• Radiation Oncology Journal
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• 제16권4호
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• pp.377-388
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• 1998

목적 : 방사선조사에 의한 apoptosis에서 나타나는 각종 세포주기관련 유전자들의 발현 양상을 RNA와 단백 수준에서 분석하여 방사선조사에 의한 apoptosis에서의 세포주기 조절의 변화를 규명함으로서 방사선치료의 기전에 대한 분자생물학적 이해를 도모하고자 본 연구를 시행하였다. 대상 및 방법 : promyelocytic leukemia 세포주인 HL60 세포주를 배양하여 선형가속기(6MV X-선)로 세포에 8Gy의 방사선을 조사하였다. 조사후 다양한 시간 간격으로 Apoptotic DNA Fragmentation Assay법을 이용하여 apoptosis를 확인하고 동시에 세포주기관련 유전자들(cyclinA, cyclin B, cyclin C, cyclin Dl, cyclin E, cdc2, CDK2, CDK4, $p16^{INK4a}$, $p21^{WAF1}$, $p27^{KIP1}$, E2F, PCNA와 Rb)을 단백질과 RNA 수준에서 분석하기위해 western blot analysis와 반정량적 RT-PCR을 시행하였다. 결과: 8 Gy의 방사선 조사에 의해 HL60세포에서 apoptosis가 관찰 되었다. 방사선 조사군에서 cyclin A단백은 조사후 48시간까지 시간이 갈수록 증가하였으며, cyclin E, E2F, CDK2 및 Rb 단백은 증가되었다가 다시 감소를 보였다. Rb단백의 증가는 대부분 비활성형인 ppRb (phosphorylated Rb protein)의 양적변화에 의한 것이었다. cyclin Dl, PCNA, COC2, CDK4, $p16^{INK4a}$단백은 발현의 차이를 보이지 않았으며 $p21^{WAF1}$과 $p27^{KIP1}$ 단백은 검출되지 않았다. cyclin A, B, C mRNA는 방사선 조사 직후 감소하였다가 12시간부터 발현이 증가되었으며 cyclin Dl mRNA는 조사후 바로 증가하여 48시간에 다시 감소하였다. cyclin E mRNA는 조사 후 시간이 경과함에 따라 감소하였다. CDK2 mRNA는 3시간째는 감소하다가 6시간부터 많은 증가를 보였으며 CDK4 mRNA는 조사후 6-12시간에 급격한 발현증가를 보였다. $p16^{INK4a}$ RNA는 발현의 변화가 없었으며, $p21^{WAF1}$ 및 $p27^{KIP1}$ RNA의 발현은 관찰되지 않았다. 결론 : 이상의 결과로 미루어볼 때, 방사선 조사에 의한 HL60세포의 apoptosis와 세포의 Gl/S transition는 밀접한 관계가 있는 것으로 생각되며 Rb단백의 증가와 활성형 Rb단백의 감소 현상도 관련이 있는 것으로 사료된다. 이는 E2F의 비정상적인 과발현 및 cyclin E/CDK2의 발현 증가와 관련이 있는 것으로 추측된다. 또한 $p21^{WAF1}$ 및 $p27^{KIP1}$는 방사선에 의한 apoptosis에는 관여되지 않는 것으로 사료된다.

• 생명과학회지
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• 제29권11호
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• pp.1218-1226
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• 2019

흰점박이꽃무지는 약용으로서 매우 유용하고 중요한 곤충종이다. 선행연구를 통해 전사체 분석결과를 바탕으로 인실리코(in silico) 분석을 실시하여 전사체유래 항균 펩타이드를 스크리닝하고 선발하여 항균활성 및 용혈활성을 확인하였다. 그 결과 선발된 항균 펩타이드들은 박테리아와 칸디다 진균에 강한 항균활성을 나타낸 반면 적혈구에 대한 용혈활성은 전혀 없었다. 선발한 펩타이드들 중 프로테티아마이신 2로 명명한 양이온 항균 펩타이드를 이용하여 항균활성뿐만 아니라 마우스의 대식세포 Raw264.7 세포주를 이용하여 프로테티아마이신 2의 항염증활성을 확인하였다. 그 결과 프로테티아마이신 2는 LPS로 유도된 Raw264.7 세포들의 산화질소 생성을 감소시키는 결과를 보여주었다. 또한 프로테티아마이신 2가 산화질소 합성효소(iNOS), 고리형 산소화 효소-2(COX-2)의 발현을 감소시킨다는 것을 실시간 역전사 중합효소 연쇄반응(qRT-PCR) 방법과 웨스턴 블랏(western blot)을 통해 확인하였다. 더욱이 Raw264.7 세포에서 전염증성 사이토카인($TNF-{\alpha}$, IL-6, $IL-1{\beta}$)의 발현이 MAPKs와 $NF-{\kappa}B$ 신호전달과정을 통해 약화되어짐을 알 수 있었다. 게다가 프로테티아마이신 2가 LPS와 상호작용을 통해 결합한다는 것을 확인하였다. 종합하여 보면 프로테티아마이신 2는 항균활성과 함께 LPS로 유도된 Raw264.7 세포에서 항염증활성도 가지고 있음을 나타내었다.

• Maxillofacial Plastic and Reconstructive Surgery
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• 제30권3호
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• pp.217-224
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• 2008

Angiogenesis plays an important role in bone development and postnatal bone fracture repair. Vascular endothelial growth factor (VEGF) and vascular endothelial growth factor receptors (VEGFRs) have been thought to be primarily involved in promoting angiogenesis. It is well known that VEGF and its receptors have been reported to play an important role in the regulation of the interaction between angiogenesis and osteogenesis during bone repair processes. Dexamethasone, a potent synthetic glucocorticoid, promotes phenotype markers of osteoblast differentiation, such as ALP and osteocalcin. It stimulates in vitro osteogenesis of human bone marrow osteogenic stromal cells. Dexamethasone has been reported to suppress VEGF gene expression in some cells. However, our previous study demonstrated VEGF quantification increased in a time-dependent manner in periosteal-derived osteogenesis under dexamethasone. So, the purpose of this study was to examine the angiogenic phenotypes in cultured human periosteal-derived cells under high-dose dexamethasone. Periosteal-derived cells were cultured using a technique previously described. After passage 3, the periosteal-derived cells were further cultured for 28 days in an osteogenic inductive culture medium containing ascorbic acid, ${\beta}$-glycerophosphate and high-dose dexamethasone, We evaluated the expression of VEGF isoforms, VEGFR-1, VEGFR-2, and neuropilin-1, ALL VEGF isoforms ($VEGF_{121},\;VEGF_{165},\;VEGF_{189}$, and $VEGF_{206}$) expression was observed by RT-PCR analysis. VEGFR-1, VEGFR-2 and neuropilin-1 expression increased up to day 14, particularly during the early stage of mineralization. Our results suggest the involvement of direct VEGFs/VEGFRs system on periosteal-derived cells during early mineralization phase under high-dose of dexamethasone. These also suggest that VEGF might act as an autocrine growth molecule during osteoblastic differentiation of cultured human periosteal-derived cells.

• Journal of Animal Science and Technology
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• 제49권5호
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• pp.577-584
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• 2007

본 연구는 후대검정 한우 295두의 혈액으로부터 genomic DNA를 추출하여 PCR 방법에 의한 증폭과 염기서열 분석을 통하여 ADSF/resistin 유전자의 단일염기다형을 발굴하고 이들과 한우 육질관련형질과의 상관관계를 분석하기 위하여 실시하였다. 확보된 DNA로부터 염기서열을 결정한 결과 promoter와 4개의 exon영역에서는 SNP를 찾지 못하였으나 intron 영역에서 7개의 SNP를 발굴하였다. 발굴된 SNP의 출현 빈도는 0.027에서 0.16까지 그 차이가 많았다. 육질형질과의 상관분석에서 이들 SNP 중 intron 2에서 발굴된 764A ins 만이 근내지방도와 상관관계가 발견되었다(P<0.05). 근내지방도는 유전력이 매우 높기 때문에 이번에 발굴된 ADSF/resistin 유전자의 SNP 764A ins와 같이 유전표지인자를 이용하는 것이 근내 지방도의 개량을 위해 우수한 결과를 가지는 것으로 사료된다. 가축의 경제형질의 경우 다수의 유전자가 관여하기 때문에 한 개의 유전자를 이용한 가축의 선발 또는 개량에 이용한다는 것은 제한적일 수 있다. 따라서 다수의 관련 유전자를 이용한 다형현상과 경제형질과의 연관성 연구가 동반될 때 육질개선 및 가축개량에 실질적인 증대효과가 있을 것으로 사료된다.

• 동아시아식생활학회지
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• 제21권5호
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• pp.739-745
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• 2011

과실류는 대표적인 신선식품으로 식용되고 있으며, 소득의 증가에 따라 소비량이 크게 늘어나고 있다. 특히, 포도에 함유되어 있는 레스베라트롤과 같은 건강기능 성분의 효과가 과학적으로 밝혀지고 있어 포도는 단순한 영양 자원이나 기호 식품의 측면을 넘어서는 관심을 받고 있다. 이에 따라 본 연구에서는 포도의 대표적인 건강기능 성분인 레스베라트롤이 강화된 생식용 포도를 효과적으로 생산하는 방법을 찾고자 포도 수확 후 자외선을 처리하는 방법을 적용하였다. 본 연구를 통해 도출한 연구의 결론은 다음과 같다. 포도 세포에 외부에서 적당한 스트레스를 가할 경우, 레스베라트롤 생합성 유전자가 발현되어 레스베라트롤의 함량이 증가하는 현상을 확인하였다. 이 현상은 유전자 조작이 이루어지는 과정이 아니며, 단지 포도 세포의 대사를 조절하는 현상이므로 신선식품의 건강기능성분을 강화하는 효과적인 방법으로 사용될 수 있다. 특히, 자외선 조사는 간단한 장치와 단순한 처리로 가능한 방법이기 때문에 포도 산지에서 효과적으로 활용할 수 있는 방법으로 판단된다. 실제로, 수확이후 포도에 자외선을 조사하였을 때 품종에 무관하게 약 5배까지 레스베라트롤을 증폭할 수 있는 것으로 확인되었다. 본 연구에서 제시한 방법은 유전자 조작법이 아니라 세포대사조절에 의한 기법에 해당하므로 현장 활용이 크게 기대된다.

• Asian-Australasian Journal of Animal Sciences
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• 제21권4호
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• pp.489-493
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• 2008

In 2004, Jorgensen and coworkers proposed the MUC4 gene as a candidate gene of enterotoxigenic Escherichia coli (ETEC) F4ab/ac receptor in piglets and a mutation of $G{\rightarrow}C$ in intron 7 of MUC4 was identified to be associated with the ETEC F4ab/ac adhesion phenotypes. In this study, we used 310 piglets of three breeds (Landrace, Large White and Chinese Songliao Black) to analyze the relationship between this mutation and the F4ab/ac adhesion phenotype. The results show that the genotypes at this site and the ETEC F4ab/ac adhesion phenotypes were not completely consistent, although they are very strongly associated. Among the individuals with genotype CC, which was identified as a resistant genotype to F4ab/ac adhesion, only 72.1% (124/172) were non-adhesive to ETEC F4ab and 77.9% (134/172) were non-adhesive to ETEC F4ac infections. This suggests that this mutation may not be the causative mutation for ETEC F4ab/ac adhesion, rather, the actual causative mutation may be in another gene closely linked to MUC4, or at aother site within the MUC4 gene. Our results also suggest that the receptors of F4ab and F4ac may be determined by two different but closely linked loci. In order to screen other genes related to F4ab/ac adhesion in piglets, the mRNA profiles from six full sib piglets, of which three were adhesive to ETEC F4ab/ac and three non-adhesive, were analyzed by suppression subtractive hybridization (SSH). One up-regulated gene, Ep-CAM, was selected for further analysis based on its role in the intestinal epithelial cells adhesion. Using real-time RT-PCR, we found that the Ep-CAM gene was significantly up-regulated in the piglets adhesive to F4ab/ac. It was mapped to SSC3q11-q14 by radiation hybrid mapping.

• Asian-Australasian Journal of Animal Sciences
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• 제23권3호
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• pp.292-301
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• 2010

Follicle-stimulating hormone (FSH) is a pituitary glycoprotein hormone that is encoded by separate alpha- and betasubunit genes. It plays a key role in stimulating and regulating ovarian follicular development and egg production in chicken. FSH signal transduction is mediated by the FSH receptor (FSHR) that exclusively interacts with the beta-subunit of FSH, but characterization of prokaryotic expression of the FSHb gene and its effect on the expression of the FSHR gene in local chickens have received very little attention. In the current study, the cDNA fragment of the FSHb gene from Dagu chicken was amplified using reverse transcription polymerase chain reaction (RT-PCR), and inserted into the pET-28a (+) vector to construct the pET-28a-FSHb plasmid. After expression of the plasmid in E. coli BL21 (DE3) under inducing conditions, the recombination protein, $FSH{\beta}$ subunit, was purified and injected into the experimental hens and the effect on the mRNA expression levels of the FSHR gene was investigated. Sequence comparison showed that the coding region of the FSHb gene in the local chicken shared 99%-100% homology to published nucleotides in chickens; only one synonymous nucleotide substitution was detected in the region. The encoded amino acids were completely identical with the reported sequence, which confirmed that the sequences of the chicken FSHb gene and the peptides of the $FSH{\beta}$ subunit are highly conserved. This may be due to the critical role of the normal function of the FSHb gene in hormonal specificity and regulation of reproduction. The results of gene expression revealed that a recombinant protein with a molecular weight of about 19 kDa was efficiently expressed and it was identified by Western blotting analysis. After administration of the purified $FSH{\beta}$ protein, significantly higher expression levels were demonstrated in uterus, ovary and oviduct samples (p<0.05). These observations suggested that the expressed $FSH{\beta}$ protein possesses biological activity, and has a potential role in regulation of reproductive physiology in chickens.

• 한국축산식품학회지
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• 제38권4호
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• pp.780-793
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• 2018

This study was conducted to compare the anti-allergic effects of a whey protein concentrate (WPC) and WPC hydrolysate. WPC hydrolysate was prepared using enzymatic digestion for 8 h with trypsin and ${\alpha}$-chymotrypsin, after which it was freeze-dried. The allergic parameters assessed in rat basophilic leukemia (RBL)-2H3 cells were degranulation and release of ${\beta}$-hexosaminidase, release of tumor necrosis factor $(TNF)-{\alpha}$, and changes in the expression of $IL-1{\beta}$, IL-4, and IL-10 by real time polymerase chain reaction (PCR). During preparation of the WPC hydrolysate, hydrolysis increased rapidly from 0 to 10 min and then gradually increased slowly from 1 h onwards, achieving a final degree of hydrolysis of 78.50%. The SDS-PAGE analysis revealed a reduction in the intensity of several protein bands in the WPC hydrolysate compared to the WPC. IgE-induced ${\beta}$-hexosaminidase release from RBL-2H3 cells was decreased to a higher degree following treatment with the hydrolysate compared to WPC treatment. W500 ($500{\mu}g/mL$ WPC) showed the least inhibition of ${\beta}$-hexosaminidase release, but there was no significant difference between W500 and W1000 ($1,000{\mu}g/mL$) (p<0.05). H1000 ($1,000{\mu}g/mL$ WPC hydrolysate) inhibited ${\beta}$-hexosaminidase release by 39%. Compared to the control, treatment with H1000 decreased $TNF-{\alpha}$ secretion to 11.87 pg/mL. The gene expression levels of IL-1${\beta}$, IL-4, and IL-13 were all significantly decreased in hydrolysate (p<0.05). In the case of $IL-1{\beta}$ and IL-4, the expression levels in W1000 treated cells were decreased by 73.67% and 65%, respectively, and that of IL-13 was decreased by 66.43% compared to the control.

• Korean Journal of Acupuncture
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• 제25권2호
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• pp.159-177
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• 2008

Objectives : Crohn's disease is a severe chronic inflammation that is treated mainly by immunosuppression, which often has serious side effects. There is need to develop new therapeutic methods or drugs that have few side effects in order to treat this disease. Acupuncture with Moxi-tar at Cheonchu (ST25) has anti-inflammatory properties, but the mechanism of its anti-inflammatory actions is unclear. We investigated the protective effects and speculated the mechanisms of acupuncture with Moxi-tar at ST25 on trinitrobenzene sulfonic acid (TNBS) induced colitis in mice which is a well known Crohn's disease animal model. Methods : 5 % TNBS was treated at day 1 and day 7 into rectum of mice. To investigate therapeutic effects of acupuncture with Moxi-tar at ST25, acupuncture was carried out on day 3, and day 6. For the data analysis, we observed macroscopic and microscopic findings of the colon. Weight and width of the colon, degree of damage, changes of body weight, and myeloperoxygenase (MPO) activity were checked. For analysing protein expression, we carried out immunohistochemical staining and Western blot. For analysing mRNA expression, RT-PCR was carried out. Results : TNBS induced damages on the colon of mice, while acupuncture of Moxi-tar at ST25 suppressed TNBS mediated damages similar to those on the colons of mice in the control (not treated with TNBS) group. The average body weight of TNBS treated mice (77.4%) was decreased compared with that of the control mice (105%), and acupuncture with Moxi-tar at ST25 suppressed the loss of body weight caused by TNBS (from 77.4% to 95.3%). TNBS induced infiltration of immune cells in all layers of the colon while acupuncture with Moxi-tar at ST25 suppressed infiltration of immune cells caused by TNBS. Furthermore, acupunctured with Moxi-tar at ST25 suppressed macro-, micro- colonic damages caused by TNBS. Acupunctured with Moxi-tar at ST25 dramatically improved the clinical and histopathological symptoms such as the increase in weight of the distal colon and the MPO activity in TNBS-induced colitis. Acupuncture with Moxi-tar at ST25 down-regulated the nuclear transcription factor kappa B ($NF-{\kappa}B$) activity and suppressed tumor necrosis factor-a (TNF-${\alpha}$), interleukin-$1{\beta}$ (IL-1${\beta}$), and intracellular adhesion molecule-1 (ICAM-1) expressions caused by TNBS. Conclusions : Acupuncture with Moxi-tar at ST25 helps recovery from the TNBS-induced colonic damage by down-regulation of $NF-{\kappa}B$ activity and suppressing of TNF-${\alpha}$, IL-1${\beta}$, and ICAM-1 expressions. This may be an important method for the treatment of Crohn's disease.

• 한국수소및신에너지학회논문집
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• 제17권4호
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• pp.379-388
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• 2006

Two mesophilic trickling bed bioreactors filled with two different types of media, hydrophilic- and hydrophobic-cubes, were designed and conducted for hydrogen production under the anaerobic fermentation of sucrose. Each bioreactor consisted of the column packed with polymeric cubes and inoculated with heat-treated sludge obtained from anaerobic digestion tank. A defined medium containing sucrose was fed by the different hydraulic retention time(HRT), and recycle rate. Hydrogen concentrations in gas-phase were constant, averaging 40% of biogas throughout the operation. Hydrogen production rate was increased till $10.5\;L{\cdot}h^{-1}{\cdot}L^{-1}$ of bioreactor when influent sucrose concentrations and recycle rates were varied. At the same time, the hydrogen production rate with hydrophobic media application was higher than its hydrophilic media application. No methane was detected when the reactor was under a normal operation. The major fermentation by-products in the liquid effluent of the both trickling biofilters were acetate, butyrate and lactate. In order to run in the long term operation of both reactor filled with hydrophilic and hydrophobic media, biofilm accumulation on hydrophilic media and biogas produced should be controlled through some process such as periodical backwashing or gas-purging. Four sample were collected from each reactor on the opposite hydrogen production rate, and their bacterial communities were compared by terminal restriction fragment length polymorphism (T-RFLP) analysis of PCR products generated using bacterial 16s rRNA gene primers (8f and 926r). It was expressed a marked difference in bacterial communities of both reactors. The trickling bed bioreactor with hydrophobic media demonstrates the feasibility of the process to produce hydrogen gas. A likely application of this reactor technology can be hydrogen gas recovery from pre-treatment of high carbohydrate-containing wastewaters.