• Journal of Yeungnam Medical Science
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• 제7권1호
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• pp.39-50
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• 1990

세포막의 정보전달기전중 phosphoinositide system은 정보가 전달될때 phospholipase C 효소의 작용으로 phosphatidyl inositol bisphosphate로부터 inositol triphosphate($IP_3$)와 diacylglycerol이 생성되며 $IP_3$는 다시 $IP_3$kinase에 의해 inositol tetrakisphosphate($IP_4$)로 되어 이차전령 물로서 작용한다. 본 연구는 $IP_3$kinase효소가 $Ca^{2+}$와 calmodulin에 의해 활성화되는 성질을 이용하여 calmodulin을 정제하고 $IP_3$kinase효소와의 친화도를 비교 관찰하였다. Calmodulin정제는 phenyl-Sepharose resin을 이용하여 column chromatography를 시행하여 정제확인하였으며 분자량이 17,000임을 SDS-polyacrylamide gel 전기영동으로 확인하였다. 정제된 calmodulin을 affigel column에 결합시킨 gel에 소의 뇌로부터 분리한 $IP_3$kinase효소가 담긴 시료를 calmodulin-affigel column에 적용하여 결합 및 유출정도를 비교하였으며 $Ca^{2+}$이 든 buffer에서 친화도가 가장 컸으며 유출은 EGTA용액에서 일부 유출되었으며 calmodulin/$Ca^{2+}$이 든 buffer에선 강한 유출정도를 관찰하였다. 그러나 calmodulin/$Ca^{2+}$는 $IP_3$kinase효소의 활성을 증가시키며 calmodulin이 단백질이어서 정제면에서 효소와의 분리가 쉽지않아 여러 다른 detergent를 적용하였으나 0.2% chaps buffer에서 집중된 유출을 관찰하였다.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
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• pp.95-95
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• 2003

Inositol은 세포의증식 및 정보전달과정에 관여하는 phosphatidylinositol (PI)의 구성성분으로서 중요한 세포내 기능을 수행한다. P1는 세포내에서 특이적 인산화효소에 의하여 Pl4-phosphate(PIP), Pl4,5-phosphate($PIP_2$)로 변환되며 PIP$_2$는 phospholipase C(PLC)에 의하여 세포내 second messengers인 1,2-diacylglycerol (DAG)와 inositol 1,4,5-triphosphate ($IP_3$)로 변환된다. 이렇게 생산된 DAG와 IP3는 각각 protein kinase C의 활성과 $Ca^{2++}$의 동원에 관여하여 다양한 세포내 신호전달에 관여하는 것으로 보고되고 있다. 또한 mouse에서 $IP_3$의 작용에 의한 $Ca^{2++}$의 상승은 난모세포의 성숙분열이 촉진되고 돼지 난포세포에 있어서도 PI대사가 일어나고 있는 것으로 보고되고 있다. 본 연구에서는 돼지 미성숙 난모세포의 성숙과 단위발생에 미치는 inositol의 영향을 확인하기 위하여 실시하였다. inositol의 농도가 난포란의 성숙에 미치는 영향을 검토하기 위하여 난포란을 각각 $150 \mu mole$, $250 \mu mole$, $350 \mu mole$, inositol을 포함하는 Whitten's 배양액에서 44시간 성숙시킨 결과 $93.57 \pm 4.21, 93.91 \pm 2.71, 92.96 \pm 3.58%$가 성숙되어 대조구의 $87.10 \pm 4.21$보다 유의적(P<0.05)으로 차이를 나타내었다. 난포란의 등급에 따른 inositol의 영향을 확인하기 위하여 형태적으로 난구세포가 치밀한 난포란과 난구세포가 치밀하지 않은 난포란을 $250 \mu mole$ inositol을 포함하는 Whitten's 배양액에서 성숙을 유도한 결과 난구세포가 치밀한 난포란과 난구세포가 치밀하지 않은 난포란에서 inositol을 첨가하였을 때 성숙율은 각각 $95.35 \pm 2.22와 63.55 \pm 8.12$로 inositol을 첨가하지 않은 대조구보다($89.21 \pm 3.69와 48.56 \pm 8.99$) 유의적인 차이를 보였다. 난구세포가 inositol에 의한 성숙에 미치는 영향을 확인하기 위하여 난구세포를 제거한 난포란을 inositol을 포함하는 배지에서 성숙을 유도한 결과 inositol을 첨가하지 않은 처리구보다 양호한 성숙율을 보였다.

• 한국발생생물학회지:발생과생식
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• 제23권1호
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• pp.1-9
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• 2019

The ability of oocytes to undergo normal fertilization and embryo development is acquired during oocyte maturation which is transition from the germinal vesicle stage (GV), germinal vesicle breakdown (GVBD) to metaphase of meiosis II (MII). Part of this process includes redistribution of inositol 1, 4, 5-triphosphate receptor (IP3R), a predominant $Ca^{2+}$ channel on the endoplasmic reticulum membrane. Type 1 IP3R (IP3R1) is expressed in mouse oocytes dominantly. At GV stage, IP3R1 are arranged as a network throughout the cytoplasm with minute accumulation around the nucleus. At MII stage, IP3R1 diffuses to the entire cytoplasm in a more reticular manner, and obvious clusters of IP3R1 are observed at the cortex of the egg. This structural reorganization provides acquisition of $[Ca^{2+}]_i$ oscillatory activity during fertilization. In this review, general properties of IP3R1 in somatic cells and mammalian oocyte are introduced.

• Journal of Animal Science and Technology
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• 제51권4호
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• pp.289-294
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• 2009

본 연구는 한우 inositol 1,4,5-triphosphate receptor type1(IP3R1) 유전자를 대상으로 SNP를 발굴하고, 도체형질과의 관련성 분석을 위하여 수행하였다. PCR 및 염기서열 결정법을 통해 IP3R1 유전자내 3개의 SNP를 발굴하였고, 이중 intron 29에 위치하는 SNP의 경우 미 보고된 신규 SNP로 확인되었다. 발굴된 3개의 SNP를 대상으로 표현형 기록치를 보유한 후대검정우 583두에 대하여 유전자형 분석 및 관련성 분석을 수행하였다. 분석결과 3개의 SNP 중 g.1428617A>G SNP가 생시체중(P<0.05) 및 도체중(P<0.01)과 유의적인 상관관계가 있음을 확인할 수 있었다. 이러한 결과는 추후 한우 개량을 위한 유전자 마커로 활용이 가능할 것으로 판단된다.

• The Korean Journal of Physiology and Pharmacology
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• 제16권1호
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• pp.31-36
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• 2012

The receptor activator of NF-${\kappa}B$ ligand (RANKL) signal is an activator of tumor necrosis factor receptor-associated factor 6 (TRAF6), which leads to the activation of NF-${\kappa}B$ and other signal transduction pathways essential for osteoclastogenesis, such as $Ca^{2+}$ signaling. However, the intracellular levels of inositol 1,4,5-trisphosphate ($IP_3$) and $IP_3$-mediated cellular function of RANKL during osteoclastogenesis are not known. In the present study, we determined the levels of $IP_3$ and evaluated $IP_3$-mediated osteoclast differentiation and osteoclast activity by RANKL treatment of mouse leukemic macrophage cells (RAW 264.7) and mouse bone marrow-derived monocyte/macrophage precursor cells (BMMs). During osteoclastogenesis, the expression levels of $Ca^{2+}$ signaling proteins such as $IP_3$ receptors ($IP_3Rs$), plasma membrane $Ca^{2+}$ ATPase, and sarco/endoplasmic reticulum $Ca^{2+}$ ATPase type2 did not change by RANKL treatment for up to 6 days in both cell types. At 24 h after RANKL treatment, a higher steady-state level of $IP_3$ was observed in RAW264.7 cells transfected with green fluorescent protein (GFP)-tagged pleckstrin homology (PH) domains of phospholipase C (PLC) ${\delta}$, a probe specifically detecting intracellular $IP_3$ levels. In BMMs, the inhibition of PLC with U73122 [a specific inhibitor of phospholipase C (PLC)[ and of $IP_3Rs$ with 2-aminoethoxydiphenyl borate (2APB; a non-specific inhibitor of $IP_3Rs$) inhibited the generation of RANKL-induced multinucleated cells and decreased the bone-resorption rate in dentin slice, respectively. These results suggest that intracellular $IP_3$ levels and the $IP_3$-mediated signaling pathway play an important role in RANKL-induced osteoclastogenesis.

• Journal of Applied Biological Chemistry
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• 제63권3호
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• pp.235-241
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• 2020

An essential component of the hemostatic process during vascular damage is platelet activation. However, many cardiovascular diseases, such as atherosclerosis, thrombosis, and myocardial infarction, can develop due to excessive platelet activation. Isoscopoletin, found primarily in plant roots of the genus Artemisia or Scopolia, has been studied to demonstrate potential pharmacological effects on Alzheimer's disease and anticancer, but its mechanisms and role in relation to thrombus formation and platelet aggregation have not yet been discovered. This research investigated the effect of isoscopoletin on collagen-induced human platelet activation. As a result, isoscopoletin strongly increased cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) levels in a concentration-dependent manner. In addition, isoscopoletin greatly phosphorylated inositol 1,4,5-triphosphate receptor (IP₃R) and vasodilator-stimulated phosphoprotein (VASP), known substrates of cAMP-dependent kinase and cGMP dependent kinase. Phosphorylation of IP₃R by isoscopoletin induced Ca²⁺ inhibition from the dense tubular system Ca²⁺ channels, and VASP phosphorylation was involved in fibrinogen binding inhibition by inactivating αIIb/β₃ in the platelet membrane. Isoscopoletin finally reduced thrombin-induced fibrin clot production and finally reduced thrombus formation. Therefore, this research suggests that isoscopoletin has strong antiplatelet effects and is likely to be helpful for thrombotic diseases involving platelets by acting as a prophylactic and therapeutic agent.

• Journal of Applied Biological Chemistry
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• 제62권4호
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• pp.425-431
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• 2019

Platelet activation is essential for hemostatic process on blood vessel damage. However, excessive platelet activation can cause some cardiovascular diseases including atherosclerosis, thrombosis, and myocardial infarction. Scoparone is commonly encountered in the roots of genus Artemisia or Scopolia, and has been studied for its potential pharmacological properties including immunosuppression and vasorelaxation, but antiplatelet effects of scoparone have not been reported yet. We investigated the effect of scoparone on human platelet activation prompted by an analogue of thromboxane A₂, U46619. As the results, scoparone dose-dependently increased cyclic adenosine monophosphate (cAMP) levels as well as cyclic guanosine monophosphate (cGMP) levels, both being aggregation-inhibiting molecules. In addition, scoparone strongly phosphorylated inositol 1, 4, 5-triphosphate receptor (IP3R) and vasodilator-stimulated phosphoprotein (VASP), substrates of cAMP dependent kinase and cGMP dependent kinase. Phosphorylation of IP₃R by scoparone resulted in inhibition of Ca²⁺ mobilization in calcium channels in a dense tubular system, and phosphorylation of VASP by scoparone led to an inability of fibrinogen being able to bind to αIIb/β3. Finally, scoparone inhibited thrombin-induced fibrin clotting, thereby reducing thrombus formation. Therefore, we suggest that scoparone has a strong antiplatelet effect and is highly probable to prevent platelet-derived vascular disease.

• 생약학회지
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• 제52권1호
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• pp.26-33
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• 2021

During blood vessel damage, an essential step in the hemostatic process is platelet activation. However, it is important to properly control platelet activation, as various cardiovascular diseases, such as stroke, atherosclerosis, and myocardial infarction, are also caused by excessive platelet activation. Found primarily in the roots of plants of the genus Artemisia or Scopolia, isoscopoletin has been studied to demonstrate its potential pharmacological effects against Alzheimer's disease and anticancer, but the mechanisms and roles involved in thrombus formation and platelet aggregation are insufficient. This study investigated the effect of isoscopoletin on U46619-induced human platelet activation. As a result, isoscopoletin significantly increased the levels of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) dose-dependently. In addition, isoscopoletin significantly phosphorylated inositol 1, 4, 5-triphosphate receptor (IP3R) and vasodilator-stimulated phosphprotein (VASP), which are known substrates for cAMP-dependent kinases and cGMP-dependent kinases. Phosphorylated IP3R by isoscopoletin inhibited Ca2+ mobilization from the dense tubular system Ca2+ channels to cytosol, and phosphorylated VASP was involved in the inhibition of fibrinogen binding through αIIb/β3 inactivation in the platelet membrane. Isoscopoletin finally reduced thrombin-induced fibrin clotting production. Therefore, this study suggests that isoscopoletin has a potent antiplatelet effect and may be helpful for platelet-related thrombotic diseases.

• 한국가축번식학회지
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• 제20권4호
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• pp.395-412
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• 1997

포유동물의 난자는 제 2 감수분열 중기에서 배란되어 수정된다. 일반적으로 포유동물에서의 수정은 inositol triphosphate (IP3) 또는 cyclic adenosine diphosphoribose (cADPr)에 의해 조절되어지는 세포질내 칼슘변화에 의해 일어난다. 난활성이 일어나면 세포질내 고농도의 MPF (maturation promotion factor)는 감소하고, 전핵이 형성되고, cytoskeleton이 재구성되고, 단백질합성이 조절되어진다. 이러한 모든 과정이 정상적으로 일어난 후 수정란은 종특이한 발달경로로 배발달을 시작한다. 본 논문에서는 돼지난자의 인위적인 난활성 유도방법에 대해서 설명하고자 한다. 신호전달(signal transduction) 경로에 의한 하나의 기전이 돼지난자에서 확인되었는데, 이 경로의 자극은 난활성으로 이루어진다. 또한 전기자극에 의한 난활성방법에 의해 12일까지 정상배 발달이 이루어진다고 보고되고 있다. 향후 연구에서는 정자가 어떠한 기전에 의해서 배발달을 시작하게 하는지를 밝혀내야 한다고 본다.

• Journal of Photoscience
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• 제6권2호
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• pp.71-75
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• 1999

Tempting to further understand the molecular mechanism of pharmacological action of ethanol, we evaluated effects of phosphatidylethanol (PET) on inositol 1,4,5-triphosphate (IP3) level and protein kinase C (PKC) activity in cultured NG108-15 cells. PET increased intracellular concentration of IP$_3$. PET incorporation into membranes of NG108-15 cells had no effect on the phosphorylation of the PKC-specific substrate MBP$\_$4-14/, thus indicates that PET does not affect PKC activity in this system.