Rianú: Multi-tissue tracking software for increased throughput of engineered cardiac tissue screening.

Background: The field of tissue engineering has provided valuable three-dimensional species-specific models of the human myocardium in the form of human Engineered Cardiac Tissues (hECTs) and similar constructs. However, hECT systems are often bottlenecked by a lack of openly available software that can collect data from multiple tissues at a time, even in multi-tissue bioreactors, which limits throughput in phenotypic and therapeutic screening applications. Methods: We developed Rianú, an open-source web application capable of simultaneously tracking multiple hECTs on flexible end-posts. This software is operating system agnostic and deployable on a remote server, accessible via a web browser with no local hardware or software requirements. The software incorporates object-tracking capabilities for multiple objects simultaneously, an algorithm for twitch tracing analysis and contractile force calculation, and a data compilation system for comparative analysis within and amongst groups. Validation tests were performed using in-silico and in-vitro experiments for comparison with established methods and interventions. Results: Rianú was able to detect the displacement of the flexible end-posts with a sub-pixel sensitivity of 0.555 px/post (minimum increment in post displacement) and a lower limit of 1.665 px/post (minimum post displacement). Compared to our established reference for contractility assessment, Rianú had a high correlation for all parameters analyzed (ranging from R2=0.7514 to R2=0.9695), demonstrating its high accuracy and reliability. Conclusions: Rianú provides simultaneous tracking of multiple hECTs, expediting the recording and analysis processes, and simplifies time-based intervention studies. It also allows data collection from different formats and has scale-up capabilities proportional to the number of tissues per field of view. These capabilities will enhance throughput of hECTs and similar assays for in-vitro analysis in disease modeling and drug screening applications.

Nutritional screening in a cancer prehabilitation programme: A cohort study.

BACKGROUND: Cancer patients are often malnourished pre-operatively. The present study aimed to establish whether current screening was appropriate for use in prehabilitation and investigate any association between nutritional risk, functionality and quality of life (QoL). METHODS: This cohort study used routinely collected data from September 2020 to August 2021 from patients in a Prehab4cancer programme. Included patients were aged ≥ 18 years, had colorectal, lung or oesophago-gastric cancer and were scheduled for surgery. Nutritional assessment included Patient-Generated Subjective Global Assessment (PG-SGA) Short-Form and QoL with a sit-to-stand test. Association between nutritional risk and outcomes was analysed using adjusted logistic regression. RESULTS: From 928 patients referred to Prehab4Cancer service over 12 months, data on nutritional risk were collected from 526 patients. Pre-operatively, 233 out of 526 (44%) patients were at nutritional risk (score ≥ 2). During prehabilitation, 31% of patients improved their PG-SGA and 74% of patients maintained or improved their weight. Odds ratios (OR) with confidence intervals (CI) showed that patients with better QoL using EuroQol-5 Dimensions (OR = 0.05, 95% CI = 0.01, 0.45, p = 0.01), EuroQol Visual Analogue Scale (OR = 0.96, 95% CI = 0.93, 1.00, p = 0.04) or sit-to-stand (OR = 0.96, 95% 0.93, 1.00, p = 0.04) were less likely to be nutritional at risk. CONCLUSIONS: Almost half of patients in Prehab4Cancer programme assessed using PG-SGA were at risk of malnutrition. However, almost half of the sample did not have their risk assessed. Patients at risk of malnutrition were more likely to have a poorer QoL and sit-to-stand test than those who were not at risk.

A lithium-isotope perspective on the evolution of carbon and silicon cycles.

The evolution of the global carbon and silicon cycles is thought to have contributed to the long-term stability of Earth's climate1-3. Many questions remain, however, regarding the feedback mechanisms at play, and there are limited quantitative constraints on the sources and sinks of these elements in Earth's surface environments4-12. Here we argue that the lithium-isotope record can be used to track the processes controlling the long-term carbon and silicon cycles. By analysing more than 600 shallow-water marine carbonate samples from more than 100 stratigraphic units, we construct a new carbonate-based lithium-isotope record spanning the past 3 billion years. The data suggest an increase in the carbonate lithium-isotope values over time, which we propose was driven by long-term changes in the lithium-isotopic conditions of sea water, rather than by changes in the sedimentary alterations of older samples. Using a mass-balance modelling approach, we propose that the observed trend in lithium-isotope values reflects a transition from Precambrian carbon and silicon cycles to those characteristic of the modern. We speculate that this transition was linked to a gradual shift to a biologically controlled marine silicon cycle and the evolutionary radiation of land plants13,14.

Health, lifestyle and sociodemographic characteristics are associated with Brazilian dietary patterns: Brazilian National Health Survey.

This study aimed to identify Brazilian dietary patterns and their associations with health, lifestyle and sociodemographic characteristics. Data from the Brazilian National Health Survey conducted in 2013 were used. A questionnaire was applied containing 22 items related to dietary consumption. Dietary patterns were determined through factor analysis (FA). Poisson regression models, with robust variance, were used to identify associations between dietary patterns and independents variables. Statistical significance was defined as p-value<0.05. Data were analysed for 60,202 adults (estimated population size: 146,308,458). FA identified three dietary patterns: healthy, protein, and western. The younger age group (18-24 years) had a lower adherence to the healthy pattern (PR:0.53; 95%CI:0.49-0.58) and greater adherence to the protein (PR:1.52; 95%CI:1.42-1.62) and western (PR:1.80; 95%CI:1.68-1.93) patterns compared to the elderly (≥60 years). Women had a greater association with the healthy pattern (PR:1.32; 95%CI:1.28-1.38) and lower association with the protein pattern (PR:0.80; 95%CI:0.77-0.82) compared to men. Illiterate participants showed lower adherence to the healthy (PR:0.58; 95%CI:0.53-0.63) and western (PR:0.54; 95%CI:0.48-0.62) patterns compared to those with higher educational levels. Smokers had lower adherence to the healthy (PR:0.76; 95%CI:0.71-0.81) and higher adherence to the protein (PR:1.14; 95%CI:1.11-1.19) patterns compared to non-smokers. Participants with poor/very poor self-rated health status had a lower adherence to the healthy (PR:0.79; 95%CI:0.73-0.86) and western (PR:0.81; 95%CI:0.73-0.89) patterns compared to those in a very good/good self-rated health status. Multimorbidity was positively associated with the healthy pattern (PR:1.18; 95%CI:1.11-1.26) and inversely associated with the protein pattern (PR:0.88; 95%CI:0.80-0.96) compared to participants without comorbidities. We suggest that strategies to promote healthy eating should consider health, lifestyle and sociodemographic characteristics in the Brazilian population.

A modern scleractinian coral with a two-component calcite-aragonite skeleton.

One of the most conserved traits in the evolution of biomineralizing organisms is the taxon-specific selection of skeletal minerals. All modern scleractinian corals are thought to produce skeletons exclusively of the calcium-carbonate polymorph aragonite. Despite strong fluctuations in ocean chemistry (notably the Mg/Ca ratio), this feature is believed to be conserved throughout the coral fossil record, spanning more than 240 million years. Only one example, the Cretaceous scleractinian coral Coelosmilia (ca. 70 to 65 Ma), is thought to have produced a calcitic skeleton. Here, we report that the modern asymbiotic scleractinian coral Paraconotrochus antarcticus living in the Southern Ocean forms a two-component carbonate skeleton, with an inner structure made of high-Mg calcite and an outer structure composed of aragonite. P. antarcticus and Cretaceous Coelosmilia skeletons share a unique microstructure indicating a close phylogenetic relationship, consistent with the early divergence of P. antarcticus within the Vacatina (i.e., Robusta) clade, estimated to have occurred in the Mesozoic (ca. 116 Mya). Scleractinian corals thus join the group of marine organisms capable of forming bimineralic structures, which requires a highly controlled biomineralization mechanism; this capability dates back at least 100 My. Due to its relatively prolonged isolation, the Southern Ocean stands out as a repository for extant marine organisms with ancient traits.

Adult human cardiac stem cell supplementation effectively increases contractile function and maturation in human engineered cardiac tissues.

BACKGROUND: Delivery of stem cells to the failing heart is a promising therapeutic strategy. However, the improvement in cardiac function in animal studies has not fully translated to humans. To help bridge the gap between species, we investigated the effects of adult human cardiac stem cells (hCSCs) on contractile function of human engineered cardiac tissues (hECTs) as a species-specific model of the human myocardium. METHODS: Human induced pluripotent stem cell-derived cardiomyoctes (hCMs) were mixed with Collagen/Matrigel to fabricate control hECTs, with an experimental group of hCSC-supplemented hECT fabricated using a 9:1 ratio of hCM to hCSC. Functional testing was performed starting on culture day 6, under spontaneous conditions and also during electrical pacing from 0.25 to 1.0 Hz, measurements repeated at days 8 and 10. hECTs were then frozen and processed for gene analysis using a Nanostring assay with a cardiac targeted custom panel. RESULTS: The hCSC-supplemented hECTs displayed a twofold higher developed force vs. hCM-only controls by day 6, with approximately threefold higher developed stress and maximum rates of contraction and relaxation during pacing at 0.75 Hz. The spontaneous beat rate characteristics were similar between groups, and hCSC supplementation did not adversely impact beat rate variability. The increased contractility persisted through days 8 and 10, albeit with some decrease in the magnitude of the difference of the force by day 10, but with developed stress still significantly higher in hCSC-supplemented hECT; these findings were confirmed with multiple hCSC and hCM cell lines. The force-frequency relationship, while negative for both, control (- 0.687 Hz- 1; p = 0.013 vs. zero) and hCSC-supplemented (- 0.233 Hz- 1;p = 0.067 vs. zero) hECTs, showed a significant rectification in the regression slope in hCSC-supplemented hECT (p = 0.011 vs. control). Targeted gene exploration (59 genes) identified a total of 14 differentially expressed genes, with increases in the ratios of MYH7/MHY6, MYL2/MYL7, and TNNI3/TNNI1 in hCSC-supplemented hECT versus controls. CONCLUSIONS: For the first time, hCSC supplementation was shown to significantly improve human cardiac tissue contractility in vitro, without evidence of proarrhythmic effects, and was associated with increased expression of markers of cardiac maturation. These findings provide new insights about adult cardiac stem cells as contributors to functional improvement of human myocardium.

Epidemiology, risk factors, and opportunities for prevention of cardiovascular disease in individuals of South Asian ethnicity living in Europe.

South Asian (SA) individuals represent a large, growing population in a number of European countries. These individuals, particularly first-generation SA immigrants, are at higher risk of developing type 2 diabetes, atherogenic dyslipidaemia, and coronary heart disease than most other racial/ethnic groups living in Europe. SAs also have an increased risk of stroke compared to European-born individuals. Despite a large body of conclusive evidence, SA-specific cardiovascular health promotion and preventive interventions are currently scarce in most European countries, as well as at the European Union level. In this narrative review, we aim to increase awareness among clinicians and healthcare authorities of the public health importance of cardiovascular disease among SAs living in Europe, as well as the need for tailored interventions targeting this group - particularly, in countries where SA immigration is a recent phenomenon. To this purpose, we review key studies on the epidemiology and risk factors of cardiovascular disease in SAs living in the United Kingdom, Italy, Spain, Denmark, Norway, Sweden, and other European countries. Building on these, we discuss potential opportunities for multi-level, targeted, tailored cardiovascular prevention strategies. Because lifestyle interventions often face important cultural barriers in SAs, particularly for first-generation immigrants; we also discuss features that may help maximise the effectiveness of those interventions. Finally, we evaluate knowledge gaps, currently available risk stratification tools such as QRISK-3, and future directions in this important field.

Cardiac Tissue Engineering Models of Inherited and Acquired Cardiomyopathies.

The lack of biomimetic in vitro models of the human heart has posed a critical barrier to progress in the field of modeling cardiac disease. Human engineered cardiac tissues (hECTs)-autonomous, beating structures that recapitulate key aspects of native cardiac muscle physiology-offer an attractive alternative to traditional in vitro models. Here we describe the use of hECTs to advance our understanding and modeling of cardiac diseases in order to test therapeutic interventions, with a focus on contractile dysfunction in the setting of inherited and acquired forms of cardiomyopathies. Four major procedures are discussed in this chapter: (1) preparation of hECTs from human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) on single-tissue and multitissue bioreactors; (2) data acquisition of hECT contractile function on both of these platforms; (3) hECT modeling of hereditary phospholamban-R14 deletion-dilated cardiomyopathy; and (4) cryo-injury and doxorubicin-induced hECT models of acquired cardiomyopathy.

Exosomal microRNA-21-5p Mediates Mesenchymal Stem Cell Paracrine Effects on Human Cardiac Tissue Contractility.

RATIONALE: The promising clinical benefits of delivering human mesenchymal stem cells (hMSCs) for treating heart disease warrant a better understanding of underlying mechanisms of action. hMSC exosomes increase myocardial contractility; however, the exosomal cargo responsible for these effects remains unresolved. OBJECTIVE: This study aims to identify lead cardioactive hMSC exosomal microRNAs to provide a mechanistic basis for optimizing future stem cell-based cardiotherapies. METHODS AND RESULTS: Integrating systems biology and human engineered cardiac tissue (hECT) technologies, partial least squares regression analysis of exosomal microRNA profiling data predicted microRNA-21-5p (miR-21-5p) levels positively correlate with contractile force and calcium handling gene expression responses in hECTs treated with conditioned media from multiple cell types. Furthermore, miR-21-5p levels were significantly elevated in hECTs treated with the exosome-enriched fraction of the hMSC secretome (hMSC-exo) versus untreated controls. This motivated experimentally testing the human-specific role of miR-21-5p in hMSC-exo-mediated increases of cardiac tissue contractility. Treating hECTs with miR-21-5p alone was sufficient to recapitulate effects observed with hMSC-exo on hECT developed force and expression of associated calcium handling genes (eg, SERCA2a and L-type calcium channel). Conversely, knockdown of miR-21-5p in hMSCs significantly diminished exosomal procontractile and associated calcium handling gene expression effects on hECTs. Western blots supported miR-21-5p effects on calcium handling gene expression at the protein level, corresponding to significantly increased calcium transient amplitude and decreased decay time constant in comparison to miR-scramble control. Mechanistically, cotreating with miR-21-5p and LY294002, a PI3K inhibitor, suppressed these effects. Finally, mathematical simulations predicted the translational capacity for miR-21-5p treatment to restore calcium handling in mature ischemic adult human cardiomyocytes. CONCLUSIONS: miR-21-5p plays a key role in hMSC-exo-mediated effects on cardiac contractility and calcium handling, likely via PI3K signaling. These findings may open new avenues of research to harness the role of miR-21-5p in optimizing future stem cell-based cardiotherapies.

Assessing equine prospective memory in a Y-maze apparatus.

Eight horses were tested in a Y-maze to see if they would approach the side on which they had seen and heard food being delivered into a food trough. The horses were tested in five experimental contexts: immediate release (IR), delayed release 3s (3DR), 6s (6DR), 9s (9DR) and 12s (12DR) after food delivery. Individual subjects performed five IR trials, followed by five each of 3DR, 6DR, 9DR and 12DR trials. A re-rest was performed 1 week later. Data were analysed by log-linear analysis of frequencies. The results showed that the horses were capable of achieving the correct choice during IR trials, were successful during the DR trials and improved marginally during the re-test. The findings suggest that horses could have a trainable form of prospective memory and benefit from incremental memory-training exercises. However, there is a risk of overestimating equine cognitive ability and unnecessary reinforcement delays are likely to lead to welfare and training setbacks within equitation.

Weighted boots influence performance in show-jumping horses.

This study investigated the effects of weighted boots on horses (n=6) jumping a 1.25 m oxer fence. The horses had similar training experience and were assigned to two groups of three subjects (groups G1 and G2). All horses performed 10 jumping efforts: G1 horses made attempts 1-5 without boots and 6-10 with boots; G2 made attempts 1-5 with boots and 6-10 without boots. Data were available via sagittal plane S-VHS recordings and t test analyses focussed on limb-placement dimensions. There were no differences among performances of the horses in the horizontal plane, but there were significant differences in the vertical plane. All horses achieved significantly greater hindlimb elevation with the weighted boots (1.60 m) compared with no boots (1.46 m; P<0.05). Although not measured directly, the significantly greater elevation during the jump stride flight phase appears to be a consequence of increased kinetic energy associated with the horses' hindlimbs.

The relationship between visual memory and rider expertise in a show-jumping context.

Individuals develop visual skills whilst participating in sport. The aim of this study was to assess the effect of reported riding expertise on the recall of visual information relating to show-jumps. Relevant (F1) and irrelevant (F2) points of focus were identified in 22 photographs of show-jumps. Participants were students (n=40) with varying levels of horse-riding ability. After viewing each photograph for 4s, the task was to identify F1s or F2s from four alternatives viewed for 10s. F1s were recalled significantly more than F2s (P<0.001). Riding expertise did not affect overall recall but only intermediate/advanced riders recalled F1s significantly better than F2s (P<0.05 and P<0.001, respectively). Recall of F1s, but not of F2s, was significantly correlated with riding expertise (P<0.05). It was concluded that the training of riders in visual attention techniques might improve ridden performance and could reduce the number of accidents associated with lack of experience and/or rider error.

Boots on horses: limb protection or hyperflexion training aids in the showjumping horse.

Showjumping riders regularly employ various schooling strategies to control the horse's jump stride kinematics (JSK). Strategies include plyometric training regimes with fences of different heights and widths set at specific distances. Gymnastic grids teach the horse to jump cleanly. Rapping, once used almost routinely, is no longer in vogue. However, the use of performance enhancing (PE) boots on the distal hind limbs to alter equine JSK has become popular. There are two broad categories of PE boots: weighted and pressure. Some riders use so-called weighted boots on the horses' hind limbs during training and in competition to improve the jump stride. The application of so-called pressure boots may be little more than an adaptation of this technique. It appears that the PE boots induce hyperflexion of the hind limbs and incline the horse to jump fences cleanly. In the absence of scientific appraisal, it is unclear if such boots are acceptable and innovative training aids within equitation.

Innovative use of an automated horse walker when breaking in young horses.

There is an inherent element of risk associated with "backing" and riding the previously unbroken horse. If training proceeds too quickly, conflict behaviors may result from the simultaneous application of too many cues. Automated horse walkers (AHW) facilitate the exercising of several horses concurrently at walk or trot for warm-up, cool-down, fitness programs, and rehabilitation purposes. The objective of this study was to investigate if backing the horse within the AHW was an appropriate training method. Ten horses (3-year-olds) took part in this study. They began training within the AHW with a simple bridle and protective boots. A handler subsequently long-reined the horses within the AHW when they wore rollers, side reins, and a saddle. When considered appropriate, the handler went from jumping beside the horse to lying over the saddle to sitting astride the horse within the AHW. The horses habituated to this innovative approach quickly without evidence of conflict behavior. The handler rode the horses from the AHW after approximately 4 riding episodes of this innovative training system.

Producer or purchaser: different expectations may lead to equine wastage and welfare concerns.

Horses are individual, each having differential characteristics such as height, color, breeding, conformation, and temperament. These bio-characteristics often influence potential purchasers when buying horses. This study sought to investigate if producers and potential purchasers placed similar emphasis on equine bio-characteristics. Sport-horse stakeholders--n = 1377 (792 producers and 585 potential purchasers)--rated various equine bio-characteristics on a Likert psychometric response scale during a questionnaire-based survey. The study analyzed responses, using the Wilcoxan test for statistical significance. The findings indicated consensus between producers and potential purchasers for equine soundness, conformation, and movement. Producers attached significantly greater importance to gender, color, pedigree details, and performance records of the horse and the horse's siblings. In contrast, potential purchasers rated equine temperament and presence (aesthetic appeal) as significantly more important attributes. Shortcomings in suitability for purpose of the horse (such as temperament) could lead to unnecessary wastage and welfare concerns. Producers need to understand consumer expectations/demands to maximize profitability and to avoid wastage and the production of unsuitable horses.

Facial hair whorls (trichoglyphs) and the incidence of motor laterality in the horse.

Several species demonstrate obvious motor laterality (sidedness, handedness) in their motor function. Motor laterality in the horse affects locomotion and subsequently equine performance during training and may have inherent safety implications for equitation. Some of the most commonly used identification features in the horse are hair whorls (trichoglyphs), since their specific location and character vary to some degree in every horse. We investigated the relationship between the hair flow of single facial hair whorls and the incidence of lateralised motor bias in 219 horses when under saddle in ridden work. The horses exhibited significant differences in motor preferences with 104 left-lateralised (LL) horses, 95 right-lateralised (RL) horses compared to only 20 well-balanced (WB) horses (chi(2)=36.9, d.f.=2, P<0.01). There was also a significant difference in the frequency distribution of single facial hair whorl patterns in the horses consisting of 114 horses with counter-clockwise (CC) whorls, 82 horses with clockwise (C) whorls and 23 horses, which had radial (R) whorls (chi(2)=38.87, d.f.=2, P<0.01). Overall there was a statistically significant association between motor behaviour and facial hair whorl patterns in the horses (chi(2)=69.4, d.f.=4, P>0.001). The RL horses had significantly more C facial hair whorls and the LL horses had significantly more CC facial hair whorls than would be expected purely by chance alone (P<0.05). The findings may provide trainers with a useful tool when attempting to identify simple, non-invasive and reliable predictors of motor laterality in the horse. Furthermore, given that efficient targeted training of performance horses during ridden work may produce WB equine athletes, the findings could assist trainers when designing individual-specific training programmes for young horses.

Equine learning behaviour.

Scientists and equestrians continually seek to achieve a clearer understanding of equine learning behaviour and its implications for training. Behavioural and learning processes in the horse are likely to influence not only equine athletic success but also the usefulness of the horse as a domesticated species. However given the status and commercial importance of the animal, equine learning behaviour has received only limited investigation. Indeed most experimental studies on equine cognitive function to date have addressed behaviour, learning and conceptualization processes at a moderately basic cognitive level compared to studies in other species. It is however, likely that the horses with the greatest ability to learn and form/understand concepts are those, which are better equipped to succeed in terms of the human-horse relationship and the contemporary training environment. Within equitation generally, interpretation of the behavioural processes and training of the desired responses in the horse are normally attempted using negative reinforcement strategies. On the other hand, experimental designs to actually induce and/or measure equine learning rely almost exclusively on primary positive reinforcement regimes. Employing two such different approaches may complicate interpretation and lead to difficulties in identifying problematic or undesirable behaviours in the horse. The visual system provides the horse with direct access to immediate environmental stimuli that affect behaviour but vision in the horse is of yet not fully investigated or understood. Further investigations of the equine visual system will benefit our understanding of equine perception, cognitive function and the subsequent link with learning and training. More detailed comparative investigations of feral or free-ranging and domestic horses may provide useful evidence of attention, stress and motivational issues affecting behavioural and learning processes in the horse. The challenge for scientists is, as always, to design and commission experiments that will investigate and provide insight into these processes in a manner that withstands scientific scrutiny.