• 생명과학회지
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• 제22권5호
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• pp.697-701
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• 2012

본 연구에서는 항응고제로서의 역할을 가지면서 또한 혈액응고와는 관련 없는 종양세포의 전이 등을 조절하는 것으로 알려진 activated protein C (APC)가 종양세포의 사멸에는 어떠한 영향을 미치는 지에 대한 연구를 수행하였다. Tumor necrosis factor (TNF)-${\alpha}$와 cyclohexamide를 병합 처리하거나 FAS를 처리하게 되면 인간 신장암세포인 Caki에서는 유의적인 세포사멸이 일어난다. 하지만, APC는 이러한 세포사멸에 아무런 영향을 미치지 못하였다. 또한 TRAIL을 인간 뇌 암세포인 T98G와 유방암세포인 MDA231세포에 처리하여 세포사멸을 일으켰을 때에도 APC는 세포사멸을 조절하지 못하였다. 그러나, TRAIL에 대한 민감도를 증가시키기 위한 kahweol과 TRAIL의 병합처리나, kahweol과 malatonin의 병합처리에 의한 신장암세포의 사멸은 APC에 의해 유의적으로 억제되는 것을 확인하였다. 따라서, 이는 APC가 항암치료의 효율성을 조절 할 수 있는 가능성을 가짐을 의미한다.

• Korean Chemical Engineering Research
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• 제47권4호
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• pp.501-505
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• 2009

본 논문에서는 혈관내피세포에서 활성화된 단백질 C(Activated Protein C, APC)의 항염증 작용에서 수용성 EPCR(Soluble Endothelial Protein C Receptor, sEPCR)의 효과를 관찰하였다. sEPCR은 APC가 매개하는 항염증 작용에 있어 내피세포막의 보호효과를 저해하고, 혈관내피세포에 대한 백혈구의 부착저해 효과를 억제하며, 혈관내피세포를 관통하는 백혈구의 이동을 저해하는 효과를 억제한다. 그리고 흥미롭게도 sEPCR은 내피세포에서 TNF-alpha에 의한 세포부착단백질의 발현을 억제하는 APC의 기전을 저해함으로써 APC가 가지는 항염증 효과를 억제한다. 이것은 APC의 Gla 도메인이 내피세포의 수용체인 EPCR에 결합할 수 있는 부위에 sEPCR이 상호작용함으로써 더 이상 APC이 세포막에 존재하는 EPCR과 결합을 못함으로써 APC의 항염증 작용은 억제되는 것을 의미한다. 이 결과는 향후 중증 패혈증 및 염증질환을 효과적으로 치료할 수 있는 신약개발에 중요한 단서를 제공할 것이고 내피세포에서 아직 명확하게 밝혀지지 않은 APC의 항염증 작용의 기전을 밝히는 데 좋은 정보를 제공할 것이다.

• 한국환경과학회:학술대회논문집
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• 한국환경과학회 2008년도 추계학술발표회 발표논문집
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• pp.538-544
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• 2008

Activated protein C (APC) is thought to exert antiinflammatory activities through the endothelial protein C receptor (EPCR)-dependent cleavage of protease activated receptor 1 (PAR-1) in endothelial cells. Since thrombin cleaves PAR-1 with $\sim$3-4-orders of magnitude higher efficiency, and PAR-1 is a target for proinflammatory activities of thrombin, it is not understood how APC can elicit protective responses through the cleavage of PAR-1. In this study, we demonstrate that EPCR is associated with caveolin-1 in endothelial lipid rafts, but its occupancy by protein C leads to its dissociation from caveolin-1 and subsequent recruitment of PAR-1 to protective signaling pathways through the coupling of PAR-1 to Gi-protein. When EPCR is bound by protein C, the PAR-1-dependent protective response in endothelial cells can be mediated by either thrombin or APC. These results provide a new paradigm for understanding the mechanism through which PAR-1 and EPCR participate in cellular signaling events in endothelial cells.

• The Korean Journal of Physiology and Pharmacology
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• 제27권5호
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• pp.427-436
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• 2023

Mitotic arrest deficient 2 like 2 (Mad2L2, also known as Mad2B), the human homologue of the yeast Rev7 protein, is a regulatory subunit of DNA polymerase ζ that shares high sequence homology with Mad2, the mitotic checkpoint protein. Previously, we demonstrated the involvement of Mad2B in the cisplatin-induced DNA damage response. In this study, we extend our findings to show that Mad2B is recruited to sites of DNA damage in human cancer cells in response to cisplatin treatment. We found that in undamaged cells, Mad2B exists in a complex with Polζ-Rev1 and the APC/C subunit Cdc27. Following cisplatin-induced DNA damage, we observed an increase in the recruitment of Mad2B and Cdc20 (the activators of the APC/C), to the complex. The involvement of Mad2B-Cdc20-APC/C during DNA damage has not been reported before and suggests that the APC/C is activated following cisplatin-induced DNA damage. Using an in vitro ubiquitination assay, our data confirmed Mad2B-dependent activation of APC/C in cisplatin-treated cells. Mad2B may act as an accelerator for APC/C activation during DNA damage response. Our data strongly suggest a role for Mad2B-APC/C-Cdc20 in the ubiquitination of proteins involved in the DNA damage response.

• Biomolecules & Therapeutics
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• 제31권3호
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• pp.340-349
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• 2023

Mad2B (Mad2L2), the human homolog of the yeast Rev7 protein, is a regulatory subunit of DNA polymerase ζ that shares sequence similarity with the mitotic checkpoint protein Mad2A. Previous studies on Mad2B have concluded that it is a mitotic checkpoint protein that functions by inhibiting the anaphase-promoting complex/cyclosome (APC/C). Here, we demonstrate that Mad2B is activated in response to cisplatin-induced DNA damage. Mad2B co-localizes at nuclear foci with DNA damage markers, such as proliferating cell nuclear antigen and gamma histone H2AX (γ-H2AX), following cisplatin-induced DNA damage. However, unlike Mad2A, the binding of Mad2B to Cdc20 does not inhibit the activity of APC/C in vitro. In contrast to Mad2A, Mad2B does not localize to kinetochores or binds to Cdc20 in spindle assembly checkpoint-activated cells. Loss of the Mad2B protein leads to damaged nuclei following cisplatin-induced DNA damage. Mad2B/Rev7 depletion causes the accumulation of damaged nuclei, thereby accelerating apoptosis in human cancer cells in response to cisplatin-induced DNA damage. Therefore, our results suggest that Mad2B may be a critical modulator of DNA damage response.

• BMB Reports
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• 제46권11호
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• pp.544-549
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• 2013

High mobility group box 1 (HMGB1) is involved in the pathogenesis of vascular diseases. Unlike activated protein C (APC), the activation of PAR-1 by thrombin is known to elicit proinflammatory responses. To determine whether the occupancy of EPCR by the Gla-domain of APC is responsible for the PAR-1-dependent antiinflammatory activity of the protease, we pretreated HUVECs with the PC zymogen and then activated PAR-1 with thrombin. It was found that thrombin downregulates the HMGB1-mediated induction of both TNF-${\alpha}$ and IL-6 and inhibits the activation of both p38 MAPK and NF-${\kappa}B$ in HUVECs pretreated with PC. Furthermore, thrombin inhibited HMGB1-mediated hyperpermeability and leukocyte adhesion/migration by inhibiting the expression of cell adhesion molecules in HUVECs if EPCR was occupied. Collectively, these results suggest the concept that thrombin can initiate proinflammatory responses in vascular endothelial cells through the activation of PAR-1 may not hold true for normal vessels expressing EPCR under in vivo conditions.

• Journal of Microbiology and Biotechnology
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• 제15권4호
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• pp.694-700
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• 2005

Defects in the mitotic spindle or in the attachment of chromosomes to the spindle are believed to release an activated form of spindle checkpoint complex that inhibits APC-dependent ubiquitination and subsequently arrests the cell cycle at metaphase. When the spindle assembly is disrupted, the fission yeast mitotic arrest deficient (mad) mutants fail to arrest and rapidly lose viability. To enhance our understanding of the molecular mechanisms for the pathway of checkpoint function, the functional characterizations of Mad 1 p from Schizosaccharomyces pombe involved in this process have been carried out. Yeast two-hybrid and various deletion analyses of S. pombe Mad1 p reveal that the C terminus of Mad1p is critical for the binding of Mad2p and maintenance of Mad 1 p-Mad2p interaction. In addition, it was found. that the Mad1p region (residues 206-356) is essential for Mad1p-other checkpoint components. Mad1p truncating this region is sufficient to bind Mad2p but abolishes the checkpoint function, indicating that the checkpoint function is necessary for interaction of Mad 1 p-other checkpoint components. The possible functions of S. pombe Mad1p at the cell cycle checkpoint are discussed.

• IMMUNE NETWORK
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• 제3권1호
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• pp.1-7
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• 2003

Type II collagen (CII), major component of hyaline cartilage, has been considered as an auto-antigen in rheumatoid arthritis (RA). However, the clinical and biological significances with regard to the CII autoimmunity need to be clarified in human RA. The presence of antibodies to CII has been identified in sera, synovial fluid, and cartilage of patients with RA. In our study, the increased titer of IgG anti-CII in sera was well correlated with C-reactive protein, suggesting that this antibody may reflect the inflammatory status of RA. The titer of anti-CII antibodies (anti-CII Abs) tended to be higher in early stages of diseases. In our extending study, among 997 patients with RA, 269 (27.0%) were positive for circulatory IgG antibody to CII, those levels were fluctuated over time. It is hard to assess the significant amount of T cell responses to CII and CII (255~274) in RA. By using a sensitive method of antigen specific mixed lymphocyte culture, we can detect the presence of CII-reactive T cells in peripheral blood mononuclear cells of RA patients. Sixty seven (46.9%) of 143 patients showed positive CII reactive T cell responses to CII or CII (255~274). The frequencies of CII reactive T cells were more prominent in inflamed synovial fluid (SF) than in peripheral blood. These T cells could be clonally expanded after consecutive stimulation of CII with feeding of autologous irradiated antigen presenting cells (APC). Moreover, the production of Th1-related cytokine, such as IFN-${\gamma}$, was strongly up-regulated by CII reactive T cells. These data suggest that T cells responding to CII, which are probably presenting the IFN-${\gamma}$ producing cells, may play an important role in the perpetuation of inflammatory process in RA. To evaluate the effector function of CII reactive T cells, we investigated the effect of CII reactive T cells and fibroblasts-like synoviocytes (FLS) interaction on the production of pro-inflammatory cytokines. When the CII reactive T cells were co-cultured with FLS, the production of IL-15 and TNF-${\alpha}$ from FLS were significantly increased (2 to 3 fold increase) and this increase was clearly presented in accord to the expansion of CII reactive T cells. In addition, the production of IFN-${\gamma}$ and IL-17, T cell derived cytokines, were also increased by the co-incubation of CII reactive T cells with FLS. We also examined the impact of CII reactive T cells on chemokines production. When FLS were co-cultured with CII stimulated T cells, the production of IL-8, MCP-1, and MIP-1${\alpha}$ were significantly enhanced. The increased production of these chemokines was strongly correlated with increase the frequency of CII reactive T cells. Conclusively, immune response to CII was frequently found in RA. Activated T cells in response to CII contributed to increase the production of proinflammatory cytokines and chemokines, which were critical for inflammatory responses in RA. The interaction of CII-reactive T cells with FLS further augmented this phenomenon. Taken together, our recent studies have suggested that autoimmunity to CII could play a crucial role not only in the initiation but amplification/perpetuation of inflammatory process in human RA.