• 농약과학회지
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• 제9권1호
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• pp.88-96
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• 2005

생물검정과 RAPD 분석을 통해서 paraquat 저항성 생태형 망초를 선발하고, 저항성 발현 기구에 있어 Paraquat의 흡수와 이행, 그리고 결합친화력 차이가 관여하고 있는지를 조사하였다. 생물검정에 의하여 선발한 paraquat 저항성 생태형 망초는 RAPD 분석 결과 감수성 생태형 망초와의 유전적 관계에 있어서는 서로 먼 유연관계에 있음이 확인되었다. 엽록소 함량을 50% 감소시키는 paraquat의 농도로 나타낸 저항성지수는 저항성 생태형이 감수성 생태형에 비하여 약 7.8배 높았다. 저항성 및 감수성 생태형 간 epicuticular wax의 함량은 비슷한 수준이었고, cuticle의 함량은 저항성 생태형이 감수성 생태형에 비하여 약 1.5배 정도 높게 나타났지만, 이러한 차이는 cuticle을 통한 paraquat의 흡수량과 이행에 영향을 끼치지는 않았다. 세포벽에 대한 결합친화력은 저항성 생태형이 감수성 생태형에 비하여 7.4배 높게 나타났으며, 엽록체포막에 대해서는 감수성 생태형이 저항성 생태형에 비하여 약 1.5배 높은 결합친화력을 보였다. paraquat의 주요 작용점인 thylakoid 막에 대한 결합친화력에서는 저항성 생태형이 감수성 생태형에 비하여 약 16.9배 정도의 차이를 나타내어 그 차이가 매우 크게 나타났다. 이상의 결과로부터 paraquat에 대한 망초의 저항성 기작은 paraquat의 세포막 및 thylakoid 막 결합에 의한 작용점으로부터 격리가 어느 정도 관여하고 있는 것으로 생각된다.

• Journal of Radiation Protection and Research
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• 제15권2호
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• pp.41-49
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• 1990

Wistar rat에 $^{88}SrCl_2$를 꼬리 정맥에 주사하여 체내 기관과 혈액 내 분포, 잔존율을 조사하였고 착화제와 유기산을 투석하여 혈장 단백질에 결합하는 Sr 양의 변화를 측정하였다. 혈액내에서 Sr은 혈장에 60%, 세포에 40% 부착되어 이동하였다. 혈장에 존재하는 Sr 중 약 50%정도는 혈장 단백질과 결합한 상태였고, 세포에는 세포 표면에 가볍게 부착되어 있었다. Erythrocyte나 granulocyte보다 lymphocyte에 많은 양의 Sr이 부착되어 있었다. 투여후 초기 1시간 이내에 혈액 내에서 급격히 감소하여 뼈에 침착되었다. 이때 각 기관에서도 Sr의 잔존율은 24시간 이내에 크게 감소하였고, 뼈로 침착된 Sr은 24시간 이후에 서서히 감소하였다. 착화제 EDTA, EGTA 및 DTPA를 투여한 경우, 혈장 단백질에 결합하는 Sr의 양은 대조군의 57%에서 27-33%로 감소하였으며 citrate 및 oxalate의 투여시는 이 값이 19%와 40%로 각각 감소하였다.

• Journal of Microbiology and Biotechnology
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• 제27권10호
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• pp.1844-1854
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• 2017

Trichomonas vaginalis is a pathogen that triggers severe immune responses in hosts. T. vaginalis ${\alpha}$-actinin 2 ($Tv{\alpha}$-actinin 2) has been used to diagnose trichomoniasis. $Tv{\alpha}$-actinin 2 was dissected into three parts; the N-terminal, central, and C-terminal portions of the protein (#1, #2, and #3, respectively). Western blot of these $Tv{\alpha}$-actinin 2 proteins with pooled patients' sera indicated that #2 and #3, but not #1, reacted with those sera. Immunofluorescence assays of two different forms of T. vaginalis (trophozoites and amoeboid forms), using anti-$Tv{\alpha}$- actinin 2 antibodies, showed localization of $Tv{\alpha}$-actinin 2 close to the plasma membranes of the amoeboid form. Fractionation experiments indicated the presence of $Tv{\alpha}$-actinin 2 in cytoplasmic, membrane, and secreted proteins of T. vaginalis. Binding of fluorescence-labeled Trichomonas to vaginal epithelial cells and prostate cells was decreased in the antibody blocking experiment using anti-$Tv{\alpha}$-actinin 2 antibodies. Pretreatment of T. vaginalis with anti-$rTv{\alpha}$-actinin 2 antibodies also resulted in reduction in its cytotoxicity. Flow cytometry, ligand-binding immunoblotting assay, and observation by fluorescence microscopy were used to detect the binding of recombinant $Tv{\alpha}$-actinin 2 to human epithelial cell lines. Specifically, the truncated N-terminal portion of $Tv{\alpha}$-actinin 2, $Tv{\alpha}$-actinin 2 #1, was shown to bind directly to vaginal epithelial cells. These data suggest that ${\alpha}$-actinin 2 is one of the virulence factors responsible for the pathogenesis of T. vaginalis by serving as an adhesin to the host cells.

• 한국가축번식학회지
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• 제18권2호
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• pp.95-99
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• 1994

우수성이 이미 입증된 oxytocin antagonist-I(AI)이 발정된 쥐의 자궁내 옥시토신 수용체 수와 결합친화력이 어떤 영향을 미치는 지를 알아보는 것이 본 연구의 목적이었다. 마취된 흰쥐들에게 5$\mu\textrm{g}$의 AI를 투여하였으며, 30분과 4시간후에 도살하였다. 자궁조직을 회수하여 잘게 잘라 냉동시킨 다음 다시 자궁조직을 분쇄한 후 단계적인 초고속 원심분리를 거쳐 옥시토신 수용체가 있는 세포막을 추출하였다. 옥시토신 수용체 분석은 높은 활동성을 보이는 옥시토신 길항제와 방사선 동위원소가 붙지 않은 옥시토신이 경쟁하는 포화상태에서 실시하였다. 옥시토신 수용체의 수와 결합친화력은 nonlinear curve fitting 방법에 의해 계산되었다. 연구 결과, AI이 투여된 흰쥐들은 수용체의 수와 결합력에 있어서 control과 유의한 차이를 보이지 않았다(p>0.05). 결론적으로 AI은 옥시토신 수용체의 수와 결합친화력을 변화시키지 않고 다만 옥시토신과 경쟁력으로 억제하는 작용을 옥시토신 수용체에서 나타낸다는 것을 이 연구에서 입증하였다.

• Clinical and Experimental Reproductive Medicine
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• 제18권2호
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• pp.181-187
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• 1991

Epidermal growth facter(EGF)는 내열성이 강하고 분자량이 6045 dalton인 단쇄상의 polypeptide로써, Cohen에 의해 생쥐의 악하선에서 처음 발견된 이래, 여러학자들에 의해 많은 연구가 되어왔다. 인체의 EGF는 urogastrone이라고도 불리우며, 인체의 소변에서 처음 검출되었고 분자구조 및 생리작용이 생쥐의 EGF와 매우 유사한 것으로 판명되었다. EGF의 자세한 작용기전은 확실히 규명되어있지는 않지만 세포의 증식과 분화를 촉진시키며 위산의 분비를 억제시킨다고 알려져 있다. 또한 EGF receptor는 분자량이 170,000${\sim}$180,000dalton인 세포표면의 polypeptide로써 인체, 쥐, 닭, 소 등의 세포막조직에 특이하게 결합되어 있다. 최근 수년동안 몇몇 학자들에 의해 EGF가 배아와 태아 및 태반의 성장을 촉진시키고 chorionic gonadotrophin과 placental lactogen의 분비를 증진하는데 기여할 것이라고 가정되어 왔다. 그러나 아직까지 배아착상에 대한 EGF의 작용여부에 관해서는 발표된 문헌이 없어 저자는 radioreceptor assay를 이용하여 EGF receptor binding과 토끼의 배아착상과의 관계를 규명하고자 임신경과에 따른 착상부위와 비착상부위의 자궁 및 태아측 태반과 모체측 태반을 분리취득하고 receptor binding assay를 시행하여 다음과 같은 결론을 얻었다. 1. 전임신군과 비임신군의 자궁조직의 membrane fraction으로부터 specific한 EGF receptor binding이 관찰되었다. 2. 착상전 임신 3일에 자궁조직의 EGF receptor수는 4.72 +0.16($10\;mol/{\mu}g$)로 비임신시보다 의의있게 증가되어 있었고(p<0.01), 착상시기인 임신 7일에는 착상된 부위에서 20.33+6.58로 훨씬 더 높은 측정치를 나타내었다(p<0.05). 3. 착상이후 가장 먼저 취득된 임신 14일의 태아측 태반은 모체측 태반의 1.39+0.49에 비해 훨씬 높은 11.94+1.97의 EGF receptor 측정치를 보였다 (p<0.01). 4. 이상의 소견들로 보아 EGF가 토끼의 배아착상에 밀접한 관련이 있을 것으로 추측되며, 이러한 착상전후의 EGF의 작용은 태아측으로부터 일 것으로 예상된다.

• IMMUNE NETWORK
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• 제6권3호
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• pp.128-137
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• 2006

Background: Apoptosis is a physiologic phenomenon involved in development, elimination of damaged cells, and maintenance of cell homeostasis. Deregulation of apoptosis may cause diseases, such as cancers, immune diseases, and neurodegenerative disorders. The mouse myeloma cell P3-X63-Ag8.653 (v653) is an HGPRT deficient $(HGPRT^-)$ mutant strain. High dependency on de novo transcription and translation of aminopterin induced apoptosis of this cell seems to be an ideal experimental system for searching apoptosis-induced genes. Methods & Results: For searching apoptosis-related genes we carried out GE-array (dot blot), Affymetrix GeneChip analysis, Northern analysis and differential display-PCR techniques. The chip data were analyzed with three different programs. 66 genes were selected through Affymetrix GeneChip analyses. All genes selected were classified into 8 groups according to their known functions. They were Genes of 1) Cell growth/maintenance/death/enzyme, 2) Cell cycle, 3) Chaperone, 4) Cancer/disease-related genes, 5) Mitochondria, 6) Membrane protein/signal transduction, 7) Nuclear protein/nucleic acid binding/transcription binding and 8) Translation factor. Among these groups number of genes were the largest in the genes of cell growth/maintenance/death/enzyme. Expression signals of most of all groups were peaked at 3 hour of apoptosis except genes of Nuclear protein/nucleic acid binding/transcription factor which showed maximum signal at 1 hour. Conclusion: This study showed induction of wide range of proapoptotic factors which accelerate cell death at various stage of cell death. In addition apoptosis studied in this research can be classified as a type 2 which involves cytochrome c and caspase 9 especially in early stages of death. But It also has progressed to type 1 in late stage of the death process.

• Toxicological Research
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• 제31권1호
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• pp.11-16
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• 2015

Our body's immune system has defense mechanisms against pathogens such as viruses and bacteria. Immune responses are primarily initiated by the activation of toll-like receptors (TLRs). In particular, TLR4 is well-characterized and is known to be activated by gram-negative bacteria and tissue damage signals. TLR4 requires myeloid differentiation factor 2 (MD2) as a co-receptor to recognize its ligand, lipopolysaccharides (LPS), which is an extracellular membrane component of gram-negative bacteria. Gambogic acid is a xanthonoid isolated from brownish or orange resin extracted from Garcinia hanburyi. Its primary effect is tumor suppression. Since inflammatory responses are related to the development of cancer, we hypothesized that gambogic acid may regulate TLR4 activation. Our results demonstrated that gambogic acid decreased the expression of pro-inflammatory cytokines ($TNF-{\alpha}$, IL-6, IL-12, and $IL-1{\beta}$) in both mRNA and protein levels in bone marrow-derived primary macrophages after stimulation with LPS. Gambogic acid did not inhibit the activation of Interferon regulatory factor 3 (IRF3) induced by TBK1 overexpression in a luciferase reporter gene assay using IFN-${\beta}$-PRD III-I-luc. An in vitro kinase assay using recombinant TBK1 revealed that gambogic acid did not directly inhibit TBK1 kinase activity, and instead suppressed the binding of LPS to MD2, as determined by an in vitro binding assay and confocal microscopy analysis. Together, our results demonstrate that gambogic acid disrupts LPS interaction with the TLR4/MD2 complex, the novel mechanism by which it suppresses TLR4 activation.

• Molecules and Cells
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• 제19권1호
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• pp.39-45
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• 2005

RPK118 is a sphingosine kinase-1-binding protein that has been implicated in sphingosine 1 phosphate-mediated signaling. It contains a PX (phox homology) domain and two pseudo-kinase domains, and co-localizes with sphingosine kinase-1 on early endosomes. In this study we identified a novel RPK118-binding protein, PRDX3 (peroxiredoxin-3), by yeast two-hybrid screening. The interaction between these proteins was confirmed by pull-down assays and co-immunoprecipitation experiments. Deletion studies showed that RPK118 interacted with PRDX3 through its pseudokinase domains, and with early endosomes through its PX domain. Double immunofluorescence experiments demonstrated that PRDX3 co-localized with RPK118 on early endosomes in COS7 cells. PRDX3 is a member of the antioxidant family of proteins synthesized in the cytoplasm and functioning in mitochondria. Our findings indicate that RPK118 is a PRDX3-binding protein that may be involved in transporting PRDX3 from the cytoplasm to its mitochondrial site of function or to other membrane structures via endosome trafficking.

• Journal of Ginseng Research
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• 제29권4호
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• pp.159-166
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• 2005

Ginsenosides, a group of Panax ginseng saponins, exert the lowering effects of plasma cholesterol levels in animals. We had reported earlier that ginsenoside Rb2 upregulate low-density lipoprotein receptor (LDLR) expression via a mechanism that is dependent of the activation of sterol response element binding protein 2 (SREBP-2) expression. This study was conducted to determine the effects of ginsenoside Rb2 on the expression of the hepatic LDLR expression at cellular levels using HepG2 cells, and to evaluate whether the sterol response element binding protein 1 (SREBP-l) was involved in the regulation of LDLR expression. Incubation of HepG2 cells in serum-free medium supplemented with cholesterol $(10{\mu}g/ml)$ for 8 hours decreased the mRNAs of LDLR mRNA by $12\%$ and SREBP-l mRNA by $35\%$. Ginsenoside Rb2 antagonized the repressive effects of cholesterol and increased both LDLR and SREBP-l mRNA expression to 1.5- and 2-fold, respectively. Furthermore, Western blot and confocal microscopic analyses with SREBP-l polyclonal antibody revealed that ginsenoside Rb2 enhanced the maturation of the SREBP-1 from the inactive precursor form in ER membrane to the active transcription factor form in nucleus. These results suggest that ginsenoside Rb2 upregulates LDLR expression via a mechanism that is dependent of the activation of not only SREBP-2 expression, but also SREBP-1 expression and maturation, and also indicate that the pharmacological value of ginsenoside Rb2 may be distinguished from that of lovastatin which is reported that it upregulate LDLR through SREBP-2 only, not through SREBP-1.

• 약학회지
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• 제30권1호
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• pp.31-41
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• 1986

Sarcolemmal membrane fraction from canine ventricle was isolated from the discarded pellet after the first homogenization in the isolation procedure of sarcoplasmic reticulum (Method 1) and the protein yield, purity, and sidedness of this preparation were compared to those of sarcolemmal fraction prepared by method of Lee et al. (Method 2) and a slight modification of original protocol of Jones et al. (Method 3). Method 1 differed from Method 2 essentially only in that vigorous homogenization was carried out by omnimixer and homogenization medium containing 30mM Tris-maleate was used in the first step. The sarcolemmal fraction was enriched from 45 to 50 and 29-fold in [$^3H$] ouabain, [$^3H$] DHA, [$^3H$] QNB binding and $Na^+$, $K^+$-ATPase activity, respectively, compared to homogenate. Total $Na^+$, $K^+$-ATPase activity of highly sarcolemma enriched fraction was 144.6$\pm$16.4$\mu\textrm{mol}$ Pi/mg protein/hr, which was about 85%, of total ATPase activity, and the yield of the preparation was 15.7 mg protein per 100g of starting ventricular tissue. The sarcolemmal preparation supported $^{45}Ca^{2+}$-uptake in the presence of ATP but this uptake was not dependent on oxalate. Sarcolemmal $Na^+$, $K^+$-ATPase activity and detectable [$^3H$] ouabain binding were increased about 32% and 35%, respectively, by pretreatment of sarcolemmal fraction with optimal concentration of sodium dodecylsulfate (0.3-0.4mg/mg protein), suggesting that this preparation contained about 24% of sealed rightside-out vesicles, 26% of sealed inside-out vesicles, and 5001o of freely permeable (leaky) form. This procedure showed the highest protein yield and leaky population, compared to Method 2 and 3. On the other hand, sarcolemmal fraction prepared by Method 2 and 3 showed low value in protein yield but comtained high population of inside-out (46%) and rightside-out (49%) vesicles, respectively, compared to present procedure (Method 1). The results indicate that vigorous homogenization decreases the population of sealed sarcolemmal vesicles but increases the sarcolemmal protein yield per gram tissue and that this procedure is available for further purification of sarcolemmal fraction and for the receptor binding study of sarcolemma.