• 한국동물생명공학회지
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• 제37권4호
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• pp.292-297
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• 2022

Direct injection of CRISPR/Cas9 into zygotes enables the production of genetically modified nonhuman primates (NHPs) essential for modeling specific human diseases, such as Usher syndrome, and for developing novel therapeutic strategies. Usher syndrome is a rare genetic disease that causes loss of hearing, retinal degeneration, and problems with balance, and is attributed to a mutation in MYO7A, a gene that encodes an uncommon myosin motor protein expressed in the inner ear and retinal photoreceptors. To produce an Usher syndrome type 1B (USH1B) rhesus macaque model, we disrupted the MYO7A gene in developing zygotes. Identification of appropriately edited MYO7A embryos for knockout embryo transfer requires sequence analysis of material recovered from a trophectoderm (TE) cell biopsy. However, the TE biopsy procedure is labor intensive and could adversely impact embryo development. Recent studies have reported using cell-free DNA (cfDNA) from embryo culture media to detect aneuploid embryos in human in vitro fertilization (IVF) clinics. The cfDNA is released from the embryo during cell division or cell death, suggesting that cfDNA may be a viable resource for sequence analysis. Moreover, cfDNA collection is not invasive to the embryo and does not require special tools or expertise. We hypothesized that selection of appropriate edited embryos could be performed by analyzing cfDNA for MYO7A editing in embryo culture medium, and that this method would be advantageous for the subsequent generation of genetically modified NHPs. The purpose of this experiment is to determine whether cfDNA can be used to identify the target gene mutation of CRISPR/Cas9 injected embryos. In this study, we were able to obtain and utilize cfDNA to confirm the mutagenesis of MYO7A, but the method will require further optimization to obtain better accuracy before it can replace the TE biopsy approach.

• Journal of Ginseng Research
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• 제44권3호
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• pp.435-441
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• 2020

Background: As a process of aging, skeletal muscle mass and function gradually decrease. It is reported that ginsenoside Rb1 and Rb2 play a role as AMP-activated protein kinase activator, resulting in regulating glucose homeostasis, and Rb1 reduces oxidative stress in aged skeletal muscles through activating the phosphatidylinositol 3-kinase/Akt/Nrf2 pathway. We examined the effects of Rb1 and Rb2 on differentiation of the muscle stem cells and myotube formation. Methods: C2C12 myoblasts treated with Rb1 and/or Rb2 were differentiated and induced to myotube formation, followed by immunoblotting for myogenic marker proteins, such as myosin heavy chain, MyoD, and myogenin, or immunostaining for myosin heavy chain or immunoprecipitation analysis for heterodimerization of MyoD/E-proteins. Results: Rb1 and Rb2 enhanced myoblast differentiation through accelerating MyoD/E-protein heterodimerization and increased myotube hypertrophy, accompanied by activation of Akt/mammalian target of rapamycin signaling. In addition, Rb1 and Rb2 induced the MyoD-mediated transdifferentiation of the rhabdomyosarcoma cells into myoblasts. Furthermore, co-treatment with Rb1 and Rb2 had synergistically enhanced myoblast differentiation through Akt activation. Conclusion: Rb1 and Rb2 upregulate myotube growth and myogenic differentiation through activating Akt/mammalian target of rapamycin signaling and inducing myogenic conversion of fibroblasts. Thus, our first finding indicates that Rb1 and Rb2 have strong potential as a helpful remedy to prevent and treat muscle atrophy, such as age-related muscular dystrophy.

• 한방재활의학과학회지
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• 제19권4호
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• pp.19-35
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• 2009

Objectives : This experimental study was designed to investigate the effect of KangwhalSokdan-tang(Jianghuoxuduan-tang) on the muscle regeneration of atrophied rat muscle by hindlimb suspension. Materials and methods : In this study, Sprague-Dawley rats weighing about 250g were subjected to hindlimb suspension and divided into total four groups: Normal group(n=6), Control group(n=6), Hindlimb non-treatment group(n=6), Hindlimb treatment group(n=6). Experiments were seperately tried two times. The first trial was studied by the following two groups; The first was normal group(n=6). The second was group(n=18) for hindlimb suspension during 2 weeks (control I group). The second trial after 2 weeks hindlimb suspension was studied by the following three groups; The third group(n=6) was expired immediately after 2 weeks hindlimb suspension. The forth group(n=6) was given free activity during 2 weeks after 2 weeks hindlimb suspension. The fifth group(n=6) was administrated of KST during 2 weeks after 2 weeks hindlimb suspension. In order to investigate degree of muscle atrophy, body weight and gastrocnemius muscle mass were compared. To analyze muscle regeneration factors(expression of IGF-1, Myogenin, MyoD), Western blot was used. Results : The results were analyzed by statistical process as follows, 1. In body weight, all hindlimb suspension groups were lower than normal group, but tendency of increase was shown in KST group compared to non-treatment group after 2 weeks hindlimb suspension. 2. In gastrocnemius muscle mass, KST group on both side was significantly higher than non-treatment group after 2 weeks hindlimb suspension. 3. In case of IGF-I, Type I of KST group was significantly increased than non-treatment group, but Type II was not shown significance. 4. There was no significantly difference in Myogenin. 5. In MyoD, Type I of KST group was significantly increased than control group, and Type II of KST group was significantly increased than non-treatment group. Conclusions : In summary, this study demonstrates that KST administration has an effect to prevent muscle atrophy and contribute muscle regeneration and proliferation. And also it is suggested that IGF-I and MyoD is major factors of myogenesis expression to KST adminstration after hindlimb suspension.

• 생명과학회지
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• 제27권12호
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• pp.1479-1485
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• 2017

일반적으로 노화, 영양부족 및 다양한 만성질환에 의하여 유발되는 근위축은 근육 단백질 합성 억제 및 분해증가를 통하여 근섬유 및 근육의 밀도를 감소시키는 것으로 알려져 있다. 본 연구에서는 근위축과 관련된 in vitro 실험을 위한 C2C12 근아세포에서 근관세포로의 분화과정을 확립하고, 분화가 유발된 C2C12 근관세포를 대상으로 dexamethasone 및 hydrogen peroxide에 의한 근위축 유발 및 관련 단백질들의 발현 변화를 조사하였다. 먼저 C2C12 근아세포에 분화배지를 처리하였을 경우 근관세포로 분화가 유발되었으며, 분화와 관련된 단백질인 myogenin 및 myoD의 발현이 증가하는 것으로 나타났다. 분화가 유발된 C2C12 근관세포에 세포독성이 없는 조건의 dexamethasone 및 hydrogen peroxide를 처리하였을 경우 근관의 지름이 감소하였으며, 이러한 현상은 musclespecific ubiquitin ligases인 MAFbx/atrogin-1 및 MuRF1의 발현 증가와 함께 muscle-specific transcription factor인 myogenin 및 MyoD의 발현 감소와 관련이 있다는 것을 확인하였다. 본 연구 결과는 근위축과 관련된 in vitro 실험 모델의 구축을 위한 최적의 분화조건 확립과 함께 dexamethasone 및 hydrogen peroxide를 근위축 유도제로 사용할 수 있는 가능성 을 제시하는 것이다.

• Journal of Plant Biotechnology
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• 제38권1호
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• pp.69-76
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• 2011

This study describes the effect of myo-inositol on sustained cell division and plant regeneration from cotyledon-derived protoplast of cabbage (Brassica oleracea var. capitata). Freshly isolated protoplasts were cultured in modified Murashige and Skoog (MS) medium removed ammonia ions and containing $0.4\;mg\;l^{-1}$ thiamine HCl, $100\;mg\;l^{-1}$ myo-inositol, $2\;mgl^{-1}$ 2,4-D, $0.5\;mgl^{-1}$ BA, $30\;gl^{-1}$ sucrose and several concentrations of myo-inositol (2, 4, 6, 8, 10% (w/v)) as an osmotic stabilizer. After 3 weeks of culture in the dark at $25^{\circ}C$, the plating efficiency of cabbage protoplasts reached to $22.5{\pm}2.9%$ when cultured in modified MS medium supplemented with $2\;mgl^{-1}$ 2,4-D, $0.5\;mgl^{-1}$ BA, $30\;gl^{-1}$ sucrose and 8% (w/v) of myo-inositol at a density of $2{\times}10^5$ protoplasts/ml. Rapidly growing cell colonies after 3 weeks of culture were transferred to the same culture medium removed osmoticum. To induce shoot regeneration from calluses, calluses with about 2 mm in diameter were transferred to the MS medium containing $2\;mgl^{-1}$ BA and $0.5\;mgl^{-1}$ NAA. After further three weeks of incubation onto the medium in the light, green shoots were formed on the surface of calluses at a frequency of 30%. Upon transfer to half-strength MS basal medium, roots were formed onto the bottom of regenerated shoots without auxin treatments. These regenerated plantlets were successfully acclimatized to soil transfer, grown to normal mature plants. The cabbage protoplast culture system established in this study could be applied for production of somatic hybrids or cybrids by asymmetric protoplast fusion and mass proliferation of elite somatic clones of cabbage.

• 공업화학
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• 제5권4호
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• pp.573-579
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• 1994

본 연구에 사용된 비이온성 계면활성제는 1-O-oleoyl glycerol, 2-O-oleoyl-myo-inositol 및 methyl 2-O-oleoyl-${\alpha}$-glucopyranoside였다. 이들 3종류의 비이온성 계면환성제에 대한 용액성질과 가용화과정은 상평형에 의하여 관찰하였다. 그 결과 게면활성제/물 2성분계와 3wt.%계면활성제/물/cyclohexane 3성분계는 각각 온도에 의존하는 것으로 나타났다. 계면활성제/물/cyclohexane계에서 3상영역은 $27^{\circ}C{\sim}32^{\circ}C$, $36^{\circ}C{\sim}45^{\circ}C$와 $38^{\circ}C{\sim}52^{\circ}C$의 온도 범위에서 각각 나타났으며 3상영역의 부근에서 물 및 오일의 가용화가 최대가 되었다. 계면활성제/물계의 온도변화에 따른 상거동을 살펴 본 결과 1-O-oleoyl glycerol은 hexagonal 액정상이, 2-O-oleoyl-myo-inositol 및 methyl 2-O-oleoyl-${\alpha}$-glucopyranoside는 lamella형태의 액정상이 관찰되었다.

• Molecules and Cells
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• 제38권9호
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• pp.781-788
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• 2015

Mutations of MYO15A are generally known to cause severe to profound hearing loss throughout all frequencies. Here, we found two novel MYO15A mutations, c.3871C>T (p.L1291F) and c.5835T>G (p.Y1945X) in an affected individual carrying congenital profound sensorineural hearing loss (SNHL) through targeted resequencing of 134 known deafness genes. The variant, p.L1291F and p.Y1945X, resided in the myosin motor and IQ2 domains, respectively. The p.L1291F variant was predicted to affect the structure of the actin-binding site from three-dimensional protein modeling, thereby interfering with the correct interaction between actin and myosin. From the literature analysis, mutations in the N-terminal domain were more frequently associated with residual hearing at low frequencies than mutations in the other regions of this gene. Therefore we suggest a hypothetical genotype-phenotype correlation whereby MYO15A mutations that affect domains other than the N-terminal domain, lead to profound SNHL throughout all frequencies and mutations that affect the N-terminal domain, result in residual hearing at low frequencies. This genotype-phenotype correlation suggests that preservation of residual hearing during auditory rehabilitation like cochlear implantation should be intended for those who carry mutations in the N-terminal domain and that individuals with mutations elsewhere in MYO15A require early cochlear implantation to timely initiate speech development.

• Asian-Australasian Journal of Animal Sciences
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• 제28권6호
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• pp.782-787
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• 2015

The myogenic regulatory factors is a family of transcription factors that play a key role in the development of skeletal muscle fibers, which are the main factors to affect the meat taste and texture. In the present study, we performed candidate gene analysis to identify single-nucleotide polymorphisms in the MyoD, Myf5, MyoG, and Mrf4 genes using polymerase chain reaction-single strand conformation polymorphism in 360 Erlang Mountain Chickens from three different housing systems (cage, pen, and free-range). The general linear model procedure was used to estimate the statistical significance of association between combined genotypes and muscle fiber traits of chickens. Two polymorphisms (g.39928301T>G and g.11579368C>T) were detected in the Mrf4 and MyoD gene, respectively. The diameters of thigh and pectoralis muscle fibers were higher in birds with the combined genotypes of GG-TT and TTCT (p<0.05). Moreover, the interaction between housing system and combined genotypes has no significant effect on the traits of muscle fiber (p>0.05). Our findings suggest that the combined genotypes of TT-CT and GG-TT might be advantageous for muscle fiber traits, and could be the potential genetic markers for breeding program in Erlang Mountain Chickens.

• 식물조직배양학회지
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• 제28권3호
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• pp.171-178
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• 2001

딸기의 기내대량번식 체계를 확립하기 위하여 딸기 잎절편체로부터 부정아 형성에 미치는 2,4-D, BAP, 한천, 자당 및 미오이노시톨의 영향에 관하여 실험하였다. 부정아 형성은 MS배지에 0.1 mg/L 2,4-D와 3 mg/L BAP, 0.4% 한천 첨가배지에서 가장 촉진적이었으며, 2,4-D 단독배지에서는 부정아 형성이 유도되지 아니하였다. 0.4% 한천을 첨가한 배지에서 부정아 형성은 1%한천 첨가배지에 비하여 약 3배가 촉진되었으나 투명화 현상이 심하였다. 따라서 딸기 잎 절편을 0.05mg/L 2,4-D, 1 mg/L BAP, 0.8% 한천,30 g/L 자당 및 100mg/L의 미오이노시톨에서 배양하였을 때 건전한 유식물체가 형성되었다.

• 한국동물학회:학술대회논문집
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• 한국동물학회 1998년도 한국생물과학협회 학술발표대회
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• pp.228.2-228
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• 1998

No Abstract, See Full Text