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Meat produced in vitro has been proposed as a humane, safe and environmentally beneficial alternative to slaughtered animal flesh as a source of nutritional muscle tissue. The basic methodology of an in vitro meat production system (IMPS) involves culturing muscle tissue in a liquid medium on a large scale. Each component of the system offers an array of options which are described taking into account recent advances in relevant research. A major advantage of an IMPS is that the conditions are controlled and manipulatable. Limitations discussed include meeting nutritional requirements and large scale operation. The direction of further research and prospects regarding the future of in vitro meat production will be speculated.Industrial relevanceThe development of an alternative meat production system is driven by the growing demand for meat and the shrinking resources available to produce it by current methods. Implementation of an in vitro meat production system (IMPS) to complement existing meat production practices creates the opportunity for meat products of different characteristics to be put onto the market. In vitro produced meat products resembling the processed and comminuted meat products of today will be sooner to develop than those resembling traditional cuts of meat. While widening the scope of the meat industry in practices and products, the IMPS will reduce the need for agricultural resources to produce meat.
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Possibilities for an in vitro meat production system
I. Datar, M. Betti
Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, Alberta, Canada T6G 2P5
abstractarticle info
Article history:
Received 28 June 2009
Accepted 11 October 2009
Editor Proof Receive Date 26 October 2009
In vitro meat
Myocyte culturing
Meat substitutes
Meat produced in vitro has been proposed as a humane, safe and environmentally benecial alternative to
slaughtered animal esh as a source of nutritional muscle tissue. The basic methodology of an in vitro meat
production system (IMPS) involves culturing muscle tissue in a liquid medium on a large scale. Each
component of the system offers an array of options which are described taking into account recent advances
in relevant research. A major advantage of an IMPS is that the conditions are controlled and manipulatable.
Limitations discussed include meeting nutritional requirements and large scale operation. The direction of
further research and prospects regarding the future of in vitro meat production will be speculated.
Industrial relevance: The development of an alternative meat production system is driven by the growing
demand for meat and the shrinking resources available to produce it by current methods. Implementation of
an in vitro meat production system (IMPS) to complement existing meat production practices creates the
opportunity for meat products of different characteristics to be put onto the market. In vitro produced meat
products resembling the processed and comminuted meat products of today will be sooner to develop than
those resembling traditional cuts of meat. While widening the scope of the meat industry in practices and
products, the IMPS will reduce the need for agricultural resources to produce meat.
© 2009 Elsevier Ltd. All rights reserved.
1. Introduction ............................................................... 14
2. Basic methodology ............................................................ 14
3. Cells................................................................... 15
3.1. Native muscle tissue formation................................................... 15
3.2. Possible cell types ......................................................... 15
3.2.1. Proposed cell types .................................................... 15
3.2.2. Adult stem cells ..................................................... 15
3.2.3. Dedifferentiated cells ................................................... 16
3.3. Replicative ability ......................................................... 16
4. Scaffold ................................................................. 16
4.1. Shape............................................................... 16
4.2. Texture and microstructure .................................................... 16
4.3. Composition ........................................................... 17
4.4. Scaffold removal. ......................................................... 17
5. Culture conditions ............................................................ 17
5.1. Growth media........................................................... 17
5.2. Regulatory factors ......................................................... 18
5.3. Contraction ............................................................ 18
6. Bioreactor ................................................................ 18
6.1. Medium perfusion ......................................................... 18
6.2. Oxygen carriers .......................................................... 18
7. Control and manipulation ........................................................ 18
Innovative Food Science and Emerging Technologies 11 (2010) 1322
Corresponding author. Tel.: +1 780 248 1598.
E-mail address: (M. Betti).
1466-8564/$ see front matter © 2009 Elsevier Ltd. All rights reserved.
Contents lists available at ScienceDirect
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8. Limitations to the methodology ......................................................19
8.1. Nutritional value ..........................................................19
8.2. Metabolism ............................................................19
8.3. Large-scale operation........................................................19
9. Conclusion and future prospects ......................................................20
Acknowledgements.............................................................. 20
References ..................................................................20
1. Introduction
In light of the sizable negative effects of livestock production,
establishment of an in vitro meat production system (IMPS) is
becoming increasingly justiable. Current meat production methods
are a major source of pollution and a signicant consumer of fossil
fuels, land and water resources. World meat production at present is
contributing between 15% and 24% of total current greenhouse gas
emissions; a great proportion of this percentage is due to deforesta-
tion to create grazing land (Steinfeld et al., 2006). The production of
beef requires 15 500 m
/ton of water, while chicken requires
3918 m
/ton (Hoekstra & Chapagain, 2007) and with a growing
population and great proportion of which facing starvation, it no
longer makes sense to contribute staple crops toward inefcient meat
production, where 1 kg poultry, pork and beef requires 2 kg, 4 kg and
7 kg of grain, respectively (Rosegrant, Leach, & Gerpacio, 1999).
Satisfying the demand for meat in the future will be a challenge if we
intend on maximizing the use of agricultural resources and reducing
greenhouse gas production, as Fiala (2008) calculates the amount of
total meat consumed worldwide in 2030 to be 72% higher than that
consumed in 2000 following current consumption patterns.
It is suspected that myocyte culturing would have a reduced water,
energy and land requirement because a) solely muscle tissue is
cultivated, bypassing the development of by-products and non-
skeletal muscle tissues; b) for the same mass of meat, tissue
cultivation is anticipated to be faster than growth to a slaughter-
ready age and c) in vitro meat production systems are capable of
increasing in volume vertically, making deforestation to create
pasture unnecessary. The controlled conditions would theoretically
eliminate product losses from infected and diseased animals. As a
method with little to no waste products or by-products and a
minimized land and resource requirement, myocyte culturing could
possibly alleviate the environmental burden exhibited by today's
meat harvesting techniques.
To provide meat globally, modern industrial meat production has
become a complex landscape of trade where feed production, animal
husbandry, processing and consumption may all take place in
different countries (Burke, Oleson, McCullough, & Gaskell, 2009).
The comparatively minimal land requirement of an in vitro meat
production system allows meat production and processing to take
place domestically in countries which would normally rely on
imported meats. By bringing the stages of the meat production
process closer together spatially and temporally, meat supply can be
better determined by demand.
Humans are taxonomically omnivorous and meat provides several
essential nutrients unavailable in plant sources. Meat is specically
valuable as a source of omega-3 fatty acids, vitamin B
, and highly
bioavailable iron (Bender, 1992). The health benets of meat are
countered by its association with cancer and cardiovascular disease
(Demeyer, Honikel, & De Smet, 2008), though these are the result of
overconsumption and high saturated fat content, not the muscle
While it is possible that a cultured meat product could consist of a
variety of animal cell types, meat is being dened here as primarily
skeletal muscle tissue. An in vitro meat production system involves
culturing muscle-like tissue in a liquid medium, therefore bypassing
animal husbandry and slaughter. The controlled conditions of the IMPS
are impossible to achieve by traditional livestockmethods and therefore
allow for a safer, healthier product. Myocyte culturing prevents the
spread of animal-borne disease which may or may not affect meat
products. Moreover, by reducing the amount of close quarter human
animal interaction, the incidence of epidemic zoonoses developing will
decline. The employment of aseptic technique throughout the culturing
process ensures that the meat product is free from contamination.
Controlled conditions also offer the capacity for manipulation to create
meat products with different nutritional, textural and taste proles. This
can be accomplished by co-culturing with different cell types, medium
supplementation or genetic engineering.
Considering the benets of an IMPS, is not surprising that a
number of parties have proposed (and patented) the methodology for
actualizing this idea (Vein, 2004; Van Eelen, van Kooten, & Westerhof,
1999; Edelman, McFarland, Mironov, & Matheny, 2005). This paper
introduces the techniques so far proposed. As of yet none of these
processes, though detailed, have been tested. This is partially because
livestock animal cell lines have not been well-established in vitro
(Talbot & Blomberg, 2008) and because growing muscle cells ex vivo
on a large scale it is certainly a vast and unexplored undertaking. The
technical demands of large scale production are unseen in the world
of medical research, where most efforts in growing tissue ex vivo have
been directed. The nutritional composition of ex vivo engineered
muscle tissue has not yet been paid much attention. As a result,
establishment of an IMPS is faced with many unique challenges so far
unexplored in the eld of tissue engineering.
Breakthroughs in relevant research since the publishing of these
patents and proposals have widely altered the scope of designing an
IMPS. Because the proposed systems at present approximate to a
schematic with modiable elements, the discrete elements will be
attended to individually, taking into account recent relevant scientic
developments. This will be followed by a discussion of some
drawbacks and limitations of the methodology, concluded with a
conjecture of the future of in vitro meat production, and suggestions
for further research.
2. Basic methodology
The methods so far proposed have a common set of elements. Each
proposal employs the growth of myoblasts or myosatellite cells on a
scaffold in a suspension culture medium within a bioreactor. Neither
of these cell types need to be stimulated to muscle cell lineages but
both have limited regenerative capacity. Van Eelen et al. (1999)
proposed growth of myocytes on a collagen meshwork, while
Edelman et al. (2005) suggest collagen beads as the scaffold. The
scaffold-based method can only produce a thin myocyte layer of 100
200 µm thick on the scaffold in static culture due to diffusional
limitations (Carrier et al., 1999). As a result, the products of these
methods lack the structure of native muscle tissue, and therefore
could only be put towards processed meat products (Edelman et al.,
2005). To create a three-dimensional meat product, Van Eelen et al.
(1999) suggest layering several conuent sheets of myocyte culture.
14 I. Datar, M. Betti / Innovative Food Science and Emerging Technologies 11 (2010) 1322
An alternative method of creating a three-dimensional productis the
expansion in volume of an explant of muscle tissue of animal origin.
Benjaminson, Gilchriest, and Lorenz (2002) were able to expand the
surface area of an explant of sh by growth in a medium containing
crude cell mixture; the resulting product was prepared and well-rated
by a food panel. This method also faces diffusional limitations and is
unlikely to translate well into a large scale operation.
3. Cells
It is worth mentioning that non-skeletal muscle cell types may have
relevance as a cultured meat product, but in vitro meat is being dened
here as a culture of primarily skeletal muscle tissue therefore cell types
destined to create this desired tissue type are discussed below.
3.1. Native muscle tissue formation
During embryological development, committed muscle tissue
formation as seen in Fig. 1a) begins with mononucleated myoblasts
of limited proliferation capacity (Benjaminson et al., 2002). Myoblasts
fuse into a multinucleated myotube, which matures into a non-
proliferative myober (Campion, 1984). Postnatally, increases in
number of myobers and number of nuclei per myober are kept
minimal, except in instances requiring repair or regeneration. In these
cases, myosatellite cells are responsible for generating new myobers
or contributing additional myonuclei to existing ones (Fig. 1b; Le
Grand & Rudnicki, 2007). Located between the basal lamina and
sarcolemma of an associated myober, mononucleated myosatellite
cells are normally in a quiescent, non-dividing state (Hill, Wernig, &
Goldspink, 2003). When activated in vivo by weight-bearing stress or
injury, myosatellite cells asymmetrically divide into self-renewing
myoblasts and committed myobers (Benjaminson et al., 2002;Le
Grand & Rudnicki, 2007).
Myosatellite cells are a very small proportion (15%) of the cell
population of muscle tissue, and this percentage is dependent on
muscle ber composition and organism age (Allen, Temm-Grove,
Sheehan, & Rice, 1997). As an organism ages, the regenerative
potential of its myosatellite cell population also decreases rapidly.
As a result, the cells that offer optimal regenerative potential and
myober morphology in vitro (longer myobers and greater myober
density) must be harvested from neonates (Delo et al., 2008).
3.2. Possible cell types
3.2.1. Proposed cell types
At the time when Van Eelen et al. (1999) and Edelman et al. (2005)
made their IMPS proposals, the two viable cell options were
embryonic stem (ES) cells or myosatellite cells. In theory, after the
ES cell line is established, its unlimited regenerative potential
eliminates the need to harvest more cells from embryos, however,
the slow accumulation of genetic mutations over time may determine
a maximum proliferation period for a useful long-term ES culture
(Amit et al., 2000). While ES cells are an attractive option for their
unlimited proliferative capacity, these cells must be specically
stimulated to differentiate into myoblasts and may inaccurately
recapitulate myogenesis (Bach, Stem-Straeter, Beier, Bannasch, &
Stark, 2003). In addition to this, there are so far no provenbovine,
porcine, caprine nor ovine ES cell lines that have been established to
the degree of a biological reagent like that of human, monkey or
mouse ES cells (Talbot & Blomberg, 2008). Efforts invested into
establishing ungulate stem-cell lines over the past two decades have
been generally unsuccessful with difculties arising in the recogni-
tion, isolation and differentiation of these cells (Keefer, Pant,
Blomberg, & Talbot, 2007).
Although myosatellite cells have the disadvantage of being a rare
muscle tissue cell type with limited regenerative potential, Bach et al.
(2003) indicate that they are the preferred source of primary myoblasts
because they recapitulate myogenesis more closely than immortal
myogenic cell lines. Myosatellite cells have been isolated and charac-
terized from the skeletal muscle tissue of cattle (Dodson, Martin,
Brannon, Mathison, & McFarland, 1987), chicken (Yablonka-Reuveni,
Quinn, & Nameroff, 1987), sh (Powell, Dodson, & Cloud, 1989), lambs
(Dodson, McFarland, Martin, & Brannon, 1986), pigs (Blanton, Grand,
McFarland, Robinson, & Bidwell, 1999; Wilschut, Jaksani, Van Den
Dolder, Haagsman, & Roelen, 2008), and turkeys (McFarland, Doumit, &
Minshall, 1988). Each animal species has its own benets and
limitations as a cell source, and myosatellite cells isolated from different
muscles have different capabilities to proliferate, differentiate, or be
regulated by growth modiers (Burton, Vierck, Krabbenhoft, Byrne, &
Dodson, 2000). Wilschut et al. (2008) have shown that porcine muscle
progenitor cells have the potential for multilineage differentiation into
adipogenic, osteogenic and chondrogenic lineages, which may play a
role in the development of co-cultures.
Recent advances in tissue engineering and cell biology offer some
alternate cell options which may have practical applications with
multilineage potential allowing for co-culture development and with
suitability for large-scale operations.
3.2.2. Adult stem cells
Myosatellite cells are one example of an adult stem-cell type with
multilineage potential (Asakura, Komaki, & Rudnicki, 2001). Adult
Fig. 1. a) Prenatal myogenesis: Stem cells give rise to proliferative muscle precursor cells,
(myoblasts), which lose proliferative capability upon fusinginto multinucleate myotubes.
Myotubesundergo morphological changesto become mature non-proliferativemyobers.
b) Postnatal/posthatchmyogenesis is for repair andregeneration of existingmuscle tissue.
Myober-associated myosatellite cells respond to weight-bearing stress or injury by
asymmetrically dividing into a self-renewing daughter cell and a nonproliferative
myober-committed cell.Committed cells can fusewith other committedcells to produce
new myobers or add nuclei to e xisting myobers.
15I. Datar, M. Betti / Innovative Food Science and Emerging Technologies 11 (2010) 1322
stem cells have been isolated from several different adult tissues
(Wagers & Weissman, 2004), but another cell type relevant to in vitro
meat production are adipose tissue-derived adult stem cells (ADSCs),
a rare population of multipotent cells found in adipose tissue (Gimble,
Katz, & Bunnell, 2007). Kim et al. (2006) have noted that these highly
expandable cells can be obtained relatively non-invasively from
subcutaneous fat and subsequently transdifferentiated to myogenic,
osteogenic, chondrogenic or adipogenic cell lineages.
The greatest concern and matter of debate regarding adult stem cells
is their proneness to malignant transformation in long-term culture
(Lazennec & Jorgensen, 2008). Rubio et al. (2005) have found that
adipose tissue-derived adult stem cells immortalize at high frequency
and undergo spontaneous transformation in long-term (45months)
culturing, while evidence of adult stem cells remaining untransformed
have also been reported (Bernardo et al., 2007). In an in vitro meat
production system, re-harvesting of adultstem cells to minimizethe risk
of spontaneous transformation may be necessary. With this in mind,
harvesting ADSCs from subcutaneous fat is far less invasive than
collection of myosatellite cells from muscle tissue and samples can be
taken from certain organisms without causing substantial harm.
3.2.3. Dedifferentiated cells
Dedifferentiation is the reversion of a terminally differentiated cell
into a multipotent cell type. It has been reported by Matsumoto et al.
(2007) that mature adipocytes can be dedifferentiated in vitro into a
multipotent preadipocyte cell line termed dedifferentiated fat (DFAT)
cells. Following this, DFAT cells are capable of being transdifferen-
tiated into skeletal myocytes (Kazama, Fujie, Endo, & Kano, 2008). By
oating a piece of glass on top of a suspension of mature adipocytes,
buoyant mononucleated adipocytes attach to the upper surface and
either a) release their fat droplet and assume the broblast-like shape
of a DFAT cell or b) asymmetrically divide to produce one broblast-
shaped DFAT daughter cell (Matsumoto et al., 2007). Termed the
ceiling culturemethod, the process certainly seems achievable on an
industrial scale. Because mature adipocytes are the most common cell
type in adipose tissue, tissue samples will have high cell yields
compared to other cell options.
Cell dedifferentiation appears to be an attractive alternative to the
use of stem cells but Rizzino (2007) has put forth the argument that
many of the claims of transdifferentiation, dedifferention and multi-
potency of once terminally differentiated cells may be due to
abnormal processes resulting in cellular look-alikes.
3.3. Replicative ability
An IMPS requires many cell divisions to mass culture muscle
tissue, but most cells have a nite number of divisions in culture
before natural cell death; this number is termed the Hayick limit.
Edelman et al. (2005) put forth the three possible means for
overcoming this limitation in an IMPS: a) regularly replenishing the
culture, b) using an immortal cell line or c) immortalizing a cell line.
Most cell types will require the rst method though embryonic stem
cells fall into the second category. Use of the third strategy is
controversial because immortalization of cell lines requires genetic
manipulation. It is also important to draw parallels between the
genetic modications that would facilitate large-scale production and
the mutations exhibited by cancerous cells.
Increasing the regenerative potential of cells without immortaliz-
ing them is one additional possibility. The Hayick limit is determined
by telomere length. Telomeres are the guanine-rich repeats found at
the ends of linear chromosomes. Due to the linearity of the
chromosome and the mechanism by which replication occurs,
telomeres are shortened with every round of DNA replication (and
cell division). Telomere length therefore corresponds to the number
of divisions a cell type is capable of. This explains why a neonatal
source is best for harvesting myosatellite cells; very early in
development the cells still have long telomeres. Telomerase is a
ribozyme capable of lengthening telomeres, naturally found in
immortal cell lines. Many different cell types have been immortalized
with ectopic telomerase expression and have showed no signs of the
growth deregulation associated with cancer cells (Harley, 2002). Un-
fortunately, this helpful genetic alteration may be subject to consumer
4. Scaffold
Scaffolding mechanisms differ in shape, composition and char-
acteristics to optimize muscle cell and tissue morphology. Myoblasts
are anchorage-dependent cells, capable of spontaneous contraction.
An ideal scaffold would have a large surface area for growth and
attachment, be exible to allow for contraction, maximize medium
diffusion and be easily dissociated from the meat culture. A scaffold
that closely mimics the in vivo situation is best; myotubes
differentiate optimally on scaffold with a tissue-like stiffness (Engler
et al., 2004). The best materials would be natural and edible, though
inedible scaffold materials cannot be disregarded. Development of
new biomaterials offer additional characteristics that may be
benecial for in vitro muscle tissue growth, such as fullling the
requirement of contraction for proliferation and differentiation (De
Deyne, 2000).
4.1. Shape
As mentioned before, Edelman et al. (2005) proposed beads made
of edible collagen as a substrate while Van Eelen et al. (1999)
proposed a collagen meshwork described as a collagen spongeof
bovine origin. The tribeculate structure of the sponge allows for
increased surface area and diffusion, but may impede harvesting of
the tissue culture. Other possible scaffold forms include large elastic
sheets or an array of long, thin laments. Conformation choice is
based primarily on maximizing surface area, which increases diffusion
and the amount of anchorage-dependent tissue that can be grown.
4.2. Texture and microstructure
Texturized surfaces can attend to specic requirements of muscle
cells, one of which is myober alignment. To mimic native muscle
architecture, Lam, Sim, Zhu, and Takayama (2006) cultured myoblasts
on a substrate with a wavy micropatterned surface and found that the
wave pattern aligned differentiated muscle cells while promoting
myoblast fusion to produce aligned myotubes. Myober organization is
important for the functional characteristics of muscle and the textural
characteristics of meat. Micropatterned surfaces could allow muscle
tissue cultured by the scaffold-based technique to assume a two-
dimensional structure more similar to that of meat of native origin.
Electrospinning is the process of using electrical charge to extract
very ne bers from liquids. Riboldi, Sampaolesi, Neuenschwander,
Cossu, and Mantero (2005) have suggested electrospun microbrous
meshwork membranes as a scaffold for skeletal myocytes, as the
membranes offer high surface area to volume ratio and some elastic
properties. Electrospinning creates very smooth bers, which may not
translate well into a good adhesive surface; Riboldi et al. (2005) have
shown thatcoating electrospun polymer berswith extracellularmatrix
proteins, such as collagen or bronectin, promotes attachment by
ligandreceptor binding interactions. Electrospinning shows promise
for scaffold formation because the process is simple, controllable,
reproducible and capable of producing polymerswith special properties
by co-spinning (Riboldi et al., 2005). Microbre organization can also
affect myober morphology. Electrobers can be spun with nanometer
to micrometer width; this corresponds to the wave periodicity (6 µm
was found to be optimal by Lam et al., 2006) found to be benecial for
myocyte alignment on a micropatterned surface. Knowing the effects of
16 I. Datar, M. Betti / Innovative Food Science and Emerging Technologies 11 (2010) 1322
micropatterning, introducing electrospun bers aligned at the correct
periodicity could theoretically align myobers.
Beyond facilitating attachment and alignment, a scaffold capable of
increasing nutrient diffusion and medium circulation would prove
invaluable to the process. Noting that tissue thickness cannot exceed
100200 µm without experiencing cell death, Borenstein et al. (2002)
microfabricated a synthetic vascular network. Edelman et al. (2005)
acknowledge that a cast of an existing vascularization network, such
as that in native muscle tissue, can be used to create a collagen
network mimicking native vessel architecture. Taking this a step
further, Borenstein et al. (2002) created articial networks of
channels in sheets of biodegradable, biocompatible polymer, then
seeded the network with endothelial cells. Following dissolution of
the polymer mold, successful proliferation could theoretically leave
behind a network of endothelial tissue onto which one could grow
myocytes. A synthetic vascular system would then require a
circulation pumping system and a soluble oxygen carrier in the
medium to be fully functional. Unfortunately, at this moment creation
of these articial vascular networks does not translate well into mass
production due to the microfabrication processes required.
4.3. Composition
Several different polymers could sufce as the scaffold material for
an in vitro meat production system. Examples of edible, naturally
derived polymers are collagen, cellulose, alginate or chitosan. These
polymers would be safe to leave in the meat product and could add a
textural quality. Edelman et al. (2005) have suggested that porous
beads made of these polymers capable of undergoing surface area
changes under different pH and temperature conditions could fulll
the contraction requirement of myoblast cells.
By contrast, inedible polymers confer some interesting qualities
that can aid muscle tissue formation. The aforementioned micro-
patterned surfaces which can aid in myober alignment are inedible,
as are the thermoresponsive coatings described below. Jun, Jeong, and
Shin (2009) have found that growing myoblasts on electrically
conductive bers induces their differentiation, forming more myo-
tubes of greater length without the addition of electrical stimulation.
Use of inedible scaffolding systems necessitates simple and non-
destructive techniques for removal of the culture from the scaffold.
4.4. Scaffold removal
A technical challenge of growing by the scaffold-based technique is
removal of the scaffolding system. Removal of conuent cultured cell
sheets is conventionally done enzymatically or mechanically, but these
two methods damage the cells and the extracellular matrix they may be
producing (Canavan, Cheng, Graham, Ratner, & Castner, 2005). Da Silva,
Mano, and Reis (2007) note that thermoresponsive coatings which
change from hydrophobic to hydrophilic at lowered temperatures can
release cultured cells and extracellular matrix as an intact sheet upon
cooling. Termed thermal liftoff,this method results in undamaged
sheets that maintain the ability to adhere if transferred onto another
substrate (da Silva et al., 2007). This opens the possibility of stacking
sheets to create a three-dimensional product.
Lam, Huang, Birla, and Takayama (2009) have presented a method
for detaching culture as a conuent sheet from a non-adhesive
micropatterned surface using the biodegradation of selective attach-
ment protein laminin.
It should be noted that culturing on a scaffold may not result in a
two-dimensional conuent sheetof culture. After scaffold removal,
the contractile forces exerted by the cytoskeleton of the myocyte are
no longer balanced by adhesion to a surface, causing the cell lawn to
contract and aggregate, forming a detached multicellular spheroid
(da Silva et al., 2007). To remove the culture as a sheet, a hydrophilic
membrane or gel placed on the apical surface of the culture before
detachment can provide physical support. Lam et al. (2009) found
that use of a brin hydrogel was ideal for skeletal muscle tissue
because cells can migrate, proliferate and produce their own
extracellular matrix within it while degrading excess brin. These
two-dimensional sheets could be stacked as suggested by Van Eelen
et al. (1999), to create a three-dimensional product.
5. Culture conditions
Perhaps the most difcult task in designing an in vitro meat
production system is determining the best culture medium formula-
tion. The medium should support and promote growth while being
made of affordable, edible components available in large quantities.
Medium composition will be a substantial cost determinant if not
solely for the fact that large quantities will be required.
5.1. Growth media
Myoblast culturing usually takes place in animal sera, a costly
media that does not lend itself well to consumer acceptance or large-
scale use. Animal sera are from adult, newborn or fetal source, with
fetal bovine serum being the standard supplement for cell culture
media (Coecke et al., 2005). Because of its in vivo source, it can have a
large number of constituents in highly variable composition and
potentially introduce pathogenic agents (Shah, 1999). The harvest of
fetal bovine serum also raises ethical concern. Commercially available
serum replacements and serum-free culture media offer some more
realistic options for culturing mammalian cells in vitro. Serum-free
media reduce operating costs and process variability while lessening
the potential source of infectious agents (Froud, 1999).
Serum-free media have been developed to support in vitro
myosatellite cell cultures from the turkey (McFarland, Pesall, Norberg,
& Dvoracek, 1991), sheep (Dodson & Mathison, 1988)andpig(Doumit,
Cook, & Merkel, 1993). Variations among different serum-free media
outline the fact that satellite cells from different species have different
requirements and respond differentially to certain additives (Dodson,
McFarland, Grant, Doumit, & Velleman, 1996). Ultroser G is an example
of a commercially available serum substitute specially designed to
replace fetal bovine serum for growth of anchorage-dependent cells in
vitro. It has a consistent composition containing growth factors, binding
proteins, adhesin factors, vitamins, hormones and mineral trace
elements, all necessary for eukaryotic cell growth (Duque et al., 2003).
It has one-fth the protein content of serum (Pope, Harrison, Wilson,
Breen, & Cummins, 1987), yet growth of mammalian skeletal myocytes
with serum substitute Ultroser G showed more maturation over the
same period of time, longer lasting viability and longer myotubes with
more localized nuclei (Benders, van Kuppevelt, Oosterhof, & Veerkamp,
1991). While Ultroser G has many benecial effects on the growth and
maturation of muscle tissue in vitro, its costliness may make this an
unlikely candidate for scale up. In addition, while the Ultroser G and
other commercially available serum replacements may have advanta-
geous effects on the growth of tissues, their exact formulations are
protected by commercial copyright and an evaluation of their suitability
on a large scale can only be determined by costand effects. In most cases,
serum-free media are supplemented with puried proteins of animal
origin (Merten, 1999).
Benjaminson et al. (2002), in their investigations with sh
explants, found that mushroom extracts were comparable to serum
as a growth medium in promoting explant surface area expansion. A
cheap, rich serum is necessary for an in vitro meat production system;
it is possible that amino acid-rich mushroom extracts could be applied
here. The development of an appropriate serum-free media com-
pletely free of any animal-derived components appears ideal, but the
potential for allergens from plant-derived proteins are a risk factor to
be mindful of.
17I. Datar, M. Betti / Innovative Food Science and Emerging Technologies 11 (2010) 1322
5.2. Regulatory factors
Creating an optimal cocktail of hormones, and growth factors is a
complex undertaking, one which requires an extensive amount of
investigation. Extrinsic regulatory factor selection must be specicto
the chosen cell type and species, as myosatellite cells for instance, of
different species respond differentially to the same regulatory factors
(Burton et al., 2000). It is also likely that the formulation may be
required to change over the course of the culturing process. For
instance, the proliferation period may require one certain combina-
tion of growth factors and hormones while the differentiation and
maturation period may require a different set.
A multitude of regulatory factors have been identied as being
capable of inducing myosatellite cell proliferation (Cheng et al., 2006),
and the regulation of meat animal-derived myosatellite cells by
hormones, polypeptide growth factors and extracellular matrix
proteins has also been investigated (Dodson et al., 1996; Doumit
et al., 1993). Puried growth factors or hormones may be supple-
mented into the media from an external source such as transgenic
bacterial, plant or animal species which produce recombinant
proteins (Houdebine, 2009). Alternatively, a sort of synthetic
paracrine signalling system can be arranged so that co-cultured cell
types (a feeder layer) can secrete growth factors which can promote
cell growth and proliferation in neighbouring cells. Co-cultured
hepatocytes for instance could provide insulin-like growth factors
which stimulate myoblast proliferation and differentiation (Cen,
Zhang, Huang, Yang, & Xie, 2008) as well as myosatellite cell pro-
liferation in several meat-animal species in vitro (Dodson et al., 1996)
Human growth hormone, routine produced from a transgenic bacte-
rial source can be supplemented into the medium to stimulate pro-
duction of insulin-like growth factors by hepatocytes.
Alternatively, autocrine growth factor signalling can play a role, as
certain muscle cell-secreted growth factors such as insulin-like growth
factor II stimulate myocyte maturation (Wilson, Hsieh, & Rotwein,
2003). Similarly, growth factor production can occur in genetically
engineered muscle cells.
5.3. Contraction
Regular contraction is a necessity for skeletal muscle. It promotes
differentiation and healthy myober morphology while preventing
atrophy. Muscle in vivo is innervated, allowing for regular, controlled
contraction. An in vitro system would necessarily culture denervated
muscle tissue, so contraction must be stimulated by alternate means.
Cha, Park, Noh, and Suh (2006) have found that administration of
cyclic mechanical strain to a highly porous scaffold sheet promotes
differentiation and alignment of smooth muscle cells. Edelman et al.
(2005) and Van Eelen et al. (1999) proposed mechanical stretching of
scaffolds and expandable scaffold beads to fulll the requirement of
providing contraction. This in mind, de Deyne (2000) noted that
external mechanical contraction is less effective than electrical
stimulation in promoting muscle development. Electrical stimulation,
feasible on a large scale, induces contraction internally as opposed to
passively and aids in differentiation and sarcomere formation. As
mentioned before, even growth on electrically conductive bers
without application of electrical stimulation sufced in reaping the
benets of induced contraction (Jun et al., 2009).
6. Bioreactor
Achieving adequate perfusion of the cultured tissue is key to
producing large culture quantities. To construct viable tissue greater
than 100200 µm in thickness it is necessary to have adequate oxygen
perfusion during cell seeding and cultivation on the scaffold (Radisic
et al., 2008). Adequate oxygen perfusion is mediated by a) bioreactors
which increase mass transport between culture medium and cells,
along with b) the use of oxygen carriers to mimic hemoglobin-
provided oxygen supply.
6.1. Medium perfusion
The design of a bioreactor is inten ded to promote the growth of tissue
cultures which accurately resemble native tissue architecture while
providing an environment which allows for increased culture volumes.
The cylindrical wall of rotating wall vessel bioreactors rotates at a
speed that balances centrifugal force, drag force and gravitational
force, leaving the three-dimensional culture submerged in the
medium in a perpetual free fall state (Carrier et al., 1999). This creates
a laminar ow of the medium which improves diffusion with high
mass transfer rates at minimal levels of shear stress, producing three-
dimensional tissues with structures very similar to those in vivo
(Martin, Wendt, & Herberer, 2004).
Direct perfusion bioreactors ow medium through a porous scaffold
with gas exchange taking place in an external uid loop (Carrier et al.,
2002). This type of bioreactor appears more appropriate for scaffold-
based myocyte cultivation. They offer high mass transfer but also
signicant shear stress, so determining an appropriate ow rate is
essential (Martin et al., 2004). Direct perfusion bioreactors are also used
for high-density, uniform myocyte cell seeding (Radisic et al., 2003).
Another method of increasing medium perfusion is by vasculariz-
ing the tissue being grown. Levenberg et al. (2005) had induced
endothelial vessel networks in skeletal muscle tissue constructs by
using a co-culture of myoblasts, embryonic broblasts and endothelial
cells co-seeded onto a highly porous biodegradable scaffold. This is
certainly a method that can increase the diffusional limitation of
tissue thickness beyond 100-200 µm in vitro.
6.2. Oxygen carriers
Cell viability and density positively correlate with the oxygen
gradient in statically grown tissue cultures (Radisic et al., 2008).
Oxygen carriers can be supplemented to the medium to maintain high
oxygen concentrations in solution, similar to that of blood. There are
two distinct varieties of oxygen carrier: those which are modied
versions of hemoglobin and those which are chemically inert,
articially produced peruorochemicals (PFCs) (Lowe, 2006a).
Though many chemically modied hemoglobins have been devel-
oped, their bovine or human source makes them an unlikely candidate
for scale up. Alternatively, human hemoglobin has been produced by
genetically modied plants (Dieryck et al., 1997) and microorganisms
(Zuckerman, Doyle, Gorczynski, & Rosenthal, 1998). Efforts to produce
heme proteins and blood substitute components using the microorgan-
isms Escherichia coli,Pichia pastis,andAspergillus niger are underway as
these organisms are already used to commercially produce human
pharmaceuticals and food additives (Lowe, 2006b).
Peruorochemicals (PFCs) dissolve large volumes of oxygen and
therefore can perform the same function as hemoglobin. To be miscible
in aqueous conditions PFCs must be emulsied andemulsications have
been available for use in vitro and in vivo (Lowe, 2006a). Shine et al.
(2005) note that medical PFCs are extremely potent greenhouse gases
on a per molecule basis and though they presently make a trivial
contribution to climate change, increasing their use requires careful
consideration. Development of an articial blood is an active area of
research and many applicable options are likely to arise with time.
7. Control and manipulation
An IMPS offers a level of control unattainable by traditional livestock
methods of producing meat. Myocyte culturing prevents the uncon-
trollable, unpredictable complications present in livestock production
includingthe spread of disease among animals and the development of
zoonoses. The diseases of concern in industrial agriculture which have
18 I. Datar, M. Betti / Innovative Food Science and Emerging Technologies 11 (2010) 1322
become increasingly more difcult to contain with expanding interna-
tional tradeavian u, swine u, foot-and-mouth disease, and bovine
spongiform encephalopathywill not threaten an IMPS. In addition, the
problems caused by pre-slaughter environment: pale, soft, exudative
(PSE) and dark, rm, dry meat (DFD), would not exist in the products of
myocyte culturing.
While the controlled conditions in vitro allow the likelihood of
unpredictable complications to be minimized, they can be manipu-
lated to intentionally create products with differing qualities. With the
advent of functional and enriched foods, consumers are more willing
to try products that have been altered to have particular nutritional
characteristics (Korhonen, 2002; Burdock, Carabin, & Grifths, 2006).
By co-culture, medium formulation or genetic engineering, it is
theoretically possible to create products with different tastes, textures
and nutrient proles.
Fat content is an example of a nutritional quality that can be
manipulated. The greatest criticism against meat is the high saturated
fat content which contributes to cardiovascular disease. In vitro,
saturated fat content is determined by the amount of adipocyte co-
culture present; without co-culture, pure myocyte culture would
produce a product rich in nutritionally benecial fatty acids from
phospholipids. Furthermore, genetic manipulation of myocytes could
allow the fatty acid composition to be altered to enrich for particularly
benecial fatty acids such as those which are omega-3 polyunsaturated.
While in theory nutrient content can be altered, one major
obstacle likely to postpone the development of an IMPS is ensuring
that the product has the full complement of nutrients available in
meat harvested in vivo.
8. Limitations to the methodology
The scientic development of an IMPS is most hindered by the facts
that a) a major tissue will be cultured in the absence of in vivo
homeostatic regulation and b) that the process needs to be carried out
on a large scale. The lack of homeostasis affects the nutritional value of
the meat product, as often other organ systems are involved in nutrient
absorption and distribution in the live organism. Muscle tissue is also a
highly metabolic tissue and the products of metabolism need to be
removed or recycled at a pace matching the provision of reactants.
Large-scale tissue culturing is a concept crucial to thedevelopment of an
IMPS but is as of yet unexplored in the eld of tissue engineering.
8.1. Nutritional value
In addition to having high protein content with the full complement
of amino acids, meat is an exclusive sourceof several essentialnutrients.
It is necessary that an in vitro grown meat product meets if not exceeds
the nutritional value of traditional meat products to be competitive on
the market. Nutrients found in meat in vivo which are not synthesized
by muscle cells must be supplemented.
For instance, the essential vitamin B12 is synthesized exclusively
by certain species of gut-colonizing bacteria and is therefore found
solely in food products of animal origin. Supplementation of
crystalline vitamin B
produced commercially by biosynthetic
microbial fermentation would be necessary in an in vitro meat
product grown in an aseptic environment.
Iron in meat is present as the Fe(II) ion in the highly bioavailable
form of heme, the prosthetic group found in myoglobin. To provide
iron to growing myocytes in a bioavailable form, Fe(III) ions bound to
the plasma binding protein transferrin will have to be supplemented
to the culture medium. By transferrin-mediated iron transport, iron
can enter the myocyte mitochondria and be incorporated into heme
synthesis and subsequent myoglobin synthesis (Aisen, Enns, &
Wessling-Resnick, 2001). It must be noted that levels of transferrin
must be closely monitored, so as not to allow free ferric or ferrous ions
to be present in the medium, as they can readily catalyze the
production of damaging reactive oxygen species in aerobic environ-
ments (Papanikolaou & Pantopoulos, 2005). In addition, Graber and
Woodworth (1986) found that myoglobin levels are not signicant in
culture until a stable population of myotubes has formed, a fact which
can help determine the optimal growth time necessary before harvest.
Vitamin B
and heme iron are two especially important and
exclusively meat-source nutrients of several. A great challenge in
producing a competitive in vitro grown meat product is ensuring that
all necessary nutrients are present. Dietary minerals and vitamins not
synthesized by myocytes will often require binding proteins in
medium and effective transport mechanisms for entry into the cells.
Knowledge of the complex metabolism of each crucial vitamin and
mineral is necessary to develop a nutritionally valuable meat product.
While determining the proper nutrient prole will be a major hurdle
to overcome, it comes with the knowledge of how to manipulate the
culturing system to make nutritionally tailored products.
8.2. Metabolism
An IMPS lacks the organ systems that maintain homeostasis in an
organism and therefore metabolism needs to be carefully and strictly
monitored. Myocytes must be metabolizing aerobically to prevent
acidication of the culture medium with lactic acid. Oxygen must be
readily available to myocyte culture to prevent hypoxia and the
acidication of medium, two situations which are damaging to the
cells. As mentioned above, adequate oxygenation is dependent on the
ability of the bioreactor to enhance perfusion and the availability of
oxygen in the medium. Adequately providing the chemical and physical
requirements for metabolism while removing damaging waste products
can be a difcult task, with the possibility of unforeseen shortcomings.
For instance, co-culturing hepatocytes to convert lactate back to the
glucose via the Cori cycle can be one method of preventing lactic
acidication of medium in the eventof anaerobic conditions. This can be
complicated, however, by the individual metabolic needs of hepato-
cytes, which may not coincide well with those of myocytes.
Also important to consider is that muscle tissue and meat are
biochemically (and therefore qualitatively) dissimilar. The metabolic
reactions that proceed post-slaughter: anaerobic glycolysis, lactic acid
accumulation, protein denaturation and enzymatic proteolysis, are
responsible for producing the textural quality, taste and appearance of
meat. One uncertainty concerning metabolism is whether these
typical post-slaughter reactions will occur in cultured meat after
harvest in like manner to properly convert cultured muscle tissue to
meat as it is traditionally dened.
8.3. Large-scale operation
The development of a large-scale IMPS facility with the capability
to mass produce meat at a rate comparable to traditional slaughter-
houses is greatly hindered by the lack of investigation into large-scale
culturing. Presently, largepieces of cultured tissue are measured on
a centimetre scale with most relevant tissue engineering efforts have
been put towards the medical application of in vivo tissue repair.
Vladimir Mironov (Medical University of South Carolina, personal
communication, 19 April 2009) has stated that an industrial scale
bioreactor would need to be at least three to ve storeys to produce
industrially relevant amounts of cultured meat. The best source of
inspiration for mass production at this time would be to look towards
the pharmaceutical industry and microbial biotechnology where
living organisms in bioreactors manufacturepuried chemicals.
Though the mass production of puried molecules is quite different
from the mass production of cultured tissue, some of the technology
and methodology may prove relevant.
While stem-cell lines are dened by the ability to be propagated in
culture indenitely while maintaining broad plasticity, a great criticism
of stem cells in long-term culturing is spontaneous transformation.
19I. Datar, M. Betti / Innovative Food Science and Emerging Technologies 11 (2010) 1322
9. Conclusion and future prospects
Developments in regenerative medicine, stem-cell research and
biomaterial engineering in the past decade have yielded highly
applicable, viable options which can aid in the scientic development
of an IMPS. One possible in vitro meat production scheme is shown in
Fig. 2. With the scaffold-technique being a practicable method for
scale up, the materials and techniques now available make the IMPS
seem more achievable, however, there remain several aspects of the
concept yet to be investigated.
Because nearly all research applicable to the development of an IMPS
is intended for biomedical application, many of the investigations into
multipotent and myogenic cell lineages have been done with human,
mouse or ratcell lines, with relatively few studies describing cell lines of
agriculturally relevant animals. Adult stem cells and dedifferentiated
cells are recently innovated cell types which require further exploration
into their safety and suitability for an in vitro meat production system.
The biological aspects of an IMPS need to be further investigated to
ensure that the resulting tissue closely mimics native muscle tissue
morphologically and nutritionally. In addition, relevant studies have so
far focused on small-scale applications, with pertinent biological
research focused towards tissue replacement in vivo. Undoubtedly, a
major hindrance in development of the science of an IMPS is based
around application on a massproduction scale, where little research has
been done. Many of the cell lines, scaffold materials and medium
components described are unreasonable for scale up economically, and
are often complicated by other issues such as environmental impact and
safety. It is absolutely necessary that anin vitro meatproduction system
be developed according to the rules and regulations of good cell culture
practice (GCCP) (Coecke et al., 2005) as well as current good
manufacturing practice (cGMP) as pertaining to food and drug
The greatest stumbling block comes with the commercial
implementation of an IMPS, where cost-effectiveness and consumer
acceptance determine if cultured myocyte tissue will become a
signicant meat alternative on the market. A preliminary economics
study reviewing the nancial viability of in vitro grown meat
estimated the cost of manufacturing to be Euro 3500/ton, but note
that because such technology has not yet been developed, this
estimation could be largely inaccurate (eXmoor Pharma Concepts,
2008). In vitro meat production on an industrial scale is feasible only
when a relatively cost-effective process creating a product qualita-
tively competitive with existing meat products is established and
provided with governmental subsidization like that provided to other
Culturedmeat can certainlyhave an applicationin ground, processed
foods such as hamburgers or hotdogs as a main component or as an
additive. In this form, the textural shortcomings of the in vitro grown
productwill not compromise the nalprocessed productand one would
expect greater consumer acceptance. The second goal of cultured meat
is to create three-dimensional products resemblingtraditional cuts with
proper textural characteristics.
Thanks to Jason G. Matheny, director of New Harvest for his support
and Vladimir Mironov of the Medical University of South Carolina and
Bernard A.J. Roelen of the Utrecht University for helpful comments on
the manuscript. The Authors also express their gratitude to Prof. Carlo
Pelanda (University of Georgia) and to his book "Futurizzazione"
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... Cells are typically grown on an edible scaffold that is designed to give them the structure and texture of meat [35]. The ideal result is that cultured seafood becomes indistinguishable from whole fish meat in terms of sensory experience and nutritional value of traditional seafood products [36]. Cell-based meat could help minimize the environmental issues and the effect of climate change due to current meat production. ...
... The combination of improved cell culture and tissue engineering has facilitated the production of meat outside the body of an animal [58]. With the objective of producing meat, which is physiologically and biologically similar to conventional farm meat, these muscle tissues are produced by culturing stem cells of an organism in the controlled environment of a bioreactor with appropriate media and nutrients to differentiate itself into the muscle cell and/or fat cells and proliferate to increase the mass of meat in the scaffold [36]. The schematic representation of networking and future prospective of cellular aquaculture is represented in Figure 2. ...
... The schematic representation of networking and future prospective of cellular aquaculture is represented in Figure 2. For in vitro meat production, cells, such as embryonic stem cells, and adult stem cells (i.e., microsatellite cells) are used [36,59]. The selection of stem cell type differs based on the objective, i.e., for increased regenerative capacity, satellite cells are preferred over other cells, muscle cells are preferred for the production of protein-rich meat [60], and adipose tissue-derived adult stem cells are selected for its ability to produce myogenic, osteogenic, and adipogenic cell lineages [61]. ...
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Aquaculture plays an important role as one of the fastest-growing food-producing sectors in global food and nutritional security. Demand for animal protein in the form of fish has been increasing tremendously. Aquaculture faces many challenges to produce quality fish for the burgeoning world population. Cellular aquaculture can provide an alternative, climate-resilient food production system to produce quality fish. Potential applications of fish muscle cell lines in cellular aquaculture have raised the importance of developing and characterizing these cell lines. In vitro models, such as the mouse C2C12 cell line, have been extremely useful for expanding knowledge about molecular mechanisms of muscle growth and differentiation in mammals. Such studies are in an infancy stage in teleost due to the unavailability of equivalent permanent muscle cell lines, except a few fish muscle cell lines that have not yet been used for cellular aquaculture. The Prospect of cell-based aquaculture relies on the development of appropriate muscle cells, optimization of cell conditions, and mass production of cells in bioreactors. Hence, it is required to develop and characterize fish muscle cell lines along with their cryopreservation in cell line repositories and production of ideal mass cells in suitably designed bioreactors to overcome current cellular aquaculture challenges.
... Since this pair of papers, the literature on IVM has evolved in a number of directions. Early publications largely consisted of overviews of the general IVM production process (Edelman et al., 2005;Hopkins and Dacey, 2008;Datar and Betti, 2010;Bhat and Fayaz, 2011;Post, 2012), environmental impacts (Tuomisto and Teixeira de Mattos, 2011;Tuomisto et al., 2014), or ethics (Pluhar, 2010;Welin and Van der Weele, 2012). These articles tended to present IVM in overall positive terms. ...
... It is also interesting to note that recent review papers of IVM deviate little if at all from earlier summary papers. Bhat et al. (2020) reads like that of Datar and Betti (2010). Stephens et al. (2019) mention that current industry challenges are essentially the same as those at the first IVM conference in 2008. ...
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This review essay documents continuities between (industrial) animal agriculture and cellular agriculture and raises key questions about whether or not the technology might be able to deliver on its promise of food system transformation. It traces how industrial history, connections to the livestock industry, and disavowal are extended through the innovation of cellular agriculture. In particular, it is shown that cellular agriculture has had connections to (industrial) animal agriculture since its very beginning and at nearly every step since then. I argue that cellular agriculture can be positioned as the epitome of (industrial) animal agriculture in terms of history, material practices, and ideology. Such a critique of cellular agriculture has become somewhat commonplace but while a number of papers have raised similar concerns individually, there exists no sustained focus on such similarities to make this point holistically. Such connections are important in framing the future of cellular agriculture and the fate of farmed animals and the environment. Carefully considering the continuities between cellular agriculture and animal agriculture is crucial when considering whether promoting cellular agricultural is a prudent approach to addressing problems associated with animal agriculture. The cumulative number and extent of connections covered in this essay leads to questions of who will benefit with the advent of cellular agriculture.
... However, it has been indicated that the consumption of meat and meat products will double by 2050, and beef consumption will increase by 200% in developing countries (FAO, 2011;Chriki and Hocquette, 2020). This increase could be attributed to several reasons, including (a) the processing of meat into ready-to-eat products (such as, sausage, polonies, and patties), (b) the application of innovative technologies to increase production and improve qualities, such as tenderness, leanness (fat content), palatability, freshness, shelf life, and safety, (c) awareness on its inherent essential micronutrients, such as iron, zinc, calcium, vitamin A, vitamin B1, and other bioactive compounds (Datar and Betti, 2010). Beyond this, the shift toward high meat consumption has been attributed to an increase in income and the human population, which has been forecast to reach 9 billion in 2050 (Nellemann et al., 2009;Alexandratos and Bruinsma, 2012;OECD-FAO, 2021). ...
... Furthermore, evidence has shown that a majority of people will support the production of cultured meat if it is healthy and safe (Haagsman et al., 2009;Goodwin and Shoulders, 2013), such as vegetarians and environmentalists, since its production will eliminate any cruel or suffering associated with factory farmed animals (Schaefer and Savulescu, 2014). In addition, other researchers have shown that the production of cultured beef will improve the biochemical composition of meat and reduce public concern about animal welfare, greenhouse emissions, foodborne disease, and biodiversity loss (Edelman et al., 2005;Datar and Betti, 2010;Canon, 2011;Goffman, 2012;Chriki and Hocquette, 2020). Furthermore, the willingness displayed by some of the respondents to eat cultured beef when commercially available also gave them the assurance that its production would enhance meat production. ...
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The creation and growing popularity of cultured meat has raised mixed reactions among consumers about its originality, acceptability, edibility, and nutritional quality across the world. The perception and reaction of consumers to novel meat are influenced by a variety of factors, such as geographical location, media coverage, educational status, culture, and religion. Therefore, this study was designed to examine the perceptions of consumers on the consumption of natural vs. cultured beef in the Eastern Cape Province, South Africa. A total of 255 respondents were interviewed using structured questionnaires, and the data were analyzed using descriptive statistics and X2 tests. Interviewees included representatives from University (educated), urban (literate), and rural (semi-literate) communities. The results revealed the majority (63%) of the respondents had not heard about the concept of cultured beef production, of which 27% of them were men and 36% were women. More than half (53%) of the respondents indicated their willingness to eat cultured beef if offered to them after explaining the concept and process of making cultured beef to them. Among all factors that were analyzed, the participant level of education was found to significantly influence their willingness to eat cultured beef when available commercially. It is therefore concluded that the majority of consumers in this study supported the concept of cultured meat as an alternative way to complement conventional meat production and would be willing to eat it when provided.
... Considering the traction of cultured meat products (growing cells to generate meat-like tissue structures in the laboratory) as an alternative protein source for human consumption, the glycoprofile workflow described in this study can also be used in establishing the critical quality attributes (CQA) of cultured meat products [39][40][41]. Given the considerable risk of cell line contamination and product adulteration in this growing industry, such techniques can help to establish a stringent quality control of these cultured meat samples in the future [42,43]. ...
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It is estimated that food fraud, where meat from different species is deceitfully labelled or contaminated, has cost the global food industry around USD 6.2 to USD 40 billion annually. To overcome this problem, novel and robust quantitative methods are needed to accurately characterise and profile meat samples. In this study, we use a glycomic approach for the profiling of meat from different species. This involves an O-glycan analysis using LC-MS qTOF, and an N-glycan analysis using a high-resolution non-targeted ultra-performance liquid chromatography-fluorescence-mass spectrometry (UPLC-FLR-MS) on chicken, pork, and beef meat samples. Our integrated glycomic approach reveals the distinct glycan profile of chicken, pork, and beef samples; glycosylation attributes such as fucosylation, sialylation, galactosylation, high mannose, α-galactose, Neu5Gc, and Neu5Ac are significantly different between meat from different species. The multi-attribute data consisting of the abundance of each O-glycan and N-glycan structure allows a clear separation between meat from different species through principal component analysis. Altogether, we have successfully demonstrated the use of a glycomics-based workflow to extract multi-attribute data from O-glycan and N-glycan analysis for meat profiling. This established glycoanalytical methodology could be extended to other high-value biotechnology industries for product authentication.
... Scaffolds are the framework for cells to adhere, grow and attain tissue maturity by mimicking the native three dimensional tissue (Handral et al., 2020). Scaffolds for cultured meat production must have biologically active, large surface area, flexibility and porosity to support tissue maturation and should be edible without any allergic responses when digested (Datar and Betti, 2010). The fibrous nature of the scaffold is important to enhance the organoleptic properties of printed meat products. ...
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Three-dimensional (3D) printing is an innovative technology adopted to develop customized products. 3D bioprinting is the utilization of 3D printing-like techniques to combine cells, growth factors and biomaterials to fabricate products that maximally imitate natural tissue characteristics. The technology has its major advancements in medical, drug and cosmetics field and is in its infancy stage in food and agricultural fields. 3D bioprinting applications in plant science field including investigation of cell dynamics, cultivation of cells and fabrication of customized plant culture systems is a matter of study present days. The major research in 3D bioprinting in food industry is done in meat applications. Attempts are being made to culture meat cells in suitable medium and to convert it into printable form that can be further processed. Researches are progressing to develop real textured food using plant tissue regeneration. Further, development of 3D printable food packaging systems from bio materials is also a matter of research. The review throws light on the current developments and methods of 3D bioprinting process in food and agriculture sector.
... Cultured meat is meat grown from animal cells, mimicking the process by which cells grow, divide and differentiate in vivo to produce a product with the same nutritional and organoleptic properties as its conventional counterpart. Different cell types have been considered for cultured meat, including embryonic stem cells (ESCs), induced pluripotent stem cells (IPSCs), mesenchymal stem cells (MSCs), and satellite cells (SCs) [71,72]. More recently, progenitor cells or proliferating adult cells such as fibroblasts, myoblasts, etc. are being adopted. ...
Due to environmental and ethical concerns, meat analogs represent an emerging trend to replace traditional animal meat. However, meat analogs lacking specific sensory properties (flavor, texture, color) would directly affect consumers' acceptance and purchasing behavior. In this review, we discussed the typical sensory characteristics of animal meat products from texture, flavor, color aspects, and sensory perception during oral processing. The related strategies were detailed to improve meat-like sensory properties for meat analogs. However, the upscaling productions of meat analogs still face many challenges (e.g.: sensory stability of plant-based meat, 3D scaffolds in cultured meat, etc.). Producing safe, low cost and sustainable meat analogs would be a hot topic in food science in the next decades. To realize these promising outcomes, reliable robust devices with automatic processing should also be considered. This review aims at providing the latest progress to improve the sensory properties of meat analogs and meet consumers' requirements.
Cell-based meat has attracted great attention in recent years as a novel product of future food biomanufacturing and a breakthrough in the global food industry. Previous reports mainly focus on the relatively independent investigation of the nature and consumer acceptance of cultured meat, and there is limited research upon its commercialization, safety, and quality control. Based on the existing literature, we overview current cultured meat startups distribution, product varieties, investment, and financing status. Furthermore, the challenges of commercializing cultured meat products are systematically discussed from the aspects of key technologies, safety and supervision, and market expectation. Finally, some strategies and prospects related to the marketing of cultured meat are put forward. Although some cultured meat startups’ development and financing results are exciting, the greatest obstacles to the market promotion of cultured meat products are the large-scale production, safety assessment, improvement of a supervision system, and product-based market survey influenced by technology challenges.
The Federal Constitution dictates that anyone who causes suffering to an animal by imposing suffering on it due to mistreatment, infringes and incurs a crime provided for in Article 32 of Law No. 9605/1998. However, in practice this is not what happens, under this perspective a vast legal support has been consolidated that aims to recognize the individual value of animal life, seeking to bring ethical and moral aspects that preserve and protect animal life. Cell-based meat is an alternative to conventional meat that does not require the rearing and slaughter of animals. With the need for increased meat production, it grows along with the dependence on the availability of large areas of pasture, amount of water and energy to support for creating number of animals, which, in turn, leads to an increase in pasture areas greenhouse effect and carbon dioxide concentration, and especially to aspects related to ethics and animal welfare. Thus, alternatives are needed to meet the world demand for animal protein, but above all respecting the animals, and among the options is cell meat, a new technology for food production. Therefore, it is extremely important that professionals involved in the conventional meat production chain have knowledge about the process, so that they can assume new roles in the chain of cellular meat processing. This review aims to bring information and clarification to veterinarians, zootechnicians and other Brazilian professionals running the system. Since cellular meat seems to be a close reality, and knowledge about its processing must be disseminated widely to reach working professionals and intend to work in the meat production chain, demystifying taboos to add value to the development of sustainable alternatives and consequently new opportunities.
In response to a growing population and rising food demand, the food industry has come up with a wide array of alterations, innovations, and possibilities for making meat in vitro. In addition to revolutionizing the meat industry, this advancement also has profound effects on the environment, health, and welfare of animals. Thus, rather than using slaughtered animals, animal cells are employed to generate cell-based meat, with the cells' proliferation and differentiation taking place in the culture environment. The primary goal of this paper is to examine the overall mechanism and numerous approaches involved in the creation of cell-based meat. It also covers upcoming issues like technical, consumer, and regulatory issues, environmental concerns, the economy, cost of the product, health and safety concerns, and ethical, religious, and societal taboos. Finally, it assesses the future prospects of cell-based meat production.
Contractile myocytes provide a test of the hypothesis that cells sense their mechanical as well as molecular microenvironment, altering expression, organization, and/or morphology accordingly. Here, myoblasts were cultured on collagen strips attached to glass or polymer gels of varied elasticity. Subsequent fusion into myotubes occurs independent of substrate flexibility. However, myosin/actin striations emerge later only on gels with stiffness typical of normal muscle (passive Young's modulus, E ~12 kPa). On glass and much softer or stiffer gels, including gels emulating stiff dystrophic muscle, cells do not striate. In addition, myotubes grown on top of a compliant bottom layer of glass-attached myotubes (but not softer fibroblasts) will striate, whereas the bottom cells will only assemble stress fibers and vinculin-rich adhesions. Unlike sarcomere formation, adhesion strength increases monotonically versus substrate stiffness with strongest adhesion on glass. These findings have major implications for in vivo introduction of stem cells into diseased or damaged striated muscle of altered mechanical composition.
In a series of experiments, cultured myotubes were exposed to passive stretch or pharmacological agents that block contractile activation. Under these experimental conditions, the formation of Z lines and A bands (morphological structures, resulting from the specific structural alignment of sarcomeric proteins, necessary for contraction) was assessed by immunofluorescence. The addition of an antagonist of the voltage-gated Na+ channels [tetrodotoxin (TTX)] for 2 days in developing rat myotube cultures led to a nearly total absence of Z lines and A bands. When contractile activation was allowed to resume for 2 days, the Z lines and A bands reappeared in a significant way. The appearance of Z lines or A bands could not be inhibited nor facilitated by the application of a uniaxial passive stretch. Electrical stimulation of the cultures increased sarcomere assembly significantly. Antagonists of L-type Ca2+ channels (verapamil, nifedipine) combined with electrical stimulation led to the absence of Z lines and A bands to the same degree as the TTX treatment. Western blot analysis did not show a major change in the amount of sarcomeric alpha -actinin nor a shift in myosin heavy chain phenotype as a result of a 2-day passive stretch or TTX treatment. Results of experiments suggest that temporal Ca2+ transients play an important factor in the assembly and maintenance of sarcomeric structures during muscle fiber development.
In this report, the possibility of using in vitro cell culturing techniques for the production of meat is investigated. There are several reasons for investing such a system. One application could be to reduce animal suffering, environmental polution or land use associated with current meat production methods. Meat is an important nutritional and social factor for the human race and meat consumption in the world is expected to increase dramatically during the coming decades. Another application could be a continuous meat supply for long-term manned space missions in the far future. It is probably possible to produce meat in vitro by culturing loose myosatellite cells on a substrate. After differentiation muscle cells could be harvested and used as processed meat. The culturing of actual muscle tissue is also an option, as long as a nutrient-and oxygen perfusion system of some kind can be established. Detailed proposals exist for the former method, and experiments have been performed on the latter method. It is not known whether in vitro cultured meat would be well accepted by the consumer, but the initial reaction seems to be moderately negative. Especially genetic manipulation might hinder consumer acceptance. To establish an sustainable in vitro meat culturing system on an industrial scale, a great deal of research is still needed on the use of culture media.
Skeletal muscle satellite cell culture techniques have provided a means for coupling muscle structural and physiological observations to cellular and molecular explanations. Intrinsic properties of these myogenic cells have been studied in vitro, and it has become apparent that the satellite cell has properties that are distinctly different from those of embryonic or fetal myogenic cells. This chapter describes two types of satellite cell culture procedures—(1) monolayer mass cultures of dissociated satellite cells and (2) single muscle fiber cultures with their associated satellite cells—and discusses their application. In both cases, cells and fibers are isolated directly from experimental subjects. The general cellular processes that have been studied in satellite cell culture systems include activation from the quiescent state, migration, proliferation, and differentiation or return to quiescence. There are advantages and disadvantages to each of the culture systems and one single culture system may not be adequate for addressing all experimental questions.
Blood substitutes are oxygen-carrying fluids that aim to provide an alternative to the transfusion of blood. Strategies for developing such substitutes have involved the production of materials based on the naturally occurring respiratory pigment, haemoglobin (Hb), or synthetic, chemically inert, fluorinated liquids called perfluorochemicals (PFCs). Commercial products in both categories have been developed and some approved for clinical use, primarily to facilitate oxygen supply to tissues during surgery or therapy. The latest research is focused on using microbial and plant ‘cell factories’ to express recombinant Hb, understanding the properties of polymeric Hbs from invertebrate animals, and the use of feedback from stakeholders to inform the development of new educational materials to assist patients to make informed choices on future transfusion options.
This was published as Methods in Cell Science, which was incorporated into Cytotechnology. Primary and clonal culture systems have been devised and refined for animal-derived satellite cells. Satellite cell (SC) culture development includes efficient cell isolation techniques, establishment of effective plating and growth conditions, formulation of media requirements and thorough evaluation of experimental limitations. As the field of muscle cell culture has expanded, the number of animal species from which satellite cells have been isolated has increased. The focus of this paper is to compare and contrast SC culture conditions presently used by a variety of researchers and to introduce a new source of SC from wapiti (elk).
Recent advances in the food and nutrition sciences support the concept that the diet has a significant role in the modulation of various functions in the body. The diet and/or its components may contribute to an improved state of well-being, a reduction of risks related to certain diseases and even an improvement in the quality of life. These new concepts have led to the introduction of a new category of health-promoting foodstuffs, i.e. functional foods. The concern about health embraces a number of driving issues, needs and opportunities which may be approached by designing specific diets from various food raw materials. These tailor-made products provide physiological benefits that are targeted at particular consumer groups. The functionality of functional foods is based on bioactive components, which may be contained naturally in the product but usually require formulation by appropriate technologies in order to optimise the desired beneficial properties. To this end, it is often necessary to develop and apply novel technologies, e.g. membrane separation, high hydrostatic pressure and supercritical fluid extraction techniques. Also the minimal processing concept could be employed in this context. This review discusses the current technological options available and the future challenges faced in the area. Particular attention is paid to the exploitation of bovine colostrum and milk-derived bioactive compounds for the development of functional foods.
We describe our protocol for isolating myogenic cells from the semimembranous muscles of lamb. Cultured cells display properties similar to myogenic satellite cells isolated from other species; similar characteristics include the ability to proliferate, differentiate, and fuse to form myotubes in primary cell culture. Using the described procedures, we provide the first documentation of satellite like cells existing in postnatal skeletal muscle of a ruminant animal species.