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Advances in the understanding and management of glenoid bone loss in shoulder instability have led to the development of alternative bony reconstruction techniques to the Latarjet using free bone grafts, but little is known about surgeon adoption of these procedures. This study sought to characterize surgeon variation in the use of glenoid bone reconstruction procedures for shoulder instability and ascertain reasons underlying procedure choice.A 9-question survey was created and distributed to 160 shoulder surgeons members of the PacWest Shoulder and Elbow Society, of whom 65 (41%) responded. The survey asked questions regarding fellowship training, years in practice, surgical volume, preferred methods of glenoid bone reconstruction, and reasons underlying treatment choice.All surgeons completed a fellowship, with an equal number of sports medicine fellowship-trained (46%) and shoulder and elbow fellowship-trained (46%) physicians. The majority had been in practice for at least 6 years (6-10 years: 25%;10 years: 59%). Most (78%) performed ≤10 glenoid bony reconstructions per year, and 66% indicated that bony procedures represented10% of their total annual shoulder instability case volume. The open Latarjet was the preferred primary reconstruction method (69%), followed by open free bone block (FBB) (22%), arthroscopic FBB (8%), and arthroscopic Latarjet (1%). Distal tibia allograft (DTA) was the preferred graft (74%) when performing an FBB procedure, followed by iliac crest autograft (18%), and distal clavicle autograft (6%). The top 5 reasons for preferring Latarjet over FBB were the sling effect (57%), the autologous nature of the graft (37%), its robust clinical evidence (22%), low cost (17%), and availability (11%). The top 5 reasons for choosing an FBB procedure were less anatomic disruption (58%), lower complication rate (21%), restoration of articular cartilage interface (16%), graft versatility (11%), and technical ease (11%). Only 20% of surgeons indicated always performing a bony glenoid reconstruction procedure in the noncontact athlete with less than 20% glenoid bone loss. However, that percentage rose to 62% when considering a contact athlete with the same amount of bone loss.Although open Latarjet continues to be the most popular glenoid bony primary reconstruction procedure in shoulder instability, nearly 30% of shoulder surgeons in the western United States have adopted FBB techniques as their preferred treatment modality--with DTA being the most frequently used graft. High-quality comparative clinical effectiveness research is needed to reduce decisional conflict and refine current evidence-based treatment algorithms.

PremiseEvolution of separate sexes from hermaphroditism often proceeds through gynodioecy, but genetic constraints on this process are poorly understood. Genetic (co-)variances and between-sex genetic correlations were used to predict evolutionary responses of multiple reproductive traits in a sexually dimorphic gynodioecious species, and predictions were compared with observed responses to artificial selection.MethodsSchiedea (Caryophyllaceae) is an endemic Hawaiian lineage with hermaphroditic, gynodioecious, subdioecious, and dioecious species. We measured genetic parameters of Schiedea salicaria and used them to predict evolutionary responses of 18 traits in hermaphrodites and females in response to artificial selection for increased male (stamen) biomass in hermaphrodites or increased female (carpel, capsule) biomass in females. Observed responses over two generations were compared with predictions in replicate lines of treatments and controls.ResultsIn only two generations, both stamen biomass in hermaphrodites and female biomass in females responded markedly to direct selection, supporting a key assumption of models for evolution of dioecy. Other biomass traits, pollen and ovule numbers, and inflorescence characters important in wind pollination evolved indirectly in response to selection on sex allocation. Responses generally followed predictions from multivariate selection models, with some responses unexpectedly large due to increased genetic correlations as selection proceeded.ConclusionsResults illustrate the power of artificial selection and utility of multivariate selection models incorporating sex differences. They further indicate that pollen and ovule numbers and inflorescence architecture could evolve in response to selection on biomass allocation to male versus female function, producing complex changes in plant phenotype as separate sexes evolve.

Individuals with specific language impairment (SLI) struggle with language acquisition despite average non-verbal intelligence and otherwise typical development. One SLI account focuses on grammar acquisition delay. The current study aimed to detect novel rare genetic variants associated with performance on a grammar assessment, the Test of Early Grammatical Impairment (TEGI), in English-speaking children. The TEGI was selected due to its sensitivity and specificity, consistently high heritability estimates, and its absence from all but one molecular genetic study. We performed whole exome sequencing (WES) in eight families with SLI (n = 74 total) and follow-up Sanger sequencing in additional unrelated probands (n = 146). We prioritized rare exonic variants shared by individuals with low TEGI performance (n = 34) from at least two families under two filtering workflows: (1) novel and (2) previously reported candidate genes. Candidate variants were observed on six new genes (PDHA2, PCDHB3, FURIN, NOL6, IQGAP3, and BAHCC1), and two genes previously reported for overall language ability (GLI3 and FLNB). We specifically suggest PCDHB3, a protocadherin gene, and NOL6 are critical for ribosome synthesis, as they are important targets of SLI investigation. The proposed SLI candidate genes associated with TEGI performance emphasize the utility of precise phenotyping and family-based genetic study.

The Indian Ocean Observing System (IndOOS), established in 2006, is a multinational network of sustained oceanic measurements that underpin understanding and forecasting of weather and climate for the Indian Ocean region and beyond. Almost one-third of humanity lives around the Indian Ocean, many in countries dependent on fisheries and rain-fed agriculture that are vulnerable to climate variability and extremes. The Indian Ocean alone has absorbed a quarter of the global oceanic heat uptake over the last two decades and the fate of this heat and its impact on future change is unknown. Climate models project accelerating sea level rise, more frequent extremes in monsoon rainfall, and decreasing oceanic productivity. In view of these new scientific challenges, a 3-yr international review of the IndOOS by more than 60 scientific experts now highlights the need for an enhanced observing network that can better meet societal challenges, and provide more reliable forecasts. Here we present core findings from this review, including the need for 1) chemical, biological, and ecosystem measurements alongside physical parameters; 2) expansion into the western tropics to improve understanding of the monsoon circulation; 3) better-resolved upper ocean processes to improve understanding of air–sea coupling and yield better subseasonal to seasonal predictions; and 4) expansion into key coastal regions and the deep ocean to better constrain the basinwide energy budget. These goals will require new agreements and partnerships with and among Indian Ocean rim countries, creating opportunities for them to enhance their monitoring and forecasting capacity as part of IndOOS-2.

Intestinal passenger T leukocytes are responsible for graft-versus-host disease (GvHD) in intestinal transplantation (ITx). We hypothesized that ex vivo fludarabine treatment of the bowel graft would diminish the risk of GvHD and improve overall survival post-transplant. We performed isolated heterotopic small bowel transplantations from Lewis (LEW) to Brown Norway (BN) rat strains, which generated GvHD signs from the fourth day post-transplant. These symptoms included rash, weight loss, piloerection, and diarrhea. The grafts of one of the experimental groups were immersed and sealed in cold Celsior preservation solution with 1000 µm fludarabine for 1 h, prior to its implantation into recipient animals. No histological signs of intestinal tissue alterations were observed after fludarabine treatment. Fludarabine-treated bowel recipients showed significantly later and milder clinical signs of GvHD and reduced total donor cell chimerism, as determined by flow cytometry using strain-specific anti-HLA antibodies. Additionally, fludarabine treatment prolonged recipients' overall survival (13.5 days ± 0.3 days vs. 9.2 days ± 0.5). We conclude that active modification of the intestinal leukocyte composition is advantageous in our ITx animal model. Immunosuppression with fludarabine during the surgical procedure, which could be translated directly to the clinic, protects bowel recipients from GvHD and improves overall post-transplant survival.

In skeletal muscle fibers, mitochondria are densely packed adjacent to myofibrils because adenosine triphosphate (ATP) is needed to fuel sarcomere shortening. However, despite this close physical and biochemical relationship, the effects of mitochondrial dynamics on skeletal muscle contractility are poorly understood. In this study, we analyzed the effects of Mitochondrial Division Inhibitor 1 (mdivi-1), an inhibitor of mitochondrial fission, on the structure and function of both mitochondria and myofibrils in skeletal muscle tissues engineered on micromolded gelatin hydrogels. Treatment with mdivi-1 did not alter myotube morphology, but did increase the mitochondrial turbidity and oxidative capacity, consistent with reduced mitochondrial fission. Mdivi-1 also significantly increased basal, twitch, and tetanus stresses, as measured using the Muscular Thin Film (MTF) assay. Finally, mdivi-1 increased sarcomere length, potentially due to mdivi-1-induced changes in mitochondrial volume and compression of myofibrils. Together, these results suggest that mdivi-1 increases contractile stress generation, which may be caused by an increase in maximal respiration and/or sarcomere length due to increased volume of individual mitochondria. These data reinforce that mitochondria have both biochemical and biomechanical roles in skeletal muscle and that mitochondrial dynamics can be manipulated to alter muscle contractility.

Mitochondria are the major source of cellular energy (ATP), as well as critical mediators of widespread functions such as cellular redox balance, apoptosis, and metabolic flux. The organelles play an especially important role in the maintenance of cardiac homeostasis; their inability to generate ATP following impairment due to ischemic damage has been directly linked to organ failure. Methods to quantify mitochondrial content are limited to low throughput immunoassays, measurement of mitochondrial DNA, or relative quantification by untargeted mass spectrometry. Here, we present a high throughput, reproducible and quantitative mass spectrometry multiple reaction monitoring based assay of 37 proteins critical to central carbon chain metabolism and overall mitochondrial function termed ‘MitoPlex’. We coupled this protein multiplex with a parallel analysis of the central carbon chain metabolites (219 metabolite assay) extracted in tandem from the same sample, be it cells or tissue. In tests of its biological applicability in cells and tissues, “MitoPlex plus metabolites” indicated profound effects of HMG-CoA Reductase inhibition (e.g., statin treatment) on mitochondria of i) differentiating C2C12 skeletal myoblasts, as well as a clear opposite trend of statins to promote mitochondrial protein expression and metabolism in heart and liver, while suppressing mitochondrial protein and ii) aspects of metabolism in the skeletal muscle obtained from C57Bl6 mice. Our results not only reveal new insights into the metabolic effect of statins in skeletal muscle, but present a new high throughput, reliable MS-based tool to study mitochondrial dynamics in both cell culture and in vivo models.

The genome tridimensional (3D) organization and its role towards the regulation of key cell processes such as transcription is currently a main question in biology. Interphase chromosomes are spatially segregated into “territories,” epigenetically-defined large domains of chromatin that interact to form “compartments” with common transcriptional status, and insulator-flanked domains called “topologically associating domains” (TADs). Moreover, chromatin organizes around nuclear structures such as lamina, speckles, or the nucleolus to acquire a higher-order genome organization. Due to recent technological advances, the different hierarchies are being solved. Particularly, advances in microscopy technologies are shedding light on the genome structure at multiple levels. Intriguingly, more and more reports point to high variability and stochasticity at the single-cell level. However, the functional consequences of such variability in genome conformation are still unsolved. Here, I will discuss the implication of the cell-to-cell heterogeneity at the different scales in the context of newly developed imaging approaches, particularly multiplexed Fluorescence in situ hybridization methods that enabled “chromatin tracing.” Extensions of these methods are now combining spatial information of dozens to thousands of genomic loci with the localization of nuclear features such as the nucleolus, nuclear speckles, or even histone modifications, creating the fast-moving field of “spatial genomics.” As our view of genome organization shifts the focus from ensemble to single-cell, new insights to fundamental questions begin to emerge. Fil: Cardozo Gizzi, Andres Mauricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Grupo Vinculado Centro de Investigación en Medicina Traslacional Severo R. Amuchástegui - Cimetsa | Universidad Nacional de Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Grupo Vinculado Centro de Investigación en Medicina Traslacional Severo R. Amuchástegui - Cimetsa | Instituto de Investigación Médica Mercedes y Martín Ferreyra. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Grupo Vinculado Centro de Investigación en Medicina Traslacional Severo R. Amuchástegui - Cimetsa; Argentina. Instituto Universitario de Ciencias Biomédicas de Córdoba; Argentina

β2 -adrenoceptor agonists improve autophagy and re-establish proteostasis in cardiac cells; therefore, suggesting autophagy as a downstream effector of β2 -adrenoceptor signaling pathway. Here, we used the pharmacological and genetic tools to determine the autophagy effect of sustained β2 -adrenoceptor activation in rodents with neurogenic myopathy, which display impaired skeletal muscle autophagic flux. Sustained β2 -adrenoceptor activation using Formoterol (10 μg kg-1 day-1 ), starting at the onset of neurogenic myopathy, prevents disruption of autophagic flux in skeletal muscle 14 days after sciatic nerve constriction. These changes are followed by reduction of the cytotoxic protein levels and increased skeletal muscle cross-sectional area and contractility properties. Of interest, sustained administration of Formoterol at lower concentration (1 μg kg-1 day-1 ) induces similar improvements in skeletal muscle autophagic flux and contractility properties in neurogenic myopathy, without affecting the cross-sectional area. Sustained pharmacological inhibition of autophagy using Chloroquine (50 mg kg-1 day-1 ) abolishes the beneficial effects of β2 -adrenoceptor activation on the skeletal muscle proteostasis and contractility properties in neurogenic myopathy. Further supporting an autophagy mechanism for β2 -adrenoceptor activation, skeletal muscle-specific deletion of ATG7 blunts the beneficial effects of β2 -adrenoceptor on skeletal muscle proteostasis and contractility properties in neurogenic myopathy in mice. These findings suggest autophagy as a critical downstream effector of β2 -adrenoceptor signaling pathway in skeletal muscle.

In ventricular myocardium, extracellular matrix (ECM) remodeling is a hallmark of physiological and pathological growth, coincident with metabolic rewiring of cardiac myocytes. However, the direct impact of the biochemical and mechanical properties of the ECM on the metabolic function of cardiac myocytes is mostly unknown. Furthermore, understanding the impact of distinct biomaterials on cardiac myocyte metabolism is critical for engineering physiologically-relevant models of healthy and diseased myocardium. For these reasons, we systematically measured morphological and metabolic responses of neonatal rat ventricular myocytes cultured on fibronectin- or gelatin-coated polydimethylsiloxane (PDMS) of three elastic moduli and gelatin hydrogels with four elastic moduli. On all substrates, total protein content, cell morphology, and the ratio of mitochondrial DNA to nuclear DNA were preserved. Cytotoxicity was low on all substrates, although slightly higher on PDMS compared to gelatin hydrogels. We also quantified oxygen consumption rates and extracellular acidification rates using a Seahorse extracellular flux analyzer. Our data indicate that several metrics associated with baseline glycolysis and baseline and maximum mitochondrial function are enhanced when cardiac myocytes are cultured on gelatin hydrogels compared to all PDMS substrates, irrespective of substrate rigidity. These results yield new insights into how mechanical and biochemical cues provided by the ECM impact mitochondrial function in cardiac myocytes. STATEMENT OF SIGNIFICANCE: Cardiac development and disease are associated with remodeling of the extracellular matrix coincident with metabolic rewiring of cardiac myocytes. However, little is known about the direct impact of the biochemical and mechanical properties of the extracellular matrix on the metabolic function of cardiac myocytes. In this study, oxygen consumption rates were measured in neonatal rat ventricular myocytes maintained on several commonly-used biomaterial substrates to reveal new relationships between the extracellular matrix and cardiac myocyte metabolism. Several mitochondrial parameters were enhanced on gelatin hydrogels compared to synthetic PDMS substrates. These data are important for comprehensively understanding matrix-regulation of cardiac myocyte physiology. Additionally, these data should be considered when selecting scaffolds for engineering in vitro cardiac tissue models.

The deleterious effects of statins on skeletal muscle are well known, but the mechanism associated with these effects remains unresolved. Statins are associated with mitochondrial damage, which may contribute to muscle myopathy. Here we demonstrate that simvastatin induces mitophagy in skeletal muscle cells and hypothesized that attenuating this process by silencing the mitophagy adapter p62/sequestosome-1 (SQSTM1) might mitigate myotoxicity. Surprisingly, silencing p62/SQSTM1 in differentiated C2C12 muscle cells exacerbated rather than attenuated myotoxicity. This inhibition of mitophagy in the face of statin challenge correlated with increased release of cytochrome c to the cytosol, activation of caspase-3, and lactate dehydrogenase (LDH) release. Correspondingly, targeted knockdown of Parkin, a canonical E3 ubiquitin ligase important for mitophagy, mirrored the effects of p62/SQSTM1 silencing. To corroborate these findings in vivo, we treated Parkin knockout mice with simvastatin for 2 wk. In line with our findings in vitro, these mitophagy-compromised mice displayed reduced spontaneous activity, loss of grip strength, and increased circulating levels of muscle damage marker LDH. Our findings demonstrate that mitophagy is an important mechanism to resist statin-induced skeletal muscle damage.—Ramesh, M., Campos, J. C., Lee, P., Song, Y., Hernandez, G., Sin, J., Tucker, K. C., Saadaeijahromi, H., Gurney, M., Ferreira, J. C. B., Andres, A. M. Mitophagy protects against statin-mediated skeletal muscle toxicity.

Animal studies have demonstrated beneficial effects of therapeutic hypothermia on myocardial function, yet exact mechanisms remain unclear. Impaired autophagy leads to heart failure and mitophagy is important for mitigating ischemia/reperfusion injury. This study aims to investigate whether the beneficial effects of therapeutic hypothermia are due to preserved autophagy and mitophagy. Under general anesthesia, the left anterior descending coronary artery of 19 female farm pigs was occluded for 90 minutes with consecutive reperfusion. 30 minutes after reperfusion, we performed pericardial irrigation with warm or cold saline for 60 minutes. Myocardial tissue analysis was performed one and four weeks after infarction. Therapeutic hypothermia induced a significant increase in autophagic flux, mitophagy, mitochondrial mass and function in the myocardium after infarction. Cell stress, apoptosis, inflammation as well as fibrosis were reduced, with significant preservation of systolic and diastolic function four weeks post infarction. We found similar biochemical changes in human samples undergoing open chest surgery under hypothermic conditions when compared to the warm. These results suggest that autophagic flux and mitophagy are important mechanisms implicated in cardiomyocyte recovery after myocardial infarction under hypothermic conditions. New therapeutic strategies targeting these pathways directly could lead to improvements in prevention of heart failure.

Language is a uniquely human ability, and failure to attain this ability can have a life-long impact on the affected individuals. This is particularly true for individuals with specific language impairment (SLI), which is defined as an impairment in normal language development in the absence of any other developmental disability. Although SLI displays high heritability, family-based linkage studies have been hampered by an unclear mode of Mendelian segregation, variable disease penetrance, and heterogeneity of diagnostic criteria. We performed genome-wide parametric linkage analysis and homozygosity mapping in 14 consanguineous families from Pakistan segregating SLI. Linkage analysis revealed a multipoint LOD score of 4.18 at chromosome 2q in family PKSLI05 under a recessive mode of inheritance. A second linkage score of 3.85 was observed in family PKSLI12 at a non-overlapping locus on chromosome 2q. Two other suggestive linkage loci were found in family PKSLI05 on 14q and 22q with LOD scores of 2.37 and 2.23, respectively, that were also identified in homozygosity mapping. Reduction to homozygosity was observed on chromosomes 2q, 5p, 8q, 14q, 17q, and 22q. Each homozygosity region occurred in multiple PKSLI families. We report new SLI loci on chromosomes 2 and 8 and confirm suggestive SLI linkage loci on chromosomes 5, 14, 17, and 22 reported previously in the population of Robinson Crusoe Island. These findings indicate that linkage and homozygosity mapping in consanguineous families can improve genetic analyses in SLI and suggest the involvement of additional genes in the causation of this disorder.

Coxsackievirus B is a significant human pathogen and is a leading cause of myocarditis. We and others have observed that certain enteroviruses including coxsackievirus B cause infected cells to shed extracellular vesicles containing infectious virus. Recent reports have shown that vesicle-bound virus can infect more efficiently than free virus. Though microRNAs are differentially regulated in cells following infection, few have been associated with the vesicles shed from infected cells. Here we report exclusive trafficking of specific microRNAs into viral vesicles compared to vesicles from non-infected cells. We found that the most highly-expressed unique microRNA in viral vesicles was miR-590-5p, which facilitates prolonged viral replication by blocking apoptotic factors. Cells over-expressing this miR were significantly more susceptible to infection. This may be a mechanism by which coxsackievirus B boosts subsequent rounds of infection by co-packaging virus and a select set of pro-viral microRNAs in extracellular vesicles.