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박태진 ․ 이상민 ․ 조준환 ․ 남수호․ 신주은 ․ 김대환 ․ 이인순 ․ 박흠대 대구대학교 산업기술연구소 2018 産業技術硏究 Vol.29 No.2
Broccoli (Brassica oleracea var. italica) is well known of excellent antioxidant food so that we researched on its anti-aging and callus differentiation from the tissue of broccoli stem by MS medium with exo-phytohormone as IAA, kinetin. The functional materials of callus tissue were extracted with water and 70% EtOH after 16 days progressed callus formation. As a result, callus differentiation and formation were showed higher efficiency at B5 condition medium (0.1 mg/L IAA, 0.4 mg/L). The callus cultured on B3, B6 and B7 medium were observed the reaction of enzymatic browning to callus tissue. The anti-aging effect on the extract was experiment with DPPH radical scavenging and catalase(CAT). The activity of DPPH radical scavenging was showed high value in water extract of B1, B6, B8, B9 and was increased depending on sample concentration. However, in case of the extract of 70% EtOH, DPPH activity was showed different effect depending on the sample kind. CAT activity was found the highest enzyme unit in the extract of natural broccoli and B1(no add exo-phytohormone). It is to express that a broccoli itself is great food for anti-aging. Therefore, callus formation of broccoli considers having no economic efficiency to do development of anti-aging product.
Park, Jin-Mo,Min, Sung-Hun,Hong, Joo-Hee,Lee, E-Nok,Son, Hyeong-Hoon,Park, Hum-Dai,Koo, Deog-Bon The Korean Society of Animal Reproduction 2011 Reproductive & developmental biology Vol.35 No.3
Autophagy is a process of intracellular bulk protein degradation, in which the accumulated proteins and cytoplasmic organelles are degraded. It plays important roles in cellular homeostasis, apoptosis, and development, but its role during early embryo development remains contentious. Therefore, in the present study, we investigated the effects of 3-methyladenine (3-MA) on early embryonic development in pigs, we also investigated several indicators of developmental potential, including mitochondrial distribution, genes expressions (autophagy-, apoptosis- related genes), apoptosis and ER-stress, which are affected by 3-MA. After in vitro maturation and fertilization, presumptive pig embryos were cultured in PZM-3 medium supplemented with 3-MA for 2 days at $39^{\circ}C$ 5% $CO_2$ in air. Developmental competence to the blastocyst stage in the presence of 3-MA was gradually decreased according to increasing concentration. Thus, all further experiments were performed using 2 mM 3-MA. Blastocysts that developed in the 3-MA treated group decreased LC3-II intensity and expressions of autophagy related genes than those of the untreated control, resulting in down-regulates the autophagy. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) showed that the number of containing fragmented DNA at the blastocyst stage increased in the 3-MA treated group compared with control ($6.0{\pm}1.0$ vs $3.3{\pm}0.6$, p<0.05). Also, the expression of the pro-apoptotic gene Bax increased in 3-MA treated group, whereas expression of the anti-apoptotic gene Bcl-XL decreased. Mito Tracker Green FM staining showed that blastocysts derived from the 3-MA treated group had lower mitochondrial integrity than that of the untreated control, resulting in decrease the embryonic qualities of preimplantation porcine blastocysts. Then, the expression of the spliced form of pXBP-1 product (pXBP-1s) increased in 3-MA treated group, resulting increase of ER-stress. Taken together, these results indicate that inhibition of autophagy by 3-MA is closely associated with apoptosis and ER-stress during preimplantation periods of porcine embryos.
Induced Pluripotent Stem Cell Generation using Nonviral Vector
Park, Si-Jun,Shin, Mi-Jung,Seo, Byoung-Boo,Park, Hum-Dai,Yoon, Du-Hak,Ryoo, Zae-Young The Korean Society of Animal Reproduction 2011 Reproductive & developmental biology Vol.35 No.4
Induced pluripotent stem (iPS) cells have been generated from mouse and human somatic cells by etopic expression of transcription factors. iPS cells are indistinguishable from ES cells in terms of morphology and stem cell marker expression. Moreover, mouse iPS cells give rise to chimeric mice that are competent for germline transmission. However, mice derived from iPS cells often develop tumors. Furthermore, the low efficiency of iPS cell generation is a big disadvantage for mechanistic studies. Nonviral plasmid.based vectors are free of many of the drawbacks that constrain viral vectors. The histone deacetylase inhibitor valproic acid (VPA) has been shown to improve the efficiency of mouse and human iPS cell generation, and vitamin C (Vc) accelerates gene expression changes and establishment of the fully reprogrammed state. The MEK inhibitor PD0325901 (Stemgent) has been shown to increase the efficiency of the reprogramming of human primary fibroblasts into iPS cells. In this report, we described the generation of mouse iPS cells devoid of exogenous DNA by the simple transient transfection of a nonviral vector carrying 2A-peptide-linked reprogramming factors. We used VPA, Vc, and the MEK inhibitor PD0325901 to increase the reprogramming efficiency. The reprogrammed somatic cells expressed pluripotency markers and formed EBs.
고추(Capsicum annuum) callis와 검은 무늬병 유발균의 elicitor를 이용한 항균물질 생산
박지은,구부연,박재영,권혜인,문슬아,조은별,이인순,박흠대 대구대학교 산업기술연구소 2014 産業技術硏究 Vol.25 No.1
We described the effects of antimicrobial activity on a produced material by the co-culture of elicitor inducing black rot's disease and callus of Capsicum annuum that callus was induced in MS medium containing IAA 0.5 mg/ml and kinetin 2.0 mg/ml. the bacterial elicitor used 0.5% Yeast extract and the fungal elicitor made by Alternaria alternata KACC 40019 with PDB medium. In order to produce antimicrobial agent protective balck rot's disease, we co-cultured the fungal elicitor or bacterial elicitor with the red pepper callus tissue for 7days repectively. The result, an antifungal agent was produced extracellular materials into medium during co-culture red pepper callus with the fungal elicitor. The antifungal activity of the extracellular material was showed 23% inhibition rate on Alternaria alternata KACC 40019 on the other hand the intracellular material was showed 2% inhibition rate. A chitinase activity of extracellular material was also showed higher value than a chitinase of the intracellular material. The extracellular material produced by elicitor was had more 15% of andtibacterial activity than the intracellular material. As a result, a biotic stress as a fungal elicitor and a bacterial elicitor brings a defence system in the red pepper callus and induce to synthesis in extracellular material an antimicrobial agent as a chitinase and phytoalexin.
Seo, Byoung-Boo,Park, Hum-Dai The Korean Society of Animal Reproduction 2011 Reproductive & developmental biology Vol.35 No.4
Mitochondria diseases have been reported to involve structural and functional defects of complex I-V. Especially, many of these diseases are known to be related to dysfunction of mitochondrial proton-translocating NADH-ubiquinone oxidoreductase (complex I). The dysfunction of mitochondria complex I is associated with neurodegenerative disorders, such as Parkinson's disease, Huntington's disease, and Leber's hereditary optic neuropathy (LHON). Mammalian mitochondrial proton-translocating NADH-quinone oxidoreductase (complex I) is largest and consists of at least 46 different subunits. In contrast, the NDI1 gene of Saccharomyces cerevisiae is a single subunit rotenone-insensitive NADH-quinone oxidoreductase that is located on the matrix side of the inner mitochondrial membrane. The Saccharomyces cerevisiae NDI1 gene using a recombinant adeno-associated virus vector (rAAV-NDI1) was successfully expressed in AML12 mouse liver hepatocytes and the NDI1-transduced cells were able to grow in media containing rotenone. In contrast, control cells that did not receive the NDI1 gene failed to survive. The expressed Ndi1 enzyme was recognized to be localized in mitochondria by confocal immunofluorescence microscopic analyses and immunoblotting. Using digitonin-permeabilized cells, it was shown that the NADH oxidase activity of the NDI1-transduced cells was not affected by rotenone which is inhibitor of complex I, but was inhibited by antimycin A. Furthermore, these results indicate that Ndi1 can be functionally expressed in the AML12 mouse liver hepatocytes. It is conceivable that the NDI1 gene is powerful tool for gene therapy of mitochondrial diseases caused by complex I deficiency. In the future, we will attempt to functionally express the NDI1 gene in mouse embryonic stem (mES) cell.
Shin, Mi-Jung,Park, Seo-Jin,Pare, Hum Dai,Ryoo, Zae-Young The Korean Society of Animal Reproduction 2010 Reproductive & developmental biology Vol.34 No.2
The spontaneous mutant circling mouse (cir/cir) shows a circling behavior and hearing loss. We produced transgenic mice overexpressing transmembrane inner ear (tmie) gene, the causative gene, for the phenotypic rescue of the circling mouse. Through the continuous breeding with circling mice, the cir/cir homozygous mice carrying the transgene (cir/cir-tg) were produced. The rescued cir/cir-tg mice were able to swim in the water with proper orientation and did not show any circling behavior like wild type mice. Western blot and immunohistochemical analysis exhibited that the transgenic tmie was expressed in the inner ear. Inner and outer hair cells were recovered in the cochlea and spiral ganglion neurons were also recovered in the rescued mice. Auditory brainstem response (ABR) test demonstrated that the cir/cir-tg mice are able to respond to sound. This study demonstrates that tmie transgene can recover the hearing impairment and abnormal behavior in the circling mouse.