Critical considerations for the development of potency tests for therapeutic applications of mesenchymal stromal cell-derived small extracellular vesicles

Mesenchymal stromal/stem cells (MSCs) have been widely tested against many diseases, with more than 1000 registered clinical trials worldwide. Despite many setbacks, MSCs have been approved for the treatment of graft-versus-host disease and Crohn disease. However, it is increasingly clear that MSCs exert their therapeutic functions in a paracrine manner through the secretion of small extracellular vesicles (sEVs) of 50–200 nm in diameter. Unlike living cells that can persist long-term, sEVs are non-living and non-replicative and have a transient presence in the body. Their small size also renders sEV preparations highly amenable to sterilization by filtration. Together, acellular MSC-sEV preparations are potentially safer and easier to translate into the clinic than cellular MSC products. Nevertheless, there are inherent challenges in the development of MSC-sEV drug products. MSC-sEVs are products of living cells, and living cells are sensitive to changes in the external microenvironment. Consequently, quality control metrics to measure key identity and potency features of MSC-sEV preparations have to be specified during development of MSC-sEV therapeutics. The authors have previously described quantifiable assays to define the identity of MSC-sEVs. Here the authors discuss requirements for prospective potency assays to predict the therapeutic effectiveness of the drug substance in accordance with International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use guidelines. Although potency assays should ideally reflect the mechanism of action (MoA), this is challenging because the MoA for the reported efficacy of MSC-sEV preparations against multiple diseases of diverse underlying pathology is likely to be complex and different for each disease and difficult to fully elucidate. Nevertheless, robust potency assays could be developed by identifying the EV attribute most relevant to the intended biological activity in EV-mediated therapy and quantifying the EV attribute. Specifically, the authors highlight challenges and mitigation measures to enhance the manufacture of consistent and reproducibly potent sEV preparations, to identify and select the appropriate EV attribute for potency assays despite a complex “work-in-progress” MoA and to develop assays likely to be compliant with regulatory guidance for assay validation.     

Mps1 promotes poleward chromosome movements in meiotic prometaphase

In prophase of meiosis I, homologous chromosomes pair and become connected by cross-overs. Chiasmata, the connections formed by cross-overs, enable the chromosome pair, called a bivalent, to attach as a single unit to the spindle. When the meiotic spindle forms in prometaphase, most bivalents are associated with one spindle pole and then go through a series of oscillations on the spindle, attaching to and detaching from microtubules until the partners of the bivalent become bioriented—attached to microtubules from opposite sides of the spindle. The conserved kinase, Mps1, is essential for the bivalents to be pulled by microtubules across the spindle in prometaphase. Here we show that MPS1 is needed for efficient triggering of the migration of microtubule-attached kinetochores toward the poles and promotes microtubule depolymerization. Our data support the model Mps1 acts at the kinetochore to coordinate the successful attachment of a microtubule and the triggering of microtubule depolymerization to then move the chromosome.     

Caring for US Children: Barriers to Effective Treatment in Children with the Disease of Obesity

In 2020, impediments to pediatric obesity (PO) treatment remain pervasive, even though these barriers are clearly documented in medical literature. Providers must invest considerable resources to overcome these barriers to care. Notable barriers include gaps in medical education, misperceptions of the disease, weight bias and stigma, exclusion of coverage in health plans, and thus an unsustainable financial framework. Hence, this review offers an updated social‐ecological framework of accessibility to care, wherein each barrier to care or variable is interdependent on the other and each is critical to creating forward momentum. The sum of all these variables is instrumental to overall smooth function, configured as a wheel. To treat PO effectively, all variables must be adequately addressed by stakeholders throughout the health care system in order to holistically comprehend and appreciate undertakings to advance the burgeoning field of PO medicine.     

Interrupted incubation: How dabbling ducks respond when flushed from the nest

Nesting birds must provide a thermal environment sufficient for egg development while also meeting self‐maintenance needs. Many birds, particularly those with uniparental incubation, achieve this balance through periodic incubation recesses, during which foraging and other self‐maintenance activities can occur. However, incubating birds may experience disturbances such as predator or human activity which interrupt natural incubation patterns by compelling them to leave the nest. We characterized incubating mallard Anas platyrhynchos and gadwall Mareca strepera hens’ responses when flushed by predators and investigators in Suisun Marsh, California, USA. Diurnal incubation recesses initiated by investigators approaching nests were 63% longer than natural diurnal incubation recesses initiated by the hen (geometric mean: 226.77 min versus 142.04 min). Nocturnal incubation recesses, many of which were likely the result of predators flushing hens, were of similar duration regardless of whether the nest was partially depredated during the event (115.33 [101.01;131.68] minutes) or not (119.62 [111.96;127.82] minutes), yet were 16% shorter than natural diurnal incubation recesses. Hens moved further from the nest during natural diurnal recesses or investigator‐initiated recesses than during nocturnal recesses, and the proportion of hen locations recorded in wetland versus upland habitat during recesses varied with recess type (model‐predicted means: natural diurnal recess 0.77; investigator‐initiated recess 0.82; nocturnal recess 0.31). Hens were more likely to take a natural recess following an investigator‐initiated recess earlier that same day than following a natural recess earlier that same day, and natural recesses that followed an investigator‐initiated recess were longer than natural recesses that followed an earlier natural recess, suggesting that hens may not fulfill all of their physiological needs during investigator‐initiated recesses. We found no evidence that the duration of investigator‐initiated recesses was influenced by repeated visits to the nest, whether by predators or by investigators, and trapping and handling the hen did not affect investigator‐initiated recess duration unless the hen was also fitted with a backpack‐harness style GPS–GSM transmitter at the time of capture. Hens that were captured and fitted with GPS–GSM transmitters took recesses that were 26% longer than recesses during which a hen was captured but a GPS–GSM transmitter was not attached. Incubation interruptions had measurable but limited and specific effects on hen behavior.     

Multicomponent shell traits are consistent with an individual recognition function of the appearance of common murre (Uria aalge) eggs: A biological replication study

In dense breeding colonies, and despite having no nest structure, common murres (or guillemots: Uria aalge) are still able to identify their own eggs. Each female murre''s egg is thought to be recognized individually by the shell''s avian‐perceivable traits. This is because the eggshells’ visible traits conform to expectations of the identity‐signaling hypothesis in that they show both high intraindividual repeatability and high interindividual variability. Identity signaling also predicts a lack of correlation between each of the putative multicomponent recognition traits, yielding no significant relationships between those eggshell traits that are generated by mutually exclusive physiological factors. Using a multivariate analysis across eggshell size and shape, avian‐perceivable background coloration, spot (maculation) shape, and spot density, we detected no unexpected statistical correlations between Icelandic common murre egg traits lacking known physiological or mathematical relationships with one another. These results biologically replicate the conclusions of a recent eggshell trait study of Canadian common murres using similar methodology. We also demonstrate the use of static correlations to infer identity signaling function without direct behavioral observations, which in turn may also be applied to rare or extinct species and provide valuable insight into otherwise unknown communicative and behavioral functions.     

Planting Design Effects on Avian Seed Dispersers in a Tropical Forest Restoration Experiment

Seed dispersal often limits tropical forest regeneration and animals disperse most rainforest tree seeds. This presents two important questions for restoration ecologists: (1) which animals are common seed dispersers? and (2) which restoration techniques attract them? Fourteen restoration sites were planted with four tree species in three designs, (1) controls (no planting, natural regeneration) (2) islands (trees planted in small patches), and (3) plantations (trees planted continuously over a large patch). We sampled birds in November, February, and April 2007–2008 with mist nets, in February and July 2009 with observations, and in July 2008 with both techniques. We documented 30 seed species from fecal samples of captured birds. All identified seed species were early‐successional forms. Four tanager species, three thrushes, two saltators, two flycatchers, and one finch were categorized as common seed dispersers, based on their high likelihood of dispersing seeds. Common dispersers were generalist species with small gape widths (<15 mm). Common dispersers were captured significantly more often in plantations than controls in most seasons and more often in plantations than islands during one season. Common disperser observations were significantly greater in plantations than controls during two periods and in plantations compared with islands in one period. Results indicate that plantation‐style planting is the conservative strategy to maximize attractiveness to common dispersers in tropical restoration sites. Island planting is an alternative when resources are limited although disperser activity may be lower in some seasons than in plantations. Additional research should investigate how to attract large, forest‐associated dispersers.     

Finite element analysis of Stryker Xia pedicle screw in artificial bone samples with and without supplemental cement augmentation

A validated, using in vitro biomechanical testing, finite element model was used to evaluate the affects of (1) cement augmentation and (2) an intact posterior cortex in osteoporotic bone. The presence of augmentation and/or a posterior cortical cortex increased the stabilization of the pedicle screw 2–5 fold. Placement of cement influenced failure load and toggle; with distal placement having the largest increase in failure load and decrease in cephalad–caudad toggle. The presence of posterior cortex caused a decrease in the amount of toggle, a proximal shift of the center of rotation and an increase in the maximum failure force.     

Neighboring Parenchyma Cells Contribute to Arabidopsis Xylem Lignification,while Lignification of Interfascicular Fibers Is Cell Autonomous

Lignin is a critical structural component of plants, providing vascular integrity and mechanical strength. Lignin precursors (monolignols) must be exported to the extracellular matrix where random oxidative coupling produces a complex lignin polymer. The objectives of this study were twofold: to determine the timing of lignification with respect to programmed cell death and to test if nonlignifying xylary parenchyma cells can contribute to the lignification of tracheary elements and fibers. This study demonstrates that lignin deposition is not exclusively a postmortem event, but also occurs prior to programmed cell death. Radiolabeled monolignols were not detected in the cytoplasm or vacuoles of tracheary elements or neighbors. To experimentally define which cells in lignifying tissues contribute to lignification in intact plants, a microRNA against CINNAMOYL CoA-REDUCTASE1 driven by the promoter from CELLULOSE SYNTHASE7 (ProCESA7:miRNA CCR1) was used to silence monolignol biosynthesis specifically in cells developing lignified secondary cell walls. When monolignol biosynthesis in ProCESA7:miRNA CCR1 lines was silenced in the lignifying cells themselves, but not in the neighboring cells, lignin was still deposited in the xylem secondary cell walls. Surprisingly, a dramatic reduction in cell wall lignification of extraxylary fiber cells demonstrates that extraxylary fibers undergo cell autonomous lignification.