• Clinical and Experimental Vaccine Research
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• 제12권4호
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• pp.328-336
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• 2023

Purpose: Human peripheral blood mononuclear cell (PBMC)-based in vitro systems can be of great value in the development and assessment of vaccines but require the right medium for optimal performance of the different cell types present. Here, we compare three commonly used media for their capacity to support innate and adaptive immune responses evoked in PBMCs by Toll-like receptor (TLR) ligands and whole inactivated virus (WIV) influenza vaccine. Materials and Methods: Human PBMCs were cultured for different periods of time in Roswell Park Memorial Institute (RPMI), Dulbecco's minimal essential medium (DMEM), or Iscove's modified DMEM (IMDM) supplemented with 10% fetal calf serum. The viability of the cells was monitored and their responses to TLR ligands and WIV were assessed. Results: With increasing days of incubation, the viability of PBMCs cultured in RPMI or IMDM was slightly higher than that of cells cultured in DMEM. Upon exposure of the PBMCs to TLR ligands and WIV, RPMI was superior to the other two media in terms of supporting the expression of genes related to innate immunity, such as the TLR adaptor protein gene MyD88 (myeloid differentiation factor 88), the interferon (IFN)-stimulated genes MxA (myxovirus resistance protein 1) and ISG56 (interferon-stimulated gene 56), and the leukocyte recruitment chemokine gene MCP1 (monocyte chemoattractant protein-1). RPMI also performed best with regard to the activation of antigen-presenting cells. As for adaptive immunity, when stimulated with WIV, PBMCs cultured in RPMI or IMDM contained higher numbers of IFNγ-producing T cells and secreted more immunoglobulin G than PBMCs cultured in DMEM. Conclusion: Taken together, among the different media assessed, RPMI was identified as the optimal medium for a human PBMC-based in vitro vaccine evaluation system.

• Asian-Australasian Journal of Animal Sciences
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• 제23권1호
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• pp.25-32
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• 2010

The present study aimed to examine the effects of ${\gamma}$-linolenic acid (GLA) supplementation to in vitro culture (IVC) medium on in vitro developmental competence, freezability and morphology of in vitro matured and fertilized bovine embryos. In vitro produced (IVP) bovine zygotes were cultured in IVC medium supplemented with 0 (negative control), 15, 31, 62, 125, 250, 500 or 1,000 ppm GLA, 250 ppm linoleic acid albumin (LAA) and without any supplement as a control. Day 6 blastocysts derived from culture control were cultured in IVC medium containing either 62, 250 GLA or 250 LAA for 24 h, and at Day 7 were subjected to freezing or morphological examination by electron microscope. GLA 15 showed a tendency to have a higher cleavage rate at Day 2 (70.3%) than other groups. The hatching rate at Day 9 in LAA (38.2%) was significantly higher than the control and all treatment groups (p<0.05), while the blastocyst rate in LAA (32.4%) did not differ from those of 15 (30.5%), 31 (27.1%), and 62 GLA (33.1%) or the control (35.1%). GLA in concentrations of 125, 250, 500, and 1,000 ppm had significantly detrimental effect on the blastocyst rate compared to 15, 31 and 62 ppm GLA, LAA, and control groups (p<0.05). In contrast, the highest post-thaw survival rate (100%) was observed in the control group (p<0.01). Large lipid droplets were observed in the cytoplasm of trophoblastic cells, even in the control, but were abundant in GLA groups. Taking the results of the study into consideration, the addition of GLA to the culture medium for IVP bovine embryos at the dose of 15 ppm increased the developmental competence of zygotes and enhanced the cleavage rate up to Day 2. However, blastulation rate and post-thaw survival were not increased when GLA was added to the culture media.

• 한국수정란이식학회지
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• 제15권2호
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• pp.191-196
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• 2000

the present study was carried out to develop a completely defined culture system and determine if high NaCl concentrations in defined (PVA added) or semi-defined (BSA added) medium is toxic to bovine embryos. Oocytes from slaughterhouse ovaries were matured and fertilized in vitro. After 30 h of insemination, only 2-cell stage embryos were selected and cultured for this experiment. The culture media used were as follows : TLP(114 mM of NaCl) + BSA (3 mg/ml), TLP + PVA (1 mg/ml), mTLP(96 mM of NaCl) + BSA, mTLP + PVA. Six to ten embryos were placed into a 30$\mu$1 drop of each medium and the embryos were examined at 10 day post-insemination without medium renewal. The experiment was replicated 4 times. All data were analyzed by chi-square. There were no significant differences among TLP-BSA, mTLP-BSA and mTLP-PVA in blastocyst development (21.6, 17.2 and 20.2%), respectively. Also, no differences were obtained in hatching rates (11.7, 9.9 and 12.2%), respecitively. However, there were significant differences between TLP-PVA (1.7% and 0.6%) and other group in blastocyst formation and hatching rates, respectively (p<0.01). Development of in vitro produced embryos cultured in BSA containing medium was not affected by high NaCl concentration, but in the completely defined medium, embryonic development was highly affected by NaCl. This study shows that reduced NaCl concentration in completely defined medium is beneficial for development of bovine pre-implantation embryos in vitro.

• Journal of Plant Biotechnology
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• 제4권1호
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• pp.23-27
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• 2002

To establish an in vitro mass production system of grape anthocyanin pigments through callus and cell suspension culture, the effects of nitrogen source and sucrose on fresh weight and anthocyanin production in cell suspension culture of 'Sheridan' grape level were studied. When the medium was devoid of $NO_3^-$, cell fresh weight was either remained stable (1% sucrose) or slightly decreased with culture time (2,3, and 4% sucrose). When $NH_4^-$ was lacking, 3% sucrose was most favorable for cell growth. When $NH_4^-$ was supplied as N source, the anthocyanin content of 2% sucrose containing medium was maintained 2 times higher than other levels till day 8 in culture, then that of 3 and 4% sucrose which peaked at day 12 thereafter. The anthocyanin content was low than $NO_3^-$-free media. Total anthocyanin content in $NH_4^-$-free medium was just about a half of that of $NH_4^+$ medium. Anthocyanin production of 2% sucrose in $NH_4^+$ medium was maintained about 3-fold till day 8, then decreased thereafter. In $NH_4^+$ medium, pH decreased gradually with final pH of 3.5 to 4.0, while pH in $NH_4^+$-free medium increased with final pH of 6.5 to 7.5.

• 생명과학회지
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• 제13권5호
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• pp.732-739
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• 2003

본 연구는 난구세포가 제거된 돼지 미성숙 난자와 포유동물 정자의 공배양이 체외핵성숙에 미치는 영향을 조사하기 위하여 실시하였다. 난구세포가 부착된 미성숙난자를 직경 3-5 mm난포로부터 채취하여 난구세포를 제거한 후 정자가 첨가된 tissue culture medium 199에서 배양하였다. 배양 후 48시간에 난자의 핵성숙을 핵막붕괴, 제일차 감수분열 중기, 제일차 감수분열 후기-말기, 제이차감수분열 중기로 판정하였다. 배양 후 제이차감수분열 중기로 성숙한 난자의 비율이 무첨가구에 비하여 정자가 첨가된 배양액에서 성숙시킨 미성숙난자에서 유의적인(P<0.01) 증가가 있었다$(31.9\pm1.8%\; vs\; 14.9\pm1.0%)$. 난자의 성숙 단계에 따른 정자의 노출시간과 정자가 유래한 종에 따라서는 차이가 없었다. 그러나 정자의 첨가농도에 따라 유의적인 차이가 인정되었다. 본 연구는 포유동물의 정자는 미성숙난자의 체외성숙을 촉진하는 물질을 갖고 있으며, 촉진 효과는 농도에 의존하는 것으로 암시하고 있다.

• Plant Resources
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• 제7권3호
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• pp.222-225
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• 2004

In Japan, propagation of cyclamen is mainly from seedlings. However, seeds are expensive and germination is slow and non..uniform. Therefore, to achieve genetically uniform propagation, multiplication must be vegetative. The rooted tuberous stems without shoots as sources of explants were cultured on the media containing BA and sucrose. After 30 days cultivation, tuberous roots were produced from the root ends attached to a tuberous stem and its capability was dependent on the type of media. The highest percentage of tuberous root formation was observed in Culture on the medium of 1/3 MS containing 0.05mgL$^{-1}$ NAA, 0.5mg L$^{-1}$ BA and 5% sucrose. Growth rates of the tuberous roots were greatly influenced by the cutting positions of a root in explants. The highest growth of was observed if small amount of root end was cut at initiation of tissue culture.

• 식물조직배양학회지
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• 제22권3호
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• pp.131-135
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• 1995

잡종 사기나무의 기내 대량증식을 목적으로 줄기절간 유래의 기내 및 기외시료를 이용하여 다경줄기 유도에 관여하는 식물생장 조절물질의 효과에 대하여 조사하였다. 기외시료의 측아를 제거한 다음 20에서 30$\mu$M 의 zeatin이 함유된 WPM 배지에 6주간 배양하였을때 각각 11개와 13개의 다경줄기가 유도되었다. 그러나 기외시료의 측아가 붙어있는 상태에서 기내배양하여 2주후에 측아를 제거하고 동일 배지에 배양하였을 경우에는 BA가 1.0에서 2.0 $\mu$M 함유된 배지에서도 13개와 15개의 다경줄기를 유도할 수 있었으며, 특히 이때 생산된 줄기의 상태는 아주 건전한 것으로 나타났다. 기내줄기를 사용하였을때는 20 $\mu$M의 zeatin을 처리하였을때 가장 많은 줄기를 생산할 수 있었으며, 생산된 조직배양묘는 일정기간 순화를 시킨 후 온실이나 포지에 식재가 가능하였다.

• 농업과학연구
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• 제12권2호
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• pp.163-165
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• 1985

식물병원균(植物病原菌)에 대(對)해서 항균작용(抗菌作用)을 갖고 물에 잘 녹지 않는 지방산(脂肪酸)의 항균작용(抗菌作用)을 검정(檢定)하는데 있어서 공시균(供試菌)의 균사생장(菌絲生長)이나 혹은 포자발아(胞子發芽)를 위한 배지중(培地中)에 검정(檢定)하기 위한 지방산(脂肪酸)을 고루 확산(擴散)시킨다는것이 필요(必要)하다. 이와같은 실험안내(實驗室內)의 실험(實驗)에 있어서 하나의 확산제(擴散劑)로서 물과 잘 혼합(混合)하며 지방(脂肪)이나 유지(油脂)나 또는 왁스 등에 용제(溶劑)로서 쓰이고 있는 아세톤을 선발했다. 공시균(供試菌)에 대(對)해 배지중(培地中)에 첨가(添加)하여 실제로 사용(使用)할 수 있는 아세톤의 농도(濃度)를 찾기 위한 이 연구(硏究)에서 10%의 아세톤을 함유(含有)하는 배지(培地)위에서 각각(各各)의 공시균(供試菌)의 균사생장(菌絲生長)이나 포자발아(胞子發芽)가 크게 저해(沮害)된 반면에 5%의 아세톤을 함유(含有)하는 배지(培地)에서는 약간 저해(沮害)되었다. 5%아세톤을 함유(含有)하는 배지(培地)에서 균(菌)이 영향(影響)을 받는다 해도 5%아세톤을 함유(含有)하는 배지(培地)의 이용(利用)이 식물병원균(植物病原菌)에 대(對)한 지방산(脂肪酸)의 항균작용(抗菌作用)을 검정(檢定)하는데 있어서 오류(誤謬)를 범할 것으로는 생각되지 않는다.

• Parasites, Hosts and Diseases
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• 제42권1호
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• pp.27-34
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• 2004

We investigated the optimal culture conditions for Cryptosporidium muris in a human stomach adenocarcinoma (AGS) cell line by determining the effects of medium pH and of selected supplements on the development of C. muris. The optimum pH of the culture medium required for the development of C. muris was determined to be 6.6. The number of parasites significantly increased during cultivation for 72 hr (p < 0.05) at this level. On the other hand, numbers decreased linearly after 24 hr of incubation at pH 7.5. When cultured in different concentrations of serum, C. muris in media containing 5% FBS induced 4-7 times more parasites than in 1% or 10% serum. Of the six medium supplements examined, only 1 mM pyruvate enhanced the number of C. muris in vitro. Transmission electron microscopic observation showed the developmental stages of C. muris in the cytoplasm of the cells, not in an extracytoplasmic location. The growth of C. muris in AGS cells provides a means of investigating its biological characteristics and of testing its response to therapeutic agents. However, a more optimized culture system is needed for the recovery of oocysts on a large scale in vitro.

• 한국식물학회:학술대회논문집
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• 한국식물학회 1995년도 식물학심포지움 식물로부터 유용 2차대사산물의 생산 PRODUCTION OF USEFUL SECONDARY METABOLITES FROM PLANTS
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• pp.40-56
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• 1995

During the past two decades, China has seen her great progress in plant biotechnology. Since the Chinese market of herb medicine is huge, while the plant resources are shrinking, particular emphasis has been placed in plant tissue and cell cultures of medicinal plants, this includes fast propagation, protoplast isolation and regeneration, cell suspension cultures and large scale fermentation. To optimize culture conditions for producing secondary compounds in vitro, various media, additives and elicitors have been tested. Successful examples of large scale culture for the secondary metabolite biosynthesis are quite limited : Lithospermum ery throrhizon and Arnebia euchroma for shikonin derivatives, Panax ginseng, P. notoginseng, P. quinquefolium for saponins, and a few other medicinal plants. Recent development of genetic transformation systems of plant cells offered a new approach to in vitro production of secondary compounds. Hairy root induction and cultures, by using Ri-plasmid, have been reported from a number of medicinal plant species, such as Artemisia annua that produces little artemisinin in normal cultured cells, and from Glycyrrhiza uralensis. In the coming five years, Chinese scientists will continue their work on large scale cell cultures of a few of selected plant species, including Taxus spp. and A. annua, for the production of secondary metabolites with medicinal interests, one or two groups of scientists will be engaged in molecular cloning of the key enzymes in plant secondary metabolism.