• Biomolecules & Therapeutics
• /
• 제31권3호
• /
• pp.264-275
• /
• 2023

Parkinson's disease (PD) is a common neurodegenerative disorder characterized by tremors, bradykinesia, and rigidity. PD is caused by loss of dopaminergic (DA) neurons in the midbrain substantia nigra (SN) and therefore, replenishment of DA neurons via stem cell-based therapy is a potential treatment option. Astrocytes are the most abundant non-neuronal cells in the central nervous system and are promising candidates for reprogramming into neuronal cells because they share a common origin with neurons. The ability of neural progenitor cells (NPCs) to proliferate and differentiate may overcome the limitations of the reduced viability and function of transplanted cells after cell replacement therapy. Achaete-scute complex homolog-like 1 (Ascl1) is a well-known neuronal-specific factor that induces various cell types such as human and mouse astrocytes and fibroblasts to differentiate into neurons. Nurr1 is involved in the differentiation and maintenance of DA neurons, and decreased Nurr1 expression is known to be a major risk factor for PD. Previous studies have shown that direct conversion of astrocytes into DA neurons and NPCs can be induced by overexpression of Ascl1 and Nurr1 and additional transcription factors genes such as superoxide dismutase 1 and SRY-box 2. Here, we demonstrate that astrocytes isolated from the ventral midbrain, the origin of SN DA neurons, can be effectively converted into DA neurons and NPCs with enhanced viability. In addition, when these NPCs are inducted to differentiate, they exhibit key characteristics of DA neurons. Thus, direct conversion of midbrain astrocytes is a possible cell therapy strategy to treat neurodegenerative diseases.

• 생명과학회지
• /
• 제33권9호
• /
• pp.736-745
• /
• 2023

노화란 세포 및 생리 기능이 점진적으로 손상되는 복잡한 과정이다. 알츠하이머, 동맥경화 및 갱년기와 같은 노화와 관련된 질병은 노화가 진행이 되면서 발생된다. 노화와 관련된 질환은 다양한 원인에 의해 발생된다. 그 중 유전적인 변화 없이 유전자 발현을 조절하는 후성유전의 변화는 노화, 그리고 노화와 관련된 질환의 발생에 중요한 조절자로 알려져있다. 이 리뷰에서는 후성유전의 변화가 노화 및 노화와 관련된 질병의 발전과 진행에 어떠한 역할을 하는지에 대해 서술하였다. 노화 중에 일어나는 유전적 변화의 분자적 기전과 이러한 변화가 노화와 관련된 질병에 미치는 영향, 특히 노화와 관련된 질환과 관련된 유전자 발현 양식을 조절하는 RNA 메틸화, DNA 메틸화 및 miRNA에 대해 중점적으로 초점을 맞추었다.

• 한국발생생물학회:학술대회논문집
• /
• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
• /
• pp.82-82
• /
• 2003

Offspring have been produced from somatic cells in a number of species. This biotechnology introduced a new phenomenon in reprogramming and differentiation of somatic cell, namely totipotency. However, birth of oversized calves and perinatal abnormalities such as increased gestation length, lack of spontaneous parturition, higher incidence of dystocia, and reduced perinatal viability of offspring are frequently observed in pregnancies of cloned bovine fetuses. Disturbance of feto-placenta has been proposed as likely causes for abnomal growth. However. Little is known the mechanism responsible for the perinatal problems. Therefore, we focused on gestation length in somatic cell nuclear recipient cows. To solve this issues, placental tissues of control and cloned bovine were obtained by a cesarean section (C-section) before 5 days of paturition. Total RNA from control and cloned bovine placenta was extractd by TRIzol reagent. GeneFishing DEG kits (Seegene) were used to identify differentially expression genes. Total RNA (3 ug) were synthesized by M-MLV reverse transcriptase (200 u/ul) with 10 uM dT-annealing control primer (ACP1) at 42C for 90 min. Then, first-strand cDNA (50 ng) was amplified using the 5 uM arbitary ACP (1-20) and 10 uM dT-ACP2 primers. Some specific expression genes were amplified, Now, we are cloning and sequencing. These finding strongly can be support to solve the problems for parturition delay in nuclear transfer cows, suggest that placenta specific proteins are key indicators for the aberration of gestation and placental function in cows.

• 한국동물생명공학회지
• /
• 제38권3호
• /
• pp.109-120
• /
• 2023

Background: Pluripotent stem cells (PSCs) including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) offer the immense therapeutic potential in stem cell-based therapy of degenerative disorders. However, clinical trials of human ESCs cause heavy ethical concerns. With the derivation of iPSCs established by reprogramming from adult somatic cells through the transgenic expression of transcription factors, this problems would be able to overcome. In the present study, we tried to differentiate porcine iPSCs (piPSCs) into endothelial cells (ECs) for stem cell-based therapy of vascular diseases. Methods: piPSCs (OSKMNL) were induced to differentiation into ECs in four differentiation media (APEL-2, APEL-2 + 50 ng/mL of VEGF, EBM-2, EBM-2 + 50 ng/mL of VEGF) on cultured plates coated with matrigel^® (1:40 dilution with DMEM/F-12 medium) for 8 days. Differentiation efficiency of these cells were exanimated using qRT-PCR, Immunocytochemistry, Western blotting and FACS. Results: As results, expressions of pluripotency-associated markers (OCT-3/4, SOX2 and NANOG) were higher observed in all porcine differentiated cells derived from piPSCs (OSKMNL) cultured in four differentiation media than piPSCs as the control, whereas endothelial-associated marker (CD-31) in the differentiated cells was not expressed. Conclusions: It can be seen that piPSCs (OSKMNL) were not suitable to differentiate into ECs in the four differentiation media unlike porcine epiblast stem cells (pEpiSCs). Therefore, it would be required to establish a suitable PSCs for differentiating into ECs for the treatment of cardiovascular diseases.

• IMMUNE NETWORK
• /
• 제23권4호
• /
• pp.28.1-28.20
• /
• 2023

Lipid accumulation in macrophages is a prominent phenomenon observed in atherosclerosis. Previously, intimal foamy macrophages (FM) showed decreased inflammatory gene expression compared to intimal non-foamy macrophages (NFM). Since reprogramming of lipid metabolism in macrophages affects immunological functions, lipid profiling of intimal macrophages appears to be important for understanding the phenotypic changes of macrophages in atherosclerotic lesions. While lipidomic analysis has been performed in atherosclerotic aortic tissues and cultured macrophages, direct lipid profiling has not been performed in primary aortic macrophages from atherosclerotic aortas. We utilized nanoflow ultrahigh-performance liquid chromatography-tandem mass spectrometry to provide comprehensive lipid profiles of intimal non-foamy and foamy macrophages and adventitial macrophages from Ldlr^-/- mouse aortas. We also analyzed the gene expression of each macrophage type related to lipid metabolism. FM showed increased levels of fatty acids, cholesterol esters, phosphatidylcholine, lysophosphatidylcholine, phosphatidylinositol, and sphingomyelin. However, phosphatidylethanolamine, phosphatidic acid, and ceramide levels were decreased in FM compared to those in NFM. Interestingly, FM showed decreased triacylglycerol (TG) levels. Expressions of lipolysis-related genes including Pnpla2 and Lpl were markedly increased but expressions of Lpin2 and Dgat1 related to TG synthesis were decreased in FM. Analysis of transcriptome and lipidome data revealed differences in the regulation of each lipid metabolic pathway in aortic macrophages. These comprehensive lipidomic data could clarify the phenotypes of macrophages in the atherosclerotic aorta.

• 한국간담췌외과학회지
• /
• 제27권4호
• /
• pp.342-349
• /
• 2023

Backgrounds/Aims: Liver organoids have emerged as a powerful tool for studying liver biology and disease and for developing new therapies and regenerative medicine approaches. For organoid culture, Matrigel, a type of extracellular matrix, is the most commonly used material. However, Matrigel cannot be used for clinical applications due to the presence of unknown proteins that can cause immune rejection, batch-to-batch variability, and angiogenesis. Methods: To obtain human primary hepatocytes (hPHs), we performed 2 steps collagenase liver perfusion protocol. We treated three small molecules cocktails (A83-01, CHIR99021, and HGF) for reprogramming the hPHs into human chemically derived hepatic progenitors (hCdHs) and used hCdHs to generate liver organoids. Results: In this study, we report the generation of liver organoids in a collagen scaffold using hCdHs. In comparison with adult liver (or primary hepatocyte)-derived organoids with collagen scaffold (hALO_C), hCdH-derived organoids in a collagen scaffold (hCdHO_C) showed a 10-fold increase in organoid generation efficiency with higher expression of liver- or liver progenitor-specific markers. Moreover, we demonstrated that hCdHO_C could differentiate into hepatic organoids (hCdHO_C_DM), indicating the potential of these organoids as a platform for drug screening. Conclusions: Overall, our study highlights the potential of hCdHO_C as a tool for liver research and presents a new approach for generating liver organoids using hCdHs with a collagen scaffold.

• 한국가금학회지
• /
• 제49권3호
• /
• pp.145-156
• /
• 2022

본 연구는 한국 토종닭을 대상으로 부계의 텔로미어 길이가 수정 능력 및 배아 발생 능력에 미치는 영향을 알아보고, 부계의 텔로미어 길이와 자식의 생산능력 및 자식의 텔로미어 길이와의 연관성을 살펴보고자 하였다. 시험에 공시된 닭은 토종 종계 수컷 22수와 이들로부터 생산된 자식 329수를 대상으로 하였다. 부계의 번식 능력으로는 수정율, 배자발육중지율 및 부화율을 조사하였고, 자손의 생산 능력으로는 생존율, 체중 및 증체량을 분석하였다. 텔로미어 길이는 34주령 부계와 12주령 자식의 백혈구 세포 내 telomeric DNA 함량을 양적 형광접합보인법으로 분석하였다. 분석 결과, 부계의 텔로미어 길이에 따른 수정율, 배자발육중지율 및 부화율의 차이는 없는 것으로 나타났고, 부계의 텔로미어 길이와 수정률 및 배자발육중지율 간에는 낮은 부(-)의 상관을, 부화율과는 낮은 정(+)의 상관을 보이나 이들의 상관계수는 유의하지 않은 것으로 나타났다(P>0.05). 부계의 텔로미어 길이와 자식의 생존율 간의 상관 계수는 0.17로 유의하지 않은 낮은 정의 상관이 추정되었다(P>0.05). 부계의 텔로미어 길이에 따른 자식의 성장 능력 분석에서 부계 텔로미어 길이가 긴 집단의 자식들이 상대적으로 낮은 성장 능력을 보이고 이들 간의 상관은 거의 모든 주령에서 유의한 부의 상관 계수가 추정되었다(P<0.05). 부계의 텔로미어 길이와 자식의 텔로미어 길이 간의 상관 계수는 0.075로 추정되어 부계와 자식 간 텔로미어 길이의 연관성이 없는 것으로 나타났다. 이상의 결과로부터 부계의 텔로미어 길이와 번식 능력, 자식의 초기 생존율 및 자식의 텔로미어 길이 간에는 거의 연관성이 없는 것으로 보여지고, 부계의 텔로미어 길이와 자식의 성장 능력 간에는 부의 상관이 있는 것으로 생각된다. 이는 배아 초기 단계에서 텔로미어의 reprogramming에 의해 발생 시 모든 개체의 텔로미어 길이가 재 신장되고 이후 연령이 증가함에 따라 감축의 정도가 달라지기 때문에 나타난 결과로 사료된다.

• Reproductive and Developmental Biology
• /
• 제36권4호
• /
• pp.261-267
• /
• 2012

The technique of SCNT is now well established but still remains inefficient. The in vitro development of SCNT embryos is dependent upon numerous factors including the recipient cytoplast and karyoplast. Above all, the metaphase of the second meiotic division (MII) oocytes have typically become the recipient of choice. Generally high level of MPF present in MII oocytes induces the transferred nucleus to enter mitotic division precociously and causes NEBD and PCC, which may be the critical role for nuclear reprogramming. In the present study we investigated the in vitro development and pregnancy of White-Hanwoo SCNT embryos treated with caffeine (a protein kinase phosphatase inhibitor). As results, the treatment of 10 mM caffeine for 6 h significantly increased MPF activity in bovine oocytes but does not affect the developmental competence to the blastocyst stage in bovine SCNT embryos. However, a significant increase in the mean cell number of blastocysts and the frequency of pregnant on 150 days of White-Hanwoo SCNT embryos produced using caffeine treated cytoplasts was observed. These results indicated that the recipient cytoplast treated with caffeine for a short period prior to reconstruction of SCNT embryos is able to increase the frequency of pregnancy in cow.

• 한국수정란이식학회지
• /
• 제26권1호
• /
• pp.79-84
• /
• 2011

This study was performed to identify the differentially methylated region (DMR) and to examine the mRNA expression of the imprinted H19 gene in day 35 of SCNT pig fetuses. The fetus and placenta at day 35 of gestation fetuses after natural mating (Control) or of cloned pig by somatic cell nuclear transfer (SCNT) were isolated from a uterus. To investigate the mRNA expression and methylation patterns of H19 gene, tissues from fetal liver and placenta including endometrial and extraembryonic tissues were collected. The mRNA expression was evaluated by real-time PCR and methylation pattern was analyzed by bisulfite sequencing method. Bisulfite analyses demonstrated that the differentially methylated region (DMR) was located between -1694 bp to -1338 bp upstream from translation start site of the H19 gene. H19 DMR (-1694 bp to -1338 bp) exhibits a normal mono allelic methylation pattern, and heavily methylated in sperm, but not in oocyte. In contrast to these finding, the analysis of the endometrium and/or extraembryonic tissues from SCNT embryos revealed a complex methylation pattern. The DNA methylation status of DMR Region In porcine H19 gene upstream was hypo methylated in SCNT tissues but hypermethylated in control tissues. Furthermore, the mRNA expression of H19 gene in liver, endometrium, and extraembryonic tissues was significantly higher in SCNT than those of control (p<0.05). These results suggest that the aberrant mRNA expression and the abnormal methylation pattern of imprinted H19 gene might be closely related to the inadequate fetal development of a cloned fetus, contributing to the low efficiency of genomic reprogramming.

• 한국정보과학회논문지:정보통신
• /
• 제35권3호
• /
• pp.215-226
• /
• 2008

센서네트워크가 다양하고 동적인 영역으로 발전해감에 따라 임무 갱신과 같은 기능을 위해 신뢰성 있는 멀티캐스팅 기술이 새롭게 요구되고 있다. 기존의 연구들은 NAK 기반 방식을 사용하고 있지만, 마지막 패킷 문제 등을 안고 있다. 본 논문은 묵시적 ACK와 간접 복구를 이용하는 ACK 기반 오류 제어 기법인 RM2I를 제안한다. 묵시적 ACK는, 송신 노드로부터 패킷을 받은 수신 노드가 다음 노드로 포워딩할 때 송신 노드도 이를 간접적으로 받게 되는데, 이 수신 내용을 ACK로 해석하는 것을 말한다. 간접 복구는 어떤 노드가 상위 노드가 아닌 이웃노드로부터 패킷을 간접적으로 받았을 때 이를 상위 노드로부터 받은 것으로 해석하여 오류 복구에 활용하는 것을 말한다. NS-2 시뮬레이터를 이용하여 다양한 환경 및 인자에서 RM2I의 에너지 성능을 분석 및 검증하였다. 실험 결과, 묵시적 ACK는 ACK의 수를 줄여 제어 부하를 감소시키고, 간접 복구는 재전송 횟수를 줄여 데이타 전송 부하를 감소시켰다. 또한, NAK 기반 오류 복구 기법의 이론적 에너지 성능 상한선을 계산하여 이와 비교하였다. 비교 결과, 에지 노드에서의 부하를 제외하면 어떤 NAK 기반 기법과도 견줄 수 있는 에너지 효율성을 제공한다는 것을 알 수 있었다.