Prevalence of neutralizing antibodies against adeno-associated virus (AAV) types 2, 5, and 6 in cystic fibrosis and normal populations: Implications for gene therapy using AAV vectors

Adeno-associated virus (AAV) vectors are promising candidates for gene therapy directed to the lungs, in particular for treatment of cystic fibrosis (CF). In animal models of lung gene transfer, neutralizing antibodies in serum made in response to vector exposure have been associated with a partial to complete block to repeat transduction by vectors with the same capsid type, thus transduction by AAV vectors might be inefficient in humans previously exposed to the same AAV type. AAV type 2 (AAV2) has been used in clinical trials of lung gene transfer, but AAV5 and AAV6 have been shown to mediate more efficient transduction in rodent lungs and in cultured human airway epithelia compared with that of AAV2. Here we have measured neutralizing antibodies against AAV type 2, 5, and 6 vectors in serum from children and adults with CF, and from normal adults. About 30% of adults were seropositive for AAV2, 20-30% were seropositive for AAV6, and 10-20% were seropositive for AAV5. CF children were seropositive for AAV type 2, 5, or 6 at rates of 4-15%. All individuals seropositive for AAV6 were also seropositive for AAV2, and the AAV6 titer was low compared with the AAV2 titer. AAV5-positive sera were lower both in titers and rates than those seen for AAV6. The results indicate that AAV type 2, 5 or 6 exposure is low in CF and control populations and even lower in CF children.     

Dynamic analysis of MAPK signaling using a high-throughput microfluidic single-cell imaging platform

Cells have evolved biomolecular networks that process and respond to changing chemical environments. Understanding how complex protein interactions give rise to emergent network properties requires time-resolved analysis of cellular response under a large number of genetic perturbations and chemical environments. To date, the lack of technologies for scalable cell analysis under well-controlled and time-varying conditions has made such global studies either impossible or impractical. To address this need, we have developed a high-throughput microfluidic imaging platform for single-cell studies of network response under hundreds of combined genetic perturbations and time-varying stimulant sequences. Our platform combines programmable on-chip mixing and perfusion with high-throughput image acquisition and processing to perform 256 simultaneous time-lapse live-cell imaging experiments. Nonadherent cells are captured in an array of 2,048 microfluidic cell traps to allow for the imaging of eight different genotypes over 12 h and in response to 32 unique sequences of stimulation, generating a total of 49,000 images per run. Using 12 devices, we carried out >3,000 live-cell imaging experiments to investigate the mating pheromone response in Saccharomyces cerevisiae under combined genetic perturbations and changing environmental conditions. Comprehensive analysis of 11 deletion mutants reveals both distinct thresholds for morphological switching and new dynamic phenotypes that are not observed in static conditions. For example, kss1Δ, fus3Δ, msg5Δ, and ptp2Δ mutants exhibit distinctive stimulus-frequency-dependent signaling phenotypes, implicating their role in filtering and network memory. The combination of parallel microfluidic control with high-throughput imaging provides a powerful tool for systems-level studies of single-cell decision making.     

Risk Stratification Algorithm for Orthopedic Trauma Patients at Risk for Fat Embolism Syndrome

BackgroundFat 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.MethodsElectronic 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).Results18 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).ConclusionFindings 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 evidenceLevel 3.     

The relationship between sleep duration and obesity in Turkish children and adolescents

Aim: To determine the relationship between sleep duration and obesity in Turkish children and adolescents.
Methods: This study was conducted in Turkey with 5358 children aged 6 to 17 years. Height, weight, waist circumference (WC), mid-upper arm circumference (MUAC), triceps skinfold thickness were measured. Body mass index (BMI), arm fat area were calculated. Self-reported sleep duration by parents were obtained.
Results: As sleep duration increased, BMI, which was significantly higher in girls sleeping ≤8 h, decreased (p < 0.05). WC, MUAC, BMI were significantly higher in boys sleeping ≤8 h versus males sleeping ≥10 h. Boys sleeping ≤10 h in 6.0–17.0-years had significantly higher risk of overweight/obesity. In 6.0 to 17.0 years, the risk of overweight/obesity in boys sleeping 9–10 h, 8–9 h and ≤8 h were 1.86-, 1.74- and 2.06-times higher respectively, versus children sleeping ≥10 h (p < 0.05).
Conclusion: Sleep duration may be an important factor for obesity and providing ≥10 h of sleep is recommended as a prevention strategy for obesity.     

The influence of cyclic tensile strain on multi-compartment collagen-GAG scaffolds for tendon-bone junction repair

ABSTRACT

Background: Orthopedic injuries often occur at the interface between soft tissues and bone. The tendon-bone junction (TBJ) is a classic example of such an interface. Current clinical strategies for TBJ injuries prioritize mechanical reattachment over regeneration of the native interface, resulting in poor outcomes. The need to promote regenerative healing of spatially-graded tissues inspires our effort to develop new tissue engineering technologies that replicate features of the spatially-graded extracellular matrix and strain profiles across the native TBJ.

Materials and Methods: We recently described a biphasic collagen-glycosaminoglycan (CG) scaffold containing distinct compartment with divergent mineral content and structural alignment (isotropic vs. anisotropic) linked by a continuous interface zone to mimic structural and compositional features of the native TBJ.

Results: Here, we report application of cyclic tensile strain (CTS) to the scaffold via a bioreactor leads to non-uniform strain profiles across the spatially-graded scaffold. Further, combinations of CTS and matrix structural features promote rapid, spatially-distinct differentiation profiles of human bone marrow-derived mesenchymal stem cells (MSCs) down multiple osteotendinous lineages. CTS preferentially upregulates MSC activity and tenogenic differentiation in the anisotropic region of the scaffold. This work demonstrates a tissue engineering approach that couples instructive biomaterials with cyclic tensile stimuli to promote regenerative healing of orthopedic interfaces.     

Epidermal growth factor receptor genomic variation in NSCLC patients receiving tyrosine kinase inhibitor therapy: a systematic review and meta-analysis

Introduction  

The objective of this analysis was to examine the relationship between genomic variation and health outcomes in studies performed in non-small cell lung cancer (NSCLC) patients treated with single agent epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) using a systematic review with statistical pooling of data.