scholarly journals Characterization of Gelatin Hydrogels Cross-Linked with Microbial Transglutaminase as Engineered Skeletal Muscle Substrates

2021 ◽  
Vol 8 (1) ◽  
pp. 6
Author(s):  
Divya Gupta ◽  
Jeffrey W. Santoso ◽  
Megan L. McCain
Keyword(s):  
Skeletal Muscle ◽  
Elastic Modulus ◽  
Ambient Air ◽  
In Vitro Models ◽  
Microbial Transglutaminase ◽  
Conventional Culture ◽  
Muscle Tissues ◽  
Physical Features ◽  
Phosphate Buffered Saline

Engineered in vitro models of skeletal muscle are essential for efficiently screening drug safety and efficacy. However, conventional culture substrates poorly replicate physical features of native muscle and do not support long-term culture, which limits tissue maturity. Micromolded gelatin hydrogels cross-linked with microbial transglutaminase (gelatin-MTG hydrogels) have previously been shown to induce C21C2 myotube alignment and improve culture longevity. However, several properties of gelatin-MTG hydrogels have not been systematically characterized, such as changes in elastic modulus during incubation in culture-like conditions and their ability to support sarcomere maturation. In this study, various gelatin-MTG hydrogels were fabricated and incubated in ambient or culture-like conditions. Elastic modulus, mass, and transmittance were measured over a one- or two-week period. Compared to hydrogels in phosphate buffered saline (PBS) or ambient air, hydrogels in Dulbecco’s Modified Eagle Medium (DMEM) and 5% CO2 demonstrated the most stable elastic modulus. A subset of gelatin-MTG hydrogels was micromolded and seeded with C2C12 or primary chick myoblasts, which aligned and fused into multinucleated myotubes with relatively mature sarcomeres. These data are important for fabricating gelatin-MTG hydrogels with predictable and stable mechanical properties and highlight their advantages as culture substrates for engineering relatively mature and stable muscle tissues.

scholarly journals Bioengineered human myobundles mimic clinical responses of skeletal muscle to drugs

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Lauran Madden ◽  
Mark Juhas ◽  
William E Kraus ◽  
George A Truskey ◽  
Nenad Bursac
Keyword(s):  
Skeletal Muscle ◽  
Acetylcholine Receptors ◽  
In Vitro Models ◽  
Calcium Handling ◽  
Muscle Tissues ◽  
Clinical Responses ◽  
And Function ◽  
Electrical Stimuli ◽  
Toxic Myopathy

Existing in vitro models of human skeletal muscle cannot recapitulate the organization and function of native muscle, limiting their use in physiological and pharmacological studies. Here, we demonstrate engineering of electrically and chemically responsive, contractile human muscle tissues (‘myobundles’) using primary myogenic cells. These biomimetic constructs exhibit aligned architecture, multinucleated and striated myofibers, and a Pax7+ cell pool. They contract spontaneously and respond to electrical stimuli with twitch and tetanic contractions. Positive correlation between contractile force and GCaMP6-reported calcium responses enables non-invasive tracking of myobundle function and drug response. During culture, myobundles maintain functional acetylcholine receptors and structurally and functionally mature, evidenced by increased myofiber diameter and improved calcium handling and contractile strength. In response to diversely acting drugs, myobundles undergo dose-dependent hypertrophy or toxic myopathy similar to clinical outcomes. Human myobundles provide an enabling platform for predictive drug and toxicology screening and development of novel therapeutics for muscle-related disorders.

scholarly journals 3D in vitro models of skeletal muscle: myopshere, myobundle and bioprinted muscle construct

2021 ◽  
Vol 52 (1) ◽  
Author(s):  
Frederic Dessauge ◽  
Cindy Schleder ◽  
Marie-Hélène Perruchot ◽  
Karl Rouger
Keyword(s):  
Skeletal Muscle ◽  
Spatial Organization ◽  
3D Culture ◽  
Cell Types ◽  
In Vitro Models ◽  
Skeletal Muscle Function ◽  
Muscle Tissues ◽  
Culture Models ◽  
Living Tissues

AbstractTypical two-dimensional (2D) culture models of skeletal muscle-derived cells cannot fully recapitulate the organization and function of living muscle tissues, restricting their usefulness in in-depth physiological studies. The development of functional 3D culture models offers a major opportunity to mimic the living tissues and to model muscle diseases. In this respect, this new type of in vitro model significantly increases our understanding of the involvement of the different cell types present in the formation of skeletal muscle and their interactions, as well as the modalities of response of a pathological muscle to new therapies. This second point could lead to the identification of effective treatments. Here, we report the significant progresses that have been made the last years to engineer muscle tissue-like structures, providing useful tools to investigate the behavior of resident cells. Specifically, we interest in the development of myopshere- and myobundle-based systems as well as the bioprinting constructs. The electrical/mechanical stimulation protocols and the co-culture systems developed to improve tissue maturation process and functionalities are presented. The formation of these biomimetic engineered muscle tissues represents a new platform to study skeletal muscle function and spatial organization in large number of physiological and pathological contexts.

scholarly journals Skeletal muscle-on-a-chip: an in vitro model to evaluate tissue formation and injury

Lab on a Chip ◽  
2017 ◽  
Vol 17 (20) ◽  
pp. 3447-3461 ◽  
Author(s):  
Gaurav Agrawal ◽  
Aereas Aung ◽  
Shyni Varghese
Keyword(s):  
Skeletal Muscle ◽  
Muscle Injury ◽  
In Vitro Model ◽  
Three Dimensional ◽  
Tissue Formation ◽  
Microfluidic Platform ◽  
Muscle Tissues ◽  
Vitro Model

We introduce a microfluidic platform in which we culture three-dimensional skeletal muscle tissues, while evaluating tissue formation and toxin-induced muscle injury.

scholarly journals A Cylindrical Molding Method for the Biofabrication of Plane-Shaped Skeletal Muscle Tissue

Micromachines ◽  
2021 ◽  
Vol 12 (11) ◽  
pp. 1411
Author(s):  
Minghao Nie ◽  
Ai Shima ◽  
Kenta Fukushima ◽  
Yuya Morimoto ◽  
Shoji Takeuchi
Keyword(s):  
Skeletal Muscle ◽  
Muscle Tissue ◽  
Myogenic Differentiation ◽  
Skeletal Muscle Tissue ◽  
Differentiation Markers ◽  
Animal Products ◽  
Molding Method ◽  
Muscle Tissues ◽  
Central Pillar

Muscle tissues can be fabricated in vitro by culturing myoblast-populated hydrogels. To counter the shrinkage of the myoblast-populated hydrogels during culture, a pair of anchors are generally utilized to fix the two ends of the hydrogel. Here, we propose an alternative method to counter the shrinkage of the hydrogel and fabricate plane-shaped skeletal muscle tissues. The method forms myoblast-populated hydrogel in a cylindrical cavity with a central pillar, which can prevent tissue shrinkage along the circumferential direction. By eliminating the usages of the anchor pairs, our proposed method can produce plane-shaped skeletal muscle tissues with uniform width and thickness. In experiments, we demonstrate the fabrication of plane-shaped (length: ca. 10 mm, width: 5~15 mm) skeletal muscle tissue with submillimeter thickness. The tissues have uniform shapes and are populated with differentiated muscle cells stained positive for myogenic differentiation markers (i.e., myosin heavy chains). In addition, we show the assembly of subcentimeter-order tissue blocks by stacking the plane-shaped skeletal muscle tissues. The proposed method can be further optimized and scaled up to produce cultured animal products such as cultured meat.

scholarly journals Electrophysiological analysis of healthy and dystrophic 3D bioengineered skeletal muscle tissues

2021 ◽  
Author(s):  
Christine T. Nguyen ◽  
Majid Ebrahimi ◽  
Penney M. Gilbert ◽  
Bryan Andrew Stewart
Keyword(s):  
Skeletal Muscle ◽  
Muscle Cell ◽  
Muscle Tissue ◽  
Three Dimensional ◽  
Dystrophin Gene ◽  
Muscle System ◽  
Electrophysiological Analysis ◽  
Cytoskeletal Network ◽  
Muscle Tissues

Recently, methods for creating three-dimensional (3D) human skeletal muscle tissues from myogenic cell lines have been reported. Bioengineered muscle tissues are contractile and respond to electrical and chemical stimulation. In this study we provide an electrophysiological analysis of healthy and dystrophic 3D bioengineered skeletal muscle tissues. We focus on Duchenne muscular dystrophy (DMD), a fatal muscle disorder involving the skeletal muscle system. The dystrophin gene, which when mutated causes DMD, encodes for the Dystrophin protein, which anchors the cytoskeletal network inside of a muscle cell to the extracellular matrix outside the cell. Here, we enlist a 3D in vitro model of DMD muscle tissue, to evaluate an understudied aspect of DMD, muscle cell electrical properties uncoupled from presynaptic neural inputs. Our data shows that electrophysiological aspects of DMD are replicated in the 3D bioengineered skeletal muscle tissue model. Furthermore, we test a block co-polymer, poloxamer 188, and demonstrate capacity for improving the membrane potential in DMD muscle. Therefore, this study serves as the baseline for a new in vitro method to examine potential therapies directed at muscular disorders.

scholarly journals RT-QuIC detection of CWD prion seeding activity in white-tailed deer muscle tissues

2021 ◽  
Author(s):  
Manci Li ◽  
Marc D. Schwabenlander ◽  
Gage R. Rowden ◽  
Jeremy M. Schefers ◽  
Michelle Carstensen ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
North America ◽  
Skeletal Muscles ◽  
Economic Losses ◽  
Food Chains ◽  
Sensitive Assay ◽  
Wasting Disease ◽  
Muscle Tissues ◽  
Highly Sensitive

Chronic wasting disease (CWD) is a prion disease circulating in wild and farmed cervid populations throughout North America (United States and Canada), Europe (Finland, Norway, Sweden), and South Korea. CWD is an immediate threat to cervid heritage and the disease is causing substantial economic losses across multiple sectors. In North America, hunting and farming industries focused on the processing and consumption of white-tailed deer (WTD) venison are particularly vulnerable to CWD, as millions of WTD are consumed annually. Real-time quaking-induced conversion (RT-QuIC) is a highly sensitive assay amplifying misfolded CWD prions in vitro and has facilitated CWD prion detection in a variety of tissues and excreta. To date, no study has comprehensively examined CWD prion content across bulk skeletal muscle tissues harvested from individual CWD infected WTD. Here, we use RT-QuIC to quantify prion-seeding activity in a variety of skeletal muscles from both wild and farmed CWD-positive WTD. We successfully detected CWD prions in muscles commonly used for consumption (e.g., backstrap, tenderloin, etc.) as well as within tongue and neck samples of WTD. Our results help to establish the utility of RT-QuIC for monitoring CWD prions in venison and suggest that the method is useful for preventing CWD prions from entering animal and human food chains. Moreover, our work indicates that CWD prions are more widely distributed across skeletal muscles of infected WTD than previously reported.

Lipid peroxidation of skeletal muscle: an in vitro study

1983 ◽  
Vol 3 (7) ◽  
pp. 609-619 ◽  
Author(s):  
M. J. Jackson ◽  
D. A. Jones ◽  
R. H. T. Edwards
Keyword(s):  
Skeletal Muscle ◽  
Lipid Peroxidation ◽  
Free Radical ◽  
Vitamin E ◽  
Hydroxyl Radicals ◽  
Muscle Tissue ◽  
In Vitro Study ◽  
Muscle Tissues ◽  
In Vitro Technique

The process of lipid peroxidation of skeletal muscle has been examined in vitro by monitoring the autoxidation of skeletal-muscle homogenates. Skeletal-muscle tissue has been shown to have considerable capacity for autoxidation and the process has been found to be initiated by a free-radical-mediated mechanism, critically dependent on the presence of free iron in the homogenate. The initiating radicals have not been firmly identified, but the results suggest that neither superoxide or hydroxyl radicals are involved. An in vitro technique for assessment of the antioxidant capacity of muscle tissue has also been developed which is capable of demonstrating differences between muscle tissues with differing vitamin E contents.

scholarly journals Decellularized Skeletal Muscles Support the Generation of In Vitro Neuromuscular Tissue Models

2021 ◽  
Vol 11 (20) ◽  
pp. 9485
Author(s):  
Paolo Raffa ◽  
Maria Easler ◽  
Francesca Cecchinato ◽  
Beatrice Auletta ◽  
Valentina Scattolini ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Three Dimensional ◽  
Culture Conditions ◽  
In Vitro Models ◽  
Easy Access ◽  
3D In Vitro Model ◽  
Multiple Cell ◽  
Vitro Model

Decellularized skeletal muscle (dSkM) constructs have received much attention in recent years due to the versatility of their applications in vitro. In search of adequate in vitro models of the skeletal muscle tissue, the dSkM offers great advantages in terms of the preservation of native-tissue complexity, including three-dimensional organization, the presence of residual signaling molecules within the construct, and their myogenic and neurotrophic abilities. Here, we attempted to develop a 3D model of neuromuscular tissue. To do so, we repopulated rat dSkM with human primary myogenic cells along with murine fibroblasts and we coupled them with organotypic rat spinal cord samples. Such culture conditions not only maintained multiple cell type viability in a long-term experimental setup, but also resulted in functionally active construct capable of contraction. In addition, we have developed a customized culture system which enabled easy access, imaging, and analysis of in vitro engineered co-cultures. This work demonstrates the ability of dSkM to support the development of a contractile 3D in vitro model of neuromuscular tissue fit for long-term experimental evaluations.

In Vitro Model of Human Skeletal Muscle Tissues with Contractility Fabricated by Immortalized Human Myogenic Cells

2020 ◽  
Vol 4 (11) ◽  
pp. 2000121
Author(s):  
Takunori Nagashima ◽  
Stacy Hadiwidjaja ◽  
Saki Ohsumi ◽  
Akari Murata ◽  
Takumi Hisada ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Human Skeletal Muscle ◽  
In Vitro Model ◽  
Myogenic Cells ◽  
Muscle Tissues ◽  
Vitro Model

scholarly journals Study of the Expression and Function of Calcium-Sensing Receptor in Human Skeletal Muscle

2021 ◽  
Vol 22 (14) ◽  
pp. 7282
Author(s):  
Cecilia Romagnoli ◽  
Preeti Sharma ◽  
Roberto Zonefrati ◽  
Gaia Palmini ◽  
Elena Lucattelli ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Satellite Cells ◽  
Human Skeletal Muscle ◽  
Myogenic Differentiation ◽  
Physiological Role ◽  
Calcium Sensing Receptor ◽  
Calcium Sensing ◽  
Muscle Tissues ◽  
And Function

Skeletal muscle has an outstanding capacity for regeneration in response to injuries, but there are disorders in which this process is seriously impaired, such as sarcopenia. Pharmacological treatments to restore muscle trophism are not available, therefore, the identification of suitable therapeutic targets that could be useful for the treatment of skeletal reduced myogenesis is highly desirable. In this in vitro study, we explored the expression and function of the calcium-sensing receptor (CaSR) in human skeletal muscle tissues and their derived satellite cells. The results obtained from analyses with various techniques of gene and protein CaSR expression and of its secondary messengers in response to calcium (Ca2+) and CaSR drugs have demonstrated that this receptor is not present in human skeletal muscle tissues, neither in the established satellite cells, nor during in vitro myogenic differentiation. Taken together, our data suggest that, although CaSR is a very important drug target in physiology and pathology, this receptor probably does not have any physiological role in skeletal muscle in normal conditions.

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