• 한국발생생물학회지:발생과생식
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• 제14권1호
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• pp.27-34
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• 2010

A fertilized oocyte can get the competence for implantation through cleavage and stage-specific gene expression. These are under the control of autonomous and exogenous regulators including physiological culture condition. Endogenous and exogenous growth factors are considered as critical regulators of cleaving embryos during travel the oviduct and uterus. In this study, an effort was made to evaluate comprehensively the quality of embryos for implantation, grown in media enriched with EGF and PAF. The study evaluated developmental rates on given time, blastulation and hatching rates, and adhesion rates. Developmental rates of blastocyst to the hatching stage were significantly high in PAF treated group compared to the control in a dose-dependent manner but not in EGF group. Implantation rates were significantly high both PAF and EGF in a dose-dependent manner. H7, a PKC inhibitor, blocked the process of hatching of the blastocysts but combined treatment of EGF and PAF enhanced the hatching and implantation of blastocsyts. Based on these results it is suggested that EGF and PAF support acquirement of implantation competence at blastocyst stage through a PKC pathway.

• Parasites, Hosts and Diseases
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• 제52권4호
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• pp.355-365
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• 2014

The enteric protozoan parasite Entamoeba histolytica is the causative agent of human amebiasis. During infection, adherence of E. histolytica through Gal/GalNAc lectin on the surface of the amoeba can induce caspase-3-dependent or -independent host cell death. Phosphorylinositol 3-kinase (PI3K) and protein kinase C (PKC) in E. histolytica play an important function in the adhesion, killing, or phagocytosis of target cells. In this study, we examined the role of amoebic PI3K and PKC in amoeba-induced apoptotic cell death in Jurkat T cells. When Jurkat T cells were incubated with E. histolytica trophozoites, phosphatidylserine (PS) externalization and DNA fragmentation in Jurkat cells were markedly increased compared to those of cells incubated with medium alone. However, when amoebae were pretreated with a PI3K inhibitor, wortmannin before being incubated with E. histolytica, E. histolytica-induced PS externalization and DNA fragmentation in Jurkat cells were significantly reduced compared to results for amoebae pretreated with DMSO. In addition, pretreatment of amoebae with a PKC inhibitor, staurosporine strongly inhibited Jurkat T cell death. However, E. histolytica-induced cleavage of caspase-3, -6, and -7 were not inhibited by pretreatment of amoebae with wortmannin or staurosporin. In addition, we found that amoebic PI3K and PKC have an important role on amoeba adhesion to host compartment. These results suggest that amebic PI3K and PKC activation may play an important role in caspase-independent cell death in Entamoeba-induced apoptosis.

• 한국해양바이오학회지
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• 제2권2호
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• pp.102-107
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• 2007

산업의 발달과 함께 환경오염에 대한 국민적인 관심도는 날로 증가하고 있다. PCB는 우리 주변에 널리 퍼져 있고 먹이사슬을 통해 체내에도 축적되어 인체의 위해성이 우려되는 대표적인 환경오염물질이다. PCB의 노출은 성장기의 두뇌에서 가장 큰 신경독성을 나타내며 영아 및 유아는 상대적으로 높게 노출되어 위험집단으로 분류된다. 본 연구는 PCB의 신경독성에 구조-활성관계가 미치는 영향을 분석하고 PCB에 의한 독성을 최소화 할 수 있는 방안으로서 해양활성물질의 사용가능성을 이해하고자 하였다. PCB노출에 따른 신경세포의 신호전달 체계변화를 분석하기 위하여 Protein Kinase C (PKC)의 변화를 측정하였다. PKC의 전체적인 활성을 [$^3H$]PDBu로 분석한 결과 ortho-position(PCB-105, -123)을 가지고 있는 PCB가 non-ortho (pCB-77, -81) 구조보다 신경에 미치는 영향은 더 높았다. Westem blot 결과 PKC isofonn 중에는 PKC-beta II 및 epsilon의 경우 ortho-position PCB에서 더 높은 활성을 보였다. 이러한 PKC의 변화는 성장기 신경세포에서 신호전달기작의 변화에 많은 영향을 미치므로 이를 예방하거나 차단 할 수 있는 물질을 발견하고자 다양한 키토산을 처리하였다. 그 결과 1백만 달톤 이상의 고분자 키토산의 경우 PCB에 의한 신호전달 기작 변화를 억제할 수 있음을 보였다. 본 연구는 환경오염 등에 의한 독성예방에 키토산의 활용가능성을 제시하였다.

• The Korean Journal of Physiology and Pharmacology
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• 제1권2호
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• pp.185-193
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• 1997

The characteristics of $Na^+$-dependent cycloleucine uptake was investigated in OK cells with regard to substrate specificity and regulation by protein kinase C (PKC). Inhibition studies with different synthetic and natural amino acids showed a broad spectrum affinity to neutral amino acids regardless of their different side chains including branched or aromatic, indicating that the $Na^+$-dependent cycloleucine uptake in OK cells is mediated by System $B^o$ or System $B^o$-like transporter rather than the classical System A or ASC. Phorbol 12-myristate 13-acetate (PMA) and phorbol 12,13-dibutyrate, but not $4{\alpha}-PMA$ elicited a time-dependent biphasic stimulation of $Na^+$-dependent cycloleucine uptake, which produced early transient peak at 30 min and late sustained peak at 180min. Both the early and late stimulations by PMA were due to an increase in Vmax and not due to a change in Km. PKC inhibitors blocked both the early and late stimulation by PMA, while protein synthesis inhibitors blocked the late stimulation only. These results suggest the existence and regulation by PKC of System $B^o$ or System $B^o$-like broad spectrum transport system for neutral amino acids in OK cells.

• 한국미생물·생명공학회지
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• 제33권3호
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• pp.189-193
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• 2005

RBL-2H3 cell lysates에 anti-protein phosphatase(PP) 1, 2A, 2B 항체를 첨가한 후 immunoprecipitation을 실시한 결과 PP2B를 가해준 샘플에서만 HRF를 확인하였다. 역으로 monoclonal anti-HRF 항체를 가한 후 immunoprecipitation을 실시한 결과 PP1, 2A는 검출되지 않았으나 PP2B의 경우는 regulatory subunit(19 kDa), catalyic subunit(60 kDa) 모두 확인할 수 있었다. Affinity chromatography를 통해서도 PP2B가 HRF의 탈인산화에 관여함을 확인하였다 즉 19kDa의 PP2B regulatory subunit과 60kDa의 catalytic subunit 모두가 확인되었으며 외부 $Ca^{2+}$이온 첨가 여부에 따른 차이는 관찰할 수 없었다. 결론적으로 RBL-2H3 cell에서 PP2B는 PP1이나 PP2A에 비해 상대적으로 그 존재량은 적으나 HRF와 상호작용하는 phosphatase로서 검출된 반면 PP1이나 PP2A는 검출되지 않았다.

• The Korean Journal of Physiology and Pharmacology
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• 제6권1호
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• pp.33-39
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• 2002

It has been suggested that $Ca^{2+}$ sensitization mechanisms might contribute to myogenic tone, however, specific mechanisms have not yet been fully identified. Therefore, we investigated the role of protein kinase C (PKC)- or RhoA-induced $Ca^{2+}$ sensitization in myogenic tone of the rabbit basilar vessel. Myogenic tone was developed by stretch of rabbit basilar artery. Fura-2 $Ca^{2+}$ signals, contractile responses, PKC immunoblots, translocation of PKC and RhoA, and phosphorylation of myosin light chains were measured. Stretch of the resting vessel evoked a myogenic contraction and an increase in the intracellular $Ca^{2+}$ concentration $([Ca^{2+}]_i)$ only in the presence of extracellular $Ca^{2+}$. Stretch evoked greater contraction than high $K^+$ at a given $[Ca^{2+}]_i.$ The stretch-induced increase in $[Ca^{2+}]_i$ and contractile force were inhibited by treatment of the tissue with nifedipine, a blocker of voltage-dependent $Ca^{2+}$ channel, but not with gadolinium, a blocker of stretch-activated cation channels. The PKC inhibitors, H-7 and calphostin C, and a RhoA-activated protein kinase (ROK) inhibitor, Y-27632, inhibited the stretch-induced myogenic tone without changing $[Ca^{2+}]_i.$ Immunoblotting using isoform-specific antibodies showed the presence of $PKC_{\alpha}$ and $PKC_{\varepsilon}$ in the rabbit basilar artery. $PKC_{\alpha},$ but not $PKC_{\varepsilon},$ and RhoA were translocated from the cytosol to the cell membrane by stretch. Phosphorylation of the myosin light chains was increased by stretch and the increased phosphorylation was blocked by treatment of the tissue with H-7 and Y-27632, respectively. Our results are consistent with important roles for PKC and RhoA in the generation of myogenic tone. Furthermore, enhanced phosphorylation of the myosin light chains by activation of $PKC_{\alpha}$ and/or RhoA may be key mechanisms for the $Ca^{2+}$ sensitization associated with myogenic tone in basilar vessels.

• Asian-Australasian Journal of Animal Sciences
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• 제19권7호
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• pp.958-963
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• 2006

The effect of ginsenosides (GS) on germ cell proliferation was evaluated with a chicken ovarian germ-somatic cell coculture model and the mechanism involving protein kinase C (PKC) pathway was investigated. Ovarian cells were cultured in serum-free McCoy's 5A medium and challenged with GS alone or in combinations with PKC activator (phorbol 12-myristate 13-acetate, PMA) or inhibitor ($H_7$) for 48 h. The number of germ cells was counted and the proliferating cells were identified by immunocytochemistry of proliferating cell nuclear antigen (PCNA). Results showed that GS significantly increased germ cell proliferation and this stimulating effect was further increased by PMA, but inhibited by H7, in a dose-dependent manner. Moreover, GS-elevated PCNA expression and the PCNA -labeling index of germ cells displayed similar changes with the increased numbers of germ cells. These results indicated that GS stimulated proliferation of ovarian germ cells with involvement of the PKC-mediated system.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1996년도 춘계학술대회
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• pp.219-219
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• 1996

Mucin release from hamster tracheal surface epithelial(HTSE) cells can be stimulated by extracellular ATP via activation of P$_2$ purinoceptors located on the cell surface which appears to be coupled to phospholipase C via G proteins. However, our preliminary data indicate that the ATP-induced mucin release involves, in part, activation of PKC, but not an increase in the intracellular Ca++ level, suggesting the presence of another pathway which is separate from the PLC-PKC pathway, In this study, we intended to confirm the previous observation and subsequently identify an additional mechanism. Confluent HTSE cells were metabolically labeled with either $^3$H-glucosamine or $^3$H-arachidonic acid(AA), and release of either $^3$H-mucin or $^3$H-AA was quantified following various treatments. $^3$H-mucin was assayed using the sepharose CL-4B gel-filtration method, whereas $^3$H-AA liberation was measured by counting $^3$H-radioactivity in the chase medium. We found that: (1)Desensitization of PKC by pretreatment with PMA completely abolished the mucin releasing effect of PMA but partially inhibited the ATP-induced mucin release; (2) ATP increases release of $^3$H-AA in a dose-dependent fashion; (3) mepacrine, an inhibitor of PLA$_2$, attenuates ATP-induced mucin release in a dose-dependent fashion. These results confirm our previous notion that the PLC-PKC pathway is responsible, in part, for ATP-induced mucin release. Furthermore, activation of PLA$_2$ appears to be an additional pathway which is involved in ATP-induced mucin release.

• Molecules and Cells
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• 제23권2호
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• pp.138-144
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• 2007

Previously we showed that the human GM3 synthase gene was expressed during the induction of megakaryocytic differentiation in human leukemia K562 cells by phorbol 12-myristate 13-acetate (PMA). In this study we found that treatment of PMA-induced K562 cells with $G{\ddot{o}}6976$, a specific inhibitor of PKC, and U0126, an inhibitor of the extracellular signal-regulated kinase (ERK) reduced expression of GM3 synthase, whereas wortmannin, an inhibitor of phosphoinositide 3-kinase (PI3K) did not. Moreover, activation of ERK and cAMP response element binding protein (CREB) was prevented by pretreatment with $G{\ddot{o}}6976$ and U0126. PMA stimulated the promoter activity of the 5'-flanking region from -177 to -83 region of the GM3 synthase gene, and mutation or deletion of a CREB site located around -143 of the promoter reduced PMA-stimulated promoter activity, as did the inhibitors $G{\ddot{o}}6976$ and U0126. Our results demonstrate that induction of GM3 synthase during megakaryocytic differentiation in PMA-stimulated human leukemia K562 cells depends upon the PKC/ERK/CREB pathway.

• The Korean Journal of Physiology and Pharmacology
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• 제5권6호
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• pp.503-510
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• 2001

Long-term treatment of cultured bovine adrenal medullary chromaffin (BAMC) cells with arachidonic acid $(100\;{\mu}M),$ angiotesnin II (100 nM), prostaglandin $E_2\;(PGE_2;\;10\;{\mu}M),$ veratridine $(2\;{\mu}M)$ or KCl (55 mM) for 24 hrs increased both norepinephrine and epinephrine levels in the supernatant. Pretreatment with staurosporine (10 nM), a protein kinase C (PKC) inhibitor, completely blocked increases of norepinephrine and epinephrine secretion induced by arachidonic acid, angiotensin II, $PGE_2,$ veratridine or KCl. In addition, K252a, another PKC inhibitor whose structure is similar to that of staurosporine, effectively attenuated both norepinephrine and epinephrine secretion induced by arachidonic acid. However, K252a did not affect the catecholamine secretion induced by angiotensin II, $PGE_2,$ veratridine or KCl. Our results suggest that staurosporine may inhibit long-term catecholamine secretion induced by various secretagogues in a mechanism other than inhibiting PKC signaling. Furthermore, long-term secretion of catecholamines induced by arachidonic acid may be dependent on PKC pathway.