• The Korean Journal of Malacology
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• v.32 no.4
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• pp.241-247
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• 2016

Macrophage Migration Inhibitory Factor (MIF) are well-defined role as unique cytokine and critical mediator in acute and chronic inflammatory diseases, autoimmune diseases. In this study, we isolated and characterized a full-length of MIF cDNA from the abalone (Haliotis discus hannai). The full-length cDNA of abMIF was of 1264 bp, consisting of a 5'-terminal UTR of 143 bp, an open reading frame of 360 bp and a 3-terminal UTR of 761 bp. The abalone MIF cDNA encodes a 119-amino acid polypeptide with a calculated molecular mass of 13.4 kDa and isoelectric point of 9.07. Multiple alignments and phylogenetic analysis with the deduced abalone MIF protein and showed strong homology with disk abalone (Haliotis discusdiscus). The deduced amino acid sequence of abMIF exhibited homology with other reported MIFs, such as 80%, with that of other disk abalone H. discus discus MIF gene. Quantitative real-time PCR (qRT-PCR) analysis indicated that abMIF was highly expression observed in hapatopacreas, intestine, foot, and gonad of normal conditioned abalone. Even though AbMIF mRNA level in hemocytes was low under the normal condition, it was sharply up-regulated and reached the maximum at 6 h post-infection with Vibrio parahaemolyticus, and then decreased at 24 h post-infection. This result indicates that abMIF plays an important role in responding in the innate immune system.

• The Korean Journal of Physiology and Pharmacology
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• v.20 no.5
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• pp.477-485
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• 2016

CG200745 is a novel inhibitor of histone deacetylases (HDACs), initially developed for treatment of various hematological and solid cancers. Because it is water-soluble, it can be administered orally. We hypothesized that the HDAC inhibitor, CG200745, attenuates cardiac hypertrophy and fibrosis in deoxycorticosterone acetate (DOCA)-induced hypertensive rats. For establishment of hypertension, 40 mg/kg of DOCA was subcutaneously injected four times weekly into Sprague-Dawley rats. All the rats used in this study including those in the sham group had been unilaterally nephrectomized and allowed free access to drinking water containing 1% NaCl. Systolic blood pressure was measured by the tail-cuff method. Blood chemistry including sodium, potassium, glucose, triglyceride, and cholesterol levels was analyzed. Sections of the heart were visualized after trichrome and hematoxylin and eosin stain. The expression of hypertrophic genes such as atrial natriuretic peptide A (Nppa) and atrial natriuretic peptide B (Nppb) in addition to fibrotic genes such as Collagen-1, Collagen-3, connective tissue growth factor (Ctgf), and Fibronectin were measured by quantitative real-time PCR (qRT-PCR). Injection of DOCA increased systolic blood pressure, heart weight, and cardiac fibrosis, which was attenuated by CG200745. Neither DOCA nor CG200745 affected body weight, vascular contraction and relaxation responses, and blood chemistry. Injection of DOCA increased expression of both hypertrophic and fibrotic genes, which was abrogated by CG200745. These results indicate that CG200745 attenuates cardiac hypertrophy and fibrosis in DOCA-induced hypertensive rats.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.12
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• pp.5303-5307
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• 2016

Objective: Brain tumors cause great mortality and morbidity worldwide, and success rates with surgical treatment remain very low. Several recent studies have focused on introduction of novel effective medical therapeutic approaches. Genistein is a member of the isoflavonoid family which has proved to exert anticancer effects. Here we assessed the effects of genistein on the expression of MMP-2 and VEGF in low and high grade gliomas in vitro. Materials and Methods: High and low grade glioma tumor tissue samples were obtained from a total of 16 patients, washed with PBS, cut into small pieces, digested with collagenase type I and cultured in DMEM containing 10% FBS. When cells reached passage 3, they were exposed to genistein and MMP-2 and VEGF gene transcripts were determined by quantitative real time PCR (qRT-PCR). Results: Expression of MMP-2 demonstrated 580-fold reduction in expression in low grade glioma cells post treatment with genistein compared to untreated cells (P value= 0.05). In cells derived from high grade lesions, expression of MMP-2 was 2-fold lower than in controls (P value> 0.05). Genistein caused a 4.7-fold reduction in VEGF transcript in high grade glioma cells (P value> 0.05) but no effects were evident in low grade glioma cells. Conclusion. Based on the data of the present study, low grade glioma cells appear much more sensitive to genistein and this isoflavone might offer an appropriate therapeutic intervention in these patients. Further investigation of this possibility is clearly warranted.

• BMB Reports
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• v.52 no.11
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• pp.653-658
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• 2019

MYB-type transcription factors (TFs) play important roles in plant growth and development, and in the rapid responses to unfavorable environmental conditions. We recently reported the isolation and characterization of a rice (Oryza sativa) MYB TF, OsMYB102, which is involved in the regulation of leaf senescence by downregulating abscisic acid (ABA) biosynthesis and the downstream signaling response. Based on the similarities of their sequences and expression patterns, OsMYB102 appears to be a homolog of the Arabidopsis thaliana AtMYB44 TF. Since AtMYB44 is a key regulator of leaf senescence and abiotic stress responses, it is important to examine whether AtMYB44 homologs in other plants also act similarly. Here, we generated transgenic Arabidopsis plants expressing OsMYB102 (OsMYB102-OX). The OsMYB102-OX plants showed a delayed senescence phenotype during dark incubation and were more susceptible to salt and drought stresses, considerably similar to Arabidopsis plants overexpressing AtMYB44. Real-time quantitative PCR (RT-qPCR) revealed that, in addition to known senescence-associated genes, genes encoding the ABA catabolic enzymes AtCYP707A3 and AtCYP707A4 were also significantly upregulated in OsMYB102-OX, leading to a significant decrease in ABA accumulation. Furthermore, protoplast transient expression and chromatin immunoprecipitation assays revealed that OsMYB102 directly activated AtCYP707A3 expression. Based on our findings, it is probable that the regulatory functions of AtMYB44 homologs in plants are highly conserved and they have vital roles in leaf senescence and the abiotic stress responses.

• Journal of Microbiology and Biotechnology
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• v.32 no.10
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• pp.1344-1354
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• 2022

Laryngeal cancer is one of the highest incidence, most prevalently diagnosed head and neck cancers, making it critically necessary to probe effective targets for laryngeal cancer treatment. Here, real-time quantitative reverse transcription PCR (qRT-PCR) and western blot analysis were used to detect gene expression levels in laryngeal cancer cell lines. Fluorescence in situ hybridization (FISH) and subcellular fractionation assays were used to detect the subcellular location. Functional assays encompassing Cell Counting Kit-8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU), transwell and wound healing assays were performed to examine the effects of target genes on cell proliferation and migration in laryngeal cancer. The in vivo effects were proved by animal experiments. RNA-binding protein immunoprecipitation (RIP), RNA pulldown and luciferase reporter assays were used to investigate the underlying regulatory mechanisms. The results showed that KIF26B antisense RNA 1 (KIF26B-AS1) propels cell proliferation and migration in laryngeal cancer and regulates the toll-like receptor 4 (TLR4) signaling pathway. KIF26B-AS1 also recruits FUS to stabilize TLR4 mRNA, consequently activating the TLR4 signaling pathway. Furthermore, KIF26B-AS1 plays an oncogenic role in laryngeal cancer via upregulating TLR4 expression as well as the FUS/TLR4 pathway axis, findings which offer novel insight for targeted therapies in the treatment of laryngeal cancer patients.

• Korean Journal of Pharmacognosy
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• v.53 no.2
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• pp.102-110
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• 2022

Alzheimer's disease (AD) is the most common form of dementia, and the accumulation of β-amyloid (Aβ) in the brain triggers AD, followed by hyperphosphorylation of tau protein, neurofibrillary tangles, and synapses loss, neuronal cell death, and cognitive decline occur in a chain. In APPswe neuronal cell line, 50 ㎍/ml of Campbell early (Vitis labruscana B.) leaves 50% ethanol extract (VLL) treatment inhibited the secretion of Aβ1-42 by about 63% and the secretion of Aβ1-40 by about 50%. VLL did not target the enzymatic activity of the amyloidogenic pathway and decreased the protein expression of APP. As a result of RT-qPCR (Reverse transcription-quantitative real-time PCR) of the APPswe cell line treated with VLL, it is thought that the protein expression of APP was reduced by inhibiting the transcription process of the APP gene. In addition, VLL inhibited acetylcholinesterase (AChE) enzyme activity in vitro by 27.6% and 54.7%, respectively, at 50 and 100 ㎍/ml concentrations. We found that VLL inhibited the production of Aβ, a dementia-inducing substance, by suppressing the transcription of the APP gene, and that VLL inhibited AChE activity. We suggest that VLL has the potential as a natural drug material that modulates the alleviation of dementia symptoms.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.4
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• pp.471-478
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• 2014

Investigating gene expression of immune cells of whole blood or peripheral blood mononuclear cells (PBMC) under polyinosinic:polycytidylic acid (poly I:C) stimulation is valuable for understanding the immune response of organism to RNA viruses. Quantitative real-time PCR (qRT-PCR) is a standard method for quantification of gene expression studies. However, the reliability of qRT-PCR data critically depends on proper selection of reference genes. In the study, using two different analysis programs, geNorm and NormFinder, we systematically evaluated the gene expression stability of six candidate reference genes (GAPDH, ACTB, B2M, RPL4, TBP, and PPIA) in samples of whole blood and PBMC with or without poly I:C stimulation. Generally, the six candidate genes performed a similar trend of expression stability in the samples of whole blood and PBMC, but more stably expressed in whole blood than in PBMC. geNorm ranked B2M and PPIA as the best combination for gene expression normalization, while according to NormFinder, TBP was ranked as the most stable reference gene, followed by B2M and PPIA. Comprehensively considering the results from the two programs, we recommended using the geometric mean of the three genes, TBP, PPIA and B2M, to normalize the gene expression of whole blood and PBMC with poly I:C stimulation. Our study is the first detailed survey of the gene expression stability in whole blood and PBMC with or without poly I:C stimulation and should be helpful for investigating the molecular mechanism involved in porcine whole blood and PBMC in response to poly I:C stimulation.

• BMB Reports
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• v.51 no.11
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• pp.602-607
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• 2018

Aberrant expression of microRNAs (miRNAs) plays important roles in carcinogenesis and tumor progression. However, the expression and biological role of miR-301b in triple-negative breast cancer (TNBC) remains unclear. Here we aimed to evaluate the roles and mechanisms of miR-301b in TNBC cells. miR-301b expression was assessed in TNBC specimens and cell lines by quantitative Real-Time PCR (qRT-PCR). TNBC cells were transfected with miR-301b mimics, inhibitors or Cylindromatosis (CYLD) small interfering RNA (siRNA) using Lipofectamine 2000. The functional roles of miR-301b were determined by cell proliferation, colony formation, and apoptosis assays. Western blots and qRT-PCR were used to measure the expression of mRNAs and proteins in the cells. We found that miR-301b was upregulated in TNBC specimens and cell lines. Overexpression of miR-301b promoted cell proliferation in TNBC cells, while inhibited the apoptosis induced by 5-FU. CYLD was downregulated by miR-301b at both mRNA and protein levels in TNBC cells. Dual-luciferase report assay confirmed that miR-301b downregulated CYLD by direct interaction with the 3'-untranslated region(3'-UTR) of CYLD mRNA. $NF-{\kappa}B$ activation was mechanistically associated with miR-301b-mediated downregulation of CYLD. However, inhibition of miR-301b reversed all the effects of miR-301b. In conclusion, miR-301b plays an oncogenic role in TNBC possibly by downregulating CYLD and subsequently activating $NF-{\kappa}B$ p65, and this may provide a novel therapeutic approach for TNBC.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.10
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• pp.5915-5920
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• 2013

Aim: Although aberrant miRNA expression has been documented, altered miR-101 expression in cervical cancer and its carcinogenic effects and mechanisms remain unexplored. The aim of our study was to investigate the role of miR-101 alteration in cervical carcinogenesis. Methods: Expression of miR-101 was examined by quantitative real-time reverse transcriptase PCR (qRT-PCR) in Hela cells. After modulating miR-101 expression using miR-101 mimics, cell growth, apoptosis and proliferation, and migration were tested separately by MTT or flow cytometry and cell wound healing assay and protein expression was detected by qRT-PCR. The expression of COX-2 in Hela cell was also examined by immunohistochemical staining and the correlation with miR-101 expression was analysed. Results: The miR-101 demonstrated significantly low expression in Hela cell. When we transfected miR-101 mimics into Hela cells, the modulation of miR-101 expression remarkably influenced cell proliferation, cycling and apoptosis: 1) The expression of microRNA-101 tended to increase after transfection; 2) Overexpression of miR-101 was able to promote cell apoptosis, the apoptosis rate being markedly higher (97.6%) than that seen pre-transfection (12.2%) (P<0.05); 3) The miR-101 negatively regulates cell migration and invasion, scratch results being lower ($42.7um{\pm}2um$) than that observed pre-transfection ($181.4um{\pm}2um$); 4) miRNA-101 inhibits the proliferation of Hela cells as well as the level of COX-2 protein, which was negatively correlated with miR-101 expression. Conclusions: Overexpression of miR-101 has obvious inhibitory effects on cell proliferation, migration and invasion. Thus reduced miR-101 expression could participate in the development of cervical cancer at least partly through loss of inhibition of target gene COX-2, which probably occurs in a relative late phase of carcinogenesis. Our data suggest an important role of miR-101 in the molecular etiology of cancer and indicate potential application of miR-101 in cancer therapy.

• Tuberculosis and Respiratory Diseases
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• v.71 no.1
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• pp.8-14
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• 2011

Background: MicroRNAs (miRNAs) have demonstrated their potential as biomarkers for lung cancer diagnosis. In recent years, miRNAs have been found in body fluids such as serum, plasma, urine and saliva. Circulating miRNAs are highly stable and resistant to RNase activity along with, extreme pH and temperatures in serum and plasma. In this study, we investigated serum miRNA profiles that can be used as a diagnostic biomarker of non-small cell lung cancer (NSCLC). Methods: We compared the expression profile of miRNAs in the plasma of patients diagnosed with lung cancer using an miRNA microarray. The data from this assay were validated by quantitative real-time PCR (qRT-PCR). Results: Six miRNAs were overexpressed and three miRNAs were underexpressed in both tissue and serum from squamous cell carcinoma (SCC) patients. Sixteen miRNAs were overexpressed and twenty two miRNAs were underexpressed in both tissue and serum from adenocarcinoma (AC) patients. Of the four miRNAs chosen for qRT-PCR analysis, the expression of miR-23a was consistent with microarray results from AC patients. Receiver operating characteristic (ROC) curve analyses were done and revealed that the level of serum miR-23a was a potential marker for discriminating AC patients from chronic obstructive pulmonary disease (COPD) patients. Conclusion: Although a small number of patients were examined, the results from our study suggest that serum miR-23a can be used in the diagnosis of AC.