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Overcoming the challenge of creating thick, tissue-resembling muscle constructs is paramount in the field of cultivated meat production. This study investigates the remarkable potential of random cellulose acetate nanofibers (CAN) as a transformative scaffold for muscle tissue engineering (MTE), specifically in the context of cultivated meat applications. Through a comparative analysis between random and aligned CAN, utilizing C2C12 and H9c2 myoblasts, we unveil the unparalleled capabilities of random CAN in facilitating muscle differentiation, independent of differentiation media, by exploiting the YAP/TAZ-related mechanotransduction pathway. In addition, we have successfully developed a novel process for stacking cell-loaded CAN sheets, enabling the production of a three-dimensional meat product. C2C12 and H9c2 loaded CAN sheets were stacked (up to four layers) to form a ~300–400 μm thick tissue 2 cm in length, organized in a mesh of uniaxial aligned cells. To further demonstrate the effectiveness of this methodology for cultivated meat purposes, we have generated thick and viable constructs using chicken muscle satellite cells (cSCs) and random CAN. This groundbreaking discovery offers a cost-effective and biomimetic solution for cultivating and differentiating muscle cells, forging a crucial link between tissue engineering and the pursuit of sustainable and affordable cultivated meat production.

Simple Summary: Canine hip dysplasia is a painful condition common in large dog breeds, leading to joint instability and osteoarthritis. Accurate diagnosis is crucial for guiding breeding decisions to help reduce its prevalence. However, evaluating hip health status can be challenging, as minor positioning changes during X-rays can distort images and hinder proper hip joint assessment. In this study, we developed an artificial intelligence tool for the automatic evaluation of pelvic alignment in X-rays. By detecting subtle asymmetries in bone structure, the tool determines whether a dog's hips are properly aligned. Our findings showed that the artificial intelligence tool performed as accurately as an expert human examiner in detecting misalignment. This automated approach could improve the reliability of canine hip dysplasia screenings, saving veterinarians time and reducing misdiagnosis risks due to human error. Ultimately, this technology has the potential to enhance medical care for dogs and support breeders in making more informed choices, contributing to canine health improvements and reducing canine hip dysplasia incidence in future generations. Canine hip dysplasia (CHD) screening relies on accurate positioning in the ventrodorsal hip extended (VDHE) view, as even mild pelvic rotation can affect CHD scoring and impact breeding decisions. This study aimed to assess the association between pelvic rotation and asymmetry in obturator foramina areas (AOFAs) and to develop a computer vision model for automated AOFA measurement. In the first part, 203 radiographs were analyzed to examine the relationship between pelvic rotation, assessed through asymmetry in iliac wing and obturator foramina widths (AOFWs), and AOFAs. A significant association was found between pelvic rotation and AOFA, with AOFW showing a stronger correlation (R2 = 0.92, p < 0.01). AOFW rotation values were categorized into minimal (n = 71), moderate (n = 41), marked (n = 37), and extreme (n = 54) groups, corresponding to mean AOFA ± standard deviation values of 33.28 ± 27.25, 54.73 ± 27.98, 85.85 ± 41.31, and 160.68 ± 64.20 mm2, respectively. ANOVA and post hoc testing confirmed significant differences in AOFA across these groups (p < 0.01). In part two, the dataset was expanded to 312 images to develop the automated AOFA model, with 80% allocated for training, 10% for validation, and 10% for testing. On the 32 test images, the model achieved high segmentation accuracy (Dice score = 0.96; Intersection over Union = 0.93), closely aligning with examiner measurements. Paired t-tests indicated no significant differences between the examiner and model's outputs (p > 0.05), though the Bland–Altman analysis identified occasional discrepancies. The model demonstrated excellent reliability (ICC = 0.99) with a standard error of 17.18 mm2. A threshold of 50.46 mm2 enabled effective differentiation between acceptable and excessive pelvic rotation. With additional training data, further improvements in precision are expected, enhancing the model's clinical utility. [ABSTRACT FROM AUTHOR]

Abstract Background The repulsive guidance molecule a (RGMa) is a GPI-anchor axon guidance molecule first found to play important roles during neuronal development. RGMa expression patterns and signaling pathways via Neogenin and/or as BMP coreceptors indicated that this axon guidance molecule could also be working in other processes and diseases, including during myogenesis. Previous works from our research group have consistently shown that RGMa is expressed in skeletal muscle cells and that its overexpression induces both nuclei accretion and hypertrophy in muscle cell lineages. However, the cellular components and molecular mechanisms induced by RGMa during the differentiation of skeletal muscle cells are poorly understood. In this work, the global transcription expression profile of RGMa-treated C2C12 myoblasts during the differentiation stage, obtained by RNA-seq, were reported. Results RGMa treatment could modulate the expression pattern of 2,195 transcripts in C2C12 skeletal muscle, with 943 upregulated and 1,252 downregulated. Among them, RGMa interfered with the expression of several RNA types, including categories related to the regulation of RNA splicing and degradation. The data also suggested that nuclei accretion induced by RGMa could be due to their capacity to induce the expression of transcripts related to ‘adherens junsctions’ and ‘extracellular-cell adhesion’, while RGMa effects on muscle hypertrophy might be due to (i) the activation of the mTOR-Akt independent axis and (ii) the regulation of the expression of transcripts related to atrophy. Finally, RGMa induced the expression of transcripts that encode skeletal muscle structural proteins, especially from sarcolemma and also those associated with striated muscle cell differentiation. Conclusions These results provide comprehensive knowledge of skeletal muscle transcript changes and pathways in response to RGMa.

Electrospinning emerged as a promising technique to produce scaffolds for cultivated meat in function of its simplicity, versatility, cost-effectiveness, and scalability. Cellulose acetate (CA) is a biocompatible and low-cost material that support cell adhesion and proliferation. Here we investigated CA nanofibers, associated or not with a bioactive annatto extract (CA@A), a food-dye, as potential scaffolds for cultivated meat and muscle tissue engineering. The obtained CA nanofibers were evaluated concerning its physicochemical, morphological, mechanical and biological traits. UV-vis spectroscopy and contact angle measurements confirmed the annatto extract incorporation into the CA nanofibers and the surface wettability of both scaffolds, respectively. SEM images revealed that the scaffolds are porous, containing fibers with no specific alignment. Compared with the pure CA nanofibers, CA@A nanofibers showed increased fiber diameter (420 ± 212 nm vs. 284 ± 130 nm). Mechanical properties revealed that the annatto extract induces a reduction of the stiffness of the scaffold. Molecular analyses revealed that while CA scaffold favored C2C12 myoblast differentiation, the annatto-loaded CA scaffold favored a proliferative state of these cells. These results suggest that the combination of cellulose acetate fibers loaded with annatto extract may be an interesting economical alternative for support long-term muscle cells culture with potential application as scaffold for cultivated meat and muscle tissue engineering.

Animals with muscle hypertrophy phenotype are targeted by the broiler industry to increase the meat production and the quality of the final product. Studies characterizing the molecular machinery involved with these processes, such as quantitative trait loci studies, have been carried out identifying several candidate genes related to this trait; however, validation studies of these candidate genes in cell culture is scarce. The aim of this study was to evaluate SAP30 as a candidate gene for muscle development and to validate its function in cell culture in vitro. The SAP30 gene was downregulated in C2C12 muscle cell culture using siRNA technology to evaluate its impact on morphometric traits and gene expression by RNA-seq analysis. Modulation of SAP30 expression increased C2C12 myotube area, indicating a role in muscle hypertrophy. RNA-seq analysis identified several upregulated genes annotated in muscle development in treated cells (SAP30-knockdown), corroborating the role of SAP30 gene in muscle development regulation. Here, we provide experimental evidence of the involvement of SAP30 gene as a regulator of muscle cell hypertrophy.

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Background and Aim: Papillomaviruses (PVs) infections have been documented in numerous animal species across different regions worldwide. They often exert significant impacts on animal health and livestock production. Scientists have studied natural products for over half a century due to their diverse chemical composition, acknowledging their value in fighting cancer. Acorns (Quercus ilex) are believed to have several unexplored pharmacological properties. This study aimed to evaluate the in vivo safety and cancer chemopreventive activity of an infusion extract of Q. ilex in a transgenic mouse model of human PV (HPV)-16, which developed squamous cell carcinomas through a multistep process driven by HPV16 oncogenes. Materials and Methods: Q. ilex extract was prepared by heating in water at 90°C and then characterized by mass spectrometry. Phenolic compounds from this extract were administered in drinking water to female mice in three different concentrations (0.03, 0.06, and 0.09 g/mL) over a period of 28 consecutive days. Six groups (n = 6) were formed for this study: group 1 (G1, wildtype [WT], water), group 2 (G2, HPV, water), group 3 (G3, WT, 0.09 g/mL), group 4 (G4, HPV, 0.03 g/mL), group 5 (G5, HPV, 0.06 g/ mL), and group 6 (G6, HPV, 0.09 g/mL). Throughout the experiment, humane endpoints, body weight, food intake, and water consumption were recorded weekly. Following the experimental period, all mice were sacrificed, and blood, internal organs, and skin samples were collected. Blood was used to measure glucose and microhematocrit and later biochemical parameters, such as creatinine, urea, albumin, alanine aminotransferase, and total proteins. Histological analysis was performed on skin and organ samples. Results: The administration of Q. ilex extract resulted in a statistically significant increase in relative organ weight among HPV transgenic animals, indicating adaptive biological response to the tested concentrations. Moreover, a reduction in characteristic skin lesions was observed in animals treated with the 0.06 and 0.09 g/mL extract. Conclusion: These results provide a favorable chemopreventive profile for Q. ilex extract at concentrations of 0.06 and 0.09 g/mL. This study highlights the potential of Q. ilex extract as a safe and effective therapeutic strategy against HPV16-associated lesions in transgenic mouse models. The limitation of our study was the durability of transgenic animals. As a more sensitive species, we must always be careful with the durability of the test. We intend to study concentrations of 0.06 and 0.09 g/mL for longer to further investigate their possible effects. [ABSTRACT FROM AUTHOR]