Morc2a variants cause hydroxyl radical-mediated neuropathy and are rescued by restoring GHKL ATPase.

Mutations in the Microrchidia CW-type zinc finger 2 (MORC2) GHKL ATPase module cause a broad range of neuropathies, such as Charcot-Marie-Tooth disease type 2Z; however, the aetiology and therapeutic strategy are not fully understood. Previously, we reported that the Morc2a p.S87L mouse model exhibited neuropathy and muscular dysfunction through DNA damage accumulation. In the present study, we analysed the gene expression of Morc2a p.S87L mice and designated the primary causing factor. We investigated the pathological pathway using Morc2a p.S87L mouse embryonic fibroblasts and human fibroblasts harbouring MORC2 p.R252W. We subsequently assessed the therapeutic effect of gene therapy administered to Morc2a p.S87L mice. This study revealed that Morc2a p.S87L causes a protein synthesis defect, resulting in the loss of function of Morc2a and high cellular apoptosis induced by high hydroxyl radical levels. We considered the Morc2a GHKL ATPase domain as a therapeutic target because it simultaneously complements hydroxyl radical scavenging and ATPase activity. We used the adeno-associated virus (AAV)-PHP.eB serotype, which has a high CNS transduction efficiency, to express Morc2a or Morc2a GHKL ATPase domain protein in vivo. Notably, AAV gene therapy ameliorated neuropathy and muscular dysfunction with a single treatment. Loss-of-function characteristics due to protein synthesis defects in Morc2a p.S87L were also noted in human MORC2 p.S87L or p.R252W variants, indicating the correlation between mouse and human pathogenesis. In summary, CMT2Z is known as an incurable genetic disorder, but the present study demonstrated its mechanisms and treatments based on established animal models. This study demonstrates that the Morc2a p.S87L variant causes hydroxyl radical-mediated neuropathy, which can be rescued through AAV-based gene therapy.

Review of technology and materials for the development of cultured meat.

Cultured meat production technology suggested that can solve the problems of traditional meat production such as inadequate breeding environment, wastewater, methane gas generation, and animal ethics issues. Complementing cultured meat production methods, sales and safety concerns will make the use of cultured meat technology easier. This review contextualizes the commercialization status of cultured meat and the latest technologies and challenges associated with its production. Investigation was conducted on materials and basic cell culture technique for cultured meat culture is presented. The development of optimal cultured meat technology through these studies will be an innovative leap in food technology. The process of obtaining cells from animal muscle, culturing cells, and growing cells into meat are the basic processes of cultured meat production. The substances needed to production of cultured meat were antibiotics, digestive enzymes, basal media, serum or growth factors. Although muscle cells have been produced closer to meat due to the application of scaffolds materials and 3 D printing technology, still a limit to reducing production costs enough to be used as foods. In addition, developing edible materials is also a challenge because the materials used to produce cultured meat are still not suitable for food sources.

Generation of myostatin-knockout chickens mediated by D10A-Cas9 nickase.

Many studies have been conducted to improve economically important livestock traits such as feed efficiency and muscle growth. Genome editing technologies represent a major advancement for both basic research and agronomic biotechnology development. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technical platform is a powerful tool used to engineer specific targeted loci. However, the potential occurrence of off-target effects, including the cleavage of unintended targets, limits the practical applications of Cas9-mediated genome editing. In this study, to minimize the off-target effects of this technology, we utilized D10A-Cas9 nickase to generate myostatin-knockout (MSTN KO) chickens via primordial germ cells. D10A-Cas9 nickase (Cas9n)-mediated MSTN KO chickens exhibited significantly larger skeletal muscles in the breast and leg. Degrees of skeletal muscle hypertrophy and hyperplasia induced by myostatin deletion differed by sex and muscle type. The abdominal fat deposition was dramatically lower in MSTN KO chickens than in wild-type chickens. Our results demonstrate that the D10A-Cas9 technical platform can facilitate precise and efficient targeted genome engineering and may broaden the range of applications for genome-edited chickens in practical industrialization and as animal models of human diseases.

Effects of self-carbon dioxide-generation material for active packaging on pH, water-holding capacity, meat color, lipid oxidation and microbial growth in beef during cold storage.

BACKGROUND: Active packaging refers to the mixing of additive agents into packaging materials with the purpose of maintaining or extending food product quality and shelf life. The aim of this study was to develop an easy and cheap active packaging for beef. Beef loin samples were divided into three packaging groups (C, ziplock bag packaging; T1, vacuum packaging; T2, active packaging) and stored at 4 °C for 21 days. RESULTS: The water-holding capacity was significantly (P < 0.05) higher in C and T2 than in T1 for up to 7 days of storage. The TBARS value was significantly (P < 0.05) lower in T1 and T2 after 7 days of storage. The counts of some microorganism were significantly (P < 0.05) lower in T1 and T2 after 7 days of storage; the total bacterial count and Escherichia coli count were lowest in T2 at the end of storage. CONCLUSION: These results indicate that active packaging using self-CO2 -generation materials can extend the shelf life similarly to that observed with vacuum packaging, and that the active packaging method can improve the quality characteristics of beef during cold storage. © 2017 Society of Chemical Industry.

The effect of clove bud powder at a spice level on antioxidant and quality properties of emulsified pork sausage during cold storage.

BACKGROUND: Clove bud is a widely used spice in meat and meat products, and it contains high level of phenolic compounds. The effectiveness of the clove as a spice has not been fully studied at a general level of addition in the meat products. Therefore, in the present study, the antioxidant, antimicrobial, and nitrite scavenging abilities of clove bud powder (CBP) was assessed at spice level (0.1% and 0.2%) in emulsified pork sausage, during 6 weeks of cold storage. RESULTS: CBP had DPPH radical scavenging ability, but CBP addition at 0.1% and 0.2% did not decrease the TBARS value. An antimicrobial effect of CBP was also not observed during the cold storage. However, residual nitrite at storage weeks 4 and 6 was shown to be lower (P < 0.05). Addition of CBP decreased CIE L* and a* values, but it produced unacceptable sensory properties. Texture profile analysis was not affected by the addition of CBP in emulsified pork sausage (P > 0.05). CONCLUSIONS: The positive effect on nitrite scavenging could be expected by the addition of 0.2% CBP as a spice. However, antioxidant and antimicrobial abilities were not observed, as well as improvement in the quality of characteristics, in emulsified pork sausage. © 2016 Society of Chemical Industry.

Assessing Individual Muscle Characteristics to Enhance Frozen-Thawed Meat Quality.

This study assessed previous research aimed at mitigating the adverse effects of freeze-thawing on meat quality. Specifically, it focuses on assessing the physicochemical alterations in meat resulting from freezing, freeze-thawing, or technologies to minimize these alterations. Recent studies have focused on conventional freeze-thaw technology applicable across various livestock species and muscle types. However, recent research has indicated the necessity for developing freeze-thaw technology considering the unique characteristics of individual muscles. In this review, we summarize previous studies that have compared alterations in the physicochemical properties of primary muscles owing to freezing or freeze-thawing. Despite the introduction of various technologies to significantly reduce the adverse effects on meat quality resulting from freeze-thawing, it is essential to consider the unique characteristics (proximate composition, pH, and muscle fiber characteristics) of individual muscles or cuts to develop enhanced the freeze-thaw processing technology.

Comparative review of muscle fiber characteristics between porcine skeletal muscles.

Meat derived from skeletal muscles of animals is a highly nutritious type of food, and different meat types differ in nutritional, sensory, and quality properties. This study was conducted to compare the results of previous studies on the muscle fiber characteristics of major porcine skeletal muscles to the end of providing basic data for understanding differences in physicochemical and nutritional properties between different porcine muscle types (or meat cuts). Specifically, the muscle fiber characteristics between 19 major porcine skeletal muscles were compared. The muscle fibers that constitute porcine skeletal muscle can be classified into several types based on their contractile and metabolic characteristics. In addition, the muscle fiber characteristics, including size, composition, and density, of each muscle type were investigated and a technology based on these muscle fiber characteristics for improving meat quality or preventing quality deterioration was briefly discussed. This comparative review revealed that differences in muscle fiber characteristics are primarily responsible for the differences in quality between pork cuts (muscle types) and also suggested that data on muscle fiber characteristics can be used to develop optimal meat storage and packaging technologies for each meat cut (or muscle type).

Longitudinal and transversal intramuscular variation of muscle fiber and meat quality characteristics in bovine M. longissimus thoracis et lumborum.

This study evaluates longitudinal and transversal intramuscular variations in muscle fiber and meat quality characteristics in bovine M. longissimus thoracis et lumborum (LTL). The LTL muscles (n = 5) from the left side of the beef carcass were cut at intervertebral segment intervals (between 1st thoracic vertebra (TV) and 6th lumbar vertebra (LV)). The pennation angle demonstrated an increasing trend from the anterior to posterior regions regardless of the medial (M-zone) and lateral (L-zone) regions (P < 0.05). The M -zone had a higher pennation angle than the L-zone in the TV and 1st LV (P < 0.05). The cross-sectional area (CSA) of muscle fibers, excluding type I, was larger in the posterior region than the anterior region (P < 0.05). A larger CSA of type I/IIA, IIA, IIAX, and IIX was observed on the lateral side than on the medial side of the 13th TV (P < 0.05). Fiber types were more oxidative (types I and IIA) in the anterior region and more glycolytic (types IIA/IIX and IIX) in the posterior region. Fat content was higher in the anterior region than in the posterior region (P < 0.05). The lowest redness, yellowness, and Warner-Bratzler shear force values were observed in the middle of the muscle, whereas the lightness value was lower in the posterior region regardless of the transversal region (P < 0.05). Therefore, bovine LTL muscles exhibit unique morphological properties and contribute to understanding meat quality associated with morphological and muscle fiber characteristics in relation to their intramuscular variations.

Comparison of Chemical Composition, Quality, and Muscle Fiber Characteristics between Cull Sows and Commercial Pigs: The Relationship between Pork Quality Based on Muscle Fiber Characteristics.

This study aims to compare the chemical composition, quality, and muscle fiber characteristics of cull sows and commercial pigs, investigating the effect of changes in muscle fiber characteristics on pork quality. The proximate composition, color, pH, water-holding capacity (drip loss and cooking loss), protein solubility, total collagen content, and muscle fiber characteristics of cull sows (n=20) and commercial pigs (n=20) pork were compared. No significant differences were found between cull sows and commercial pigs in terms of proximate composition, drip loss, protein solubility, or total collagen content of their meat (p<0.05). However, cull sow pork exhibited a red color and a higher pH (p<0.05). This appears to be the result of changes in muscle fiber number and area composition (p<0.05). Cull sow meat also displayed better water-holding capacity as evident in a smaller cooking loss (p<0.05), which may be related to an increase in muscle fiber cross-sectional area (p<0.05). In conclusion, muscle fiber composition influences the pork quality; cull sow pork retains more moisture when cooked, resulting in minimal physical loss during processing and can offer more processing suitability.

Identification of myosin heavy chain isoforms in porcine longissimus dorsi muscle by electrophoresis and mass spectrometry.

Myosin heavy chain (MHC) isoforms have been considered as makers for muscle fiber types in relation to meat quality, whereas MHC isoforms in porcine skeletal muscle have not been fully identified. The improved technique of SDS-PAGE and 2DE were used to separate porcine MHC isoforms. Western blotting with monoclonal antibodies including BA-F8 (anti-MHC slow/I), SC-71 (anti-MHC 2a and 2x), 10F5 (anti-MHC 2b), and BF-35 (anti-MHC slow/I and 2a) and MS were used to confirm MHC migration rate and identify MHC isoforms from separated bands and spots. Up to 45% w/v of glycerol, 8% w/v of acrylamide content, and 25 h of electrophoretic time at 70 V allowed a clear separation of MHC isoforms. Major MHC isoforms such as slow, 2a, 2x, and 2b were clearly separated by SDS-PAGE. A total of 23 MHC spots were separated and identified by 2DE and MS. Therefore, four MHC isoforms such as slow/I, 2a, 2x, and 2b could be identified by the improved SDS-PAGE technique, 2DE and MS. Therefore, these techniques allow more accurate and accessible analysis in muscle fiber typing and in relationship between MHC isoforms, muscle fiber characteristics, and pork quality.

Effects of nitrite-rich and pigment-rich substitutes for sodium nitrite on the quality characteristics of emulsion-type pork sausages during cold storage.

This study was conducted to evaluate the effects of nitrite-rich (celery powder; CP) and pigment-rich (purple sweet potato powder, PSP; red beet powder, RB) substitutes for synthetic sodium nitrite (CON) on the quality characteristics of emulsion-type pork sausages during four weeks of cold storage. Natural substitutes decreased the pH, lightness, and textural properties of pork sausages during storage (P < 0.05). Pigment-rich substitutes showed a decreased antioxidant effect after two weeks of storage when compared to the nitrite-rich groups (CON and CP; P < 0.05). Pigment-rich substitutes also accelerated the discoloration of pork sausages by increasing yellowness (RB and PSP) and decreasing redness (PSP) during storage (P < 0.05). However, these two pigment-rich substitutes showed different trends in redness (higher in PSP and lower in RB) and yellowness (higher in RB and lower in PSP) when compared to the nitrite-rich groups (CON and CP). Different types (nitrite-rich and pigment-rich) of natural substitutes for sodium nitrite had different effects on the quality characteristics of emulsion-type pork sausages throughout the four weeks of storage evaluated in this study.

Effect of freezing raw meat on the physicochemical characteristics of beef jerky.

The effect of freezing raw meat on the quality characteristics of beef jerky was evaluated in the present study. Jerky was made using different types of raw beef (fresh, frozen, and frozen-thawed) with different curing times (6 h and 12 h). Frozen-thawed beef had a lower moisture content than fresh or frozen beef due to higher exudate loss (P < 0.05). Jerky made using frozen and frozen-thawed beef showed lower drying yield and higher shear force than jerky prepared using fresh beef (P < 0.05). Freezing raw beef decreased the fat content and increased the redness, yellowness, chroma, and hue values of jerky (P < 0.05). The microstructure of beef jerky was showed to increase the deformation and contraction of muscle fibers due to freezing. Longer curing times increased the moisture content of jerky made using frozen meat (P < 0.05). Jerky made using frozen or frozen-thawed meat was tough due to excessive fat and moisture loss.

Analysis of commercial fetal bovine serum (FBS) and its substitutes in the development of cultured meat.

Fetal bovine serum (FBS) is an extremely important culture growth supplement, accounting for approximately 60 % of cell-culture-media costs; therefore, lowering FBS-acquisition costs for the industrialization of cultured meat is imperative. This study attempted to produce an FBS substitute using discarded livestock by-products, with particular focus on formulating a product with a composition similar to that of FBS to improve effectiveness. However, to date, no study has precisely analyzed the commercial components of FBS, and this study is the first to compare the chemical composition of FBS and commercially available horse serum purchased from the United States or Europe with that of FBS substitutes developed by our team. This study analyzed the chemical composition of the FBS products purchased by our team over the past 3 years via blood, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and independent composition analyses. While the composition and quality of commercial FBS products are known to vary, the FBS composition of our purchased products was relatively uniform regardless of company, brand, or country of origin. In contrast, FBS substitutes obtained from three major livestock species (cattle, pig, and chicken) clearly exhibited differences in composition, a phenomenon that was also observed upon comparing with FBS as well as among different species. Therefore, to replace commercial FBS entirely, the production of a proportionately effective substitute product comprising an equal or similar composition is required, and the results of this study can be a steppingstone to achieving this. In addition, FBS substitutes manufactured using inexpensive slaughter by-products as raw materials are expected to ultimately reduce the unit cost of cultured meat production.

Improved culture procedure for bovine muscle satellite cells for cultured meat.

Many researchers and companies around the world are reported to have developed cultured meat, but their specific techniques have rarely been disclosed. Thus, the purpose of this study is to provide an improved procedure for cultured meat. There are four major steps in this cultured meat production: muscle cell isolation, proliferation, differentiation, and validation. The improved isolation enabled the efficient removal of unnecessary cells and tissues compared to previous procedures. In addition, proper use of basal media can improve the proliferation efficiency by about 2-fold. During the differentiation process, improved procedure was performed by using 10 % horse serum-containing media after 3 days of initial differentiation for myotube induction. This method demonstrated significantly enhanced myotube formation, up to 2.6-fold increase in area and up to 1.9-fold increase in fusion index compared to the previous method. This study provides a simple, improved procedure to enable more effective cultured meat production compared to previous procedures and is expected to help produce inexpensive and safe cultured meat.

Proteolysis and changes in meat quality of chicken pectoralis major and iliotibialis muscles in relation to muscle fiber type distribution.

The proteolysis trends and meat quality of the chicken pectoralis major (PM) and iliotibialis (IL) muscles stored at 4°C for 7 d were investigated. After 7 d of storage, the purge loss was higher (P < 0.05) in PM than in IL muscle. The difference in the composition of muscle fibers between PM (100% fast type) and IL (88.85% fast and 11.15% slow types) resulted in differences in proteolysis. Fructose-bisphosphate aldolase, troponin I, myosin heavy chain, and malate dehydrogenase exhibited the same tendencies, but pyruvate kinase, creatine kinase, L-lactate dehydrogenase, and triosephosphate isomerase exhibited different tendencies in the 2 muscles. The activity of cathepsin B was higher in PM than in IL during storage (P < 0.05). These results indicate that the proteolysis trend and changes in meat quality during cold storage are dependent on the different muscle fiber characteristics.

Comparison of Meat Quality Characteristics and Proteolysis Trends associated with Muscle Fiber Type Distribution between Duck Pectoralis Major and Iliotibialis Muscles.

This study was conducted to evaluate the proteolysis trends and change in meat quality during 10 days of cold storage in duck M. pectoralis major (PM) and M. iliotibialis (IL). Duck IL had a higher pH and greater degree of lightness but lower cooking loss than PM (p<0.05). During the 10-day cold storage, the pH value of PM declined significantly (p<0.05), while the meat quality traits of IL were not affected by cold storage (p>0.05). In PM, the redness increased from day 1 to day 5, while cooking loss was lower on day 10 compared to day 5 (p<0.05). There were no significant differences in the activities of cathepsin B and proteasome 20S during cold storage (p>0.05). The activity of calpains declined gradually during 10 days of storage (p<0.05), and the activity of calpains in PM was higher than that in IL (p<0.05). A total of 5,155 peptides were detected and derived from 34 proteins of duck PM muscle, whereas 4,222 peptides derived from 32 proteins were detected from duck IL muscle. Duck PM muscle was composed only of fast type of muscle fiber, whereas IL muscle was composed of both slow and fast types. The proteins responsible for glycolysis or myofibrillar proteins were closely related to changes in meat color or water-holding capacity during cold storage. These results suggest that changes in meat quality characteristics during cold storage are closely related to protein degradation, which is also related to the distribution of muscle fiber types.

A Comparative Study on the Taste Characteristics of Satellite Cell Cultured Meat Derived from Chicken and Cattle Muscles.

This study investigated the amino acid and nucleotide-related compound composition and taste characteristics of cultured muscle tissue (CMT) obtained by culturing satellite cells isolated from chicken and cattle and compared them to those of traditional meat (TM). The content of all amino acids except valine and tyrosine was significantly different between CMT and TM (p<0.05). The amount of glutamic acid was not significantly different between CMT and TM in cattle, but the glutamic acid in chicken CMT was lower than that of TM (p<0.05). Among the nucleotide-related compounds, only the content of inosine-5'-monophosphate (IMP) was significant, and the amount of IMP in CMT derived from chicken and cattle was significantly lower than that of TM (p<0.05). There were significant differences in the taste characteristics assessed by an electronic tongue system, and the umami, bitterness, and sourness values of CMT were significantly lower than those of TM from both chicken and cattle (p<0.05). The results of the present study suggest that it is necessary to develop a satellite cell culture method that could increase the umami and bitterness intensity of CMT and adjust the composition of the growth medium to produce cultured meat with a taste similar to that of TM.

Comparison of Meat Quality and Muscle Fiber Characteristics between Porcine Skeletal Muscles with Different Architectures.

This study aimed to compare the similarities, physicochemical properties, and muscle fiber characteristics of porcine skeletal muscles. Fourteen types of muscles were collected from nine pig carcasses at 24 h post-mortem and classified by muscle architecture into two main groups, namely parallel and pennate. The muscles were further differentiated into three subtypes per group. These included fan-shaped, fusiform, and strap for the parallel group, and unipennate, bipennate, and multipennate for the pennate group. Parallel-fibered muscles, which were composed of larger I, IIA, IIX, and IIXB fibers and a lower density of IIA fibers, showed higher redness and yellowness values than pennate-fibered muscles (p<0.05). However, the relative fiber area was not significantly different between the parallel and pennate groups (p>0.05). In the subtypes of parallel architecture, the strap group showed lower moisture content and higher redness values than the other subtypes and had considerably higher amounts of oxidative fibers (I and IIA; 72.3%) than the fan-shaped and fusiform groups (p<0.05). In the pennate group, unipennate showed comparatively lower moisture content and higher lightness than other pennate subtypes and was composed of smaller I, IIA, and IIX fibers than the bipennate and multipennate groups (p<0.05). Finally, a different trend of muscle clustering by hierarchical cluster analysis was found between physicochemical properties and muscle fiber characteristics. These results suggest that the physicochemical properties and muscle fiber characteristics of porcine skeletal muscles are not significantly dependent on morphological properties but are rather related to the intrinsic properties of the individual muscles.

Review of the Current Research on Fetal Bovine Serum and the Development of Cultured Meat.

The purpose of this review is to summarize studies that investigate blood and the main components of fetal bovine serum (FBS) in vertebrates, including major livestock, and review the current research on commercializing cultured meat. Detailed research on FBS is still lacking; however, some studies have shown that FBS consists of proteins, carbohydrates, growth factors, cytokines, fats, vitamins, minerals, hormones, non-protein nitrogen, and inorganic compounds. However, there are few studies on how the composition of FBS differs from blood or serum composition in adult animals, which is probably one of the main reasons for not successfully replacing FBS. Moreover, recent studies on the development of FBS replacers and serum-free media have shown that it is difficult to conclude whether FBS has been completely replaced or serum-free media have been developed successfully. Our review of the industrialization of cultured meat reveals that many basic studies on the development of cultured meat have been conducted, but it is assumed that the study to reduce or replace ingredients derived from fetuses such as FBS has not yet been actively developed. Therefore, developing inexpensive and edible media is necessary for the successful industrialization of cultured meat.

Frozen/thawed meat quality associated with muscle fiber characteristics of porcine longissimus thoracis et lumborum, psoas major, semimembranosus, and semitendinosus muscles.

To evaluate the relationship between muscle fiber characteristics and the quality of frozen/thawed pork meat, four different muscles, M. longissimus thoracis et lumborum (LTL), M. psoas major (PM), M. semimembranosus (SM), and M. semitendinosus (ST), were analyzed from twenty carcasses. Meat color values (lightness, redness, yellowness, chroma, and hue) changed due to freezing/thawing in LTL, which showed larger IIAX, IIX, and IIXB fibers than found in SM (P < 0.05). SM and ST showed a significant decrease in purge loss and an increase in shear force caused by freezing/thawing (P < 0.05). Compared with LTL, SM contains more type IIXB muscle fibers and ST had larger muscle fibers I and IIA (P < 0.05). PM was the most stable of all muscles, since only its yellowness and chroma were affected by freezing/thawing (P < 0.05). These results suggest that pork muscle fiber characteristics of individual cuts must be considered to avoid quality deterioration during frozen storage.