Hyuk Cheol Kwon;Hyun Su Jung;Do Hyun Kim;Jong Hyeon Han;Seo Gu Han;Dong Hyun Keum;Seong Joon Hong;Sung Gu Han
Animal Bioscience
/
v.36
no.11
/
pp.1757-1768
/
2023
Objective: The number of bovine mammary epithelial cells (BMECs) is closely associated with the quantity of milk production in dairy cows; however, the optimal levels and the combined effects of hormones and essential amino acids (EAAs) on cell proliferation are not completely understood. Thus, the purpose of this study was to determine the optimal combination of individual hormones and EAAs for cell proliferation and related signaling pathways in BMECs. Methods: Immortalized BMECs (MAC-T) were treated with six hormones (insulin, cortisol, progesterone, estrone, 17β-estradiol, and epidermal growth factor) and ten EAAs (arginine, histidine, leucine, isoleucine, threonine, tryptophan, lysine, methionine, phenylalanine, and valine) for 24 h. Results: Cells were cultured in a medium containing 10% fetal bovine serum (FBS) as FBS supplemented at a concentration of 10% to 50% showed a comparable increase in cell proliferation rate. The optimized combination of four hormones (insulin, cortisol, progesterone, and 17β-estradiol) and 20% of a mixture of ten EAAs led to the highest cell proliferation rate, which led to a significant increase in cell cycle progression at the S and G2/M phases, in the protein levels of proliferating cell nuclear antigen and cyclin B1, cell nucleus staining, and in cell numbers. Conclusion: The optimal combination of hormones and EAAs increased BMEC proliferation by enhancing cell cycle progression in the S and G/2M phases. Our findings indicate that optimizing hormone and amino acid levels has the potential to enhance milk production, both in cell culture settings by promoting increased cell numbers, and in dairy cows by regulating feed intake.
The present study investigated the efficient methods to produce in vitro Hanwoo embryos, and to improve the pregnancy rate. The developmental rate, total cell number and ICM ratio of in vitro embryos were compared amongst different culture media. Comparisons were also made on the status of recipients, pregnancy rate along with day of transfer after the estrus. Development of embryos into blastocyst stage in IVMD101 supplemented with 5% fetal bovine serum (FBS) group was significantly higher (34.2%) than that of TCM-199 supplemented with 5% FBS (26.8%) and IVMD101 without FBS (25.9%) (p<0.05). The development rate to blastocyst stage was significantly faster in IVMD101(5% FBS) than that of other groups ($0.2{\sim}2.3%$) (p<0.05). The average number of inner cell mass and trophectoderm were similar among treatment groups, which were $36.0{\sim}44.7$ and $83.3{\sim}106.7$. However, total cell number in IVMD101(5% FBS and 0% FBS) was significantly higher than that of TCM199(5% FBS). There were no differences in the pregnancy rate among treatment groups (32.0%, 33.9% and 28.6%, respectively). However, the pregnancy rate of Day 6 embryos cultured in IVMD101(5% FBS) was significantly (p<0.01) higher than IVMD101 without FBS and TCM-199 + 5% FBS (38.0% vs. 17.2% and 32.4%, respectively). No significant difference was observed for the pregnancy rate between heifer and cow transferred with Day 6 embryos cultured in IVMD 101(5% FBS) (42.7% and 39.3%, respectively). However, there was a significant difference of pregnancy rate (p<0.05) in heifer between one and two embryos transferred (31.4% and 41.9%). There was no difference of pregnancy rate among transfer days after estrus between heifer and cow, but the pregnancy rate of transfer to heifer with day 6 after estrus was significantly higher (p<0.05) than that of day 7 and 8 (22.2% vs. 49.0% and 38.7% respectively). Based on the above findings, there is a possibility to produce in vitro produced embryos cultured in IVMD101(5% FBS) showed higher blastocyst rate and the increased cell number. In terms of the pregnancy rate of in vitro produced embryos, the highest pregnancy rate was observed when two embryos were cultured in IVMD101(5% FBS) and transferred.
Fetal bovine serum (FBS), which contains various nutrients, comprises 20% of the growth medium for cell-cultivated meat. However, ethical, cost, and scientific issues, necesitates identification of alternatives. In this study, we investigated commercially manufactured serum-free media capable of culturing Hanwoo satellite cells (HWSCs) to identify constituent proliferation enhancing factors. Six different serum-free media were selected, and the HWSC proliferation rates in these serum-free media were compared with that of control medium supplemented with 20% FBS. Among the six media, cell proliferation rates were higher only in StemFlexTM Medium (SF) and Mesenchymal Stem Cell Growth Medium DXF (MS) than in the control medium. SF and MS contain high fibroblast growth factor 2 (FGF2) concentrations, and we found upregulated FGF2 protein expression in cells cultured in SF or MS. Activation of the fibroblast growth factor receptor 1 (FGFR1)-mediated signaling pathway and stimulation of muscle satellite cell proliferation-related factors were confirmed by the presence of related biomarkers (FGFR1, FRS2, Raf1, ERK, p38, Pax7, and MyoD) as indicated by quantitative polymerase chain reaction, western blotting, and immunocytochemistry. Moreover, PD173074, an FGFR1 inhibitor suppressed cell proliferation in SF and MS and downregulated related biomarkers (FGFR1, FRS2, Raf1, and ERK). The promotion of cell proliferation in SF and MS was therefore attributed to FGF2, which indicates that FGFR1 activation in muscle satellite cells may be a target for improving the efficiency of cell-cultivated meat production.
Surface-modified poly(L-lactic acid) (PLLA) films and scaffolds were treated with plasma discharge in oxygen gas and subsequently subjected to $in$$situ$ grafting of acrylic acid (AA) in order to increase the cell compatibility. The surface of AA-grafted PLLA was converted to hydroxyapatite (HA)-deposited PLLA in stimulated body fluid (SBF). After the samples were immersed in phosphate-buffered saline (PBS), fetal bovine serum (FBS), normal saline, or cell medium, the water contact angles were significantly reduced on the surface of HA-deposited PLLA. Chondrocyte and osteoblast showed a higher attachment and cell proliferation on HA-deposited surfaces and in particular, it was confirmed that chondrocyte was considerably influenced by HA. However, osteoblast showed better cell proliferation on the surfaces immersed in FBS, cell medium or HA-deposited surface. In addition, the cell proliferation in 3D scaffolds was much higher than that on film type, irrespective of chondrocyte and osteoblast. Therefore, such surface-modified PLLAs are expected to be useful as organic-inorganic hybrid scaffolds in the regeneration of cartilage and bone.
PURPOSE. These days, mesenchymal stem cells (MSCs) have received worldwide attention because of their potentiality in tissue engineering for implant dentistry. The purpose of this study was to evaluate various growth inducing factors in media for improvement of acquisition of bone marrow mesenchymal stem cells (BMMSCs) and colony forming unit-fibroblast (CFU-F). MATERIALS AND METHODS. The mouse BMMSCs were freshly obtained from female C3H mouse femur and tibia. The cells seeded at the density of $10^6$/dish in media supplemented with different density of fetal bovine serum (FBS), $1{\alpha}$, 25-dihydroxyvitamin (VD3) and recombinant human epidermal growth factor (rhEGF). After 14 days, CFU-F assay was conducted to analyze the cell attachment and proliferation, and moreover for VD3, the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay was additionally conducted. RESULTS. The cell proliferation was increased with the increase of FBS concentration (P<.05). The cell proliferation was highest at the density of 20 ng/mL rhEGF compared with 0 ng/mL and 200 ng/mL rhEGF (P<.05). For VD3, although the colony number was increased with the increase of its concentration, the difference was not statistically significant (P>.05). CONCLUTION. FBS played the main role in cell attachment and growth, and the growth factor like rhEGF played the additional effect. However, VD3 did not have much efficacy compare with the other two factors. Improvement of the conditions could be adopted to acquire more functional MSCs to apply into bony defect around implants easily.
The aim of present experiment was to examine hatching rate as in vitro indicator of viability of porcine embryos before early stage embryo transfer such as zygotes or 2-cell stage embryos. Cumulus-oocyte complexes (COCs) collected from ovaries were matured in North Carolina State University 23 (NCSU-23) containing 10% porcine follicular fluid (pFF), 10 ng/ml epidermal growth factor (EGF), $10{\mu}g/ml$ follicle stimulating hormone (FSH), $35{\mu}g/ml$ luteinizing hormone (LH), and 1mg/ml cysteine. After 24 hours, the COCs were transferred to the same medium without hormones. After 65h of maturation, oocytes were exposed to phosphate buffered saline (PBS) with 7% ethanol (v/v) for 7 minutes, and then the oocytes were washed and cultured in tissue culture medium (TCM) 199 containing 5 ug/ml cytochalasin B for 5h at $38.5^{\circ}C$ in an atmosphere of 5% $CO_2$ and 95% air with high humidity. After cytochalasin B treatment, the presumptive parthenotes were cultured in porcine zygote medium (PZM)-5 and cleavage of the parthenotes was assessed at 72h of activation, Normally cleaved parthenotes were cultured for an additional 8 days to evaluate their ability to develop to blastocyst and hatching stages. The fetal bovine serum (FBS) were added at Day 4 or 5 with concentrations of 2.5, 5 or 10%. The blastocyst rates were ranged within $39.1{\sim}70%$ in each treatment. However hatching rate was dramatically decreased in non-addition group. In this experiment, embryo viability in female reproductive tract may be estimated before embryo transfer with in vitro culture adding FBS by hatching ability.
The purpose of this study was to determine toxic effect of sucrose and trehalose prior to cryopreservation on nuclear maturation and embryonic development in immature bovine oocytes. All cryoprotectant was prepared in tissue culture medium 199-HEPES (TCM 199-HEPES) with 10% fetal bovine serum (FBS). Immature oocytes were exposed to 1.2M ethylene glycol (EG) and 0.1M sucrose or 1.2M EG and 0.1M trehalose for 3 min and then were exposed to 3.2 M EG and 0.25 M sucrose or 3.2 M EG and 0.25 M trehalose for 1 min. Oocytes treated with cryoprotectants were exposed to 0.25 M sucrose or 0.25 M trehalose for 5 min and then 0.1 M sucrose or 0.1 M trehalose for 5 min. Depending on type of sugar added to cryopreservation solution, oocytes were allocated to sucrose group and trehalose group, respectively. Oocytes exposed to TCM 199-HEPES with 10% FBS were considered as control. Oocytes were cultured in TCM 199 supplemented with 10% FBS, 5 ng/ml epidermal growth factor, 0.01 IU/ml luteinizing hormone, and $1\;{\mu}g/ml$ estradiol for 24 h in $39^{\circ}C$, 5% $CO_2$. Nuclear maturation was assessed by staining oocytes with 1% aceto-orcein. Oocytes were fertilized in vitro and were cultured in TCM 199 supplemented with 10% FBS, 5 mM sodium pyruvate, and antibiotics in $39^{\circ}C$, 5% $CO_2$. The rates of cleavage and blastocyst, and cell number in blastocyst were assessed. Metaphase II rates were not different among experimental groups regardless of type of sugar. The cleavage rate of trehalose group (73.3%) was significantly higher (p<0.05) than those of sucrose group (62.8%) and control group (60.8%). The blastocyst rate was significantly higher in trehalose group (p<0.05). Mean cell number in blastocyst were not different among experimental groups, although cell number of blastocyst in trehalose group was significantly higher on day 7 (p<0.05). In conclusion, sucrose and trehalose were not toxic to immature bovine oocytes prior to cryopreservation. In particular, trehalose was more effective on embryonic development.
Bovine oocytes with compact and complete cumulus cells were cultured for up to 24h in TCM199 buffered with 25mmol/l hepes and supplemented with 10% FBS (fetal bovine serum), 1mg/ml $17{\beta}$-estradiol, 20IU/ml hCG(human chorionic gonadotropin). All of the oocytes were divided into at 6 groups depending upon incubation times (control, 0 hour, 6 hours, 12 hours, 16 hours, 18 hours). To all experimental media, $200{\mu}g/ml$ puromycin was added at different incubation times mentioned above. Following these culture times, in vitro insemination was conducted with frozen-thawed bovine spermatozoa in medium BO (Brackett and Oliphant medium for in vitro insemination) with $10{\mu}g/ml$ BSA(bovine serum albumin) and 10 mg/ml heparin added. After 22h culture, the oocytes were fixed with acetic alcohol solution and stained with orcein acetic solution to evaluate sperm nuclear progression. Addition of puromycin after 0, 6 and 12 h of culture resulted in near of oocyte maturation at the M1 stage. Contrariwise, puromycin addition after 12 h of culture led to restoration of nuclear progression to M2 stage. On the one hand, puromycin affected the synthesis of Cyclin B protein that may be involved in the oocyte maturation and sperm capacitation for in vitro fertilization. The present study suggests the participation of protein synthesis, cyclin B, in the oocyte development from M1 to M2 stages in vitro.
Human mesenchymal stem cells(hMSC), that have been reported to be present in bone marrow, adipose tissues, dermis, muscles and peripheral blood, have the potential to differentiate along different lineages including those forming bone, cartilage, fat, muscle and neuron. Therefore, hMSC are attractive candidates for cell and gene therapy. The optimal conditions for hMSC expansion require medium supplemented with fetal bovine serum(FBS). Some forms of cell therapy will involve multiple doses, raising a concern over immunological reactions caused by medium-derived FBS proteins. Previously, we have shown that hADSC can be cultured in human serum(HS) during their isolation and expansion, and that they maintain their proliferative capacity and ability for multilineage differentiation and promote engraftment of peripheral blood-derived CD34 cells mobilized from bone marrow in NOD/SCID mice. In this study we determined whether hADSC grown in HS maintain surface markers expression similar with cells grown in FBS during culture expansion and compared gene expression profile by Affymetrix microarray. Flow cytometry analysis showed that HLA-DR, CD117, CD29 and CD44 expression in HS-cultured hADSC during culture expansion were similar with that in FBS-cultured cells. However, the gene expression profile in HS-cultured hADSC was significantly different from that in FBS-cultured cells. Therefore, these data indicated that HS-cultured hADSC should be used in vivo animal study of hADSC transplantation for direct extrapolation of preclinical data into clinical application.
Cell cycle activation and progression in vascular proliferative disease represent potent therapeutic targets. Luteolin, which occurs as glycosylated forms in celery, green pepper, perilla leaf, and camomile tea, has demonstrated antimutagenic, antitumorigenic, antioxidant, and antiinflammatory properties. In this study, we investigated the effect of luteolin on the proliferation of primary cultured rat aortic vascular smooth muscle cells induced by 5% fetal bovine serum. Luteolin at concentrations of 5, 20, and $50{\mu}M$ significantly inhibited this proliferation by 29.6, 50.8, and 83.1%, respectively. The incorporation of $[^3H]$-thymidine into DNA was also inhibited by 25.8, 57.6, and 81.0%, respectively. Flow cytometry analysis of DNA content revealed that FBS-inducible cell cycle progression was blocked by luteolin. Luteolin showed no cytotoxicity in VSMCs in this experimental condition according to WST-1 assays. Luteolin may represent a potential anti-proliferative agent for treatment of angioplasty restenosis and atherosclerosis.
본 웹사이트에 게시된 이메일 주소가 전자우편 수집 프로그램이나
그 밖의 기술적 장치를 이용하여 무단으로 수집되는 것을 거부하며,
이를 위반시 정보통신망법에 의해 형사 처벌됨을 유념하시기 바랍니다.
[게시일 2004년 10월 1일]
이용약관
제 1 장 총칙
제 1 조 (목적)
이 이용약관은 KoreaScience 홈페이지(이하 “당 사이트”)에서 제공하는 인터넷 서비스(이하 '서비스')의 가입조건 및 이용에 관한 제반 사항과 기타 필요한 사항을 구체적으로 규정함을 목적으로 합니다.
제 2 조 (용어의 정의)
① "이용자"라 함은 당 사이트에 접속하여 이 약관에 따라 당 사이트가 제공하는 서비스를 받는 회원 및 비회원을
말합니다.
② "회원"이라 함은 서비스를 이용하기 위하여 당 사이트에 개인정보를 제공하여 아이디(ID)와 비밀번호를 부여
받은 자를 말합니다.
③ "회원 아이디(ID)"라 함은 회원의 식별 및 서비스 이용을 위하여 자신이 선정한 문자 및 숫자의 조합을
말합니다.
④ "비밀번호(패스워드)"라 함은 회원이 자신의 비밀보호를 위하여 선정한 문자 및 숫자의 조합을 말합니다.
제 3 조 (이용약관의 효력 및 변경)
① 이 약관은 당 사이트에 게시하거나 기타의 방법으로 회원에게 공지함으로써 효력이 발생합니다.
② 당 사이트는 이 약관을 개정할 경우에 적용일자 및 개정사유를 명시하여 현행 약관과 함께 당 사이트의
초기화면에 그 적용일자 7일 이전부터 적용일자 전일까지 공지합니다. 다만, 회원에게 불리하게 약관내용을
변경하는 경우에는 최소한 30일 이상의 사전 유예기간을 두고 공지합니다. 이 경우 당 사이트는 개정 전
내용과 개정 후 내용을 명확하게 비교하여 이용자가 알기 쉽도록 표시합니다.
제 4 조(약관 외 준칙)
① 이 약관은 당 사이트가 제공하는 서비스에 관한 이용안내와 함께 적용됩니다.
② 이 약관에 명시되지 아니한 사항은 관계법령의 규정이 적용됩니다.
제 2 장 이용계약의 체결
제 5 조 (이용계약의 성립 등)
① 이용계약은 이용고객이 당 사이트가 정한 약관에 「동의합니다」를 선택하고, 당 사이트가 정한
온라인신청양식을 작성하여 서비스 이용을 신청한 후, 당 사이트가 이를 승낙함으로써 성립합니다.
② 제1항의 승낙은 당 사이트가 제공하는 과학기술정보검색, 맞춤정보, 서지정보 등 다른 서비스의 이용승낙을
포함합니다.
제 6 조 (회원가입)
서비스를 이용하고자 하는 고객은 당 사이트에서 정한 회원가입양식에 개인정보를 기재하여 가입을 하여야 합니다.
제 7 조 (개인정보의 보호 및 사용)
당 사이트는 관계법령이 정하는 바에 따라 회원 등록정보를 포함한 회원의 개인정보를 보호하기 위해 노력합니다. 회원 개인정보의 보호 및 사용에 대해서는 관련법령 및 당 사이트의 개인정보 보호정책이 적용됩니다.
제 8 조 (이용 신청의 승낙과 제한)
① 당 사이트는 제6조의 규정에 의한 이용신청고객에 대하여 서비스 이용을 승낙합니다.
② 당 사이트는 아래사항에 해당하는 경우에 대해서 승낙하지 아니 합니다.
- 이용계약 신청서의 내용을 허위로 기재한 경우
- 기타 규정한 제반사항을 위반하며 신청하는 경우
제 9 조 (회원 ID 부여 및 변경 등)
① 당 사이트는 이용고객에 대하여 약관에 정하는 바에 따라 자신이 선정한 회원 ID를 부여합니다.
② 회원 ID는 원칙적으로 변경이 불가하며 부득이한 사유로 인하여 변경 하고자 하는 경우에는 해당 ID를
해지하고 재가입해야 합니다.
③ 기타 회원 개인정보 관리 및 변경 등에 관한 사항은 서비스별 안내에 정하는 바에 의합니다.
제 3 장 계약 당사자의 의무
제 10 조 (KISTI의 의무)
① 당 사이트는 이용고객이 희망한 서비스 제공 개시일에 특별한 사정이 없는 한 서비스를 이용할 수 있도록
하여야 합니다.
② 당 사이트는 개인정보 보호를 위해 보안시스템을 구축하며 개인정보 보호정책을 공시하고 준수합니다.
③ 당 사이트는 회원으로부터 제기되는 의견이나 불만이 정당하다고 객관적으로 인정될 경우에는 적절한 절차를
거쳐 즉시 처리하여야 합니다. 다만, 즉시 처리가 곤란한 경우는 회원에게 그 사유와 처리일정을 통보하여야
합니다.
제 11 조 (회원의 의무)
① 이용자는 회원가입 신청 또는 회원정보 변경 시 실명으로 모든 사항을 사실에 근거하여 작성하여야 하며,
허위 또는 타인의 정보를 등록할 경우 일체의 권리를 주장할 수 없습니다.
② 당 사이트가 관계법령 및 개인정보 보호정책에 의거하여 그 책임을 지는 경우를 제외하고 회원에게 부여된
ID의 비밀번호 관리소홀, 부정사용에 의하여 발생하는 모든 결과에 대한 책임은 회원에게 있습니다.
③ 회원은 당 사이트 및 제 3자의 지적 재산권을 침해해서는 안 됩니다.
제 4 장 서비스의 이용
제 12 조 (서비스 이용 시간)
① 서비스 이용은 당 사이트의 업무상 또는 기술상 특별한 지장이 없는 한 연중무휴, 1일 24시간 운영을
원칙으로 합니다. 단, 당 사이트는 시스템 정기점검, 증설 및 교체를 위해 당 사이트가 정한 날이나 시간에
서비스를 일시 중단할 수 있으며, 예정되어 있는 작업으로 인한 서비스 일시중단은 당 사이트 홈페이지를
통해 사전에 공지합니다.
② 당 사이트는 서비스를 특정범위로 분할하여 각 범위별로 이용가능시간을 별도로 지정할 수 있습니다. 다만
이 경우 그 내용을 공지합니다.
제 13 조 (홈페이지 저작권)
① NDSL에서 제공하는 모든 저작물의 저작권은 원저작자에게 있으며, KISTI는 복제/배포/전송권을 확보하고
있습니다.
② NDSL에서 제공하는 콘텐츠를 상업적 및 기타 영리목적으로 복제/배포/전송할 경우 사전에 KISTI의 허락을
받아야 합니다.
③ NDSL에서 제공하는 콘텐츠를 보도, 비평, 교육, 연구 등을 위하여 정당한 범위 안에서 공정한 관행에
합치되게 인용할 수 있습니다.
④ NDSL에서 제공하는 콘텐츠를 무단 복제, 전송, 배포 기타 저작권법에 위반되는 방법으로 이용할 경우
저작권법 제136조에 따라 5년 이하의 징역 또는 5천만 원 이하의 벌금에 처해질 수 있습니다.
제 14 조 (유료서비스)
① 당 사이트 및 협력기관이 정한 유료서비스(원문복사 등)는 별도로 정해진 바에 따르며, 변경사항은 시행 전에
당 사이트 홈페이지를 통하여 회원에게 공지합니다.
② 유료서비스를 이용하려는 회원은 정해진 요금체계에 따라 요금을 납부해야 합니다.
제 5 장 계약 해지 및 이용 제한
제 15 조 (계약 해지)
회원이 이용계약을 해지하고자 하는 때에는 [가입해지] 메뉴를 이용해 직접 해지해야 합니다.
제 16 조 (서비스 이용제한)
① 당 사이트는 회원이 서비스 이용내용에 있어서 본 약관 제 11조 내용을 위반하거나, 다음 각 호에 해당하는
경우 서비스 이용을 제한할 수 있습니다.
- 2년 이상 서비스를 이용한 적이 없는 경우
- 기타 정상적인 서비스 운영에 방해가 될 경우
② 상기 이용제한 규정에 따라 서비스를 이용하는 회원에게 서비스 이용에 대하여 별도 공지 없이 서비스 이용의
일시정지, 이용계약 해지 할 수 있습니다.
제 17 조 (전자우편주소 수집 금지)
회원은 전자우편주소 추출기 등을 이용하여 전자우편주소를 수집 또는 제3자에게 제공할 수 없습니다.
제 6 장 손해배상 및 기타사항
제 18 조 (손해배상)
당 사이트는 무료로 제공되는 서비스와 관련하여 회원에게 어떠한 손해가 발생하더라도 당 사이트가 고의 또는 과실로 인한 손해발생을 제외하고는 이에 대하여 책임을 부담하지 아니합니다.
제 19 조 (관할 법원)
서비스 이용으로 발생한 분쟁에 대해 소송이 제기되는 경우 민사 소송법상의 관할 법원에 제기합니다.
[부 칙]
1. (시행일) 이 약관은 2016년 9월 5일부터 적용되며, 종전 약관은 본 약관으로 대체되며, 개정된 약관의 적용일 이전 가입자도 개정된 약관의 적용을 받습니다.