• The Korean Journal of Physiology and Pharmacology
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• 제19권5호
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• pp.467-472
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• 2015

Histone deacetylase (HDAC) has been recognized as a potentially useful therapeutic target for cardiovascular disorders. However, the effect of the HDAC inhibitor, trichostatin A (TSA), on vasoreactivity and hypertension remains unknown. We performed aortic coarctation at the inter-renal level in rats in order to create a hypertensive rat model. Hypertension induced by abdominal aortic coarctation was significantly suppressed by chronic treatment with TSA (0.5 mg/kg/day for 7 days). Nicotinamide adenine dinucleotide phosphate-driven reactive oxygen species production was also reduced in the aortas of TSA-treated aortic coarctation rats. The vasoconstriction induced by angiotensin II (Ang II, 100 nM) was inhibited by TSA in both endothelium-intact and endothelium-denuded rat aortas, suggesting that TSA has mainly acted in vascular smooth muscle cells (VSMCs). In cultured rat aortic VSMCs, Ang II increased p66shc phosphorylation, which was inhibited by the Ang II receptor type I ($AT_1R$) inhibitor, valsartan ($10{\mu}M$), but not by the $AT_2R$ inhibitor, PD123319. TSA ($1{\sim}10{\mu}M$) inhibited Ang II-induced p66shc phosphorylation in VSMCs and in HEK293T cells expressing $AT_1R$. Taken together, these results suggest that TSA treatment inhibited vasoconstriction and hypertension via inhibition of Ang II-induced phosphorylation of p66shc through $AT_1R$.

• Asian Pacific Journal of Cancer Prevention
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• 제13권7호
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• pp.3471-3476
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• 2012

Background and Objective: Histone deacetylase (HDAC) inhibitors represent a promising class of potential anticancer agents for treatment of human malignancies. In this study, we investigated the effect of trichostatin A (TSA), one such HDAC inhibitor, in combination with docetaxel (TXT), a cytotoxic chemotherapy agent or erlotinib, a novel molecular target therapy drug, on lung cancer A549 cells. Methods: A549 cells were treated with TXT, erlotinib alone or in combination with TSA, respectively. Cell viability, apoptosis, and cell cycle distribution were evaluated using MTT (3- (4, 5-dimethylthiazol-2-yl) -2, 5-diphenyltetrazolium bromide) assay, Hochst33258 staining and flow cytometry. Moreover, immunofluorescent staining and Western blot analysis were employed to examine alterations of ${\alpha}$-tubulin, heat shock protein 90 (hsp90), epidermal growth factor receptor (EGFR), and caspase-3 in response to the different exogenous stimuli. Results: Compared with single-agent treatment, co-treatment of A549 cells with TSA/TXT or TSA/erlotinib synergistically inhibited cell proliferation, induced apoptosis, and caused cell cycle delay at the $G_2/M$ transition. Treatment with TSA/TXT or TSA/erlotinib led to a significant increase of cleaved caspase-3 expression, also resulting in elevated acetylation of ${\alpha}$-tubulin or hsp90 and decreased expression of EGFR, which was negatively associated with the level of acetylated hsp90. Conclusions: Synergistic anti-tumor effects are observed between TXT or erlotinib and TSA on lung cancer cells. Such combinations may provide a more effective strategy for treating human lung cancer.

• Asian Pacific Journal of Cancer Prevention
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• 제16권8호
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• pp.3543-3549
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• 2015

Background: Bladder cancer is one of the most common cancers worldwide. Gene expression profiling using microarray technologies improves the understanding of cancer biology. The aim of this study was to determine the gene expression profile in Egyptian bladder cancer patients. Materials and Methods: Samples from 29 human bladder cancers and adjacent non-neoplastic tissues were analyzed by cDNA microarray, with hierarchical clustering and multidimensional analysis. Results: Five hundred and sixteen genes were differentially expressed of which SOS1, HDAC2, PLXNC1, GTSE1, ULK2, IRS2, ABCA12, TOP3A, HES1, and SRP68 genes were involved in 33 different pathways. The most frequently detected genes were: SOS1 in 20 different pathways; HDAC2 in 5 different pathways; IRS2 in 3 different pathways. There were 388 down-regulated genes. PLCB2 was involved in 11 different pathways, MDM2 in 9 pathways, FZD4 in 5 pathways, p15 and FGF12 in 4 pathways, POLE2 in 3 pathways, and MCM4 and POLR2E in 2 pathways. Thirty genes showed significant differences between transitional cell cancer (TCC) and squamous cell cancer (SCC) samples. Unsupervised cluster analysis of DNA microarray data revealed a clear distinction between low and high grade tumors. In addition 26 genes showed significant differences between low and high tumor stages, including fragile histidine triad, Ras and sialyltransferase 8 (alpha) and 16 showed significant differences between low and high tumor grades, like methionine adenosyl transferase II, beta. Conclusions: The present study identified some genes, that can be used as molecular biomarkers or target genes in Egyptian bladder cancer patients.

• Biomolecules & Therapeutics
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• 제24권4호
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• pp.387-394
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• 2016

Sepsis, a serious clinical problem, is characterized by a systemic inflammatory response to infection and leads to organ failure. Toll-like receptor (TLR) signaling is intimately implicated in hyper-inflammatory responses and tissue injury during sepsis. Histone deacetylase (HDAC) inhibitors have been reported to exhibit anti-inflammatory properties. The aim of this study was to investigate the hepatoprotective mechanisms of trichostatin A (TSA), a HDAC inhibitor, associated with TLR signaling pathway during sepsis. The anti-inflammatory properties of TSA were assayed in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. Polymicrobial sepsis was induced in mice by cecal ligation and puncture (CLP), a clinically relevant model of sepsis. The mice were intraperitoneally received TSA (1, 2 or 5 mg/kg) 30 min before CLP. The serum and liver samples were collected 6 and 24-h after CLP. TSA inhibited the increased production of tumor necrosis factor (TNF)-${\alpha}$ and interleukin (IL)-6 in LPS-stimulated RAW264.7 cells. TSA improved sepsis-induced mortality, attenuated liver injury and decreased serum TNF-${\alpha}$ and IL-6 levels. CLP increased the levels of TLR4, TLR2 and myeloid differentiation primary response protein 88 (MyD88) protein expression and association of MyD88 with TLR4 and TLR2, which were attenuated by TSA. CLP increased nuclear translocation of nuclear factor kappa B and decreased cytosolic inhibitor of kappa B ($I{\kappa}B$) protein expression, which were attenuated by TSA. Moreover, CLP decreased acetylation of $I{\kappa}B$ kinase (IKK) and increased association of IKK with $I{\kappa}B$ and TSA attenuated these alterations. Our findings suggest that TSA attenuates liver injury by inhibiting TLR-mediated inflammatory response during sepsis.

• Asian Pacific Journal of Cancer Prevention
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• 제13권4호
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• pp.1395-1399
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• 2012

Background: Histone deacetylase (HDAC) inhibitors have been reported to induce cell growth arrest, apoptosis and differentiation of tumor cells. The present study aimed to examine the effects of trichostatin A (TSA), one such inhibitor, on the cell cycle, apoptosis and invasiveness of osteosarcoma cells. Methods: MG-63 cells were treated with TSA at various concentrations. Then, cell growth and apoptosis were determined by 3-(4, 5-dimethyl-2-thiazolyl)-2H-tetrazolium bromide (MTT) and TUNEL assays, respectively; cell cycling was assessed by flow cytometry; invasion assays were performed with the transwell Boyden Chamber system. Results: MTT assays revealed that TSA significantly inhibited the growth of MG-63 cells in a concentration and time dependent manner. TSA treated cells demonstrated morphological changes indicative of apoptosis and TUNEL assays revealed increased apoptosis of MG-63 cells after TSA treatment. Flow cytometry showed that TSA arrested the cell cycle in G1/G2 phase and annexin V positive apoptotic cells increased markedly. In addition, the invasiveness of MG-63 cells was inhibited by TSA in a concentration dependent manner. Conclusion: Our findings demonstrate that TSA inhibits the proliferation, induces apoptosis and inhibits invasiveness of osteosarcoma cells in vitro. HDAC inhibitors may thus have promise to become new therapeutic agents against osteosarcoma.

• Reproductive and Developmental Biology
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• 제35권2호
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• pp.159-165
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• 2011

To explore the role of histone deactylase (HDAC) activation in an in vivo model of hypertrophy, we studied the effects of Trichostatin A (TSA). TSA subjected to thoracic aortic banding (TAB)-induced pressure stress in mice. In histological observations, TAB in treated mice showed a significant hypertrophic response, whereas the sham operation remained nearly normal structure with partially blunted hypertrophy. TSA treatment had no effect (measured as HW/BW) on sham-operated animals. TAB animals treated with vehicle manifested a robust ~50% hypertrophic response (p<0.05 vs sham). TAB mice treated with 2 mg/kg/day TSA manifested a blunted growth responses, which was significantly diminished (p<0.05) compared with vehicle-treated TAB mice. TAB mice treated with a lower dose of TSA (0.5 mg/kg/day) manifested a similar blunting of hypertrophic growth (~25% increase in heart mass). Furthermore, to determine activity duration of TSA in vitro, 1 nM TSA was added to H9c2 cells. Histone acetylation was initiated at 4 hr after treatment, and it was peak up to 18 hr, then followed by significantly reduced to 30 hr. We also analyzed the expression of p53 following TSA treatment, wherein p53 expression was elevated at 4 hr, and it was maintained to 24 hr after treatment. ERK was activated at 8 hr, and maintained till 30 hr after treatment suggesting an intracellular signaling interaction between TSA and p53 expression Taken together, it is suggested that HDAC activation is required for pressure-overload growth of the heart. Eventually, these data suggest that histone acetylation may be a novel target for therapeutic intervention in pressure-overloaded cardiac hypertrophy.

• International Journal of Oral Biology
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• 제35권4호
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• pp.209-214
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• 2010

Cells that have endogenous multipotent properties can be used as a starting source for the generation of induced pluripotent cells (iPSC). In addition, small molecules associated with epigenetic reprogramming are also widely used to enhance the multi- or pluripotency of such cells. Skinderived precursor cells (SKPs) are multipotent, sphereforming and embryonic neural crest-related precursor cells. These cells can be isolated from a juvenile or adult mammalian dermis. SKPs are also an efficient starting cell source for reprogramming and the generation of iPSCs because of the high expression levels of Sox2 and Klf4 in these cells as well as their endogenous multipotency. In this study, valproic acid (VPA), a histone deacetylase (HDAC) inhibitor, was tested in the generation of iPSCs as a potential enhancer of the reprogramming potential of SKPs. SKPs were isolated from the back skins of 5-6 week old C57BL/6 X DBA/2 F1 mice. After passage 3, the SKPs was treated with 2 mM of VPA and the quantitative real time RT-PCR was performed to quantify the expression of Oct4 and Klf4 (pluripotency specific genes), and Snai2 and Ngfr (neural crest specific genes). The results show that Oct4 and Klf4 expression was decreased by VPA treatment. However, there were no significant changes in neural crest specific gene expression following VPA treatment. Hence, although VPA is one of the most potent of the HDAC inhibitors, it does not enhance the reprogramming of multipotent skin precursor cells in mice.

• Tuberculosis and Respiratory Diseases
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• 제56권5호
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• pp.465-484
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• 2004

배 경 : 인슐린 양 성장 인자(IGF) 결합 단백질-3(IGFBP-3)은 IGF와 결합하여 IGF의 세포 분열 촉진 및 항세포 고사 기전을 억제할 뿐 아니라 IGF와는 독립적으로 세포고사를 유도함으로써 비소세포성 폐암 세포주의 성장을 억제한다. 방 법 : 본 연구에서 저자들은 IGFBP-3 promoter의 hyper-methylation이 IGFBP-3 단백 발현에 어떠한 역할을 하는가를 연구하였다. 또한 비소세포성 폐암 세포주에서 methylation된 IGFBP-3 promoter에서 유전자 발현을 억제하는 기전을 연구하였다. 결 과 : 본 연구에 사용된 15 종의 비소세포성 폐암 세포주 중 7종 (46.7%)에서 IGFBP-3 promoter의 methylation 이 관찰되었으며, 23명의 I기 환자 검체 중 16 (69.7%), 9명의 II기 환자 검체중 7 (77.8%), 5명의 IIIA 환자 검체중 4 (80%), 6명의 IIIB 환자 검체중 4 (66.7 %), 그리고 6 명의 IV기 환자검체중 6명 모두에서 (100%) promoter 의 methylation 이 관찰되었다. 이 비소세포성 폐암 세포주에서 promoter methylation 상태는 IGFBP-3 단백 및 mRNA 발현양상과 잘 일치하였으며, IGFBP-3의 발현이 억제되었던 비소세포성 폐암 세포주들 중 일부의 세포에서 demethylating 약제인 5'-aza-2'-deoxycytidine (5'-aza-dC) 처리 후 그 발현이 회복되었다. IGFBP-3 promoter 활성도에 중요한 역할을 하는 Sp-1/Sp-3 결합 요소는 IGFBP-3 단백 발현이 억제된 비소세포성 폐암 세포주에서 methylation되어 있었으며, 이 요소의 methylation 은 Sp-1 전사 인자의 결합을 억제하였다. ChIP assay 결과에서 IGFBP-3 promoter의 methylation 상태는 Sp-1/Sp-3 결합 요소에 Sp-1, methyl-CpG binding protein-2 (MeCP2), 그리고 histone deacetylase (HDAC)의 결합에 영향을 주며, 이는 5'-aza-dC 처리에 의하여 역전 되었다. Sp-1/Sp-3 결합 요소를 포함하고 있는 IGFBP-3 promoter의 in vitro methylation은 promoter activity를 현저히 감소시켰으며 이는 MeCP2 단백을 동시에 발현 시켰을 때 더욱 억제되며 5'-aza-dC 처리시 회복되었다. 결 론 : 이러한 결과들은 IGFBP-3 promoter의 methylation이 IGFBP-3 발현을 억제하는 하나의 기전이며, HDAC의 모집을 유도함으로서 MeCP2가 IGFBP-3 발현 억제에 중요한 역할을 함을 보이는 것이다. 이런 현상은 비소세포성 폐암에서 진단 당시의 진행된 병기와도 관계가 있어 IGFBP-3 promoter의 methylation 상태가 비소세포성 폐암의 발암 기전 및 진행에 중요한 역할을 하고 있음을 보이고 있으며, 나아가 조기 진단 및 암 예방영역에서 하나의 생물학적 지표로도 사용될 수 있을 것으로 생각된다.

• Molecules and Cells
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• 제28권1호
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• pp.1-5
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• 2009

Vascular endothelial growth factor (VEGF) is essential for many angiogenic processes both in normal and pathological conditions. However, the signaling pathways involved in VEGF-induced angiogenesis are incompletely understood. The protein kinase D1 (PKD1), a newly described calcium/calmodulin-dependent serine/threonine kinase, has been implicated in cell migration, proliferation and membrane trafficking. Increasing evidence suggests critical roles for PKD1-mediated signaling pathways in endothelial cells, particularly in the regulation of VEGF-induced angiogenesis. Recent studies show that class IIa histone deacetylases (HDACs) are PKD1 substrates and VEGF signal-responsive repressors of myocyte enhancer factor-2 (MEF2) transcriptional activation in endothelial cells. This review provides a guide to PKD1 signaling pathways and the direct downstream targets of PKD1 in VEGF signaling, and suggests important functions of PKD1 in angiogenesis.

• Asian Pacific Journal of Cancer Prevention
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• 제15권5호
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• pp.2169-2177
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• 2014

Matrine, a main active component extracted from dry roots of Sophora flavecens, has been reported to exert antitumor effects on A549 human non-small lung cancer cells, but its mechanisms of action remain unclear. To determine effects of matrine on proliferation of A549 cells and assess possible mechanisms, MTT assays were employed to detect cytotoxicity, along with o flow cytometric analysis of DNA content of nuclei of cells following staining with propidium iodide to analyze cell cycle distribution. Western blotting was performed to determined expression levels of Bax, Bcl-2, VEGF and HDAC1, while a microarray was used to assessed changes of miRNA profiles. In the MTT assay, matrine suppressed growth of human lung cancer cell A549 in a dose- and timedependent manner at doses of 0.25-2.5 mg/ml for 24h, 48h or 72h. Matrine induced cell cycle arrest in G0/G1 phase and decreased the G2/M phase, while down-regulating the expression of Bcl2 protein, leading to a reduction in the Bcl-2/Bax ratio. In addition, matrine down regulated the expression level of VEGF and HDAC1 of A549 cells. Microarray analysis demonstrated that matrine altered the expression level of miRNAs compared with untreated control A549 cells. In conclusion, matrine could inhibit proliferation of A549 cells, providing useful information for understanding anticancer mechanisms.