• Molecules and Cells
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• 제44권1호
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• pp.38-49
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• 2021

Airway mucus secretion is an essential innate immune response for host protection. However, overproduction and hypersecretion of mucus, mainly composed of the gel-forming MUC5AC protein, are significant risk factors for patients with asthma and chronic obstructive pulmonary disease (COPD). The transforming growth factor β (TGFβ) signaling pathway negatively regulates MUC5AC expression; however, the underlying molecular mechanism is not fully understood. Here, we showed that TGFβ significantly reduces the expression of MUC5AC mRNA and its protein in NCI-H292 cells, a human mucoepidermoid carcinoma cell line. This reduced MUC5AC expression was restored by a TGFβ receptor inhibitor (SB431542), but not by the inhibition of NF-κB (BAY11-7082 or Triptolide) or PI3K (LY294002) activities. TGFβ-activated Smad3 dose-dependently bound to MUC5AC promoter. Notably, TGFβ-activated Smad3 recruited HDAC2 and facilitated nuclear translocation of HDAC2, thereby inducing the deacetylation of NF-κB at K310, which is essential for a reduction in NF-κB transcriptional activity. Both TGFβ-induced nuclear translocation of Smad3/HDAC2 and deacetylation of NF-κB at K310 were suppressed by a Smad3 inhibitor (SIS3). These results suggest that the TGFβ-activated Smad3/HDAC2 complex is an essential negative regulator for MUC5AC expression and an epigenetic regulator for NF-κB acetylation. Therefore, these results collectively suggest that modulation of the TGFβ1/Smad3/HDAC2/NF-κB pathway axis can be a promising way to improve lung function as a treatment strategy for asthma and COPD.

• 한국식품과학회지
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• 제54권2호
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• pp.155-162
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• 2022

본 연구는 돈태반 효소 가수분해물의 면역증진 효과를 확인하고자 Cy를 이용한 면역 저하 모델에서 농도별 돈태반 효소 가수분해물을 투여한 실험군의 주간 체중과 조직 중량, 혈중 면역세포(백혈구, 과립구, 림프구, 중간구) 함량, 혈중 cytokine 및 immunoglobulin 함량, 자연살해세포 활성, 비장 조직 분석을 수행하였다. 체중은 대조군과 비교하여 돈태반 효소 가수분해물을 투여한 실험군 중 중농도 투여군(1.03 mg/kg BW, total nitrogen)과 고농도 투여군(2.07 mg/kg BW, total nitrogen)에서 다소 증가하는 경향을 보였고, 조직 중량은 돈태반 효소 가수분해물을 투여한 실험군이 Cy만을 단독 투여한 대조군에 비해 다소 높았으며, 이 중 고농도 투여군은 비장과 흉선 조직 중량 모두에서 대조군보다 유의적으로 높게 조사되었다. 각 실험군별 비장 조직을 이용한 자연살해세포 활성 분석 결과 정상군에 비해 대조군은 유의하게 감소하였으나 돈태반 효소 가수분해물을 고농도로 투여한 실험군과 양성대조군은 대조군에 비해 증가하는 경향을 보여 정상군과 유사한 수준을 보였다. 일반 혈액학적 분석(CBC analysis)에서 돈태반 효소 가수분해물을 투여한 실험군은 대조군에 비해 백혈구와 과립구, 림프구 및 중간구에서 모두 높은 함량을 보이는 것으로 나타났는데, 특히 고농도 투여군은 백혈구의 경우 양성 대조군인 HemoHIM 투여군과 유사한 수준으로, 과립구와 중간구의 경우 양성대조군보다 더 높은 함량을 보이는 것으로 조사되었다. 혈중 cytokine과 immunoglobulin의 함량을 분석한 결과 돈태반효소 가수분해물을 투여한 실험군에서 혈중 TNF-α와 IL-1β, IL-2, IL-12 및 IgG의 함량을 대조군과 비교하여 유의하게 증가시키거나 증가시키는 경향을 보였다. 또한 조직학적 분석에서 비장조직은 대조군에서 관찰되던 백색수질의 붕괴와 적색수질에서의 세포 응축현상은 돈태반 효소 가수분해물을 투여한 실험군에서 점차 호전되는 경향을 보였다. 이러한 결과를 바탕으로 돈태반효소 가수분해물은 Cy로 인한 세포와 조직 손상을 감소시키고 혈중 면역 관련 인자들의 함량을 증가시켜 면역력을 증진시키는데 긍정적인 영향을 미치는 것으로 판단되며, 추후 이를 활용한 건강 기능성 개발 및 의약품 개발에 따른 활용가치가 매우 높을 것으로 생각된다.

• 생명과학회지
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• 제32권1호
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• pp.36-43
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• 2022

점막면역체계는 병원체에 대항하여 정밀하게 설계된 생물체의 방어체계로 그 중 위장관은 최전방에서 가장 중요한 기능을 하는 기관이다. 숙주 감염을 일으키는 병원성 미생물 가운데 식중독을 유발하는 살모넬라는 구강경로로 체내로 들어와 파이어판을 통해서 침입한다. 다양한 혈청형의 살모넬라 균주는 여러 톨유사수용체의 리간드를 통해서 숙주의 선천면역을 자극하지만, 숙주 특이성과 병원성에 따라 장내 점막에서 다양한 후천면역반응을 유도하기도 한다. 우리는 살모넬라 균주 중 비교적 연구가 덜 되어있는 S. enteritidis에 의해 감염되었을 때 장 내에서 일어나는 면역반응을 연구하였다. 우선 마우스에 처리할 적정 농도의 살모넬라 농도를 정하기 위해 살모넬라의 농도를 다르게 하여 실험을 진행하였고 고농도의 살모넬라 처리군에서 더 낮은 생존율과 높은 몸무게 감소율을 관찰하였다. 반수치사량의 반에 해당하는 농도의 살모넬라를 마우스에 감염시켰을 때 장 내 수지상세포와 T 세포에 유의적인 차이가 없는 것을 확인하였다. 하지만 동일한 조건의 마우스에서 회장조직을 염색했을 때 뮤신 분비의 증가와 술잔세포의 감소를 확인할 수 있었다. 또한 밀착연접단백질 유전자의 상대적인 발현량을 분석하였고 살모넬라 처리군에서 Claudin의 유의적인 감소를 확인할 수 있었다. 이어 살모넬라 감염 후 DSS를 처리한 실험에서 살모넬라 처리군에서 생존율이 낮아짐을 확인하여 S. enteritidis가 염증반응을 악화시킨다는 것을 알 수 있었다.

• BMB Reports
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• 제55권2호
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• pp.81-86
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• 2022

Macrophages are a major cellular component of innate immunity and are mainly known to have phagocytic activity. In the tumor microenvironment (TME), they can be differentiated into tumor-associated macrophages (TAMs). As the most abundant immune cells in the TME, TAMs promote tumor progression by enhancing angiogenesis, suppressing T cells and increasing immunosuppressive cytokine production. N-myc downstream-regulated gene 2 (NDRG2) is a tumor suppressor gene, whose expression is down-regulated in various cancers. However, the effect of NDRG2 on the differentiation of macrophages into TAMs in breast cancer remains elusive. In this study, we investigated the effect of NDRG2 expression in breast cancer cells on the differentiation of macrophages into TAMs. Compared to tumor cell-conditioned medium (TCCM) from 4T1-mock cells, TCCM from NDRG2-over-expressing 4T1 mouse breast cancer cells did not significantly change the morphology of RAW 264.7 cells. However, TCCM from 4T1-NDRG2 cells reduced the mRNA levels of TAM-related genes, including MR1, IL-10, ARG1 and iNOS, in RAW 264.7 cells. In addition, TCCM from 4T1-NDRG2 cells reduced the expression of TAM-related surface markers, such as CD206, in peritoneal macrophages (PEM). The mRNA expression of TAM-related genes, including IL-10, YM1, FIZZ1, MR1, ARG1 and iNOS, was also downregulated by TCCM from 4T1-NDRG2 cells. Remarkably, TCCM from 4T1-NDRG2 cells reduced the expression of PD-L1 and Fra-1 as well as the production of GM-CSF, IL-10 and ROS, leading to the attenuation of T cell-inhibitory activity of PEM. These data showed that compared with TCCM from 4T1-mock cells, TCCM from 4T1-NDRG2 cells suppressed the TAM differentiation and activation. Collectively, these results suggest that NDRG2 expression in breast cancer may reduce the differentiation of macrophages into TAMs in the TME.

• Nutrition Research and Practice
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• 제16권6호
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• pp.685-699
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• 2022

BACKGROUND/OBJECTIVES: Platycodon grandiflorum (PG) has long been known as a medicinal herb effective in various diseases, including bronchitis and asthma, but is still more widely used for food. Fermentation methods are being applied to increase the pharmacological composition of PG extracts and commercialize them with high added value. This study examines the hydrolyzed and fermented PG extract (HFPGE) fermented with Lactobacillus casei in RAW 264.7 cells, and investigates the effect of amplifying the immune and the probable molecular mechanism. MATERIALS/METHODS: HFPGE's total phenolic, flavonoid, saponin, and platycodin D contents were analyzed by colorimetric analysis or high-performance liquid chromatography. Cell viability was measured by the MTT assay. Phagocytic activity was analyzed by a phagocytosis assay kit, nitric oxide (NO) production by a Griess reagent system, and cytokines by enzyme-linked immunosorbent assay kits. The mRNA expressions of inducible nitric oxide synthase (iNOS) and cytokines were analyzed by reverse transcription-polymerase chain reaction, whereas MAPK and nuclear factor (NF)-κB activation were analyzed by Western blots. RESULTS: Compared to PGE, HFPGE was determined to contain 13.76 times and 6.69 times higher contents of crude saponin and platycodin D, respectively. HFPGE promoted cell proliferation and phagocytosis in RAW 264.7 cells and regulated the NO production and iNOS expression. Treatment with HFPGE also resulted in increased production of interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, C-X-C motif chemokine ligand10, granulocyte-colony-stimulating factor, granulocyte-macrophage colony-stimulating factor, and monocyte chemoattractant protein-1, and the mRNA expressions of these cytokines. HFPGE also resulted in significantly increasing the phosphorylation of NF-κB p65, extracellular signal-regulated kinase, and c-Jun N-terminal kinase. CONCLUSIONS: Taken together, our results imply that fermentation and hydrolysis result in the extraction of more active ingredients of PG. Furthermore, we determined that HFPGE exerts immunostimulatory activity via the MAPK and NF-κB signaling pathways.

• 대한임상검사과학회지
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• 제54권3호
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• pp.217-223
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• 2022

다양한 건강기능식품 중 프로바이오틱스는 전 세계적으로 가장 방대한 시장을 이루고 있으며, 장 건강이 우리 몸의 전반적인 면역작용을 조절하며 질병을 예방하고, 나아가 치료에 도움을 줄 수 있다는 연구결과에 따라 지속적으로 프로바이오틱스에 대한 관심이 증가하고 있다. 본 연구에서는 무화과식초가 가진 항균능을 이용하여 유제품과 드레싱 소스를 개발하여 그 선호도를 조사하고 대장세포에 미치는 영향을 분석하고자 하였다. 일반우유에 5% 무화과식초를 첨가하였을 때 마시는 요거트와 리코타치즈에 대한 가장 만족도가 높았다. 제조된 유제품에서 분리한 균은 Leuconostoc lactis이며 1.0×10⁷~1.0×10⁸ CFU/mL 이상의 세균수가 측정되어 식품의 기준 및 규격의 기준치를 만족하였다. 인체의 소장 상피세포와 흡사하게 증식되는 Caco-2 세포주에서 세포생존율을 관찰한 결과, 유청을 10% 농도로 처리하였을 때 세포생존율이 19% 가량 유의적으로 증가하였다. 농도별로 유청을 처리하였을 때 최대 58%의 항산화능을 보였다. 또한 유청에서 분리된 Leuconostoc lactis는 우수한 부착능이 관찰됨에 따라 장건강에 도움을 주는 유제품임을 확인하였다.

• Clinical and Experimental Reproductive Medicine
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• 제50권1호
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• pp.44-52
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• 2023

Objective: The DNA integrity of spermatozoa that attach to fallopian tube (FT) cells is higher than spermatozoa that do not attach. FT epithelial cells can distinguish normal and abnormal sperm chromatin. This study investigated the effects of sperm with a high-DNA fragmentation index (DFI) from men with unexplained repeated implantation failure (RIF) on the Toll-like receptor (TLR) signaling pathway in human FT cells in vitro. Methods: Ten men with a RIF history and high-DFI and 10 healthy donors with low-DFI comprised the high-DFI (>30%) and control (<30%) groups, respectively. After fresh semen preparation, sperm were co-cultured with a human FT epithelial cell line (OE-E6/E7) for 24 hours. RNA was extracted from the cell line and the human innate and adaptive immune responses were tested using an RT2 profiler polymerase chain reaction (PCR) array. Results: The PCR array data showed significantly higher TLR-1, TLR-2, TLR-3, TLR-6, interleukin 1α (IL-1α), IL-1β, IL-6, IL-12, interferon α (IFN-α), IFN-β, tumor necrosis factor α (TNF-α), CXCL8, GM-CSF, G-CSF, CD14, ELK1, IRAK1, IRAK2, IRAK4, IRF1, IRF3, LY96, MAP2K3, MAP2K4, MAP3K7, MAP4K4, MAPK8, MAPK8IP3, MYD88, NFKB1, NFKB2, REL, TIRAP, and TRAF6 expression in the high-DFI group than in the control group. These factors are all involved in the TLR-MyD88 signaling pathway. Conclusion: The MyD88-dependent pathway through TLR-1, TLR-2, and TLR-6 activation may be one of the main inflammatory pathways activated by high-DFI sperm from men with RIF. Following activation of this pathway, epithelial cells produce inflammatory cytokines, resulting in neutrophil infiltration, activation, phagocytosis, neutrophil extracellular trap formation, and apoptosis.

• 대한한방부인과학회지
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• 제22권1호
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• pp.263-278
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• 2009

Purpose: The purpose of this study is to research trends in the study of breast cancer in Traditional Chinese Medicine (TCM) and to establish the further direction for its study. Methods: We reviewed TCM papers published in the last 29 years (1979-2008). Results: 1. We researched 49 papers and the patterns of study were as follows: in vitro studies were 27 papers (55.1%), in vivo studies were 9 papers (18.4%) and clinical studies were 19 papers (38.8%). 2. In vitro studies on breast cancer research in TCM were focused on cytotoxicity (17 papers) and apoptosis (8 papers). Most of in vivo studies (6 papers) were done for the purpose of inducing growth suppression of tumor cell after administration of the test drug. Each drug acted on this effect through various types of mechanism. 3. Unlike in vitro and in vivo studies, clinical studies on growth suppression of tumor cell were rare (4 papers). Most of the studies were focused on reduction of side effect of chemotherapy or synergistic effect with chemotherapy (7 papers), immune regulation (7 papers), and improvement of quality of life (6 papers). 4. Among the treatment method we reviewed, 'Runing Ⅱ(Ⅱ號方)' was the only medication that further studied as clinical trial after experimental study. 5. Since almost all studies have defects like poorly designed model or insufficient data description, it was difficult to make any definite conclusion about these studies. Conclusion: More subsequent clinical studies based on experimental study will be needed afterwards. Strict and high-level study design with detailed description will be needed in further study.

• Journal of Gastric Cancer
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• 제23권3호
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• pp.476-486
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• 2023

Purpose: The optimal tumor mutational burden (TMB) value for predicting treatment response to programmed cell death-1 (PD-1) checkpoint inhibitors in advanced gastric cancer (AGC) remains unclear. We aimed to investigate the optimal TMB cutoff value that could predict the efficacy of PD-1 checkpoint inhibitors in AGC. Materials and Methods: Patients with AGC who received pembrolizumab or nivolumab between October 1, 2020, and July 27, 2021, at Samsung Medical Center in Korea were retrospectively analyzed. The TMB levels were measured using a next-generation sequencing assay. Based on receiver operating characteristic curve analysis, the TMB cutoff value was determined. Results: A total 53 patients were analyzed. The TMB cutoff value for predicting the overall response rate (ORR) to PD-1 checkpoint inhibitors was defined as 13.31 mutations per megabase (mt/Mb) with 56% sensitivity and 95% specificity. Based on this definition, 7 (13.2%) patients were TMB-high (TMB-H). The ORR differed between the TMB-low (TMB-L) and TMB-H (8.7% vs. 71.4%, P=0.001). The progression-free survival and overall survival (OS) for 53 patients were 1.93 (95% confidence interval [CI], 1.600-2.268) and 4.26 months (95% CI, 2.992-5.532). The median OS was longer in the TMB-H (20.8 months; 95% CI, 2.292-39.281) than in the TMB-L (3.31 months; 95% CI, 1.604-5.019; P=0.049). Conclusions: The TMB cutoff value for predicting treatment response in AGC patients who received PD-1 checkpoint inhibitor monotherapy as salvage treatment was 13.31 mt/Mb. When applying the programmed death ligand-1 status to TMB-H, patients who would benefit from PD-1 checkpoint inhibitors can be selected.

• IMMUNE NETWORK
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• 제22권2호
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• pp.18.1-18.15
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• 2022

Dysfunction of mitochondrial metabolism is implicated in cellular injury and cell death. While mitochondrial dysfunction is associated with lung injury by lung inflammation, the mechanism by which the impairment of mitochondrial ATP synthesis regulates necroptosis during acute lung injury (ALI) by lung inflammation is unclear. Here, we showed that the impairment of mitochondrial ATP synthesis induces receptor interacting serine/threonine kinase 3 (RIPK3)-dependent necroptosis during lung injury by lung inflammation. We found that the impairment of mitochondrial ATP synthesis by oligomycin, an inhibitor of ATP synthase, resulted in increased lung injury and RIPK3 levels in lung tissues during lung inflammation by LPS in mice. The elevated RIPK3 and RIPK3 phosphorylation levels by oligomycin resulted in high mixed lineage kinase domain-like (MLKL) phosphorylation, the terminal molecule in necroptotic cell death pathway, in lung epithelial cells during lung inflammation. Moreover, the levels of protein in bronchoalveolar lavage fluid (BALF) were increased by the activation of necroptosis via oligomycin during lung inflammation. Furthermore, the levels of ATP5A, a catalytic subunit of the mitochondrial ATP synthase complex for ATP synthesis, were reduced in lung epithelial cells of lung tissues from patients with acute respiratory distress syndrome (ARDS), the most severe form of ALI. The levels of RIPK3, RIPK3 phosphorylation and MLKL phosphorylation were elevated in lung epithelial cells in patients with ARDS. Our results suggest that the impairment of mitochondrial ATP synthesis induces RIPK3-dependent necroptosis in lung epithelial cells during lung injury by lung inflammation.