONCLUSIONS: We conclude that the use of TM revision shells for complex acetabular reconstruction yields satisfactory results.

Habitat loss and modification due to gas development in the Fayetteville shale.

Hydraulic fracturing and horizontal drilling have become major methods to extract new oil and gas deposits, many of which exist in shale formations in the temperate deciduous biome of the eastern United States. While these technologies have increased natural gas production to new highs, they can have substantial environmental effects. We measured the changes in land use within the maturing Fayetteville Shale gas development region in Arkansas between 2001/2002 and 2012. Our goal was to estimate the land use impact of these new technologies in natural gas drilling and predict future consequences for habitat loss and fragmentation. Loss of natural forest in the gas field was significantly higher compared to areas outside the gas field. The creation of edge habitat, roads, and developed areas was also greater in the gas field. The Fayetteville Shale gas field fully developed about 2% of the natural habitat within the region and increased edge habitat by 1,067 linear km. Our data indicate that without shale gas activities, forest cover would have increased slightly and edge habitat would have decreased slightly, similar to patterns seen recently in many areas of the southern U.S. On average, individual gas wells fully developed about 2.5 ha of land and modified an additional 0.5 ha of natural forest. Considering the large number of wells drilled in other parts of the eastern U.S. and projections for new wells in the future, shale gas development will likely have substantial negative effects on forested habitats and the organisms that depend upon them.