• Nutrition Research and Practice
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• 제15권6호
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• pp.673-685
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• 2021

BACKGROUND/OBJECTIVES: Obesity is associated with the impaired regulation of T cells characterized by increased numbers of Th1 and Th17 cells and the dysregulation of vitamin D metabolism. Both obesity and vitamin D have been reported to affect autophagy; however, a limited number of studies have investigated the effects of vitamin D on T cell autophagy in obese mice. Therefore, we aimed to determine whether in vitro treatment with vitamin D affects the proliferation, function, and autophagy of T cells from obese and control mice. MATERIALS/METHODS: Five-week-old male C57BL/6 mice were fed control or high-fat diets (10% or 45% kcal fat: CON or HFDs, respectively) for 12 weeks. Purified T cells were stimulated with anti-CD3 and anti-CD28 monoclonal antibodies and cultured with either 10 nM 1,25(OH)₂D₃ or 0.1% ethanol (vehicle control). The proliferative response; expression of CD25, Foxp3, RORγt, and autophagy-related proteins (LC3A/B, SQSTM1/P62, BECLIN-1, ATG12); and the production of interferon (IFN)-γ, interleukin (IL)-4, IL-17A, and IL-10 by T cells were measured. RESULTS: Compared with the CON group, T cell proliferation tended to be lower, and the production of IFN-γ was higher in the HFD group. IL-17A production was reduced by 1,25(OH)2D₃ treatment in both groups. The LC3 II/I ratio was higher in the HFD group than the CON group, but P62 did not differ. We observed no effect of vitamin D treatment on T cell autophagy. CONCLUSIONS: Our findings suggest that diet-induced obesity may impair the function and inhibit autophagy of T cells, possibly leading to the dysregulation of T cell homeostasis, which may be behind the aggravation of inflammation commonly observed in obesity.

• Reproductive and Developmental Biology
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• 제37권4호
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• pp.269-279
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• 2013

Low efficiency of somatic cell nuclear transfer (SCNT) is attributed to incomplete reprogramming of transfered nuclei into oocytes. Trichostatin A (TSA), histone deacetylase inhibitor and 5-aza-2'deoxycytidine (5-aza-dC), DNA methylation inhibitor has been used to enhance nuclear reprogramming following SCNT. However, it was not known molecular mechanism by which TSA and 5-aza-dC improve preimplantation embryo and fetal development following SCNT. The present study investigates embryo viability and gene expression of cloned porcine preimplantation embryos in the presence and absence of TSA and 5-aza-dC as compared to embryos produced by parthenogenetic activation. Our results indicated that TSA treatment significantly improved development. However 5-aza-dC did not improve development. Presence of TSA and 5-aza-dC significantly improved total cell number, and also decreased the apoptotic and autophagic index. Three apoptotic-related genes, Bak, Bcl-xL, and Caspase 3 (Casp3), and three autophagic-related genes, ATG6, ATG8, and lysosomal-associated membrane protein 2 (LAMP2), were measured by real time RT-PCR. TSA and 5-aza-dC treatment resulted in high expression of anti-apoptotic gene Bcl-xL and low pro-apoptotic gene Bak expression compared to untreated NT embryos or parthenotes. Furthermore, LC3 protein expression was lower in NT-TSA and NT-5-aza-dC embryos than those of NT and parthenotes. In addition, TSA and 5-aza-dC treated embryos displayed a global acetylated histone H3 at lysine 9 and methylated DNA H3 at lysine 9 profile similar to the parthenogenetic blastocysts. Finally, we determined that several DNA methyltransferase genes Dnmt1, Dnmt3a and Dnmt3b. NT blastocysts showed higher levels Dnmt1 than those of the TSA and 5-aza-dC blastocysts. Dnmt3a is lower in 5-aza-dC than NT, NTTSA and parthenotes. However, Dnmt3b is higher in 5-aza-dC than NT and NTTSA. These results suggest that TSA and 5-aza-dC positively regulates nuclear reprogramming which result in modulation of apoptosis and autophagy related gene expression and then reduce apoptosis and autophagy. In addition, TSA and 5-aza-dC affects the acetylated and methylated status of the H3K9.

• 생명과학회지
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• 제29권12호
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• pp.1305-1313
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• 2019

감초(Glycyrrhiza inflata)의 뿌리에서 분리된 Licochalcone (LC)은 항염증 및 항종양과 같은 많은 약리학적 효과를 가지고 있다. 현재까지 LCC는 구강암과 방광암에서 연구되었지만 폐암에서의 연구는 밝혀진 바 없다. 또한, 암에서 LCC에 의해 유도된 autophagy에 대한 연구는 없었다. 본 연구는 gefitinib-민감성 또는 내성을 갖는 폐암 세포에 대한 LCC의 효과 및 작용 메커니즘을 조사하기 위해 고안되었다. MTT 분석 데이터는 LCC가 비소세포폐암 세포주인 HCC827 (gefitinib-민감성) 및 HCC827GR (gefitinib-내성)에서 세포생존율을 유의하게 억제함을 보여주었다. 흥미롭게도, Annexin V/7-aminoactinomycin D 이중 염색 및 세포주기 분석에서 가장 높은 농도의 LCC 처리는 apoptosis를 유도하는 비율이 약 10%였다. LCC는 비소세포폐암 세포주에서 세포주기 G2/M 관련 단백질인 cyclin B1 및 cdc2의 발현을 감소시킴으로써 G2/M 정지를 야기하였다. LCC의 처리는 autophagy marker 단백질인 microtubule-associated protein 1 light chain 3 (LC3) 및 autophagy과정에 관여하는 단백질인 autophagy-related gene (Atg)5의 발현을 증가시킴으로써 autophagy를 유도하였다. 또한, LCC는 reactive oxygen species (ROS)의 생성을 증가시켰으며, ROS 억제제인 N-acetyl-L-cysteine (NAC)에 의해 세포생존율이 부분적으로 회복되었다. Western blotting 분석에서, NAC과 LCC의 동시처리에 의해 cdc2의 발현이 증가하고 LC3의 발현은 감소되었다. 이러한 결과는 LCC가 비소세포폐암에서 ROS-의존적 G2/M 정지 및 autophagy를 유도함으로써 항종양 효과에 기여할 수 있음을 나타낸다. 결론적으로, LCC 치료는 비소세포폐암에 대한 잠재적 치료제로서 유용할 수 있다.

• 대한치과마취과학회지
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• 제14권2호
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• pp.101-106
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• 2014

Background: Remifentanil, an ultra-short-acting mu-opioid receptor agonist, is unique from other opioids because of its esterase-based metabolism, minimal accumulation, and very rapid onset and offset of clinical action. Remifentanil can prevent the inflammatory response and can suppress inducible nitric oxide synthase expression in a septic mouse model. However, the effects of remifentanil on human keratinocyte and autophagy have yet to be fully elucidated during hypoxia-reoxygenation. Here we investigated whether remifentanil confers protective effect against hypoxia-reoxygenation in human keratinocyte and, if so, whether autophagy mediates this effect. Methods: The human keratinocytes were cultured under 1% oxygen tension. The cells were gassed with 94% $N_2$, and 5% $CO_2$ and incubated for 24 h at $37^{\circ}C$. To determine whether the administration of affects human keratinocytes hypoxia-reoxygenation injury, cells were then exposed to various concentrations of remifentanil (0.01, 0.1, 0.5 and 1 ng/ml) for 2 h. After remifentanil treatment, to simulate reoxygenation and recovery, the cells were reoxygenated for 12 h at $37^{\circ}C$. Control group did not receive remifentanil treatment. Normoxia group did not receive hypoxia and remifentanil treatment for 36 h. 3-MA group was treated 3-methyladenine (3-MA) for 1h before remifentanil treatment. Cell viability was measured using a quantitative colorimetric assay with MTT, showing the mitochondrial activity of living cells. Cells were stained with fluorescence and analyzed with Western blot analysis to find out any relations with activation of autophagy. Results: Prominent accumulation of autophagic specific staining MDC was observed around the nuclei in RPT group HaCaT cells. Similarly, AO staining, red fluorescent spots appeared in RPT group HaCaT cells, while the Normoxia, control and 3-MA groups showed mainly green cytoplasmic fluorescence. We here examined activation of autophagy related protein under H/R-induced cells by Western blotting analysis. Atg5, Beclin-1, LC3-II (microtubule-associated protein 1 light chain 3 form II) and p62 was elevated in RPT group cells. But they were decreased when autophagy was suppressed by 3-MA (Fig. 5). Conclusions: Although the findings of this study are limited to an in vitro interpretation, we suggest that remifentanil may have a beneficial effect in the recovery of wound from hypoxia-reoxygenation injury.

• 원예과학기술지
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• 제34권1호
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• pp.102-111
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• 2016

본 연구의 목적은 BrDSR(Drought Stress Resistance in B. rapa) 유전자의 기능을 명확히 밝히고, 배추에서 건조 스트레스 반응 유전자들을 분석하는데 있다. 내혼계배추('CT001')와 BrDSR 완전장(438bp의 오픈리딩프레임)을 지닌 pSL100 vector를 재료로 아그로박테리아를 이용한 배추 형질전환을 수행하였다. PCR 분석을 통해 4개체의 형질전환체를 확보하였고, 이들의 BrDSR 발현량은 건조 스트레스 조건에서 비형질전환체 대비 약 1.9-3.4배 정도 더 큰 것으로 분석되었다. 또한 표현형 분석에서도 BrDSR이 과발현된 형질전환체들은 건조 스트레스에 저항성을 보이며 정상적인 생장을 하였다. 기 구축된 건조 스트레스 반응 유전자의 상호발현 네트워크를 기반으로 BrDSR과 밀접한 관련이 있는 유전자들을 분석하기 위해 B. rapa 135K cDNA microarray 데이터를 분석하였다. 그 결과, 환경 스트레스와 관련하여 식물체에서 잎의 노화와 자가소화에 관련된 것으로 보고된 'dark inducible 2(DIN2, AT3G60140)'와 'autophagy 8h(ATG8H, AT3G06420)' 유전자가 확인되었다. 위 결과들을 근거로 BrDSR 유전자는 건조 스트레스에 대한 저항성 향상에 중요한 역할을 할 것으로 판단되었다.