• 한국발생생물학회지:발생과생식
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• 제25권1호
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• pp.15-24
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• 2021

Benzo[a]pyrene (B[a]P) is a potent carcinogen and is classified as an endocrine-disrupting chemical. In mammalian testes, Sertoli cells support spermatogenesis. Therefore, if these cells are negatively affected by exposure to xenotoxic chemicals, spermatogenesis can be seriously disrupted. In this context, we evaluated whether mouse testicular TM4 Sertoli cells are susceptible to the induction of cytotoxicity-mediated cell death after exposure to B[a] P in vitro. In the present study, while B[a]P and B[a]P-7,8-diol were not able to induce cell death, exposure to BPDE resulted in cell death. BPDE-induced cell death is accompanied by the activation of caspase-3 and caspase-7. Depolarization of the mitochondrial membrane and cytochrome c release from mitochondria were observed in benzo[a]pyrene-7,8-diol-9,10-epoxide (BPDE)-treated cells. These results indicate that TM4 cells are susceptible to apoptosis in a caspase-dependent manner. Western blot and reverse transcription-polymerase chain reaction (RT-PCR) analyses showed that aryl hydrocarbon receptor (AhR) expression was almost undetectable in TM4 cells and that its expression was not altered after B[a]P treatment. This indicates that TM4 cells are nearly AhR-deficient. In TM4 cells, the CYP1A1 protein and its activity were not present. From these results, it is clear that AhR may be a prerequisite for CYP1A1 expression in TM4 cells. Therefore, TM4 cells can be referred to as CYP1A1-deficient cells. Thus, TM4 Sertoli cells are believed to have a rigid and protective cellular machinery against genotoxic agents. In conclusion, it is suggested that tolerance to B[a]P cytotoxicity is associated with insufficient AhR and CYP1A1 expression in testicular Sertoli cells.

• 한국발생생물학회지:발생과생식
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• 제11권1호
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• pp.49-54
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• 2007

Ethane 1,2-dimethane sulfonate(EDS)은 Leydig cells(LC)만을 선별적 사멸을 유도하는 약물로서 가역적인 테스토스테론(testosterone, T) 결핍 흰쥐 모델을 만드는데 널리 사용된다. 본 연구에서는 수컷 흰쥐 뇌하수체의 생식소자극호르몬인 LH와 FSH의 발현에 미치는 EDS 투여 효과를 조사하였다. 성숙한 수컷 흰쥐(SD strain, $300{\sim}350\;g$ B.W.)에 EDS(75 mg/kg, i.p.)를 1회 복강주사하고 주사 후 0, 1, 2, 3, 4, 5, 6 그리고 7주가 경과한 날 희생시켰다. 뇌하수체로부터 total RNA를 추출한 후 뇌하수체 glycoprotein hormone common alpha subunit($C{\alpha}$), LH beta subunit($LH{\beta}$), FSH beta subunit($FSH{\beta}$) 그리고 GnRH 수용체(GnRH-R)의 발현 변화를 semi-quantitative RT-PCR로 측정하였다. 그 결과, $C{\alpha}$ 전사수준은 주사 후 1주부터 급격히 상승하여 주사 후 4주까지 유의하게 높게 유지되다가 5주 후부터 control 수준으로 회귀하였다. $LH{\beta}$ 전사 수준은 주사 후 2주부터 유의하게 상승하여 주사 후 4주에 최고 수준에 도달하였으며, 5주 후부터 control 수준으로 감소하였다. $FSH{\beta}$ 전사수준은 주사 후 2주부터 유의하게 상승하여 주사 후 3주에 최고 수준에 도달하였으며, 4주 후부터 감소하여 5주 후에 최소치를 보였다. 유사하게, GnRH-R 전사 수준도 주사 후 2주부터 유의하게 상승하여 주사 후 3주에 최고 수준에 도달하였으며, 5주 후부터 control 수준으로 감소하였다. 본 연구는 EDS 주사에 의해 수컷 흰쥐 뇌하수체 전엽에서의 생식소 자극호르몬 subunit들과 GnRH-R의 발현 변화가 가역적으로 유도될 수 있음을 보여준 것이다. EDS 주사 모델은 수컷 흰쥐에서의 시상하부-뇌하수체 신경내분비 축의 호르몬 조절에 대한 기작을 이해하는데 도움이 될 것이다.

• 대한본초학회지
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• 제31권2호
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• pp.63-69
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• 2016

Objectives : The aerial parts of Agrimonia pilosa Ledeb (Agrimoniae Herba; AH) has been traditionally used as a Korean medicine to treatment of abdominal pain, sore throat, headaches, bloody discharge, parasitic infections and eczema. In this study, we investigated the effect of AH ethanol extract on lipopolysaccharide (LPS)-induced inflammation in RAW264.7 macrophage cells.Methods : AH was extracted by 30% ethanol (AH-E). Raw264.7 cells were treated with AH-E extract at different concentrations for 30 min and then stimulated with LPS (1㎍/㎖) or without for indicated times. Cell viability was measured by MTT assay, and nitric oxide (NO) production was measured by Griess assay. The expression of inflammatory mediators, iNOS and COX-2 and inflammatory cytokines, TNF-α, IL-1β, and IL-6 was detected by RT-PCR, and the phosphorylation of ERK1/2, p38 and JNK MAP kinases (MAPKs) was analyzed by Western blot. Also, the expression of NF-κB in nuclear and cytosol was detected by Western blot.Results : AH-E extract significantly decreased LPS-induced NO production in RAW264.7 cells. AH-E extract inhibited the mRNA expression of iNOS, COX-2, TNF-α, IL-1β, and IL-6 in LPS-stimulated cells with a dose-dependent manner. In addition, the phosphorylation of ERK, p38 and JNK MAPKs was also inhibited by AH-E extract. AP-E extract inhibited the nuclear translocation of NF-κB in LPS-stimulated cells.Conclusions : Our results suggest that AH-E extract has an anti-inflammatory activity in macrophages-mediated inflammation.

• 동의생리병리학회지
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• 제29권2호
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• pp.168-173
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• 2015

Bangpungtongseong-san(BPTSS, Fangfengtongsheng-san in Chineses) is a traditional herbal formula comprising 18 medicinal herbs. In the present study, we performed the simultaneous analysis for four compounds of BPTSS and examined anti-allergic effects in human keratinocytes and mouse splenocytes. The column for separation of four compounds was used Luna C18 column and maintained at 40℃. The mobile phase for gradient elution consisted of two solvent systems. The analysis was carried out at a flow rate of 1.0 mL/min with PDA detection at 254 and 280 nm. To evaluate production and expression of Th2 chemokines, ELISA and RT-PCR were conducted in tumor necrosis factor (TNF)-α and interferon (IFN)-γ-stimulated HaCaT cells with or without BPTSS or silymarin, a positive control for skin inflammation. To measure Th2 cytokines, primary mouse splenocytes were treated with BPTSS and performed ELISA for interleukin (IL)-4, 5, 13. Calibration curves were acquired with r2>0.9999. The contents of geniposide, liquiritin, baicalin, and glycyrrhizin in BPTSS were 5.06 ㎎/g, 7.33 ㎎/g, 27.56 ㎎/g, and 7.81 ㎎/g, respectively. BPTSS reduced TARC and RANTES production and mRNA expression in TNF-α and IFN-γ-treated HaCaT cells. BPTSS inhibited IL-4, 5, and 13 production in mouse splenocytes. Our data will be a helpful information to upgrade quality control and anti-allergic effects of BPTSS.

• Animal cells and systems
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• 제15권4호
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• pp.310-314
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• 2011

The Banna miniature pig (BNMP) is a representative miniature pig breed in China. Even though BNMP dwarfism is obvious, its underlying causative mutations remain unknown. In this study, the BNMP and Large White pig (LWP) serum growth hormone (GH) and insulin-like growth factor (IGF-1) levels were detected by ELISA and compared. BNMP serum IGF-1 levels were significantly lower than LWP levels (P<0.05). The miniature condition may arise from mutations in the GH and GH receptor (GHR) genes. Therefore, GH and GHR cDNA from the BNMP were cloned into a pMD18-T vector by RT-PCR using the total RNA obtained from the BNMP's pituitary and liver tissues. Sequencing results indicated that the open reading frame of the BNMP GH gene is composed of a 26-residue signal peptide and a 191-residue mature peptide. The coding sequence of the BNMP GHR gene contained 639 amino acids, including a signal peptide that is 18 amino acids long. Two amino acid substitutions, A09V and R22Q, were found in the signal peptide of the GH gene. Additionally, the S104P mutation was found in the BNMP's mature GH protein. Four mutations in the cytoplasmic domain of GHR may influence the downstream signal transduction of GHR, which needs further experimental evidence.

• 동아시아식생활학회지
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• 제25권5호
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• pp.813-821
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• 2015

Asthma is an increasing global health problem, and novel strategies to prevent or ameliorate the condition are needed. The purpose of the present study was to evaluate the anti-inflammatory and anti-asthma effects of Adenophora triphylla var. japonica extracts. We investigated the molecular mechanism underlying the effects of 80% ethanol extracts (AE) of A. triphylla on lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. AE treatment inhibited pro-inflammatory cytokines such as TNF-${\alpha}$ and IL-6 as well as nitric oxide (NO) production in LPS-stimulated RAW 264.7 cells. In particular, NO and pro-inflammatory cytokine production was suppressed more effectively by aerial parts (AE-A) than roots (AE-R) of A. triphylla. Quantitative RT-PCR assay showed that AE reduced mRNA levels of iNOS and COX-2. We also evaluated the anti-asthmatic effects of AE-A in an ovalbumin (OVA)-induced BALB/c mouse model. AE-A supplementation significantly reduced the amounts of airway eosinophils, IL-4 and IL-13 levels in BALF, and IgE levels in serum as compared with untreated, OVA-induced mice. These results suggest that AE-A can be considered as a therapeutic agent to potentially relieve asthma.

• Journal of Animal Science and Technology
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• 제60권7호
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• pp.18.1-18.12
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• 2018

Background: Xanthosine treatment has been previously reported to increase mammary stem cell population and milk production in cattle and goats. However, the underlying molecular mechanisms associated with the increase in stem cell population and milk production remain unclear. Methods: Primiparous Beetal goats were assigned to the study. Five days post-partum, one mammary gland of each goat was infused with xanthosine (TRT) twice daily ($2{\times}$) for 3 days consecutively, and the other gland served as a control (CON). Milk samples from the TRT and CON glands were collected on the 10th day after the last xanthosine infusion and the total RNA was isolated from milk fat globules (MEGs). Total RNA in MFGs was mainly derived from the milk epithelial cells (MECs) as evidenced by expression of milk synthesis genes. Significant differentially expressed genes (DEGs) were subjected to Gene Ontology (GO) terms using PANTHER and gene networks were generated using STRING db. Results: Preliminary analysis indicated that each individual goat responded to xanthosine treatment differently, with this trend being correlated with specific DEGs within the same animal's mammary gland. Several pathways are impacted by these DEGs, including cell communication, cell proliferation and anti-microbials. Conclusions: This study provides valuable insights into transcriptomic changes in milk producing epithelial cells in response to xanthosine treatment. Further characterization of DEGs identified in this study is likely to delineate the molecular mechanisms of increased milk production and stem or progenitor cell population by the xanthosine treatment.

• Tuberculosis and Respiratory Diseases
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• 제69권3호
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• pp.207-211
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• 2010

Novel influenza A (H1N1) virus is a common pathogen of febrile respiratory infection recently. Here, we report the case of a 63-year-old male patient who presented with 3 days' ongoing cough and fever. He was diagnosed with novel influenza A (H1N1) pneumonia by real-time reverse-transcriptase-polymerase-chain-reaction (rRT-PCR). During treatment for novel influenza A (H1N1), his symptoms and radiologic findings improved initially, but multiple lung nodules developed subsequently and found on chest x-ray (on the 5th hospital day). Mycobacterium abscessus was isolated repeatedly from sputum and bronchoalveolar lavage fluid. To our knowledge, this is the first reported case of Mycobacterium abscessus lung disease in a patient with H1N1 influenza pneumonia.

• 대한의생명과학회지
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• 제22권4호
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• pp.220-226
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• 2016

Hypertriglyceridemia induces atherosclerosis and accordingly is a major causative factor in cardiovascular diseases. Macrophages that develop into foam cells are a crucial component in the development of atherosclerosis. Monocytes can be differentiated into M1 or M2 macrophages. M1 macrophages promote inflammatory responses, whereas M2 macrophages exhibit anti-inflammatory activity. Recently, we found that triglyceride (TG)-treated THP-1 monocytes express a variety of macrophage-specific surface markers, indicating that TG treatment could trigger the differentiation of monocytes into macrophages. In this study, we investigated whether TG-induced macrophages express the M1 or the M2 macrophage phenotype. THP-1 cells were treated with various concentrations of TG for different times and the expression of M1- and M2-specific markers was evaluated by RT-PCR. We found increased expression of M1 markers (CD40, CD80, and CD86) in TG-treated THP-1 cells in a TG dose- and time-dependent manner. The expression of M2 markers (CD163, CD200R, and CD206) showed variable responses to TG treatment. Taken together, our results indicate that TG treatment triggers the differentiation of monocytes into M1 macrophages, rather than into M2 macrophages, suggesting that TG contributes to pro-inflammatory responses.

• Journal of Microbiology and Biotechnology
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• 제13권6호
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• pp.897-905
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• 2003

Phospholipase D (PLD) and ADP-ribosylation factor (ARF) were partially purified on a series of column chromatography, and their biochemical properties were characterized to understand the regulatory mechanism of PLD activation by ARF protein in the antigen-induced immune responses in guinea pigs. Heparin Sepharose and high-Q Sepharose column chromatographies were used for the purification of PLD, and Sephadex G-25, DEAE Sephacel, Source 15 PHE (HIC), Superdex-75, and Uno-Q column chromatographies were used for the purification of ARF. The purified PLD and ARF proteins were identified with anti-rabbit PLD- or ARF-specific antibodies, showing about 64 or 85 kDa for the molecular mass of PLD and 29 or 35 kDa for the sizes of ARF. Partial cDNA of ARF3 was cloned by RT-PCR in guinea pig lung tissue and its nucleotides and amino acids were sequenced. Guinea pig ARF3 showed 92% of nucleotides sequence identity and 100% of amino acid sequence homology with human ARF3. The ARF-regulated PLD activity was measured in the oleate or ARFs-containing mixed lipid vesicles. The purified and recombinant ARF (rARF) activities were assessed with the $GTP{\gamma}S$ binding assay. The PLD activity was induced by oleate in a dose-dependent manner. The purified ARF and recombinant ARF3 increased PLD activity in guinea pig lung tissues. These data show that the activity of membrane-bound PLD can be regulated by the cytosolic ARF proteins, suggesting that ARF proteins in guinea pig lung can act as a regulatory factor in controlling the PLD activity in allergic reaction.