• Biomolecules & Therapeutics
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• v.25 no.2
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• pp.202-212
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• 2017

Doxorubicin (DOX) is a highly effective chemotherapeutic agent; however, the dose-dependent cardiotoxicity associated with DOX significantly limits its clinical application. In the present study, we investigated whether Rb1 could prevent DOX-induced apoptosis in H9C2 cells via aryl hydrocarbon receptor (AhR). H9C2 cells were treated with various concentrations ($-{\mu}M$) of Rb1. AhR, CYP1A protein and mRNA expression were quantified with Western blot and real-time PCR analyses. We also evaluated the expression levels of caspase-3 to assess the anti-apoptotic effects of Rb1. Our results showed that Rb1 attenuated DOX-induced cardiomyocytes injury and apoptosis and reduced caspase-3 and caspase-8, but not caspase-9 activity in DOX-treated H9C2 cells. Meanwhile, pre-treatment with Rb1 decreased the expression of caspase-3 and PARP in the protein levels, with no effects on cytochrome c, Bax, and Bcl-2 in DOX-stimulated cells. Rb1 markedly decreased the CYP1A1 and CYP1A2 expression induced by DOX. Furthermore, transfection with AhR siRNA or pre-treatment with AhR antagonist CH-223191 significantly inhibited the ability of Rb1 to decrease the induction of CYP1A, as well as caspase-3 protein levels following stimulation with DOX. In conclusion, these findings indicate that AhR plays an important role in the protection of Ginsenoside Rb1 against DOX-triggered apoptosis of H9C2 cells.

• KSBB Journal
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• v.18 no.4
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• pp.289-293
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• 2003

Doxorubicin is an anthracycline-family polyketide compound with a very potent anti-cancer activity, typically produced by Streptomyces peucetius. In order to increase doxurubicin productivity, a semi-industrial doxorubicin-producing Streptomyces strain named BR-Dox was cultured in a R2YE liquid medium containing CaCO$_3$, and then converted to a cell wall-free protoplast using lysozyme treatment method, followed by PEG-mediated cell wall regeneration. Among several protoplast-regenerated Streptomyces BR-Dox strains, two independent isolates named BR-Dox4 and BR-Dox6 were visually selected using thin layer chromatography (TLC) based on the pigment overproducing phenotype. Comparing with Streptomyces BR-Dox parental strain, two protoplast-regenerated strains, BR-Dox4 and BR-Dox6 exhibited 25.2％ and 12.2％ higher doxorubicin productivity analyzed by high pressure liquid chromatography (HPLC), respectively. This result suggests that a protoplast-regeneration of an antibiotics-producing Streptomyces strain should be a promising strain development approach for antibiotics overproduction in Streptomyces species.

• Journal of Microbiology and Biotechnology
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• v.24 no.8
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• pp.1065-1072
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• 2014

Doxorubicin, produced by Streptomyces peucetius ATCC 27952, is tightly regulated by dnrO, dnrN, and dnrI regulators. Genome mining of S. peucetius revealed the presence of the IclR (doxR) type family of transcription regulator mediating the signal-dependent expression of operons at the nonribosomal peptide synthetase gene cluster. Overexpression of doxR in native strain strongly repressed the drug production. Furthermore, it also had a negative effect on the regulatory system of doxorubicin, wherein the transcript of dnrI was reduced to the maximum level in comparision with the other two. Interestingly, the overexpression of the same gene also had strong inhibitory effects on the production of actinorhodin (blue pigment) and undecylprodigiosin (red pigment) in Streptomyces coelicolor M145, herboxidiene production in Streptomyces chromofuscus ATCC 49982, and spinosyn production in Saccharopolyspora spinosa NRRL 18395, respectively. Moreover, DoxR exhibited pleiotropic effects on the production of blue and red pigments in S. coelicolor when grown in different agar media, wherein the production of blue pigment was inhibited in R2YE medium and the red pigment was inhibited in YEME medium. However, the production of both blue and red pigments from S. coelicolor harboring doxR was halted in ISP2 medium, whereas S. coelicolor produced both pigmented antibiotics in the same plate. These consequences demonstrate that the on and off production of these antibiotics was not due to salt stress or media compositions, but was selectively controlled in actinomycetes.

• Bulletin of the Korean Chemical Society
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• v.34 no.1
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• pp.154-158
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• 2013

Development of dual functional liposome has been studied for cancer theragnostics. Therefore, we focused on ultrasound-sensitive liposomes with doxorubicin (DOX) and gadolinium (Gd) as a theragnostic carrier having a potential for cancer therapy and diagnosis. In this study, Gd(III)-DOTA-modified sonosensitive liposomes (GL) was developed using chemically synthesized Gd(III)-DOTA-DPPE lipid. Sonosensitivity of GL to 1 MHz ultrasound induced 25% of DOX release. The relaxivities ($r_1$) of GL were $7.33-10.34\;mM^{-1}s^{-1}$, which was higher than that of MR-bester$^{(R)}$. Intracellular delivery of DOX from GL by ultrasound irradiation was evaluated according to ultrasound intensity, resulting in increase of uptake of DOX released from ultrasound-triggered GLs compared to GL3 or Doxil$^{(R)}$ without ultrasound. Taken together, this study shows that the paramagnetic and sonosensitive liposomes, GL, is a novel and highly effective delivery system for drug with the potential for broad applications in human disease.

• Journal of Life Science
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• v.24 no.7
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• pp.798-803
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• 2014

Cathepsin D (CtsD), an aspartyl peptidase, is involved in apoptosis, resulting in the release of cytochrome C from mitochondria in cells. Here, we investigated microRNA regulation of CtsD expression in 3T3-L1 cells First, we observed the expression of CtsD in cells in response to doxorubicin (Dox). As expected, the level of CtsD mRNA was increased in 3T3-L1 cells exposed to Dox in a dose-dependent manner. Cellular viability of ectopically expressed CtsD cells was also decreased. Next, we used the miRanda program to search for particular microRNA targeting CtsD. MiR-145 was selected as a putative controller for CtsD because miR-145 had a high mirSVR score. In a reporter assay, the luciferase activity of cells containing the CtsD 3'-UTR region was decreased in cells transfected with miR-145 mimic compared to that of a control. The level of CtsD expression was down-regulated in preadipocytes ectopically expressing miR-145 and up-regulated by an miR-145 inhibitor. Cells also suppressed miR-145 expression when exposed to Dox. The miR-145 inhibitor reduced the cellular viability of 3T3-L1 cells. Taken together, these data suggest that miR-145 regulates CtsD-mediated cell death in adipocytes. These findings may have valuable implications concerning the molecular mechanism of CtsD-mediated cell death in obesity, suggesting that CtaD could be a useful therapeutic tool for the prevention and treatment of obesity by regulating fat cell numbers.

• Journal of Life Science
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• v.24 no.7
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• pp.777-783
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• 2014

Cathepsin D (CtsD), an aspartyl peptidase, is involved in apoptosis, resulting in the release of cytochrome C from mitochondria in cells. Here, we investigated microRNA regulation of CtsD expression in 3T3-L1 cells. First, we observed the expression of CtsD in cells in response to doxorubicin (Dox). As expected, the level of CtsD mRNA increased in 3T3-L1 cells exposed to Dox in a dose-dependent manner. The cellular viability of ectopically expressed CtsD cells was decreased. Next, we used the miRanda program to search for particular microRNA targeting CtsD. MiR-145 was selected as a putative controller of CtsD because it had a high mirSVR score. In a reporter assay, the luciferase activity of cells containing the CtsD 3'-UTR region decreased in cells transfected with a miR-145 mimic compared to that of a control. The level of CtsD expression was down-regulated in preadipocytes ectopically expressing miR-145 and up-regulated by an miR-145 inhibitor. Cells also suppressed miR-145 expression when exposed to Dox. The miR-145 inhibitor reduced the cellular viability of 3T3-L1 cells. Taken together, these data suggest that miR-145 regulates CtsD-mediated cell death in adipocytes. These findings may have valuable implications concerning the molecular mechanism of CtsD-mediated cell death in obesity, suggesting that CtsD could be a useful therapeutic tool for the prevention and treatment of obesity by regulating fat cell numbers.

• Molecules and Cells
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• v.38 no.11
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• pp.959-965
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• 2015

Inducible and reversible gene silencing in desired types of cells is instrumental for deciphering gene functions using cultured cells or in vivo models. However, efficient conditional gene knockdown systems remain to be established. Here, we report the generation of an inducible expression system for short hairpin RNA (shRNA) targeted to PTEN, a well-documented dual-specificity phosphatase involved in tumor suppression and ontogenesis. Upon induction by doxycycline (DOX), the reverse tetracycline transcriptional activator (rtTA) switched on the concomitant expression of GFP and a miR-30 precursor, the subsequent processing of which released the embedded PTEN-targeted shRNA. The efficacy and reversibility of PTEN knockdown by this construct was validated in normal and neoplastic cells, in which PTEN deficiency resulted in accelerated cell proliferation, suppressed apoptosis, and increased invasiveness. Transgenic mice harboring the conditional shRNA-expression cassette were obtained; GFP expression and concurrent PTEN silencing were observed upon ectopic expression of rtTA and induction with Dox. Therefore, this study provides novel tools for the precise dissection of PTEN functions and the generation of PTEN loss of function models in specific subsets of cells during carcinogenesis and ontogenesis.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.8
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• pp.3213-3222
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• 2015

Background: Cancer metastasis depends on cell motility which is driven by cycles of actin polymerization and depolymerization. Reactive oxygen species (ROS) and metabolic oxidative stress have long been associated with cancer. ROS play a vital role in regulating actin dynamics that are sensitive to oxidative modification. The current work aimed at studying the effects of sub-lethal metabolic oxidative stress on actin cytoskeleton, focal adhesion and cell migration. Materials and Methods: T47D human breast cancer cells were treated with 2-deoxy-D-glucose (2DG), L-buthionine sulfoximine (BSO), or doxorubicin (DOX), individually or in combination, and changes in intracellular total glutathione and malondialdehyde (MDA) levels were measured. The expression of three major antioxidant enzymes was studied by immunoblotting, and cells were stained with fluorescent-phalloidin to evaluate changes in F-actin organization. In addition, cell adhesion and degradation ability were measured. Cell migration was studied using wound healing and transwell migration assays. Results: Our results show that treating T47D human breast cancer cells with drug combinations (2DG/BSO, 2DG/DOX, or BSO/DOX) decreased intracellular total glutathione and increased oxidized glutathione, lipid peroxidation, and cytotoxicity. In addition, the drug combinations caused a reduction in cell area and mitotic index, prophase arrest and a decreased ability to form invadopodia. The formation of F-actin aggregates was increased in treated T47D cells. Moreover, combination therapy reduced cell adhesion and the rate of cell migration. Conclusions: Our results suggest that exposure of T47D breast cancer cells to combination therapy reduces cell migration via effects on metabolic oxidative stress.

• Mycobiology
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• v.33 no.1
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• pp.12-14
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• 2005

The culture conditions for the enhanced mycelial growth of Ramaria botrytis was investigated. The optimal temperature and pH for the mycelial growth were $24^{\circ}C$ and 5.0, respectively. It was shown that starch was best of several carbon sources in Czapek-Dox medium as a minimal medium for the enhanced mycelial growth. Organic nitrogen sources were better than inorganic ones for mycelial growth. The appropriate vitamin and mineral salt were biotin and FeCl3, respectively. When this strain was cultured with $FeCl_3$ for 30 days, 19.23 g/l of dry mycelium of R. botrytis was obtained.

• Research in Plant Disease
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• v.15 no.3
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• pp.209-216
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• 2009

During the period of June, 2005 to May, 2008, 44 host plants infected with powdery mildew were collected in the Jeon-ju and Jang-su districts of Jeonbuk province and in the Jang-sung district of Jeonnam province. The hyperparasites, Ampelomyces were confirmed in 12 plant species. Most of the pycnidium shapes of Ampelomyces were circular or oval shaped, and the sizes were different even within the same host plant, and also the color of pycnidium was ranged from light brown to dark brown. Ampelomyces species were isolated from 4 hosts including Impatiens balsamina L., Cucurbita pepo, Rudbeckia laciniata var. elatier and Youngia sonchifolia, and thus the most appropriate 12 Ampelomyces strains for the current experiment were selected. When analyzing the selected 12 strains' incubational and nutritional characteristics, the malt extract agar was the most appropriate media. When investigating the effect of osmotic pressure on the spore germination, 0.15M NaCl concentration was the optimum germination concentration. When the isolated Ampelomyces sp. was tested in-vitro, it was found to be effective to control in other plant pathogens, isolated Ampelomyces showed no pathogenicity to the plant. strains isolated . studied on rDNA ITS sequence analysis. The rDNA ITS sequence data of Ampelomyces sp. isolate BSLAH16 from Impatiens balsamina L. were analyzed and identified.